Test Methods for Evaluating Solid Waste Physical/Chemical Methods SW-846 3rd Edition Update 4a Draft


DRAFT UPDATE IVA
Cover Sheet
THIS PACKET CONTAINS NEW AND REVISED MATERIAL
BEING CONSIDERED FOR INCLUSION IN:
TEST METHODS FOR EVALUATING SOLID WASTE
PHYSICAL/CHEMICAL METHODS
(SW-846) THIRD EDITION
Contents:
1.	Cover sheet. (What you are currently reading)
2.	Instructions. Read this section! It explains how Draft Update IVA relates;
to the rest of your SW-846.
3.	Draft Update IVA Table of Contents. The Table of Contents (dated
January 1998) lists all of the methods (Third Edition, Updates I, II, IIA,
IIB, III, and Draft Update IVA) in the order in which they will appear in
the manual when Update IVA is finalized.
4.	Revised Chapter Two: Choosing the Right Method
5.	Revised Chapter Three and new/revised methods for inorganic analyses.
6.	Revised Chapter Four and new/revised methods for organic analyses.
7.	Revised Chapter Five and a new method for miscellaneous analyses.

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INSTRUCTIONS
SW-846, a methods manual, is a "living" document that changes when new data and
advances in analytical techniques are incorporated into the manual as new or revised
methods. To date, the Agency has formally issued proposed and promulgated Updates
I, H, HA, BOB, and 111, This package contains Draft Update IVA. For specific and
important information regarding this update, please read the section below entitled
"About Draft Update IVA,"
These instructions describe how to get your basic manual up-to-date and what to do
with your Draft Update IVA package. Additional updates will be released by the
Agency in the future. New instructions, to supersede these, will be included with each
of those new update releases. In general, final updates should always be incorporated
into SW-846 in chronological order (e.g. Update I should be incorporated before
Update II).
The following definitions are provided to you as a guide:
New subscribers are defined as individuals who have recently (6-8 weeks) placed an order
with the GPO and have received new copies of the 4 (four) volume set of the Third Edition,
a copy of Final Update I, a copy of Final Update WHA, a copy of Final Update HB, a copy
of Update HI, and a copy of Draft Update IVA.
Previous subscribers are defined as individuals that have received copies of the Third
Edition and other SW-846 Updates (including proposed Updates) in the past and have just
received their Draft Update IVA package in the mail.
Instructions -1
Draft Update IVA
January 1998

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BACKGROUND INFORMATION
A number of SW-846 update packages have been released to the public since the original Third
Edition was released. The dates and labels on these packages can be confusing. The following is
a brief summary of what new subscribers and previous subscribers should check upon receipt of the
Draft Update IVA package:
NEW SUBSCRIBERS - If you are a new subscriber, you should perform the following tasks before
addressing your new Draft Update IVA:
•	Place the original Third Edition of SW-846 (September 1986) in the properly labeled four
3-ring notebooks according to the instructions in Update in.
Incorporate Final Update I (My 1992) into the manual according to the instructions in
Update m.
*	Incorporate Final Updates II (September 1994) and HA (August 1993) into the manual
according to the instructions in Update HL
•	Incorporate Final Update IIB (January 1995) into the manual according to the instructions
in Update m.
*	Incoiporate Final Update HI (December 1996) into the manual according to the instructions
in Update m
PREVIOUS SUBSCRIBERS - If you are a previous subscriber, it is important to establish exactly
what is currently contained in your manual before addressing Draft Update IVA. If your manual
is properly updated, the ONLY white pages in the document should be dated September 1986 (Third
Edition), July 1992 (Final Update I), August 1993 (Final Update HA), September 1994 (Final
Update II), January 1995 (Final Update KB), and December 1996 (Final Update III). Remove (and
recycle or archive) any white pages from your manual that have any other dates. There may also
be colored pages (e.g., pink pages for Proposed Update E3) inserted in the manual. Remove all
yellow, blue, green, or pink pages from the manual. These colored pages represent versions of
methods and chapters that are not final. (Some individuals may chose to keep their copies of
colored versions in separate binders.)
Instructions - 2
Draft Update IVA
January 1998

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TIPDATE HISTORY OF SW-846
The table below can be used as an aid to understanding the update history of SW-846, Third Edition,
Finalized updates are printed in bold and underlined. An individual or organization that has held
an SW-846 GPO subscription for several years may have received copies of any or all of the
updates.
•
A BRIEF HISTORY OF THE SW-846, THIRD EDITION AND ITS UPDATES
Package
Date Listed on Methods
Color of Paper
Status of PackaEe
Third Edition
September 1986
White
Finalized (Promulgated)
Proposed Update I
December 1987
Green
Obsolete
Final Update I
(Released by accident)
November 1990
White
Obsolete! Neva-formally
finalized.
Proposed Update n (Released
by accident)
November 1990
Blue
Obsolete! Never formally
proposed.
Final Update I
July 1992
White
Finalized (Promulgated)
Proposed Update II
November 1992
Yellow
Obsolete
Proposed Update HA*
(Available from EPA by
request only.)
October 1992
White
Obsolete
Pinal Update HA* ("Included
with Final Update U.)
August 1993
White
Finalized (Promulgated)
Final UDdate FT
September 1994
White
Finalized (Promulgated)
Final TIndate HB**
January 1995
White
Finalized (Promulgated)
Proposed Update HI
January 1995
Pink
Proposed
Final Update HI
December 1996
White
Finalized (Promulgated)
Draft Update IVA
January 1998
Salmon
Draft
* Contains only Method 4010.
** Contains only a revised Table of Contents, a revised Chapter Six, and revised Methods 9040B and 9045C
Instructions - 3
Draft Update IVA
January 1998

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ABOUT DRAFT UPDATE IVA
Draft Update IVA has been issued by the EPA's Office of Solid Waste and contains methods which
are being considered for inclusion someday in the SW-846 methods manual. "Die Draft Update IVA
package includes 15 revised methods, four revised chapters, a revised Table of Contents, and 13 new
methods. In addition, Draft Update IVA involves the removal or integration of 44 other methods.
Please see the section below entitled "Removal or Integration of Some Methods in SW-846."
In order to distinguish this draft update from other SW-846 updates, it has been printed on salmon-
colored paper. (Final updates are printed on white paper.) The date "January 1998" is found in the
lower right-hand corner of each page in the Draft Update IVA package. This date should be used
to definitively distinguish the Draft Update IVA versions of SW-846 methods from previous and
future versions.
A notice announcing the availability of Draft Update IVA has been published in the Federal
Register and invites public comment on its content EPA has published the notice and made this
draft update available for informational purposes only, and is not at this time formally proposing
to revise SW-846 by adding Update IVA to it or to incorporate the update in the RCRA regulations
for required uses of SW-846 methods. EPA is only making the Agency-reviewed methods of
Update IVA available to the public early, for guidance purposes.
The aforementioned notice fully explains the Agency's plans regarding this update, and EPA
encourages the public to read the notice. As explained in the notice, several regulations under
subtitle C of RCRA currently require that certain SW-846 methods be employed. However, any
reliable analytical method may be used to meet other requirements in 40 CFR parts 260 through 270.
The methods of Update IVA fall in the category of "any reliable method." The methods may
currently be used in all applications for which the use of SW-846 methods is not mandatory. The
methods of Draft Update IVA, however, cannot be used for compliance with required uses of SW-
846 methods. The Agency also recommends that the regulated community obtain permission from
the appropriate regulating entity, if required under state or local regulations, before using these
methods for non-mandatory applications.
A copy of the Federal Register announcing the availability of Draft Update IVA (and copies of
other Federal Register documents) can be accessed from EPA's web ate at the following two
Internet locations:
http://www.epa.gov/epahome/rules.html
http://www.epa.gov/fedrgstr
Instructions - 4
Draft Update IVA
January 1998

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REQUEST FOR PUBLIC COMMENTS ON DRAFT UPDATE IVA
Hie EPA is requesting public comment on the methods in Update IVA (e.g., requesting suggestions
or recommendations regarding the content of the methods). Comments on the suggested changes
to the Draft Update IVA methods package, and other topics addressed in the Update IVA Federal
Register notice, must be submitted within the time period specified in the "Dates" section of the
notice. SW-846 methods not contained in this package are not open to comment.
The Agency is interested in comments on the content of all sections or parts of new methods to SW-
846 found in Draft Update IVA. Regarding the revised methods and chapters, the Draft Update IVA
Federal Register notice contains a table identifying those parts of each document which are open
for comment EPA is interested in comments from the public on only the identified parts because
other parts of the methods have not been significantly revised from the promulgated version
currently in SW-846. Please see the Draft Update IVA Federal Register for this information before
developing and submitting comments on revised methods found in Draft Update IVA.
HOW TO SUBMIT COMMENTS
As explained in the "Addresses" section of the Draft Update IVA Federal Register Notice,
comraenters must send an original and two copies of their comments referencing docket number
F-98-4TMA-FFFFF to: RCRA Information Center (RIC), Office of Solid Waste (5305G), U.S.
Environmental Protection Agency Headquarters (EPA, HQ), 401 M Street, S.W., Washington, DC
20460. Courier deliveries of comments should be submitted to the RIC at the address listed below.
Comments may also be submitted electronically through the Internet to the following address:
RCRA-docket@epamail.epa.gov. Comments in electronic format should be identified by the docket
number F-98-4TMA-FFFFF. Submit electronic comments as an ASCII (TEXT) file and avoid the
use of special characters and any form of encryption. EPA's Office of Solid Waste (OSW) also
accepts comments and data on diskettes in WordPerfect 6.1 file format. On the disk label, specify
the coramenter's name and the word processing software and version/edition.
Commenters should not submit electronically any confidential business information (CBI). An
original and two copies of the CBI must be submitted under separate cover to: Regina Magbie,
RCRA CBI Document Control Officer, Office of Solid Waste (5305W), U.S. EPA, 401 M Street,
S.W., Washington, DC 20460.
Public comments and supporting materials are available for viewing in the RIC, located at Crystal
Gateway One, 1235 Jefferson Davis Highway, First Floor, Arlington, Virginia. The RIC is open
from 9 a.m. to 4 p.m., Monday through Friday, except for Federal holidays. To review docket
materials or make photocopies, the public must make an appointment by calling 703-603-9230.
Instructions - 5
Draft Update IVA
January 1998

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REMOVAL OR INTEGRATION OF SOME METHODS IN SW-846
The Agency is also requesting comment on the removal of one obsolete headspace method and the
removal of 43 atomic absorption methods. The individual methods are being deleted as part of Draft
Update IVA because their inclusion is redundant given that their procedures and target analytes have
been fully integrated into revised Method 7000B and new Method 7010, the general methods for
the techniques. The table to follow is a list of the methods being considered for deletion from the
manual.
METHODS BEING CONSIDERED FOR REMOVAL FROM SW-846
Method No.
Method Title
38iOw
Headspace
7020(b)
Aluminum (Atomic Absorption, Direct Aspiration)
7040(b)
Antimony (Atomic Absorption, Direct Aspiration)
7041)
Iron (Atomic Absorption, Direct Aspiration)
7381te)
Iron (Atomic Absorption, Furnace Technique)
Instructions - 6
Draft Update IVA
January 1998

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7420w
Lead (Atomic Absorption, Direct Aspiration)
7421
lithium (Atomic Absorption, Direct Aspiration)
7450w
Magnesium (Atomic Absorption, Direct Aspiration)
7460(b)
Manganese (Atomic Absorption, Direct Aspiration)
7461(c)
Manganese (Atomic Absorption, Furnace Technique)
7480(b)
Molybdenum (Atomic Absorption, Direct Aspiration)
748 l(c)
Molybdenum (Atomic Absorption, Furnace Technique)
7520w
Nickel (Atomic Absorption, Direct Aspiration)
7521(c)
Nickel (Atomic Absorption, Furnace Method)
7550°°
Osmium (Atomic Absorption, Direct Aspiration)
7610w
Potassium (Atomic Absorption, Direct Aspiration)
7740(c)
Selenium (Atomic Absorption, Furnace Technique)
7760A(b)
Silver (Atomic Absorption, Direct Aspiration)
7761(c)
Silver (Atomic Absorption, Furnace Technique)
7770^'
Sodium (Atomic Absorption, Direct Aspiration)
778(f)
Strontium (Atomic Absorption, Direct Aspiration)
7840w
Thallium (Atomic Absorption, Direct Aspiration)
7841(c)
Thallium (Atomic Absorption, Furnace Technique)
7B70W
Tin (Atomic Absorption, Direct Aspiration)
7910Cb)
Vanadium (Atomic Absorption, Direct Aspiration)
7911
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In addition, Chapter Eleven is being considered for deletion from SW-846. If Chapter Eleven is
removed, the manual will amply refer to the most current version of the ground-water monitoring
guidance published by the Office of Solid Waste. (See the Draft Update IVA Federal Register
notice for more information regarding the removal of Chapter Eleven.)
HANDLING OF DRAFT UPDATE IVA
Since these are only draft methods and chapters, the material in Draft Update IVA does not change
anything in the official version of the manual (Le., SW-846, Third Edition, as updated by Final
Updates I, n, HA, IIB, and III). The user should not remove any white pages from the manual at
this time. Regarding the placement and storage of this draft update, the Agency recommends one
of the following:
1. The subscriber may place the salmon sheets in the manual (without removing the white
pages of methods) in die order that they appear in the Draft Update IVA Table of Contents.
Due to the volume of material, the subscriber can split the material in any volume (e.g.,
Volume IIB) into two parts and place one part into an extra binder supplied by the
subscriber.
2. Instead of inserting die draft methods into the manual with the final methods, the subscriber
may instead place the entire Draft Update IVA package into a separate binder supplied by
the subscriber.
IN SUMMARY
To summarize these instructions, please note the following important points:
This package contains Draft Update IVA. The USEPA is issuing these methods for possible
future inclusion in SW-846. The draft methods are reliable methods, approved for use in
all applications for which the use of SW-846 methods is not mandatory. The methods of
Draft Update IVA cannot be used for compliance with required uses of SW-846 methods.
The public may submit comments to the EPA regarding these methods in paper and/or
electronic format SW-846 methods not contained in this package are not open to comment.
Do not remove any white pages from your copy of SW-846 and replace them with Draft
Update IVA pages.
If you have properly inserted all other updates, Draft Update IVA will not fit in the existing
four 3-ring binder note books provided with the manual. You may insert (without replacing
any white pages) Draft Update IVA into the manual and split some of a volume into a
separate binder that you supply. You may also keep Draft Update IVA in its own separate
binder that you supply.
Instructions - 8
Draft Update IVA
January 1998
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ASSISTANCE
After reading these instructions, if you need help due to difficulties understanding the status of the
package or have technical questions regarding the methods, you may telephone the Methods
Information Communication Exchange (MICE) at 703-821-4690 or send an E-mail to:
mice@lan828.ehsg.saic.com.
If you have questions concerning your SW-846 U.S. Government Printing Office (GPO)
subscription, you should telephone the GPO at 202-512-1806. If you did not purchase your SW-846
from the GPO, the GPO will not be able to help you.
SW-846 AVAILABILITY ON CD-ROM
A CD-ROM version (Version 2.0) of Test Methods for Evaluating Solid Waste, Physical/Chemical
Methods (SW-846) has been developed by EPA in cooperation with the National Technical
Information Service (NllS). On a single disc, it includes all text and figures found in the final
version of SW-846 as updated by Updates I, n, DA, IIB, and HI. (It does not include Draft Update
IVA.) It can be used for word searching (e.g, analytes, keywords); and to cut and paste or export
text and diagrams to update or develop laboratory standard operating procedures (SOPs). To order
by phone, call NTIS at (800) 553-6847 and request order number PB97-501928FCD for a single
user copy, PB97-502512FCD for a 5-user LAN copy, or PB97-502520FCD for unlimited users.
To receive information by fax from NTIS about this CD-ROM, call (703) 487-4140 and enter
publication number code 8698.
A CD-ROM version of Draft Update IV on CD-ROM is planned for the future. To place an early
order by phone, call NTIS at (800) 553-6847 and request order number PB97-501936 (single user),
PB97-502538FCD (5-user LAN), or PB97-502546FCD (unlimited users).
Instructions - 9
Draft Update IVA
January 1998
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TABLE OF CONTENTS
VOLUME ONE
SECTJON A
DISCLAIMER
ABSTRACT
TABLE OF CONTENTS
METHOD INDEX AND CONVERSION TABLE
PREFACE
ACKNOWLEDGEMENTS
PART 1 METHODS FOR ANALYTES AND PROPERTIES
CHAPTER ONE - QUALITY CONTROL
1.0
Introduction
2.0
OA Project Plan
3.0
Field Operations
4.0
Laboratory Operations
5.0
Definitions
6.0
References
CHAPTER TWO - CHOOSING THE CORRECT PROCEDURE
2.1 Guidance Regarding Flexibility Inherent to SW-846 Methods and the Precedence of SW-846
Quality Control Criteria
2.2 Required Information
2.3 Implementing the Guidance
2.4 Characteristics
2.5 Ground Water
2.6 References
CHAPTER THREE - INORGANIC ANALYTES
3.1 Sampling Considerations
3.2 Sample Digestion Methods
Method 3005A:
Method 301 OA:
Method 3015A:
Method 3020A:
Acid Digestion of Waters for Total Recoverable or Dissolved Metals
for Analysis by FLAA or ICP Spectroscopy
Add Digestion of Aqueous Samples and Extracts for Total Metals for
Analysis by FLAA or ICP Spectroscopy
Microwave Assisted Acid Digestion of Aqueous Samples and Extracts
Acid Digestion of Aqueous Samples and Extracts for Total Metals for
Analysis by GFAA Spectroscopy
CONTENTS -1
Revision 5
January 1998
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Method 3031:
Method 3040A:
Method 3050B:
Method 3051A:
Acid Digestion of Oils for Metals Analysis by Atomic Absorption or
ICP Spectrometry
Dissolution Procedure for Oils, Greases, or Waxes
Acid Digestion of Sediments, Sludges, and Soils
Microwave Assisted Add Digestion of Sediments, Sludges, Soils, and
Oils
Microwave Assisted Acid Digestion of Siliceous and Organically
Based Matrices
Alkaline Digestion for Hexavalent Chromium
Method 3052:
Method 3060A:
Methods for Determination of Inorganic Analytes
Method 4500:
Method 601 OB:
Method 6020A:
Method 6200:
Method 6500:
Method
Method
Method
Method
Method
Method
6800:
7000B:
7010:
7061A:
7062:
7063:
Method 7195:
Method 7196A:
Method 7197:
Method 7198:
Method 7199:
Method
7470A:
Method
7471B:
Method
7472:
Method
7473:
Method
7474:
Method
7580:
Method
7741A:
Method
7742:
Method
9000:
Method
9001:
Mercury in Soil by Immunoassay
Inductively Coupled Plasma-Atomic Emission Spectrometry
Inductively Coupled Plasma - Mass Spectrometry
Field Portable X-Ray Fluorescence Spectrometry for the Determination
of Elemental Concentrations in Soil and Sediment
Dissolved Inorganic Anions in Aqueous Matrices by Capillary Ion
Electrophoresis
Elemental and Speciated Isotope Dilution Mass Spectrometry
Flame Atomic Absorption Spectrophotometry
Graphite Furnace Atomic Absorption Spectrophotometry
Arsenic (Atomic Absorption, Gaseous Hydride)
Antimony and Arsenic (Atomic Absorption, Borohydride Reduction)
Arsenic in Aqueous Samples and Extracts by Anodic Stripping
Voltammetry (ASV)
Chromium, Hexavalent (Coprecipitation)
Chromium, Hexavalent (Colorimetric)
Chromium, Hexavalent (Chelation/Extraction)
Chromium, Hexavalent (Differential Pulse Polarography)
Determination of Hexavalent Chromium in Drinking Water,
Groundwater and Industrial Wastewater Effluents by Ion
Chromatography
Mercury in Liquid Waste (Manual Cold-Vapor Technique)
Mercury in Solid or Semisolid Waste (Manual Cold-Vapor Technique)
Mercury in Aqueous Samples and Extracts by Anodic Stripping
Voltammetry (ASV)
Mercury in Solids and Solutions by Thermal Decomposition,
Amalgamation, and Atomic Absorption Spectrophotometry
Mercury in Sediment and Tissue Samples by Atomic
Fluorescence Spectrometry
White Phosphorus (P4) by Solvent Extraction and Gas
Chromatography
Selenium (Atomic Absorption, Gaseous Hydride)
Selenium (Atomic Absorption, Borohydride Reduction)
Determination of Water in Waste Materials by Karl Fischer Titration
Determination of Water in Waste Materials by Quantitative Calcium
Hydride Reaction
CONTENTS-2
Revision 5
January 1998
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NOTE: A suffix of "A" in the method number indicates revision one (the method has
been revised once), A suffix of "B" in the method number indicates revision two (the
method has been revised twice). A suffix of "C" in the method number indicates
revision three (the method has been revised three times). In order to properly
document the method used for analysis, the entire method number including the
suffix tetter designation (e.g., A, B, or C) must be identified by the analyst. A
method reference found within the RCRA regulations and the text of SW-846 methods
and chapters refers to the latest promulgated revision of the method, even though the
method number does not include the appropriate letter suffix.
CONTENTS - 3
Revision 5
January 1998
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VOLUME ONE
SECTION B
DISCLAIMER
ABSTRACT
TABLE OF CONTENTS
METHOD INDEX AND CONVERSION TABLE
PREFACE
ACKNOWLEDGEMENTS
CHAPTER ONE. REPRINTED - QUALITY CONTROL
1.0
Introduction
2.0
OA Project Plan
3.0
Field Operations
4.0
Laboratory Operations
5.0
Definitions
6.0
References
CHAPTER FOUR - ORGANIC ANALYTES
4.1 Sampling Considerations
4.2 Sample Preparation Methods
4.2.1 Extractions and Preparations
Organic Extraction and Sample Preparation
Separatory Funnel Liquid-Liquid Extraction
Continuous Liquid-Liquid Extraction
Solid-Phase Extraction (SPE)
Soxhlet Extraction
Automated Soxhlet Extraction
Extraction of Semivolatile Analytes Collected Using Method 0010
(Modified Method 5 Sampling Train)
Pressurized Fluid Extraction (PFE)
Ultrasonic Extraction
Supercritical Fluid Extraction of Total Recoverable Petroleum
Hydrocarbons
Supercritical Fluid Extraction of Polynuclear Aromatic Hydrocarbons
Supercritical Fluid Extraction of Polychlorinated Biphenyls (PCBs) and
Organochlorine Pesticides
Waste Dilution
Waste Dilution for Volatile Organics
Sample Preparation for Volatile Organic Compounds
Volatile Organic Compounds in Soils and Other Solid Matrices Using
Equilibrium Headspace Analysis
CONTENTS-4 Revision 5
January 1998
Method
3500B:
Method
3510C:
Method
3520C:
Method
3535A:
Method
3540C:
Method
3541:
Method
3542:
Method
3545A:
Method
3550B:
Method
3560:
Method
3561:
Method
3562:
Method
3580A:
Method
3585:
Method
5000:
Method
5021:
-------
Method 5030B:
Method 5031:
Method 5032:
Method 5035:
Method 5041A:
Purge-and-Trap for Aqueous Samples
Volatile, Nonpurgeable, Water-Soluble Compounds by Azeotropic
Distillation
Volatile Organic Compounds by Vacuum Distillation
Closed-System Purge-and-Trap and Extraction for Volatile Organics
in Soil and Waste Samples
Analysis for Desorption of Sorbent Cartridges from Volatile Organic
Sampling Train (VOST)
4.2.2
Cleanup
Method 3600C:
Method 361 OB:
Method 3611B:
Method 3620B:
Method 3630C:
Method 3640A:
Method 3650B:
Method 3660B:
Method 3665A:
Cleanup
Alumina Cleanup
Alumina Column Cleanup and Separation of Petroleum Wastes
Florisil Cleanup
Silica Gel Cleanup
Gel-Permeation Cleanup
Acid-Base Partition Cleanup
Sulfur Cleanup
Sulfuric Acid/Permanganate Cleanup
Determination of Organic Analytes
4.3.1
Gas Chromatographic Methods
Method 8000B:
Method 8011:
Method 8015B:
Method 8021B:
Method 8031:
Method 8032A:
Method 8033:
Method 8041:
Method 8061A:
by
Method
Method
Method
Method
Method
Method
Method
8070A:
8081B:
8082A:
8091:
8100:
8111:
8121:
Method 8131:
Method 8141B:
Method 8151 A:
Determinative Chromatographic Separations
1,2-Dibromoethane and 1,2-Dibromo-3-chloropropane
Microextpaction and Gas Chromatography
Nonhalogenated Organics Using GC/FID
Aromatic and Halogenated Voiatifes by Gas Chromatography Using
Photoionization and/or Electrolytic Conductivity Detectors
Acrylonitrile by Gas Chromatography
Acrylamide by Gas Chromatography
Acetonitrile by Gas Chromatography with Nitrogen-Phosphorus
Detection
Phenols by Gas Chromatography
Phthalate Esters by Gas Chromatography with Electron Capture
Detection (GC/ECD)
Nitrosamines by Gas Chromatography
Organochlorine Pesticides by Gas Chromatography
Polychlorinated Biphenyis (PCBs) by Gas Chromatography
Nitroaromatics and Cyclic Ketones by Gas Chromatography
Polynuclear Aromatic Hydrocarbons
Haloethers by Gas Chromatography
Chlorinated Hydrocarbons by Gas Chromatography: Capillary
Column Technique
Aniline and Selected Derivatives by Gas Chromatography
Organophosphorus Compounds by Gas Chromatography
Chlorinated Herbicides by GC Using Methyl ation or
Pentafluoro be nzylation Derivatization
CONTENTS - 5
Revision 5
January 1998
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4.3.2
Gas Chromatographic/Mass Spectroscopic Methods
Method 8260B:
Method 8270D:
Method 827SA:
Method 8280B:
Method 8290A:
Appendix A;
Volatile Organic Compounds by Gas Chromatography/Mass
Spectrometry (GC/MS)
Semivolatile Organic Compounds by Gas Chromatography/Mass
Spectrometry (GC/MS)
Semivolatile Organic Compounds (PAHs and PCBs) in Soils/Sludges
and Solid Wastes Using Thermal Extraction/Gas
Chromatography/Mass Spectrometry (TE/GC/MS)
Polychlorinated Dibenzo-p-Dioxins and Polychlorinated Dibenzofurans
by High Resolution Gas Chromatography/Low Resolution Mass
Spectrometry (HRGC/LRMS)
Polychlorinated Dibenzodioxins (PCDDs) and Polychlorinated
Dibenzofurans (PCDFs) by High-Resolution Gas
Chromatography/High-Resolution Mass Spectrometry (HRGC/HRMS)
Procedures for the Collection, Handling, Analysis, and
Reporting of Wipe Tests Performed within the Laboratory
4.3.3
High Performance Liquid Chromatographic Methods
Method 8310:
Method 8315A:
Appendix A:
Method 8316:
Method 8318:
Method 8321B:
Method 8325:
Method 8330A:
Method 8331:
Method 8332:
Polynuclear Aromatic Hydrocarbons
Determination of Carbonyl Compounds by High Performance Liquid
Chromatography (HPLC)
Recrystallization of 2,4-Dinitrophenylhydrazine (DNPH)
Acrylamide, Acrylonitrile and Acrolein by High Performance Liquid
Chromatography (HPLC)
N-Methylcarbamates by High Performance Liquid Chromatography
(HPLC)
Solvent-Extractable Nonvolatile Compounds by High Performance
Liquid Chromatography/Thermospray/Mass Spectrometry
(HPLC/TS/MS) or Ultraviolet (UV) Detection
Solvent Extractable Nonvolatile Compounds by High Performance
Liquid Chromatography/Particte Beam/Mass Spectrometry
(HPLC/PB/MS)
Nitroaromatics and Nitramines by High Performance Liquid
Chromatography (HPLC)
Tetrazene by Reverse Phase High Performance Liquid
Chromatography (HPLC)
Nitroglycerine by High Performance Liquid Chromatography
4.3.4
Infrared Methods
Method 8410:
Method 8430:
Method 8440:
Gas Chromatography/Fourier Transform Infrared (GC/FT-IR)
Spectrometry for Semivolatile Organics: Capillary Column
Analysis of Bis(2-chloroethyl) Ether and Hydrolysis Products by Direct
Aqueous Injection GC/FT-IR
Total Recoverable Petroleum Hydrocarbons by Infrared
Spectrophotometry
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4.3.5
Miscellaneous Spectrometric Methods
4.4
Method 8520: Continuous Measurement of Formaldehyde in Ambient Air
Immunoassay Methods
Method 4000:
Method 401 OA:
Method 4015:
Method 4020:
Method 4030:
Method 4035:
Method 4040
Method 4041
Method 4042:
Method 4050
Method 4051
Method 4670
Immunoassay
Screening for Pentachlorophenol by Immunoassay
Screening for 2,4-Dichlorophenoxyacetic Acid by Immunoassay
Screening for Polychlorinated Biphenyls by Immunoassay
Soil Screening for Petroleum Hydrocarbons by Immunoassay
Soil Screening for Polynuclear Aromatic Hydrocarbons by
Immunoassay
Soil Screening for Toxaphene by Immunoassay
Soil Screening for Chtordane by immunoassay
Soil Screening for DDT by Immunoassay
TNT Explosives in Soil by Immunoassay
Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) in Soil by Immunoassay
Triazine Herbicides as Atrazine in Water by Quantitative Immunoassay
4.5 Miscellaneous Screening Methods
Method 3820: Hexadecane Extraction and Screening of Purgeable Organics
Method 8515: Colorimetric Screening Method for Trinitrotoluene (TNT) in Soil
Method 9074: Turbidimetric Screening Method for Total Recoverable Petroleum
Hydrocarbons in Soil
Method 9078: Screening Test Method for Polychlorinated Biphenyls in Soil
Method 9079: Screening Test Method for Polychlorinated Biphenyls in Transformer
Oil
NOTE: A suffix of "A" in the method number indicates revision one (the method has
been revised once). A suffix of "B" in the method number indicates revision two (the
method has been revised twice). A suffix of "C" in the method number indicates
revision three (the method has been revised three times). In order to properly
document the method used for analysis, the entire method number Including the
suffix letter designation (e.g., A, B, or C) must be Identified by the analyst. A
method reference found within the RCRA regulations and the text of SW-846 methods
and chapters refers to the latest promulgated revision of the method, even though the
method number does not include the appropriate letter suffix.
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VOLUME ONE
SECTION C
DISCLAIMER
ABSTRACT
TABLE OF CONTENTS
METHOD INDEX AND CONVERSION TABLE
PREFACE
CHAPTER ONE, REPRINTED - QUALITY CONTROL
1.0 Introduction
2.0 OA Project Plan
3.0 Field Operations
4.0 Laboratory Operations
5.0 Definitions
6.0 References
CHAPTER FIVE - MISCELLANEOUS TEST METHODS
Method 5050:
Method 9010B;
Method 9012A:
Method 9013:
Method 9014:
Method
Method
Method
Method
Method
Method
Method
Method
Method
Method
Method
Method
9020B:
9021:
9022;
9023:
9030B:
9031:
9034:
9035:
9036:
9038:
9056:
9057:
Method 9060:
Method 9065:
Method 9066:
Method 9067:
Method 9070:
Method 9071A:
Bomb Preparation Method for Solid Waste
Total and Amenable Cyanide: Distillation
Total and Amenable Cyanide (Automated Colorimetric, with Off-Line
Distillation)
Cyanide Extraction Procedure for Solids and Oils
Titrimetric and Manual Spectrophotometric Determinative Methods for
Cyanide
Total Organic Halides (TOX)
Purgeable Organic Halides (POX)
Total Organic Halides (TOX) by Neutron Activation Analysis
Extractable Organic Halides (EOX) in Solids
Acid-Soluble and Acid-Insoluble Sulfides: Distillation
Extractable Sulfides
Titrimetric Procedure for Acid-Soluble and Acid-Insoluble Sulfides
Sulfate (Colorimetric, Automated, Chloranilate)
Sulfate (Colorimetric, Automated, Methylthymol Blue, AAII)
Sulfate (Turbidimetric)
Determination of Inorganic Anions by Ion Chromatography
Determination of Chloride from HCI/CI2 Emission Sampling Train
(Methods 0050 and 0051) by Anion Chromatography
Total Organic Carbon
Phenoiics (Spectrophotometric, Manual 4-AAP with Distillation)
Phenoiics (Colorimetric, Automated 4-AAP with Distillation)
Phenoiics (Spectrophotometric, MBTH with Distillation)
Total Recoverable Oil & Grease (Gravimetric, Separatory Funnel
Extraction)
Oil and Grease Extraction Method for Sludge and Sediment Samples
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Method 9075:
Method 9076:
Method 9077:
Method A:
Method B:
Method C:
Method 9131:
Method 9132:
Method 9210:
Method 9211
Method 9212
Method 9213
Method 9214
Method 9215
Method 9216:
Method 9250:
Method 9251:
Method 9253:
Method 9320:
Test Method for Tot a! Chlorine in New and Used Petroleum Products
by X-Ray Fluorescence Spectrometry (XRF)
Test Method for Total Chlorine in New and Used Petroleum Products
by Oxidative Combustion and Microcoulometry
Test Methods for Total Chlorine in New and Used Petroleum
Products (Field Test Kit Methods)
Fixed End Point Test Kit Method ...
Reverse Titration Quantitative End Point Test Kit Method
Direct Titration Quantitative End Point Test Kit Method
Total Coliform: Multiple Tube Fermentation Technique
Total Coliform: Membrane-Filter Technique
Potentiometric Determination of Nitrate in Aqueous Samples with ion-
Selective Electrode
Potentiometric Determination of Bromide in Aqueous Samples with
Ion-Selective Electrode
Potentiometric Determination of Chloride in Aqueous Samples with
Ion-Selective Electrode
Potentiometric Determination of Cyanide in Aqueous Samples and
Distillates with Ion-Selective Electrode
Potentiometric Determination of Fluoride in Aqueous Samples with
Ion-Selective Electrode
Potentiometric Determination of Sulfide in Aqueous Samples and
Distillates with Ion-Selective Electrode
Potentiometric Determination of Nitrite in Aqueous Samples with Ion-
Selective Electrode
Chloride (Colorimetric, Automated Ferricyanide AAI)
Chloride (Colorimetric, Automated Ferricyanide AAII)
Chloride (Titrimetric, Silver Nitrate)
Radium-228
SIX-PROPERTIES
Method 1030:
Ignitability of Solids
Method 1120:
Dermal Corrosion
Method 1312:
Synthetic Precipitation Leaching Procedure
Method 1320:
Multiple Extraction Procedure
Method 1330A:
Extraction Procedure for Oily Wastes
Method 9041A:
pH Paper Method
Method 9045C:
Soil and Waste pH
Method 9050A:
Specific Conductance
Method 9080:
Cation-Exchange Capacity of Soils (Ammonium Acetate)
Method 9081:
Cation-Exchange Capacity of Soils (Sodium Acetate)
Method 9090A:
Compatibility Test for Wastes and Membrane Liners
Method 9095A:
Paint Filter Liquids Test
Method 9096:
Liquid Release Test (LRT) Procedure
Appendix A:
Liquid Release Test Pre-Test
Method 9100:
Saturated Hydraulic Conductivity, Saturated Leachate Conductivity,

and Intrinsic Permeability
Method 9310:
Gross Alpha and Gross Beta
Method 9315:
Alpha-Emitting Radium Isotopes
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PART II CHARACTERISTICS
CHAPTER SEVEN - CHARACTERISTICS INTRODUCTION AND REGULATORY DEFINITIONS
7.1 Ignitability
7.2 Corrosivity
7.3 Reactivity
Test Method to Determine Hydrogen Cyanide Released from Wastes
Test Method to Determine Hydrogen Sulfide Released from Wastes
7.4 Toxicity Characteristic Leaching Procedure
CHAPTER EIGHT - METHODS FOR DETERMINING CHARACTERISTICS
8.1 Ignitability
Method 1010:
Method 1020A:
8.2 Corrosivity
Method 9040B:
Method 1110:
8.3 Reactivity
8.4 Toxicity
Method 131 OA:
Method 1311:
Pensky-Martens Closed-Cup Method for Determining Ignitability
Setaflash Closed-Cup Method for Determining Ignitability
pH Electrometric Measurement
Corrosivity Toward Steel
Extraction Procedure (EP) Toxicity Test Method and Structural
Integrity Test
Toxicity Characteristic Leaching Procedure
NOTE: A suffix of "A" in the method number indicates revision one (the method has
been revised once). A suffix of "B" in the method number indicates revision two (the
method has been revised twice). A suffix of "C" in the method number indicates
revision three (the method has been revised three times). In order to properly
document the method used for analysis, the entire method number Including the
suffix letter designation (e.g., A, B, or C) must be identified by the analyst. A
method reference found within the RCRA regulations and the text of SW-846 methods
and chapters refers to the latest promulgated revision of the method, even though the
method number does not include the appropriate letter suffix.
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VOLUME TWO
DISCLAIMER
ABSTRACT
TABLE OF CONTENTS
METHOD INDEX AND CONVERSION TABLE
PREFACE
CHAPTER ONE, REPRINTED -- QUALITY CONTROL
1.0
Introduction
2.0
OA Project Plan
3.0
Field Operations
4.0
Laboratory Operations
5.0
Definitions
6.0
References
PART 111 SAMPLING
CHAPTER NINE - SAMPLING PLAN
9.1 Design and Development
9.2 Implementation
CHAPTER TEN - SAMPLING METHODS
Method 0010:
Appendix A:
Appendix B:
Method 0011:
Method 0020:
Method 0023A:
Method 0030
Method 0031
Method 0040
Method 0050
Method 0051
Method 0060
Method 0061
Method 0100;
from
and
Modified Method 5 Sampling Train
Preparation of XAD-2 Sorbent Resin
Total Chromatographable Organic Material Analysis
Sampling for Selected Aldehyde and Ketone Emissions
Stationary Sources
Source Assessment Sampling System (SASS)
Sampling Method for Polychlorinated Dibenzo-p-Dioxins
Polychlorinated Dibenzofuran Emissions from Stationary Sources
Volatile Organic Sampling Train
Sampling Method for Volatile Organic Compounds (SMVOC)
Sampling of Principal Organic Hazardous Constituents from
Combustion Sources Using Tedlar® Bags
Isokinetic HCI/CU Emission Sampling Train
Midget Impinger HCI/CI2 Emission Sampling Train
Determination of Metals in Stack Emissions
Determination of Hexavalent Chromium Emissions from Stationary
Sources
Sampling for Formaldehyde and Other Carbonyl Compounds in Indoor
Air
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PART IV MONITORING
CHAPTER ELEVEN - GROUND WATER MONITORING
11.1 Background and Objectives
11.2 Relationship to the Regulations and to Other Documents
11.3 Revisions and Additions
11.4 Acceptable Designs and Practices
11.5 Unacceptable Designs and Practices
CHAPTER TWELVE - LAND TREATMENT MONITORING
12.1 Background
12.2 Treatment Zone
12.3 Regulatory Definition
12.4 Monitoring and Sampling Strategy
12.5 Analysis
12.6 References and Bibliography
CHAPTER THIRTEEN - INCINERATION
13.1 Introduction
13.2 Regulatory Definition
13.3 Waste Characterization Strategy
13.4 Stack-Gas Effluent Characterization Strategy
13.5 Additional Effluent Characterization Strategy
13.6 Selection of Specific Sampling and Analysis Methods
13.7 References
NOTE: A suffix of "A" in the method number indicates revision one (the method has
been revised once). A suffix of "B" in the method number indicates revision two (the
method has been revised twice). A suffix of "C" in the method number indicates
revision three (the method has been revised three times). In order to properly
document the method used for analysis, the entire method number including the
suffix letter designation (e.g., A, B, or C) must be identified by the analyst. A
method reference found within the RCRA regulations and the text of SW-846 methods
and chapters refers to the latest promulgated revision of the method, even though the
method number does not include the appropriate letter suffix.
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CHAPTER TWO
CHOOSING THE CORRECT PROCEDURE
SW-846 analytical methods are written as quantitative trace analytical methods to
demonstrate that a waste does not contain anaiytes of concern that cause it to be managed as a
hazardous waste. As such, these methods typically contain relatively stringent quality control (QC)
criteria appropriate to trace analyses. However, if a particular application does not require data of
this quality, less stringent QC criteria may be used. The purpose of this chapter is to aid the analyst
in choosing the appropriate methods for sample analyses, based upon the sample matrix and the
anaiytes to be determined. The ultimate responsibility for producing reliable analytical results lies
with the entity subject to the regulation. Therefore, members of the regulated community are
advised to refer to this chapter and to consult with knowledgeable laboratory personnel when
choosing the most appropriate suite of analytical methods. In addition, analysts and data users are
advised that, except where explicitly specified in a regulation, the use of SW-846 methods is not
mandatory in response to Federal testing requirements.
Section 2,1 provides guidance regarding the analytical flexibility inherent to SW-846 methods
and the precedence of various QC criteria. Section 2.2 reviews the information required to choose
the correct combination of methods for an analytical procedure. Section 2.3 provides useful
information on implementing the method selection guidance for organic analyses. Section 2.4
provides guidance on characteristic analyses and Section 2.5 provides guidance on the
determination of anaiytes in ground water.
2.1 GUIDANCE REGARDING FLEXIBILITY INHERENT TO SW-846 METHODS AND THE
PRECEDENCE OF SW-846 QUALITY CONTROL CRITERIA
The specific products and instrument settings cited in SW-846 methods represent those
products and settings used during method development or subsequently evaluated by the Agency
for use in the method. Glassware, reagents, supplies, equipment and settings other than those
listed in this manual may be employed, provided that method performance appropriate for the
intended RCRA application has been documented. Such performance includes consideration of
precision, accuracy {or bias), recovery, representativeness, comparability, and sensitivity (detection,
quantitation, or reporting limits) relative to the data quality objectives for the intended use of the
analytical results. In response to this inherent flexibility, if an alternative analytical procedure is
employed, then EPA expects the laboratory to demonstrate and document that the procedure is
capable of providing appropriate performance for its intended application. This demonstration must
not be performed after the fact, but as part of the laboratory's initial demonstration of proficiency with
the method. The documentation should be in writing, maintained in the laboratory, and available for
inspection upon request by authorized representatives of the appropriate regulatory authorities. The
documentation should include the performance data as well as a detailed description of the
procedural steps as performed (i.e., a written standard operating procedure).
Given this allowance for flexibility, EPA wishes to emphasize that this manual also contains
procedures for "method-defined parameters," where the analytical result is wholly dependant on the
process used to make the measurement. Examples include the use of the toxicity characteristic
leaching procedure (TCLP) to prepare a leachate, and the flash point, pH, paint filter liquids, and
conrosivity tests. In these instances, changes to the specific methods may change the end result
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and incorrectly identify a waste as nonhazardous. Therefore, when the measurement of such
method-defined parameters is required by regulation, those methods are no| subject to the flexibility
afforded in other methods.
Analysts and data users are advised that even for those analytes that are not method-
defined, different procedures may produce some difference in results. Common examples include
the differences in recoveries of phenolic compounds extracted from water by separatory funnel
(Method 3510) and continuous liquid-liquid (Method 3520) extraction techniques, differences in
recoveries of many compounds between Soxhlet (Method 3540) and ultrasonic (Method 3550)
extraction techniques, and differences resulting from the choice of acid digestion of metals (Method
3050) or microwave digestion (Method 3051). Where practical, the Agency has included guidance
in the individual methods regarding known potential problems, and analysts are advised to review
this information carefully in choosing or modifying analytical procedures. Chapter One describes a
variety of QC procedures that may be used to evaluate the quality of the analytical results. Additional
QC procedures may be described in the individual methods. The results of these QC procedures
should be used by the analyst to evaluate if the choice of the analytical procedures and/or any
modifications are appropriate to generate data of the quality necessary to satisfy the data quality
needs of the intended application.
The performance data included in the SW-846 methods are not intended to be used as
absolute QC acceptance criteria for method performance. The data are intended to be guidance,
by providing typical method performance in typical matrices, to assist the analyst in selection of the
appropriate method for the intended application. In addition, it is the responsibility of the laboratory
to establish actual operating parameters and in-house QC acceptance criteria, based on its own
laboratory SOPs and in-house QC program, to demonstrate appropriate performance of the methods
used in that laboratory for the RCRA analytical applications for which they are intended.
The regulated community is further advised that the methods here or from other sources
need only be used for those specific analytes of concern that are subject to regulation or other
monitoring requirements. The fact that a method provides a long list of analytes does not mean that
each of those analytes is subject to any or all regulations, or that all of those analytes must be
analyzed each time the method is employed, or that all of the analytes can be analyzed using a
single sample preparation procedure. It is EPA's intention that the target analyte list for any
procedure includes those analytes necessary to meet the data quality objectives of the project, i.e.,
those analytes subject to monitoring requirements and set out in a RCRA permit (or other applicable
regulation), plus those analytes used in the methods for QC purposes, such as surrogates, internal
standards, system performance check compounds, etc. Additional analytes, not included on the
analyte list of a particular method(s) but needed for a specific project, may be analyzed by that
particular method(s), if appropriate performance can be demonstrated for the analytes of concern
in the matrices of concern at the levels of concern.
2.1.1 Trace Analysis vs. Macroanalvsis
Through the choice of sample size and concentration procedures, the methods presented
in SW-846 were designed to address the problem of "trace" analyses (<1000 ppm), and have been
developed for an optimized working range. These methods are also applicable to "minor" (1000 ppm
- 10,000 ppm) and "major" (>10,000 ppm) analyses, as well, through use of appropriate sample
preparation techniques that result in analyte concentrations within that optimized range. Such
sample preparation techniques include:
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1) adjustment of size of sample prepared for analysis (for homogeneous samples),
2) adjustment of injection volumes,
3) dilution or concentration of sample,
4) elimination of concentration steps prescribed for "trace" analyses, and
5) direct injection (of samples to be analyzed for volatile constituents).
The performance data presented in each of these methods were generated from "trace"
analyses, and may not be applicable to "minor" and "major" analyses. Generally, extraction
efficiency improves as concentration increases,
CAUTION: Great care should be taken when performing trace analyses after the analysis of
concentrated samples, given the possibility of contamination.
2.1.2 Choice of Apparatus and Preparation of Reagents
Since many types and sizes of glassware and supplies are commercially available, and since
it is possible to prepare reagents and standards in many different ways, the apparatus, reagents, and
volumes specified in these methods may be replaced by any similar types as long as this substitution
does not affect the overall quality of the analyses.
2.1.3 Quality Control Criteria Precedence
Chapter One contains general quality control (QC) guidance for analyses using SW-846
methods. QC guidance specific to a given analytical technique (e.g., extraction, cleanup, sample
introduction, or analysis) may be found in Methods 3500, 3600, 5000, 7000, and 8000. Method-
specific QC criteria may be found in Sec. 8.0 of each individual method (or in Sec. 11.0 of air
sampling methods). When inconsistencies exist between the information in these locations, method-
specific QC criteria take precedence over both technique-specific criteria and those criteria given in
Chapter One, and technique-specific QC criteria take precedence over the criteria in Chapter One.
2.2 REQUIRED INFORMATION
In order to choose the correct combination of methods to comprise the appropriate analytical
procedure, some basic information is required.
2.2.1 Physical Stated of Sample
The phase characteristics of the sample must be known. There are several general
categories of phases into which the sample may be categorized, including:
There may be a substantial degree of overlap between the phases listed above and it may
be useful to further divide these phases in certain instances. A multiphase sample may be a
Aqueous
Sludge
TCLP or EP Extract
Solid
Ground Water
Oil or other Organic Liquid
Stack Sampling (VOST) Condensate
Multiphase Sample
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combination of aqueous, organic liquid, sludge, and/or solid phases, and generally must undergo a
phase separation as the first step in the analytical procedure.
2.2.2 Analvtes
Analytes may be divided into various classes based on the determinative methods which are
used to identify and quantify them. The most basic differentiation is between organic (e.g., carbon-
containing) analytes and inorganic (e.g., metals and anions) analytes.
Table 2-1 is an alphabetical list of analytes cited within the SW-846 organic determinative
methods (excludes immunoassay and other screening methods). These analytes have been
evaluated by those methods. The methods may also be applicable to other analytes that are similar
to those listed. Tables 2-2 through 2-32 list the analytes for each organic determinative method.
Table 2-33 indicates which methods are applicable to inorganic analytes.
2.2.3 Detection Limits
Some regulations may require a specific sensitivity or detection limit for an analysis, as in the
determination of analytes for the Toxicity Characteristic (TC). Drinking water detection limits, for
those specific organic and metallic analytes covered by the National Primary Drinking Water
Regulations, are desired in the analysis of ground water.
2.2.4 Analytical Objective
Knowledge of the analytical objective will be useful in the choice of sample preparation
procedures and in the selection of a determinative method. This is especially true when the sample
has more than one phase. Knowledge of the analytical objective may not be possible or desirable
at all management levels, but that information should be transmitted to the analytical laboratory
management to ensure that the correct techniques are used during the analytical effort.
2.2.5 Detection and Monitoring
The strategy for detection of compounds in environmental or process samples may be
contrasted with the strategy for collecting monitoring data. Detection samples define initial
conditions. When there is little information available about the composition of the sample source,
e.g., a well or process stream, mass spectral identification of organic analytes leads to fewer false
positive results. Thus, the most practical form of detection for organic analytes is often mass
spectral identification. However, where the sensitivity requirements exceed those that can be
achieved using mass spectral method (e.g., GC/MS or HPLC/MS), it may be necessary to employ
a more sensitive detection method (e.g., electron capture). In these instances, the risk of false
positive results may be minimized by confirming the results through a second analysis with a
dissimilar detector or chromatographic column. Thus, the choice of technique for organic analytes
may be governed by the detection limit requirements and potential interferants.
Similarly, the choice of technique for metals is governed by the detection limit requirements
and potential interferants.
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In contrast, monitoring samples are analyzed to confirm existing and on-going conditions,
tracking the presence or absence of known constituents in an environmental or process matrix. In
well-defined matrices and under stable analytical conditions, less compound-specific detection
modes may be used, as the risk of false positive results is less.
2.2.6 Sample Containers. Preservations, and Holding Times
Appropriate sample containers, sample preservation techniques, and sample holding times
for aqueous matrices are listed in Table 2-34, near the end of this chapter. Similar information may
be found in Table 3-1 of Chapter Three (inorganic analytes) and Table 4-1, of Chapter Four (organic
analytes). Samples must be extracted and analyzed within the specified holding times for the results
to be considered reflective of total concentrations. Analytical data generated outside of the specified
holding times must be considered to be minimum values only. Such data may be used to
demonstrate that a waste is hazardous where it shows the concentration of a constituent to be
above the regulatory threshold but cannot be used to demonstrate that a waste is not hazardous.
2.3 IMPLEMENTING THE GUIDANCE
The choice of the appropriate sequence of methods depends on the information required and
on the experience of the analyst. Figure 2-1 summarizes the organic analysis options available.
Appropriate selection is confirmed by the quality control results. The use of the recommended
procedures, whether they are approved or mandatory, does not release the analyst from
demonstrating the correct execution of the method.
2.3.1 Extraction and Sample Preparation Procedures for Organic Analytes
Methods for preparing samples for organic analytes are shown in Table 2-35. Method 3500
and associated methods should be consulted for further details on preparing the sample for analysis.
2.3.1.1 Aoueous Samples
Methods 3510 and 3520 may be used for extraction of the semivolatile organic
compounds from aqueous samples. The choice of a preparative method depends on the
sample. Method 3510, a separatory funnel liquid-liquid extraction technique, is appropriate
for samples which will not form a persistent emulsion interface between the sample and the
extraction solvent. The formation of an emulsion that cannot be broken up by mechanical
techniques will prevent proper extraction of the sample. Method 3520, a continuous liquid-
liquid extraction technique, may be used for any aqueous sample and will minimize emulsion
formation.
Method 3535 is solid-phase extraction technique that has been tested for
organochlorine pesticides and phthalate esters and may be applicable to other semivolatile
and extractable compounds as well. The aqueous sample is passed through a solid sorbent
material which traps the analytes. They are then eluted from the solid-phase sorbent with
a small volume of organic solvent. This technique may be used to minimize the volumes of
organic solvents that are employed, but may not be appropriate for aqueous samples with
high suspended solids contents.
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2.3.1.1.1 Basic or Neutral Extraction of Semivolatile Analvtes
The solvent extract obtained by performing Method 3510, 3520, or 3535 at
a neutral or basic pH will contain the neutral organic compounds and the organic
bases of interest. Refer to Table 1 in the extraction methods <3510 and/or 3520) for
guidance on the requirements for pH adjustment prior to extraction and analysis.
2.3.1.1.2 Acidic Extraction of Phenols and Acid Analvtes
The solvent extract obtained by performing Method 3510, 3520, or 3535 at
a pH less than or equal to 2 will contain the phenols and acid extractable organics
of interest.
2.3.1.2 Solid Samples
Soxhlet extraction (Methods 3540 and 3541), ultrasonic extraction (Method 3550),
and accelerated solvent extraction (Method 3545) may be used with solid samples.
Consolidated samples should be ground finely enough to pass through a 1 mm sieve. In
limited applications, waste dilution (Methods 3580 and 3585) may be used if the entire
sample is soluble in the specified solvent.
Methods 3540, 3541, 3545, and 3550 are neutral-pH extraction techniques and
therefore, depending on the analysis requirements, acid-base partition cleanup (Method
3650) may be necessary. Method 3650 will only be needed if chromatographic interferences
are severe enough to prevent detection of the analytes of interest. This separation will be
most important if a GC method is chosen for analysis of the sample. If GC/MS is used, the
ion selectivity of the technique may compensate for chromatographic interferences.
There are two extraction procedures for solid samples that employ supercritical fluid
extraction (SFE). Method 3560 is a technique for the extraction of petroleum hydrocarbons
from various solid matrices using carbon dioxide at elevated temperature and pressure.
Method 3561 may be used to extract polynuclear aromatic hydrocarbons (PAHs) from solid
matrices using supercritical carbon dioxide.
2.3.1.3 Oils and Organic Liquids
Method 3580, waste dilution, may be used to prepare oils and organic liquid samples
for analysis of semivolatile and extractable organic analytes by GC or GC/MS. Method 3585
may be employed for the preparation of these matrices for volatiles analysis by GC or
GC/MS. To avoid overloading the analytical detection system, care must be exercised to
ensure that proper dilutions are made. Methods 3580 and 3585 give guidance on performing
waste dilutions.
To remove interferences for semivolatiles and extractables, Method 3611 (Alumina
cleanup) may be performed on an oil sample directly, without prior sample preparation.
Method 3650 is the only other preparative procedure for oils and other organic liquids.
This procedure is a back extraction into an aqueous phase. It is generally introduced as a
cleanup procedure for extracts rather than as a preparative procedure. Oils generally have
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a high concentration of semivolatile compounds and, therefore, preparation by Method 3650
should be done on a relatively small aliquot of the sample. Generally, extraction of 1 mL of
oil will be sufficient to obtain a saturated aqueous phase and avoid emulsions.
2,3.1.4 Sludoe Samples
Determining the appropriate methods for analysis of sludges is complicated because
of the lack of precise definitions of sludges with respect to the relative percent of liquid and
solid components. There is no set ratio of liquid to solid which enables the analyst to
determine which of the three extraction methods cited is the most appropriate. Sludges may
be classified into three categories: liquid sludges, solid sludges, and emulsions, but with
appreciable overlap.
If the sample is an organic sludge (solid material and organic liquid, as opposed to
an aqueous sludge), the sample should be handled as a multiphase sample.
2.3.1.4.1 Liquid Sludges
Use of Method 3510 or Method 3520 may be applicable to sludges that
behave like and have the consistency of aqueous liquids. Ultrasonic extraction
(Method 3550) and Soxhlet (Method 3540) procedures will, most likely, be ineffective
because of the overwhelming presence of the liquid aqueous phase.
2.3.1.4.2 Solid Sludges
Soxhlet extraction (Methods 3540 and 3541), accelerated solvent (Method
3545) extraction, and ultrasonic extraction (Method 3550) will be more effective when
applied to sludge samples that resemble solids. Samples may be dried or
centrifuged to form solid materials for subsequent determination of semivolatile
compounds.
Using Method 3650, Acid-Base Partition Cleanup, on the extract may be
necessary, depending on whether chromatographic interferences prevent
determination of the analytes of interest.
2.3.1.4.3 Emulsions
Attempts should be made to break up and separate the phases of an
emulsion. Several techniques are effective in breaking emulsions or separating the
phases of emulsions, including:
1. Freezing/thawing: Certain emulsions will separate if exposed to
temperatures below 0°C.
2. Salting out: Addition of a salt to make the aqueous phase of an emulsion too
polar to support a less polar phase promotes separation.
3. Cerrtrifugation: Centrifugal force may separate emulsion components by
density.
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4. Addition of water or ethanol: Emulsion polymers may be destabilized when
a preponderance of the aqueous phase is added.
5. Forced filtering through glass wool: Many emulsions can be broken by
forcing the emulsion through a pad of Pyrex glass wool in a drying column
using a slight amount of air pressure (using a rubber bulb usually provides
sufficient pressure).
If techniques for breaking emulsions fail, use Method 3520. If the emulsion
can be broken, the different phases (aqueous, solid, or organic liquid) may then be
analyzed separately.
2.3.1.5 Multiphase Samples
Choice of the procedure for separating multiphase samples is highly dependent on
the objective of the analysis. With a sample in which some of the phases tend to separate
rapidly, the percent weight or volume of each phase should be calculated and each phase
should be individually analyzed for the required analytes.
An alternate approach is to obtain a homogeneous sample and attempt a single
analysis on the combination of phases. This approach will give no information on the
abundance of the analytes in the individual phases other than what can be implied by
solubility.
A third alternative is to select phases of interest and to analyze only those selected
phases. This tactic must be consistent with the sampling/analysis objectives or it will yield
insufficient information for the time and resources expended. The phases selected should
be compared with Figure 2-1 and Table 2-35 for further guidance.
2.3.2 Cleanup Procedures
Each category in Table 2-36, Cleanup Methods for Organic Analyte Extracts, corresponds
to one of the possible determinative methods available in the manual. Cleanups employed are
determined by the analytes of interest within the extract. However, the necessity of performing
cleanup may also depend upon the matrix from which the extract was developed. Cleanup of a
sample may be done exactly as instructed in the cleanup method for some of the analytes. There
are some instances when cleanup using one of the methods may only proceed after the procedure
is modified to optimize recovery and separation. Several cleanup techniques may be possible for
each analyte category. The information provided is not meant to imply that any or all of these
methods must be used for the analysis to be acceptable. Extracts with components which interfere
with spectral or chromatographic determinations are expected to be subjected to cleanup
procedures.
The analyst's discretion must determine the necessity for cleanup procedures, as there are
no clear cut criteria for indicating their use. Method 3600 and associated methods should be
consulted for further details on extract cleanup.
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2.3.3 Determinative Procedures
The determinative methods for organic analytes have been divided into three categories, as
shown in Table 2-37: gas chromatography/mass spectrometry (GC/MS); specific detection methods,
i.e., gas chromatography (GC) with specific non-MS detectors; and high performance liquid
chromatography (HPLC). This division is intended to help an analyst choose which determinative
method will apply. Under each analyte column, SW-846 method numbers have been indicated, if
appropriate, for the determination of the analyte. A blank has been left if no chromatographic
determinative method is available.
Generally, the MS procedures are more specific but less sensitive than the appropriate gas
chromatographic/specific detection method.
Method 8000 gives a general description of the techniques of gas chromatography and high
performance liquid chromatography. Method 8000 should be consulted prior to application of any
of the gas chromatographic methods.
Method 8081 (organochlorine pesticides), Method 8082 (polychlorinated biphenyls), Method
8141 (organophosphorus pesticides), and Method 8151 (chlorinated herbicides), are preferred over
GC/MS because of the combination of selectivity and sensitivity of the flame photometric, nitrogen-
phosphorus, and electron capture detectors.
Method 8260 is a GC/MS method for volatile analytes, which employs a capillary column.
A variety of sample introduction techniques may be used with Method 8260, including Methods 5021,
5030, 5031,5035, and 3585. A GC with a selective detector is also useful for the determination of
volatile organic compounds in a monitoring scenario, as described in Sec. 2.2.5.
Method 8270 is a GC/MS method for semivoiatile analytes, which employs a capillary column.
Table 2-37 lists several GC and HPLC methods that apply to only a small number of analytes.
Methods 8031 and 8033 are GC methods for acrolein, acrylonitrile, and acetonitrile. Methods 8315
and 8316 are HPLC methods for these three analytes. Method 8316 also addresses acrylamide,
which may be analyzed by Method 8032.
HPLC methods have been developed for other types of analytes, most notably carbamates
(Method 8318); azo dyes, phenoxy acid herbicides, carbamates, and organophosphorus pesticides
(Method 8321); PAHs (Method 8310); explosives (Methods 8330, 8331, and 8332); and some volatile
organics (Methods 8315 and 8316).
Method 8430 utilizes a Fourier Transform Infrared Spectrometer (FT-IR) coupled to a gas
chromatograph to determine bis(2-chloroethyl) ether and its hydrolysis products. The sample is
introduced by direct aqueous injection. Method 8440 may be employed for the determination of total
recoverable petroleum hydrocarbons (TRPH) in solid samples by infrared (IR) spectrophotometry.
The samples may be extracted with supercritical carbon dioxide, using Method 3560.
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2.4 CHARACTERISTICS
Figure 2-2 outlines a sequence for determining if a waste exhibits one or more of the
characteristics of a hazardous waste.
2,4.1 EP and TCLP extracts
The leachate obtained from using either the EP (Figure 2-3A) or the TCLP (Figure 2-3B) is
an aqueous sample, and therefore, requires further solvent extraction prior to the analysis of
semivolatile compounds.
The TCLP leachate is solvent extracted with methylene chloride at a pH > 11 and at a pH <2
by either Method 3510 or 3520. Method 3510 should be used unless the formation of emulsions
between the sample and the solvent prevent proper extraction, if this problem is encountered,
Method 3520 should be employed.
The solvent extract obtained by performing either Method 3510 or 3520 at a basic or neutral
pH will contain the base/neutral compounds of interest. Refer to the specific determinative method
for guidance on the pH requirements for extraction prior to analysis. Method 5031 (Azeotropic
Distillation) may be used as an effective preparative method for pyridine.
Due to the high concentration of acetate in the TCLP extract, it is recommended that purge-
and-trap be used to introduce the volatile sample into the gas chromatograph.
2.5 GROUND WATER
Appropriate analysis schemes for the determination of analytes in ground water are
presented in Figures 2-4A, 2-4B, and 2-4C. Quantitation limits for the inorganic analytes should
correspond to the drinking water limits which are available.
2.5.1 Special Techniques for Inorganic Analvtes
All atomic absorption analyses should employ appropriate background correction systems
whenever spectral interferences could be present. Several background correction techniques are
employed in modem atomic absorption spectrometers. Matrix modification can complement
background correction in some cases. Since no approach to interference correction is completely
effective in all cases, the analyst should attempt to verily the adequacy of correction. If the
interferant is known (e.g., high concentrations of iron in the determination of selenium), accurate
analyses of synthetic solutions of the interferant (with and without analyte) could establish the
efficacy of the background correction. If the nature of the interferant is not established, good
agreement of analytical results using two substantially different wavelengths could substantiate the
adequacy of the background correction.
To reduce matrix interferences, all graphite furnace atomic absorption (GFAA) analyses
should be performed using techniques which maximize an isothermal environment within the furnace
cell. Data indicate that two such techniques, L'vov platform and the Delayed Atomization Cuvette
(DAC), are equivalent in this respect, and produce high quality results.
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All furnace atomic absorption analysis should be carried out using the best matrix modifier
for the analysis. Some examples of modifiers are listed below. (See also the appropriate methods.)
Elementfs)
Modifier(s)
Pb
Cd
Sb
TI
As and Se
Nickel nitrate, palladium
Phosphoric acid, ammonium phosphate, palladium
Ammonium phosphate, palladium
Ammonium nitrate, palladium
Platinum, palladium
ICP. AA, and GFAA calibration standards must match the acid composition and strength of
the adds contained in the samples. Acid strengths of the calibration standards should be stated in
the raw data. When using a method which permits the use of internal standardization, and the
internal standardization option is being used, matrix matching is not required.
2.6 ADDITIONAL GUIDANCE REGARDING INORGANIC ANALYSES
Methods for preparing different sample matrices for inorganic analytes are shown in Table
2-38. Guidance regarding the use of leaching and digestive methods for inorganic analysis is
provided in Table 2-39.
2.7 REFERENCES
1. Barcelona, M.J. 'TOC Determinations in Ground Water3'; Ground Water 1984, 22m. 18-24.
2. Riggin, R.; et al. Development and Evaluation of Methods for Total Organic Halide and
Purgeable Organic Halide in Wastewater: U.S. Environmental Protection Agency. Office of
Research and Development. Environmental Monitoring and Support Laboratory, ORD
Publication Offices of Center for Environmental Research Information: Cincinnati, OH, 1984;
EPA-600/4-84-008.
3. McKee, G.; et al. Determination of Inorganic Anions in Water bv Ion Chromatography:
(Technical addition to Methods for Chemical Analysis of Water and Wastewater, EPA 600/4-
79-020), U.S. Environmental Protection Agency. Environmental Monitoring and Support
Laboratory. ORD Publication Offices of Center for Environmental Research Information:
Cincinnati, OH, 1984; EPA-600/4-84-017.
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TABLE 2-1
DETERMINATIVE METHODS FOR ORGANIC ANALYTES
Analyte Applicable Method(s)
Acenaphthene 8100, 8270, 8275, 8310, 8410
Acenaphthytene 8100, 8270, 8275, 8310, 8410
Acetaldehyde 8315
Acetone 8015, 8260, 8315
Acetonitrile 8015, 8033, 8260
Acetophenone 8270
2-Acetylaminofluorene 8270
1-Acetyl-2-thiourea 8270
Acifluorfen 8151
Acrolein (Propenal) 8015, 8260, 8315, 8316
Acrylamide 8032, 8316
Acrylonitrile 8015,8031,8260,8316
Alachlor 8081
Aldicarb (Temik) 8318, 8321
Aldicarb sulfone 8318, 8321
Aldicarb sulfoxide 8321
Aldrin 8081, 8270
Ally! alcohol 8015, 8260
Ally! chloride 8021, 8260
2-Aminoanthraquinone . 8270
Aminoazobenzene 8270
4-Aminobiphenyi 8270
Aminocarb , 8321
2-Amino-4,6-dinitrotoluene (2-Am-DNT)
4-Amino-2,6-dinitrotoluene (4-Am-DNT)
3-Amino-9-ethylcarbazole
Anilazine
Aniline 8131
o-Anisidine
Anthracene 8100, 8270, 8275, 8310
Aramite
Aroclor-1016 (PCB-1016) 8082
Aroclor-1221 (PCB-1221) 8082
Aroclor-1232 (PCB-1232) 8082
Aroclor-1242 (PCB-1242) 8082
Aroclor-1248 (PCB-1248) 8082
Aroclor-1254 (PCB-1254) 8082
Aroclor-1260 (PCB-1260) 8082
Aspon
Asuiam
Atrazine
Azinphos-ethyl
Azinphos-methyl 8141
Barban 8270
Baygon (Propoxur) 8318
Bendiocarb
8330
8330
8270
8270
8270
8270
8410
8270
8270
8270
8270
8270
8270
8270
8270
8141
8321
8141
8141
8270
8321
8321
8321
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TABLE 2-1. (Continued)
Anafyte Applicable Method(s)
Benefin 8091
Benomyl 8321
Bentazon , 8151
Benzal chloride 8121
Benzaldehyde 8315
Benz(a)anthracene 8100,8270,8275,8310,8410
Benzene I 8021, 8260
Benzenethiol (Thiophenol) 8270
Benzidine 8270, 8325
Benzo(b)fluoranthene 8100,8270,8275,8310
BenzoQ)fluoranthene 8100
Benzo(k)fIuoranthene 8100, 8270, 8275, 8310
Benzoic acid 8270, 8410
Benzo(g,h,i)perylene 8100, 8270, 8275, 8310
Benzo(a)pyrene 8100, 8270, 8275, 8310, 8410
p-Benzoquinone 8270
Benzotrichloride 8121
Benzoylprop ethyl 8325
Benzyl alcohol 8270
Benzyl chloride 8021, 8121, 8260
a-BHC (a-Hexachlorocyclohexane) 8081,8121,8270
P-BHC (P-Hexachlorocyclohexane) ... 8081, 8121, 8270
5-BHC (6-Hexachlorocyclohexane) 8081, 8121, 8270
y-BHC (Lindane, Y-Hexachlorocyclohexane) 8081, 8121, 8270
Bis(2-chloroethoxy)methane . . 8111, 8270, 8410
Bis(2-chloroethyl) ether 8111, 8270, 8410, 8430
Bis(2-chloroethyl)sulfide 8260
Bis(2-chloroisopropyl) ether 8021, 8111, 8270, 8410
Bis(2-n-butoxyethyl) phthalate 8061
Bis(2-ethoxyethyt) phthalate 8061
Bis(2-ethyihexyl) phthalate 8061, 8270, 8410
Bis(2-methoxyethyI) phthalate 8061
Bis(4-methyl-2-pentyl)-phthalate 8061
Bolstar (Sulprofos) 8141
Bromacil : 8321
Bromoacetone 8021, 8260
4-Bromoaniline 8131
Bromobenzene 8021, 8260
Bromochloromethane 8021, 8260
2-Bromo-6-chloro-4-nitroaniline 8131
Bromodichloromethane 8021, 8260
2-Bromo-4,6-dinitroaniline 8131
Bromoform 8021, 8260
Bromomethane 8021, 8260
4-Bromophenyl phenyl ether 8111,8270,8275,8410
Bromoxynil 8270
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TABLE 2-1. (Continued)
Analyte Applicable Method(s)
Butana!
1-Butanol (n-Butyl alcohol)
n-Butanol
2-Butanone (Methyl ethyl ketone, MEK)
Butralin .......
n-Butyl alcohol (1-Butanol)
8315
8015
8260
8015, 8260
.... 8091
..8015
t-Buty! alcohol 8015
n-Butylbenzene 8021, 8260
sec-Butylbenzene 8021,8260
tert-Butylbenzene 8021,8260
Butyl benzyl phthalate ".... 8061, 8270, 8410
2-sec-Buty1-4,6-dinitrophenol (DNBP, Dinoseb) 8041, 8151, 8270, 8321
Caffeine 8321
Captafol 8081, 8270
Captan ..... 8270
Carbaryl (Sevin) 8270, 8318, 8321, 8325
Carbendazim 8321
Carbofuran (Furaden) 8270, 8318, 8321
Carbon disulfide 8260
Carbon tetrachloride 8021,8260
Carbophenothion 8141, 8270
Chloral hydrate 8260
Chloramben 8151
Chlordane (NOS) 8270
a-Chlordane 8081
Y-Chlordane 8081
Chlorfenvinphos ............ 8141, 8270
Chloroacetonitrile 8260
2-Chloroaniline 8131
3-Chloroaniline 8131
4-Chloroaniline 8131, 8270, 8410
Chlorobenzene 8021, 8260
Chlorobenzilate 8081, 8270
2-Chlorobiphenyl 8082, 8275
2-Chloro-1,3-butadiene (Chloroprene) 8021,8260
1-Chlorobutan e 8260
Chlorodibromomethane (Dibromochloromethane) 8021, 8260
2-Chloro-4,6-dinitroanllin e 8131
1-Chloro-2,4-dinitrobenzene 8091
1-Chloro-3,4-di nitrobenzene 8091
Chloroethane 8021, 8260
2-Chloroethano l 8021, 8260, 8430
2-(2-Ch!oroethoxy)ethanol 8430
2-Chloroethyl vinyl ether 8021, 8260
Chloroform 8021, 8260
1-ChIorohexane 8260
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TABLE 2-1. (Continued)
Analyte Applicable Method(s)
Chloromethane 8021, 8260
5-Chloro-2-methylaniline 8270
Chtoromethyi methyl ether 8021
2-Chloro-5-methylpheno l 8041
4-Chloro-2-methyIphenol 8041
4-Chloro-3-methyIphenoI 8041, 8270, 8410
3-(Chloromethyl)pyridine hydrochloride . 8270
1 -Chloro naphthalene 8270, 8275
2-Chlononaphthalene 8121, 8270, 8410
Chloroneb 808
2-Chloro-4-nitroaniline . 813
4-Chloro-2-nitroaniline 813
1 -Chloro-2-nitroberizene 809
1 -Chloro-4-nitrobenzene 809
2-Chloro-6-nitrotoluene 809
4-Chloro-2-nitrotoluene 809
4-Chloro-3-nitrotoluene 809
2-Chlorophenol 8041,8270,8410
3-Chlorophenol 8041
4-Chlorophenol 8041, 8410
4-Chloro-1,2-phenylenediam ine 8270
4-Chloro-1,3-phenylenediamine 8270
4-Chiorophenyl phenyl ether 8111, 8270, 8410
2-Chlorophenyl 4-nitrophenyl ether 8111
3-Chlorophenyl 4-nitrophenyl ether 8111
4-Chlorophenyl 4-nitrophenyl ether 8111
o-Chlorophenyl thiourea 8325
Chloroprene (2-Chloro-1,3-butadiene) 8021, 8260
3-ChIoropropionitrile 8260
Chloropropham 8321
Chloropropylate 8081
Chlorothalonil 8081
2-Chlorotoluene 8021, 8260
4-Chlorotoluene 8021, 8260
Chloroxuron 8321
Chlorpyrifos 8141
Chlorpyrifos methyl 8141
Chrysene 8100, 8270, 8275, 8310, 8410
Coumaphos 8141, 8270
Coumarin Dyes 8321
p-Cresidine 8270
o-Cresol (2-MethyIphenoI) 8041, 8270, 8410
m-Cresol (3-Methylphenol) 8041, 8270
p-Cresol (4-Methyiphenol) 8041, 8270, 8275, 8410
Crotonaldehyde 8015, 8260, 8315
Crotoxyphos 8141, 8270
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TABLE 2-1. (Continued)
Analyte Applicable Method(s)
Cyelohexanone 8315
2-CyclohexyW,6-dinitrophenol 8041, 82T0
2,4-D 8151,8321
Daiapon 8151, 8321
2,4-DB 8151, 8321
DBCP (1,2-Dibromo-3-chloropropane) 8011, 8021, 8081, 8260, 8270
2,4-D, butoxyethanol ester ! 8321
DCM (Dichloromethane, Methylene chloride) 8021,8260
DCPA 8081
DCPA diacid 8151
4,4-DDD :. 8081, 8270
4,4'-DDE 8081, 8270
4,4-DDT 8081, 8270
DDVP (Dichlorvos, Dichlorovos) 8141, 8270, 8321
2,2',3,3,4,4'5,5,,6,6'-Decachlorobiphenyl 8275
Decanal 8315
Demeton-O, and Demeton-S 8141, 8270
2,4-D, ethylhexyi ester 8321
Diailate 8081, 8270
Diamyt phthalate 8061
2,4-Diamlnotoluene 8270
Diazinon 8141
Dibenz(a,h)acridine 8100
Dibenz(a,j)acridine 8100, 8270
Dibenz(a, h)anthracene 8100,8270,8275,8310
7H-Dibenzo(c,g) ca rbazole 8100
Dibenzofuran 8270, 8275, 8410
Dibenzo(a,e)pyrene 8100, 8270
Dibenzo(a,h)pyrene 8100
Dibenzo(a,i)pynene 8100
Dibenzothiophene 8275
Dibromochloromethane (Chlorodibromomethane) 8021, 8260
1,2-Dibromo-3-chloroprapane (DBCP) 8011, 8260, 8270
1,2-Dibromoethane (EDB, Ethylene dibromide) 8011, 8021, 8260
Dibromofluoromethane 8260
Dibromomethane 8021, 8260
2,6-Dibromo-4-nitroaniline 8131
2,4-Dibromophenyl 4-nitrophenyl ether 8111
Di-n-butyl phthalate 8061,8270,8410
Dicamba 8151, 8321
Dichlone 8081, 8270
3,4-Dichloroaniline 8131
1.2-Dichlorobenzen e 8021, 8121, 8260, 8270, 8410
1.3-Dichlorobenzen e 8021, 8121, 8260, 8270, 8410
1.4-Dichlorobenzen e 8021,8121,8260,8270,8410
3,3'-Dichlorobenzidine 8270,8325
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TABLE 2-1. (Continued)
Analyte Applicable Method(s)
3.5-Dichlorobenzoic acid 8151
2,3-Dichlorobiphenyl 8082, 8275
3,3'-Dichlorobiphenyl ... 8275
cis-1,4-Dichloro-2-butene 8260
trans-1,4-Dichloro-2-butene .. 8260
Dichlorodifluoromethane 8021, 8260
1.1-Dichloroethane 8021, 8260
1.2-Dichloroethane 8021, 8260
1.1-Dichloroethene (Vinylidene chloride) 8021,8260
cis-1,2-Dichloroethene 8021, 8260
trans-1,2-Dichloroethene 8021, 8260
Dichlorofenthion 8141
Dichioromethane (DCM, Methylene chloride) 8021, 8260
2.6-Dich!oro-4-nitroaniline 8131
2.3-Dichloronitrobenzene 8091
2.4-Dichloronitrobenzen e 8091
3.5-Dichloronitrobenzen e 8091
3.4-Dichloronitrobenzen e 8091
2.5-Dichloronitrobenzen e 8091
2.3-Dichlorophenol 8041
2.4-Dichlorophenol . 8041, 8270, 8410
2.5-Dichlorophenol 8041
2.6-Dichlorophenol 8041, 8270
3.4-Dichiorophenol 8041
3.5-Dichloropheno! 8041
2.4-Dichlorophenol 3-methyI-4~nitrophenyl ether 8111
2.6-Dichtorophenyl 4-nitrophenyl ether 8111
3.5-Dichloropherryl 4-nitrophenyl ether 8111
2,5-Dichlorophenyl 4-nitrophenyl ether 8111
2,4-Dichlorophenyl 4-nitrophenyl ether 8111
2.3-Dichlorophenyl 4-nitrophenyl ether 8111
3.4-Dichlorophenyl 4-nitrophenyl ether 8111
Dichloroprop (Dichlorprop) 8151, 8321
1.2-Dichloropropane 8021, 8260
1.3-Dichloropropane 8021,8260
2.2-Dichloropropane 8021,8260
1.3-Dichloro-2-propano l 8021, 8260
1,1-Dichloroprop ene 8021, 8260
cis-1,3-Dichloropropene : 8021, 8260
trans-1,3-Dichloropropene 8021, 8260
Dichlorovos (DDVP, Dichlorvos) 8141, 8270, 8321
Dichlorprop (Dichloroprop) 8151, 8321
Dichlorvos (DDVP, Dichlorovos) 8141, 8270, 8321
Dicrotophos 8141, 8270
Dicofol ..... 8081
Dicyclohexyl phthalate 8061
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TABLE 2-1. (Continued)
Analyte Applicable Method(s)
Dieldrin 8081, 8270
1,2,3,4-Diepoxybutane 8260
Diesel range organics (DRO) 8015, 8440
Diethylene glycol 8430
Diethyl ether 8015, 8260
Diethyl phthalate 8061, 8270, 8410
Diethyl stilbestrol 8270
Diethyl sulfate 8270
Dihexyl phthalate 8061
Diisobuty! phthalate 8061
Dimethoate 8141, 8270, 8321
3,3'-Dimethoxybenzidine 8270, 8325
Dimethylaminoazobenzene 8270
2,5-Dimethylbenzaldehyde 8315
7,12-Dimethylbenz(a)anthracene 8270
3,3-Dimethylbenzidine 8270, 8325
a,a-Dimethylphenethylamine 8270
2.3-Dimethylpheno I 8041
2.4-Dimethylpheno I 8041, 8270
2.5-Dimethylpheno l 8041
2.6-Dimethyl phenol 8041
3,4-Dimethylphenol 8041
Dimethyl phthalate 8061, 8270, 8410
Dinitramine 8091
2,4-DinitroanIiine 8131
1.2-Dinitrobenzene 8091, 8270
1.3-Dinitrobenzene (1,3-DNB) , 8091, 8270, 8330
1.4-Dinitrobenzene 8091, 8270
4,6-Dinitro-2-methylphenol 8270, 8410
2.4-Dinitropheno l 8041, 8270, 8410
2.5-Dinitrophenol 8041
2,4-Dinitrotoluene (2,4-DNT) 8091, 8270, 8330, 8410
2.6-Dinitrotoluene (2,6-DNT) 8091, 8270, 8330, 8410
Dinocap 8270
Dinonyl phthalate " 8061
Dinoseb (2-sec-Butyl-4,6-dinitrophenol, DNBP) 8041, 8151, 8270, 8321
Di-n-octyl phthalate 8061, 8270, 8410
Dioxacarb 8318
1.4-Dioxane 8015, 8260
Dioxathion 8141
Di-n-propyl phthalate 8410
Diphenylamine 8270
5.5-Diphenylhydantoi n 8270
1,2-Diphenylhydrazine 8270
Disperse Blue 3 8321
Disperse Blue 14 8321
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TABLE 2-1. (Continued)
Analyte Applicable Method(s)
Disperse Brown 1 ,. 8321
Disperse Orange 3 8321
Disperse Orange 30 8321
Disperse Red 1 8321
Disperse Red 5 8321
Disperse Red 13 8321
Disperse Red 60 8321
Disperse Yellow 5 ... 8321
Disuffoton 8141, 8270, 8321
Diuron 8321, 8325
1.3-DNB (1,3-Dinitrobenzene) 8091, 8270, 8330 '
DNBP (2-sec-Sutyl-4,6-dinitrophenol, Dinoseb) 8151, 8270, 8321
2.4-DNT (2,4-Dinitrotoluene) 8091, 8270, 8275, 8330, 8410
2,6-DNT (2,6-Dinitrotoluene) 8091, 8270, 8330, 8410
EDB (1,2-Dibromoethane, Ethylene dibromide) 8011, 8021, 8260
Endosulfan I 8081, 8270
Endosulfan II 8081, 8270
Endosulfan sulfate 8081, 8270
Endrin 8081, 8270
Endrin aldehyde 8081, 8270
Endrin ketone 8081, 8270
Epichlorohydrin 8021, 8260
EPN 8141,8270
Ethanol 8015, 8260
Ethion 8141, 8270
Ethoprop 8141
Ethyl acetate 8015, 8260
Ethylbenzene 8021, 8260
Ethyl carbamate 8270
Ethyl cyanide (Propionitrile) 8015, 8260
Ethylene dibromide (EDB, 1,2-Dibromoethane) 8011, 8021, 8260
Ethylene glycol 8015,8430
Ethylene oxide 8015, 8260
Ethyl methacrylate 8260
Ethyl methanesulfonate ..... 8270
Etridiazole 8081
Famphur 8141, 8270, 8321
Fenitrothion 8141
Fensulfothion 8141, 8270, 8321
Fenthion 8141, 8270
Fenuron 8321
Fluchloralin 8270
Fluometuron 8321
Fiuoranthene 8100, 8270, 8275, 8310, 8410
Fluorene . 8100, 8270, 8275, 8310, 8410
Fluorescent Brig htener 61 8321
TWO-19 Revision 4
January 1998
-------
TABLE 2-1, (Continued)
Analyte Applicable Method(s)
Fluorescent Brightener 236 8321
Fonophos 8141
Formaldehyde 8315
Furaden (Carbofuran) 8270,8318,8321
Gasoline range organics (GRO) 8015
Halowax-1000 8081
Halowax-1001 8081
Halowax-1013 8081
Halowax-1014 8081
Halowax-1051 8081
Halowax-1099 " 8081
Heptachfor 8081, 8270
2,2',3,3',4,4',5-Heptachlorobiphenyl 8082, 8275
2,2't3,4,4,,5,5'-Heptachlorobiphenyl 8082, 8275
2,2',3,4,4',5',6-Heptachlorobiphenyf 8082
2,2',3,4',5,5',6-Heptachlorobiphenyl 8082, 8275
Heptachlor epoxide 8081, 8270
Heptanal 8315
Hexachlorobenzene 8081, 8121, 8270, 8275, 8410
2,2',3,3,4,4'-Hexachlorobiphenyl 8275
2,2\3,4,4\5,-Hexachlorobipheny[ 8082, 8275
2,2',3,4,5,5,-Hexachlorobiphenyl 8082
2,2',3,5,5',6-Hexachlorobiphenyl 8082
2,2',4,4',5,5'-Hexachlorobiphenyl 8082
Hexachlorobutadiene 8021, 8121, 8260, 8270, 8410
a-Hexachlorocyclohexane (a-BHC) 8081,8121,8270
P-Hexachlorocyclohexane (p-BHC) 8081, 8121, 8270
5-Hexachlorocyclohexane (5-BHC) 8081,8121,8270
y-Hexachlorocyclohexane (y-BHC, Lindane) 8081,8121,8270
Hexachlorocyclopentadiene 8081, 8121, 8270, 8410
Hexachioroethane 8121, 8260, 8270, 8410
Hexachlorophene 8270
Hexachioropropene 8270
Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) 8330
Hexameihylphosphoramide (HMPA) 8141, 8270
Hexanal 8315
2-Hexanone 8260
Hexyl 2-ethyIhexyl phthalate 8061
HMPA (Hexamethy! phosphoramide) 8141, 8270
HMX (Octahydro-1,3,5,7-tetranitro-1,3,57-tetrazocine) 8330
1,2,3,4,6,7,8-HpCDD 8280, 8290
HpCDD, total 8280, 8290
1.2.3.4.6.7.8-HpCD F 8280,8290
1.2.3.4.7.8.9-HpCD F 8280,8290
HpCDF, total . 8280, 8290
1,2,3,4,7,8-HxCDD ..... 8280, 8290
TWO-20
Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte Applicable Method(s)
1.2.3.6.7.8-HxCDD . 8280, 8290
1.2.3.7.8.9-HxCDD 8280, 8290
HxCDD, total 8280, 8290
1,2,3,4,7,8-HxCDF 8280, 8290
1.2.3.6.7.8-HxCDF 8280, 8290
1.2.3.7.8.9-HxCD F 8280, 8290
2,3,4,6,7,8-HxCDF : 8280, 8290
HxCDF 8280, 8290
Hydroquinone 8270
3-Hydroxycarbofuran 8318, 8321
5-Hydroxydicamba 8151
2-Hydroxypropionitiile 8260
lndeno(1,2,3-cd)pyrene 8100, 8270, 8275, 8310
lodomethane (Methyl iodide) 8260
Isobutyl alcohol (2-Methyl-1-propanol) 8015, 8260
Isodrin 8081, 8270
Isophorane 8270, 8410
Isopropalin 8091
Isopropyl alcohol (2-Propanol) . 8015, 8260
Isopropylbenzene 8021, 8260
p-lsopropyltoluene 8021, 8260
Isosafrole 8270
Isovaleraldehyde 8315
Kepone 8270
Lannate (Methomyl) 8318, 8321
Leptophos 8141, 8270
Lindane (y-Hexachlorocyclohexane, y-BHC) 8081,8121,8270
Linuron (Lorox) 8321,8325
Lorox (Linuron) 8321, 8325
Malathion 8141, 8270
Maleic anhydride 8270
Malononitrile 8260
MCPA 8151, 8321
MCPP 8151, 8321
Merphos 8141, 8321
Mestranol 8270
Mesurol (Methiocarb) 8318, 8321
Methacrylonitrile 8260
Methanol 8015, 8260
Methapyrilene 8270
Methiocarb (Mesurol) 8318, 8321
Methomyl (Lannate) 8318, 8321
Methoxychlor 8081, 8270
Methyl acrylate 8260
2-Methyl-1-propanol (Isobutyl alcohol) 8015, 8260
Methyl-t-butyl ether 8260
TWO-21
Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte Applicable Method(s)
3-Methylcholanthrene , 8100, 8270
2-Methyl-4,6-dinitrophenoI 8041
4,4'-Methylenebis(2-chloroaniline) 8270
4,4'-Methylenebis(N1N-dimethylanrline) 8270
Methyl ethyl ketone {MEK, 2-Butanone) 8015, 8260
Methylene chloride (Dichloromethane, DCM) 8021, 8260
Methyl iodide (lodomethane) : 8260
Methyl isobutyl ketone (MIBK, 4-Methyl-2-pentanone) 8015, 8260
Methyl methacrylate 8260
Methyl methanesuifonate 8270
2-Methylnaphthalene 8270, 8410
Methyl parathion (Parathion, methyl) 8270,8141, 8321
4-Methyl-2-perrtanone (MIBK, Methyl isobutyl ketone) 8015, 8260
2-Methylphenol (o-Cresol) 8041,8270,8410
3-Methylphenol (m-Cresol) 8041, 8270
4-Methylphenol (p-Cresol) 8041, 8270, 8410
2-Methylpyridine (2-Picoline) 8015,8260,8270
Methyl-2,4,6-trinitrophenylnitramine (Tetryl) 8330
Mevinphos 8141, 8270
Mexacarbate 8270, 8321
MIBK (Methyl isobutyl ketone, 4-MethyI-2-pentanone) 8015, 8260
Minex 8081, 8270
Monocrotophos 8141,8270,8321
Monuron 8321, 8325
Naled 8141, 8270, 8321
Naphthalene 8021, 8100, 8260, 8270, 8275, 8310, 8410
NB (Nitrobenzene) 8091, 8260, 8270, 8330, 8410
1,2-Naphthoquinone 8091
1,4-Naphthoquinone 8270, 8091
1-Naphthylamin e 8270
2-Naphthylamin e 8270
Neburon 8321
Nicotine 8270
5-Nitroacenaphthen e 8270
2-Nitroanilin e 8131, 8270, 8410
3-Nitroaniline 8131, 8270, 8410
4-Nitroanilin e 8131, 8270, 8410
5-Nitro-o-anisidin e 8270
Nitrobenzene (NB) 8091, 8260, 8270, 8330, 8410
4-Nitrobiphenyl 8270
Nitrofen 8081, 8270
Nitroglycerin 8332
2-Nitropheno l 8041, 8270, 8410
3-Nitropheno l 8041
4-Nitropheno l 8041, 8151, 8270, 8410
4-NitrophenyI phenyl ether 8111
TWO - 22 Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte Applicable Method(s)
2-Nitropropane 8260
NitroquinoIine-1-oxide 8270
N-Nitrosodi-n-butylamine 8015, 8260, 8270
N-Nitrosodiethylamine ..... ..... 8270
N-Nitrosodimethylamine 8070, 8270, 8410
N-Nitrosodi-n-butylamine (N-Nitrosodibutylamine) 8015, 8260, 8270
N-Nitrosodiphenylamine :.... 8070, 8270, 8410
N-Nitrosodi-rvpropylamine 8070, 8270, 8410
N-Nitrosomethylethylamine 8270
N-Nitrosomorpholine 8270
N-Nitrosopiperidine : 8270
N-Nitrosopyrrolidine 8270
2-Nitrotoluene (o-Nitrotoluene, 2-NT) 8091, 8330
3-NitrotoIuene (m-Nitrotoluene, 3-NT) 8091, 8330
4-Nitrotoluene (p-Nitrotoluene, 4-NT) 8091, 8330
o-Nitrotoluene (2-Nitrotoluene, 2-NT) 8091, 8330
m-Nitrotoluene (3-Nitrotoluene, 3-NT) 8091,8330
p-Nitrotoluene (4-Nitrotoluene, 4-NT) 8091,8330
5-Nitro-o-toluidine 8270
frans-Nonachlor 8081
2,2'3,3'4,4'5,5'6-NonachlorobiphenyI . 8082, 8275
Nonanal 8315
2-NT (2-Nitrotoluene, o-Nitrotoluene) 8091, 8330
3-NT (3-Nitrotoluene, m-Nitrotoluene) 8091, 8330
4-NT (4-Nitrotoluene, p-Nitrotoluene) 8091, 8330
OCDD 8280, 8290
OCDF 8280, 8290
2,2,,3,3',4,4,5,5'-Octachlorobiphenyl 8275
Octahydro-1,3,5,7-tetranitro-1t3,5,7-tetrazocjne (HMX) 8330
Octamethyl pyrophosphoramide 8270
Octanal 8315
Oxamyl 8321
4,4'-Oxydianiline 8270
Paraldehyde 8015, 8260
Parathion 8270
Parathion, ethyl 8141
PCB-1016 (Aroclor-1016) . 8082, 8270
PCB-1221 (Aroclor-1221) 8082, 8270
PCB-1232 (Aroclor-1232) 8082, 8270
PCB-1242 (Aroclor-1242) 8082, 8270
PCB-1248 (Aroclor-1248) 8082, 8270
PCB-1254 (Aroclor-1254) 8082, 8270
PCB-1260 (Aroclor-1260) . 8082, 8270
PCNB 8081
1,2,3,7,8-PeCDD 8280, 8290
PeCDD, total 8280, 8290
TWO-23 Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte
Applicable Method(s)
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCDF
PeCDF, total
Pendimethaline (Penoxalin) ...
Penoxalin (Pendimethaline) ...
Pentachlorobenzene
2I2',3,4,5'-Pentachlorobiphenyl
2,2,,4,5,5,-Pentachlorobiphenyl
2,3,3',4',6-PentachlorobiphenyI
2,3',4,4',5-Pentachlorobiphenyl
Pentachloroethane
Pentachloronitrobenzene
Pentachlorophenol
Pentafluorobenzene
Pentanal (Valeraldehyde)
2-Pentanone
Permethrin (c/s and trans)
Perthane
Phenacetin
Phenanthrene
Phenobarbital
Phenoi
1,4-Phenylenediamine
Phorate
Phosalone
Phosmet
Phosphamidon
Phthaiic anhydride
Picloram
2-Picoline (2-Methylpyridine) ..
Piperony! sulfoxide
Profluralin
Promecarb
Pronamide
Propachlor
Propanal (Propionaldehyde)...
1-Propano l
2-Propanol (Isopropyl alcohol)
Propargyl alcohol
Propenal (Acrolein) .........
Propham
B-Propiolactone
Propionaldehyde (Propanal)...
Propionitrile (Ethyl cyanide) ...
Propoxur(Baygon)
n-Propylamine
8041
8100,8270, 8275, 8310
8041, 8270
8280, 8290
8280, 8290
8280, 8290
8091
8091
8121,8270
8082
8082, 8275
8082
8275
8260
. 8091,8270
8151, 8270, 8410
8260
8315
8015,8260
8081
8081
8270
8410
8270
8410
8270
8321
8270
8270
8270
8270
8151
8270
8270
8091
8318
8270
8321
8321
8260
8260
8260
8315
8321
8260
8315
8260
8321
8260
8141, 8270
8141
8141
8015, 8260
8081
8315
8015
8015
8260
8015
8318
TWO-24
Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte
Applicable Method(s)
n-Propylbenzene
Propylthiouracil
Prothiophos (Tokuthion)
Pyrene
Pyridine
RDX (Hexahydro-1,3,5-trinitro-1,3,5-triazine)
Resorcinoi
Ronnel
Rotenone
Safrole
Sevln (Carbaryl)
Siduron
Simazine
Silvex (2,4,5-TP)
Solvent Red 3
Solvent Red 23
Stirophos (Tetrachlorvinphos)
Strobane
Strychnine
Styrene
Sulfallate
Sulfotepp
Sulprofos (Bolstar)
2,4,5-T
2,4,5-T, butoxyethanol ester
2,4,5-T, butyl ester
2,3,7,8-TCDD
TCDD, total
2,3,7,8-TCDF
TCDF, total
Tebuthiuron
Temik (Aldicarb)
Terbufos
1.2.3.4-Tetrachlorobenzen e
1.2.3.5-Tetrachlorobenzene
1,2,4,5-Tetrachlorobenzene
2,2,,3,5,-TetrachlorobiphenyI
2,2,,4,5,-Tetrachlorobiphenyl
2,2\5,5'-Tetrachlorobiphenyl
2,3'.4,4'-Tetrachlorobiphenyl
1,1,1,2-T etrachloroethane
1,1,2,2-Tetrachloroethane
Tetrachloroethene
2.3.4.5-Tetrachlorophenol
2.3.4.6-Tetrachloropheno l
2,3,5,6-T etrachlorophenol
.. 8021, 8260
........ 8270
8141
8100, 8270, 8275, 8310, 8410
"8015, 8260
8330
8270
8141
. .. .. 8325
8270
8270, 8318, 8321, 8325
8321, 8325
8141
8151, 8321
8321
8321
8141, 8270
8081
8270, 8321
8021, 8260
8270
8141
8141
8151, 8321
8321
8321
8280, 8290
8280, 8290
8280, 8290
8280, 8290
8321
8318, 8321
8141, 8270
8121
8121
8121, 8270
8082, 8275
8275
8082, 8275
8082, 8275
8021, 8260
8021, 8260
8021, 8260
8041
8041, 8270
8041
TWO-25
Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte Applicable Method(s)
2.3.4.5-Tetrachloronitrobenzene 8091
2.3.5.6-Tetrachloronitrobenzene . 8091
Tetrachlorvinphos (Stirophos) 8141, 8270
Tetraethyi dithiopyrophosphate 8270
Tetraethyl pyrophosphate (TEPP) ...... 8141, 8270
Tetrazene 8331
Tetryl (Methyl-2,4,6-trinitrophenylnitramine) 8330
8321
8270
8270
8330
8330
8141
8141
8315
8315
8315
8260
8270
8270
8270
8321
8131
8131
8260
8410
8275
8275
8275
8121
8260
8260
8260
8260
8321
8141
8091
8091
Thiofanox
Thionazin (Zinophos) 8141
Thiophenol (Benzenethiol)
1.3.5-TNB (1,3,5-Trinitrobenzene) : 8270
2.4.6-TNT (2,4,6-T rinitrobenzene)
TOCP (Tri-o-cresylphosphate) ..
Tokuthion (Prothiophos) ......
m-Tolualdehyde
o-Toiualdehyde
p-Tolualdehyde
Toluene 8021
Toluene diisocyanate
o-Toluidine 8015, 8260
Toxaphene 8081
2.4.5-TP (Silvex) ! 8151
2.4.6-Trichloroanilin e
2,4,5-T richloroaniline
1.2.3-Trichlorobenzene 8021, 8121
1.2.4-Trichtorobenzen e 8021,8121,8260,8270,8275
2,2', 5-Trichlorobiphenyl 8082
2,3',5-T richlorobiphenyl
2,4',5-T richlorobiphenyl 8082
1.3.5-T richtorobenzene
1.1.1-Trichloroethane 8021
1.1.2-Trichloroethane 8021
Trichloroethene 8021
Trichlorofluoromethane 8021
Trichlorfon 7 8141
Trichloronate
1.2.3-TrichIoro~4-nitrobenzen e
1.2.4-Trichloro-5-nitrobenzene
2.4.6-T richloronitrobenzene 8091
2.3.4-Trichloropheno l 8041
2.3.5-T richlorophenol 8041
2.3.6-Trichloropheno l 8041
2.4.5-Trichloropheno l 8041, 8270, 8410
2.4.6-T richlorophenol 8041, 8270, 8410
2,4,6-Trichlorophenyl 4-nitrophenyl ether 8111
2,3,6-Trichlorophenyl 4-nitrophenyl ether 8111
TWO - 26 Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte Applicable Method(s)
2,3,5-Trichtorophenyl 4-nitrophenyl ether 8111
2,4,5-Trichlorophenyl 4-nitrophenyt ether , 8111
3,4,5-Trichlorophenyl 4-nitrophenyl ether , 8111
2.3.4-Trichlorophenyl 4-nitrophenyl ether 8111
1.2.3-Trichloropropane 8021, 8260
O.O.O-Triethyl phosphorothioate 8270
Trifluralin 8091, 8081, 8270
2.4.5-Trimethylaniiine 8270
1.2.4-Trimethylbenzen e 8021, 8260
1.3.5-Trimethylbenzen e 8021,8260
Trimethylphosphate ..." 5270
1.3.5-Trinitrobenzene (1,3,5-TNB) 8270, 8330
2.4.6-Trinitrobenzene (2,4,6-TNT) 8330
Tris-BP (Tris-(2,3-dibromopropyl) phosphate) 8270, 8321
Tri-o-cresylphosphate (TOCP) 8141
Tri-p-toiyI phosphate 8270
Tris-(2,3-dibromopropyl) phosphate (Tris-BP) 8270, 8321
Valeraldehyde (Pentanal) 8315
Vinyl acetate 8260
Vinyl chloride 8021, 8260
Vinylidene chloride (1,1-Dichloroethene) .... 8021, 8260
o-Xylene 8021, 8260
m-Xylene 8021, 8260
p-Xylene 8021, 8260
Zinophos (Thionazin) .... 8141, 8270
TWO-27
Revision 4
January 1998
-------
TABLE 2-2
METHOD 8011 (MICROEXTRACTiON AND GAS CHROMATOGRAPHY)
1,2-Dibromo-3-chloropropane (DBCP)
1,2-DIbromoethane (EDB)
TABLE 2-3
METHOD 8015 (GC/FID) - NONHALOGENATED VOLATILES
Acetone
Acetonitrile
Acrolein
Acrylonitrile
Ally! alcohol
1-Butanol (n-Butyl alcohol)
t-Butyl alcohol
Crotonaldehyde
Diethyl ether
1,4-Dioxane
Ethanol
Ethyl acetate
Ethylene glycol
Ethylene oxide
Isobutyl alcohol
Isopropyl alcohol
Methanol
Methyl ethyl ketone (MEK)
Methyl isobutyl ketone (MIBK)
N-Nitroso-di-n-butylamine
Paraldehyde
2-Pentanone
2-Picoline
1-Propanol
Propionitrile
Pyridine
o-Toluidine
Gasoline range organics (GRO)
Diesel range organics (DRO)
TWO-28
Revision 4
January 1998
-------
TABLE 2-4
METHOD 8021 (GC, PHOTOIONIZATION AND ELECTROLYTIC
CONDUCTIVITY DETECTORS) - AROMATIC AND HALOGENATED VOLATILES
Allyl chloride
cis-1,2-Dichloroethene
Benzene
trans-1,2-Dichloroethene
Benzyl chloride
1,2-Dichloropropane
Bis(2-chloroisopropyl)
1,3-Dichloropropane
ether
2,2-Dichloropropane
Bromoacetone
1,3-DichIoro-2-propanol
Bromobenzene
1,1 -Dichloropropene
Bromochloromethane
cis-1,3-Dichloropropene
Bromodichloromethane
trans-1,3-Dichloropropene
Bromoform
Epichlorhydrin
Bromomethane
Ethylbenzene
n-Butylbenzene
Hexachlorobutadiene '
sec-Butylbenzene
Isopropylbenzene
tert-Butylbenzene
p-lsopropyltoluene
Cartoon tetrachloride
Methylene chloride
Chlorobenzene
Naphthalene
Chlorodibromo methane
n-Propylbenzene
Chloroethane
Styrene
2-Chloroethanol
1,1,1,2-T etrachloroethane
2-Chloroethyl vinyl ether
1,1,2,2-T etrachloroethane
Chloroform
Tetrachloroethene
Chloromethyl methyl ether
Toluene
Chloroprene
1,2,3-T richlorobenzene
Chloromethane
1,2,4-T ri chlorobenzene
2-Chlorotoluene
1,1,1-T richloroethane
4-Chlorotoluene
1,1,2-T richloroethane
1,2-Dibromo-3-chloropropane
Trichloroethene
1,2-Dibromoethane
Trichlorofluoromethane
Dibromomethane
1,2,3-Trichloropropane
1,2-Dichlorobenzene
1,2,4-T rimethylbenzene
1,3-Dichlorobenzene
1,3,5-T rimethylbenzene
1,4-Dichlorobenzene
Vinyl chloride
Dichlorodifluoromethane
o-Xylene
1,1-Dichloroethane
m-Xylene
1,2-Dichloroethane
p-Xylene
1,1-Dichloroethene

TABLE 2-5
METHODS 8031 AND 8032 (GC) AND 8033 (GC WITH
NITROGEN-PHOSPHORUS DETECTION)
Method 8031: Acrylonitrile
Method 8032: Acrylamide
Method 8033: Acetonitrile
TWO-29
Revision 4
January 1998
-------
TABLE 2-6
METHOD 8041 (GC) - PHENOLS
2-Chlono-6-meihylpherioI

2,4-Dinitrophenol
4-Chloro-2-methy1phenol

2,5-Dinitrophenol
4-Chloro-3-methylphenol

Dinoseb
2-Chlorophenol

2-Methy!-4,6-dinitrophenol
3-ChIorophenoI

2-Methylphenol (o-Cresol)
4-Chlorophenol

3-MethyIphenol (m-Cresol)
2-CyelohexyI-4,6-dinitro-

4-Methylphenol (p-Cresol)
phenol

2-Nitraphenol
2,3-DichIorophenol

3-Nitrophenol
2,4-DichIorophenol

4-Nitrophenol
2,5-Dichlorophenol

Fentachlorophenol
2,6-Dichlorophenol

Phenol
3,4-Dichlorophenol

2,3,4,5-Tetrachlorophenol
3,5-Dichlorophenol

2,3,4,6-Tetrachlorophenol
2,3-Dimethylphenol

2,3,5,6-Tetrachlorophenol
2,4-Dimethyiphenol

2,3,4-TrichlorophenoI
2,5-Dimethylphenol

2,3,5-T richiorophenol
2,6-DimethylphenoI

2,3,6-Trichiorophenol
3,4-Dimethylphenol

2.4.5-Trichlorophenol
2.4.6-Trichlorophenol

TABLE 2-7
METHOD 8061 (GC/ECD) -
PHTHALATE ESTERS
Bis(2-n-butoxyethyl) phthalate

Dicyclohexyl phthalate
Bis(2-ethoxyethyl) phthalate

Dihexyl phthalate
Bis(2-ethylhexyl) phthalate

Diisobutyi phthalate
Bis(2-methoxyethyl) phthalate

Di-n-butyl phthalate
Bis(4-methyl-2-pentyl)-

Diethyl phthalate
phthalate

Dinonyl phthalate
Butyl benzyl phthalate

Dimethyl phthalate
Diamyl phthalate

Di-n-octyl phthalate
Hexyl 2-ethylhexyl phthalate
TABLE 2-8
METHOD 8070 (GC) - NITROSAMINES
N-Nitrosodimethylamine
N-Nitrosodiphenylamine
N-Nitrosodi-n-propylamine
TWO-30
Revision 4
January 1998
-------
TABLE 2-9
METHOD 8081 (GC) - ORGANOCHLORINE PESTICIDES AND PCBs
Alachlor
Dichlone
Hexachlorobenzene
Aidrin
Dicofol
Hexachlorocyclo-
a-BHC
Dieldrin
pentadiene
p-BHC
Endosulfan I
Isodrin
5-BHC
Endosulfan II
Methoxychlor
Y-BHC (Lindane)
Endosulfan sulfate
Mirex
Captafol
Endrin
Nitrofen
Chlorobenzilate
Endrin aldehyde
^rans-Nonachlor
a-Chlordane
Endrin ketone
PCNB
y-Chlordane
Etridiazole
Permethrin (c/s and
Chlordane (NOS)
Halowax-1000
trans)
Chloroneb
Halowax-1001
Perthane
Chloropropyiate
Halowax-1013
Propachlor
Chlorothalonil
Halowax-1014
Strobane
DBCP
Halowax-1051
Toxaphene
DCPA
Halowax-1099
Trifluralin
4,4-DDD
Heptachlor

4,4'-DDE
Heptachlor

4,4'-DDT
epoxide

Diallate

TWO-31
Revision 4
January 1998
-------
TABLE 2-10
METHOD 8082 (GC) - POLYCHLORINATED BIPHENYLS
Aroclor 1016
Aroclor 1221
Aroclor 1232
Aroclor 1242
Aroclor 1248
Aroclor 1254
Aroclor 1260
2-Chiorobiphenyl
2,3-Dichlorobiphenyl
2,2',5-T richlorobiphenyl
2,4',5-Trichlorobiphenyl
2,2,,315'-Tetrachlorobiphenyl
2,2',5,5-Tetrachlorobiphenyl
2,3',4,4-Tetrachlorobiphenyl
2,2\3,4,5'-PentachlorobiphenyI
2,2',4,515'-Pentachlorobiphenyl
2,3,3',4',6-Pentachlorobiphenyt
2,2\3,4,4\5,-Hexachlorobiphenyi
2,2',3,4,5,5-Hexachlorobiphenyl
2,2',3,5,5',6-Hexachlorobiphenyl.
2,2',414\5,5,-Hexachlorobiphenyl
2,2',3,3',4,4',5-Heptachlorobiphenyl
2,2'l314,4,,5,5'-Heptachlorobiphenyl
2,2',314,4\5',6-Heptachloro-
biphenyl
2,2\3,4,,5,5,,6-Heptachlorobiphenyl
2,2,I313,,4,4,,5,5\6-N6nachloro-
biphenyl
TABLE 2-11
METHOD 8091 (GC) - NITROAROMATICS AND CYCLIC KETONES
Benefiri
Butralin
1-Chloro-2,4-diriitrobenzene
1 -Chloro-3,4-dinitrobenzene
1-Chloro-2-nitrobenzene
1-Chloro-4-nitrobenzene
2-Chloro-6-nitrotoluene
4-Chloro-2-nitrotolue ne
4-Ch loro-3-nitrotolue ne
2.3-Dichloronitrobenzene
2.4-Dichloronitrobenzene
3.5-Dichloronitrobenzene
3.4-Dichloronitrobenzene
2.5-DichIoronitrobenzene
Dinitramine
1.2-Dinitrobenzerie
1.3-Dinitrobenzene
1.4-Dinitrobenzene
2,4-Dinitrotoluerie
2,6-Dinitrotoluene
Isopropalin
1,2-Naphthoquinone
1,4-Naphthoquinone
Nitrobenzene
2-NitrotoJuene
3-Nitrotoluene
4-Nitrotoluene
Penoxalin [PendimethalinJ
Pentachloronitrobenzene
Profluralin
2.3.4.5-TetrachIoronitrobenzene
2.3.5.6-Tetrachloronitrobenzene
1.2.3-Trichloro-4-nitrobenzene
1.2.4-Trichloro-5-nitrobenzene
2,4,6-Trichlononitrobenzene
Trifluralin
TWO-32
Revision 4
January 1998
-------
TABLE 2-12
METHOD 8100 - POLYNUCLEAR AROMATIC HYDROCARBONS
Acenaphthene
Acenaphthylene
Anthracene
Benz(a)anthracerie
Benzo(b)fluoranthene
Benzo(j)fluoranthene
Benzo(k)fluoranthene
Benzo(g,h,i)peryIene
Benzo(a)pyrene
Chrysene
Dibenz(a,h)acridine
Dibenz(a ,j)acridine
Q'benz(a,h)anthracene.
7H-Dibenzo(c,g)carbazoIe
Dibenzo(a,e)pyrene
Dibenzo(a,h)pyrene
Dibenzo(a,i)pyrene
Fluoranthene
Fluorene
Indeno(1,2,3-cd)pyrene
3-Methyicholanthrene
Naphthalene
Phenanthrene
Pyrene
TABLE 2-13
METHOD 8111 (GC) - HALOETHERS
Bis(2-chIoroethoxy)methane
Bis(2-chioroethyl) ether
Bis(2-chIoroisopropyl) ether
4-Bromophenyl phenyl ether
4-Chlorophenyi phenyl ether
2-Chlorophenyl 4-nitrophenyl ether
3-Chlorophenyl 4-nitrophenyl ether
4-Chlorophenyf 4-nitrophenyl ether
2,4-Dibnomophenyf 4-nitrophenyl
ether
2.4-Dichlorophenyl 3-methyl-4-
nitrophenyl ether
2,6-Dichlorophenyf 4-nitrophenyl
ether
3.5-Dichlorophenyl 4-nitrophenyl
ether
2,5-Dichlorophenyl 4-nitrophenyl
ether
2,4-Dichlorophenyl 4-nitrophenyl
ether
2.3-Dichlorophenyi 4-nitrophenyl
ether
3.4-Dichloropheny! 4-nitrophenyl
ether
4-Nitrophenyl phenyl ether
2,4,6-Trichlorophenyl 4-nitrophenyl
ether
2,3,6-Trichlorophenyi 4-nitrophenyl
ether
2,3,5-Trichlorophenyf 4-nitrophenyl
ether
2,4,5-Trichiorophenyl 4-nitrophenyl
ether
3,4,5-Trichlorophenyl 4-nitrophenyl
ether
2,3,4-Trichlorophenyl 4-nitrophenyl ether
TWO-33
Revision 4
January 1998
-------
TABLE 2-14
METHOD 8121 (GC) - CHLORINATED HYDROCARBONS
Benzal chloride
Benzotrichloride
Benzyl chloride
2-Chloronaphthalene
1 »2-Dichlorobenzene
1.3-Dichlorobenzene
1.4-Dichlorobenzene
Hexachlorobenzene
Hexachlorobutadiene
a-Hexachlorocyclohexane
[a-BHC]
P-Hexachlorocyclohexane
P-BHq
5-Hexachlorocyclohexane
[5-BHC]
Y-Hexachlorocyclohexane [y-BHC]
Hexachlorocyclopentadiene
Hexachloroethane
Pentachlorobenzene
1.2.3.4-Tetrachlorobenzene
1.2.3.5-Tetrachlorobenzene
1,2,4,5-Tetrachlorobenzene
1.2.3-T richlorobenzene
1.2.4-T richloroberizene
1.3.5-Trichlorobenzene
TABLE 2-15
METHOD 8131 (GC) - ANILINE AND SELECTED DERIVATIVES
Aniline
4-Bromoanillne
2-Bromo-6-chloro-4-nitroanilne
2-Bromo-4,6-dintroaniline
2-Chloroaniline
3-Chloroaniline
4-Chloroaniline
2-Chloro-4,6-dinitroaniIine
2-Chloro-4-nitroaniline
4-ChIoro-2-nitroaniIine
2,6-Dibromo-4-nitroaniline
3,4-Dichloroaniline
2,6-Dichloro-4-nitroaniline
2,4-Dinitroaniline
2-Nitroaniline
3-Nitroaniiine
4-Nitroaniline
2,4,6-Trichloroaniline
2,4,5-Trichloroaniline
TWO-34
Revision 4
January 1998
-------
TABLE 2-16
METHOD 8141 (GC) - ORGANOPHOSPHORUS COMPOUNDS
Aspon
Fenthion
Atrazirie
Fonophos
Azinphos-ethyi
Hexamethyt phosphoramide (HMPA)
Azinphos-methyt
Leptophos
Bolstar (Sulprofos)
Malathion
Carbophenothion
Merphos
Chlorfenvinphos
Mevinphos
Chlorpyrifos
Monocrotophos
Chlorpyrifos methyl
Naled
Coumaphos
Parathion, ethyl
Crotoxyphos
Parathion, methyl
Demeton-O, and -S
Phorate
Diazinon
Phosmet
Dichlorofenthion
Phosphamidon
Dichlorvos (DDVP)
Ronnel
Dicrotophos
Simazine
Dimethoate
Stirophos (Tetrachlorvinphos)
Dioxathion
Sulfotepp
Disulfoton
Tetraethyl pyrophosphate (TEPP)
EPN
Terbufos
Ethion
Thionazin (Zinophos)
Ethoprop
Tokuthion (Prothiophos)
Famphur
Trichlorfon
Fenitrothion
Trichloronate
Fensulfothion
Tri-o-cresyl phosphate (TOCP)
TABLE 2-17
METHOD 8151 (GC USING METHYLATION OR PENTAFLUOROBENZYLATION
DERIVATIZATON) - CHLORINATED HERBICIDES
Acifluorfen
Dicamba
MCPP
Bentazon
3,5-Dichlorobenzoic
4-Nitrophenol
Chlorannben
acid
Pentachlorophenol
2,4-D
Dichloroprop
Picloram
Dalapon
Dinoseb
2,4,5-TP (Silvex)
2,4-DB
5-Hydroxydicamba
2,4,5-T
DCPA diacid
MCPA

TWO-35
Revision 4
January 1998
-------
TABLE 2-18
METHOD 8260 (GC/MS)- VOLATILE ORGANIC COMPOUNDS
Acetone
Dibromomethane
Methylene chloride
Acetonitrile
1,2-Dichlorobenzene
Methyl acrylate
Acrolein (Propenal)
1,3-Dichlorobenzene
Methyl methacrylate
Acryionitrile
1,4-Dichlorobenzene
4-Methyl-2-pentanone
Allyl alcohol
cis-1,4-Dichloro-
(MIBK)
Altyl chloride
2-butene
Naphthalene
Benzene
trans-1,4-Dichloro-2-
Nitrobenzene
Benzyl chloride
butene
2-Nitropropane
Bis(2-chloroethyl)-
Dichlorodifluoromethane
N-Nitroso-di-n-
suifide
1,1-Dichloroethane
butylamine
Bromoacetone
1,2-Dichloroethane
Paraldehyde
Bromobenzene
1,1-Dichloroethene
Perrtachloroethane
Bromochloromethane
cis-1,2-Dichloroethene
Pentafluorobenzene
Bromodichloromethane
trans-1,2-DichIoro-
2-Pentanone
Bromoform
ethene
2-Picoline
Bromomethane
1,2-Dichloropropane
1-Propanol
n-Butanol
1,3-Dichloropropane
2-Propanol
2-Butanone (MEK)
2,2-Dichloropropane
Propargyl alcohol
t-Butyl alcohol
1,3-Dichloro-2-propanol
B-Propiolactone
n-Butylbenzene
1,1 -Dichloropropene
Propionitrile (Ethyl
sec-Butylbenzene
cis-1,3-Dichloropropene
cyanide)
tert-Butylbenzene
trans-1,3-Dichloro-
n-Propylamine
Carbon disulfide
propene
n-Propylbenzene
Cartoon tetrachloride
1,2,3,4-Diepoxybutane
Pyridine
Chloral hydrate
Diethyl ether
Styrene
Chloroacetonitrile
1,4-Dioxane
1,1,1,2-Tetrachloro-
Chlorobenzene
Epichlorohydrin
ethane
1-ChlorobLitane
Ethanol
1,1,2,2-T etrachloro-
Chlorodibromomethane
Ethyl acetate
ethane
Chloroethane
Ethylbenzene
Tetrachloroethene
2-Chloroethanol
Ethylene oxide
Toluene
2-Chloroethyl vinyl
Ethyl methacrylate
o-Toluidine
ether
Hexachlorobutadiene
1,2,3-T richlorobenzene
Chloroform
Hexachloroethane
1,2,4-T richlorobenzene
1-Chiorohexane
2-Hexanone
1,1,1 -T richloroethane
Chloromethane
2-Hydroxypropionitrile
1,1,2-T richloroethane
Chioroprene
iodomethane
Tricftloroethene
3-Chloropropionitrile
Isobutyl alcohol
T richlorofluoromethane
2-Chlorotoluene
Isopropylbenzene
1,2,3-T richloropropane
4-Chlorotoluene
p-lsopropyltoluene
1,2,4-T rimethylbenzene
Crotonaldehyde
Malononitrile
1,3,5-T rimethylbenzene
1,2-Dibromo-3-
Methacrylonitrile
Vinyl acetate
chloropropane
Methanol
Vinyl chloride
1,2-Dibromoethane
Methyl-t-butyl ether
o-Xylene
Dibrornofluoromethane

m-Xylene
p-Xylene
TWO-36
Revision 4
January 1998
-------
TABLE 2-19
METHOD 8270 (GC/MS) - SEMI VOLATILE ORGANIC COMPOUNDS
Acenaphthene
Acenaphthylene
Acetophenone
2-Acetylamiriofluorene
1 -Acetyl-2-thiourea
Aldrin
2-Aminoanthraquinone
Aminoazobenzene
4-Aminobiphenyi
3-Amino-9-ethyl-
carbazole
Anilazine
Aniline
o-Anisidine
Anthracene
Aramite
Aroclor-1016
Aroclor-1221
Aroclor-1232
Aroclor-1242
Aroclor-1248
Aroclor-1254
Aroclor-1260
Azinphos-methyl
Barban
Benz(a)anthracene
Benzidine
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzoic acid
Benzo(g,h,i)perylene
Benzo(a)pyrene
p-Benzoquinone
Benzyl alcohol
a-BHC
0-BHC
5-BHC
y-BHC (Lindane)
Bis(2-chloroethoxy)-
methane
Bis(2-chloroethyl)
ether
Bis(2-chloroisopropyl)
ether
Bis(2-ethylhexyl)
phthalate
4-Bromophenyl phenyl
ether
Bromoxynil
Butyl benzyl phthalate
Captafol
Captan
Carbaryl
Carbofuran
Carbophenothion
Chiordane (NOS)
Chlorfenvinphos
4-Chloroaniline
Chlorobenzilate
5-Chloro-2-methyl-
aniline
4-Chloro-3-methyiphenol
3-(Chloromethyl)-
pyridine hydro-
chloride
1 -Chloronaphthalene
2-ChloronaphthaIene
2-Chlorophenol
4-Chloro-1,2-phenylene-
diamine
4-Chloro-1,3-phenylene-
diamine
4-ChlorophenyI phenyl
ether
Chrysene
Coumaphos
p-Cresidine
Crotoxyphos
2-Cyclohexyi-4,6-
dinitrophenol
4,4-DDD
4,4-DDE
4,4'-DDT
Demeton-O
Demeton-S
Diallate (cis or trans)
2,4-DiaminotoIuene
Dibenz(a,j)acridine
Dibenz(a, h)anthracene
Dibenzofuran
Dibenzo (a, e)pyrene
1,2-Dibromo-3-
chloropropane
Di-n-buty! phthalate
Dichlone
1,2-Dichlorobenzene
1.3-Dichlorobenzene
1.4-Dichlorobenzene
3.3-Dichjorobenzidine
2.4-Dichlorophenol
2,6-Dichlorophenol
Dichlorovos
Dicrotophos
Dieldrin
Diethyl phthalate
Diethylstilbestrol
Diethyl sulfate
Dimethoate
3,3'-Dimethoxybenzidine
Dimethylaminoazobenzene
7,12-Dimethylbenz(a)-
anthracene
3,3'-Dimethylbenzidine
a,a-Dimethytphenethyl-
amine
2,4-Dimethytpheno!
Dimethyl phthalate
1.2-Din'rtrobenzene
1.3-Dinitrobenzene
1.4-Dinitrobenzene
4,6-Dinitro-2-methyl-
phenol
2,4-Dinitrophenol
2.4-DinitrotoIuene
2,6-Dinitrotoluene
Dinocap
Dinoseb
Diphenylamine
5.5-Diphenylhydantoin
1,2-Diphenylhydrazine
Di-n-octyl phthalate
Disulfoton
Endosulfan I
Endosulfan II
Endosulfan sulfate
Endrin
Endrin aldehyde
Endrin ketone
EPN
Ethion
Ethyl carbamate
Ethyl methanesulfonate
Famphur
Fensulfothion
TWO-37
Revision 4
January 1998
-------
TABLE 2-19 (CONTINUED)
Fenthion
Naphthalene
Phosphamidion
Fluchloralin
1,4-Naphthoquinone
Phthalic anhydride
FJuoranthene
1-Naphthylamine
2-Picoline (2-Methylpyridine)
Fluorene
2-Naphthylamine
Piperonyl sulfoxide
Heptachlor
Nicotine
Pronamide
Heptachlor epoxide
5-Nitroacenaphthene
Propylthiouradl
Hexachlorobenzene
2-Nitroaniline
Pyrene
Hexachlorobutadiene
3-NKroaniline
Resordnol
Hexachlorocyclo-
4-Nitroaniline
Safrole
pentadiene
5-Nitro-o-anisidine
Strychnine
Hexachioroethane
Nitrobenzene
Sulfaliate
Hexachlorophene
4~Nitrobiphenyl
Terbufos
Hexachloropropene
Nitrofen
1,2,4,5-Tetrachloro
Hexamethylphosphoramide
2-Nitrophenol
benzene
Hydroquinone
4-N'rtrophenol
2,3.4,6-Tetrachloro-
lndeno(1,2,3-ed)pyrene
Nitroquinoline-1 -oxide
phenol
Isodrin
N-Nitrosodi-n-
Tetrachforvinphos
Isophorone
butylamine
Tetraethyl dithio-
Isosafrole
N-Nitrosodiethylamine
pyrophosphate
Kepone
N-Nitrosodimethylamine
Tetraethyl
leptophos
N-Nitrosodiphenylamine
pyrophosphate
Malathion
N-Nitrosodi-n-propyl-
Thionazine
Maleic anhydride
amine
Thiophenoi
Mestranol
N-Nitrosomethylethyl-
(Benzenethiol)
Methapyrilene
amine
Toluene diisocyanate
Methoxychlor
N-Nitrosomorpholine
o-Toluidine
3-Methytcholanthrene
N-Nitrosopiperidine
Toxaphene
4,4-Methylenebis-
N-Nitrosopyrrolidine
1,2,4-Trichlorobenzene
(2-chloroaniline)
5-Nitroo-toluidine
2,4,5-TrichlorophenoI
4,4-MethyIenebis-
Octamethyl pyrophos-
2,4,6-T richlorophenol
(N, N-dimethylaniline)
phoramide
0,0,0-Triethyl
Methyl methanesulfonate
4,4'-Oxydianiline
phosphorothioate
2-MethylnaphthaIene
Parathion
Trifluralin
Methyl parathion
Pentachlorobenzene
2,4,5-Trimethylaniline
2-MethyIphenol
Pentachloronitrobenzene
Trimethyl phosphate
3-MethylphenoI
Pentachiorophenol
1,3,5-T rinitrobenzene
4-MethyIphenol
Phenacetin
T ris(2,3-dibromopropyl)
Mevlriphos
Phenanthrene
phosphate
Mexacarbate
Phenobartital
Tri-p-tolyl phosphate
Mirex
Phenol

Monocrotophos
1,4-Phenylenediamine

Naled
Phorate

Phosalone

Phosmet

TWO-38
Revision 4
January 199B
-------
TABLE 2-20
METHOD 8275 (TE/GC/MS) - SEMIVOLATILE ORGANIC COMPOUNDS
Acenaphthene
Pyrene
2,3',4,4',5-Penta-
Acenaphthylene
1,2,4-T richlorobenzene
chlorobiphenyl
Anthracene
2-ChlorobiphenyI
2,2',3,4,4',5'-
Be nz(a) a nth racen e
3,3'-Dichlorobipheny!
Hexachlorobiphenyl
Benzo(a)pyrene
2,2',5-Trichloro-
2,2', 3,3', 4,4'-
Benzo(b)fluoranthene
biphenyl
Hexachlorobiphenyi
Benzo(g,h,i)perylene
2,3',5-TrichIoro-
2,2',3,4',5,5',6-
Benzo(k)fluoranthene
biphenyl
Heptachlorobiphenyl
4-Bromophenyl phenyl ether
2,4',5-Trichloro-
2,2',3,4,4',5,5'-
1-Chloronaphthalene
biphenyl
Heptachlorobiphenyl
Chrysene
2,2',5,5-Tetrachloro-
2,2",3,3',4,4',5-
Dibenzofuran
biphenyl
Heptachlorobiphenyl
2,2',3,3',4,4',5,5'-
Dibenz(a,h)anthracene
2,2*4,5-Tetrachloro-
Dibenzothiophene
biphenyl
Octachlorobiphenyl
Fluoranthene
2,2'3,5-Tetrachloro-
2,2',3,3'4,4',5,5',6-
Fluorene
biphenyl
Nonachlorobiphenyl
Hexachlorobenzene
2,3,,414'-Tetrachloro-
2,2\3,3'4,4',5,5',6f6'-
lndeno(1,2,3-cd)pyrene
biphenyl
Decachlorobiphenyl
Naphthalene
2,2',4,5,5'-Penta-

Phenanthrene
chlorobiphenyl

TABLE 2-21
METHODS 8280 (HRGC/LRMS) AND 8290 (HRGC/HRMS) -
POLYCHLORINATED DIBENZO-p-DlOXINS (PCDDs)
AND POLYCHLORINATED DIBENZOFURANS (PCDFs)
2,3,7,8-TCDD
TCDD, total
1,2,3,7,8-PeCDD
PeCDD, total
1,2,3,4,7,8-HxCDD
1.2.3.6.7.8-HxCDD
1.2.3.7.8.9-HxCDD
HxCDD, total
1,2,3,4,6,7,8-HpCDD
HpCDD, total
OCDD
2,3,7,8-TCDF
TCDF, total
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCDF
«"¦* W> tit
PeCDF, total
1,2,3,4,7,8-HxCDF
1,2,3,6,7,8-HxCDF
1,2,3,7,8.9-HxCDF
2,3,4,6,7,8-HxCDF
HxCDF, total
1.2.3.4.6.7.8-HpCDF
1.2.3.4.7.8.9-HpCDF
HpCDF, total
OCDF
TWO-39
Revision 4
January 1998
-------
TABLE 2-22
METHOD 8310 (HPLC) - POLYNUCLEAR AROMATIC HYDROCARBONS
Acenaphthene
Aceriaphthylene
Anthracene
ienzo(a)anthracene
Benzo(a)pyrer»e
Benzo(b)fluoranthene
Benzo(g,h,i)perylene
Benzo(k)fluoranthene
Chrysene
Dibenzo (a, h)anthracene
Fluoranthene
Fluorene
lndeno{1,2,3-cd)pyrene
Naphthalene
Phenanthrene
Pyrene
TABLE 2-23
METHOD 8315 - CARBONYL COMPOUNDS
Aceialdehyde
Acetone
Acrolein
Benzaldehyde
Butanal (Butyraldehyde)
Crotonaldehyde
Cyclohexanone
Decanal
2,5-Dimethylbenzaldehyde
Formaldehyde
Heptanal
Hexanal (Hexaldehyde)
Isovaleraldehyde
Nonanal
Octanal
Pentanal (Valeraldehyde)
Propanal
(Propionaldehyde)
m-Tolualdehyde
o-Tolualdehyde
p-Tolualdehyde
TWO-40
Revision 4
January 1998
-------
TABLE 2-24
METHOD 8316 (HPLC)
Acrylamide
Aerylonitrile
Acrolein
TABLE 2-25
METHOD 8318 (HPLC) - N-METHYLCARBAMATES
Aldicarb (Temik)
Aldicarb sulfone
Carbaryl (Sevin)
Carbofuran (Furadan)
Dioxacarb
3-Hydroxycarbofuran
Methiocarb (Mesurol)
Methomyl (Lannate)
Promecarb
Propoxur (Baygon)
TWO-41
Revision 4
January 1998
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TABLE 2-26. METHOD 8321 (HPLC/TS/MS) - NONVOLATILE ORGANIC COMPOUNDS
Azo Dves
Disperse Red 1
Disperse Red 5
Disperse Red 13
Disperse Yellow 5
Disperse Orange 3
Disperse Orange 30
Disperse Brown 1
Solvent Red 3
Solvent Red 23
Chlorinated Phenoxvacid Compounds
2,4-D
2,4-D, butoxyethanol ester
2,4-D, ethylhexy! ester
2,4-DB
Dalapon
Dicamba
Dichlorprop
Dinoseb
MCPA
MCPP
Silvex (2,4,5-TP)
2,4,5-T
2,4,5-T, butyl ester
2,4,5-T, butoxyethanol ester
Alkaloids
Strychnine
Caffeine
Oroanophosphoais Compounds
Asulam
Fensulfothion
Dichlorvos
Dimethoate
Disulfoton
Parathion methyl
Merphos
Methomyl
Monocrotophos
Famphur
Naled
Phorate
Trichlorfon
Thiofanox
Tris(2,3-dibromopropyl) phosphate
(Tris-BP)
Anthraauinone Dves
Disperse Blue 3
Disperse Blue 14
Disperse Red 60
Coumarin Dyes
Fluorescent Briahteners
Fluorescent Brightener 61
Fluorescent Brightener 236
Carbamates
Aldicarb
Aldicarb sulfone
Aldicarb sulfoxide
Aminocarb
Barban
Benomyl
Bromacil
Bendiocarb
Carbaryl
Carbendazim
Carbofuran
3-Hydroxycarbofuran
Chloroxuron
Chloropropham
Diuron
Fenuron
Fluometuron
Linuron
Methiocarb
Methomyl
Mexacarbate
Monuron
Neburon
Oxamyl
Propachlor
Propham
Propoxur
Siduron
Tebuthiuron
TWO-42
Revision 4
January 1998
-------
TABLE 2-27
METHOD 8325 (HPLC/PB/MS) - NONVOLATILE ORGANIC COMPOUNDS
Benzidine
Benzoylprop ethyl
Carbaryl
o-Chlorophenyl thiourea
3,3-Dichlorobenzidine
3,3'-Dimethoxybenzidine
3,3'-Dimethylbenzidine
Diuron
Linuron (Lorox)
Monuron
Rotenone
Siduron
TABLE 2-28
METHOD 8330 (HPLC) - NITROAROMATICS AND NITRAMINES
4-Amino-2,6-din'rtrotoluene
(4-Am-DNT)
2-Amino-4,6-dinitrotoluene
(2-Am-DNT)
1.3-Dinitrobenzene (1,3-DNB)
2.4-Dinitrotoluene (2,4-DNT)
2,6-DinitrotoIuene (2,6-DNT)
Hexahydro-1,3,5-trinitro-
1,3,5-triazine (RDX)
Methyl-2,4,6-trinitraphenyl-
nitramine (Tetryl)
Nitrobenzene (NB)
2-Nitrotoluene (2-NT)
3-Nitrotoluene (3-NT)
4-Nitrotoluene (4-NT)
Octahydro-1,3,5,7-tetranitro-
1,3,5,7-tetrazocine (HMX)
1.3.5-Trinitrobenzene (1,3,5-TNB)
2.4.6-Trinitrotoluene (2,4,6-TNT)
TABLE 2-29
METHOD 8331 (REVERSE PHASE HPLC)
Tetrazene
TABLE 2-30
METHOD 8332 (HPLC)
Nitroglycerine
TWO-43
Revision 4
January 1998
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TABLE 2-31
METHOD 8410 - SEMIVOLATILE ORGANIC COMPOUNDS
Acenaphthene
Aceriaphthylene
Anthracene
Benzo(a)anthracene
Benzo(a)pyrene
Benzoic acid
Bis(2-chIoroethoxy)methane
Bis(2-chIoroethyl) ether
Bis(2-chloroisopropyl) ether
Bis(2-ethyfhexyl) phthalate
4-Bromophenyl phenyl ether
Butyi benzyl phthalate
4-Chloroaniline
4-Chloro-3-methylphenol
2-Chloronaphthalene
2-Chlorophenol
4-Chlorophenol
4-Chlorophenyl phenyl ether
Chrysene
Dibenzofuran
Di-n-butyl phthalate
1.2-Dichlorobenzene
1.3-Dichlorobenzene
1.4-Dichlorobenzene
2,4-Dichlorophenol
Diethyl phthalate
Dimethyl phthalate
4,6-Dinitro-2-methylphenol
2,4-Dinitrophenol
2,4-Dinitrotoluene
2,6-Dinitrotoluene
Di-n-octyl phthalate
Di-n-propyl phthalate
Fluoranthene
Fluorene
Hexachlorobenzene
1,3-HexachIorobutadiene
Hexachlorocydopentadiene
Hexachloroethane
Isophorone
2-Methylnaphthalene
2-Methylphenol
4-Methylphenol
Naphthalene
2-Nitroaniline
3-Nitroaniline
4-Nitroaniline
Nitrobenzene
2-Nitrophenol
4-Nitrophenol
N-N'rtrosodimethylamine
N-Nitrosodiphenylamine
N-Nitroso-di-n-propylamine
Pentachiorophenol
Phenanthrene
Phenol
Pyrene
1.2.4-T richlorobenzene
2.4.5-T richlorophenol
2.4.6-T richlorophenol
TABLE 2-32
METHOD 8430 (GC/FT-IR) - BIS(2-CHLOROETHYL) ETHER
AND ITS HYDROLYSIS PRODUCTS
Bis(2-chloroethyl) ether
2-Chloroethanol
2-(2-Chloroethoxy) eth anol
Diethylene glycol
Ethylene glycol
TWO-44
Revision 4
January 1998
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TABLE 2-33. DETERMINATIVE METHODS FOR INORGANIC ANALYTES
Anaiyte Applicable Method(s)
Aluminum 6010, 6020, 6800, 7000, 7010
Antimony 6010, 6020, 6200, 6800, 7000, 7062
Arsenic 6010, 6020, 6200, 7010, 7061, 7062, 7063
Barium .... 6010, 6020, 6200, 6800, 7000, 7010
Beryllium 6010,6020,7000,7010
Boron 6800
Bromide 6500, 9056, 9211
Cadmium 6010, 6020, 6200, 6800, 7000, 7010
Calcium ..... 6010, 6020, 6200, 6800, 7000
Chloride 6500, 9056, 9057, 9212, 9250, 9251, 9253
Chromium 6010, 6020, 6200, 6800, 7000, 7010
Chromium, hexavalent 7195, 7196, 7197, 7198, 7199
Cobalt 6010, 6020, 6200, 7000, 7010
Copper 6010, 6020, 6200, 6800, 7000, 7010
Cyanide 9010, 9012, 9013, 9213
Fluoride 6500, 9056, 9214
Iron 6010, 6020, 6200, 6800, 7000, 7010
Lead 6010,6020,6200,6800,7000,7010
Lithium 6010, 7000
Magnesium 6010, 6020, 6800, 7000
Manganese 6010,6020,6200,7000,7010
Mercury 4500, 6020, 6200, 6800, 7470, 7471, 7472, 7473, 7474
Molybdenum 6010,6200,6800,7000,7010
Nickel ........ 6010, 6020,, 6200, 6800, 7000, 7010
Nitrate 6500, 9056, 9210 •
Nitrite ........ 6500, 9056, 9216
Osmium 7000
Phosphate ............. 6500, 9056
Phosphorus 6010
Phosphorus, white 7580
Potassium 6010, 6020, 6200, 6800, 7000
Rubidium 6200
Selenium 6010, 6020, 6200, 6800, 7010, 7741, 7742
Silver 6010, 6020, 6200, 6800, 7000, 7010
Sodium 6010,6020,7000
Strontium 6010, 6200, 6800, 7000
Sulfate 6500, 9035, 9036, 9038, 9056
Sulfide 9030, 9031, 9215
Thallium 6010, 6020, 6200, 6800, 7000, 7010
Thorium 6200
Tin 6200, 7000
Titanium 6200
Vanadium 6010, 6020, 6200, 6800, 7000, 7010
Zinc 6010, 6020, 6200, 6800, 7000, 7010
Zirconium 6200
January 1998
-------
TABLE 2-34
CONTAINERS, PRESERVATION TECHNIQUES, AND HOLDING TIMES
FOR AQUEOUS MATRICES*
Name
Container
Preservation
Maximum holding time
Inorganic Tests:
Chloride
Cyanide, total and
amenable
to chlorination
Hydrogen ion (pH)
Nitrate
Sulfate
Sulfide
P, G
P, G
P,G
P, G
P, G
P, G
None required 28 days
Cool to 4°C; if oxidizing 14 days
agents present add 5 mL
0.1N NaAs02 per L or
0.06 g of ascorbic acid
per L; adjust pH>12 with
50% NaOH.
See Method 9010 for
other interferences.
None required 24 hours
Cool to 4°C 48 hours
Cool to 4°C 28 days
Cool to 4°C, add zinc 7 days
acetate
Metals:
Chromium VI
Mercury
Metals, except chromium VI
and mercury
Organic Tests:
G
G
G
Cool to 4°C
HN03 to pH<2
HN03 to pH<2
24 hours
28 days
6 months
Acrolein and acrylonitrile
G, PTFE-lined
Cool to 4°C,
* A
14 days

septum
0.008% Na2S203 ,

Adjust pH to 4-5

Benzidines
G, PTFE-lined
Cool to 4°C,
7 days until extraction,

cap
0.008% Na2S2033
40 days after extraction
Chlorinated hydrocarbons
G, PTFE-lined
Cool to 4°C,
7 days until extraction,

cap
0.008% Na2S2033
40 days after extraction
Dioxins and Furans
G, PTFE-lined
Cool to 4°C,
30 days until extraction,

cap
0.008% Na2S2033
45 days after extraction
Haloethers
G, PTFE-lined
Cool to 4°C,
7 days until extraction,

cap
0.008% Na2S2033
40 days after extraction
Nitroaromatics and
G, PTFE-lined
Cool to 4°C,
7 days until extraction,
cyclic ketones
cap
0.008% Na2S2033,
40 days after extraction

store in dark

NKrosamines
G, PTFE-lined
Cool to 4°C,
7 days until extraction,

cap
0.008% Na2S2033,
40 days after extraction

store in dark

(continued on next page)
TWO-46
Revision 4
January 1998
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TABLE 2-34 (continued)
Name Container1 Preservation Maximum holding time
Oil and grease
G
Cool to 4°C,
28 days

add 5 mL diluted HCI

Organic carbon, total (TOO)
P,G
Cool to 4°C,
28 days

store in dark2

Organochlorine pesticides
G, PTFE-lined
Cool to 4°C
7 days until extraction,

cap
Cool to 4°C4
40 days after extraction
Organophosphorus

7 days until extraction,
pesticides
G, PTFE-lined

40 days after extraction

cap

PCBs
G, PTFE-lined
Cool to 4°C
7 days until extraction,

cap

40 days after extraction
Phenols
G, PTFE-lined
Cool to 4°C,
7 days until extraction,

cap
0,008% Na2S2033
40 days after extraction
Phthalate esters
G, PTFE-lined
Cool to 4°C
7 days until extraction,

cap

40 days after extraction
Polynuclear aromatic
G, PTFE-lined
Cool to 4°C,
7 days until extraction,
hydrocarbons
cap
0.008% Na2S2033,
40 days after extraction

store in dark

Purgeable aromatic
G, PTFE-lined
Cool to 4°C,
14 days
hydrocarbons
septum
0.008% Na2S2032,3

Purgeable Halocarbons
G, PTFE-lined
Cool to 4°C,
14 days

septum
0.008% Na2S2033

Total organic halides (TOX)
G, PTFE-lined
Cool to 4°C, Adjust to
28 days

cap
pH<2 with H2S04

Radiological Tests:

Alpha, beta and radium
P, G
HN03 to pH<2
6 months
A Table originally excerpted, in part, from Table II, 49 FR 28, October 26,1984, and revised as appropriate
for SW-846. See Chapter Three, Chapter Four, or the individual methods for more information.
1 Polyethylene (P) or Glass (G)
2 Adjust to pH<2 with H2S04, HCI or solid NaHSO<. Free chlorine must be removed prior to adjustment
3 Free chlorine must be removed by the appropriate addition of Na2S203.
4 Adjust samples to pH 5-8 using NaOH or H2S04
TWO-47
Revision 4
January 1998
-------
TABLE 2-35
PREPARATION METHODS FOR ORGANIC ANALYTES
(Note: Footnote text is located on the last page of the table.)

Matrix
Analyte Type
Aqueous1
Solids
Sludges and
Emulsions1,2
Organic
Liquids,
Tars, Oils
Acid Extractable
3510
3520
(pHs2)
3540
3541
3545
3550
3520
(pH s 2)
3650
35803
Acrolein, Acrylonitrile, and
Acetonitrile
5031
5031
5031
3585
Acrylamide
80324

Aniline and Selected Derivatives
3510
3520
(pH >11}
5031"
3540
3541
3545
3550
3520
(pH >11)
35803
Aromatic Volatiles
5021
5030
5032
5021
5032
5035
5030
5032
3585
Base/Neutral Extractable
3510
3520
(pH >11)
3540
3541
3545
3550
3520
{pH >11)
3650
35803
Carbamates
8318s
8321
8318s
8321
8318s
8318s
Chlorinated Herbicides
8151s
(pH s 2)
8321
8151®
8321
8151®
(pH «; 2)
35803
Chlorinated Hydrocarbons
3510
3520
(pH as
received)
3540
3541
3550
3520
(pH as
recieved)
35803
Dyes
3510
3520
3540
3541
3545
3550

Explosives
83307
8331®
83307
8331®

Formaldehyde
83159
83159

Haloethers
3510
3520
3540
3541
3545
3550

TWO-48
Revision 4
January 1998
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TABLE 2-35
PREPARATION METHODS FOR ORGANIC ANALYTES
(continued)

Matrix
Analyte Type
Aqueous1
Solids
Sludges and
Emulsions1*2
Organic
Liquids,
Tars, Oils
Halogenated Volatiles
5021
5030
5032
5021
5032
5035
5030
" 3585
Nitroaromatic and Cyclic
Ketones
3510
3520
(pH 5-9)
3540
3541
3545
3550
3520
(pH 5-9)
35803
Niirosamines
3510
3520
3540
3541
3545
3550

Non-halogenated Volatiles
5021
5031
5032
5021
5031
5032
5021
5031
5032
5032
3585
Organochlorine Pesticides
3510
- 3520
3535
(pH 5-9)
3540
3541
3545
3550
3520
(pH 5-9)
35803
Organophosphorus Pesticides
3510
3520
(pH 5-8)
3540
3541
3545
3520
(pH 5-8)
35803
Phenols
3510
3520
(pH s 2)
3540 ,
3541
3545
3550
3562
3520
(PH i 2)
3650
35803
Phthaiate Esters
3510
3520
3535
(pH 5-7)
3540
3541
3545
3550
3520
(pH 5- 7)
35803
Polychlorinated Biphenyls
3510
3520
3535
(pH 5-9)
3540
3541
3545
3562
3520
(PH 5-9)
35803
PCDDs and PCDFs
828010
829010
828010
829010
828010
829010
828010
829010
TWO-49
Revision 4
January 1998
-------
TABLE 2-35
PREPARATION METHODS FOR ORGANIC ANALYTES
(continued)

Matrix

Aqueous1

Sludges and
Organic
Analyte Type
Solids
Emulsions12
Liquids,

Tars, Oils
Polynuclear Aromatic
3510
3540
3520
35803
Hydrocarbons
3520
3541
(pH as

(pH as
3545
received)

received)
3550

3561

Volatile Organics
5021
5021
5021
3585

5030
5031
5030

5031
5032
5031

5032
5035
5032

Footnotes for Table 2-35
1 The pH at which extraction should be performed is shown in parentheses,
2 If attempts to break an emulsion are unsuccessful, these methods may be used.
3 Method 3580 is only appropriate if the sample is soluble in the specified solvent
4 Method 8032 contains the extraction, cleanup, and determinative procedures for this analyte.
5 Method 8318 contains the extraction, cleanup, and determinative procedures for these analytes.
6 Method 8151 contains the extraction, cleanup, and determinative procedures for these analytes.
7 Method 8330 contains the extraction, cleanup, and determinative procedures for these analytes.
8 Method 8331 is for Tetrazene only, and contains the extraction, cleanup, and determinative procedures for
this analyte.
9 Method 8315 contains the extraction, cleanup, and determinative procedures for this analyte.
10 Methods 8280 and 8290 contain the extraction, cleanup, and determinative procedures for these analytes.
11 Method 5031 may be used when only aniline is to be determined.
TWO-50
Revision 4
January 1998
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TABLE 2-36. CLEANUP METHODS FOR ORGANIC ANALYTE EXTRACTS
Analyte Type
Method
Acid Extractable
3650, 3640
Base/Neutral Extractable
3650, 3640
Carbamates
831 e1
Chlorinated Herbicides
81512
Chlorinated Hydrocarbons
3620

3640
Haloethers
3620

3640
Nitroaromatics & Cyclic Ketones
3620

3640
Nitrosamines
3610, 3620:3640
Organochlorine Pesticides
3620

3630

3640

3660
Organophosphorus Pesticides
3620
Phenols
3630

3640

3650

80413
Phthalate Esters
3610

3611

3620

3640
Polychlorinated Biphenyls
3620

3630

3640

3660

3665
Polychlorinated Dibenzo-p-Dioxins and
82804
Polychlorinated Dibenzofurans
82904
Polynuclear Aromatic Hydrocarbons
3610

3611

3630

3640

3650
1 Method 8318 contains the extraction, cleanup, and determinative procedures for these analytes.
2 Method 8151 contains the extraction, cleanup, and determinative procedures for these analytes.
3 Method 8041 includes a dervatization technique followed by GC/ECD analysis, if interferences
are encountered using GC/FID.
4 Methods 8280 and 8290 contain the extraction, cleanup, and determinative procedures for these
analytes.
TWO-51
Revision 4
January 1998
-------
TABLE 2-37. DETERMINATIVE METHO
3S ORGANIC ANALYTES
Analyte Type
GC/MS
Method
Specific GC
Method
HPLC
Method
Acid Extractable
8270

Acrolein, Acrylonitrile, Acetonitriie
8260
8031
80331
83152
8316
Acrylamide
8260
8032
8316
Aniline and Selected Derivatives
8270
8131
..
Aromatic Volatiles
8260
8021

Base/Neutral Extractable
8270

8325*
Carbamates

8318,8321
Chlorinated Herbicides
82703
8151
8321
Chlorinated Hydrocarbons
8270
8121

Dyes

8321
Explosives

8330,
8331, 8332
Formaldehyde

8315
Haloethers
8270
8111

Halogenated Volatiles
8260
8011, 8021

Nitroaromatics and Cyclic Ketones
8270
8091
8330s
Nitrosoamines
8270
8070

Non-halogenated Volatiles
8260
8015
8315
Organochlorine Pesticides
82703
8081

Organophosphorus Pesticides
82703
8141
8321
Phenols
8270
8041

Petroleum Hydrocarbons

8015

Phthalate Esters
8270
8061

Polychlorinated Biphenyls
82703
8082

PCDDs and PCDFs
8280
8290

Polynuclear Aromatic Hydrocarbons
8270
8100
8310
Volatile Organics
8260
8011,8015,
8021,8031,
8032, 8033
8315
8316
1 Of these analytes, Method 8033 is for acetonitriie only.
2 Of these analytes, Method 831S is for acrolein only.
3 This method is an alternative confirmation method, not the method of choice.
4 Benzidines and related compounds.
5 Nitroaromatics (see "Explosives").
TWO-52
Revision 4
January 1998
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TABLE 2-38
PREPARATION METHODS FOR INORGANIC ANALYSES1
MATRIX
METHOD
Surface Water
3005, 3010, 3015, 3020
Ground Water
3005, 3010, 3015, 3020
Extracts
3010, 3015, 3020
Aqueous samples containing suspended
solids
3010, 3015, 3020
Oils
3031, 3040, 3051, 30522
Oil Sludges
3031, 30522
Tars
3031, 30522
Waxes
3031, 3040, 30522
Paints
3031, 30522
Paint Sludges
3031, 30522
Petroleum Products
3031, 3040, 30522
Sediments
3050, 3051, 30522, 30603
Sludges
3050, 3051, 30522, 30603
Soil Samples
3050, 3051, 30522, 30603
Ashes
30522
Biological Tissues
30522
'It is the responsibility of the analyst to refer to each analytical method to determine applicability of
the chosen method to a specific waste type and target analyte,
2For total decomposition analysis ONLY.
3 For the analysis of samples for hexavalent chromium ONLY.
TWO-53
Revision 4
January 1998
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TABLE 2-39
USE OF LEACHING, EXTRACTION AND DIGESTION METHODS
FOR INORGANIC ANALYSIS
(Generally ordered by increasing strength)
METHOD
REAGENTS & CONDITIONS
USE
1310
dilute acetic add (synthetic municipal
solid waste leachate)
Simulate leaching of a waste in a
municipal solid waste landfill
1311
dilute acetic acid (synthetic municipal
solid waste leachate)
Simulate leaching of a waste in a
municipal solid waste landfill
1312
dilute H2S04 and HN03 (synthetic acid
rain)
Simulate acid rain leaching of a
waste
1320
dilute H2S04 and HN03 (synthetic acid
rain)
Simulate long-term acid rain
leaching of a waste
3040
solvent
Dissolution of oils, oily wastes,
greasses and waxes
3005
HN031 heat
Surface and ground waters
3020
HN03, heat
Aqueous samples and extracts for
GFAA work only
3010
HNOa, HCt, heat
Aqueous samples and extracts
3060A
NajCOa/NaOH, heat
Soils, sludges, sediments and some
industrial wastes for the analysis of
hexavalent chromium only.
3015
HN03, HCI (optional), pressure, heat
Aqueous samples and extracts
3050
HN03( H202i HCI (optional), heat
Sediments, soils, and sludges
3051
HNOa, HCI (optional), pressure, heat
Sludges, sediments, soils and oils
3031
Potassium permanganate, H2S04, HN03,
HCI, heat
Oils, oily sludges, tars, waxes, paint,
paint sludge
3052
HN03, HF, HCI (optional) H2Oz (optional),
heat, pressure
Siliceous, organic and other
complex matrices for total sample
decomposition
TWO -54
Revision 4
January 1998
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FIGURE 2-1
ORGANIC ANALYSIS OPTIONS FOR SOILD
AND LIQUID MATRICES
Extractable
Volatile
Organio Liquid
Oil
Soild or
Sludge
Aqueous Sample
No
Aqueous
Liquid
Solid
Organic or
Liquid Oil
Yes
No
Analysis
Procedure?
GC/MS
cleanup
needed?
HPLC
Yes
GC
HPLC
GC
Analysis
procedure?
GC/MS
/ sample \
to be analyzed
for extractable*
or vola-
\ tiles? /
Sample
Sample
Prep:
3585.
OC/MS
Procedure:
8270.
Physical
Characteristic
of Sample.
Sample Prep:
5030, 5031,
5032, 8011
Sample Prep:
5021, 5032,
5035.
Extraction
Procedure:
3510, 3520,
3535
GC/MS Analysis
Procedure:
8260
Extraction
Procedure: 3650
or 3580.
Extraction
Procedure: 3540,
3541, 3550, 3560,
3561, 3562.
HPLC Analysis Procedures:
8310, 8318, 8321, 8325, 8330,
8331, 8332.
HPLC Analysts Procedures:
Acrolein, Acrylonitrile, Acrylamide:
Carbaryl Compounds:
8316
8315
GC Analysis Procedures:
EDB and DBCP:
Noiihalogenated Volatile Organics:
Halogenated Volatile Compounds:
Acrylamide:
Acetonitrile:
8011
8015
8021
8032
8033
Cleanup Procedure:
Alumina Column:
Alumina Column for Petroleum Wastes:
Plorisil Column:
Silica Gel Column:
Gel Permeation:
Acid Base Partioning:
Sulfur.
Sulfuric Acid Cleanup:
3610
3611
3620
3630
3640
3650
3660
3665
GC Analysis Procedures:
Phenols:
Phthalate Esters:
Nitrosamines: '
Organochlorine Pesticides:
PCBs:
Nitroaromatics and Cyclic Ketones:
Polynucleor Aromatic Hydrocarbons:
Haloethers:
Chlorinated Hydrocarbons:
Oiganophosphorus Pesticides:
Chlorinated Herbicides:
8041
8061
8070
8081
8082
8091
8100
8111
8121
8141
8151
Revision 4
TWO - 55 January 1998
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FIGURE 2-2
SCHEMATIC OF SEQUENCE TO DETERMINE
IF A WASTE IS HAZARDOUS BY CHARACTERISTIC
C~ Start
DOT (49 CFR 173.300)
Is waste
a gas?
No
waste
reactive to
air and/w
•s. water? .
No
Yes

I No
Generator Knowledge
DOT (49 CFR 173.151)
Yes
What is ^
physical state
s. of waste?
Solid
Hazardous J
No
jquid
Perform Paint
RlterTest
{Method 9095)
Methods 1110 and 9040
Mo
Methods 1010 or 1020
No
TWO-56
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FIGURE 2-2
(Continued)
Non hazardous
for ignitability
characteristic
Reactive CN
and Sulfide Tests
Does waste
generate toxic
gas?
No
Nonhazardous
tor toxic gas generation
(reactivity) characteristic
/ Is total >
ooncen. of TC
constituents20 <
v TC regulatory
\ limit? /
Yes
No
Stop
' Is waste
teachable and
toxic?
{Method 1311)
es
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FIGURE 2-3A
RECOMMENDED SW-846 METHODS FOR ANALYSIS OF EP LEACHATES
Leach Method
Sample

13
10
Prep. Methods
3010,3015
(7760 Ag)
Determinative
Methods
6010
6020
7000
7010
3510/3520/3535
Neutral
Pesticides
8151/8321
Herbicides
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FIGURE 2-3B
RECOMMENDED SW-846 METHODS FOR ANALYSIS OF TCLP LEACHATES
Leach
Method
Prep.
Methods
Determinative
Methods
Sample

TCLP

3510/
35
Net
3520/
35
itral
7470
Hg
8081
Pesticides
8260
Volatile
Organics
3510/3520
(Acidic
and Basic)
8270
Semivol-
atile
Organics
8151/8321
Herbicides
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FIGURE 2-4A.
GROUND WATER ANALYSIS: ORGANIC ANALYTES
8280 or
8290
8270
Dioxins
8081
3510, 3520
or 3535
Neutral
VOA
Pesticides
8260
Herbicides
8151 or
8321
Semivolatiies
3510 or
3520
3620, 3640,
and/or 3660
Organic
Sample
1 - Optional; Cleanup required only if interferences prevent analysis.
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FIGURE 2-4B.
GROUND WATER ANALYSIS: INDICATOR ANALYTES
Indicator
Anaiytefs}
Br-.6500,
9056, 9211
Field
Teste
TOX:
9020
TOO:
9060
POC
POX;
9021
pH:
9040/9041
CN-:
9213,
9010/9012
NCy/NCV:
6500, 9056,
9210, 9216
a-:
6500, 9212,
9260/9251,
9258, 90S6
SO/: 6500
9035/9036
9038/9056
Phenolics:
9066,9067
NH,:
9210
F-: 6500,
9056,9214
Specific
Conductance:
9050
1 - Barcelona, 1984, (See Reference 11
2 - Riggin, 1984, (See Reference 2)
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FIGURE 2-4C.
GROUND WATER ANALYSIS: INORGANIC ANALYTES
GROUND WATER1
SAMPLE
7010
6020
7000
6010
7741 or 7742
Se
7470 or 7472
7061, 7062,
or 7063
As
Ag, Al, As, Ba,
Be, Cd. Co, Cr,
Cu, Mn, Ni, Pb,
Sb, Tl, Zn
Ag, Al, As, Ba,
Be, Cd, Co, Cr,
Cu, Fe, Mg, Mn,
Mo, Ni, Pb, Sb,
Se, Tl, V, Zn
Ag, Al, Ba, Be, Ca,
Cd, Co, Cr, Cu, Fe,
K, Li, Mg. Mn, Mo.
Na, NI, Os, Pb, Sb,
Sn, Sr, Tl, V, Zn
SAMPLE PREPARATION
3005 OR 3015
Ag, As, Ba, Be,
Cd, Co, Cr, Cu,
Fe, Pb, Mn, Mo,
Ni, Sb, Se, "F1,
V, Zn
SAMPLE PREPARATION
3015 OR 3020
1 When analyzing for total dissolved metals, digestion is not
necessary if the samples are filtered at the time of
collection, and then acidified to the same concentration as the standards.
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CHAPTER THREE
INORGANIC ANALYTES
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the allowed
flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified in a
regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to be
used by the analyst and the regulated community in making judgements necessary to meet the data
quality objectives or needs for the intended use of the data.
3.1 SAMPLING CONSIDERATIONS
3.1.1 Intrpduption
This manual contains procedures for the analysis of inorganic analytes in a variety of matrices.
These methods are written as specific steps in the overall analysis scheme - sample handling and
preservation, sample digestion or preparation, and sample analysis for specific inorganic
components. From these methods, the analyst must assemble a total analytical protocol which is
appropriate for the sample to be analyzed and for the information required. This introduction
discusses the options available in general terms, provides background information on the analytical
techniques, and highlights some of the considerations to be made when selecting a total analysis
protocol.
3.1.2 Definition of Terms
Optimum concentration range: A range, defined by limits expressed in concentration, below
which scale expansion must be used and above which curve correction should be considered. This
range will vary with the sensitivity of the instrument and the operating conditions employed.
Sensitivity: (a) Atomic Absorption: The concentration in milligrams of metal per liter that
produces an absorption of 1%. (b) Inductively Coupled Plasma (ICP): The slope of the analytical
curve, i.e., the functional relationship between emission intensity and concentration.
Method detection limit (MDU: The minimum concentration of a substance that can be
measured and reported with 99% confidence that the analyte concentration is greater than zero. The
MDL is determined from analysis of a sample in a given matrix containing the analyte which has
been processed through the preparative procedure.
Total recoverable metals: The concentration of metals in an unfiltered sample following
treatment with hot dilute mineral acid (Method 3005).
Dissolved metals: The concentration of metals determined in a sample after the sample is
filtered through a 0.45-pm filter (Method 3005).
Suspended metals: The concentration of metals determined in the portion of a sample that is
retained by a 0.45-pm filter (Method 3005).
Total metals: The concentration of metals determined in a sample following digestion by
Methods 3010, 3015, 3020, 3050, 3051, or 3052.
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Instrument detection limit C1DL1: The concentration equivalent to a signal due to the analyte
which is equal to three times the standard deviation of a series of 7 replicate measurements of a
reagent blank's signal at the same wavelength.
Interference check sample (ICS'S: A solution containing both interfering and analyte elements
of known concentration that can be used to verify background and inter-element correction factors.
initial calibration verification flCY) standard: A certified or independently prepared solution used
to verify the accuracy of the initial calibration. For 1CF analysis, it must be run at each wavelength
used in the analysis.
Continuing calibration verification fCC\A: Used to assure calibration accuracy during each
analysis run. It must be run for each analyte as described in the particular analytical method. At a
minimum, it should be analyzed at the beginning of the run and after the last analytical sample. Its
concentration should be at or near the mid-range levels of the calibration curve.
Calibration standards: A series of known standard solutions used by the analyst for calibration
of the instrument (i.e., preparation of the analytical curve).
Linear dynamic range: The concentration range over which the analytical curve remains linear.
Method blank: A volume of reagent water processed through each sample preparation
procedure.
Calibration blank: A volume of reagent water acidified with the same amounts of acids as were
the standards and samples.
Laboratory control standard: A volume of reagent water spiked with known concentrations of
analytes and carried through the preparation and analysis procedure as a sample. It is used to
monitor loss/recovery values.
Method of standard addition fMSAfr The standard-addition technique involves the use of the
unknown and the unknown plus one or more known amounts of standard. See Method 7000, for
detailed instructions.
Sample holding time: The storage time allowed between sample collection and sample analysis
when the designated preservation and storage techniques are employed.
Check Standard: A solution containing a known concentration of analyte derived from externally
prepared test materials. The check standard is obtained from a source external to the laboratory and
is used to check laboratory performance.
3.1.3 Sample Handling and Preservation
Sample holding times, digestion volumes and suggested collection volumes are listed in Table
3-1. The sample volumes required depend upon the number of different digestion procedures
necessary for analysis. This may be determined by the application of graphite-furnace atomic
absorption spectrophotometry (GFAA), flame atomic absorption spectrophotometry (FLAA),
inductively coupled argon plasma emission spectrometry (ICP), hydride-generation atomic absorption
spectrometry (HGAA), inductively coupled plasma mass spectrometry (ICP-MS) or cold-vapor atomic
absorption spectrometry (CVAA) techniques, each of which may require different digestion
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procedures. The indicated volumes in Table 3-1 refer to that recommended for the individual
digestion procedures and to that recommended for sample collection volumes. In alt cases for waste
testing, representative sampling must be maintained.
In the determination of trace metals, containers can introduce either positive or negative errors
in the measurement of trace metals by (a) contributing contaminants through leaching or surface
desorption, and (b) depleting concentrations through adsorption. Thus the collection and treatment
of the sample prior to analysis require particular attention. The following cleaning treatment
sequence has been determined to be adequate to minimize contamination in the sample bottle,
whether borosiiicate glass, linear polyethylene, polypropylene, or Teflon: detergent, tap water, 1:1
nitric acid, tap water, 1:1 hydrochloric acid, tap water, and reagent water.
NOTE: Chromic acid should not be used to clean glassware, especially if chromium is to be
included in the analytical scheme. Commercial, non-chromate products (e.g., Nochromix) may
be used in place of chromic add if adequate cleaning is documented by an analytical quality
control program, (Chromic add should also not be used with plastic bottles.)
3.1.4 Safety
The toxidty or cardnogenidty of each reagent used in these methods has not been precisely
defined. However, each chemical compound should be treated as a potential health hazard. From
this viewpoint, exposure to these chemicals must be reduced to the lowest possible level by
whatever means available. The laboratory is responsible for maintaining a current awareness file
of OSHA regulations regarding the safe handling of the chemicals specified in these methods. A
reference file of material data-handling sheets should also be made available to all personnel
involved in the chemical analysis. The following additional references to laboratory safety are
available:
1. "Carcinogens - Working with Carcinogens," Department of Health, Education, and Welfare,
Public Health Service, Center for Disease Control, National Institute for Occupational Safety and
Health, Publication No. 77-206, August 1977.
2. "OSHA Safety and Health Standards, General Industry," 29 CFR 1910.
3. "Proposed OSHA Safety and Health Standards, Laboratories," Occupational Safety and Health
Administration, 51 FR 26660, July 24, 1586.
4. "Safety in Academic Chemistry Laboratories," American Chemical Society Publication,
Committee on Chemical Safety.
3.1.5 Sample Preparation
Fa* all non-spedated digestion methods, great reduction in analytical variability can be achieved
by use of appropriate sample preparation procedures. Generally the reduction in subsampling
variance is accomplished by drying the sample, redudng its partide size, and homogeneously mixing
the resulting fines.
Specifically, if the sample can not be well mixed and homogenized on an as received basis,
then air or own drying at 60°C or less, crushing, sieving, grinding, and mixing should be performed
as needed to homogenize the sample until the subsampling variance is less than the data quality
objectives of the analysis. While proper sample preparation generally produces great reduction in
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analytical variability, be aware that in certain unusual circumstances there could be loss of volatile
metals (e.g. Hg, organometallics) or irreversible chemical changes (e.g., precipitation of insoluble
species, change in valence state) caused by inappropriate sample preparation procedures.
Variability inherent in the analytical determinative procedure is assessed by matrix spiking of
individually digested samples. Variability due to sample heterogeneity is assessed by analyzing
sample replicates. For most samples, sampling imprecision is much greater than analytical
imprecision. Because of this, the greatest advances in environmental monitoring are occurring in
the area of sample collection and preparation.
3.1.6 Clean Chemistry and the Analytical Blank
The significant role of the analytical blank in chemical analysis of trace metals cannot be over
stressed. Sensitive instrumentation such as inductively coupled plasma mass spectrometry (ICP-
MS), inductively coupled plasma atomic emission spectrometry (ICP-AES), and graphite-furnace
atomic absorption spectrophotometry (GFAA) requires that sample preparation be at least as
sophisticated as the instalments used in analysis. The analytical blank is normally a primary source
of error in trace element analysis. Trace analysis is as dependent on control of the analytical blank
as it is on the accuracy and precision of the instalment making the measurement. Inability to control
contamination that is external to the sample, or those contributions of the analyte coming from all
other sources than the sample, is frequently the limiting factor in trace (parts per million (ppm) to
parts per billion (ppb)) and ultra-trace (ppb to parts per trillion (ppt)) analysis. Analytical blank
contributions occur from the following four major sources (Ref. 1-5):
• the atmosphere in which the sample preparation and analysis are conducted,
• the purity of the reagents used in sample preparation, including all reagents and the
quantities added directly to the sample,
• the materials and equipment used in digestion or extraction vessels that come in contact with
the sample during the sample preparation and analysis, and
¦ the analyst's technique and skill in preparing the samples and performing the analyses.
Only under very few circumstances can the analyst ignore the contribution of the uncertainty of
the blank when calculating the uncertainty of the overall analytical result. One condition to consider
is whether the concentration of the blank is insignificant compared to the analytical level. For
example, when the blank value is less than 103-104 smaller than the sample measurement, the
uncertainty of the blank measurement is insignificant compared with the uncertainty of the analjrtical
measurement. This situation only occurs when the blank signal is extremely low compared to the
measurement, which is rarely the case when trace and ultra-trace analyses are conducted. Typically
the blank value is significant and must be subtracted from the measurement. Because the blank
concentration is closer to the detection limit of the instrument, the imprecision of the blank is large
compared with the blank measurement itself. This relationship causes the analytical blank to
frequently become the limiting factor in the overall measurement precision.
To compute the overall standard deviation for a final measurement, several sources of error and
imprecision must be combined. The standard deviation for each component of the computation of
y must be considered when determining the overall measurement uncertainty.
y(±sy) = a(±st) - bfrsj
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As shown by the following equation, the standard deviation of the result, sy is given by
combining the standard deviations of the measurements (Reference 6).
In this case, "a" represents the standard deviation of the measurement,"b" represents the
standard deviation of the blank, that must be subtracted, and "
-------
The Sample Preparation and Analysis Atmosphere
The laboratory environment in which the sample is prepared is the major source of
contamination for most elements. Some rare elements may be an exception, but for the majority of
elements of interest, contamination from airborne sources is the most significant of the four main
sources. Table 3-3 illustrates concentrations of lead in the air.
This contamination can also be seen in the comparison of 58,000 particles per liter of air
measured in a normal laboratory in Pittsburgh, Pennsylvania, and inside a clean chamber in an
adjacent laboratory five meters away. Figure 1 demonstrates the dramatic difference between the
two environments. Cost effective methods of creating clean chambers for sample preparation are
documented along with this data in the references (1).
Any laboratory air that comes in contact with the sample may deposit some portion of its
elemental content into the sample. The sample is especially vulnerable to this transfer when it is
being decomposed in acid. The acid will leach particles from the air resulting in unwanted ions in
solution, mixing with those of the sample.
To prevent air from contaminating the sample, the sample must be processed in a dean
environment. This is much easier to accomplish than it may appear at first. These precautions are
becoming state-of-the-art in many analytical and environmental laboratories. The prevention of
airborne contamination is most frequently dealt with by employing a laminar flow clean bench or a
clean laboratory facility. Instructions are referenced for the construction of both from component
parts; both are relatively inexpensive and uncomplicated, once the concepts are understood (Ref.
1).
There are many sources of airborne contamination. Several of the sources have been
described and their particle size ranges are provided in Figure 2. These diverse sources primarily
provide particulates in discrete size ranges. Depending on whether the laboratory is located in an
industrial, urban, or rural area, or near the sea, the distribution of these source particles will be
different, as will their composition. The vertical dashed line in Figure 2 indicates the particle size
cutoff, usually 0.5 |jm, for the high efficiency particulate air (HEPA) filter used to prevent particulate
contamination. Particles above this size cannot pass through a HEPA filter that is in good working
order. These filters were developed jointly by the Massachusetts Institute of Technology and Arthur
D. Little & Company, Inc., for the Manhattan Project during World War II and are in common use
today (Refs.1,11).
The definition of clean air is derived from Federal Standard 209a, which defines cleanliness
levels. Table 3-4 lists these conditions. "Laminar flow" is directed coherent air movement that does
not contain any turbulence.
A dramatic reduction in airborne contaminants can be expected by using HEPA filtered air in
laminar flow clean hoods or entire clean laboratories. Table 3-5 demonstrates the dramatic
differences in airborne contaminant concentrations in an ordinary laboratory, a clean laboratory, and
a clean hood inside a clean laboratory.
Reagent Purity
For add decomposition, leaching, and extraction, the purity of the reagents used is extremely
important to the overall level of the blank. Reagents have very different purities depending on their
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processing grade and purpose. Frequently, the analyst must purchase special reagents or purify
lesser grade reagents prior to use in order to minimize the analytical blank.
In addition to the purity of the reagents, the quantity that must be added is also significant.
When reagents are added, they bring with them elemental and molecular components that exist as
contaminants. The more reagent that must be used due to reasons other than the stoichiometric
reaction, the higher the blank. Reagents of high purity must either be purchased or produced in the
laboratory.
In the preparation of high purity reagents, there is only one significant and practical choice for
the method of purification. Sub-boiling distillation (Refs. 13,14), different from normal distillation,
uses an infrared radiation source to heat the reagent to a temperature just below the boiling point.
Not allowing the reagents to boil prevents the "irownian movemenf of solution droplets produced
when bubbles burst at the surface of the liquid. These aerosolized solution particles are carried
everywhere in the apparatus and physically transport metal ions and contaminants that should have
been left in solution. Sub-boiling distillation is a slower but very reliable method of purifying ail of the
common mineral acids and many organic reagents used in analytical methods. It relies exclusively
on the vapor pressure of the reagent, and contaminant, and can therefore be specifically optimized
for purification of the mineral adds if the object is to remove metal ions. Of all acids, nitric acid, for
a variety of reasons, can be purified to excellent quality. Because large quantities of reagents are
necessary for many laboratories and a continuous supply of these reagents is desirable, methods
for constructing a sub-boiling distillation apparatus are provided in the references; sources of these
apparatus are also provided. Purchasing sub-boiling acids from commercial sources is also an
option (1). Construction or purchase of sub-boiling reagent purification equipment is cost effective
depending on the quantity of reagents required.
Materials for Sample Preparation, Storage, and Analysis
For elemental analysis, specific, preferred materials are used for the construction of sample
vessels and instrument components that come in contact with the sample. Over the past two
decades, materials identified as being non-contaminating have become the top choices for bottles,
beakers, reaction vessels, storage containers, nebulizers, and instrument components for trace and
ultra-trace analysis. These materials are the same as those currently being used in many digestion
vessels, bomb liners, and microwave vessels. These materials are thermally durable, chemically
resistant or inert, non-contaminating, and have appropriate compression and tensile strength. Table
3-6 lists tie specific types of materials of non-contaminating nature and chemical inertness to most
add reactions, in order of preference. These materials have been evaluated and tested extensively
for their elemental contamination characteristics (Refs. 1-3,15,16).
With the exception of polyethylene, these are the most common materials used for sample
preparation vessels, both atmospheric pressure vessels and dosed vessel liners that come in
contact with the sample. These materials are the most stable to add reactions (with the exception
of quartz and glass if hydrofluoric acid is used). Fiuoropolymers are the most common and were
adapted from other chemical uses for application in pressure systems. The fiuoropolymers, TFM,
PFA and TFE or PTFE have the highest range of use temperatures of most plastics, ranging from
270 to 300 °C. They are also chemically inert to the majority of mineral acids and combinations
thereof. Sulfuric add has a boiling point of approximately 330 °C and can damage all fiuoropolymers
by melting them. Quartz and glass can safely contain sulfuric add at these high temperatures, but
borosilicate glass is not appropriate for ultra-trace elemental analysis (Refs. 3,15). Glass actually
forms a gel layer that hydrates and leaches, transferring elemental components from the glass to
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the sample solution. While these are minute quantities, there are many low level analyses where
these contributions would be detected in the blank and the sample.
Polyethylene is suitable for storage of diluted samples after decomposition, but it does not have
a thermal use temperature appropriate for decomposition. It is also not sufficiently inert to be useful
as a decomposition vessel or vessel liner, similar to polycarbonate and polypropylene. The low cost
of polyethylene and its relative inertness to cool, weakly acidic solutions make it an excellent storage
container for trace element solutions (Ref, 1).
Analytical Technique and Synergistic Equipment
The fourth significant source of analytical blank contamination is the skill of the analyst and the
appropriateness of the technique that is being performed. Analytical blank control has been
explained as the combination of atmosphere, reagent, material, and the protocol being used
correctly. Here the skill and awareness of the analyst and the way in which combinations of the
aforementioned clean chemistry tools are applied is significant to the final result of contamination
and analytical blank control. Sample preparation instrumentation may also assist in these protocols.
For example, microwave sample preparation assists each of these parameters in synergistic ways,
thus lowering the analytical blank, improving blank precision, and enhancing overall quality control
and transferability of methods. Some instrumentation and fundamental processes involved in
specific sample preparation procedures assists the analyst by incorporating useful clean chemistry
concepts into instrumentation and method structure. Such instrumentation is pertinent since
microwave methods now exist that provide sample preparation for leaching or total analysis of all
elements simultaneously. Analyst skill involving clean chemistry is assisted by the method structure
and microwave equipment as indicated below.
If a closed or controlled atmospheric microwave vessel is prepared in a clean
hood and sealed before leaving the clean environment, the sample will not be
affected by atmospheric contamination during the reaction, since it has not been
removed from a clean environment.
The vessel materials described would not normally be used by many laboratories
that can benefit, so the advantages of the fluoropolymers would not be realized
if they were not required in most microwave reaction vessels.
The time the sample spends in decomposition, leaching, or extraction is typically
reduced from hours to minutes, thus reducing the potential leaching from the
container walls
Because most microwave systems are sealed systems, evaporation of the
reagent before it reacts productively is prevented and smaller quantities of
reagents are used thus preventing excess blank accumulation.
The blank is reduced in size and is more consistent due to limiting the exposure variables. An
example of the these components working together has been provided in the literature where
analysis under different conditions has verified these conclusions (Refs. 1,18,19). The example
illustrates the isolation of the blank optimization areas: environment, reagents, materials, and
analysis skills. The skill of the analyst is kept more constant as the instrument dictates more clean,
chemically appropriate procedures.
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References
(1) Kingston, S, H. M.; Walter, P. J.; Chalk, S.; Lorentzen, E.; Link, D. In Microwave - Enhanced
Chemistry: Fundmentals, Sample Preparation, and Applications; Kingston, H. M. S,, Haswell, S. J.,
Eds.; American Chemcial Society: Washington D.C., 1997, pp 257-279.
(2) Tolg, G.; Tschopel, P. In Determination of Tram Elements-, Affaassi, Z. B„ Ed.; VCH: New York,
1994, pp 1-38.
(3) Murphy, T. J. In National Bureau of Standards Special Publication 422: Accuracy in Trace
Analysis: Sampling, Sample Handling, and Analysis] National Bureau of Standards: Gaithersburg,
MD, 1976, pp 509-539.
(4) Moody, J. R. "NBS Clean Laboratories for Trace Element Analysis", Anal. Chem. 1982, 54,
1358A-1376A.
(5) Adeloju, S. B.; Bond, A. M. "Influence of Laboratory Environment on the Precision and Accuracy
of Trace Element Analysis", Anal. Chem. 1985, 57,1728-1733.
(6) Skoog, D. A,; West, D. M,; Holler, F. J. Fundamentals of Analytical Chemistry, 6th ed.;
Saunders College Publishing: Fort Worth, 1992.
(7) Taylor, J. K. Statistical Techniques for Data Analysis; Lewis Publishers: Chelsea, Ml, 1990.
(8) Taylor, J. K. In Principles of Environmental Sampling; Keith, L. H., Ed.; American Chemical
Society: Washington, DC, 1996, pp 77-83.
(9) Rabinowrtz, M. B.; Wetherill, G. W. "Identifying Sources of Lead Contamination by Stable
Isotope Techniques", Environ. Sci. Techno!. 1972, 6, 705-709.
(10) Maienthal, E. J. In U. S. National Bureau of Standards Technical Note 545; Taylor, J. K., Ed.;
U. S. Governmental Printing Office: Washington, D. C., 1970, pp 53-54.
(11) Zief, M.; Mitchell, J. W. In Chemical Anatysisr, Elving, P. J., Ed.; John Wiley & Sons: New York,
1976; Vol. 47.
(12) Miller, G. T. Living in the Environment, Wadsworth Inc.: Belmont, CA, 1994.
(13) Kuehner, E. C.; Alvarez, R.; Paulsen, P. J.; Murphy, T. J. "Production and Analysis of Special
High-Purity Acids Purified by Sub-Boiling Distillation", Anal. Chem. 1972, 44,2050-2056.
(14) Moody, J. R.; E.S., B. "Purified Reagents for Trace Metal Analysis", Talanta 1982, 29,1003-
1010.
(15) Moody, J. R.; Lindstrom, R. M. "Selection and Cleaning of Plastic Containers for Storage of
Trace Element Samples", Anal. Chem. 1977, 49,2264-2267.
(16) Moody, J. R. The Sampling, Handling and Storage of Materials for Trace Analysis", Philos.
Trans, ft Soc. London 1982, 305, 669-680.
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(17) Kuehner, E. C.; Freeman, D. H. In Purification of Inorganic and Organic Materials; Zief, M., Ed.;
Marcel Dekker. New York, 1969, pp 297-306.
(18) Skeity, E. M.; DiStefano, F. T. "Clean Room and Microwave Digestion Techniques: Improvement
in Detection Limits for Aluminum Determination by GF-AAS", Appf. Spectrosc. 1988, 42,1302-1306.
(19) Prevatt, F. J. "Clean Chemistry for Trace Analysis", Environmental Testing and Analysis 1995,
4, 24-27.
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TABLE 3-1.
SAMPLE HOLDING TIMES, RECOMMENDED DIGESTION VOLUMES AND
RECOMMENDED COLLECTION VOLUMES FOR INORGANIC
DETERMINATIONS IN AQUEOUS AND SOLID SAMPLES
Measurement
Digestion
Volume.
(mL)*'c
Collection
Volume
_Gs!£I
Treatment/
Preservative
Holding Time''
Inorganic Analvtes (except hexavalent chromium and mercury):
Aqueous
Total 100 600
HN03 to pH <2
6 months
Dissolved
100
600
niter on site;
HNOa to pH <2
6 months
Suspended
100
600
Filter on ate
6 months
Solid
Total
Hexavalent Chromium:
Aqueous
Solid
2g
100
2.5 g
200 g
400
100 g
Mercury:
Aqueous
Total
Solid
Dissolved
Total
100
100
0.2 g
400
400
200 g
6 months
24 hours
Store at 4°± 2°C
until analyzed
One month
to extraction, 4 days
after extraction
Store at 4°±2eC
until analyzed
HNO, to pH <2
28 days
Filter;
HNOs to pH <2
28 days
28 days
Store at 4 °± 2° C
until analyzed
Unless stated otherwise.
Either glass or plastic containers may be used.
Any sample volume reduction from the reference method's instructions must be made in tfie exact
proportion as described In the method and representative sampling must be maintained.
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TABLE 3-2
EXAMPLES OF THE ANALYTICAL BLANK INFLUENCE
ON TRACE ANALYSIS OF ELEMENTS IN GLASS (Ref. 3)
CONDITIONS
Pb (ng)
Ag (ng)
Initial analysis of TEG* standard
Analysis using sub-boiled distilled acids
Analysis in class 100 hood
330±250
260±200
20±8
Analysis using sub-boiled acids in class 100 hood 2±1
970±500
207±200
3±2
TEG - Trace Element in glass, SUMS 610 - 619, ± s.
TABLE 3-3
EXAMPLES OF LEAD CONCENTRATIONS IN AIR
SITE
Downtown Air, St Louis, MO
Rural Park Air, Southeastern MO
Laboratory Air, NIST, MD
LEAD CONCENTRATION m-3)
18,84 (Ref. 9)
0.77 (Ref. 10)
0.4 (Ref.3)
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TABLE 3-4
CLEANLINESS LEVELS IN FEDERAL STANDARD 209a (Ref. 11)
CLASSA MAXIMUM CONTAMINATION IN WORK AREA (particles ft"3)
100 particles > 0.5 pm
0 particles > 5,0 pm
10,000 particles > 0.5 pm
65 particles > 5.0 jjm
100,000 particles > 0.5 Mm
700 particles > 5.0 pm
AThe standard required laminar-flow equipment to attain this level of cleanliness.
Since measurement of dust particles smaller than 0.5 pm introduces substantial errors, 0.5 pm has
been adopted as the criterion of measurement.
TABLE 3-5
PARTICULATE CONCENTRATIONS IN LABORATORY AIR (Ref. 10)
SITE CONCENTRATION (ua m^
jron Copper Lead Cadmium
Ordinary Laboratory 0.2 0.02 0.4 0.002
Dean Room 0.001 0.002 0.0002 ND
Clean Hood 0.0009 0.007 0.0003 0.0002
ND - Not Detected
100
10,000
100,000
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TABLE 3-6
NON-CONTAMINATING MATERIALS AND SPECIFICATION FOR USE
IN ULTRA-TRACE ANALYSIS AND AS DECOMPOSITION
VESSELS AND SAMPLE CONTAINERS (Ref. 11)
Listed from highest to lowest preference for use in sample containment
Fluoropolymers: PFA*, TFM, TFE*, FEP*, Tefeel*
Quartz-Synthetic
Polyethylene (suitable for storgage only, not for add digestion)
Quartz - Natural
Borosilicate Glass
* Various forms of Teflon®
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FIGURE 1
COMPARISON OF PARTICLE COUNT ANALYSIS, COUNTS VS. PARTICLE SIZE, OF A
CLEAN ROOM AND A STANDARD LABORATORY AT DUQUESNE UNIVERSITY
IN PITTSBURGH, PA (Ref. 1)
6 10
- 510
3
O
U
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FIGURE 2
PARTICLE SIZE COMPARISON CHART FOR COMMON PARTICULATES (RefS. 1, 12)
Scanning Electron Microscope
Optical Microscope
Visible to
Naked Eye
Ionic
Range
Molecular Range
Macro Molecular Range
Micro Particle Range
Macro
Particle
Ranee
small
organic
molecules
(sugars)
sea Sc It nuclei'
tobaccc smoke
carbon black
virus
fly ash
bacteria
paint pig nent
human
hair.
pollens
cement dust
furhes
millet L flour
coal
oil smoke
metallurj ;ical dus^ and fun
red
blood
cells
es
dust
photochi imical smog
i tisecticide dusts
0.001 0.01 0.10 0.5 1.0 10.0 100.0
Average particle diameter (jim)
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3.2 SAMPLE DIGESTION METHODS
The methods in SW-846 for sample digestion or dissolution are as follows1:
Method 3005 prepares ground water and surface water samples for total recoverable and
dissolved metal determinations by FLAA, ICP-AES, or ICP-MS. Hie unfiltered or filtered sample is
heated with dilute HCl and HN03 prior to metal determination.
Method 3010 prepares waste samples for total recoverable metal determinations by FLAA, ICP-
AES, or ICP-MS. The samples are vigorously digested with nitric add followed by dilution with
hydrochloric acid. The method is applicable to aqueous samples, leachates, and mobility-procedure
extracts.
Method 3015 prepares aqueous samples, mobility-procedure extracts, and wastes that contain
suspended solids for total recoverable metal determinations by FLAA, GFAA, ICP-AES, or ICP-MS.
Nitric add and hydrochloric acid are added to the sample in a Teflon digestion vessel and heated
in a microwave unit prior to metals determination.
Method 3020 prepares waste samples for total recoverable metals determinations by furnace
GFAA or ICP-MS. The samples are vigorously digested with nitric acid followed by dilution with nitric
add. The method is applicable to aqueous samples, leachates, and mobility-procedure extracts.
Method 3031 prepares waste oils, oil sludges, tars, waxes, paints, paint sludges and other
viscous petroleum products for analysis by FLAA, GFAA, and ICP-AES. The samples are vigorously
digested with nitric acid, sulfuric add, hydrochloric add, and potassium permanganate prior to
analysis.
Method 3040 prepares oily waste samples for determination of soluble metals by FLAA, and ICP-
AES methods. The samples are dissolved and diluted in organic solvent prior to analysis. The
method is applicable to the organic extract in the oily waste EP procedure and other samples high
in oil, grease, or wax content.
Method 3050 prepares waste samples for total recoverable metals determinations by FLAA and
ICP-AES, or GFAA and ICP-MS depending on the options chosen. The samples are vigorously
digested in nitric add and hydrogen peroxide followed by dilution with either nitric or hydrochloric
add. The method is applicable to soils, sludges, and solid waste samples.
Method 3051 prepares sludges, sediments, soils and oils for total recoverable metal
determinations by FLAA, GFAA, ICP-AES or ICP-MS. Nitric acid and hydrochloric add are added
to the representative sample in a fluorocarbon digestion vessel and heated in a microwave unit prior
to metals determination.
Method 3052 prepares siliceous and organically based matrices induding ash, biological tissue,
oil, oil contaminated soil, sediment, sludge, and soil for total analysis by FLAA, CVAA, GFAA, ICP-
AES, and ICP-MS. Nitric add and hydrofluoric add are added to a representative sample in a
fluorocarbon digestion vessel and heated in a microwave unit prior to analysis.
1 Please note that chlorine is an interferant in ICP-MS analyses and its use should be
discouraged except when absolutely necessary.
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Method 3060 prepares soils, sludges, sediments and similar waste materials for hexavalent
chromium determination. The samples are digested and heated to dissolve the Cr(VI) and stabilize
it against reduction to Cr(IH).
Prior to employing the above methods in this chapter, analysts are advised to consult the
disclamer statement at the front of this manual and the information In Chapter Two for guidance on
the allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to be
used by the analyst and the regulated community in making judgements necessary to meet the data
quality objectives or needs for the intended use of the data.
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METHOD 3015A
MICROWAVE ASSISTED ACID DIGESTION OF
AQUEOUS SAMPLES AND EXTRACTS
1.0 SCOPE AND APPLICATION
1.1 This microwave method is designed to perform extraction using microwave heating with
nitric acid (HNO3), or alternatively, nitric acid and hydrochloric acid (HCI). Since this method is not
intended to accomplish total decomposition of the sample, the extracted analyte concentrations may
not reflect the total content in the sample. This method is applicable to the microwave-assisted add
extraction/dissolution of available metals in aqueous samples, drinking water, mobility-procedure
extracts, and wastes that contain suspended solids for the following elements:
aChemical Abstract Service Registry Number
'Elements which typically require the addition of HO for optimum recoveries. Other
elements and matrices may be analyzed by this method if performance js
demonstrated for the analyte of interest, in the matrices of interest, at the
concentration levels of interest (see Sec. 9.0).
Element
CASRN"
Aluminum (Al)
Antimony (Sb)
Arsenic (As)
Barium (Ba)
Beryllium (Be)
Boron (B)
Cadmium (Cd)
Calcium (Ca)
Chromium (Cr)
Cobalt (Co)
Copper (Cu)
Iron (Fe)
Lead (Pb)
Magnesium (Mg)
Manganese (Mn)
Mercury (Hg)
Molybdenum (Mo)
Nickel (Ni)
Potassium (K)
Selenium (Se)
Silver (Ag)
Sodium (Na)
Strontium (Sr)
Thallium (TI)
Vanadium (V)
Zinc (Zn)
7429-90-5*
7440-36-0*
7440-38-2
7440-39-3*
7440-41-7*
7440-42-8
7440-43-9
7440-70-2
7440-47-3*
7440-48-4
7440-50-8
7439-89-6*
7439-92-1
7439-95-4*
7439-96-5
7439-97-6
7439-98-7
7440-02-0
7440-09-7
7782-49-2
7440-22-4*
7440-23-5
7440-24-6
7440-28-0
7440-62-2*
7440-66-6
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1.2 This method provides options for improving the performance for certain analytes, such
as antimony, iron, aluminum, and silver by the addition of hydrochloric add, when necessary. It is
intended to provide a rapid multi-element acid extraction prior to analysis so that decisions can be
made about materials and site clean-up levels, and as an estimate of metal toxicity. Digests
produced by the method are suitable for analysis by inductively coupled plasma mass spectrometry
(ICP-MS), inductively coupled plasma atomic emission spectrometry (ICP-AES), flame atomic
absorption spectrophotometry (FLAA), and graphite furnace atomic absorption spectrophotometry
(GFAA). However, the addition of HQ may limit the methods of detection, or increase the difficulties
of detection with some techniques.
Due to the rapid advances in microwave technology, consuN the manufacturer's recommended
instructions for guidance on their microwave digestion system. This method is generic and may be
implemented using a wide variety of laboratory microwave equipment.
2.0 SUMMARY OF METHOD
2.1 A representative 45 mL aqueous sample is extracted in 5 mL concentrated nitric acid
or, optionally, 4 mL concentrated nitric acid and 1 mL concentrated hydrochloric acid, for 20 minutes
using microwave heating with a suitable laboratory microwave unit. The temperature of the acid-
sample mixture is brought to 170 ± 5 °C in 10 minutes, and maintained at 170 ± 5 °C for 10 minutes
to accelerate the leaching process. The sample and acid(s) are placed in a fluorocarbon polymer
(such as PFA or TFM) or quartz microwave vessel or vessel liner. The vessel is sealed and heated
in the microwave unit. After cooling, the vessel contents are filtered, centrifuged, or allowed to settle
and then diluted to volume and analyzed by the appropriate determinative method.
3.0 DEFINITIONS
Refer to Chapter One and Chapter Three for a listing of applicable definitions.
4.0 INTERFERENCES
4.1 Digestion of samples which contain organics will create high pressures due to the
evolution of gaseous digestion products. This may cause venting of the vessels with potential loss
of sample components and/or analytes. When warranted by the potential reactivity of the sample,
a smaller sample size may be used, and the concentration for final calculations adjusted, but the final
water volume prior to addition of acid(s) is recommended to be 45 mL. This is recommended in
order to retain the heat characteristics of the calibration procedure if used. Variations of the method,
due to very reactive materials, are specifically addressed in Section 11.3.3. Limits of quantitation
mil change with sample quantity (dilution) and with instrumentation.
4.2 Many samples can be dissolved by this method. However, when the sample contains
suspended solids which are made up of refractory compounds, such as silicon dioxide, titanium
dioxide, alumina, and other oxides, they will not be dissolved and in some cases may sequester
target analyte elements. These bound elements are considered nonmobile in the environment and
are excluded from most aqueous pollutant transport mechanisms.
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5.0 SAFETY
5.1 The microwave unit cavity must be corrosion resistant and well ventilated. All
electronics must be well protected against corrosion for safe operation.
CAUTION: There are many safety and operational recommendations specific to the model and
manufacturer of the microwave equipment used in individual laboratories. A listing of these
specific suggestions is beyond the scope of this method. The analyst is advised to consult the
equipment manual, the equipment manufacturer, and other appropriate literature for proper and
safe operation of the microwave equipment and vessels. For further details and safety
literature, references 1,7 and 8 review methods and safety in microwave sample preparation.
5.2 The method requires microwave transparent and reagent resistant materials such as
fluorocarbon polymers (examples are PFA and TFM) or quartz to contain acids and samples. For
higher pressure capabilities, the vessel may be contained within layers of different microwave
transparent materials for strength, durability, and safety. The internal volume of the vessel should
be at least 100 mL, and the vessel must be capable of withstanding pressures of at least 30 atm
(435 psi), and capable of controlled pressure relief. These specifications are to provide an
appropriate, safe, and durable reaction vessel of which there are many adequate designs by many
suppliers.
CAUTION: The outer layers of vessels are frequently not as acid or reagent resistant as the
liner material. In order to retain the specified performance and safety requirements, these
outer layers must not be chemically degraded or physically damaged. Routine examination of
the vessel materials is necessary to ensure their safe use.
CAUTION: Another safety concern relates to the use of sealed containers without pressure
relief devices. Temperature is the important variable controlling the reaction. Pressure is
needed to attain elevated temperatures, but must be safely contained. Some digestion vessels
constructed from certain fluorocarborts may crack, burst, or explode in the unit under certain
pressures. Only fluorocarbon (such as PFA, TFM, and others) or quartz containers with
pressure relief mechanisms or containers with fluorocarbon or quartz liners and pressure relief
mechanisms are considered acceptable.
CAUTION: An aqueous sample must contain no more than 1% (V/V or g/V) oxidizable organic
material. Upon oxidation, organic material, whether liquid or solid, contributes to gaseous
digestion products. Pressure build-up above the pressure limit will result in venting of the
closed digestion vessel.
CAUTION: Laboratories should not use domestic (kitchen) type microwave ovens for this
method because of significant safety issues. When adds such as nitric and hydrochloric are
used to effect sample digestion in microwave units in open or sealed vessel(s), there is the
potential for add vapors released to corrode the safety devices that prevent the microwave
magnetron from shutting off when the door is opened. This can result in operator exposure
to microwave energy. Use of a system with isolated and corrosion resistant instrument
components and safety devices prevents this from occurring.
Users are advised not to use domestic (kitchen) type microwave ovens or sealed containers
which are not equipped with controlled pressure relief mechanisms for microwave add digestions
by this method. Use of laboratory-grade microwave equipment is required to prevent safety hazards.
For further details, consult references 1, 7, and 8.
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6.0 EQUIPMENT AND SUPPLIES
6.1 Microwave apparatus requirements
6.1.1 Hie temperature performance requirements necessitate the microwave
decomposition system to sense the temperature to within ± 2.5 °C and automatically adjust
the microwave field output power within 2 seconds of sensing. Temperature sensors should
be accurate to ± 2 °C (including the final reaction temperature of 170 ± 5 °C). Temperature
feedback control provides the primary performance mechanism for the method. Due to the
variability in sample matrix types and microwave digestion equipment (i.e., different vessel
types and microwave oven designs), temperature feedback control is preferred for reproducible
microwave heating. For further details, consult reference 7,
Alternatively, for a specific vessel type, specific set of reagent(s), and sample type, a
calibration control mechanism can be developed similar to those described in previous.
microwave methods (See EPA Method 3051). Through calibration of the microwave power
for a specific number and type of vessel, vessel load, and heat toss characteristics of a specific
vessel series, the reaction temperature profile described in Section 11.3.5 can be reproduced
(Reference 7). The calibration settings are specific for the number and type of vessel and
microwave system being used, in addition to the specific reagent combination being used.
Therefore, no specific calibration settings are provided in this method. These settings may be
developed by using temperature monitoring equipment for each specific set of microwave
equipment and vessel type. They may be used as previously described in EPA Methods 3052
and 3051. In this circumstance, the microwave system provides programmable power, which
can be programmed to within ± 12 W of the required power. Typical systems provide 600 W -
1200 W of power. Calibration control provides backward compatibility with older laboratory
microwave systems which may not be equipped for temperature monitoring or feedback control
and with lower cost microwave systems for some repetitive analyses. Older vessels with lower
pressure capabilities may not be compatible (References 4 - 8).
6.1.2 The accuracy of the temperature measurement system should be periodically
validated at an elevated temperature. This can be done using a container of silicon oil (a high
temperature oil) and an external, calibrated temperature measurement system. The oil should
be adequately stirred to ensure a homogeneous temperature, and both the microwave
temperature sensor and the external temperature sensor placed into the oil. After heating the
oil to a constant temperature of 170 ± 5°C, the temperature should be measured using both
sensors. If the measured temperatures vary by more than 1 to 2°C, the microwave
temperature measurement system should be calibrated. Consult the microwave
manufacturer's instructions about the specific temperature sensor calibration procedure (see
EPA Method 3052).
6.1.3 A rotating turntable is employed to ensure the homogeneous distribution of
microwave radiation within the unit. The speed of the tumtabie should be a minimum of 3 rpm.
Other types of equipment that are used to assist in achieving uniformity of the microwave field
may also be appropriate.
6.2 Class A or appropriate mechanical pipette, volumetric flask, or graduated cylinder, 50
or 100 mL capacity or equivalent.
6.3 Filter paper, qualitative or equivalent.
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6.4 Filler funnel, glass, polypropylene, or other appropriate material.
6.5 Analytical balance, of appropriate capacity and resolution, meeting data qualify
objectives.
7.0 REAGENTS
7.1 All acids should be sub-boiling distilled and/or high purity where possible to minimize
blank levels due to metallic contamination. Other grades may be used, provided it is first ascertained
that the reagent is of sufficient purity to permit its use without decreasing the accuracy of the
determination. If the purify of a reagent is questionable, the reagent should be analyzed to determine
the level of impurities. The reagent blank must be less than the MDL in order to be used.
7.1.1 Concentrated nitric acid (HNOj). The acid should be analyzed to determine
levels of impurity. If the method blank is less than the MDL, the acid can be used.
7.1.2 Concentrated hydrochloric acid (HCI). The add should be analyzed to
determine levels of impurity. If the method blank is less than the MDL, the acid can be used.
7.2 Reagent Water. Reagent water shall be interference free. All references to water in
the method refer to reagent water unless otherwise specified. For further details, consult Reference
2.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 All samples must have been collected using a sampling plan that addresses the
considerations discussed in Chapter Nine of SW-846. Refer to that chapter, as updated, for
guidance.
8.2 All sample containers must be prewashed with acids, water, and metal-free detergents,
if necessary, depending on the use history of the container (Reference 7). Plastic and glass
containers are both suitable. For further information, see Chapter Three.
8.3 Aqueous waste waters must be acidified to a pH < 2 with HN03.
9.0 QUALITY CONTROL
9.1 All quality control data must be maintained and available for reference or inspection for
a period of three years. This method is restricted to use by, or under supervision of, experienced
analysts.
9.2 Duplicate samples should be processed on a routine basis. A duplicate sample is a
sample brought through the whole sample preparation and analysis process. A duplicate sample
should be processed with each analytical batch or every 20 samples, whichever is the greater
number. A duplicate sample should be prepared for each matrix type (i.e., wastewaters, extracts,
etc.).
9.3 Spiked samples or standard reference materials should be included with each group
of samples processed, or every 20 samples, whichever is the greater number. A spiked sample
should also be included whenever a new sample matrix is being analyzed.
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9.4 Periodically, the accuracy of the temperature measurement system used to control the
microwave equipment should be validated per Section 6.1.2.
9.5 (Not necessary if using temperature feedback control.) Each day that samples are
extracted, the microwave-power calibration should be verified by heating 1 kg of ASTM Type II water
(at 22 ± 3 °C) in a covered, microwave-transparent vessel for 2 mifi at the setting for 490 W and
measuring the water temperature after heating per Section 10.1.5. if the power calculated (per
Section 12) differs from 490 W by more than ± 10 W, the microwave settings should be recalibrated
per Section 10.0.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Calibration of Microwave Equipment
NOTE: If the microwave unit uses temperature feedback control to control the performance
specifications of the method, then performing the calibration procedure is not necessary.
10.1.1 Calibration is the normalization and reproduction of a microwave field strength
to permit reagent and energy coupling in a predictable and reproducible manner. It balances
reagent heating and heat loss from the vessels and is equipment dependent due to the heat
retention and loss characteristics of the specific vessel. Available power is evaluated to permit
the microwave field output in watts to be transferred from one microwave system to another.
Use of calibration to control this reaction requires balancing output power, coupled
energy, and heat loss to reproduce the temperature heating profile given in section 11.3.5.
The conditions for each acid mixture and each batch containing the same specified number
of vessels must be determined individually. Only identical add mixtures and vessel models
and specified numbers of vessels may be used in a given batch.
10.1.2 For cavity type microwave equipment, calibration is accomplished by
measuring the temperature rise in 1 kg of water exposed to microwave radiation for a fixed
period of time. The analyst can relate power in watts to the partial power setting of the system.
The calibration format required for laboratory microwave systems depends on the type of
electronic system used by the manufacturer to provide partial microwave power. Few systems
have an accurate and precise linear relationship between percent power settings and absorbed
power. Where linear circuits have been utilized, the calibration curve can be determined by
a three-point calibration method (see Section 10.1.4). Otherwise, the analyst must use the
multiple point calibration method (see Section 10.1.3). Assistance in calibration and software
guidance of calibration are available in References 7 and 8.
10.1.3 Multiple point calibration involves the measurement of absorbed power over
a large range of power settings. Typically, for a 600 W unit, the following power settings are
measured: 100,99,98,97,95,90, 80, 70,60, 50, and 40% using the procedure described in
Section 10.1.5. This data is clustered about the customary working power ranges.
Nonlinearity has been encountered at the upper end of the calibration. If the system's
electronics are known to have nonlinear deviations in any region of proportional power control,
it will be necessary to make a set of measurements that bracket the power to be used. The
final calibration point should be at the partial power setting that will be used in the test. This
setting should be checked periodically to evaluate the integrity of the calibration. If a significant
change is detected (± 10 W), then the entire calibration should be re-evaluated.
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10.1.4 The three-point calibration involves the measurement of absorbed power at
three different power settings. Power is measured at 100% and 50% using the procedure
described in Section 10.1.5. From this 2-point line, determine the partial power setting that
corresponds to the power, in watte, specified in the procedure to reproduce the heating profile
specified in Section 11.3.6. Measure the absorbed power at that partial power setting. If the
measured absorbed power does not correspond to the specified power within ±10 W, use the
multiple point calibration in Section 10.1.3. This point should also be used to periodically verify
the integrity of the calibration.
10.1.5 Equilibrate a large volume of water to room temperature (22 ± 3 "C). One kg
of reagent water is weighed (1,000.0 ± 0.1 g) into a fluorocarbon beaker or a beaker made of
some other material that does not significantly absorb microwave energy (glass absorbs
microwave energy and is not recommended). The initial temperature of the water should be
22 i 3 °C measured to ± 0.05 °C. The covered beaker is circulated continuously (in the
normal sample path) through the microwave field for 2 minutes at the desired partial power
setting with the system's exhaust fan on maximum (as it will be during normal operation). The
beaker is removed and the water is vigorously stirred. Use a magnetic stirring bar inserted
immediately after microwave irradiation (irradiating with the stir bar in the vessel could cause
electrical arcing) and record the maximum temperature within the first 30 seconds to ± 0.05
°C. Use a new sample for each additional measurement. If the water is reused (after making
adjustments for any loss of weight due to heating), both the water and the beaker must have
returned to 22 ± 3 °C. Three measurements at each power setting should be made.
The absorbed power is determined by the following relationship:
Eq"1 p (K)(Cp)(m)(AT)
t
Where:
P = the apparent power absorbed by the sample in watts (W) (joule sec"1)
K = the conversion factor for thermochemical calories sec1 to watts (K= 4.184)
Cp = the heat capacity, thermal capacity, or specific heat (cal g"1 °C*1) of water
m - the mass of the water sample in grams (g)
AT = the final temperature minus the initial temperature (°C)
t = the time in seconds (s)
Using the experimental conditions of 2 minutes (120 sec) and 1 kg (1000 g) of distilled
water (heat capacity at 25 °C is 0.9997 cal g"1 °C1), the calibration equation simplifies to:
Eq. 2 p = (ATX34.86)
NOTE: Stable line voltage is necessary for accurate and reproducible calibration and operation.
The line voltage should be within manufacturer's specification. During measurement and
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operation the line voltage should not vary by more than ± 2 V (Reference 7). Electronic
components in most microwave units are matched to the system's function and output. When
any part of the high voltage circuit, power source, or control components in the system have
been serviced or replaced, it will be necessary to recheck the system's calibration. If the power
output has changed significantly (± 10 W), then the entire calibration should be re-evaluated.
11.0 SAMPLE PROCEDURE
11.1 Temperature control of closed vessel microwave instruments provides the main
feedback control performance mechanism for the method. Method control requires a temperature
sensor in one or more vessels during the entire digestion. The microwave decomposition system
should sense the temperature to within ± 2.5 °C and permit adjustment of the microwave output
power within 2 seconds.
11.2 All digestion vessels and volumetric ware must be carefully acid washed and rinsed
with reagent water. When switching between highly concentrated samples and low concentrated
samples, all digestion vessels (fluoropolymer or quartz liners) should be cleaned by leaching with
hot (1:1) hydrochloric add (greater than 80 °C, but less than boiling) for a minimum of two hours
followed by hot (1:1) nitric add (greater than 80 °C, but less than boiling) for a minimum of two
hours. The vessels should then be rinsed with reagent water and dried in a dean environment. This
deaning procedure should also be used whenever the prior use of the digestion vessels is unknown
or cross contamination from prior sample digestions in vessels is suspected. Polymeric or glass
volumetric ware and storage containers should be deaned by leaching with more dilute adds
(approximately 10% V/V) appropriate for the specific material used and then rinsed with reagent
water and dried in a dean environment.
11.3 Sample Digestion
11.3.1 Measure a 45 mL aliquot of a well-shaken, homogenized sample using an
appropriate volumetric measurement and delivery device, and quantitatively transfer the aliquot
to an appropriate vessel equipped with a controlled pressure relief mechanism.
11.3.2 Add 5 ± 0.1 mL concentrated nitric add or, alternatively, 4 ± 0.1 mL
concentrated nitric add and 1 ± 0.1 mL concentrated hydrochloric add to the vessel in a fume
hood (or fume exhausted endosure). The addition of concentrated hydrochloric add to the
nitric add is appropriate for the stabilization of certain analytes, such as Ag, Ba, and Sb and
high concentrations of Fe and A1 in solution. Improvements and optimal recoveries of antimony
and silver upon addition of HCI have been described in the literature (Reference 7). The
addition of hydrochloric add may, however, limit the detection techniques or increase the
difficulties of analysis for some detection systems.
QAI/PQN: The addition of hydrochloric add must be in the form of concentrated
hydrochloric add and not from a premixed combination of adds. A build-up of chlorine
gas, as well as other gases, will result from a premixed add solution. These gases
may be violently released upon heating. This is avoided by adding the acid in the
described manner.
CAUTION: Toxic nitrogen oxide(s) and chlorine fumes are usually produced during
digestion. Therefore, all steps involving open or the opening of microwave vessels
must be performed in a properly operating fume ventilation system.
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CAUTION: The analyst should wear protective gloves and face protection.
CAUTION: The use of microwave equipment with temperature feedback control is
required to control any unfamiliar reactions that may occur during the leaching of
samples of unknown composition. The leaching of these samples may require
additional vessel requirements such as increased pressure capabilities.
11.3.3 The analyst should be aware of the potential for a vigorous reaction,
especially with samples containing suspended solids composed of volatile or easily oxidized
organic species. When digesting a matrix of this type, if a vigorous reaction occurs upon the
addition of reagent(s), this sample represents a safety hazard. Do not leach the sample as
described in this method due to the high potential for unsafe and uncontrollable reactions.
11.3.4 Seal the vessel according to the manufacturer's directions. Property place
the vessel in the microwave system according to the manufacturer's recommended
specifications and, when applicable, connect appropriate temperature and pressure monitoring
equipment to vessels according to manufacturer's specifications.
11.3.5 This method is a performance based method, designed to achieve or
approach consistent leaching of the sample through achieving specific reaction conditions.
The temperature of each sample should rise to 170 ± 5 °C in approximately 10 minutes and
remain at 170 ±5 °C for 10 minutes, or for the remainder of the twenty-minute digestion period
(References 3, 4, 6, and 7). The time vs. temperature and pressure profiles for the leaching
of three simulated wastewater samples using Method 3015 are shown in Figure 1. The
samples are composed of approximately 0.35 g SRM 2704 (Buffalo River Sediment) mixed in
45 mL double-deionized water. The figure demonstrates the temperature and pressure profiles
for both the all-nitric digest (5 mL concentrated nitric acid), and the nitric and hydrochloric
mixed-acid digest (4 mL concentrated nitric acid and 1 mL concentrated hydrochloric acid).
Also shown is the profile for the heating of the wastewater sample without addition of adds.
When using temperature feedback control, the number of samples that may be simultaneously
digested may vary, from one sample up to the maximum number of vessels that can be
heated by the magnetron of the microwave unit according to the heating profile specified in this
section. (The number will depend on the power of the unit, the number of vessels, and the
heat loss characteristics of the vessels (Reference 7)).
11.3.5.1 Calibration control is applicable in reproducing this method
provided the power in watts versus time parameters are determined to reproduce the
specifications listed in Section 11.3.5. The calibration settings will be specific to the
quantity of reagents, the number of vessels, and the heat loss characteristics of the
vessels (Reference 7). If calibration control is being used, any vessels containing
acids for analytical blank purposes are counted as sample vessels. When fewer than
the recommended number of samples are to be digested, the remaning vessels
should be filled with 45 mL water, and the acid mixture added, so that the full
complement of vessels is achieved. This provides an energy balance, since the
microwave power absorbed is proportional to the total absorbing mass in the cavity
(Reference 7). Irradiate each group of vessels using the predetermined calibration
settings. (Different vessel types should not be mixed.)
11.3.6 At the end of the microwave program, allow the vessels to cool for a minimum
of 5 minutes before removing them from the microwave system. Cooling of the vessels may
be accelerated by internal or external cooling devices. When the vessels have cooled to
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near room temperature, determine if the microwave vessels have maintained their seal
throughout the digestion. Due to the wide variability of vessel designs, a single procedure
is not appropriate. For vessels that are sealed as discrete separate entities, the vessel
weight may be taken before and after digestion to evaluate seal integrity. If the weight loss
of the sample exceeds 1 % of the weight of the sample and reagents, then the sample is
considered compromised. For vessels with burst disks, a careful visual inspection of the
disk, in addition to weighing, may identify compromised vessels. For vessels with reseating
pressure relief mechanisms, an auditory or a physical sign that can indicate whether a vessel
has vented is appropriate.
11.3.7 Complete the preparation of the sample by carefully uncapping and venting
each vessel in a chemical fume hood (or fume exhausted enclosure). Vent the vessels using
the procedure recommended by the vessel manufacturer. Quantitatively transfer the sample
to an acid-cleaned bottle. If the digested sample contains particulates which may clog
nebulizers or interfere with injection of the sample into the Instalment, the sample may be
centrifuged (Section 11.3.7.1), allowed to settle (Section 11.3.7.2), or filtered (Section
11.3.7.3).
11.3.7.1 Centrifugation: Centrifugation at 2,000 - 3,000 rpm for 10
minutes is usually sufficient to clear the supernatant.
11.3.7.2 Settling: If undissolved material, such as Si02, Ti02, or other
refractory oxides, remains, allow the sample to stand until the supernatant is clear.
Allowing a sample to stand overnight will usually accomplish this. If it does not,
centrifuge or filter the sample.
11.3.7.3 Filtering: If necessary, the filtering apparatus must be
thoroughly cleaned and prerinsed with dilute (approximately 10% V/V) nitric acid.
Filter the sample through qualitative filter paper into a second acid-cleaned container.
11.3.8 The removal or reduction of the quantity of sample may be desirable fa*
concentration of analytes prior to analysis. The chemistry and volatility of the anaiytes of
interest should be considered and evaluated when using this alternative (Reference 7, 8).
Sample evaporation in a controlled environment with controlled purge gas and neutralizing
and collection of exhaust interactions is an alternative where appropriate. This manipulation
may be performed in the microwave system, if the system is capable of this function, or
external to the microwave system in more common apparatus(s). This option must be tested
and validated to determine analyte retention and loss and should be accompanied by
equipment validation possibly using the standard addition method and standard reference
materials. For further information, see References 7 and 8 and Method 3052.
NOTE: The final solution typically requires nitric acid to maintain appropriate sample
solution acidity and stability of the elements. Commonly, a 2% (v/v) nitric acid
concentration is desirable. Waste minimization techniques should be used to capture
reagent fumes. This procedure should be tested and validated in the apparatus and
on standards before using on unknown samples.
11.3.9 Transfer or decant the sample into volumetric ware and dilute the digest to
a known volume. The digest is now ready for analysis for elements of interest using
appropriate elemental analysis techniques and/or SW-846 methods.
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12.0 DATA ANALYSIS AND CALCULATIONS
12.1 Calculations: The concentrations determined are to be reported on the basis of the
actual volume of the original sample.
12.2 Prior to use of the method, verify that the temperature sensing equipment is properly
reading temperature. A procedure for verification is given in Section 6.1.2. This will establish the
accuracy and precision of the temperature sensing equipment, which should be carried throughout
the statistical treatment of the quality assurance data.
12.3 In calibrating the microwave unit (Section 10.0), the power absorbed (for each power
setting) by 1 kg of reagent water exposed to 120 seconds of microwave energy is determined by the
expression:
Power (in watts) = - T2) (34.86)
where: T, = Initial temperature of water (between 21 and 25 °C to nearest 0.1 °C)
T2 = Final temperature of water (to nearest 0.1 °C)
12.4 Plot the power settings against the absorbed power (calculated in Section 12.3) to
obtain a calibration relationship. Alternatively, use a microwave calibration program to analyze the
calibration data (References 7 and 8). Interpolate the data to obtain the instrument settings needed
to provide the wattage levels specified in Section 12.3.
13.0 METHOD PERFORMANCE
13.1 The fundamental analytical validation of Method 3015 with nitric add has been
performed (Reference 6). The results are shown in Table 1. Variations of 3015 including nitric acid
and hydrochloric acid have also been published in the literature (References 5, 7, 9). The method
has also been tested on a variety of matrices, including two simulated wastewater matrices, one
consisting of ~ 0.35 g sediment (SRM 2704) mixed with 45 mL double-deionized water, and the other
consisting of ~ 0.35 g soil (SRM 4355) mixed with 45 mL double-deionized water. The results are
shown in Tables 2 and 3, and are published in the literature (Reference 9).
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the
quantity or toxicity of waste at the point of generation. Numerous opportunities for pollution
prevention exist in laboratory operations. The EPA has established a preferred hierarchy of
environmental management techniques that places pollution prevention as the management option
of first choice. Whenever feasible, laboratory personnel should use pollution prevention techniques
to address their waste generation. When wastes cannot be feasibly reduced at the source, the
Agency recommends recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions, consult Less is Better: Laboratory Chemical Management for Waste Reduction.
available from the American Chemical Society's Department of Government Relations and Science
Policy, 1155 16th Street, NW, Washington, DC 20036, (202) 872-4477.
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15.0 WASTE MANAGEMENT
15,1 The Environmental Protection Agency requires that laboratory waste management
practices be consistent with all applicable rules and regulations. The Agency urges laboratories to
protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel, available from the American
Chemical Society's Department of Government Relations and Science Policy, 1155 16th Street, NW,
Washington, DC 20036, (202) 872-4477.
16.0 REFERENCES
1. Kingston, H.M.; Jassie, L.B. In Introduction to Microwave Sample Preparation: Theory and
Practice: Kingston, H. M. and Jassie, L. B., Eds.; ACS Professional Reference Book Series;
American Chemical Society: Washington, DC, 1988; Chapters 6 and 11.
2. 1985 Annual Book of ASTM Standards. Vol. 11.01; "Standard Specification for Reagent
Water*; ASTM, Philadelphia, PA, 1985, D1193-77.
3. Kingston, H. M., Final Report EPA IAG #DWI3932541-01-I, September 30,1988, Appendix
A.
4. Shannon, M., Alternate Test Procedure Application, USEPA Region V, Central Regional
Laboratory, 536 S. Clark Street, Chicago, IL 60606,1989.
5. Kingston, H. M., Walter, P. J,, "Comparison of Microwave Versus Conventional Dissolution
for Environmental Applications", Spectroscopy, Vol. 7 No. 9, 20-27,1992.
6. Sosinski, P., and Sze, C., "Absolute Accuracy Study, Microwave Digestion Method 3015
(nitric add only)*; EPA Region III Central Regional Laboratory, 1991.
7. Kingston, H. M., Haswell, S. J., Eds. Microwave Enhanced Chemistry: Fundamentals.
Sample Preparation, and Applications: ACS Professional Reference Book Series; American
Chemical Society: Washington, DC 1997.
8. Duquesne University. Analytical Sample Preparation and Microwave Chemistry Center.
Sampl&Pt&p Web. Access http://www.sampleprep.duq.edu/
9. Link, D.D., Kingston, H.M., Walter, P.J., "Development and Validation of the New EPA
Microwave-Assisted Leach Methods 3051A and 3015A.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 through 3, Figure 1, and a flow diagram of method
procedure.
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TABLE 1
RESULTS OF VALIDATION STUDY FOR METHOD 3015 (NITRIC ONLY)
(REFERENCE 6)
Element
TM-11
TM-12
T-95
T-11
57
T-1C
8
Mean
Std
Dev
Mean
Std
Dev
Mean
Std
Dev
Mean
Std
Dev
Mean
Std
Dev
Al
480
26
2800
88.

210
19
120
31
As

13
1
90
11
Ba
140
23
2400
70

200
16

11.3
0.5
26
1
Ca

12000
783
59000
999
Cd
45
2
240
8

12
1
10
2
Co
240
14
1150
36

Cr
64
4
350
10

23
1
30
6
Cu
78
4
320
9

42
4
34
4
Fe
290
16
1180
43

60
9
130
7
K

5000
784

2600
383
Mg

35000
1922
2200
110
10200
218
Mn
61
3
300
9

53
3
47
3
Na

20000
10690
2300
1056
13800
516
Ni
280
16
1290
39

61
2
Pb
280
32
1360
35

30.1
0.2
39
1
Se

65.97
2.65
13
1

V
530
26
2400
61

Zn
56
3
520
9

31
3
70
4
Element
WP9C
0#1
WP98
0 #2
WS378 #4
WS37
8 #12
Mean
Std
Dev
Mean
Std
Dev
Mean
Std
Dev
Mean
Std
Dev
Sb
18.0
0.5
110
34

Tl
55
2
7.0
0.5

19
5
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TABLE 2
COMPARISON OF ANALYTE RECOVERIES FROM "SIMULATED WASTEWATER" MIXTURE
OF ~ 0.35 G SRM 2704 (BUFFALO RIVER SEDIMENT) AND 45 ML DOUBLE-DEIONIZED
WATER USING BOTH DIGEST OPTIONS OF METHOD 3015
(REFERENCE 9)
Element
5 itiL HNOj
digest
4 mL HNOj +
1 mL HCI digest
Total Anaiyte
Concentration
Ag
0.31 ± 0.05
0.41 ± 0.09
<4
B
23.8 ± 3.1
30.6 ±8.3
_#
Be
0.81 ±0.13
0.91 ±0.19
*
Co
12.0 ± 0.30
11.5 ±0.98
14.0 ± 0.6
Hg
—
1.49 ±0.03
1.44 ±0.07
Mo
2.97 ±0.72
3.15 ±0.28
~
Ni
39.6 ±2.5
41.3 ±1.7
44.1 ± 3.0
Sr
41.9 ±1.3
49,0 ±1.6
(130)
V
6.18 ±2.5
14.6 ±2.4
95 4 4
Zn
418 ±12
412 ±31
438 ± 12
Results reported in pg/g anaiyte (mean ± 95% confidence limit).
Total concentrations are taken from NIST SRM Certificate of Analysis.
Values in parenthesis are reference concentrations.
* The total concentration of this anaiyte in SRM 2704 is not certified.
3015A- 14
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TABLE 3
COMPARISON OF ANALYTE RECOVERIES FROM "SIMULATED WASTEWATER" MIXTURE
OF -0.35 G SRM 4355 (PERUVIAN SOIL) AND 45 ML DOUBLE-DEIONIZED WATER USING
BOTH DIGEST OPTIONS OF METHOD 3015
(REFERENCE 9)
Element
5 mL HNOj
digest
4 mL HNOj +
1 mL HCI digest
Total Arialyte
Concentration
Ag
1.31 ±0.12
1.62 ± 0.11
(1.9)*
B
32.9 ± 2.1
31.812.7
(63)*
Co
10.5 ± 0.34
10.4 ± 0.41
14.8 ±0.76
Mo
0.99 ±0.06
1.1 ±0.11
(1.7)*
Ni
12.2 ± 1.2
13.1 ± 1.9
(13)*
Pb
135 ± 4
136 ±4
129 ±26
Sb
3.7 ± 0.30
5.2 ± 0.53
14.3 ± 2.2
Sr
140 ±6
143 ±7
(330)
Results reported in pg/g analyte (mean ± 95% confidence limit).
Total concentrations are taken from NiST SRM Certificate of Analysis.
* Values in parenthesis are reference concentrations.
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FIGURE 1
THE TYPICAL TEMPERATURE AND PRESSURE PROFILE FOR THE HEATING OF A
SIMULATED WASTEWATER SAMPLE (~ 0.35 G SRM 2704 + 45 ML DOUBLE-DEIONIZED
WATER) USING BOTH DIGEST OPTIONS (5 ML HN03 AND 4 ML HN03 + 1 ML HCL)
OF METHOD 3015.
temperature
U
0
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METHOD 3015A
MICROWAVE ASSISTED ACID DIGESTION OF AQUEOUS SAMPLES AND EXTRACTS
113.1 Measure a 45 ml aliquot
of sample into ffle digestion vessel.
11.3.2 Add Sto.t mL cone. HNO3
or, alternatively, 4+G.1 mL eene.
HNOg aoo itO.1 mL cone, HCl
Yes
Ves
No
11.3,7,1 - 11 3.7.3
Centrifuge, settle or
fitter samoie.
11.3.4 Seal and place vessels
in mtcro«#av* system.
11.3.6 AIJcw vessels to cool le
mom temperature.
10.1 Calibrate
microwave equipment.
112 Acto wash and water nose
all digestion vessels ana glassware.
12 0 Calculate concemraiics eased
on ordinal sample weigftt.
11.3 5 Use appropriate elemental
analysis techmaues andtar
SW-846 methods.
113.8 Remove or reduce quantity
of sample oy evaporation wrth
controlled Purge gas.
t 1.3.7 Vent each vessel.
Transfer sample to an acid-
eleaned bottle
11 3.5 Heat samples aecoreing to
time vs. temperature and pressure
profiles.
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METHOD 3051A
MICROWAVE ASSISTED ACID DIGESTION OF
SEDIMENTS. SLUDGES. SOILS. AND OILS
1.0 SCOPE AND APPLICATION
1.1 This microwave extraction method is designed to mimic extraction using conventional
heating with nitric acid (HN03), or alternatively, nitric acid and hydrochloric acid (HCI), according to
EPA Methods 200.2 and 3050. Since these methods are not intended to accomplish total
decomposition of the sample, the extracted analyte concentrations may not reflect the total content
in the sample. This method is applicable to the microwave-assisted acid extraction/dissolution* of
sediments, sludges, soils, and oils for the following elements:
"Chemical Abstract Service Registry Number
•Indicates elements which typically require the addition of HCI to achieve equivalent
results with EPA Method 3050, as noted in reference 3.
*Note: For matrices such as certain types of oils, this method may or may not
provide total sample dissolution. For other matrices, such as soils and sediments, it
should be considered an extraction method. Other elements and matrices may be
Aluminum (Al)
Antimony (Sb)
Arsenic (As)
Barium (Ba)
Beryllium (Be)
Boron (B)
Cadmium (Cd)
Calcium (Ca)
Chromium (Cr)
Cobalt (Co)
Copper (Cu)
Iron (Fe)
Lead (Pb)
Magnesium (Mg)
Manganese (Mn)
Mercury (Hg)
Molybdenum (Mo)
Nickel (Ni)
Potassium (K)
Selenium (Se)
Silver (Ag)
Sodium (Na)
Strontium (Sr)
Thallium (Tl)
Vanadium (V)
Zinc (Zn)
Element
CASRN*
7429-90-5*
7440-36-0*
7440-38-2
7440-39-3*
7440-41-7*
7440-42-8
7440-43-9
7440-70-2
7440-47-3*
7440-48-4
7440-50-8
7439-89-6*
7439-92-1
7439-95-4*
7439-96-5
7439-97-6
7439-98-7
7440-02-0
7440-09-7
7782-49-2
7440-22-4*
7440-23-5
7440-24-6
7440-28-0
7440-62-2*
7440-66-6
3051A - 1
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analyzed by this method if performance is demonstrated for the analyte of interest,
in the matrices of interest, at the concentration levels of interest (see Sec. 9.0).
1.2 This method is provided as an alternative to EPA Methods 200.2 and 3050. This
method provides options for improving the performance for certain analytes, such as antimony, iron,
aluminum, and silver by the addition of hydrochloric acid, when necessary. It is intended to provide
a rapid mufti-element add extraction or dissolution prior to analysis so that decisions can be made
about materials and site cleanup levels, the need for TCLP testing of a waste (see EPA Method
1311, Section 1.2, for further details), and whether a BOAT process is providing acceptable
performance. Digests produced by the method are suitable for analysis by flame atomic absorption
spectrophotometry (FLAA), graphite furnace atomic absorption spectrophotometry (GFAA),
inductively coupled plasma atomic emission spectrometry (ICP-AES) and inductively coupled plasma
mass spectrometry (ICP-MS). However, the addition of HCl may limit the methods of detection, or
increase the difficulties of detection with some techniques.
Due to the rapid advances in microwave technology, consult your manufacturer's
recommended instructions for guidance on their microwave digestion system.
2.0 SUMMARY OF METHOD
2.1 A representative sample of up to 0.5 g is extracted and/or dissolved in 10 mL
concentrated nitric add, or alternatively, 9 mL concentrated nitric acid and 3 mL concentrated
hydrochloric add fa* 10 minutes using microwave heating with a suitable laboratory microwave unit.
The sample and add(s) are placed in a fluorocarbon polymer (PFA or TFM) or quartz microwave
vessel or vessel liner. The vessel is sealed and heated in the microwave unit. After cooling, the
vessel contents are filtered, centrifuged, or allowed to settle and then diluted to volume and analyzed
by the appropriate determinative method.
3.0 DEFINITIONS
Please refer to Chapter One and Chapter Three for a listing of applicable definitions.
4.0 INTERFERENCES
4.1 Very reactive samples or volatile materials may create high pressures due to the
evolution of gaseous digestion products. This may cause venting of the vessels with potential loss
of sample and/or analytes. The complete decomposition of either carbonates, or carbon based
samples, may produce enough pressure to vent the vessel if the sample size is greater than 0.25
g (depending on the pressure capability of the vessel). Variations of the method to accommodate
very reactive materials are spedfically addressed in Section 11.3.3.
4.2 Many types of samples will be dissolved by this method. A lew refractory sample matrix
compounds, such as quartz, silicates, titanium dioxide, alumina, and other oxides may not be
dissolved and in some cases may sequester target analyte elements. These bound elements are
considered non-mobile in the environment and are exduded from most aqueous transport
mechanisms of pollution.
5.0 SAFETY
5.1 The microwave unit cavity must be corrosion resistant and well ventilated. All
electronics must be protected against corrosion for safe operation.
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CAUTION: There are many safety and operational recommendations specific to the
model and manufacturer of the microwave equipment used in individual laboratories. A listing
of these specific suggestions is beyond the scope of this method. The analyst is advised to
consult the equipment manual, the equipment manufacturer, and other appropriate literature
for proper and safe operation of the microwave equipment and vessels. For further details,
see reference 3 and the document of Sec. 13.2.1 for a review of safety in microwave sample
preparation.
5.2 The method requires essentially microwave transparent and reagent resistant materials
such as fluorocarbon polymers (examples are PFA or TFM) or quartz to contain acids and samples.
For higher pressure capabilities the vessel may be contained within layers of different microwave
transparent materials for strength, durability, and safety. The internal volume of the vessel should
be at least 45 mL, and the vessel must be capable of withstanding pressures of at least 30 atm (435
psi), and capable of controlled pressure relief. These specifications are to provide an appropriate,
safe, and durable reaction vessel of which there are many adequate designs by many suppliers.
CAUTION: The reagent combination (9 mL nitric acid to 3 mL hydrochloric acid) results
in greater pressures than those resulting from the use of only nitric acid. As demonstrated in
Figures 1 and 2, pressures of approximately 12 atm have been reached during the heating of
the add mixture alone (no sample in the vessel). Pressures reached during the actual
decomposition of a sediment sample (SRM 2704, a matrix with low organic content) have more
than doubled when using the 9 mL nitric and 3 mL hydrochloric acid mixture. These higher
pressures necessitate the use of vessels having higher pressure capabilities (30 atm or 435
psi). Matrices having large organic content, such as oils, can produce approximately 25 atm
of pressure inside the vessel (as described in EPA Method 3052).
CAUTION: The outer layers of vessels are frequently not as acid or reagent resistant as
the liner material. In order to retain the specified performance and safety requirements, these
outer layers must not be chemically degraded or physically damaged. Routine examination of
the vessel materials is necessary to ensure their safe use,
CAUTION: Another safety concern relates to the use of sealed containers without
pressure relief devices. Temperature is the important variable controlling the reaction.
Pressure is needed to attain elevated temperatures, but must be safely contained. Some
digestion vessels constructed from certain fluorocarbons may crack, burst, or explode in the
unit under certain pressures. Only fluorocarbon (such as PFA or TFM and others) or quartz
containers with pressure relief mechanisms or containers with fluorocarbon or quartz liners
and pressure relief mechanisms are considered acceptable.
CAUTION. Laboratories should not use domestic (kitchen) type microwave ovens for this
method because of significant safety issues. When acids such as nitric and hydrochloric are
used to effect sample digestion in microwave units in open vessel(s), or sealed vessel(s), there
is the potential for any released add vapors to corrode the safety devices that prevent the
microwave magnetron from shutting off when the door is opened. This can result in operator
exposure to microwave energy. Use of a system with isolated and corrosion resistant safety
devices prevents this from occurring.
Users are therefore advised not to use domestic (kitchen) type microwave ovens or
sealed containers which are not equipped with controlled pressure relief mechanisms for
microwave acid digestions by this method. Use of laboratory-grade microwave equipment is
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required to prevent safety hazards. For further details, consult reference 3 and the document
listed in Sec. 13.2.1.
6.0 EQUIPMENT AND SUPPLIES
6.1 Microwave apparatus requirements.
6.1.1 The temperature performance requirements necessitate the microwave
decomposition system to sense the temperature to within ± 2.5 °C and automatically adjust
the microwave field output power within 2 seconds of sensing. Temperature sensors should
be accurate to ± 2 °C (including the final reaction temperature of 175 ± 5 °C). Temperature
feedback control provides the primary performance mechanism for the method. Due to the
variability in sample matrix types and microwave digestion equipment (i.e., different vessel
types and microwave oven designs), temperature feedback control is preferred for reproducible
microwave heating. For further details consult reference 3.
Alternatively, for a specific vessel type, specific set of reagent(s), and sample type, a
calibration control mechanism can be developed. Through calibration of the microwave power
for a specific number and type of vessels, vessel load, and heat loss characteristics of a
specific vessel series, the reaction temperature profile described in Sec. 11.3.5 can be
reproduced. The calibration settings are specific for the number and type of vessels and
microwave system being used, in addition to the specific reagent combination being used.
Therefore, no specific calibration settings are provided in this method. These settings may be
developed by using temperature monitoring equipment for each specific set of microwave
equipment and vessel type. They may be used as previously described in methods such as
EPA Methods 3051, 3015, and 3052. In this circumstance, the microwave system provides
programmable power, which can be programmed to within ± 12 W of the required power.
Typical systems provide a nominal 600 W to 1200 W of power. Calibration control provides
backward compatibility with older laboratory microwave systems which may not be equipped
for temperature monitoring or feedback control and with lower cost microwave systems for
some repetitive analyses. Older vessels with lower pressure capabilities may not be
compatible (see refs. 1,2, and 3 and the documents listed in 13.3.3 and 13.3.5).
6.1.2 The accuracy of the temperature measurement system should be periodically
validated at an elevated temperature (see Section 12.2). This can be done using a container
of silicon oil (a high temperature oil) and an external, calibrated temperature measurement
system. The oil should be adequately stirred to ensure a homogeneous temperature, and both
the microwave temperature sensor and the external temperature sensor placed into the oil.
After heating the oil to a constant temperature of 180 ± 5 °C, the temperature should be
measured using both sensors. If the measured temperatures vary by more than 1 to 2 °C, the
microwave temperature measurement system should be calibrated. Consult the microwave
manufacturer's instructions about the specific temperature sensor calibration procedure.
6.1.3 A rotating turntable is employed to ensure the homogeneous distribution of
microwave radiation within the unit. The speed of the turntable should be a minimum of 3 rpm.
Other types of equipment that are used to assist in achieving uniformity of the microwave field
may also be appropriate.
6.2 Volumetric graduated cylinder, 50 or 100 mL capacity or equivalent,
6.3 Filter paper, qualitative or equivalent.
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6.4 Filter funnel, glass, polypropylene, or other appropriate material.
6.5 Analytical balance, of appropriate capacity and resolution meeting data quality
objectives.
7.0 REAGENTS
7.1 All acids should be sub-boiling distilled where possible to minimize the blank levels due
to metallic contamination. Other grades may be used, provided it is first ascertained that the reagent
is of sufficient purity to permit its use without decreasing the accuracy of the determination. If the
purity of a reagent is questionable, the reagent should be analyzed to determine the level of
impurities. The reagent blank must be less than the MDL in order to be used.
7.1.1 Concentrated nitric acid (HN03). The acid should be analyzed to determine
levels of impurity. If the method blank is less than the MDL, the acid can be used.
7.1.2 Concentrated hydrochloric acid (HCI). The acid should be analyzed to
determine levels of impurity. If the method blank is less than the MDL, the acid can be used.
7.2 Reagent Water. Reagent water shall be interference free. All references to water in
the method refer to reagent water unless otherwise specified. For further details, consult the
document listed in Sec. 13.3.3.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 All samples must have been collected using a sampling plan that addresses the
considerations discussed in Chapter Nine of SW-846. Refer to that chapter, as updated, for
guidance.
8.2 All sample containers must be prewashed with acids and water, and metal-free
detergents, if necessary, depending on the history of use of the container (Ref. 3). Plastic and glass
containers are both suitable. For further information, see Chapter Three of SW-846.
8.3 Samples must be refrigerated upon receipt and analyzed as soon as possible.
9.0 QUALITY CONTROL
9.1 All quality control data must be maintained and available for reference or inspection for
a period of three years. This method is restricted to use by, or under supervision of, experienced
analysts.
9.2 Duplicate samples should be processed on a routine basis. A duplicate sample is a
sample brought through the whole sample preparation and analysis process. A duplicate sample
should be processed with each analytical batch or every 20 samples, whichever is the greater
number. A duplicate sample should be prepared for each matrix type (i.e., soil, sludge, etc.).
9.3 Spiked samples or standard reference materials should be included with each group
of samples processed, or every 20 samples, whichever is the greater number. A spiked sample
should also be included whenever a new sample matrix is being analyzed.
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9.4 Periodically, the accuracy of the temperature measurement system used to control the
microwave equipment should be validated per Section 6.1.2.
9.5 (Not necessary if using temperature feedback control.) Each day that samples are
extracted, the microwave-power calibration should be verified by heating 1 kg of ASTM Type II water
(at 22 ±3 °C) in a covered, microwave-transparent vessel for 2 min at the setting for 490 W and
measuring the water temperature after heating per Section 10.1.5. If the power calculated (per
Section 12) differs from 490 W by more than ± 10 W, the microwave settings should be recalibrated
per Section 10.0.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Calibration of Microwave Equipment
NOTE: If the microwave unit uses temperature feedback control to control the performance
specifications of the method, then performing the calibration procedure is not necessary.
10.1.1 Calibration is the normalization and reproduction of a microwave field strength
to permit reagent and energy coupling in a predictable and reproducible manner. It balances
reagent heating and heat loss from the vessels and is equipment dependent due to the heat
retention and loss characteristics of the specific vessel. Available power is evaluated to permit
the microwave field output in watts to be transferred from one microwave system to another.
Use of calibration to control this reaction requires balancing output power, coupled
energy, and heat loss to reproduce the temperature heating profile given in Section 11.3.5.
The conditions for each acid mixture and each batch containing the same specified number
of vessels must be determined individually. Only identical add mixtures and vessel models
and specified numbers of vessels may be used in a given batch.
10.1.2 For cavity type microwave equipment, calibration is accomplished by
measuring the temperature rise in 1 kg of water exposed to microwave radiation for a fixed
period of time. The analyst can relate power in watts to the partial power setting of the system.
The calibration format required for laboratory microwave systems depends on the type of
electronic system used by the manufacturer to provide partial microwave power. Few systems
have an accurate and precise linear relationship between percent power settings and absorbed
power. Where linear circuits have been utilized, the calibration curve can be determined by
a three-point calibration method (see Section 10.1.4). Otherwise, the analyst must use the
multiple point calibration method (see Section 10.1.3). Assistance in calibration and software
guidance of calibration are available in reference 3 and the document listed in Sec. 13.3.5.
10.1.3 The multiple point calibration involves the measurement of absorbed power
over a large range of power settings. Typically, for a 600 W unit, the following power settings
are measured: 100, 99, 98, 97, 95,90, 80, 70,60, 50, and 40% using the procedure described
in Section 10.1.5. This data is clustered about the customary working power ranges. Non-
linearity has been encountered at the upper end of the calibration. If the system's electronics
are known to have nonlinear deviations in any region of proportional power control, it will be
necessary to make a set of measurements that bracket the power to be used. The final
calibration point should be at the partial power setting that will be used in the test. This setting
should be checked periodically to evaluate the integrity of the calibration. If a significant
change is detected (± 10 W), then the entire calibration should be re-evaluated.
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10.1.4 The three-point calibration Involves the measurement of absorbed power at
three different power settings. Measure the power at 100% and 50% using the procedure
described in Section 10.1.5. From this 2-point line, determine the partial power setting that
corresponds to the power, in watts, specified in the procedure to reproduce the heating profile
specified in Section 11.3.6. Measure the absorbed power at that partial power setting. If the
measured absorbed power does not correspond to the specified power within ± 10 W, use the
multiple point calibration in Section 10.1.3. This point should also be used to periodically verily
the integrity of the calibration.
10.1.5 Equilibrate a large volume of water to room temperature (22 ± 3 °C). One kg
of reagent water is weighed (1,000.0 ±0.1 g) into a fluorocarbon beaker or a beaker made of
some other material that does not significantly absorb microwave energy (glass absorbs
microwave energy and is not recommended). The initial temperature of the water should be
22 ± 3 °C measured to ± 0.05 °C. The covered beaker is circulated continuously (in the
normal sample path) through the microwave field for 2 minutes at the desired partial power
setting with the system's exhaust fan on maximum (as it will be during normal operation). Hie
beaker is removed and the water vigorously stirred. Use a magnetic stirring bar inserted
immediately after microwave irradiation (irradiating with the stir bar in the vessel could cause
electrical arcing) and record the maximum temperature within the first 30 seconds to ± 0.05
°C. Use a new sample for each additional measurement. If the water is reused (after making
adjustments for any loss of weight due to heating), both the water and the beaker must have
returned to 22 ± 3 °C. Three measurements at each power setting should be made.
The absorbed power is determined by the following relationship:
Equation 1 p. (K)(Cp)(m)(AT)
t
Where:
P = the apparent power absorbed by the sample in watts (W) (joule/sec)
K = the conversion factor for thermochemical calories sec"1 to watts (K= 4.184)
Cp = the heat capacity, thermal capacity, or specific heat [cal/(g °C)J of water
m »the mass of the water sample in grams (g)
AT = the final temperature minus the initial temperature (°C)
t s= the time in seconds (s)
Using the experimental conditions of 2 minutes (120 sec) and 1 kg (1000 g) of distilled water
[heat capacity at 25 °C is 0.9997 cal/(g °C)] the calibration equation simplifies to:
Equation 2 P=(AT)(34.86)
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NOTE: Stable line voltage is necessary for accurate and reproducible calibration and operation.
The line voltage should be within manufacturer's specification, and during measurement and
operation should not vary by more than ±2 V (Reference 3). Electronic components in most
microwave units are matched to the system's function and output. When any part of the high
voltage circuit, power source, or control components in the system have been serviced or
replaced, it will be necessary to recheck the system's calibration. If the power output has
changed significantly (±10 W), then the entire calibration should be re-evaluated.
11.0 PROCEDURE
11.1 Temperature control of closed vessel microwave instruments provides the main
feedback control performance mechanism for the method. Method control requires a temperature
sensor in one or more vessels during the entire decomposition. The microwave decomposition
system should sense the temperature to within ± 2.5 °C and permit adjustment of the microwave
output power within 2 seconds.
11.2 All digestion vessels and volumetric ware must be carefully add washed and rinsed
with reagent water. When switching between high concentration samples and low concentration
samples, all digestion vessels (fluoropolymer or quartz liners) should be cleaned by leaching with
hot (1:1) hydrochloric acid (greater than 80 °C, but less than boiling) for a minimum of two hours
followed by hot (1:1) nitric add (greater than 80 °C, but less than boiling) for a minimum of two
hours. The vessels should then be rinsed with reagent water and dried in a clean environment. This
cleaning procedure should also be used whenever the prior use of the digestion vessels is unknown
or cross contamination from prior sample digestions in vessels is suspected. Polymeric or glass
volumetric ware and storage containers should be cleaned by leaching with more dilute adds
(approximately 10% V/V) appropriate for the specific material used and then rinsed with reagent
water and dried in a dean environment.
11.3 Sample Digestion
11.3.1 Weigh a well-mixed sample to the nearest 0.001 g into an appropriate vessel
equipped with a controlled pressure relief mechanism. For soils, sediments, and sludges, use
no more than 0.500 g. For oil or oil contaminated soils, initially use no more than 0.250 g.
When large samples of oil are necessary, use of EPA Method 3052, which has sample scale-
up options, is recommended. If the sample can not be well mixed and homogenized on an as
received basis, then air or oven drying at 60°C or less, crushing, sieving, grinding, and mixing
should be performed as necessary to homogenize the sample until the subsampling variance
is less than the data quality objectives of the analysis. While proper sample preparation
generally produces great reduction in analytical variability, be aware that in certain unusual
circumstances there could be loss of volatile metals (e.g., Hg, organometailics) or irreversible
chemical changes ( e.g., predpitation of insoluble spedes, change in valence state). See
Chapter Three for more details.
11.3.2 Add 10 ± 0.1 ml concentrated nitric add or, alternatively, 9 ± 0.1 mL
concentrated nitric acid and 3 ± 0.1 mL concentrated hydrochloric acid to the Vessel in a fume
hood (or fume exhausted endosure). The addition of concentrated hydrochloric add to the
nitric acid is appropriate for the stabilization of certain analytes, such as Ag, Ba, and Sb and
high concentrations of Fe and Al in solution. Improvements and optimal recoveries of
antimony, iron, and silver from a variety of matrices upon addition of HCI are demonstrated in
Section 17.0, in Figures 3 through 7. The addition of hydrochloric add may, however, limit the
detection techniques or increase the difficulties of analysis for some detection systems.
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CAUTION: The addition of hydrochloric acid must be in the form of concentrated
hydrochloric acid and not from a premixed combination of acids as a buildup of chlorine gas,
as well as other gases, will result from a premixed acid solution. These gases may be violently
released upon heating. This is avoided by adding the acid in the described manner.
CAUTION: Toxic nitrogen oxide(s) and chlorine fumes are usually produced during
digestion. Therefore, all steps involving open or the opening of microwave vessels must be
performed in a properly operating fume ventilation system.
CAUTION: The analyst should wear protective gloves and face protection.
CAUTION: The use of microwave equipment with temperature feedback control is
required to control any unfamiliar reactions that may occur during the leaching of samples of
unknown composition. The leaching of these samples may require additional vessel
requirements such as increased pressure capabilities.
11.3.3 The analyst should be aware of the potential for a vigorous reaction,
especially with samples containing volatile or easily oxidized organic species. When digesting
a matrix of this type, initially use no more than 0.100 g of sample. If a vigorous reaction occurs
upon the addition of reagent(s), allow the sample to predigest in the uncapped digestion vessel
until the reaction ceases. Heat may be added in this step for safety considerations (for
example, the rapid release of carbon dioxide from carbonates, easily oxidized organic matter,
etc.). Once the initial reaction has ceased, the sample may continue through the digestion
procedure. However, if no appreciable reaction occurs, a sample mass of up to 0.250 g for
oils, or 0.500 g for solids, may be used.
11.3.4 Seal the vessel according to the manufacturer's directions. Properly place
the vessel in the microwave system according to the manufacturer's recommended
specifications and, when applicable, connect appropriate temperature and pressure sensors
to vessels according to manufacturer's specifications.
11.3.5 This method is a performance based method, designed to achieve or
approach consistent leaching of the sample through achieving specific reaction conditions.
The temperature of each sample should rise to 175 ±5 °C in approximately 5.5 ± 0.25 minutes
and remain at 175 ± 5 °C for 4.5 minutes, or for the remainder of the ten minute digestion
period (see Refs. 2,3, and 4 and the document listed in 13.3.4). The time versus temperature
and pressure profile is given for a standard sediment sample in Figure 2. When using
temperature feedback control, the number of samples that may be simultaneously digested
may vary, from one sample up to the maximum number of vessels that can be heated by the
magnetron of the microwave unit according to the heating profile specified previously in this
section. (The number will depend on the power of the unit, the number of vessels, and the
heat loss from the vessels (Ref. 3)).
The pressure should peak between 5 and 10 minutes for most samples (see Refs. 1
and 2 and the document listed in 13.3.4). If the pressure exceeds the pressure limits of the
vessel, the pressure should be safely and controllably reduced by the pressure relief
mechanism of the vessel.
11.3.5.1 Calibration control is applicable in reproducing this method
provided the power in watts versus time parameters are determined to reproduce the
specifications listed in 11.3.5. The calibration settings will be specific to the quantity
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of reagents, the number of vessels, and the heat loss characteristics of the vessels
(see Ref. 3 and the document listed in Sea 13.3.3). If calibration control is being used,
any vessels containing acids for analytical blank purposes are counted as sample
vessels. When fewer than the recommended number of samples are to be digested,
the remaining vessels should be filled with the same acid mixture to achieve the full
complement of vessels. This provides an energy balance, since the microwave power
absorbed is proportional to the total absorbing mass in the cavity. Irradiate each group
of vessels using the predetermined calibration settings. (Different vessel types should
not be mixed).
11.3.6 At the end of the microwave program, allow the vessels to cod for a minimum
of 5 minutes before removing them from the microwave system. Cooling of the vessels may
be accelerated by internal or external cooling devices. When the vessels have cooled to near
room temperature, determine if the microwave vessels have maintained their seal throughout
the digestion. Due to the wide variability of vessel designs, a single procedure is not
appropriate. For vessels that are sealed as discrete separate entities, the vessel weight may
be taken before and after digestion to evaluate seal integrity. If the weight loss of sample
exceeds 1% of the weight of the sample and reagents, then the sample is considered
compromised. For vessels with burst disks, a careful visual inspection of the disk, in addition
to weighing, may identify compromised vessels. For vessels with resealing pressure relief
mechanisms, an auditory or a physical sign that can indicate whether a vessel has vented is
appropriate.
11.3.7 Complete the preparation of the sample by carefully uncapping and venting
each vessel in a chemical fume hood (or fume exhausted enclosure). Vent the vessels using
the procedure recommended by the vessel manufacturer. Quantitatively transfer the sample
to an acid-cleaned bottle. If the digested sample contains particulates which may dog
nebulizers or interfere with injection of the sample into the instrument, the sample may be
centrifuged (11.3.7.1), allowed to settle (11.3.7.2), or filtered (11.3.7.3).
11,3.7.1 Centrifugaiion: Centrifugation at 2,000 - 3,000 rpm for 10 minutes
is usually sufficient to clear the supernatant.
11.3.7:2 Settling: If undissolved material, such as SiOz, Ti02, or other
refractory oxides, remains, allow the sample to stand until the supernatant is clear.
Allowing a sample to stand overnight will usually accomplish this. If it does not,
centrifuge or filter the sample.
11.3.7.3 Filtering: If necessary, the filtering apparatus must be thoroughly
cleaned and pre-rinsed with dilute (approximately 10% V/V) nitric acid. Filter the
sample through qualitative filter paper into a second aad-cleaned container.
11.3.8 The removal or reduction of the quantity of the nitric and hydrochloric acids
prior to analysis may be desirable. The chemistry and volatility of the analytes of interest
should be considered and evaluated when using this alternative (Reference 3). Evaporation
to near dryness in a controlled environment with controlled purge gas and neutralizing and
collection of exhaust interactions is an alternative where appropriate. This manipulation may
be performed in the microwave system, if the system is capable of this function, or external to
the microwave system in more common apparatus(s). This option must be tested and
validated to determine analyte retention and loss and should be accompanied by equipment
validation possibly using the standard addition method and standard reference materials. This
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alternative may be used to alter either the acid concentration and/or acid composition prior to
analysis. (For further information, see reference 3 and Method 3052).
NOTE The final solution typically requires nitric add to maintain appropriate sample
solution acidity and stability of the elements. Commonly, a 2% (v/v) nitric acid
concentration is desirable. Waste minimization techniques should be used to capture
reagent fumes. This procedure should be tested and validated in the apparatus and
on standards before using on unknown samples.
11,3.9 Transfer or decant the sample into volumetric ware and dilute the digest to a known
volume. The digest is now ready for analysis for elements of interest using appropriate elemental
analysis techniques and/or SW-846 methods.
12.0 DATA ANALYSIS AND CALCULATIONS
12.1 Calculations: The concentrations determined are to be reported on the basis of the
actual weight of the original sample.
12.2 Prior to use of the method, verify that the temperature sensing equipment is property
reading temperature. A procedure for verification is given in Section 6.1.2. This will establish the
accuracy and precision of the temperature sensing equipment, which should be carried throughout
the statistical treatment of the quality assurance data.
12.3 In calibrating the microwave unit (Section 10.0), the power absorbed (for each power
setting) by 1 kg of reagent water exposed to 120 seconds of microwave energy is determined by the
expression
Power (in watts) = (T, - T2) (34.86)
where: T, « Initial temperature of water (between 21 and 25 °C to nearest 0.1 °C)
T2 = Final temperature of water (to nearest 0.1 °C)
12.4 Plot the power settings against the absorbed power (calculated in Section 12.3) to
obtain a calibration relationship. Alternatively, use a microwave calibration program to analyze the
calibration data (see Ref. 3 and the document listed in Sec. 13.3.5).' Interpolate the data to obtain
the instrument settings needed to provide the wattage levels specified in Section 12.3.
12.5 Calculate the sample dry-weight fraction as follows:
Diy-Wt fraction = ^•
(N,) - flny
where: W1 = Wt for sample + vessel, before drying, g
W2 = Wt for sample + vessel, after drying, g
W3 = Wt for empty, dry vessel, g
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12.6 Convert the extract concentration obtained from the instrument in mg/L to mg/kg dry-
weight of sample by:
Sample concentration = ^
(W)(S)
where: C = Concentration in extract (mg/L)
D = Dilution factor
S = Solid dry-weight fraction for sample, g/g
V = Volume of extract, mL x 0.001
W = Weight of undried sample extracted, g x 0.001
13.0 METHOD PERFORMANCE
13.1 The fundamental chemical basis of Method 3051 with and without HCI has been
compared with Method 3050 in several sources (see 13.3.4 and 13.3.5). Several papers have
evaluated the teachability of NIST SRMs with this method (Ref. 1 and Sec. 13.3.5). Evaluations and
optimizations of this method are being published (Ref. 5 and 6) as well as additional leaches
performed on more matrices, which will be addressed in future literature papers. Method 3051 has
been determined to be appropriate for enhancing recoveries of certain analytes. This data is
contained in Section 17 of this method. Matrices tested include SRM 2710 (Montana Soil - Highly
Elevated Concentrations), SRM 2704 (Buffalo River Sediment), and SRM 1084a (Wear Metals in
Oil). Analytes demonstrating better recoveries upon addition of HCI include antimony, iron, and
silver.
13.2 The foBowing documents may provide additional guidance and insight on this method
and technique:
13.2.1 Kingston, H. M. and L B. Jassie, "Safety Guidelines for Microwave Systems
in the Analytical Laboratory". In Introduction to Microwave Acid Decomposition: Theory and
Practice: Kingston, H.M. and Jassie, L.B., eds,; ACS Professional Reference Book Series;
American Chemical Society: Washington, DC, 1988.
13.2.2 1985 Annual Book of ASTM Standards. Vol. 11.01; "Standard Specification
for Reagent Water; ASTM, Philadelphia, PA, 1985, D1193-77. -
13.3.3 Introduction to Microwave Sample Preparation: Theory aid Practice.
Kingston, H.M. and Jassie, L.B., Eds.; ACS Professional Reference Book Series; American
Chemical Society: Washington, DC, 1988.
13.3.4 Kingston, H.M., Waiter, P.J., "Comparison of Microwave Versus Conventional
Dissolution for Environmental Applications", Spectroscopy, Vol. 7 No. 9,20-27,1992.
13.3.5 Waiter, P. J. Special Publication IR4718: Microwave Calibration Program,
2.0 ed.; National Institutes of Standards and Technology: Gaithersburg, MD, 1991.
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13.3.6 Kingston, H.M., Walter, P.J., Chalk, S.J., Lorentzen, E.M., Link, D.D.,
"Environmental Microwave Sample Preparation: Fundamentals, Methods, and Applications".
In Microwave Enhanced Chemistry: Fundamentals. Sample Preparation, and Applications:
ACS Professional Reference Book Series; American Chemical Society: Washington, DC
1997.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity
or toxicity of waste at the pant of generation. Numerous opportunities for pollution prevention exist
in laboratory operations. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of first choice.
Whenever feasible, laboratory personnel should use pollution prevention techniques to address their
waste generation. When wastes cannot be feasibly reduced at the source, the Agency recommends
recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions, consult Less is Better: Laboratory Chemical Management for Waste Reduction.
available from the American Chemical Society's Department of Government Relations and Science
Policy, 1155 16th Street, NW, Washington, DC 20036, (202) 872-4477.
15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be consistent with all applicable rules and regulations. The Agency urges laboratories to protect the
air, water, and land by minimizing and controlling all releases from hoods and bench operations,
complying with the letter and spirit of any sewer discharge permits and regulations, and by complying
with all solid and hazardous waste regulations, particularly the hazardous waste identification rules
and land disposal restrictions. For further information on waste management, consult The Waste
Management Manual for Laboratory Personnel, available from the American Chemical Society's
Department of Government Relations and Science Policy, 115516th Street, NW, Washington, DC
20036, (202) 872-4477.
16.0 REFERENCES
1. Kingston, H.M. EPA IAG #DWI-393254-01-0 January 1-March 31,1988, quarterly report.
2. Binstock, D.A., Yeager, W.M., Grohse, P.M. and Gaskill, A. Validation of a Method for
Determining Elements in Solid Waste bv Microwave Digestion. Research Triangle Institute
Technical Report Draft, RTI Project Number 321U-3579-24, November, 1989, prepared for the
Office of Solid Waste, U.S. Environmental Protection Agency, Washington, DC 20460.
3. Kingston, H.M., Haswell, S, Microwave Enhanced Chemistry: Fundamentals. Sample
Preparation, and Applications: ACS Professional Reference Book Series; American Chemical
Society: Washington, DC 1997.
4. Binstock, D.A., Grohse, P.M., Gaskill, A., Sellers, C., Kingston, H.M., Jassie, L.B.,
"Development and Validation of a Method for Determining Elements in Solid Waste using
Microwave Digestion", J. Assoc. Off. Anal. Chem., Vol. 74, 360 - 366 , 1991.
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5. Kingston, H.M., Walter, P.J., Lorentzen, E.M.L., Lusnak, G.P. The Performance of Leaching
Studies on Soil SRM's 2710 and 2711"; Final Report to the National Institute of Standards and
Technology, Duquesne University: Pittsburgh, PA, April 5,1994.
6. Link, D.D., Kingston, H.M., Walter, P.J., "Development and Validation of the New EPA
Microwave-assisted Leach Method 3051A".
7. Link, DD., Kingston, H.M., Waiter, P.J.; Development and Validation of the EPA Microwave-
assisted Methods 301SA and 3051A: Validation Studies for Updated Microwave Leach
Methods, Proceedings for the Waste Testing and Quality Assurance Symposium; July 1997.
8. Kingston, H.M., Walter, P.J., "Comparison of Microwave verses Conventional Dissolution for
Environmental Applications", Spectroscopy, Vol. 7 No. 9,20-27,1992.
9. Walter, P.J. Special Publication IR7718: Microwave Calibration Program, 2.0 ed.; National
Institute of Standards and Technology: Gaithersburg, MD, 1991.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 through 3, Figures 1 through 7, and a flow diagram of
method procedure.
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TABLE 1
COMPARISON OF ANALYTE RECOVERIES FROM SRM 2704 (BUFFALO RIVER SEDIMENT)
USING BOTH DIGEST OPTIONS OF METHOD 3051 (Refs. 5, 6)
Element
10 mL HN03
digest
9 mL HNOa +
3 mL Ha digest
Total Analyte
Concentration
Cd
3.40 ±0.34
3.62 ±0.17
3.45 ± 0.22
Cr
84.7 ± 5.6
77.1 ± 12.6
135 ±5
Ni
45.5 ± 5.9
42.2 ± 3.2
44.1 ± 3.0
Pb
163 ±9
161 ±17
161 ± 17
Results reported in pg/g analyte (mean ± 95% confidence limit).
Total concentrations are taken from NIST SRM Certificate of Analysis.
TABLE 2
COMPARISON OF ANALYTE RECOVERIES FROM SRM 4355 (PERUVIAN SOIL)
USING BOTH DIGEST OPTIONS OF METHOD 3051 (Ref. 6).
Element
10 mL HNO3
digest
9 mL HNO3 +
3 mL HCI digest
Total Analyte
Concentration
Cd
0.86 ±0.16
0.85 ±0.17
(1.50)
Cr
14.6 ± 0.47
19.0 ± 0.69
28.9 ± 2.8
Ni
9.9 ± 0.33
11.2 ±0.44
(13)
Pb
124 ± 5.3
130 ±3.6
129 ± 26
Results reported in pg/g analyte (mean ± 95% confidence limit).
Total concentrations are taken from NIST SRM Certificate of Analysis.
Values in parenthesis are reference concentrations.
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TABLE 3
COMPARISON OF ANALYTE RECOVERIES FROM SRM 1084A (WEAR METALS !N OIL)
USING BOTH DIGEST OPTIONS OF METHOD 3051 (Ref. 6)
Element
10 mL HN03
digest
9 mL HN03 +
3 mL HCI digest
Total Analyte
Concentration
Cu
91.6 ±4.0
93.0 ±2.6
100.0 ±1.9
Cr
91.2 ± 3.3
94.3 ±3.1
98.3 ± 0.8
Mg
93.2 ± 3.6
93.5 ±2.8
99.5 ±1.7
Ni
91.6 ±3.9
92.9 ± 3.4
99.7 ±1.6
Pb
104 ±4.1
99.5 ±5.1
101.1 ±1.3
Results reported in pg/g analyte (mean ± 95% confidence limit).
Total concentrations are taken from NIST SRM Certificate of Analysis,
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FIGURE 1
TEMPERATURE AND PRESSURE PROFILES FOR THE HEATING OF DIFFERENT RATIOS
OF NITRIC ACID TO HYDROCHLORIC ACID USING METHOD 3051
U
41
2
-------
FIGURE 2
TEMPERATURE AND PRESSURE PROFILE FOR THE EXTRACTION AND DISSOLUTION OF
NIST SRM 2704 (BUFFALO RIVER SEDIMENT) USING DIFFERENT RATIOS OF NITRIC ACID
TO HYDROCHLORIC ACID
U
v
hi
s
-------
FIGURE 3
PERCENT RECOVERY OF ANTIMONY FROM NIST SRM 2710 (MONTANA SOIL) VERSUS
VARIOUS COMBINATIONS OF NITRIC AND HYDROCHLORIC ACIDS (N=6) (Refs. 6, 7)
100
"i—i—n—f—i—i—i—3—j—r
80
60
4)
5
u
4)
Jh
X>
m 40
20
0
10:0
_J 1_J L_*_J I [ I I S I 1 I S I L
Acid Ratio
-------
FIGURE 4
PERCENT RECOVERY OF ANTIMONY FROM NIST SRM 2704 (BUFFALO RIVER SEDIMENT)
VERSUS VARIOUS COMBINATIONS OF NITRIC AND HYDROCHLORIC ACIDS
(N=6) (RefS. 6, 7).
Acid Ratio (Nitric:Hydrochloric)
3051A - 20
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January 1998
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FIGURES
PERCENT RECOVERY OF IRON FROM NIST SRM 2704 (BUFFALO RIVER SEDIMENT)
VERSUS VARIOUS COMBINATIONS OF NITRIC AND HYDROCHLORIC ACIDS
(N=6) (Refs. 6, 7).
Acid Ratio (NitricHydrochloric)
3051A-21
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January 1998
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FIGURE 6
PERCENT RECOVERY OF SILVER FROM NIST SRM 2710 (MONTANA SOIL) VERSUS
VARIOUS COMBINATIONS OF NITRIC AND HYDROCHLORIC ACIDS (N=6) (Refs. 6,7)
100
~ 95
do
<
b
a?
g 90
u
-------
FIGURE 7
PERCENT RECOVERY OF ANTIMONY AND IRON, RESPECTIVELY, FROM SRM 4355
(PERUVIAN SOIL) USING BOTH DIGEST OPTIONS
(10 ML HNOa AND 9 ML HNOa + 3 ML HCL ) OF METHOD 3051
(N=6) (Refs. 6,7)
100
so
9:3 Sb
60
10:0 Fe
40
20
10;0/5b
0
Acid Ratio (NitricrHydrochloric)
3051A-23
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January 1998
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METHOD 3051A
MICROWAVE ASSISTED ACID DIGESTION OF SEDIMENTS. SLUDGES. SOILS AND OILS
10 1 Cafsbrate
mtcrowov© atjutpmen?
E

11.2 Acid W**ft and water cmm
all digestion vwuH and
Siaaawere
1
'

11.3.1 W«tgn tttquot into m«

aige»on vessel.
1
f

11.3.2 Ado 10* 0.1 mL of csnc. HNGjgor,
alternatively, *±0.1 ml. of cone. NNO3.
and 3*0 1 mL srf ©one. HCt
/n.a.2\
/ OcM a \
/ fiflopsui meson \ Yea
\ occur uDon / '
\ addition of /
\ (Mfltft? /

11.3.4 Seal veaarts ano place m
wtcrewewi syttem.

11.3 5 Heat samples accoramgte ante
of fcemperawte ano ontssure petite.
1
f

11.3 6 Ajlow vetae*s » soot
(0 rami temoeratur*.
y' Have
No / VHMI Y«
Allow sample to
in tmcaeoed
vmm). Add
fteet if necessary
tl 3.7 V«nt Men vmmI. Transfer
sample to an acid cleaned oottie,
11 3.7.1. it 3.7.3
CentrrfuQe, msM or
hWIMM
11.3 a May remote of reduce mno3
1M MCI eveporaeort mar
dryrtMs with eontmtied purpe gas.
11.3.9 Um» aeeropnate * omental
MNtiyKH twcttnKKfv en«W* SW-W<5
method!.
12.fi Calculate concfitrnmna
en original Mmgl* weight.
3051A- 24
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3.3 METHODS FOR DETERMINATION OF INORGANIC ANALYTES
This section of the manual contains analytical techniques for trace inorganic analyte
determinations. Examples of the techniques included in this section are: inductively coupled argon
plasma atomic emission spectrometry (ICP-AES), inductively coupled plasma mass spectrometry
(ICP-MS), direct-aspiration or flame atomic absorption spectrophotometry (FLAA), graphite-furnace
atonic absorption spectrophotometry (GFAA), hydride-generation atomic absorption spectrometry
(HGAA), cold-vapor atomic absorption spectrometry (CVAA), and several procedures for hexavalent
chromium analysis. Each of these is briefly discussed below in terms of advantages, disadvantages,
and cautions for analysis of wastes.
ICP allows simultaneous or rapid sequential determination of many elements in a short time. A
primary disadvantage of ICP is the occurrence of background radiation from other elements and the
plasma gases. Although all ICP instruments utilize high-resolution optics and back-ground correction
to minimize these interferences, analysis for traces of inorganic analytes in the presence of a large
excess of a single analyte is difficult. Examples would be traces of inorganic analytes in an alloy or
traces of metals in a limed (high calcium) waste. ICP and Flame AA have comparable detection
limits (within a factor of 4) except that ICP exhibits greater sensitivity for refractories (Al, Ba, etc.).
Furnace AA, in general, will exhibit lower detection limits than either ICP or FLAA. Detection limits
are drastically improved when ICP-MS is used. In general ICP-MS exhibits greater sensitivity than
either GFAA or FLAA for most elements. The greatest disadvantage of ICP-MS is isobaric elemental
interferences. These are caused by different elements forming atomic ions with the same nominal
mass-to-charge ratio. Mathematical correction for interfering ions can minimize these interferences.
Flame AAS (FLAA) direct aspiration determinations, as opposed to ICP, are normally completed
as single element analyses and are relatively free of interelement spectral interferences. Either a
nitrou s-oxide/acetylene or air/acetylene flame is used as an energy source for dissociating the
aspirated sample into the free atomic state, making analyte atoms available for absorption of light.
In the analysis of some elements, the temperature or type of flame used is critical. If the proper
flame and analytical conditions are not used, chemical and ionization interferences can occur.
Graphite furnace AAS (GFAA) replaces the flame with an electrically heated graphite furnace.
The furnace allows for gradual heating of the sample aliquot in several stages. Thus, the processes
of dissolution, drying, decomposition of organic and inorganic molecules and salts, and formation
of atoms which must occur in a flame or ICP in a few milliseconds may be allowed to occur over a
much longer time period and at controlled temperatures in the furnace. This allows an experienced
analyst to remove unwanted matrix components by using temperature programming and/or matrix
modifiers. The major advantage of this technique is that it affords extremely low detection limits.
It is the easiest to perform on relatively clean samples. Because this technique is so sensitive,
interferences can be a real problem; finding the optimum combination of digestion, heating times and
temperatures, and matrix modifiers can be a challenge for complex matrices.
Hydride AA utilizes a chemical reduction to reduce and separate arsenic or selenium selectively
from a sample digestate. The technique therefore has the advantage of being able to isolate these
two elements from complex samples which may cause interferences for other analytical procedures.
Significant interferences have been reported when any of the following is present: (1) easily reduced
metals (Cu, Ag, Hg); (2) high concentrations of transition metals (>200 mg/L); (3) oxidizing agents
(oxides of nitrogen) remaining following sample digestion.
THREE-19
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Cold-Vapor AA uses a chemical reduction to reduce mercury selectively. The procedure is
extremely sensitive but is subject to interferences from some volatile organics, chlorine, and sulfur
compounds.
Prior to employing the above methods in this chapter, analysts are advised to consult the
disclaimer statement at the front of this manual and the information in Chapter Two for guidance on
the allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to be
used by the analyst and the regulated community in making judgements necessary to meet the data
quality objectives or needs for the intended use of the data.
The following methods are included in this section:
Method 4500:
Method 601 OB:
Method 6020A:
Method 6200:
Method 6500:
Method 6800:
Method 7000B:
Method 7010:
Method 7061A:
Method 7062:
Method 7063:
Method 7195:
Method 7196A:
Method 7197:
Method 7198:
Method 7199:
Method 7470A:
Method 7471B:
Method 7472:
Method 7473:
Method 7474:
Method 7580:
Method 7741A:
Method 7742:
Method 9000:
Method 9001:
Mercury in Soil by Immunoassay
Inductively Coupled Plasma-Atomic Emission Spectrometry
Inductively Coupled Plasma - Mass Spectrometry
Field Portable X-Ray Fluorescence Spectrometry for the Determination of
Elemental Concentrations in Soil and Sediment
Dissolved Inorganic Anions in Aqueous Matrices by Capillary Ion
Electrophoresis
Elemental and Speciated Isotope Dilution Mass Spectrometry
Flame Atomic Absorption Spectrophotometry
Graphite Furnace Atomic Absorption Spectrophotometry
Arsenic (Atomic Absorption, Gaseous Hydride)
Antimony and Arsenic (Atomic Absorption, Borohydride Reduction)
Arsenic in Aqueous Samples and Extracts by Anodic Stripping Voltammetry
(ASV)
Chromium, Hexavalent (Coprecipitation)
Chromium, Hexavalent (Colorimetric)
Chromium, Hexavalent (Chelation/Extraction)
Chromium, Hexavalent (Differential Pulse Polarography)
Determination of Hexavalent Chromium in Drinking Water, Groundwater and
industrial Wastewater Effluents by Ion Chromatography
Mercury in Liquid Waste (Manual Cold-Vapor Technique)
Mercury in Solid or Semisolid Waste (Manual Cold-Vapor Technique)
Mercury in Aqueous Samples and Extracts by Anodic Stripping Voltammetry
(ASV)
Mercury in Solids and Solutions by Thermal Decomposition, Amalgamation,
and Atomic Absorption Spectrophotometry
Mercury in Sediment and Tissue Samples by Atomic Fluorescence
Spectrometry
White Phosphorus (P4) by Solvent Extraction and Gas Chromatography
Selenium (Atomic Absorption, Gaseous Hydride)
Selenium (Atomic Absorption, Borohydride Reduction)
Determination of Water in Waste Materials by Karl Fischer Titration
Determination of Water in Waste Materials by Quantitative Calcium Hydride
Reaction
THREE - 20
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METHOD 4500
MERCURY in SOU PY IMMUNOASSAY
1.0 SCOPE AND APPLICATION
1.1 This method provides a screening procedure for the determination of mercury in soils
at concentrations as low as 0.5 mg/Kg.
1.2 This procedure describes the analysis of soil samples for the detection of mercury by
an Enzyme-Linked Immunosorbent Assay (ELISA). This method is conducted as a screening
technique, the sample's concentration is estimated through comparison of the sample to a standard.
Other solid matrices (see Table 5) may be analyzed by this technique as long as the QC parameters
detailed in this method are achievable.
1.3 In conjunction with this technique, Method 7471 should be used to determine the exact
concentration of mercury when required or for confirmatory purposes. This is especially true near
regulatory or action levels. A minimum of 10% confirmatory analyses is suggested.
14 This method is restricted to use by or under the supervision of trained analysts. Each
analyst must demonstrate the ability to generate acceptable results with this method.
1.5 Tubes or reagents from different kits or separate batches are NOT interchangeable.
2.0 SUMMARY OF THE METHOD
2.1 Test kits are commercially available for this method. The manufacturer's directions
should be followed.
2.2 In general, the method is performed using an extract of a solid sample. Solid samples
are prepared by extraction with a mixture of hydrochloric and nitric acids for ten minutes and then
buffered prior to analysis. The sample is added to a tube (treated with BSA-glutathione) and
incubated at ambient temperatures for five minutes. The mercuric ions bound to the sulflhydryl
groups of the BSA-glutathione are now reacted with a reconstituted antibody specific for mercury and
incubated for five more minutes. A peroxidase conjugate is added to the sample, reacting with any
mercury specific antibody. The substrate is then added forming a color that is in proportion to the
amount of mercury originally present in the sample. The color produced is then
spectrophotometrically compared with the control standards.
3.0 DEFINITIONS
3.1 Antibody - A binding protein which is produced in response to an antigen, and which
has the ability to bond with the antigen that stimulated its production.
3.2 Cross-Reactivity - The relative concentration of an untargeted substance that would
produce a response equivalent to a specified concentration of the targeted compound. In a semi-
quantitative immunoassay, it provides an indication of the concentration of cross-reactant that would
produce a positive response. Cross-reactivity for individual compounds is often calculated as the
ratio of target substance concentration to the cross-reacting substance concentration at 50%
inhibition of the immunoassay's maximum signal x 100%.
4500 -1
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3.3 Dose-Response Curve - Representation of the signal generated by an immunoassay
(y axis) plotted against the concentration of the target compound (x axis) in a series of standards of
known concentration. When plotting a competitive immunoassay in a rectilinear format, the dose-
response will have a hyperbolic character. When the Iog,0 of concentration is used, the plot
assumes a sigmoidal shape, when the iog of signal is plotted against the logit transformation (linear
representation of calibration data) of concentration, a straight line is produced.
3.4 ELISA - Enzyme Linked Immunosorbent Assay is an enzyme immunoassay method that
uses an immobilized reagent {e.g., antibody adsorbed to a plastic tube) to facilitate the separation
of targeted anaiytes (antibody-bound components) from non-targeted substances (free reaction
components), using a washing step and an enzyme conjugate to generate the signal used for the
interpretation of results.
3.5 Enzyme Conjugate - A molecule produced by the coupling of an enzyme molecule to
an immunoassay component that is responsible for acting upon a substrate to produce a detectable
signal.
3.6 Enzyme Immunoassay - An immunoassay method that uses an enzyme conjugate
reagent to generate the signal used for interpretation of results. The enzyme mediated response
may take the form of a chnomogenic, Audiogenic, chemiluminescent or potentiometric reaction, (see
immunoassay and ELISA)
3.7 False Negatives - A negative interpretation of the sample containing the target anaiytes
at or above the action level. Ideally, an immunoassay test product should produce no false
negatives. The maximum permissible false negative rate is 5%, as measured by analyzing split
samples using both the test product and a reference method.
3.8 False Positives • A positive interpretation for a sample is defined as a positive response
for a sample that contains anaiytes below the action level.
3.9 Immunoassay - An analytical technique that uses an antibody molecule as a binding
agent in the detection and quantitation of substances in a sample, (see Enzyme Immunoassay and
ELISA)
3.10 Immunogen - A substance having a minimum size and complexity, and that is
sufficiently foreign to a genetically competent host to stimulate an immune response.
4.0 INTERFERENCES
4.1 Refer to Table 3 for a comparison of the effects of other metals on the procedure.
4.2 Consult the information provided by the manufacturer of the kit used for additional
information regarding cross reactivity with other compounds.
4.3 Temperature range in which test can be reliably conducted (refer to test-kit
instructions).
5.0 SAFETY
Refer to Chapter Three for guidance.
4500-2
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6.0 EQUIPMENT AND SUPPLIES
6.1 BiMelyze® Soil Extraction Kit and BiMelyze® Mercury Assay Tube Kit for Solid Matrices
(BioNebraska, Inc. 3820 NW 46th St., Lincoln, NE 68524) or equivalent. Each commercially
available test kit will supply or specify the apparatus and materials necessary for successful
completion of the test.
6.2 Analytical balance - capable of measuring 5 g ± 0.1 g.
6.3 Differential photometer or equivalent - capable of reading the absorbance at 405 nm.
6.4 Timer.
6.5 Permanent marking pen.
6.6 Cleaning and waste supplies - lab tissues, disposable gloves, waste container.
6.7 Micropipets - capable of accurate delivery volumes at 105 and 500 yL.
6.8 Squirt bottle - 500 mL or equivalent
6.9 Graduated cylinder - 500 mL or equivalent.
7.0 REAGENTS AND STAN DARDS
7.1 Each commercially available test kit will supply or specify the reagents necessary for
successful completion of the test.
7.2 Nitric acid, concentrated - reagent grade or equivalent.
7.3 Hydrochloric acid, concentrated - reagent grade or equivalent.
7.4 Acid mixture -Add 36 mL HCI to 18 mL of reagent water and then add 18 mL of HNOs
to the HCI/reagent water solution.
7.5 Reagent water - All references to water in this method refer to reagent water unless
otherwise specified. Reagent grade water is defined in Chapter One.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 Environmental samples may be contaminated, and should therefore be considered
hazardous and handled accordingly.
8.2 All test kits must be stored under the conditions described by the manufacturer.
8.3 Sample Collection - Sufficient sample should be collected to ensure a representative
sample. Samples should be collected in pre-cleaned glass or plastic containers.
8.4 All samples that are not immediately analyzed must be stored under the conditions
described in Chapter Three.
4500-3
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9.0 QUALITY CONTROL
9.1 Follow the manufacturer's instructions for the test kit being used for quality control
procedures specific to the test kit used. Additionally, guidance provided in Method 4000 and Chapter
One should be followed.
9.2 Use of replicate analyses, particularly when results indicate concentrations near the
action level, is recommended to refine information gathered with the tube kit.
9.3 Do not use test kits past their expiration date.
9.4 Do not use tubes or reagents designated for use with other manufacturer's test kits and
do not use tubes or reagents from separate batches of test kits.
9.5 Use the test kits within their specified storage temperature and operating temperature
limits.
9.6 Although Method 4500 is intended as a field screening method, the appropriate level
of quality assurance should accompany the application of this method to document data quality.
These include but are not limited to positive and negative controls.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Follow the instrument manufacturer's instruction when conducting the calibration.
10.2 All analyses must be accompanied by at least a reference standard (NIST 2709, NIST
2704, or equivalent).
10.3 All analyses must be accompanied by at least one control. Additional controls may be
used to refine the data.
11.0 PROCEDURE
11.1 Follow all of the manufacturer's specific instruction when conducting analyses by the
immunoassay technique. A general overview of the technique follows.
11.2 Prepare all assay solutions, standards, and controls prior to beginning the analysis.
Appropriately label all vials.
11.3 Soil Extraction
11.3.1 Weigh out 5 ± 0.1 g of soil sample and place into the extraction vessel.
11.3.2 Add a 4 mL volume of the acid mixture to the extraction vessel for all
samples, standards, and controls.
11.3.3 Cap the vessels and swirl the samples for 15 seconds of each minute during
the 10-minute extraction period and then add 7 mL of the buffer included with the test-kit.
11.3.4 Place bottle filter tops firmly onto extraction bottles. Squeeze bottle and
discard the first few drops. Add three drops (105 pL) into the corresponding dilution bottles.
4500 - 4
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11.4 Sample Analysis
11.4.1 Add diluted samples to the black line on each mercury assay tube (500 ^L)
Incubate tubes for five minutes at ambient temperature. Occasionally swirl and after five
minutes empty and then rinse tubes three times with approximately 2 mL of reagent water.
11.4.2 Add antibody to the black line on each mercury assay tube (500 fjL), incubate
for 5 minutes at ambient temperature, wash and rinse tubes with buffer three times.
11.4.3 Add conjugate to the black line on each mercury tube (500 |jL), incubate for
5 minutes at ambient temperature, wash and rinse tubes with buffer three times.
11.4.4 Add substrate to the black line on each mercury tube (500 fjL), and incubate
for 5 minutes at ambient temperature. Add three drops (105 jjL) of stop solution to each tube
in the same order as the substrate was added.
11.4.5 Read absorbance of each sample and standard at 405 nm.
11.4.5.1 Samples with absorbances less than that of a control should be
reported as "<[cdntroI concentration]." For example, if the control is 4 ppm and the
sample's absorbance is less than that of the 4 ppm control then the result should be
reported as "«4 ppm."
11.4.5.2 Samples with absorbances greater than that of a control should be
reported as ">[controi concentration]." For example, if the control is 4 ppm and the
sample's absorbance is greater than that of the 4 ppm control then the result should
be reported as ">4 ppm."
11.4.5.3 Results may be bracketed between two controls to further refine the
data. For example, if a 4 ppm control and a 20 ppm control are used, potentially three
results are possible: <4 ppm, 4-20 ppm, and >20 ppm.
12.0 DATA ANALYSIS AND CALCULATIONS
Ensure that all sample containers, dilution bottles, extraction vials, and tubes have been
labeled properly prior to analysis. Proper laboratory protocols, including documentation and
notetaking, should be conducted according to good laboratory practices.
13.0 METHOD PERFORMANCE
13.1 Table 1 displays data obtained from analyzing samples using both the tube kit and
CVAA at a Superfund Site. Sixty-nine samples were analyzed, nine samples analyzed by the tube
kit had results greater than that of the CVAA analyses. This gives a false positive rate of 13%. Two
of the samples analyzed by the tube kit gave results lower than the CVAA analyses. This gives a
false negative result of 2.8%.
13.2 Table 2 displays data concerning false positives and false negatives at the detection
limit of 0.5 ppm. Twenty samples were prepared using NIST Standard Reference Material 8407 (soil
standard) diluted gravimetrically with mercury free soils to levels of 1.0, 0.50, 0.25, and 0.00 ppm.
Four sets of the soil samples were extracted in five separate experiments. Within each of the five
experiments, a 0.5 ppm sample that gave an absorbance greater than that of the 0.5 ppm standard
would be considered a false positive. An absorbance greater than that of a 0.5 ppm standard would
4500 - 5
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be determined to be a false negative,
negative results.
None of the experiments yielded false positives or false
13.3 Table 3 displays the effects of other metals on the quantitation of mercury by ELISA
and the concentration at which cross-reactivity may be observed. Elements with values designated
"as greater than" displayed no interference at the levels shown in the table.
13.4 Table 4 provides comparison of the tube kit with eight different certified reference
materials.
13.5 Table 5 provides data for a set of analyses conducted at an abandoned battery
reclamation site.
13.6 Figure 1 displays the tube-kit's 10 month stability claims at 4°C with a control (fresh kit
every analysis period), a kit stored at room temperature (22-25°C), and a kit stored at 4°C.
13.7 The MDL at the 99% confidence level was derived from the data in Table 2. The
standard deviation of the lowest standard analyzed (0.067 = SD for the 0.25 ppm standard) was
multiplied by the t-statistic for 20 samples (2.54). The calculated MDL is 0.17 ppm. For the
purposes of this methodology the detection limit will be listed as 0.5 ppm.
13.8 The following documents may provide additional guidance and insight on this method
and technique:
13.8.1 Wylie, D.E., Lu, D., Carison, LD., Carlson, R., Babacan, K.F., Schuster, S.M.,
and Wagner, F., "Monoclonal Antibodies Specific for Mercuric Ions", Proc. Natl. Acad Sci.,
Vol. 89, pp 4104-4108, May 1992.
13.8.2 Wylie, D.E., Lu, D., Carlson, LD., Carlson, Schuster, S.M., and Wagner, F.,
"Detection of Mercuric Ions in Water by ELISA with a Mercury-Specific Antibody", Analytical
Biochemistry 194, 381-387 (1991).
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity
and/or toxicity of waste at the point of generation. Numerous opportunities for pollution prevention
exist in laboratory operation. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of first choice.
Whenever feasible, laboratory personnel should use pollution prevention techniques to address their
waste generation. When wastes cannot be feasibly reduced at the source, the Agency recommends
recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Less is Better. Laboratory Chemical Management for Waste Reduction
available from the American Chemical Society as listed in.
15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the tetter and spirit of any sewer discharge permits and regulations, and
4500 - 6
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by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society, Department of Government Relations and Science Policy, 1155 16th Street, NW,
Washington, D.C., 20036, (202) 872-4477.
16.0 REFERENCES
1. BiMelyze® Mercury Assay Kit and BiMelyze® Mercury Assay Soil Extraction Kit, BioNebraska,
Inc.
2. Schweitzer, Craig, et. al.; "Enzyme-Linked Immunoassay (EUSA) for the Detection of Mercury
in Environmental Matrices."
3. Letter to Frank Calovini, SAIC; data submission from Craig Schweitzer, BioNebraska, Inc.;
August 16,1995.
4. Letter and data submission attachments, O.M. Fordham, USEPA; from Craig Schweitzer,
BioNebraska, inc.; March 20,1995.
5. California Environmental Protection Agency, Department of Toxic Substances Control,
Environmental Technology Certification Program, "BiMelyze® Field Screening Assay for
Mercury (Tube Assay") and Soil Extraction Kit, with a Partial Evaluation of BiMelyze®
Laboratory Screening Test for Mercury ("Plate Assay")", BioNebraska, Inc. Lincoln, NB;
Contract No. 93-T0470, June 1995.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 through 5, Figure 1, and a flow diagram for this method's
procedure.
4500 - 7
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TABLE 1
CORRELATION OF IMMUNOASSAY AND CVAA ANALYSES
FOR MERCURY IN SOIL AT A SUPERFUND SITE
Hg conc. by CVAA (ppm)
Hg conc. by Immunoassay
(ppm)
Agreementa
1.0
<5
Y
0.2
<5
Y
0.02
<5
Y
0.03
<5
Y
<0.02
<5
Y
<0.02
<5
Y
0.02
<5
Y
36.2
>15
Y
7.4
5-15
Y
0.03
<5.
Y
0.3
<5 •
Y
0.03
<5
Y
0.03
<5
Y
0.1
<5
Y
<0.03
<5
Y
0.9
<5
Y
0.03
<5
Y
0.04
<5
Y
<0.02
<5
Y
39.4
>15
Y
46.5
>15
Y
18.2
>15
Y
139
>15
Y
106
>15
Y
4.7
>15
N-FP
0.4
<5
Y
(Continued)
4500 - 8
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TABLE 1 (Continued)
| Hg conc. by CVAA (ppm)
Hg conc. by Immunoassay
(ppm)
Agreement*
1.0
<5
Y
0.2
<5
Y
0.3
<5
Y
4.1
5-15
N-FP
<0.02
<5
Y
0.05
<5
Y
56.6
>15
Y
0.5
<5
Y
0.2
<5
Y
0.1
<5
Y
0.3
<5
Y
0.02
<5
Y
0.04
<5
Y
0.08
<5 '
Y
0.03
<5
Y
0.02
<5
Y
<0.01
<5
Y
0J2
<5
Y
0.06
<5
Y
<0.01
<5
Y
<0.01
<5
Y
28.1
5-15
N-FN
51.8
>15
Y
21.8
5-15
N-FN
7.7
5-15
Y
0.4
<5
Y
18.0
>15
Y
(Continued)
4500 -9
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TABLE 1 (Continued)
Hg conc. by CVAA (ppm)
Hg conc. by Immunoassay
(ppm)
Agreement"
0.8
<5
Y
2.2
<5
Y
4.4
5-1S
N-FP
1.1
5-15
N-FP
0.05
<5
Y
1.3
5-1S
N-FP
0.06
<5
Y
<0.02
<5
Y
<0.01
<5
Y
3.1
5-15
N-FP
3.4
5-15
N-FP
0.3
<5
Y
3.4
5-15
N-FP
2.0
5-15
N-FP
0.13
<5
Y
0.06
<5
Y
a Y = Yes, N = No, FN = False Negative, FP = False Positive
Source: Reference 4
4500-10
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TABLE 2
MERCURY ANALYSIS OF SOIL NEAR THE IMMUNOASSAY DETECTION LIMIT
OF 0.50 PPM
ABSORBANCE OBTAINED BY
IMMUNOASSAY CONCENTRATION BY CVAA
SAMPLE CONC. (ppm) SAMPLE CONC. (ppm)
1.0 0.50 0.25 0.00 1.0 0.50 0.25 0.00
#1)
A)
0.97
0.75
0.53
0.08
0.99
0.48
0.24
0.00
B)
0.93
0.74
0.50
0.06
1.00
0.50
0.25
0.00

C)
0.84
0.73
0.51
0.08
0.98
0.53
0.25
0.00

D)
0.88
0.70
0.51
0.08
0.98
0.49
0.24
0.00
#2)
A)
1.05
0.81
0.58
0.10
1.02
0.52
0.24
0.00
B)
0.99
0.83
0.43
0.10
0.90
0.50
0.24
0.00

C)
1.00
0.65
0.57
0.10
0.97
0.49
0.25
0.00

D)
1.08
0.88
0.48
0.12
0.94
0.53
0.25
0.00
#3)
A)
1.17
0.80
0.58
0.08
1.06
0.55
0.26
0.00
B)
1.11
0.82
0.52
0.11
1.03
0.54
0.28
0.00

C)
0.95
0.62
0.43
0.10
1.00
0.52
0.28
0.00

D)
0.99
0.80
0.51
0.11
1.03
0.54
0.27
0.00
#4)
A)
0.91
0.76
0.36
0.09
1.07
0.57
0.28
0.00
B)
0.87
0.66
0.49
0.07
1.15
0.58
0.31
0.00

C)
0.78
0.67
0.42
0.06
1.26
0.57
0.30
0.00

D)
0.90
0.69
0.39
0.06
1.16
0.57
0.28
0.00
#5)
A)
1.15
0.61
0.46
0.07
0.88
0.47
0.24
0.00
B)
1.11
0.67
0.35
0.07
0.94
0.48
0.23
0.00

C)
1.07
0.66
0.48
0.07
0.90
0.47
0.26
0.00

D)
1.09
0.54
0.50
0.09
0.88
0.47
0.24
0.00
Source: Reference 5
4500-11
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TABLE 3
CROSS-REACTIVITY OF ELEMENTS WITH THE MERCURY IMMUNOASSAY
ELEMENT
SOIL EQUIVALENT CONCENTRATION
REQUIRED TO YIELD A POSITIVE RESULT
fPDm)
Mercury
0.36
Arsenic
>55,000
Barium
>100,000
Cadmium
>82,000
Chromium
35,000
Copper
47,000
Gold
144.000
Iron
>41,000
Lead
>150,000
Nickel
>43,000
Silver
79,000
Sodium
>17,000
Strontium
>64,000
Thallium
>150,000
Zinc
>48,000
Source: Reference 3
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TABLE 4
ANALYSIS OF CERTIFIED REFERENCE SOILS USING IMMUNOASSAY
REFERENCE SAMPLE IHg] ABSORBANCE @ 405-nm INTERPRETATION
(ppm) exp.1 exp.2 exp.3
ERA Inorganic Blank Soil
<0.10
0.12
0.05
0.08
3
NIST 2709
1.40
1.01
0.64
0.47
<4
NIST 2704
1.47
0.78
0.41
0.47
<4
ERA CLP Lot #216
2.36
1.54
0.84
0.932
<4
ERA Custom Mercury Std.1

dil. 1
4
1.76
1.01
0.83
3
dil.2
15
1.99
1.45
1.59
3
dil. 3
50
2.04
1.73
2.02
>15
NIST 8408
122
2.55
2.55
2.55
>15
Source: Reference 2
1 dilutions from 107 ppm.
2 only value that gives incorrect conclusion.
3 standard reference point, no interpretation.
TABLE S
ANALYSIS OF MULTIPLE MATRICES AT AN ABANDONED BATTERY RECLAMATION SITE
USING IMMUNOASSAY
SAMPLE DESCRIPTION IMMMUNOASSAY RESULTS CVAA RESULTS
Process Room
< 5 ppm
0.83 ppm
Dust from process room
< 5 ppm
> 4.5 ppm
Groundwater - unfiltered
< 0.5 ppb
< 0,4 ppb
Soil, alkaline
< 5 ppm
0.93 ppm
Sludge from tank
> 15 ppm
4,400 ppm
Sump sludge
5 > 15 ppm
14 ppm
Cinder block
< 5 ppm
3 ppm
Cinderblock duplicate
< 5 ppm

Soil
5 > 15 ppm
14 ppm
Paint
> 15 ppm
34 ppm
Background cinderblock
< 5 ppm
1.4 ppm
Background paint
> 15 ppm
14 ppm
Debris from COz blast
> 15 ppm
19 ppm
Source: Reference 2
4500-13
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FIGURE 1
TEN-MONTH IMMUNOASSAY KIT STABILITY
2.09
1J
1J
1J
1.S
-»-KT
CONTROL |
1.6
0.8
0.45
030 ¦¦
0.4
o.oe
0.00
0.0
1
10
100
uig|Ha]0i|)6)
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METHOD 4500
MERCURY IN SOIL BY IMMUNOASSAY
Start
Stop
11.4,5 Read absorbance at 405 nm.
11,4,4 Add substrate, incubate for
5 miri. Add 3 drops of stop solution
to each tube.
11.4,3 Add conjugate, incubate for
5 mm., wash and rinse with buffer.
11,3.1 Place 5 *1- O.ig of soii sample
into extraction vessel.
11.3.2 Add 4mL of acid mixture to
the extraction vessel.
11.3.4 Place bottle filter tops
onto extraction bottles. Squeeze
bottle and discard first drops. Add
three drops to dilution bottles.
11.4.2 Add antibody to each tube,
incubate for 5 min., wash and rinse
with buffer.
11.4.1 Add diluted samples to each
mercury assay tube. Incubate for
5 min., empty and rinse tubes 3x
with reagent water.
11,3,3 Cap vessels, swirl samples
during 10 min. extraction period.
Add 7mI of buffer.
4500 -15
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METHOD 6020A
INDUCTIVELY COUPLED PLASMA - MASS SPECTROMETRY
1.0 SCOPE AND APPLICATION
1.1 Inductively coupled plasma-mass spectrometry (ICP-MS) is applicable to the
determination of sub-jjg/L concentrations of a large number of elements in water samples and in
waste extracts or digests (References 1 and 2). When dissolved constituents are required, samples
must be filtered and acid-preserved prior to analysis. No digestion is required prior to analysis for
dissolved elements in water samples. Acid digestion prior to filtration and analysis is required for
groundwater, aqueous samples, industrial wastes, soils, sludges, sediments, and other solid wastes
for which total (acid-Ieachable) elements are required.
1.2 ICP-MS has been applied to the determination of over 60 elements in various matrices.
Anaiytes for which EPA has demonstrated the acceptability of Method 6020 in a multi-laboratory
study on solid and aqueous wastes are listed below.
Element

CASRN8
Aluminum
(Al)
7429-90-5
Antimony
(Sb)
7440-36-0
Arsenic
(As)
7440-38-2
Barium
(Ba)
7440-39-3
Beryllium
(Be)
7440-41-7
Cadmium
(Cd)
7440-43-9
Calcium
(Ca)
7440-70-2
Chromium
(Ci)
7440-47-3
Cobalt
(Co)
7440-48-4
Copper
(Cu)
7440-50-8
Iron
(Fe)
7439-89-6
Lead
(Pb)
7439-92-1
Magnesium
(Mg)
7439-95-4
Manganese
(Mn)
7439-96-5
Mercury
(Hg)
7439-97-6
Nickel
(Ni)
7440-02-0
Potassium
(K)
7440-09-7
Selenium
(Se)
7782-49-2
Silver
(Ag)
7440-22-4
Sodium
(Na)
7440-23-5
Thallium
(TO
7440-28-0
Vanadium
(V)
7440-62-2
Zinc
(Zn)
7440-66-6
"Chemical Abstract Service Registry Number
Acceptability of the method for an element was based upon the multi-laboratory performance
compared with that of either furnace atomic absorption spectrophotometry or inductively coupled
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plasma-atomic emission spectrometry. It should be noted that one multi-laboratory study was
conducted in 1988 and advances in ICP-MS instrumentation and software have been made since
that time and additional studies have been added with validation and improvements in performance
of the method. Performance, in general, exceeds the multi-laboratory performance data for the listed
elements. It is expected that current performance will exceed the multi-laboratory performance data
for the listed elements (and others) that are provided in Section 13,0. Instrument detection limits,
sensitivities, and linear ranges will vary with the matrices, instrumentation, and operating conditions.
In relatively simple matrices, detection limits will generally be below 0.1 pg/L, Less sensitive
elements (like Se and As) and desensitized major elements may be 1.0 pg/L or higher.
1.3 If Method 6020 is used to determine any analyte not listed in Section 1.2, it is the
responsibility of the analyst to demonstrate the accuracy and precision of the method in the waste
to be analyzed. The analyst is always required to monitor potential sources of interferences and take
appropriate action to ensure data of known quality (see Section 9.4). Other elements and matrices
may be analyzed by this method if performance is demonstrated for the analyte of interest, in the
matrices of interest, at the concentration levels of interest in the same manner as the listed elements
and matrices (see Sec. 9.0).
1.4 Use of this method should be relegated to spectroscopists who are knowledgeable in
the recognition and in the correction of spectral, chemical, and physical interferences in ICP-MS.
1.5 An appropriate internal standard is required for each analyte determined by ICP-MS.
Recommended internal standards are ®U, ^Sc, 89Y, 103Rh, 1iSin, iSSTb, 1&Ho, and 209Bi. The lithium
internal standard should have an enriched abundance of 6Li, so that interference from lithium native
to the sample is minimized. Other elements may need to be used as internal standards when
samples contain significant native amounts of the recommended internal standards.
2.0 SUMMARY OF METHOD
2.1 Prior to analysis, samples which require total ("acid-leachable") values must be
digested using appropriate sample preparation methods (such as Methods 3005 - 3052).
2.2 Method 6020 describes the multi-elemental determination of analytes by ICP-MS in
environmental samples. The method measures ions produced by a radio-frequency inductively
coupled plasma. Analyte species originating in a liquid are nebulized and the resulting aerosol is
transported by argon gas into the plasma torch. The ions produced by high temperatures are
entrained in the plasma gas and introduced, by means of an interface, into a mass spectrometer.
The ions produced in the plasma are sorted according to their mass-to-charge ratios and quantified
with a channel electron multiplier. Interferences must be assessed and valid corrections applied or
the data flagged to indicate problems. Interference correction must Include compensation for
background ions contributed by the plasma gas, reagents, and constituents of the sample matrix.
3.0 DEFINITIONS
Refer to Chapter One and Chapter Three for a listing of applicable definitions,
4.0 INTERFERENCES
4.1 Isobaric elemental interferences in ICP-MS are caused by isotopes of different
elements forming atomic ions with the same nominal mass-to-charge ratio (m/z). A data system must
be used to correct for these interferences. This involves determining the signal for another isotope
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of the interfering element and subtracting the appropriate signal from the analyte isotope signal.
Since commercial ICP-MS instruments nominally provide unit resolution at 10% of the peak height,
very high ion currents at adjacent masses can also contribute to ion signals at the mass of interest.
Although this type of interference is uncommon, it is not easily corrected, and samples exhibiting a
significant problem of this type could require resolution improvement, matrix separation, or analysis
using another verified and documented isotope, or use of another method.
4.2 Isobaric molecular and doubly-charged ion interferences in ICP-MS are caused by
ions consisting of more than one atom or charge, respectively. Most isobaric interferences that
could affect ICP-MS determinations have been identified in the literature (References 3 and 4).
Examples include reArCr ion on the 75As signal and MoO* ions on the cadmium isotopes. While the
approach used to correct for molecular isobaric interferences is demonstrated below using the
natural isotope abundances from the literature (Reference 5), the most precise coefficients for an
instrument can be determined from the ratio of the net isotope signals observed for a standard
solution at a concentration providing suitable («1 percent) counting statistics. Because the ^Cl
natural abundance of 75.77 percent is 3,13 times the 37CI abundance of 24.23 percent, the chloride
correction for arsenic can be calculated (approximately) as follows (where the ^APCI* contribution
at m/z 75 is a negligible 0.06 percent of the '"Ar^Cf signal):
Corrected arsenic signal (using natural isotopes abundances for coefficient approximations) =
(m/z 75 signal) - (3.13) (m/z 77 signal) + (2.73) (m/z 82 signal),
where the final term adjusts for any selenium contribution at 77 m/z,
NOTE: Arsenic values cm be biased high by this type of equation when the net signal at m/z
82 is caused by ions other than ^Se4, (e.g., 81Brt-f from bromine wastes [Reference 6]).
Similarly,
Corrected cadmium signal (using natural isotopes abundances for coefficient approximations) =
(m/z 114 signal) - (0.027)(m/z 118 signal) - (1.63)(m/z 108 signal),
where last 2 terms adjust for any 114Sn* or 114MoO* contributions at m/z 114.
NOTE: Cadmium values will be biased low by this type of equation when 92ZrO* ions
contribute at m/z 108, but use of m/z 111 for Cd is even subject to direct (^ZrOhf) and
indirect (^ZrO*) additive interferences when Zr is present.
NOTE: As for the arsenic equation above, the coefficients couldfce improved. The most
appropriate coefficients for a particular instrument can be determined from the ratio of the
net isotope signals observed for a standard solution at a concentration providing suitable (<1
percent) counting precision.
The accuracy of these types of equations is based upon the constancy of the OBSERVED isotopic
ratios for the interfering species. Corrections that presume a constant fraction of a molecular ion
relative to the "parent" ion have not been found (Ref. 7) to be reliable, e.g., oxide levels can vary with
operating conditions. If a correction for an oxide ion is based upon the ratio of parent-to-oxide ion
intensities, the correction must be adjusted for the degree of oxide formation by the use of an
appropriate oxide internal standard previously demonstrated to form a similar level of oxide as the
interferent. For example, this type of correction has been reported (Ref. 7) for oxide-ion corrections
using ThQTTh* for the determination of rare earth elements. The use of aerosol desolvation and/or
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mixed gas plasmas have been shown to greatly reduce molecular interferences (Ref. 8). These
techniques can be used provided that method detection limits, accuracy, and precision requirements
for analysis of the samples can be met.
4.3 Additionally, solid phase chelation may be used to eliminate isobaric interferences from
both element and molecular sources. An on-line method has been demonstrated for environmental
waters such as sea water, drinking water and acid decomposed samples. Add decomposed
samples refer to samples decomposed by methods similar to methods 3052, 3051, 3050 or 3015.
Samples with percent levels of iron and aluminum should be avoided. The method also provides
a method for preconcentration to enhance detection limits simultaneously with elimination of isobaric
interferences. The method relies on chelating resins such as imminodiacetate or other appropriate
resins and selectively concentrates the elements of interest while eliminating interfering elements
from the sample matrix. By eliminating the elements that are direct isobaric interferences or those
that form isobaric interfering molecular masses, the mass region is simplified and these interferences
can not occur. The method has been proven effective for the certification of standard reference
materials and validated using SRMs (References 13-15). The method has the potential to be used
on-line or off-line as an effective sample preparation method specifically designed to address
interference problems.
4.4 Physical interferences are associated with the sample nebulization and transport
processes as well as with ion-transmission efficiencies. Nebulization and transport processes can
be affected if a matrix component causes a change in surface tension or viscosity. Changes in
matrix composition can cause significant signal suppression or enhancement (Ref. S). Dissolved
solids can deposit on the nebulizer tip of a pneumatic nebulizer and on the interface skimmers
(reducing the orifice size and the instrument performance). Total solid levels below 0.2% (2,000
mg/L) have been currently recommended (Ref. 10) to minimize solid deposition. An internal
standard can be used to correct for physical interferences, if it is carefully matched to the analyte so
that the two elements are similarly affected by matrix changes (Ref. 11). When intolerable physical
interferences are present in a sample, a significant suppression of the internal standard signals (to
less than 30 % of the signals in the calibrations standard) will be observed. Dilution of the sample
fivefold (1+4) will usually eliminate the problem (see Sec. 9.3).
4.5 Memory interferences or carry-over can occur when there are large concentration
differences between samples or standards which are analyzed sequentially. Sample deposition on
the sampler and skimmer cones, spray chamber design, and the type of nebulizer affect the extent
of the memory interferences which are observed. The rinse period between samples must be long
enough to eliminate significant memory interference.
5.0 SAFETY
Refer to Chapter Three for a discussion on safety related references and issues.
6.0 EQUIPMENT AND SUPPLIES
6.1 Inductively coupled plasma-mass spectrometer
6.1.1 A system capable of providing resolution, better than or equal to 1.0 amu at
10% peak height is required. The system must have a mass range from at least 6 to 240
amu and a data system that allows corrections for isobaric interferences and the application
of the internal standard technique. Use of a mass-flow controller for the nebulizer argon and
a peristaltic pump for the sample solution are recommended.
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6.1.2 Argon gas supply: high-purity grade (99.99%).
7.0 REAGENTS AND STANDARDS
7.1 Acids used in the preparation of standards and for sample processing must be of
high purity. Redistilled acids are recommended because of the high sensitivity of ICP-MS. Nitric
acid at less than 2 per cent (v/v) is required for ICP-MS to minimize damage to the interface and to
minimize isobaric molecular-ion interferences with the analytes. Many more molecular-ion
interferences are observed when hydrochloric and sulfuric acids are used (References 3 and 4).
Concentrations of antimony and silver between 50-500 pg/L require 1% (v/v) HCI for stability; for
concentrations above 500 pg/L Ag, additional HCI will be needed. Consequently, accuracy of
analytes requiring significant chloride molecular ion corrections (such as As and V) will degrade.
7.2 Reagent water All references to water in the method refer to reagent water unless
otherwise specified. Refer to Chapter One for a definition of reagent water.
7.3 Standard stock solutions for each analyte may be purchased or prepared from
ultra-high purity grade chemicals or metals (99.99 or greater purity ). See Method 6010 for
instructions on preparing standard solutions from solids.
7.3.1 Bismuth internal standard stock solution (1 mL = 100 pg Bi): Dissolve
0.1115 g Bi203 in a minimum amount of dilute HN03. Add 10 mL cone, HN03 and dilute to
1,000 mL with reagent water,
7.3.2 Hoimium internal standard stock solution (1 mL = 100 pg Ho): Dissolve
0.1757 g Ho2(C03)2-5H20 in 10 mL reagent water and 10 mL HN03. After dissolution is
complete, warm the solution to degas. Add 10 mL conc. HNOs and dilute to 1,000 mL with
reagent water.
7.3.3 Indium internal standard stock solution (1 mL = 100 pg In): Dissolve 0.1000 g
indium metal in 10 mL conc. HNOs. Dilute to 1,000 mL with reagent water.
7.3.4 Lithium internal standard stock solution (1 mL = 100 pg ®Li): Dissolve
0.6312 g 95-atom-% 6Li, Li2C03 in 10 mL of reagent water and 10 mL HNO3. After
dissolution is complete, warm the solution to degas. Add 10 mL conc. HN03 and dilute to
1,000 mL with reagent water.
7.3.5 Rhodium internal standard stock solution (1 mL = 100 pg Rh): Dissolve
0.3593 g ammonium hexachlororhodate (III) (NH^RhClg in 10 mL reagent water. Add
100 mL conc. HCI and dilute to 1,000 mL with reagent water.
7.3.6 Scandium internal standard stock solution (1 mL = 100 pg Sc): Dissolve
0.15343 g SCjOg in 10 mL (1+1) hot HN03. Add 5 mL conc. HN03and dilute to 1,000 mL
with reagent water.
7.3.7 Terbium internal standard stock solution (1 mL = 100 pg Tb): Dissolve
0.1828 g Tb2(C03)3-5H20 in 10 mL (1+1) HN03. After dissolution is complete, warm the
solution to degas. Add 5 mL conc. HN03 and dilute to 1,000 mL with reagent water.
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7.3.8 Yttrium internal standard stock solution (1 mL= 100|jg Y): Dissolve 0.2316 g
Y2(C03)3.3H20 in 10 mL (1+1) HNOa. Add 5 mL conc. HNOa and dilute to 1,000 mL with
reagent water.
7.3.9 Titanium interference stock solution (1 mL - 100 yg Ti): Dissolve 0.4133 g
(NH4)2TiF8 in reagent water. Add 2 drops conc. HF and dilute to 1,000 mL with reagent
water.
7.3.10 Molybdenum interference stock solution (1 mL = 100 pg Mo): Dissolve
0.2043 g (NH4)2Mo04 in reagent water. Dilute to 1,000 mL with reagent water.
7.3.11 Gold preservative stock solution for mercury (1 mL = 100 pg): Recommend
purchasing as high purity prepared solution of AuCI3 in dilute hydrochloric add matrix.
7.4 Mixed calibration standard solutions are prepared by diluting the stock-standard
solutions to levels in the linear range for the instrument in a solvent consisting of 1 percent (v/v)
HNOjin reagent water. The calibration standard solutions must contain a suitable concentration of
an appropriate internal standard for each analyte. internal standards may be added on-line at the
time of analysis using a second channel of the peristaltic pump and an appropriate mixing manifold.
Generally, an internal standard should be no more than 50 amu removed from the analyte.
Recommended internal standards include 6Li, ^Sc, 89Y, 103Rh, 115In, 1S9Tb, 169Ho, and ^Bi. Prior to
preparing the mixed standards, each stock solution must be analyzed separately to determine
possible spectral interferences or the presence of impurities. Care must be taken when preparing
the mixed standards that the elements are compatible and stable. Transfer the mixed standard
solutions to freshly acid-cleaned FEP fluorocarbon bottles for storage. Fresh mixed standards must
be prepared as needed with the realization that concentrations can change on aging. Calibration
standards must be initially verified using a quality control standard (see Section 7.7).
7.5 Blanks: Three types of blanks are required for the analysis. The calibration blank is
used in establishing the calibration curve. The preparation blank is used to monitor for possible
contamination resulting from the sample preparation procedure. The rinse blank is used to flush the
system between all samples and standards.
7.5.1 The calibration blank consists of the same concentrations) of the same
acid(s) used to prepare the final dilution of the calibrating solutions of the analytes [often 1
percent HNO, (v/v) in reagent water] along with the selected concentrations of internal
standards such that there is an appropriate internal standard element for each of the
analytes. Use of HCI for antimony and silver is cited in Section 7.1.
7.5.2 The preparation (or reagent) blank must be carried through the complete
preparation procedure and contain the same volumes of reagents as the sample solutions.
7.5.3 The rinse blank consists of 1 to 2 percent HN03 (v/v) in reagent water.
Prepare a sufficient quantity to flush the system between standards and samples. If mercury
is to be analyzed, the rinse blank should also contain 2 pg/mL (ppm) AuCI3 solution.
7.6 The interference check solution (ICS) is prepared to contain known concentrations
of interfering elements that will demonstrate the magnitude of interferences and provide an adequate
test of any corrections. Chloride in the ICS provides a means to evaluate software corrections for
chloride-related interferences such as ^Cl16©* on 51V* and ^Ar^cr on 75As\ Iron is used to
demonstrate adequate resolution of the spectrometer for the determination of manganese.
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Molybdenum serves to indicate oxide effects on cadmium isotopes. The other components are
present to evaluate the ability of the measurement system to correct for various molecular-ion
isobaric interferences. The ICS is used to verify that the interference levels are corrected by the data
system within quality control limits.
NOTE: The final ICS solution concentrations in Table 1 are intended to evaluate corrections
for known interferences on only the analytes in Sec. 1.2. If Method 6020 is used to
determine an element not listed in Sec. 1.2, it is the responsibility of the analyst to modify
the ICS solutions, or prepare an alternative ICS solution, to allow adequate verification of
correction of interferences on the unlisted element (see Section 9.4).
7.6.1 These solutions must be prepared from ultra-pure reagents. They can be
obtained commercially or prepared by the following procedure.
7.6.1.1 Mixed ICS solution I may be prepared by adding 13.903 g
AI(N03)3*9H20, 2.498 g CaC03 (dried at 180 °C for 1 hour before weighing), 1.000 g
Fe, 1.658 g MgO, 2.305 g Na2C03, and 1.767 g KjCOs to 25 mL of reagent water.
Slowly add 40 mL of (1+1) HN03. After dissolution is complete, warm the solution
to degas. Cool and dilute to 1,000 mL with reagent water.
7.6.1.2 Mixed ICS solution II may be prepared by slowly adding 7.444 g 85
% H3P04i 6.373 g 96% H2S04, 40.024 g 37% HCI, and 10.664 g citric acid C607H8
to 100 mL of reagent water. Dilute to 1,000 mL with reagent water.
7.6.1.3 Mixed ICS solution III may be prepared by adding 1.00 mL each of
100-pg/mL arsenic, cadmium, selenium, chromium, cobalt, copper, manganese,
nickel, silver, vanadium, and zinc stock solutions to about 50 mL reagent water. Add
2.0 mL concentrated HN03, and dilute to 100.0 mL with reagent water.
7.6.1.4 Working ICS Solutions
7.6.1.4.1 ICS-A may be prepared by adding 10.0 mL of mixed ICS
solution I (Sec. 7.6.1.1), 2.0 mL each of 100-yg/mL titanium stock solution
(Sec. 7.3.9) and molybdenum stock solution (Sec. 7.3.10), and 5.0 mL of
mixed ICS solution II (Sec. 7.6.1.2). Dilute to 100 mL with reagent water.
ICS solution A must be prepared fresh weekly.
7.6.1.4.2 ICS-AB may be prepared by adding 10.0 mL of mixed ICS
solution I (Sec. 7.6.1.1), 2.0 mL each of 100-pg/mL titanium stock solution
(Sec. 7.3.9) and molybdenum stock solution (Sec. 7.3.10), 5.0 mL of mixed
ICS solution II (Sec. 7.6.1.2), and 2.0 mL of Mixed ICS solution III (Sec.
7.6.1.3). Dilute to 100 mL with reagent water. Although the ICS solution AB
must be prepared fresh weekly, the analyst should be aware that the solution
may precipitate silver more quickly.
7.7 The quality control standard is the initial calibration verification solution (ICV), which
must be prepared in the same acid matrix as the calibration standards. This solution must be an
independent standard near the midpoint of the linear range at a concentration other than that used
for instrument calibration. An independent standard is defined as a standard composed of the
analytes from a source different from those used in the standards for instrument calibration.
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7.8 Mass spectrometer tuning solution. A solution containing elements representing all
of the mass regions of interest (for example, 10 pg/L of Li, Co, In, and Tl) must be prepared to verity
that the resolution and mass calibration of the instalment are within the required specifications (see
Section 10.1). This solution is also used to verify that the instrument has reached thermal stability
(see Section 11.4).
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 Sample collodion procedures should address the considerations described in Chapter
Nine.
8.2 See the introductory material in Chapter Three, Inorganic Analytes, for information
on sample handling, storage, holding times and preservation. Only polyethylene or fluorocarbon
(TFE or PFA) containers are recommended for use in this method.
9.0 QUALITY CONTROL
9.1 All quality control data should be maintained and be available for easy reference or
inspection.
9.2 Instrument detection limits (IDLs) in pg/L can be estimated by calculating the average
of the standard deviations of three runs on three non-consecutive days from the analysis of a
reagent blank solution with seven consecutive measurements per day. Each measurement must
be performed as though it were a separate analytical sample (i.e., each measurement must be
followed by a rinse and/or any other procedure normally performed between the analysis of separate
samples). IDLs must be determined at least every three months and kept with the instrument log
book. Refer to Chapter One for additional guidance.
9.3 The intensities of all internal standards must be monitored for every analysis. If the
intensity of any internal standard in a sample falls below 30 percent of the intensity of that internal
standard in the initial calibration standard, a significant matrix effect must be suspected. Under
these conditions, the detection limit has degraded and the correction ability of the internal
standardization technique becomes questionable. The following procedure is followed: First, make
sure the instrument Has not just drifted by observing the internal standard intensities in the nearest
dean matrix (calibration blank, Section 7.5.1). If the low internal standard intensities are also seen
in the nearest calibration Wank, terminate the analysis, correct the problem, recalibrate, verify the
new calibration, and reanalyze the affected samples. If drift has not occurred, matrix effects need
to be removed by dilution of the affected sample. The sample must be diluted fivefold (1+4) and
reanalyzed with the addition of appropriate amounts of internal standards. If the first dilution does
not eliminate the problem, this procedure must be repeated until the internal-standard intensities rise
above the 30 percent limit. Reported results must be corrected for all dilutions.
9.4 To obtain analyte data of known quality, it is necessary to measure more than the
analytes of interest in order to apply corrections or to determine whether interference corrections are
necessary. For example, tungsten oxide molecular® can be very difficult to distinguish from mercury
isotopes. If the concentrations of interference sources (such as C, CI, Mo, Zr, W) are such that, at
the correction factor, the analyte is less than the limit of quantification and the concentration of
interferents are insignificant, then the data may go uncorrected. Note that monitoring the
interference sources does not necessarily require monitoring the interferant itself, but that a
molecular species may be monitored to indicate the presence of the interferent. When correction
equations are used, all QC criteria must also be met. Extensive QC for interference corrections are
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required at all times. The monitored masses must include those elements whose hydrogen, oxygen,
hydroxyl, chlorine, nitrogen, cartoon and sulfur molecular ions could impact the analytes of interest.
Unsuspected interferences may be detected by adding pure major matrix components to a sample
to observe any impact on the analyte signals. When an interference source is present, the sample
elements impacted must be flagged to indicate (a) the percentage interference correction applied
to the data or (b) an uncorrected interference by virtue of the elemental equation used for
quantitation. The isotope proportions for an element or molecular-ion cluster provide information
useful for quality assurance.
NOTE: Only isobaric elemental, molecular, and doubly charged interference corrections
which use the observed isotopio-response ratios or parent-to-oxide ratios (provided an oxide
internal standard is used as described in Section 4.2) for each instrument system are
acceptable connections for use in Method 6020.
9.5 Dilution test (serial dilution): If the analyte concentration is within the linear dynamic
range of the instrument and sufficiently high (minimally, a factor of at least 100 times greater than
the concentration in the reagent blank, refer to Section 7.5.2), an analysis of a fivefold (1+4) dilution
must agree within ± 10% of the original determination. If not, an interference effect must be
suspected. One dilution test must be included for each twenty samples (or less) of each matrix in
a batch.
9.6 Post-digestion spike addition: An analyte spike added to a portion of a prepared
sample, or its dilution, should be recovered to within 75 to 125 percent of the known value or within
the laboratory derived acceptance criteria. The spike addition should be based on the indigenous
concentration of each element of interest in the sample. If the spike is not recovered within the
specified limits, the sample must be diluted and reanalyzed to compensate for the matrix effect.
Results must agree to within 10% of the original determination. The use of a standard-addition
analysis procedure may also be used to compensate for this effect (refer to Method 7000).
9.7 A laboratory control sample (LCS) should be analyzed for each analyte using the
same sample preparations, analytical methods and QA/QC procedures employed for the test
samples. One LCS should be prepared and analyzed for each sample batch at a frequency of one
LCS for each 20 samples or less.
9.8 Check the instrument calibration by analyzing appropriate quality control solutions as
follows:
9.8.1 Check instrument calibration using a calibration blank (Section 7.5.1) and the
initial calibration verification solution (Sections 7.7 and 11.6).
9.8.2 Verify calibration at a frequency of every 10 analytical samples with the
instrument check standard (Section 7.6) and the calibration blank (Section 7.5.1). These
solutions must also be analyzed for each analyte at the beginning of the analysis and after
the last sample.
9.8.3 The results of the initial calibration verification solution and the instrument
check stahdard must agree within ± 10% of the expected value. If not, terminate the
analysis, correct the problem, and recalibrate the instrument. Any sample analyzed under
an out-of-control calibration must be reanalyzed .
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9.8.4 The results of the calibration blank must be less than 3 times the current IDL
for each element If this is not the case, the reason for the out-of-control condition must be
found and corrected, and affected samples must be reanalyzed. If the laboratory consistently
has concentrations greater than 3 times the IDL, the IDL may be indicative of an estimated
IDL and should to re-evaluated.
9.9 Verify the magnitude of elemental and molecular-ion isobaric interferences and the
adequacy of any corrections at the beginning of an analytical run or once every 12 hours, whichever
is more frequent. Do this by analyzing the interference check solutions A and AB._ The analyst
should be aware that precipitation from solution AB may occur with some elements, specifically
silver. Refer to Section 4.0 for a discussion on interferences and potential solutions to those
interferences if additional guidance is needed.
9.10 Analyze one duplicate sample for every matrix in a batch at a frequency of one matrix
duplicate for every 20 samples.
9.10.1 The relative percent difference (RPD) between duplicate determinations must
be calculated as follows:
Pi - D2 |
RPD = x 100
(D1 + D2)/2
where:
RPD - relative percent difference.
D, = first sample value.
D2 = second sample value (duplicate)
A control limit of 20% RPD should not be exceeded for analyte values greater than 100 times
the instrumental detection limit. If this limit is exceeded, the reason for the out-of-control
situation must be found and corrected, and any samples analyzed during the out-of-control
condition must be reanalyzed.
9.11 Ultra-trace analysis requires the use of dean chemistry. Several suggestions for
reduction on the analytical blank are provided in Chapter Three.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Conduct mass calibration and resolution checks in the mass regions of interest. The
mass calibration and resolution parameters are required criteria which must be met prior to any
samples being analyzed. If the mass calibration differs more than 0.1 amu from the true value, then
the mass calibration must be adjusted to the correct value. The resolution must also be verified to
be less than 0.9 amu full width at 10 percent peak height.
10.2 Calibrate the instrument for the analytes of interest (recommended isotopes for the
analytes in Sec. 1.2 are provided in Table 2), using the calibration blank and at least a single initial
calibration standard according to the instalment manufacturer's procedure. Flush the system with
the rinse blank (Sec. 7.5.3) between each standard solution. Use the average of at least three
integrations for both calibration and sample analyses.
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NOTE: Analysts have noted improved performance in calibration stability if the instrument
is exposed to the interference check solution after cleaning sampler and skimmer cones.
Improved performance is also realized if the instrument is allowed to rinse for 5 or 10 minutes
before the calibration blank is run.
10.3 All masses which could affect data quality should be monitored to determine potential
effects from matrix components on the analyte peaks. Hie recommended isotopes to be monitored
are listed in Table 2.
10.4 Immediately after the calibration has been established, the calibration must be verified
and documented for every analyte by the analysis of the calibration verification solution (Section 7.7).
When measurements exceed ± 10% of the accepted value, the analyses must be terminated, the
problem corrected, the instrument recalibrated, and the new calibration verified. Any samples
analyzed under an out-of-control calibration must be reanalyzed. During the course of an analytical
run, the instrument may be "resloped" or recalibrated to correct for instrument drift but resloping must
not be used as an alternative to reanalyzing samples following an unacceptable QC sample, such
as a CCV. A recalibration must then be followed immediately by a new analysis of a CCV and CCB
before any further samples may be analyzed.
11.0 PROCEDURE
11.1 Solubilization and digestion procedures are presented in Chapter Three (e.g.,
Methods 3005 - 3052).
NOTE: If mercury is to be analyzed, the digestion procedure must use mixed nitric and
hydrochloric acids through all steps of the digestion. Mercury will be lost if the sample is
digested when hydrochloric acid is not present. If it has not already been added to the
sample as a preservative, Au should be added to give a final concentration of 2 mg/L (use
2.0 mL of 5.3.11 per 100 mL of sample) to preserve the mercury and to prevent it from
plating out in the sample introduction system.
11.2 Initiate appropriate operating configuration of the instruments computer according to
the instrument manufacturer's instructions.
11.3 Set up the instrument with the proper operating parameters according to the
instrument manufacturer's instructions.
11.4 Operating conditions: The analyst should follow the instructions provided by the
instrument manufacturer. Allow at least 30 minutes for the instrument to equilibrate before analyzing
any samples. This must be verified by analyzing a tuning solution (Section 7.8) at least four times
with relative standard deviations of < 5% for the analytes contained in the tuning solution.
NOTE: The instrument should have features that protect "itself from high ion currents. If not,
precautions must be taken to protect the detector from high ion currents. A channel electron
multiplier or active film multiplier suffer from fatigue after being exposed to high ion currents.
This fatigue can last from several seconds to hours depending on the extent of exposure.
During this time period, response factors are constantly changing, which invalidates the
calibration curve, causes instability, and invalidates sample analyses.
11.5 Calibrate the instrument following the procedure outlined in Section 10.0.
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11.6 Flush the system with the rinse blank solution (Sec. 7.5.3) until the signal levels
return to the DQO or method's levels of quantitation (usually about 30 seconds) before the analysis
of each sample (see Section 10.3). Nebulize each sample until a steady-state signal is achieved
(usually about 30 seconds) prior to collecting data. Analyze the calibration verification solution
(Section 7.6) and the calibration blank (Section 7.5.1) at a frequency of at least once every 10
analytical samples. Flow-injection systems may be used as long as they can meet the performance
criteria of this method.
11.7 Dilute and reanalyze samples that are more concentrated than the linear range for
an analyte (or species needed for a correction) or measure an alternate but less-abundant isotope.
The linearity at the alternate mass must be confirmed by appropriate calibration (see Sec. 10.2 and
10.4). Alternatively apply solid phase chelation chromatography to eliminate the matrix as described
in Sec. 4.3.
12.0 DATA ANALYSIS AND CALCULATIONS
12.1 The quantitative values shall be reported in appropriate units, such as micrograms
per liter (pg/L) for aqueous samples and milligrams per kilogram (mg/kg) for solid samples. If
dilutions were performed, the appropriate corrections must be applied to the sample values.
12.1.1 If appropriate, or required, calculate results for solids on a dry-weight basis
as follows:
(1) A separate determination of percent solids must be performed.
(2) The concentrations determined in the digest are to be reported on the
basis of the dry weight of the sample.
Concentration {dry weight)(mg/kg) = 9* ^
W x s
Where,
C = Digest Concentration (mg/L)
V = Final volume in liters after sample preparation
W = Weight in kg of wet sample
S= % Solids
100
Calculations must include appropriate interference corrections (see Section 4.2 for
examples), internal-standard normalization, and the summation of signals at 206,207, and
208 m/z for lead (to compensate for any differences in the abundances of these isotopes
between samples and standards).
13.0 METHOD PERFORMANCE
13.1 In an EPA multi-laboratory study (Ref. 12), twelve laboratories applied the ICP-MS
technique to both aqueous and solid samples. Table 3 summarizes the method performance data
for aqueous samples. Performance data for solid samples are provided in Table 4.
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13.2 Table 5 summarizes the method performance data for aqueous and sea water
samples with interfering elements removed and samples preconcentrated prior to analysis. Table
6 summarizes the performance data for a simulated drinking water standard.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the
quantity and/or toxicity of waste at the point of generation. Numerous opportunities for pollution
prevention exist in laboratory operation. The EPA has established a preferred hierarchy of
environmental management techniques that places pollution prevention as the management option
of first choice. Whenever feasible, laboratory personnel should use pollution prevention techniques
to address their waste generation. When wastes cannot be feasibly reduced at the source, the
Agency recommends recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Less is Better Laboratory Chemical management for Waste Reduction
available from the American Chemical Society's Department of Government Relations and Science
Policy, 1155 16th St., N.W. Washington, D.C. 20036, (202) 872-4477.
15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society at the address listed in Sec. 14.2.
16.0 REFERENCES
1. Horlick, G„ et al., Spectrochim. Acta 40B, 1555 (1985).
2. Gray, A.L., Spectrochim. Acta 40B, 1525 (1985); 41B, 151 (1986).
3. Tan, S.H., and Horiick, G., Appl. Spectrosc. 40, 445 (1986).
4. Vaughan, M.A., and Horlick, G., Appl. Spectrosc. 40,434 (1986).
5. Holden, N.E., "Table of the Isotopes," in Lide, D.R., Ed., CRC Handbook of Chemistry and
Physics, 74th Ed., CRC Press, Boca Raton, FL, 1993.
6. Hinners, T.A., Heithmar, E., Rissmann, E., and Smith, D., Winter Conference on Plasma
Spectrochemistry, Abstract THP18; p. 237, San Diego, CA (1994).
7. Lichte, F.E., et al., Anal. Chem. 59,1150 (1987).
8. Evans E.H., and Ebdon, L., J. Anal. At. Spectrom. 4, 299 (1989).
9. Beauchemin, D., et al., Spectrochim. Acta 42B, 467 (1987).
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10. Houk, R.S., Anal. Chem. 58,97A (1986),
11. Thompson, J.J., and Houk, R.S., Appl. Spectrosc. 41, 801 (1987).
12. Newbeny, W.R., Butler, L.C., Hurd, M.L., Laing, G.A., Siapanian, M.A., Aleckson, K.A., Dobb,
D.E., Rowan, J.T., and Garner, F.C., Final Report of the Multi-Laboratory Evaluation of Method
6020 CLP-M Inductively Coupled Plasma-Mass Spectrometry (1989).
13. Taylor, Daniel B., Kingston, H.M., Nogay, Donald J., Kolfer, Dagmar, and Hutton, Robert. "On-
Line Solid-phase Chelation for the Determination of Eight Metals in Environmental Waters by
Inductively Coupled Plasma Mass Spectrometry".
14. Kingston, H. M., Siriraks, A., and Riviello, J. M., Patent Number 5,126,272, "A Method and
Apparatus for Detecting Transition and Rare Earth Elements in a Matrix", U.S. Patent, Filed U.S.
Patent Office, March 1989, 31 pages, Granted June 30,1992, Patent held by US Government.
15. Kingston, H. M., Siriraks, A., and Riviello, J. M., Patent Number 5,244,634 , "A Method and
Apparatus for Detecting Transition and Rare Earth Elements in a Matrix", U.S. Patent, Filed U.S.
Patent Office, March 1989,31 pages, Granted Sept. 14, 1993, Patent held by US Government.
16. Dobb, David E., Rowan, J.T., and Cardenas, D., Lockheed Environmental Systems and
Technologies Co., Las Vegas, NV; and Butler, L.C., and Heithman, E.M., U.S.EPA, Las Vegas,
NV; "Determination of Mercury by I CP-MS".
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 through 7, and a flow diagram of the method procedure.
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TABLE 1
RECOMMENDED INTERFERENCE CHECK SAMPLE COMPONENTS
AND CONCENTRATIONS
Solution Solution A Solution AB
Component Concentration Concentration
(mg/L) (mg/L)
Al
100.0
100.0
Ca
300.0
300.0
Fe
250.0
250.0
Mg
100.0
100.0
Na
250.0
250.0
P
100.0
100.0
K
100.0
100.0
S
100.0
100.0
C
200.0
200.0
CI
2000.0
2000.0
Mo
2.0
2.0
Ti
2.0
2.0
As
0.0
0.100
Cd
0.0
0.100
Cr
0.0
0.200
Co
0.0
0.200
Cu
0.0
0.200
Mn
0.0
0.200
Hg
0.0
0.020
Ni
0.0
0.200
Se
0.0
0.100
Ag
0.0
0.050
V
0.0
0.200
Zn
0.0
0.100
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TABLE 2
RECOMMENDED ISOTOPES FOR SELECTED ELEMENTS
Element of Interest
Mass
Aluminum
27
Antimony
121, 123
Arsenic
11
Barium
138, 137,136, 135, 134
Beryllium
9
Bismuth (IS)
209
Cadmium
114,112,111,110,113,116,106
Calcium (1)
42, 43, 46, 48
Chlorine (1)
35, 37, (77, 82)a
Chromium
52, S3, 50, 54
Cobalt
59
Copper
63,65
Holmium (IS)
165
Indium (IS)
115. 113
Iron (1)
56, 54, 57, 58
Lanthanum (1)
139
Lead
208, 207, 206, 204
Lithium (IS)
6*77
Magnesium (1)
24, 25, 26
Manganese
55
Mercury
202, 200, 199, 201
Molybdenum (1)
98, 96, 92, 97, 94, (108)a
Nickel
58, 60. 62, 61. 64
Potassium (1)
39
Rhodium (IS)
103
Scandium (IS)
45
Selenium
80, 78, 82, 76, J7, 74
Silver
107. 109
Sodium (1)
23
Terbium (IS)
159
Thallium
205, 203
Vanadium
51. §S
Tin (1)
120,118
Yttrium (IS)
89
Zinc
, 64, 66, 68, 67, 70
NOTE: Method 6020 is recommended for only those analytes listed in Sec.12. Other elements are included
in this table because they are potential interferents (labeled I) in the determination of recommended analytes,
or because they are commonly used internal standards (labeled IS), isotopes are listed in descending order
of natural abundance. The most generally useful Isotopes are underlined and in boldface, although certain
matrices may require the use of alternative isotopes.
* These masses are also useful for interference correction (Section 4.2).
b Internal standard must be enriched in the *U isotope. This minimizes interference from indigenous lithium.
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TABLE 3
ICP-MS MULTI-LABORATORY PRECISION AND ACCURACY DATA FOR AQUEOUS SOLUTIONS
Element
Comparability*
Range
%RSD
Range
n*
Se
Aluminum
95 -100
11-14
14 - 14
4
Antimony
d
5.0 - 7.6
16-16
3
Arsenic
97-114
7.1-48
12-14
4
Barium
91-99
4.3 - 9.0
16-16
5
Beryllium
103-107
8.6-14
13-14
3
Cadmium
98-102
4.6 - 7.2
18-20
3
Calcium
99-107
5.7 - 23
17-18
5
Chromium
95-105
13-27
16-18
4
Cobalt
101 - 104
8.2 - 8.5
18-18
3
Copper
85 -101
6.1-27
17-18
5
Iron
91 - 900
11 -150'
10-12
5
Lead
71 - 137
11-23
17- 18
6
Magnesium
98 - 102
10-15
16-16
5
Manganese
95-101
8.8-15
18-18
4
Nickel
98-101
6.1 - 6.7
18-18
2
Potassium
101 - 114
9.9 -19
11 - 12
5
Selenium
102-107
15-25
12-12
3
Silver
104-105
5.2 - 7.7
13- 16
2
Sodium
82 - 104
24-43
9-10
5
Thallium
88-97
9.7-12
18-18
3
Vanadium
107 - 142
23-68
8-13
3
Zinc
93 -102
6.8 -17
16-18
5
Data obtained from reference 12.
8 Comparability refers to the percent agreement of mean ICP-MS values to those of the reference
technique (ICP-AES or GFAA).
b N is the range of the number of ICP-MS measurements where the analyte values exceed the limit
of quantitation (3.3 times the average IDL value). A larger number gives a more reliable comparison.
c S is the number of samples with results greater than the limit of quantitation.
d No comparability values are provided for antimony because of evidence that the reference data is
affected by an interference.
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TABLE 4
iCP-MS MULTI-LABORATORY PRECISION AND ACCURACY DATA FOR SOLID MATRICES

Comparability"
%RSD

Element
Range
Range
Nb
Sc
Aluminum
83 -101
11-39
13-14
7
Antimony
d
12-21
15-16
2
Arsenic
79 -102
12-23
16-16
7
Barium
100 -102
4.3 -17
15-16
7
Beryllium
50-87
19-34
12-- 14
5
Cadmium
93-100
6.2-25
19-20
5
Calcium
95-109
4.1-27
15-17
7
Chromium
77-98
11-32
17-18
7
Cobalt
43 -102
15-30
17-18
6
Copper
90-109
9.0-25
18- 18
7
Iron
87-99
6.7 - 21
12-12
7
Lead
90-104
5.9 - 28
15-18
7
Magnesium
89-111
7.6-37
15-16
7
Manganese
80-108
11 - 40
16-18
7
Nickel
87-117
9.2 - 29
16-18
7
Potassium
97 -137
11-62
10-12
5
Selenium
81
39
12
1
Silver
43-112
12-33
15-15
3
Sodium
100 - 146
14-77
CO
i
o
5
Thallium
91
33
18
1
Vanadium
83 -147
20-70
6-14
7
Zinc
84 -124
14-42
18-18
7
Data obtained from reference 12.
a Comparability refers to the percent agreement of mean ICP-MS values to those of the reference
technique.
b N is the range of the number of ICP-MS measurements where the analyte values exceed the limit
of quantitation (3.3 times the average IDL value).
c S is the number of samples with results greater than the limit of quantitation.
d No comparability values are provided for antimony because of evidence that the reference data is
affected by an interference.
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TABLE 5
METHOD PERFORMANCE DATA FOR AQUEOUS AND SEA WATER SAMPLES*
WITH INTERFERING ELEMENTS REMOVED
AND SAMPLES PRECONCENTRATED PRIOR TO ANALYSIS
CONCENTRATION (no/mLl8
ELEMENT
ISOTOPE
9.0 mL
27.0 mL
CERTFIED
Manganese
55
1.8±0.05
1.9±0.2
1.99±0.15
Nickel
58
0.32±0.018
0.32±0.04
0.30±0.04
Cobalt
59
0.033±0.002
0.028±0.003
0.025±0.006
Copper
63
0.68±0.03
0.63±0.03
0.68±0.04
Zinc
64
1.6±0.05
1.8±0.15
1.97±0.12
Copper
65
0.67±0.03
0.6±0.05
0.68±0.04
Zinc
66
1.6±0.06
1.8±0.2
1.97±0.12
Cadmium
112
0.020±0.0015
0.019±0.0018
0.019±0.004
Cadmium
114
0.020±0.0009
0.019±0.002
0.019±0.004
Lead
206
0.013±0.0009
0.019±0.0011
0.019±0.006
Lead
207
0.014±0.0005
0.019±0.004
0.019±0.006
Lead
208
0.014±0.0006
0.019±0.002
0.019±0.006
Data obtained from reference 12.
A The dilution of the sea-water during the adjustment of pH produced 10 mL samples containing 9 mL of sea-water and 30 mL samples
containing 27 mL of sea-water. Samples containing 9.0 mL of CASS-2, n=5; samples containing 27.0 mL of CASS-2, n=3.
B Concentration (ng/mL) ± 95% confidence limits.
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TABLE 6
ANALYSIS OF NIST SRM 1643b, TRACE METALS IN WATER*
CONCENTRATION (ng/mUB
ELEMENT
ISOTOPE
DETERMINED
CERTFIED
Manganese
55
3011.3
2812
Nickel
58
50±2
4913
Cobalt
59
27l1.3
2611
Nickel
SO
51 ±2
4913
Copper
63
23±1.0
21.910,4
Zinc
64
67±1.4
6612
Copper
65
22±0.9
21.910.4
Zinc
66
67±1.8
6612
Cadmium
111
2010.5
2011
Cadmium
112
19,910.3
2011
Cadmium
114
19.810.4
2011
Lead
206
2310.5
23.710.7
Lead
207
23.910.4
23.710.7
Lead
208
24,210.4
23.7l0,7
Data obtained from reference 12.
A 5.0 mL samples, n=5.
B Concentration (ng/mL) 1 95% confidence limits.
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TABLE 7
COMPARISON OF TOTAL MERCURY RESULTS IN HEAVILY CONTAMINATED SOILS
Soil Sample
Mercury in pg^9
ICP-MS
CVAA
1
27,8
29.2
2
442
376
3
64.7
58.2
4
339
589
5
281
454
6
23.8
21.4
7
217
183
8
157
129
9
1670
1360
10
73.5
64.8
11
2090
1830
12
96.4
85.8
13
1080
1190
14
294
258
15
3300
2850
16
301
281
17
2130
2020
18
247
226
19
2630
2080
Source: Reference 16.
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METHOD 6020A
INDUCTIVELY COUPLED PLASMA - MASS SPECTROMETRY
Start
Yes
/Arc \
/ results N
outside +/-
10% range of
actual
v value? /
No
Yes
No
Stop
7,11 Calculate
concentration.
7.8 Verify
calibration
with tCV,
7.4 Set
operating
conditions as
recommended.
7.3 Set up
and stabilize
instrument.
7.10 Dilute
extract.
7.9 Flush
system and
analyze
sample.
7.6 Calibrate
the instrument
for the analytes
of interest.
7.1 Use
appropriate
digestion
procedure
(Chapter Three.)
7.5 Perform
mass
calibration and
resolution
checks.
7.9 Analyze
check standard
and calibration
blank after each
10 samples.
7.7 Monitor
all masses that
could affect data
quality, as
recommended.
7.2 Initiate
operating
configuration of
instrument
computer.
Readjust
instrument per
manufacturers"
recorn mendations.
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METHOD 6200
FIELD PORTABLE X-RAY FLUORESCENCE SPECTROMETRY FOR THE
DETERMINATION OF ELEMENTAL CONCENTRATIONS IN SOIL AND SEDIMENT
1.0 SCOPE AND APPLICATION
1.1 Wis method is applicable to the in situ and intrusive analysis of the 26 analytes listed
in Table 1 for soil and sediment samples. Some common elements are not listed in Table 1 because
they are considered "light" elements that cannot be detected by field portable x-ray fluorescence
(FPXRF). They are: lithium, beryllium, sodium, magnesium, aluminum, silicon, and phosphorus.
Most of the analytes listed in Table 1 are of environmental concern, while a few others have
interference effects or change the elemental composition of the matrix, affecting quantitation of the
analytes of interest. Generally elements of atomic number 16 or greater can be detected and
quantitated by FPXRF.
1.2 Detection limits depend on several factors, the analyte of interest, the type of detector
used, the type of excitation source, the strength of the excitation source, count times used to
irradiate the sample, physical matrix effects, chemical matrix effects, and interelement spectral
interferences. General instrument detection limits for analytes of interest in environmental
applications are shown in Table 1. These detection limits apply to a clean matrix of quartz sand
(silicon dioxide) free of interelement spectral interferences using long (600-second) count times.
These detection limits are given for guidance only and will vary depending on the sample matrix,
which instrument is used, and operating conditions. A discussion of field performance-based
detection limits is presented in Section 13.4 of this method. The dean matrix and field performance-
based detection limits should be used for general planning purposes, and a third detection limit
discussed, based on the standard deviation around single measurements, should be used in
assessing data quality. This detection limit is discussed in Sections 9.7 and 11.3.
1.3 Use of this method is restricted to personnel either trained and knowledgeable in the
operation of an XRF instrument or under the supervision of a trained and knowledgeable individual.
This method is a screening method to be used with confirmatory analysis using EPA-approved
methods. This method's main strength is as a rapid field screening procedure. The method
detection limits (MDL) of FPXRF are above the toxicity characteristic regulatory level for most RCRA
analytes. If the precision, accuracy, and detection limits of FPXRF meet the data quality objectives
(DQOs) of your project, then XRF is a fast, powerful, cost effective technology for site
characterization.
2.0 SUMMARY OF METHOD
2.1 The FPXRF technologies described in this method use sealed radioisotope sources to
irradiate samples with x-rays. X-ray tubes are used to irradiate samples in the laboratory and are
beginning to be incorporated into field portable instruments. When a sample is irradiated with x-rays,
the source x-rays may undergo either scattering or absorption by sample atoms. This later process
is known as the photoelectric effect. When an atom absorbs the source x-rays, the incident radiation
dislodges electrons from the innermost shells of the atom, creating vacancies. The electron
vacancies are filled by electrons cascading in from outer electron shells. Electrons in outer shells
have higher energy states than inner shell electrons, and the outer shell electrons give off energy
as they cascade down into the inner shell vacancies. This rearrangement of electrons results in
emission of x-rays characteristic of the given atom. The emission of x-rays, in this manner, is termed
x-ray fluorescence.
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Three electron shells are generally involved in emission of x-rays during FPXRF analysis of
environmental samples: the K, L, and M shells. A typical emission pattern, also called an emission
spectrum, fa* a given metal has multiple intensity peaks generated from the emission of K, L, or M
shell electrons. The most commonly measured x-ray emissions are from the K and L shells; only
metals with an atomic number greater than 57 have measurable M shell emissions.
Each characteristic x-ray line is defined with the letter K, L, or M, which signifies which shell
had the original vacancy and by a subscript alpha (a) or beta (P), which indicates the higher shell
from which electrons fell to fill the vacancy and produce the x-ray. For example, a K„ line is
produced by a vacancy in the K shell filled by an L shell electron, whereas a Kg line is produced by
a vacancy in the K shell filled by an M shell electron. The transition is on average 6 to 7 times
more probable than the Kp transition; therefore, the line is approximately 7 times more intense
than the Kp line for a given element, making the K„ line the choice for quantitation purposes.
The K lines for a given element are the most energetic lines and are the preferred lines for
analysis. For a given atom, the x-rays emitted from L transitions are always less energetic than
those emitted from K transitions. Unlike the K lines, the main L emission lines (L^ and Lp) for an
element are of nearly equal intensity. The choice of one or the other depends on what interfering
element lines might be present. The L emission lines are useful for analyses involving dements of
atomic number (Z) 58 (cerium) through 92 (uranium).
An x-ray source can excite characteristic x-rays from an element only if the source energy is
greater than the absorption edge energy for the particular line group of the element, that is, the K
absorption edge, L absorption edge, or M absorption edge energy. The absorption edge energy is
somewhat greater than the corresponding line energy. Actually, the K absorption edge energy is
approximately the sum of the K, L, and M line energies of the particular element, and the L
absorption edge energy is approximately the sum of the L and M line energies. FPXRF is more
sensitive to an element with an absorption edge energy close to but less than the excitation energy
of the source. For example, when using a cadmium-109 source, which has an excitation energy of
22.1 klloelectron volts (keV), FPXRF would exhibit better sensitivity for zirconium which has a K line
energy of 15.7 keV than to chromium, which has a K line energy of 5.41 keV.
2.2 Under this method, inorganic analytes of interest are identified and quantitated using
a field portable energy-dispersive x-ray fluorescence spectrometer. Radiation from one or more
radioisotope sources or an electrically excited x-ray tube is used to generate characteristic x-ray
emissions from elements in a sample. Up to three sources may be used to irradiate a sample. Each
source emits a specific set of primary x-rays that excite a corresponding range of elements in a
sample. When more than one source can excite the element of interest, the source is selected
according to its excitation efficiency for the element of interest.
For measurement, the sample is positioned in front of the probe window. This can be done
in two manners using FPXRF instruments: in situ or intrusive. If operated in the in situ mode, the
probe window is placed in direct contact with the soil surface to be analyzed. When an FPXRF
instrument is operated in the intrusive mode, a soil or sediment sample must be collected, prepared,
and placed in a sample cup. The sample cup is then placed on. top of the window inside a protective
cover for analysis.
Sample analysis is then initiated by exposing the sample to primary radiation from the source.
Fluorescent and backscattered x-rays from the sample enter through the detector window and are
converted into electric pulses in the detector. The detector in FPXRF instruments is usually either
a solid-state detector or a gas-filled proportional counter. Within the detector, energies of the
characteristic x-rays are converted into a train of electric pulses, the amplitudes of which are linearly
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proportional to the 6n©rQy of 1h6 xn^ys. An 6i6ctronic ¦ i iultichsnndl snslyzsr ^MOA) me a s u r6 s the
pulse amplitudes, which is the basis of qualitative x-ray analysis. The number of counts at a given
energy per unit of time is representative of the element concentration in a sample and is the basis
for quantitative analysis. Most FPXRF instruments are menu-driven from software built into the units
or from personal computers (PC).
The measurement time of each source is user-selectable. Shorter source measurement times
(30 seconds) are generally used for initial screening and hot spot delineation, and longer
measurement times (up to 300 seconds) are typically used to meet higher precision and accuracy
requirements.
FPXRF instruments can be calibrated using the following methods: internally using
fundamental parameters determined by the manufacturer, empirically based on site-specific
calibration standards (SSCS), or based on Compton peak ratios. The Compton peak is produced
by backscattering of the source radiation. Some FPXRF instruments can be calibrated using multiple
methods.
3.0 DEFINITIONS
3.1 FPXRF: Field portable x-ray fluorescence.
3.2 MCA: Multichannel analyzer for measuring pulse amplitude.
3.3 SSCS: Site specific calibration standard.
3.4 £E: Fundamental parameter.
3.5 ROI: Region of interest.
3.6 SRM: Standard reference material. A standard containing certified amounts of metals
in soil or sediment.
3.7 sM: Electron Volt. A unit of energy equivalent to the amount of energy gained by an
electron passing through a potential difference of one volt.
3.8 Refer to Chapter One and Chapter Three for additional definitions.
4.0 INTERFERENCES
4.1 The total method error for FPXRF analysis is defined as the square root of the sum of
squares of both instrument precision and user- or application-related error. Generally, instrument
precision is the least significant source of error in FPXRF analysis. User- or application-related error
is generally more significant and varies with each site and method used. Some sources of
interference can be minimized or controlled by the instrument operator, but others cannot. Common
sources of user- or application-related error are discussed below.
4.2 Physical matrix effects result from variations in the physical character of the sample.
These variations may include such parameters as particle size, uniformity, homogeneity, and surface
condition. For example, if any analyte exists in the form of very fine particles in a coarser-grained
matrix, the analyte's concentration measured by the FPXRF will vary depending on how fine particles
are distributed within the coarser-grained matrix. If the fine particles "settle" to the bottom of the
sample cup, the analyte concentration measurement will be higher than if the fine particles are not
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mixed in well and stay on top of the coarser-grained particles in the sample cup. One way to reduce
such error is to grind and sieve all soil samples to a uniform particle size thus reducing sample-to-
sample particle size variability. Homogeneity is always a concern when dealing with soil samples.
Every effort should be made to thoroughly mix and homogenize soil samples before analysis. Field
studies have shown heterogeneity of the sample generally has the largest impact on comparability
with confirmatory samples.
4.3 Moisture content may affect the accuracy of analysis of soil and sediment sample
analyses. When the moisture content is between 5 and 20 percent, the overall error from moisture
may be minimal. However, moisture content may be a major source of error when analyzing
samples of surface soil or sediment that are saturated with water. This error can be minimized by
drying the samples in a convection or toaster oven. Microwave drying is not recommended because
field studies have shown that microwave drying can increase variability between FPXRF data and
confirmatory analysis and because metal fragments in the sample can cause arcing to occur in a
microwave.
4.4 Inconsistent positioning of samples in front of the probe window is a potential source
of error because the x-ray signal decreases as the distance from the radioactive source increases.
This error is minimized by maintaining the same distance between the window and each sample.
For the best results, the window of the probe should be in direct contact with the sample, which
means that the sample should be flat and smooth to provide a good contact surface.
4.5 Chemical matrix effects result from differences in the concentrations of interfering
elements. These effects occur as either spectral interferences (peak overlaps) or as x-ray
absorption and enhancement phenomena. Both effects are common in soils contaminated with
heavy metals. As examples of absorption and enhancement effects; iron (Fe) tends to absorb
copper (Cu) x-rays, reducing the intensity of the Cu measured by the detector, while chromium (Cr)
will be enhanced at the expense of Fe because the absorption edge of Cr is slightly lower in energy
than the fluorescent peak of iron. The effects can be corrected mathematically through the use of
fundamental parameter (FP) coefficients. The effects also can be compensated for using SSCS,
which contain all the elements present on site that can interfere with one another.
4.6 When present in a sample, certain x-ray lines from different elements can be very close
in energy and, therefore, can cause interference by producing a severely overlapped spectrum. The
degree to which a detector can resolve the two different peaks depends on the energy resolution of
the detector. If the energy difference between the two peaks in electron volts is less than the
resolution of the detector in electron volts, then the detector will not be able to fully resolve the
peaks.
. The most common spectrum overlaps involve the Kg line of element Z-1 with the K,, line of
element Z. This is called the K„/Kp interference. Because the K^Kp intensity ratio for a given
element usually is about 7:1, the interfering element, Z-1, must be present at large concentrations
to cause a problem. Two examples of this type of spectral interference involve the presence of large
concentrations of vanadium (V) when attempting to measure Cr or the presence of large
concentrations of Fe when attempting to measure cobalt (Co). The V K,, and Kp energies are 4.95
and 5.43 keV, respectively, and the Cr energy is 5.41 keV. The Fe and Kg energies are 6.40
and 7.06 keV, respectively, and the Co Kg energy is 6.92 keV. The difference between the V Kg and
CrK,, energies is 20 eSf, and the difference between the Fe and the Co Kg energies is 140 eV.
The resolution of the highest-resolution detectors in FPXRF instruments is 170 eV. Therefore, large
amounts of V and Fe will interfere with quantitation of Cror Co, respectively. The presence of Fe
is a frequent problem because it is often found in soils at tens of thousands of parts per million
(ppm).
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4.7 Other interferences can arise from K/L, K/M, and L/M fine overlaps, although these
overlaps are less common. Examples of such overlap involve arsenic (As) Ka/lead (Pb) L„ and sulfur
(S) Kc/Pb In the As/Pb case, Pb can be measured from the Pb line, and As can be measured
from either the As K,, or the As Kg line; in this way the interference can be corrected. If the As Kp
line is used, sensitivity will be decreased by a factor of two to five times because it is a less intense
line than the As K„ line. If the As K„ line is used in the presence of Pb, mathematical corrections
within the instrument software can be used to subtract out the Pb interference. However, because
of the limits of mathematical corrections, As concentrations cannot be efficiently calculated for
samples with Pb:As ratios of 10:1 or more. This high ratio of Pb to As may result in no As being
reported regardless of the actual concentration present.
No instrument can fully compensate for this interference. It is important for an operator to
understand this limitation of FPXRF instruments and consult with the manufacturer of the FPXRF
instrument to evaluate options to minimize this limitation. The operator's decision will be based on
action levels for metals in soil established for the site, matrix effects, capabilities of the instrument,
data quality objectives, and the ratio of lead to arsenic known to be present at the site. If a site is
encountered that contains lead at concentrations greater than ten times the concentration of arsenic
it is advisable that all critical soil samples be sent off site for confirmatory analysis by an EPA-
approved method.
4.8 If SSCS are used to calibrate an FPXRF instrument, the samples collected must be
representative of the site under investigation. Representative soil sampling ensures that a sample
or group of samples accurately reflects the concentrations of the contaminants of concern at a given
time and location. Analytical results for representative samples reflect variations in the presence and
concentration ranges of contaminants throughout a site. Variables affecting sample
representativeness include differences in soil type, contaminant concentration variability, sample
collection and preparation variability, and analytical variability, all of which should be minimized as
much as possible.
4.9 Soil physical arid chemical effects may be corrected using SSCS that have been
analyzed by inductively coupled plasma (ICP) or atomic absorption (AA) methods. However, a major
source of error can be introduced if these samples are not representative of the site or if the
analytical error is large. Another concern is the type of digestion procedure used to prepare the soil
samples for the reference analysis. Analytical results for the confirmatory method will vary
depending on whether a partial digestion procedure, such as SW-846 Method 3050, or a total
digestion procedure, such as Method 3052 is used. It is known that depending on the nature of the
soil or sediment, Method 3050 will achieve differing extraction efficiencies for different analytes of
interest. The confirmatory method should meet the project data quality objectives.
XRF measures the total concentration of an element; therefore, to achieve the greatest
comparability of this method with the reference method (reduced bias), a total digestion procedure
should be used for sample preparation. However, in the study used to generate the performance
data for this method, the confirmatory method used was Method 3050, and the FPXRF data
compared very well with regression correlation coefficients (r2 often exceeding 0.95, except for
barium and chromium. See Table 9 in Section 17.0). The critical factor is that the digestion
procedure and analytical reference method used should meet the data quality objectives (DQOs) of
the project and match the method used for confirmation analysis.
4.10 Ambient temperature changes can affect the gain of the amplifiers producing instrument
drift. Gain or drift is primarily a function of the electronics (amplifier or preamplifier) and not the
detector as most instrument detectors are cooled to a constant temperature. Most FPXRF
instruments have a built-in automatic gain control. If the automatic gain control is allowed to make
6200 -5
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periodic adjustments, the instrument will compensate for the influence of temperature changes on
its energy scale. If the FPXRF instrument has an automatic gain control function, the operator will
not have to adjust the instrument's gain unless an error message appears. If an error message
appears, the operator should follow the manufacturer's procedures for troubleshooting the problem.
Often, Ms involves performing a new energy calibration. The performance of an energy calibration
check to assess drift is a quality control measure discussed in Section 9.2.
If the operator is instructed by the manufacturer to manually conduct a gain check because of
increasing or decreasing ambient temperature, it is standard to perform a gain check after every 10
to 20 sample measurements or once an hour whichever is more frequent. It is also suggested that
a gain check be performed if the temperature fluctuates more than 10 to 20°F. The operator should
follow the manufacturer's recommendations for gain check frequency.
5.0 SAFETY
5.1 Proper training for the safe operation of the instrument and radiation training should be
completed by the analyst prior to analysis. Radiation safety for each specific instalment can be
found in the operators manual. Protective shielding should never be removed by the analyst or any
personnel other than the manufacturer. The analyst should be aware of the local state and national
regulations that pertain to the use of radiation-producing equipment and radioactive materials with
which compliance is required. Licenses for radioactive materials are of two types; (1) general license
which is usually provided by the manufacturer for receiving, acquiring, owning, possessing, using,
and transferring radioactive material incorporated in a device or equipment, and (2) specific license
which is issued to named persons for the operation of radioactive instruments as required by local
state agencies. There should be a person appointed within the organization that is solely
responsible for properly instructing all personnel, maintaining inspection records, and monitoring x-
ray equipment at regular intervals, A copy of the radioactive material licenses and leak tests should
be present with the instrument at alt times and available to local and national authorities upon
request. X-ray tubes do not require radioactive material licenses or leak tests, but do require
approvals and licenses which vary from state to state. In addition, fail-safe x-ray warning lights
should be illuminated whenever an x-ray tube is energized. Provisions listed above concerning
radiation safety regulations, shielding, training, and responsible personnel apply to x-ray tubes just
as to radioactive sources. In addition, a log of the times and operating conditions should be kept
whenever an x-ray tube is energized. Finally, an additional hazard present with x-ray tubes is the
danger of electric shock from the high voltage supply. The danger of electric shock is as substantial
as the danger from radiation but is often overlooked because of its familiarity.
5.2 Radiation monitoring equipment should be used with the handling of the instrument.
"Ttie operator and the surrounding environment should be monitored continually for analyst exposure
to radiation. Thermal luminescent detectors (TLD) in the form of badges and rings are used to
monitor operator radiation exposure. The TLDs should be worn in the area of most frequent
exposure. The maximum permissible whole-body dose from occupational exposure is 5 Roentgen
Equivalent Man (REM) per year! Possible exposure pathways for radiation to enter the body are
ingestion, inhaling, and absorption. The best precaution to prevent radiation exposure is distance
and shielding.
5.3 Refer to Chapter Three for guidance on some proper safety protocols.
6.0 EQUIPMENT AND SUPPLIES
6.1 FPXRF Spectrometer An FPXRF spectrometer consists of four major components:
(1) a source that provides x-rays; (2) a sample presentation device; (3) a detector that converts x-
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ray-generated photons emitted from the sample into measurable electronic signals; and (4) a data
processing unit that contains an emission or fluorescence energy analyzer, such as an MCA, that
processes the signals into an x-ray energy spectrum from which elemental concentrations in the
sample may be calculated, and a data display and storage system. These components and
additional, optional items, are discussed below.
6.1.1 Excitation Sources: Most FPXRF instruments use sealed radioisotope sources
to produce x-rays in order to irradiate samples. The FPXRF instrument may contain between
one and three radioisotope sources. Common radioisotope sources used for analysis for
metals in soils are iron (Fe)-55, cadmium (Cd)-109, americium (Am)-241, and curium (Cm)-
244. These sources may be contained in a probe along with a window and the detector; the
probe is connected to a data reduction and handling system by means of a flexible cable.
Alternatively, the sources, window, and detector may be included in the same unit as the data
reduction and handling system.
The relative strength of the radioisotope sources is measured in units of millicuries
(mCi). All other components of the FPXRF system being equal, the stronger the source, the
greater the sensitivity and precision of a given instrument. Radioisotope sources undergo
constant decay. In fact, it is this decay process that emits the primary x-rays used to excite
samples for FPXRF analysis. The decay of radioisotopes is measured in "half-lives." The half-
life of a radioisotope is defined as the length of time required to reduce the radioisotopes
strength or activity by half. Developers of FPXRF technologies recommend source
replacement at regular intervals based on the source's half-life. The characteristic x-rays
emitted from each of the different sources have energies capable of exciting a certain range
of analytes in a sample. Table 2 summarizes the characteristics of four common radioisotope
sources.
X-ray tubes have higher radiation output, no intrinsic lifetime limit, produce constant
output over their lifetime, and do not have the disposal problems of radioactive sources but are
just now appearing in FPXRF instruments An electrically-excited x-ray tube operates by
bombarding an anode with electrons accelerated by a high voltage. The electrons gain an
energy in electron volts equal to the accelerating voltage and can excite atomic transitions in
the anode, which then produces characteristic x-rays. These characteristic x-rays are emitted
through a window which contains the vacuum required for the electron acceleration. An
important difference between x-ray tubes and radioactive sources is that the electrons which
bombard the anode also produce a continuum of x-rays across a broad range of energies in
addition to the characteristic x-rays. This continuum is weak compared to the characteristic
X-rays but can provide substantial excitation since it covers a broad energy range. It has the
undesired property of producing background in the spectrum near the analyte x-ray lines when
it is scattered by the sample. For this reason a filter is often used between the x-ray tube and
the sample to suppress the continuum radiation while passing the characteristic x-rays from
the anode. This filter is sometimes incorporated into the window of the x-ray tube. The choice
of accelerating voltage is governed by the anode material, since the electrons must have
sufficient energy to excite the anode, which requires a voltage greater than the absorption
edge of the anode material. The anode is most efficiently excited by voltages 2 to 2.5 times
the edge energy (most x-rays per unit power to the tube), although voltages as low as 1.5
times the absorption edge energy will work. The characteristic x-rays emitted by the anode are
capable of exciting a range of elements in the sample just as with a radioactive source. Table
3 gives the recommended operating voltages and the sample elements excited for some
common anodes.
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6.1.2 Sample Presentation Device: FPXRF instruments can be operated in two
modes: in situ and intrusive. If operated in the in situ mode, the probe window is placed in
direct contact with the soil surface to be analyzed. When an FPXRF instrument is operated
in the intrusive mode, a soil or sediment sample must be collected, prepared, and placed in
a sample cup. For most FPXRF instruments operated in the intrusive mode, the probe is
rotated so that the window faces upward. A protective sample cover is placed over the
window, and the sample cup is placed on top of the window inside the protective sample cover
for analysis.
6.1.3 Detectors: The detectors in the FPXRF instruments can be either solid-state
detectors or gas-filled, proportional counter detectors. Common solid-state detectors include
mercuric iodide (Hgl2), silicon pin diode and lithium-drifted silicon Si(Li). The Hgl2 detector is
operated at a moderately subambient temperature controlled by a low power thermoelectric
cooler. The silicon pin diode detector also is cooled via the thermoelectric Peltier effect. The
Si(Li) detector must be cooled to at least -90 °C either with liquid nitrogen or by thermoelectric
coding via the Pettier effect instalments with a Si(Li) detector have an internal liquid nitrogen
dewar with a capacity of 0.5 to 1.0 liter. Proportional counter detectors are rugged and'
lightweight, which are important features of a field portable detector. However, the resolution
of a proportional counter detector is not as good as that of a solid-state detector. The energy
resolution of a detector for characteristic x-rays is usually expressed in terms of full width at
half-maximum (FWHM) height of the manganese K„ peak at 5.89 keV. The typical resolutions
of the above mentioned detectors are as follows: Hgl2-270 eV; silicon pin diode-250 eV;
Si(Li>-17Q eV; and gas-filled, proportional counter-750 eV.
During operation of a solid-state detector, an x-ray photon strikes a biased, solid-state
crystal and loses energy in the crystal by producing electron-hole pairs. The electric charge
produced is collected and provides a current pulse that is directly proportional to the energy
of the x-ray photon absorbed by the crystal of the detector. A gas-filled, proportional counter
detector is an ionization chamber filled with a mixture of noble and other gases. An x-ray
photon entering the chamber ionizes the gas atoms. The electric charge produced is collected
and provides an electric signal that is directly proportional to the energy of the x-ray photon
absorbed by the gas in the detector.
6.1.4 Data Processing Units: The key component in the data processing unit of an
FPXRF instrument is the MCA. The MCA receives pulses from the detector and sorts them
by their amptitudes (energy level). The MCA counts pulses per second to determine the height
of the peak in a spectrum, which is indicative of the target analyte's concentration. The
spectrum of element peaks are built on the MCA. The MCAs in FPXRF instruments have from
256 to 2,048 channels. The concentrations of target analytes are usually shown in parts per
million on a liquid crystal display (LCD) in the instrument: FPXRF instruments can store both
spectra and from 100 to 500 sets of numerical analytical results. Most FPXRF instruments are
menu-driven from software built into the units or from PCs. Once the data-storage memory
of an FPXRF unit is full, data can be downloaded by means of an RS-232 port and cable to a
PC.
6.2 Spare battery chargers.
6.3 Polyethylene sample cups: 31 millimeters (mm) to 40 mm in diameter with collar, or
equivalent (appropriate for FPXRF instrument).
6.4 X-ray window film: Mylar™, Kapton™, Spectrolene™, polypropylene, or equivalent; 2.5
to 6.0 micrometers (pm) thick.
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6.5 Mortar and pestle: glass, agate, or aluminum osdde; for grinding soil and sediment
samples.
6.6 Containers: glass or plastic to store samples.
6.7 Sieves: 60-mesh (0.25 rtim), stainless-steel, Nylon, or equivalent for preparing soil and
sediment samples.
6.8 Trowels: for smoothing soil surfaces and collecting soil samples.
6.9 Plastic bags: used for collection and homogenization of soil samples.
6.10 Drying oven: standard convection or toaster oven, for soil and sediment samples that
require drying.
7.0 REAGENTS AND STANDARDS
7.1 Pure Element Standards: Each pure, single-element standard is intended to produce
strong characteristic x-ray peaks of the element of interest only. Other elements present must not
contribute to the fluorescence spectrum. A set of pure element standards for commonly sought
analytes is supplied by the instrument manufacturer, if required for the instrument; not all instruments
require the pure element standards. The standards are used to set the region of interest (ROI) for
each element. They also can be used as energy calibration and resolution check samples.
7.2 Site-specific Calibration Standards: Instruments that employ fundamental parameters
(FP) or similar mathematical models in minimizing matrix effects may not require SSCS. If the FP
calibration model is to be optimized or if empirical calibration is necessary, then SSCSs must be
collected, prepared, and analyzed.
7.2; 1 The SSCS must be representative of the matrix to be analyzed by FPXRF.
These samples must be well homogenized. A minimum of ten samples spanning the
concentration ranges of the analytes of interest and of the interfering elements must be
obtained from the site. A sample size of 4 to 8 ounces is recommended, and standard glass
sampling jars should be used.
7.2.2 Each sample should be oven-dried for 2 to 4 hours at a temperature of less
than 150°C. If mercury is to be analyzed, a separate sample portion must remain undried, as
heating may volatilize the mercury. When the sample is dry, all large, organic debris and
nonrepresentative material, such as twigs, leaves, roots, insects, asphalt, and rock should be
removed. The sample should be ground with a mortar and pestle and passed through a 60-
mesh sieve. Only the coarse rock fraction should remain on the screen.
7.2.3 The sample should be homogenized by using a riffle splitter or by placing 150
to 200 grams of the dried, sieved sample on a piece of kraft or butcher paper about 1.5 by 1.5
feet in size. Each comer of the paper should be lifted alternately, rolling the soil over on itself
and toward the opposite comer. The soil should be rolled on itself 20 times. Approximately
5 grams of the sample should then be removed and placed in a sample cup for FPXRF
analysis. The rest of the prepared sample should be sent off site for ICP or AA analysis. The
method use for confirmatory analysis should meet the data quality objectives of the project.
7.3 Blank Samples: The blank samples should be from a "clean" quartz or silicon dioxide
matrix that is free of any analytes at concentrations above the method detection limits. These
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samples are used to monitor for cross-contamination and laboratory-induced contaminants or
interferences.
7.4 Standard Reference Materials: Standard reference materials (SRM) are standards
containing certified amounts of metals in soil or sediment. These standards are used for accuracy
and performance checks of FPXRF analyses. SRMs can be obtained from the National Institute of
Standards and Technology (NIST), the U.S. Geological Survey (USGS), the Canadian National
Research Council, and the national bureau of standards in foreign nations. Pertinent NIST SRMs
for FPXRF analysis include 2704, Buffalo River Sediment; 2709, San Joaquin Soil; and 2710 and
2711, Montana Soil. These SRMs contain soil or sediment from actual sites that has been analyzed
using independent inorganic analytical methods by many different laboratories.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
Sample handling and preservation procedures used in FPXRF analyses should follow the guidelines
in Chapter Three, Inorganic Analytes.
9.0 QUALITY CONTROL
9.1 Refer to Chapter One for additional guidance on quality assurance protocols. All field
data sheets and quality control data should be maintained for reference or inspection.
9.2 Energy Calibration Check: To determine whether an FPXRF instrument is operating
within resolution and stability tolerances, an energy calibration check should be run. The energy
calibration check determines whether the characteristic x-ray lines are shifting, which would indicate
drift within the instrument. As discussed in Section 4.10, this check also serves as a gain check in
the event that ambient temperatures are fluctuating greatly (> 10 to 20°F).
The energy calibration check should be run at a frequency consistent with manufacturers
recommendations. Generally, this wouid be at the beginning of each working day, after the batteries
are changed or the instrument is shut off, at the end of each working day, and at any other time
when the instrument operator believes that drift is occurring during analysis. A pure element such
as iron, manganese, copper, or lead is often used for the energy calibration check. A manufacturer-
recommended count time per source should be used for the check.
9.2.1 The instrument manufacturer's manual specifies the channel or kiloelectron
volt level at which a pure element peak should appear and the expected intensity of the peak.
The intensity and channel number of the pure element as measured using the radioactive
source should be checked and compared to the manufacturer's recommendation. If the energy
calibration check does not meet the manufacturer's criteria, then the pure element sample
should be repositioned and reanalyzed. If the criteria are still not met, then an energy
calibration should be performed as described in the manufacturer's manual. With some
FPXRF instruments, once a spectrum is acquired from the energy calibration check, the peak
can be optimized and realigned to the manufacturer's specifications using their software.
9.3 Blank Samples: Two types of blank samples should be analyzed for FPXRF analysis:
instrument blanks and method blanks. An instrument blank is used to verify that no contamination
exists in the spectrometer or on the probe window.
9.3.1 The instrument blank can be silicon dioxide, a Teflon block, a quartz block,
"clean" sand, or lithium carbonate. This instrument blank should be analyzed on each working
day before and after analyses are conducted and once per every twenty samples. An
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instrument blank should also be analyzed whenever contamination is suspected by the analyst.
The frequency of analysis will vary with the data quality objectives of the project. A
manufacturer-recommended count time per source should be used for the blank analysis. No
element concentrations above the method detection limits should be found in the'instrument
blank. If concentrations exceed these limits, then the probe window and the check sample
should be checked for contamination. If contamination is not a problem, then the instrument
must be "zeroed" by following the manufacturer's instructions.
9.3.2 A method blank is used to monitor for laboratory-induced contaminants or
interferences. The method blank can be "clean" silica sand or lithium carbonate that
undergoes the same preparation procedure as the samples. A method blank must be analyzed
at least daily. The frequency of analysis will depend on the data quality objectives of the
project. To be acceptable, a method blank must not contain any analyte at a concentration
above its method detection limit. If an analyte's concentration exceeds its method detection
limit, the cause of the problem must be identified, and all samples analyzed with the method
blank must be reanalyzed.
9.4 Calibration Verification Checks: A calibration verification check sample is used to check
the accuracy of the instrument and to assess the stability and consistency of the analysis for the
analytes of interest. A check sample should be analyzed at the beginning of each working day,
during active sample analyses, and at the end of each working day. The frequency of calibration
checks during active analysis will depend on the data quality objectives of the project. The check
sample should be a well characterized soil sample from the site that is representative of site samples
in terms of particle size and degree of homogeneity and that contains contaminants at
concentrations near the action levels. If a site-specific sample is not available, then an NIST or other
SRM that contains the analytes of interest can be used to vierify the accuracy of the instrument. The
measured value for each target analyte should be within ±20 percent (%D) of the true value for the
calibration verification check to be acceptable. If a measured value falls outside this range, then the
check sample should be reanalyzed. If the value continues to fall outside the acceptance range, the
instrument should be recalibrated, and the batch of samples analyzed before the unacceptable
calibration verification check must be reanalyzed.
9.5 Precision Measurements: The precision of the method is monitored by analyzing a
sample with low, moderate, or high concentrations of target analytes. The frequency of precision
measurements Will depend on the data quality objectives for the data. A minimum of one precision
sample should be run per day. Each precision sample should be analyzed 7 times in replicate. It
is recommended that precision measurements be obtained for samples with varying concentration
ranges to asseiss the effect of concentration on method precision. Determining method precision
for analytes at concentrations near the site action levels can be extremely important if the FPXRF
results are to be used in an enforcement action; therefore, selection of at least one sample with
target analyte concentrations at or near the site action levels or levels of concern is recommended.
A precision sample is analyzed by the instrument for the same field analysis time as used for other
project samples. The relative standard deviation (RSD) of the sample mean is used to assess
method precision. For FPXRF data to be considered adequately precise, the RSD should not be
greater than 20 percent with the exception of chromium. RSD values for chromium should not be
greater than 30 percent.
The equation for calculating RSD is as follows:
RSD - (SD/Mean Concentration) x 100
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where:
RSD
Relative standard deviation for the precision measurement for the
analyte
Standard deviation of the concentration for the analyte
Mean concentration for the analyte
SD
Mean Concentration
The precision or reproducibility of a measurement will improve with increasing count time,
however, increasing the count time by a factor of 4 will provide only 2 times better precision, so there
is a point of diminishing return. Increasing the count time also improves the detection limit, but
decreases sample throughput.
9.6 Detection Limits: Results for replicate analyses of a low-concentration sample, SSCS,
or SRM can be used to generate an average site-specific method detection and quantitation limits.
In this case, the method detection limit is defined as 3 times the standard deviation of the results for
the low-concentration samples and the method quantitation limit is defined as 10 times the standard
deviation of the same results. Another means of determining method detection and quantitation
limits involves use of counting statistics. In FPXRF analysis, the standard deviation from counting
statistics is defined as SD = (N)M, where SD is the standard deviation for a target analyte peak and
N is the net counts for the peak of the analyte of interest (i.e., gross counts minus background under
the peak). Three times this standard deviation would be the method detection limit and 10 times this
standard deviation would be the method quantitation limit if both of the above mentioned
approaches are used to calculate method detection limits, the larger of the standard deviations
should be used to provide the more conservative detection limits.
This 8D based detection limit criteria must be used by the operator to evaluate each
measurement for its useability. A measurement above the average calculated or manufacturer's
detection limit, but smaller than three times its associated SD, should not be used as a quantitative
measurement. Conversely, if the measurement is below the average calculated or manufacturer's
detection limit, but greater than three times its associated SD. It Should be coded as an estimated
value.
9.7 Confirmatory Samples: The comparability of the FPXRF analysis is determined by
submitting FPXRF-analyzed samples for analysis at a laboratory. The method of confirmatory
analysis must meet the project and XRF measurement data quality objectives. The confirmatory
samples must be splits of the well homogenized sample material. In some cases the prepared
sample cups can be submitted. A minimum of 1 sample for each 20 FPXRF-analyzed samples
should be submitted for confirmatory analysis. This frequency will depend on data quality objectives.
The confirmatory analyses can also be used to verify the quality of the FPXRF data. Hie
confirmatory samples should be selected from the lower, middle, and upper range of concentrations
measured by the FPXRF. They should also include samples with analyte concentrations at or near
the site action levels. The results of the confirmatory analysis and FPXRF analyses should be
evaluated witfi a least squares linear regression analysis. If the measured concentrations span more
than one order of magnitude, the data should be log-transformed to standardize variance which is
proportional to the magnitude of measurement. The correlation coefficient (r2) for the results should
be 0.7 or greater for the FPXRF data to be considered screening level data. If the r2 is 0.9 or greater
and inferential statistics indicate the FPXRF data and the confirmatory data are statistically
equivalent at a 99 percent confidence level, the data could potentially meet definitive level data
criteria.
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10.0 CALIBRATION AND STANDARDIZATION
10.1 Instrument Calibration: Instrument calibration procedures vary among FPXRF
instruments. Users of this method should follow the calibration procedures outlined in the operator's
manual for each specific FPXRF instrument. Generally, however, three types of calibration
procedures exist for FPXRF instalments: FP calibration, empirical calibration, and the Compton peak
ratio or normalization method. These three types of calibration are discussed below.
10.2 Fundamental Parameters Calibration: FP calibration procedures are extremely variable.
An FP calibration provides the analyst with a "standardless" calibration. The advantages of FP
calibrations over empirical calibrations include the following:
• No previously collected site-specific samples are required, although
site-specific samples with confirmed and validated analytical results for all
elements present could be used.
* Cost is reduced because fewer confirmatory laboratory results or calibration
standards are required.
However, the analyst should be aware of the limitations imposed on FP calibration by particle
size and matrix effects. These limitations can be minimized by adhering to the preparation
procedure described in Section 7.2. The two FP calibration processes discussed below are based
on an effective energy FP routine and a back scatter with FP (BFP) routine. Each FPXRF FP
calibration process is based on a different iterative algorithmic method. The calibration procedure
for each routine is explained in detail in the manufacturer's user manual for each FPXRF instrument;
in addition, training courses are offered for each instrument.
10.2.1 Effective Energy FP Calibration: The effective energy FP calibration is
performed by the manufacturer before an instrument is sent to the analyst. Although SSCS
can be used, the calibration relies on pure element standards or SRMs such as those obtained
from NIST for the FP calibration. The effective energy routine relies on the spectrometer
response to pure elements and FP iterative algorithms to compensate for various matrix
effects.
Alpha coefficients are calculated using a variation of the Sherman equation, which
calculates theoretical intensities from the measurement of pure element samples. These
coefficients indicate the quantitative effect of each matrix element on an analyte's measured
x-ray intensity. Next, the Lachance Traill algorithm is solved as a set of simultaneous
equations based on the theoretical intensities. The alpha coefficients are then downloaded
into the specific instrument.
The working effective energy FP calibration curve must be verified before sample
analysis begins on each working day, after every 20 samples are analyzed, and at the end of
sampling. This verification is performed by analyzing either an NIST SRM or an SSCS that is
representative of the site-specific samples. This SRM or SSCS serves as a calibration check.
A manufacturer-recommended count time per source should be used for the calibration check.
The analyst must then adjust the y-intercept and slope of the calibration curve to best fit the
known concentrations of target analytes in the SRM or SSCS.
A percent difference (%D) is then calculated for each target analyte. The %D should
be within ±20 percent of the certified value for each analyte. If the %D falls outside this
acceptance range, then the calibration curve should be adjusted by varying the slope of the
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line or the y-intercept value for the analyte. The SRM or SSCS is reanalyzed until the %D falls
within ±20 percent. The group of 20 samples analyzed before an out-of-control calibration
check should be reanalyzed.
The equation to calibrate %D is as follows:
%D=((CS-Ck)/Cl()x100
where:
%D = Percent difference
C* * Certified concentration of standard sample
C, = Measured concentration of standard sample
10.2.2 BFP Calibration: BFP calibration relies on the ability of the liquid nitrogen-
cooled, Si(Li) solid-state detector to separate the coherent (Compton) and Incoherent
(Rayieigh) backscatter peaks of primary radiation. These peak intensities are known to be a
function of sample composition, and the ratio of the Compton to Rayieigh peak is a function
of the mass absorption of the sample. The calibration procedure is explained in detail in the
instrument manufacturer's manual. Following is a general description of the BFP calibration
procedure.
The concentrations of all detected and quantified elements are entered into the
computer software system. Certified element results for an NIST SRM or confirmed and
validated results for an SSCS can be used. In addition, the concentrations of oxygen and
silicon must be entered; these two concentrations are not found in standard metals analyses.
The manufacturer provides silicon and oxygen concentrations for typical soil types. Pure
element standards are then analyzed using a manufacturer-recommended count time per
source. The results are used to calculate correction factors in order to adjust for spectrum
overlap of elements.
The working BFP calibration curve must be verified before sample analysis begins on
each working day, after every 20 samples are analyzed, and at the end of the analysis. This
verification is performed by analyzing either an NIST SRM or an SSCS that is representative
of the site-specific samples. This SRM or SSCS serves as a calibration check. The standard
sample is analyzed using a manufacturer-recommended count time per source to check the
calibration curve. The analyst must then adjust the y-intercept and slope of the calibration
curve to best fit the known concentrations of target analytes in the SRM or SSCS.
A %D is then calculated for each target analyte. The %D should fall within ±20 percent
of the certified value for each analyte. If the %D falls outside this acceptance range, then the
calibration curve should be adjusted by varying the slope of the line the y-intercept value for
the analyte. The standard sample is reanalyzed until the %D falls within ±20 percent. The
group of 20 samples analyzed before an out-of-control calibration check should be reanalyzed.
10.3 Empirical Calibration: An empirical calibration can be performed with SSCS, site-typical
standards, or standards prepared from metal oxides. A discussion of SSCS is included in Section
7.2; if no previously characterized samples exist for a specific site, site-typical standards can be
used. Site-typical standards may be selected from commercially available characterized soils or
from SSCS prepared for another site. The site-typical standards should closely approximate the
site's soil matrix with respect to particle size distribution, mineralogy, and contaminant analytes. If
neither SSCS nor site-typical standards are available, it is possible to make gravimetric standards
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by adding metal oxides to a "clean" sand or silicon dioxide matrix that simulates soil. Metal oxides
can be purchased from various chemical vendors. If standards are made on site, a balance capable
of weighing items to at least two decimal places is required. Concentrated ICR or AA standard
solutions can also be used to make standards. These solutions are available in concentrations of
10,000 parts per million, thus only small volumes have to be added to the soil.
An empirical calibration using SSCS involves analysis of SSCS by the FPXRF instrument and
by a conventional analytical method such as I CP or AA. A total add digestion procedure should be
used by the laboratory for sample preparation. Generally, a minimum of 10 and a maximum of 30
well characterized SSCS, site-typical standards, or prepared metal oxide standards are required to
perform an adequate empirical calibration. The number of required standards depends on the
number of analytes of interest and interfering elements. Theoretically, an empirical calibration with
SSCS should provide the most accurate data for a site because the calibration compensates for site-
specific matrix effects.
The first step in an empirical calibration is to analyze the pure element standards for the
elements of interest. This enables the instrument to set channel limits for each element for spectral
deconvolution. Next the SSCS, site-typical standards, or prepared metal oxide standards are
analyzed using a count time of 200 seconds per source or a count time recommended by the
manufacturer. This will produce a spectrum and net intensity of each analyte in each standard. The
analyte concentrations for each standard are then entered into the instrument software; these
concentrations are those obtained from the laboratory, the certified results, or the gravimetrically
determined concentrations of the prepared standards, this gives the instrument analyte values to
regress against corresponding intensities during the modeling stage. The regression equation
correlates the concentrations of an analyte with its net intensity.
The calibration equation is developed using a least squares fit regression analysis. After the
regression terms to be used in the equation are defined, a mathematical equation can be developed
to calculate the analyte concentration in an unknown sample, in some FPXRF instruments, the
software of the instrument calculates the regression equation. The software uses calculated
intercept and slope values to form a multiterm equation. In conjunction with the software in the
instrument, the operator can adjust the multiterm equation to minimize interelement interferences
and optimize the intensity calibration curve.
It is possible to define up to six linear or nonlinear terms in the regression equation. Terms can
be added and deleted to optimize the equation. The goal is to produce an equation with the smallest
regression error and the highest correlation coefficient. These values are automatically computed
by the software as the regression terms are added, deleted, or modified. It is also possible to delete
data points from the regression line if these points are significant outliers or if they are heavily
weighing the data. Once the regression equation has been selected for an analyte, the equation can
be entered into the software for quantitation of analytes in subsequent samples. For an empirical
calibration to be acceptable, the regression equation for a specific analyte should have a correlation
coefficient of 0.98 or greater or meet the DQOs of the project.
In an empirical calibration, one must apply the DQOs of the project and ascertain critical or
action levels for the analytes of interest. It is within these concentration ranges or around these
action levels that the FPXRF instrument should be calibrated most accurately. It may not be possible
to develop a good regression equation over several orders of analyte concentration.
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10.4 Compton Normalization Method: The Compton normalization method is based on
analysis of a single, certified standard and normalization for the Compton peak. The Compton peak
is produced from incoherent backscattering of x-ray radiation from the excitation source and is
present in the spectrum of every sample. The Compton peak intensity changes with differing
matrices. Generally, matrices dominated by lighter elements produce a larger Compton peak, and
those dominated by heavier elements produce a smaller Compton peak. Normalizing to the
Compton peak can reduce problems with varying matrix effects among samples. Compton
normalization is similar to the use of internal standards in organics analysis. "Hie Compton
normalization method may not be effective when analyte concentrations exceed a few percent.
The certified standard used for this type of calibration could be an NIST SRM such as 2710 or
2711. The SRM must be a matrix similar to the samples and must contain the analytes of interests
at concentrations near those expected in the samples. First, a response factor has to be determined
for each analyte. This factor is calculated by dividing the net peak intensity by the analyte
concentration. The net peak intensity is gross intensity corrected for baseline interference.
Concentrations of analytes in samples are then determined by multiplying the baseline corrected
analyte signal intensity by the normalization factor and by the response factor. The normalization
factor is the quotient of the baseline corrected Compton K„ peak intensity of the SRM divided by that
of the samples. Depending on the FPXRF instrument used, these calculations may be done
manually or by the instrument software.
11.0 PROCEDURE
11.1 Operation of the various FPXRF instruments will vary according to the manufacturers'
protocols. Before operating any FPXRF instrument, one should consult the manufacturer's manual.
Most manufacturers recommend that their instruments be allowed to warm up for 15 to 30 minutes
before analysis of samples. This will help alleviate drift or energy calibration problems later on in
analysiis.
11.2 Each FPXRF instrument should be operated according to the manufacturer's
recommendations. There are two modes in which FPXRF instruments can be operated: in situ and
intrusive. The in situ mode involves analysis of an undisturbed soil sediment or sample. Intrusive
analysis involves collection and preparation of a soil or sediment' sample before analysis. Some
FPXRF instruments can operate in both modes of analysis, while others are designed to operate in
on|y one mode. The two modes of analysis are discussed below.
11.3 For in situ analysis, one requirement is that any large or nonrepresentative debris be
removed from the soil surface before analysis. This debris includes rocks, pebbles, leaves,
Vegetation, roots, and concrete. Another requirement is that the soil surface be as smooth as
possible so that the probe window will have good contact with the surface. This may require some
leveling of the surface with a stainless-steel trowel. During the study conducted to provide data for
Ms method, this modest amount of sample preparation was found to take less than S minutes per
sample location. The last requirement is that the soil or sediment not be saturated with water.
Manufacturers state that their FPXRF instruments will perform adequately for soils with moisture
contents of 5 to 20 percent but will not perform well for saturated soils, especially if ponded water
exists on the surface. Another recommended technique for in situ analysis is to tamp the soil to
increase soil density and compactness for better repeatability and representativeness. This
condition is especially important for heavy element analysis, such as barium. Source count times
for in situ analysis usually range from 30 to 120 seconds, but source count times will vary among
instruments and depending on required detection limits.
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11.4 For intrusive analysis of surface or sediment, it is recommended that a sample be
collected from a 4- by 4-inch square that is 1 inch deep. This will produce a soil sample of
approximately 375 grams or 250 cm3, which is enough soil to fill an 8-ounce jar. The sample should
be homogenized, dried, and ground before analysis. The sample can be homogenized before or
after drying. The homogenization technique to be used after drying is discussed in Section 4.2. if
the sample is homogenized before drying, it should be thoroughly mixed in a beaker or similar
container, or if the sample is moist and has a high clay content, it can be kneaded in a plastic bag.
One way to monitor homogenization when the sample is kneaded in a plastic bag is to add sodium
fluorescein dye to the sample. After the moist sample has been homogenized, it is examined under
an ultraviolet light to assess the distribution of sodium fluorescein throughout the sample. If the
fluorescent dye is evenly distributed in the sample, homogenization is considered complete; if the
dye is not evenly distributed, mixing should continue until the sample has been thoroughly
homogenized. During the study conducted to provide data for this method, the homogenization
procedure using the fluorescein dye required 3 to 5 minutes per sample. As demonstrated in
Sections 13.5 and 13.7, homogenization has the greatest impact on the reduction of sampling
variability. It produces little or no contamination. Often, it can be used without the more labor
intensive steps of drying, grinding, and sieving given in Sections 11.5 arid 11.6. Of course, to
achieve the best data quality possible all four steps must be followed.
11.5 Once the soil or sediment sample has been homogenized, it should be dried. This can
be accomplished with a toaster oven or convection oven. A small aliquot of the sample (20 to 50
grams) is placed in a suitable container for drying. The sample should be dried fa* 2 to 4 hours in
the convection or toaster oven at a temperature not greater than 150°C. Microwave drying is not
a recommended procedure. Field studies have shown that microwave drying can increase variability
between the FPXRF data and confirmatory analysis. High levels of metals in a sample can cause
arcing in the microwave oven, and sometimes slag forms in the sample. Microwave oven drying can
also melt plastic containers used to hold the sample.
11.6 The homogenized dried sample material should be ground with a mortar and pestle and
passed through a 60-mesh sieve to achieve a uniform particle size. Sample grinding should
continue until at least 90 percent of the original sample passes through the sieve. The grinding step
normally takes an average of 10 minutes per sample. An aliquot of the sieved sample should then
be placed in a 31,0-mm polyethylene sample cup (or equivalent) for analysis. The sample cup
should be one-half to three-quarters full at a minimum. The sample cup should be covered with a
2.5 Mm Mylar (or equivalent) film for analysis. The rest of the soil sample should be placed in a jar,
labeled, and archived for possible confirmation analysis. All equipment including the mortar, pestle,
and sieves must be thoroughly cleaned so that any cross-contamination is below the MDLs of the
procedure or DQOs of the analysis.
12.0 DATA ANALYSIS AND CALCULATIONS
Most FPXRF instruments have software capable of storing all analytical results and spectra. The
results are displayed in parts per million and can be downloaded to a PC, which can provide a hard
copy printout. Individual measurements that are smaller than three times their associated SD should
not be used for quantitation.
13.0 METHOD PERFORMANCE
13.1 This section discusses four performance factors, field-based method detection limits,
precision, accuracy, and comparability to EPA-approved methods. The numbers presented in
Tables 4 through 9 were generated from data obtained from six FPXRF instruments. The soil
samples analyzed by the six FPXRF instruments were collected from two sites in the United States.
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The soil samples contained several of the target analytes at concentrations ranging from nondetect
to tens of thousands of mg/kg.
13.2 The six FPXRF instalments included the TN 9000 and TN Lead Analyzer manufactured
by TN Spectrace; the X-MET 920 with a SiLi detector and X-MET 920 with a gas-filled proportional
detector manufactured by Metorex, Inc.; the XL Spectrum Analyzer manufactured by Niton; and the
MAP Spectrum Analyzer manufactured by Scitec. The TN 9000 and TN Lead Analyzer both have
a Hgl2 detector. The TN 9000 utilized an Fe-55, Cd-109, and Am-241 source. The TN Lead
Analyzer had only a Cd-109 source. The X-Met 920 with the SiLi detector had a Cd-109 and Am-241
source. The X-MET 920 with the gas-filled proportional detector had only a Cd-109 source. The XL
Spectrum Analyzer utilized a silicon pin-diode detector and a Cd-109 source. The MAP Spectrum
Analyzer utilized a solid-state silicon detector and a Cd-109 source.
13.3 AH data presented in Tables 4 through 9 were generated using the following calibrations
and source count times. The TN 9000 and TN Lead Analyzer were calibrated using fundamental
parameters using NIST SRM 2710 as a calibration check sample. The TN 9000 was operated using
100,60, and 60 second count times for the Cd-109, Fe-55, and Am-241 sources, respectively. The
TN Lead analyzer was operated using a 60 second count time for the Cd-109 source. The X-MET
920 with the Si(Li) detector was calibrated using fundamental parameters and one well characterized
site-specific soil standard as a calibration check. It used 140 and 100 second count times for the
Cd-109 and Am-241 sources, respectively. The X-MET 920 with the gas-filled proportional detector
was calibrated empirically using between 10 and 20 well characterized site-specific soil standards.
It used 120 second times for the Cd-109 source. The XL Spectrum Analyzer utilized NIST SRM 2710
for calibration and the Compton peak normalization procedure for quantitation based on 60 second
count times for the Cd-109 source. The MAP Spectrum Analyzer was internally calibrated by the
manufacturer, the calibration was checked using a well-characterized site-specific soil standard.
It used 240 second times for the Cd-109 source.
13.4 Field-Based Method Detection Limits; The field-based method detection limits are
presented in Table 4. The field-based method detection limits were determined by collecting ten
replicate measurements on site-specific soil samples with metals concentrations 2 to 5 times the
expected method detection limits. Based on these ten replicate measurements, a standard deviation
on the replicate analysis was calculated. The method detection limits presented in Table 4 are
defined as 3 times the standard deviation for each anaiyte.
The field-based method detection limits were generated by using the count times discussed
earlier in this section. All the field-based method detection limits were calculated for soil samples
that had been dried and ground and placed in a sample cup with the exception of the MAP Spectrum
Analyzer. This instrument can only be operated in the in situ mode, meaning the samples were
moist and not ground.
Some of the analytes such as cadmium, mercury, silver, selenium, and thorium were not
detected or only detected at very low concentrations such that a field-based method detection limit
could not be determined. These analytes are not presented in Table 4. Other analytes such as
calcium, iron, potassium, and titanium were only found at high concentrations (thousands of mg/kg)
so that reasonable method detection limits could not be calculated. These analytes also are not
presented in Table 4.
13.5 Precision Measurements: The precision data is presented in Table 5. Each of the six
FPXRF instruments performed 10 replicate measurements on 12 soil samples that had anaiyte
concentrations ranging from nondetects to thousands of mg/kg. Each of the 12 soil samples
underwent 4 different preparation techniques from in situ (no preparation) to dried and ground in a
6200-18
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sample cup. Therefore, there were 48 precision data points for five of the instruments and 24
precision points for the MAP Spectrum Analyzer. The replicate measurements were taken using the
source count times discussed at the beginning of this section.
For each detectable analyte in each precision sample a mean concentration, standard
deviation, and RSD was calculated for each analyte. The data presented in Table 5 is an average
RSD for the precision samples that had analyte concentrations at 5 to 10 times the MDL for that
analyte for each instrument. Some anaiytes such as mercury, selenium, silver, and thorium were
not detected in any of the precision samples so these anaiytes are not listed in Table 5. Some
anaiytes such as cadmium, nickel, and tin were only detected at concentrations near the MDLs so
that an RSD value calculated at 5 to 10 times the MDL was not possible.
One FPXRF instrument collected replicate measurements on an additional nine soil samples
to provide a better assessment of the effect of sample preparation on precision. Table 6 shows
tfiese results. The additional nine soil samples were comprised of three from each texture and had
analyte concentrations ranging from near the detection limit of the FPXRF analyzer to thousands of
mg/kg. The FPXRF analyzer only collected replicate measurements from three of the preparation
methods,' no measurements were collected from the in situ homogenized samples. The FPXRF
analyzer conducted five replicate measurements ofthe in situ field samples by taking measurements
at five different points within the 4-inch by 4-inch sample square. Ten replicate measurements were
collected for both the intrusive undried and unground and intrusive dried and ground samples
contained in cups. The cups were shaken between each replicate measurement.
Table 6 shows that the precision dramatically improved from the in situ to the intrusive
measurements. In general there was a slight improvement in precision when the sample was dried
and ground. Two factors caused the precision for the in situ measurements to be poorer. The major
factor is soil heterogeneity. By moving the probe within the 4-inch by 4-inch square, measurements
of different soil samples were actually taking place within the square. Table 6 illustrates the
dominant effect of soil heterogeneity. It overwhelmed instrument precision when the FPXRF
analyzer was used in this mode. The second factor that caused the RSD values to be higher for the
in situ measurements is the fact that only five versus ten replicates were taken. A lesser number
of measurements caused the standard deviation to be larger which in turn elevated the RSD values.
13.6 Accuracy Measurements: Five of the FPXRF instruments (not including the MAP
Spectrum Analyzer) analyzed 18 SRMs using the source count times and calibration methods given
at the beginning of this section. The 18 SRMs included 9 soil SRMs, 4 stream or river sediment
SRMs, 2 sludge SRMs, and 3 ash SRMs. Each of the SRMs contained known concentrations of
certain target anaiytes. A percent recovery was calculated for each analyte in each SRM for each
FPXRF instrument. Table 7 presents a summary of this data. With the exception of cadmium,
chromium, and nickel, the values presented in Table 7 were generated from the 13 soil and sediment
SRMs only. The 2 sludge and 3 ash SRMs were included for cadmium, chromium, and nickel
because ofthe low or nondetectable concentrations of these three anaiytes in the soil and sediment
SRMs.
Only 12 anaiytes are presented in Table 7. These are the anaiytes that are of environmental
concern and provided a significant number of detections in the SRMs for an accuracy assessment.
No data is presented for the X-MET 920 with the gas-filled proportional detector. This FPXRF
instrument was calibrated empirically using site-specific soil samples. The percent recovery values
from this instrument were very sporadic and the data did not lend itself to presentation in Table 7.
Table 8 provides a more detailed summary of accuracy data for one FPXRF instrument (TN
9000) for the 9 soil SRMs and 4 sediment SRMs. Table 8 shows the certified value, measured
6200-19
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value, arid percent recovery for five analytes. These analytes were chosen because they are of
environmental concern and were most prevalently certified for in the SRM and detected by the
FPXRF instrument. The first nine SRMs are soil and the last 4 SRMs are sediment. Percent
recoveries for the four NIST SRMs were often between 90 and 110 percent for all analytes.
13.7 Comparability: Comparability refers to the confidence with which one data set can be
compared to another. In this case, FPXRF data generated from a large study of six FPXRF
instruments was compared to SW-846 Methods 3050 and 6010 which are the standard soil
extraction for metals and analysis by inductively coupled plasma. An evaluation of comparability was
conducted by using linear regression analysis. Three factors were determined using the linear
regression. These factors were the y-intercept, the slope of the line, and the coefficient of
determination (r2).
As part of the comparability assessment, the effects of soil type and preparation methods were
studied. Three soil types (textures) and four preparation methods were examined during the study.
The preparation methods evaluated the cumulative effect of particle size, moisture, and
homogenization on comparability. Due to the large volume of data produced during this study, linear
regression data for six analytes from only one FPXRF instrument is presented in Table 9. Similar
trends in the data were seen for all instruments.
Table 9 shows the regression parameters for the whole data set, broken out by soil type, and
by preparation method. The soil types are as follows: soil 1-sand; soil 2-loam; and soil 3—siity clay.
The preparation methods are as follows: preparation 1—in situ in the field; preparation 2-in situ,
sample collected and homogenized; preparation 3-intrusive, with sample in a sample cup but
sample still wet and not ground; and preparation 4-rsample dried, ground, passed through a 40-mesh
sieve, and placed in sample cup.
For arsenic, copper, lead, and zinc, the comparability to the confirmatory laboratory was
excellent with r2 values ranging from 0.80 to 0.99 for all six FPXRF instruments. The slopes of the
regression lines for arsenic, copper, lead, and zinc, were generally between 0.90 arid 1.00 indicating
the data would need to be corrected very little or not at all to match the confirmatory laboratory data.
The l values and slopes of the regression lines for barium and chromium were not as good as for
the other for analytes, indicating the data would have to be corrected to match the confirmatory
laboratory.
Table 9 demonstrates that there was little effect of soil type on the regression parameters for
any of the six analytes. The only exceptions were for barium in soil 1 and copper in soil 3. In both
of these cases, however, it is actually a concentration effect and not a soil effect causing the poorer
comparability. All barium and copper concentrations in soil 1 and 3, respectively, were less than 350
mg/kg.
Table 9 shows there was a preparation effect on the regression parameters for all six analytes.
With the exception of chromium, the regression parameters were primarily improved going from
preparation 1 to preparation 2. In this step, the sample was removed from the soil surface, all large
debris was removed, and the sample was thoroughly homogenized. The additional two preparation
methods did little to improve the regression parameters. This data indicates that homogenization
is the most critical factor when comparing the results. It is essential that the sample sent to the
confirmatory laboratory match the FPXRF sample as closely as possible.
Section 11.0 of this method discusses the time necessary for each of the sample preparation
techniques. Based on the data quality objectives for the project, an analyst must decide if it is worth
the extra time required to dry and grind the sample for small improvements in comparability.
6200 - 20
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Homogenization requires 3 to 5 minutes. Drying the sample requires one to two hours. Grinding and
sieving requires another 10 to 15 minutes per sample. Lastly, when grinding and sieving is
conducted, time must be allotted to decontaminate the mortars, pestles, and sieves. Drying and
grinding the samples and decontamination procedures will often dictate that an extra person be on
site so that the analyst can keep up with the sample collection crew. The cost of requiring an extra
person on site to prepare samples must be balanced with the gain in data quality and sample
throughput.
13.8 The following documents may provide additional guidance and insight on this method
and technique:
13.8.1 Hewitt, A.D, 1994. "Screening for Metals by X-ray Fluorescence
Spectrometry/Response Factor/Compton K,, Peak Normalization Analysis." American
Environmental Laboratory. Pages 24-32.
13.8.2 Piorek, S., and J.R. Pasmore. 1993. "Standardless, In Situ Analysis of
Metallic Contaminants in the Natural Environment With a PC-Based, High Resolution Portable
X-Ray Analyzer." Third International Symposium on Field Screening Methods for Hazardous
Waste arid Toxic Chemicals. Las Vegas, Nevada. February 24-26,1993. Volume 2, Pages
1135-1151.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity
and/or toxicity of waste at the point of generation. Numerous opportunities for pollution prevention
exist in laboratory operation. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of first choice.
Whenever feasible, laboratory personnel should use pollution prevention techniques to address their
waste generation. When wastes cannot be feasibly reduced at the source, the Agency recommends
recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Less is Better: Laboratory Chemical management for Waste Reduction
available from the American Chemical Society's Department of Government Relations and Science
Policy, 1155 16th Street N.W., Washington D.C. 20036, (202) 872-4477.
15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society at the address listed in Sec. 14.2.
16.0 REFERENCES
1. Metorex. X-MET 920 User's Manual.
6200 - 21
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2. Spectrace Instruments. 1994. Energy Dispersive X-ray Fluorescence Spectrometry: An
Introduction.
3. TN Spectrace. Spectrace 9000 Field Portable/Benchtop XRF Training and Applications
Manual.
4. Unpublished SITE data, recieved from PRC Environment Management, Inc.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 through 9 and a method procedure flow diagram.
6200 - 22
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TABLE 1
INTERFERENCE FREE DETECTION LIMITS
Analyte
Chemical
Abstract
Series Number
Detection Limit in
Quartz Sand
(milligrams per kilogram)
Antimony (Sb)
7440-36-0
40
Arsenic (As)
7440-38-0
40
Barium (Ba)
7440-39-3
20
Cadmium (Cd)
7440-43-9
100
Calcium (Ca)
7440-70-2
70
Chromium (Cr)
7440-47-3
150
Cobalt (Co)
7440-48-4
W
Copper (Cu)
7440-50-8
50
Iron (Fe)
7439-89-6
60
Lead (Pb)
7439-92-1
20
Manganese (Mn)
7439-96-5
70
Mercury (Hg)
7439-97-6
30
Molybdenum (Mo)
7439-93-7
10
Nickel (Ni)
7440-02-0
50
Potassium (K)
7440-09-7
200
Rubidium (Rb)
7440-17-7
10
Selenium (Se)
7782-49-2
40
Silver (Ag)
7440-22-4
70
Strontium (Sr)
7440-24-6
10
Thallium (Tl)
7440-28-0
20
Thorium (Th)
7440-29-1
10
Tin (Sri)
7440-31-5
60
Titanium (Ti)
7440-32-6
50
Vanadium (V)
7440-62-2
50
Zinc (Zn)
7440-66-6
50
Zirconium (Zr)
7440-67-7
10
Source: References 1,2, arid 3
6200-23
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TABLE 2
SUMMARY OF RADIOISOTOPE SOURCE CHARACTERISTICS
Source
Activity
ftnCO
Half-Life
(Years)
Excitation Energy
(keV)
Elemental Analysis Range
Fe-55
20-50
2.7
5.9
Sulfur to Chromium K Lines
Molybdenum to Barium L Lines
Cd-109
5-30
1.3
22.1 and 87.9
Calcium to Rhodium - K Lines
Tantalum to Lead K Lines
Barium to Uranium L Lines
Am-241
5-30
458
26.4 and 59.6
Copper to Thulium K Lines
Tungsten to Uranium L Lines
Cm-244
60-100
17.8
14.2
Titanium to Selenium K Lines
Lanthanum to Lead L Lines
Source: Reference 1,2, and 3
TABLE 3
SUMMARY OF X-RAY TUBE SOURCE CHARACTERISTICS
Anode
Material
Recommended
Voltage Range
(kV)
K-alpha
Emission
(keV)
Elemental Analysis Range
Cu
18-22
8.04
Potassium to Cobalt
Silver to Gadolinium
K Lines
L Lines
Mo
40-50
17.4
Cobalt to Yttrium
Europium to Radon
K Lines
L Lines
Ag
50-65
22.1
Zinc to Technicium
Ytterbium to Neptunium
K Lines
L Lines
Source: Reference 4
Notes: The sample elements excited are chosen by taking as the lower limit the same ratio of
excitation line energy to element absorption edge as in Table 2 (approximately 0.45) and the
requirement that the excitation line energy be above the element absorption edge as the upper
limit (L2 edges used for L lines). K-beta excitation lines were ignored.
6200 - 24
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TABLE 4
FIELD-BASED METHOD DETECTION LIMITS (mg/kg)*
Analyte
Instalment
TN
9000
TN Lead
Analyzer
X-MET 920
(SiLi
Detector)
X-MET 920
(Gas-Filled
Detector)
XL
Spectrum
Analyzer
MAP
Spectrum
Analyzer
Antimony
55
NR
NR
NR
NR
NR
Arsenic
60
50
55
50
110
225
Barium
60
NR
30
400
NR
NR
Chromium
200
460
210
110
900
NR
Cobalt
330
NR
NR
NR
NR
NR
Copper
85
115
75
100
125
525
Lead
45
40
45
100
75
165
Manganese
240
340
NR
NR
NR
NR
Molybdenum
25
NR
NR
NR
30
NR
Nickel
100
NR
NA
NA
NA
NR
Rubidium
30
NR
NR
NR
45
NR
Strontium
35
NR
NR
NR
40
NR
Tin
85
NR
NR
NR
NR
NR
Zinc
80
95
70
NA
110
NA
Zirconium
40
NR
NR
NR
25
NR
Source: Reference 4
8 MDLs are related to the total number of counts taken. See Section 13,3 for count times
used to generate this table.
NR Not reported.
NA Not applicable; analyte was reported but was not at high enough concentrations for method
detection limit to be determined.
6200 - 25
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TABLE 5
PRECISION
Analyte
Average Relative Standard Deviation for Each instrument
at 5 to 10 Times the MDL
TN
9000
TN Lead
Analyzer
X-MET 920
(SiLi
Detector)
X-MET 920
(Gas-Filled
Detector)
XL
Spectrum
Analyzer
MAP
Spectrum
Analyzer
Antimony
6.54
NR
NR
NR
NR
NR
Arsenic
5.33
4.11
3.23
1.91
12.47
6.68
Barium
4.02
NR
3.31
5.91
NR
NR
Cadmium
29.84'
NR
24.80"
NR
NR
NR
Calcium
2.16
NR
NR
NR
NR
NR
Chromium
22.25
25.78
22.72
3.91
30.25
NR
Cobalt
33.90
NR
NR
NR
NR
NR
Copper
7.03
9.11
8.49
9.12
12.77
14.86
Iron
1.78
1.67
1.55
NR
2.30
NR
Lead
6.45
5.93
5.05
7.56
6.97
12.16
Manganese
27.04
24.75
NR
NR
NR
NR
Molybdenum
6.95
NR
NR
NR
12.60
NR
Nickel
30.85®
NR
24.92*
20.92"
NA
NR
Potassium
3.90
NR
NR
NR
NR
NR
Rubidium
13.06
NR
NR
NR
32.69"
NR
Strontium
4.28
NR
NR
NR
8.86
NR
Tin
24.32"
NR
NR
NR
NR
NR
Titanium
4.87
NR
NR
NR
NR
NR
Zinc
7.27
7.48
4.26
2.28
10.95
0.83
Zirconium
3.58
NR
NR
NR
6.49
NR
Source: Reference 4
a
NR
NA
These values are biased high because the concentration of these analytes in the soil
samples was near the detection limit for that particular FPXRF instrument.
Not reported.
Not applicable; analyte was reported but was below the method detection limit.
6200 - 26
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TABLE 8
PRECISION AS AFFECTED BY SAMPLE PREPARATION
Analyte
Average Relative Standard Deviation for Each Preparation Method
In Situ-Field
Intrusive-
Undried and Unground
Intrusive-
Dried and Ground
Antimony
30.1
15.0
14.4
Arsenic
22.5
5.36
3.76
Barium
17,3
3.38
2.90
Cadmium®
41.2
30.8
28.3
Calcium
17.5
1.68
1.24
Chromium
17.6
28.5
21.9
Cobalt
28.4
31.1
28.4
Copper
26.4
10.2
7.90
Iron
10.3
1.67
1.57
Lead
25.1
8.55
6.03
Manganese
40.5
12.3
13.0
Mercury
ND
ND
ND
Molybdenum
21.6
20.1
19.2
Nickel"
29.8
20.4
18.2
Potassium
18.6
3.04
2.57
Rubidium
29.8
16.2
18,9
Selenium
ND
20.2
19.5
Silver®
31.9
31.0
29.2
Strontium
15.2
3.38
3.98
Thallium
39.0
16.0
19.5
Thorium
NR
NR
NR
Tin
ND
14.1
15.3
Titanium
13.3
4.15
3.74
Vanadium
NR
NR
NR
Zinc
26.6
13.3
11.1
Zirconium
20.2
5.63
5.18
Source: Reference 4
9 These values may be biased high because the concentration of these anaiytes in the soil
samples was near the detection limit
ND Not detected.
NR Not reported.
6200-27
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TABLE 7
ACCURACY
Analyte
Instrument
TN 9000
TN Lead Analyzer
X-MET 920 (SiLi Detector)
XL Spectrum Analyzer
n
Range
of
% Rec.
Mean
% Rec.
SD
n
Range
of
% Rec.
Mean
%
Rec.
SD
n
Range
of
% Rec.
Mean
%
Rec
SD
n
Range
of
% Rec.
Mean
%
Rec.
SD
Sb
2
100-149
124.3
NA

—a

«•»

As
5
68-115
92.8
17.3
5
44-105
83.4
23.2
4
9.7-91
47.7
39.7
5
38-535
189.8
206
Ba
9
98-198
135.3
36.9

9
18-848
168.2
262
--

Cd
2
99-129
114.3
NA

6
81-202
110.5
45.7

__
Cr
2
99-178
138.4
NA
__

7
22-273
143.1
93.8
3
98-625
279.2
300
Cu
8
61-140
95.0
28.8
6
38-107
79.1
27.0
11
10-210
111.8
72.1
8
95-480
203.0
147
Fe
6
78-155
103.7
26.1
6
89-159
102.3
28.6
6
48-94
80.4
16.2
6
26-187
108.6
52.9
Pb
11
66-138
98.9
19.2
11
68-131
97.4
18.4
12
23-94
72.7
20.9
13
80-234
107.3
39.9
Mn
4
81-104
93.1
9.70
3
92-152
113.1
33.8
__

9mm

. .

__
Ni
3
99-122
109.8
12.0

__
__

•**.
WW

3
57-123
87.5
33.5
Sr
8
110-178
132.6
23.8
__

_ •

__
7
86-209
125.1
39.5
Zn
11
41-130
94.3
24.0
10
81-133
100.0
19.7
12
46-181
106.6
34.7
11
31-199
94.6
42.5
Source: Reference 4
n Number of samples that contained a certified value for the analyte and produced a detectable concentration from the FPXRF instrument.
SD Standard deviation.
NA Not applicable; only two data points, therefore, a SD was not calculated.
%Rec. Percent recovery.
- No data.
6200 - 28
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TABlc8
ACCURACY FOR TN 9000*
Standard
Reference
Material
Arsenic
Barium
Copper
Lead
Zinc
Cert.
Cone.
Meas.
Cone.
%Rec.
Cert.
Cone.
Meas.
Cone.
%Rec.
Cert.
Cone.
Meas.
Cone.
%Rec.
Cert.
Cone.
Meas.
Cone.
%Rec.
Cert.
Cone.
Meas.
Cone.
%Rec.
RTC CRM-021
24.8
ND
NA
586
1135
193.5
4792
2908
60.7
144742
149947
103.6
546
224
40.9
RTC CRM-020
397
429
92.5
22.3
ND
NA
753
583
77.4
5195
3444
66.3
3022
3916
129.6
BCR CRM 143R
-
-
-
-
-
—
131
105
80.5
180
206
114.8
1055
1043
99.0
BCR CRM 141
—
-
—
-
-
—
32.6
ND
NA
29.4
ND
NA
81.3
ND
NA
USGS GXR-2
25.0
ND
NA
2240
2946
131.5
76.0
106
140.2
690
742
107.6
530
596
112.4
USGS GXR-6
330
294
88.9
1300
2581
198.5
66.0
ND
NA
101
80.9
80.1
118
ND
NA
NIST 2711
105
104
99.3
726
801
110.3
114
ND
NA
1162
1172
100.9
350
333
94.9
NIST 2710
626
722
115.4
707
782
110.6
2950
2834
96.1
5532
5420
98.0
6952
6476
93.2
NIST 2709
17.7
ND
NA
968
950
98.1
34.6
ND
NA
18.9
ND
NA
106
98.5
93.0
NIST 2704
23.4
ND
NA
414
443
107.0
98.6
105
106.2
161
167
103.5
438
427
97.4
CNRC PACS-1
211
143
67.7
-
772
NA
452
302
66.9
404
332
82.3
824
611
74.2
SARM-51
-
-
-
335
466
139.1
268
373
139.2
5200
7199
138.4
2200
2676
121.6
SARM-52
-
-
-•
410
527
128.5
219
193
88.1
1200
1107
92.2
264
215
81.4
Source: Reference 4
* All concentrations in milligrams per kilogram.
%Rec. Percent recovery.
ND Not detected.
NA Not applicable.
- No data.
6200 - 29
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TABLE 9
REGRESSION PARAMETERS FOR COMPARABILITY1

Arsenic
Bar
um
Copper
n
r2
Int.
Slope
n
r2
Int.
Slope
n
r2
Int.
Slope
Ml Data
824
0.94
1.62
0.94
1255
0.71
60.3
0.54
984
0.93
2.19
0.93
Soil 1
368
0.96
1.41
0.95
393
0.05
42.6
0.11
385
0.94
1.26
0.99
Soil 2
453
0.94
1.51
0.96
462
0.56
30.2
0.66
463
0.92
2.09
0.95
Soil 3
—
—
— •
—'
400
0.85
44.7
0.59
136
0.46
16.60
0.57
Prep 1
207
0.87
2.69
0.85
312
0.64
53.7
0.55
256
0.87
3.89
0.87
Prep 2
208
0.97
1.38
0.95
315
0.67
64.6
0.52
246
0.96
2.04
0.93
Prep 3
204
0.96
1.20
0.99
315
0.78
64.6
0.53
236
0.97
1.45
0.99
Prep 4
205
0.96
1.45
0.98
313
0.81
58.9
0.55
246
0.96
1.99
0.96

Lead

Zinc

Chromium

n
r2
Int.
Slope
n
r2
Int.
Slope
n
r2
Int.
Slope
Ml Data
1205
0.92
1.66
0.95
1103
0.89
1.86
0.95
280
0.70
64.6
0.42
Soil 1
357
0.94
1.41
0.96
329
0.93
1.78
0.93
—
—
—
—
Soil 2
451
0.93
1.62
0.97
423
0.85
2.57
0.90
—
—
—
—
Soil 3
397
0.90
2.40
0.90
351
0.90
1.70
0.98
186
0.66
38.9
0.50
Prep 1
305
0.80
2.88
0.86
286
0,79
3.16
0.87
105
0.80
66.1
0.43
Prep 2
298
0.97
1.41
0.96
272
0.95
1.86
0.93
77
0.51
81.3
0.36
Prep 3
302
0.98
1.26
0.99
274
0.93
1.32
1.00
49
0.73
53.7
0.45
Prep 4
300
0.96
1.38
1.00
271
0.94
1.41
1.01
49
0.75
31.6
0.56
Source: Reference 4
1 Log-transformed data
n Number of data points
r2 Coefficient of determination
Int. Y-intercept
— No applicable data
6200 - 30
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METHOD 6200
FIELD PORTABLE X-RAY FLUORESCENCE SPECTROMETRY FOR THE
DETERMINATION OF ELEMENTAL CONCENTRATIONS IN SOIL AND SEDIMENT
Start
11.2
Type of
analysis
mode.
intrusive
In situ
/ Sample N
homogenization
before
v drying? ,
No
11.3 Perform analysis.
Follow preparation
procedure to achieve
your DQOs.
11.4 Collect sample from
a 4 i 4 inch square of
soli.
11.1 Fallow manufacturers' manual
for operation of FPXRF instrumentation.
11.3 Remove debris from
soil surface and level
surface, if necessary. Tap
soil to increase density
and compactness.
Yes
Stop
11.6 Ground sample until 90%
of original sample passes
through a 60-mesh sieve.
11.6 Place sample in
polyethylene sample cup and
perform analysis.
11.5 Dry 20 - 50 grams of
sample for 2 - 4 hours at a
tamp, no greater than ISO «C.
11.4 Thoroughly mi* sample
in a beaker or plastic bag. Monitor
homogenization with sodium
fluorescein dye.
6200 - 31
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METHOD 6500
DISSOLVED INORGANIC ANIONS IN AQUEOUS MATRICES
BY CAPILLARY ION ELECTROPHORESIS
1.0 SCOPE AND APPLICATION
1.1 This test method is applicable for determination of the dissolved inorganic anions;
fluoride, bromide, chloride, nitrite, nitrate, ortho-phosphate, and sulfate in aqueous matrices using
capillary ion electrophoresis with indirect UV detection.
1.2 This test method is applicable to drinking water, wastewater and ground water for the
analysis of inorganic anions in the concentration range of 0.1 to 50 mg/L, except for fluoride, which
has a range of 0.1 to 25 mg/L. It is the user's responsibility to ensure the applicability of this test
method for other anion concentration ranges and other aqueous sample matrices.
1.3 Capillary ion electrophoresis provides a simultaneous separation and determination of
several inorganic anions using nanoliters of sample in a single injection. Only 500 |j(- of sample is
required to fill the analysis vial. Analysis time is less than 5 minutes.
2.0 SUMMARY OF METHOD
2.1 Capillary ion electrophoresis (Figs. 1 - 4) is a free zone electrophoretic technique
optimized for the analysis of anions with molecular weights less than 200. The anions migrate and
are separated according to their mobility in the electrolyte when an electrical field is applied through
the open tubular fused silica capillary. The electrolyte's electroosmotic flow (EOF) modifier
dynamically coats the inner wall of the capillary, changing the surface to a net positive charge. This
reversal of wall charge reverses the natural EOF. The modified EOF in combination with a negative
power supply augments the mobility of the analyte anions towards the anode and detector achieving
rapid analysis times. Cations migrate in the opposite direction towards the cathode and are removed
from the sample during analysis. Water and other neutral species move toward the detector at the
same rate as the EOF. The neutral species migrate slower than the analyte anions and do not
interfere with anion analysis (Figs. 2 and 3).
2.2 The sample is introduced into the capillary using hydrostatic sampling. The inlet of the
capillary, containing electrolyte, is immersed in the sample and the sample raised 10 cm for 30
seconds where 36 nanoliter volumes are siphoned into the capillary. After sample loading, the
capillary is immediately immersed bade into the electrolyte. The voltage is applied initiating the
separation process. Pressure injection may also be used as long as the performance specifications
of this method are achievable.
2.3 Anion detection is based upon the principles of indirect UV detection. The UV
absorbing electrolyte anion is displaced charge-for-charge by the separated analyte anion. The
analyte anion zone has a net decrease in background absorbance. This decrease in UV absorbance
is quantitatively proportional to analyte anion concentration (Fig. 4). Detector output polarity is
reversed to provide positive mV response to the data system, and to make the negative absorbance
peaks appear positive.
2.4 The analysis is complete once the last anion of interest is detected. The capillary is
then vacuum purged by the system of any remaining sample, and replenished with fresh electrolyte.
The system is now ready for the next analysis.
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3.0 DEFINITIONS
3.1 Capillary Ion Electrophoresis: An electrophoretic technique in which an UV absorbing
electrolyte is placed in a 75 pm fused silica capillary. Voltage is applied through the capillary causing
electrolyte and anions to migrate towards the anode and through the capillary's UV detector window.
Anions are separated based upon the anion's differential rates of migration in Hie electrical field
which is directly related to the anion's equivalent ionic conductance. Anion detection and quantitation
are based upon the principles of indirect UV detection.
3.2 Electrolyte: A combination of a UV absorbing salt and an electroosmotic flow modifier
placed inside the capillary, used as a carrier for the anaiytes, and for anion detection and
quantitation. The UV absorbing portion of the salt must be anionic and have an elect rophoretic
mobility similar to the analyte anions of interest.
3.3 Electroosmotic Flow (EOF): The direction and velocity of electrolyte solution flow within,
the capillary under an applied electrical potential (voltage); the velocity and direction of flow is
determined by electrolyte chemistry, power supply polarity and applied voltage.
3.4 Electroosmotic Flow Modifier (OFM): A cationic amine in the electrolyte that dynamically
coats the negatively charged silica wall reversing the direction of the electrolyte's natural
electroosmotic flow and directing it towards the anode and detector. This modifier augments anion
migration and enhances speed of analysis (Fig. 2).
3.5 Electrophoretic Mobility: The specific velocity of a charged analyte in the electrolyte
under specific electroosmotic flow conditions. The mobility of an analyte is directly related to the
analyte's equivalent ionic conductance and applied voltage, and is the primary mechanism of
separation.
3.6 Electropherogram: A graphical presentation of UV detector response versus time of
analysis; the x axis is the migration time which is used to qualitatively identify the anion, and the y
axis is the UV response which can be converted to time corrected peak area for quantification.
3.7 Hydrostatic Sampling: A sample introduction technique in which the capillary with
electrolyte is immersed in the sample, and both are elevated to a specific height, typically 10 cm,
above the receiving electrolyte reservoir for a preset amount of time, typically less than 60 seconds.
Nanoliters of sample are siphoned into the capillary by differential head pressure and gravity.
3.8 Indirect UV Detection: A form of UV detection in which the analyte displaces an
equivalent net charge amount of the highly UV absorbing component of the electrolyte causing a net
decrease in background absorbance. The magnitude of the decreased absorbance is directly
proportional to analyte concentration. Detector output polarity is switched in order to obtain a positive
mV response.
3.9 Migration Time: The time required for a specific analyte to migrate through the capillary
to the detector. The migration time in capillary ion electrophoresis is analogous to retention time in
chromatography.
3.10 Time Corrected Peak Area (normalized peak area): Peak area divided by migration
time. CIE principles state that peak area is dependant on migration time, i.e. for same concentration
of analyte, as migration time increases (decreases) peak area increases (decreases). Timed
corrected peak area accounts for these changes.
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3.11 Midpoint of Peak Width - CIE peaks are typically asymmetrical with the peak apex
shifting with increasing concentration, and peak apex may not be indicative of true analyte migration
time. Midpoint of peak width is the midpoint between the analyte peak's start and stop integration.
4.0 INTERFERENCES
4.1 The most difficult quantitation and possible comigration occurs when one anion is in
significant excess to other anions in close proximity. For two closely adjacent peaks reliable
quantitation can be achieved when the concentration differential is less than 100:1. As the resolution
between two anion peaks increase so does the tolerated concentration differential.
4.2 Dissolved carbonate, as HC03"1, is an anion present in all aqueous environmental
samples, especially alkaline samples. Under the defined analysis conditions, carbonate at less than
1000:1 concentration differential to the anions will not interfere with the quantitation of the anions
listed in Section 1.1.
4.3 Most monovalent organic acids and neutral organic® commonly found in wastewater
and groundwater migrate later in the electropherogram, after carbonate, and do not interfere with
the anions listed in Section 1.1. Formate, a common organic acid found in environmental samples,
migrates shortly after fluoride but before phosphate. At high formate concentrations the
quantification of fluoride may be incorrectly identified. Include 5 mg/L formate into the mixed anion
working solution to aid with fluoride identification and quantitation (Fig. 5).
4.4 Other inorganic or organic anions present in the sample will be separated and detected
yielding an anionic profile of the sample. Other matrix anions commonly found in drinking water or
wastewater do not interfere with the analysis of anions given in Section 1.1. However, unknown
matrix anions may co-migrate or be a direct interferant with the analyte anions of interest.
4.5 Divalent organic acids usually found in wastewater migrate after phosphate. At
concentrations greater than 10 mg/L, they may interfere with phosphate identification and
quantitation.
4.6 Chlorate also migrates in the phosphate region but does not interfere with phosphate
identification or quantitation at concentrations less than 3 mg/L For chlorate concentrations greater
than 3 mg/L, add 5 mg/L chlorate to the mixed anion working solution to aid in identification of
phosphate and chlorate.
4.7 As the concentration of analyte increases the analyte peak shape becomes
asymmetrical. If adjacent analyte peaks are not baseline resolved, the data system will drop a
perpendicular line between them to the baseline. This causes a decrease in peak area for both
analyte peaks and a low bias for analyte amounts. For optimal quantitation, ensure that adjacent
peaks are fully resolved, if they are not, dilute the sample 1:1 with reagent water.
5.0 SAFETY
5.1 Refer to Chapter Three for additional guidance on safety protocols.
5.2 It is the responsibility of the user to prepare, handle, and dispose of electrolyte solutions
in accordance with all applicable federal, state, and local regulations.
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WARNING — This capillary electrophoresis method uses high voltage as a means for
separating the analyte anions, and can be hazardous if not used properly. Use only those
instruments with the appropriate safety features.
6.0 EQUIPMENT AND SUPPLIES
6.1 Capillary Ion Electrophoresis System: Consists of the following components, as shown
in Fig. 1, or equivalent.
6.1.1 High Voltage Power Supply: Capable of generating voltage potential between
0 and minus 30 kV relative to ground.
6.1.2 Covered Sample Carousel: To prevent environmental contamination of the
samples during a multi-sample analysis.
6.1.3 Sample Introduction Mechanism: Capable of hydrostatic or pressure sampling
techniques.
6.1.4 Capillary Purge Mechanism: To automatically purge the capillary after every
analysis to eliminate any cross contamination from the previous sample matrix and to replenish
the capillary with fresh electrolyte, or clean the capillary with other reagents, such as sodium
hydroxide.
6.1 J UV Detector Capable of monitoring 254 nm with a time constant of 0.1 s.
6.1.6 Fused Silica Capillary: A 75 |jm (inner diameter) x 375 |im (outer diameter)
x 60 cm (length) having a polymer coating for flexibility, and a non-coated section to act as the
cell window for UV detection.
6.1.7 Constant Temperature Compartment: To keep the samples, capillary and
electrolytes at constant temperature.
6.2 Data System: Computer system capable of acquiring data at 20 points per second,
ability to express migration time or relative migration time in minutes to 3 decimal places, use
midpoint of the analyte peak width to determine the migration time of the analyte, use reference
peaks and normalized migration time relative to the reference peak for qualitative identification,
report time corrected peak area, and express results in concentration units.
6.3 Anion exchange cartridge, hydroxide form or equivalent.
6.4 Plastic syringes, 20 mL disposable.
6.5 Vacuum filtration apparatus using a 0.45 pm aqueous compatible filter.
7.0 REAGENTS AND STANDARDS
7.1 Purity of Reaoents: Unless otherwise indicated, it is intended that all reagents shall
conform to the reagent grade specification of the Analytical Reagents of the American Chemical
Society, where such specifications are available. Other grades may be used, provided it is first
ascertained that the reagent is of sufficient high purity to permit its use without lessening the
6500-4
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performance or accuracy of the determination. Detection limits of this method are limited by the
purity of the reagents.
7.2 Reagent Water: All references to water in this method refer to reagent water unless
otherwise specified. Reagent water will be interference free. Refer to Chapter One for a definition
of reagent water,
7.3 Individual Anion Solution. Stock Standard f1000 ma/L AnionV Individual stock solution
may be purchased from an appropriate vendor or may be prepared in the laboratory. Recommend
use of certified 1000 ppm stock standards.
NOTE: All weights given are for anhydrous or dried salts,
7.3.1 Bromide Solution. Standard: Dry approximately 0.5 g of sodium bromide
(NaBr) for 6 hours at 150°C and cool in a desiccator. In 100 ml volumetric flask dissolve 0.128
g of the diy salt with water, and fill to mark with water.
7.3.2 Chloride Solution. Standard: Dry approximately 0.5 g of sodium chloride
(Nad) for 1 hour at 100°C and cool in a desiccator. In 100 mL volumetric flask dissolve 0.165
g of the dry salt with water, and fill to mark with water.
7.3.3 Fl wri
-------
7.4 Mixed Anion Solution, Woffcing: Prepare a blank, and at least 3 different working
standard concentrations for the analyte anion of interest within the desired range of analysis, typically
between 0.1 and 50 mg/L. To a pre-rinsed 100 ml. volumetric flask add an appropriate aliquot of
individual anion stock standard solution (Section 7.3) and dilute with water. Add 5 mg/L formate to
all standards.
NOTE: Use 0.1 ml_ of individual anion stock standard solution (Section 7.3) per 100 mL for
1 mg/L anion.
NOTE: Anions of no interest may be omitted.
NOTE: The mid-range mixed anion working solution (Section 7,4) may be used for the
determination of migration times and resolution described in Section 10.1, and for quality
control evaluation described in Section 9.0.
7.5 Electrolyte Reagents: Although any electrolyte meeting the performance criteria of this
method may be used. This method has boen validated using a chromate-based electrolyte.
7.5.1 Chromate Concentrate: (100 mM Chromate)- in a 1 L volumetric flask dissolve
23.41 g of sodium chromate tetrahydrate (Na2Cr04*4H20) in 500 mL of water, and dilute to 1L
with water. This concentrate may be stored in a capped glass or plastic container for up to 1
year.
7.5.2 Electroosmotic Flow Modifier Concentrate: (100 mM Tetradecyltrimethyl
ammonium bromide, TTABr) - In a 100 mL volumetric flask dissolve 3.365 g of
tetradecyltrimethyl ammonium bromide (TTABr) in 70 mL of water, and dilute to 100 mL with
water.
NOTE: TTABr needs to be converted to the hydroxide form using the anion exchange
cartridge. TTAOH is commercially available from Waters Corp. (sole source).
7.5.3 Buffer Solution: (100 mM CHES/1mM Calcium Gluconate) - In a 1 L
volumetric flask dissolve 20.73 g of CHES (2-[N-Cyclohexy1amino]-Ethane Sulfonic Add) and
0.43 g of calcium gluconate in 500 mL of water, and dilute to 1 L with water. This concentrate
may be stored in a capped glass or plastic container for up to one year.
7.5.4 Sodium Hydroxide Soiution: (500 mM Sodium Hydroxide) - In a 100 mL
volumetric flask dissolve 2 g of sodium hydroxide in 50 mL of water and dilute to 100 m L with
water.
7.5.5 Electrolyte Solution. Working: (4,7 mM Chromate/4 mM TTAOH/10mM
CHES/0.1mM Calcium Gluconate) -Wash the anion exchange cartridge in the hydroxide form
using the 20 mL plastic syringe with 10 mL of 500 mM NaOH followed by 10 mL of water.
Discard the washings. Slowly pass 4 mL of the 100 mM OFM Concentrate Solution through
the cartridge into a 100 mL volumetric flask. Rinse the cartridge with 20 mL of water, adding
the washing to the volumetric flask.
NOTE: The above procedure is used to convert the TTABr to TTAOH which is used
in the electrolyte. If using commercially available 100 mM TTAOH, this step is not
necessary.
6500 -6
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Into the 100 mL volumetric flask add 4.7 mL of chromate concentrate solution and 10
mL buffer solution. Mix and dilute to 100 mL with water. The natural pH of the electrolyte
should be 9.0 ± 0.1. Filter and degass using the vacuum filtration apparatus. Store the
remaining electrolyte in a capped glass or plastic container at ambient temperature. The
electrolyte is stable for one year. This electrolyte is commercially available from Waters Corp.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 Sample collection procedures should address the considerations described in Chapter
Nine of this manual.
8.2 See the introductory material in Chapter Three, Inorganic Analytes, for information on
sample handling and preservation.
8.3 Rinse sampling containers with the sample and discard to eliminate any contamination
from the container, fill to overflowing, and cap to exclude air.
8.4 Analyze samples as soon as possible after collection. For nitrite, nitrate, and
phosphate refrigerate the sample at 4°C after collection and warm to room temperature before
dilution and analysis. Determine nitrite and nitrate within 48 hours.
8.5 Filter samples containing suspended solids through a pre-rinsed 0.45 pm aqueous
compatible membrane filter before transferring the sample to the analysis vial.
8.6 If sample dilution is required, dilute with reagent water only.
9.0 QUALITY CONTROL
9.1 All quality control data should be maintained and available for easy reference or
inspection.
9.2 For each batch of samples processed, method blanks must be carried throughout the
entire sample preparation and analytical process according to the frequency described in Chapter
One. These blanks will be useful in determining if samples are being contaminated. Refer to
Chapter One for the proper protocol when analyzing blanks.
9.3 Matrix Spike/Matrix Spike Duplicates (MS/MSDs): MS/MSDs are intralaboratory split
samples spiked with identical concentrations of target analytes. The spiking occurs prior to sample
preparation and analysis. An MS/MSD is used to document the bias and precision of a method in
a given sample matrix. MS/MSDs are to be analyzed at the frequency of one per analytical batch
as described in Chapter One. Refer to the definitions of bias and precision, in Chapter One, for the
proper data reduction protocols. Each laboratory should calculate its own acceptance criteria based
on its historical data for each matrix type. Refer to Chapter One for guidance.
9.4 A laboratory control sample shall also be processed with each sample batch. Refer to
Chapter One for more information.
9.5 Recalibrate after 15 analyses to account for any changes in migration time or response.
Use the single mixed anion working solution (Sec. 7.4). Replace the new calibration results with the
previous calibration results.
6500 - 7
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9.6 The new calibration curve is validated if the single point calibration response factor of
new recalibration generated in Section 8.5 is ± 5% of the previous calibration response factor, and
if analyte migration time is + 5% of previous migration time determined in Section 10.1.
9.7 If the calibration curve is not validated then discard the spent electrolyte and replace
with fresh electrolyte. Calibrate as described in Section 10.1.
mm: Replace the electrolyte working solution in the instrument daily.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Determination of migration times - The migration time of an anion is dependent upon
the electrolyte compositions, pH, capillary surface and length, applied voltage, the ionic strength of
the sample, and temperature. For every fresh electrolyte determine the analyte migration time in
minutes, to the third decimal place, of the mid-range mixed anion standard working solution (Section
7.4), using the analysis scheme described in (Section 11.0). Use mid-point of analyte peak width
as the determinant of analyte migration time (Fig. 5 and Table 2).
NOTE: Analyte peak apex may be used as the migration time determinant, but potential analyte
misidentification may result with asymmetrical shape at high analyte concentrations.
10.2 For each anion concentration (X-axis) plot time corrected peak area response (Y-axis).
Determine the best linear calibration line through the data points, or use the linear regression
calibration routine available in the data systems. Do not force the line through zero.
10.3 After verification of linear multiple calibration, a single point calibration can be used
between 0.1 and 50 mg/L anion. This single point calibration solution can be used for subsequent
recalibration.
11.0 PROCEDURE
11.1 Set up the capillary electrophoresis system according to the manufacturer's
instructions. Fill the electrolyte reservoirs with fresh electrolyte. Transfer the blank, standard, or
sample into a pre rinsed plastic sample analysis vial and place in the covered sample carousel.
11.2 Program the system according to manufacturer's instructions using the following
instrument settings as guidelines for analysis of standards, and samples.
11.2.1 Condition a new 75 Mm i.d. x 375 jim o.d. x 60 cm capillary with 100 mM
NaOH for 5 minutes followed by working chromate electrolyte solution A for 5 minutes.
NOTE: This conditioning step should be repeated weekly in order to regenerate the
capillary surface for optimum reproducibility.
Program the system for at least a one minute purge of the capillary with electrolyte
between each standard or sample. Using a 15 psi vacuum purge mechanism, one 60 cm
capillary volume can be displaced in 30 seconds.
11.2.2 Program the system for the hydrostatic sampling technique for 30 seconds.
Different sampling times may be used provided that samples and standards are analyzed
identically. Approximately 1.2 nL of sample per second is siphoned into a 75 Mm capillary.
6500 -8
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11.2.3 Program the system for constant current 14 pA and a run time of 5 minutes;
if an anionic profile of the sample is of interest set the time to 7 minutes. Using a capillary 60
cm in length, the field strength at 15 mv applied voltage is 250 V/cm.
11.2.4 Program ttie integrator or computer for data acquisition rate of 20 points per
second with a run time designated in Section 11.2.3. Set up data processing method
according to manufacturers instructions.
11.2.5 Monitor UV response at 254 nm. Since detector ranges are variable, the
range setting required for analysis mil depend on the concentration of anions in the sample
and should be chosen accordingly.
11.2.6 The electropherogram of the working calibration standards (Section 7.4)
should be similar to the inorganic anion electropherogram shown in Fig. 5.
11.3 Analyze all standards (Section 7.4) and samples as described in Section 11.2. Refer
to Figs. 5-9 for representative anion standard, 0.1 mg/L anion standard, drinking water, and waste
water (municipal and industrial).
12.0 DATA ANALYSIS AND CALCULATIONS
12.1 Relate the time corrected peak area lor each sample anion with the calibration curve
generated in Section 10.2 to determine mg/L concentration of anion. If the sample was diluted prior
to analysis, then multiply mg/L anion by the dilution factor to obtain the original sample concentration.
Original Sample mg/L Anion = (A x SF)
where:
A = mg/L anion determined from the calibration curve
SF - scale or dilution factor
13.0 METHOD PERFORMANCE
13.1 Figures 6-12 display representative examples of electropherograms and linearity of
calibration curves.
13.2 Tables 1-10 provide collaborative design, migration time reproducibility, comparison of
CIE with other approved EPA methods, and interteboratory reproducibility and precision for the
capillary ion electrophoresis technique.
13.3 Table 11 is entitled "Capillary Ion Electrophoresis Anion Analysis Round Robin Using
Cbromate Electrolyte (mg/L)" and provides precision data in some common environmental matrices.
13.4 The following documents may provide additional information regarding this method and
technique:
13.4.1 Romano, J., Krol, J, "Capillary Ion Electrophoresis, An Environmental Method
for the Determination of Anions in Water", J. of Chromatography. Vol. 640,1993, p. 403.
13.4.2 Romano, J., "Capillary Ion Analysis: A Method for Determining Ions in Water
and Solid Waste Leachates", Amer. Lab., May 1993, p. 48.
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13.4.3 Jones, W., "Method Development Approaches for Ion Electrophoresis", J. of
Chromatography. Vol. 640,1993, p. 387.
13.4.4 Jones, W., Jandik, P., "Various Approaches to Analysis of Difficult Sample
Matrices for Anions using Capillary Electrophoresis", J. of Chromatography. Vol. 608,1992,
p. 385.
13.4.5 Bondoux, G., Jandik, P., Jones, W.t "New Approaches to the Analysis of Low
Level of Anions in Water", 4, 9f Chromatography, Vol. 602,1992, p. 79.
13.4.6 Jandik, P., Jones, W., Weston, A., Brown, P.,"Electrophoretic Capillary ion
Analysis: Origins, Principles, and Applications", LCGC, Vol. 9, Number 9,1991, p. 634.
13.4.7 Romano, J., Jackson, P., "Optimization of Inorganic Capillary Electrophoresis
for the Analysis of Anionic Solutes in Real Samples", J. of Chromatography. Vol. 546,1991,
p. 411.
13.4.8 Jandik, P., Jones, W., "Optimization of Detection Sensitivity in the Capillary
Electrophoresis of Inorganic Anions", J of Chromatography. Vol. 546,1991, p. 431.
13.4.9 Jandik, P., Jones, W., "Controlled Changes of Selectivity in the Separation
of tons by Capillary Electrophoresis", J. of Chromatography. Vol. 546,1991, p 445.
13.4.10 Foret, R„ et.al.t "Indirect Photometric Detection in Capillary Zone
Electrophoresis", J. of Chromatography. Vol. 470,1989, p. 299.
13.4.11 Hjerte'n, S. et. al.,"Carrier-free Zone Electrophoresis, Displacement
Electrophoresis and Isoelectric Focusing in an Electrophoresis Apparatus", J. of
Chromatography. Vol. 403,1987, p. 47
13.4.12 Jandik, P., Bonn, G., "Capillary Electrophoresis of Small Molecules and tons',
VCH Publishers, 1993.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity
and/or toxicity of waste at the point of generation. Numerous opportunities for pollution prevention
exist in laboratory operation. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of first choice.
Whenever feasible, laboratory personnel should use pollution prevention techniques to address their
waste generation. When wastes cannot be feasibly reduced at the source, the Agency recommends
recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Less is Better, Laboratory Chemical management for Waste Reduction
available from the American Chemical Society, Department of Government Relations and Science
Policy, 1155 16th Street, NW, Washington, DC, 20036, (202) 872-4477.
6500 -10
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January 19%
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15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling ail releases from hoods and tench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society at the address listed in Section 14.2.
18.0 REFERENCES
1. Waters Chromatography, "Innovative Methods for Ion Analysis", Method N-601b, 1992.
2. Waters Chromatography, Validation Data for Method 6500, Millipore Corporation Waters
Chromatography Division, Ion Analysis Group; Milford, Massachusetts.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Table 1 through 11, Figures 1 through 12, and a flow diagram of
method procdures.
6500-11
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January 1999
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TABLE 1
COLLABORATIVE DESIGN AS FOUR YOUDEN PAIR SETS1
Individual Youden Pair Standards, in nrig/L

1
2
3
4
5
6
7
8
a
0.7
2.0
3.0
15.0
40.0
20.0
50.0
0.5
Br
2.0
3.0
15.0
40.0
20.0
50.0
0.7
0.5
NO;
3.0
40.0
20.0
15.0
50.0
0.5
2.0
0.7
so4
40.0
50.0
0.5
0.7
2.0
3.0
15.0
20.0
NOs
15.0
20.0
40.0
50.0
0.5
0.7
2.0
3.0
F
2.0
0.7
0.5
3.0
10.0
7.0
20.0
25.0
po4
50.0
40.0
20.0
0.5
3.0
2.0
0.7
15.0
Source: Reference 2
^The collaborative design is intended to demonstrate performance between 0.1 and 50 mg/L
anion, except for Fluoride between 0.1 and 25 mg/L. The concentrations among anions is
varied as not to have any one standard at all lowor all high anion concentrations.
6500 -12
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January 1998
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TABLE 2
ANION MIGRATION TIME REPRODUCIBILITY FROM YOUDEN PAIR STANDARDS
USING CHROMATE ELECTROLYTE AND CONSTANT CURRENT
Analyte Mid-Point Migration Time, Ave of Triplicate Samplings
Analyte
CI
Br
no2
CO
p
NO,
F
po4
1
3.132
3.226
3.275
3.405
3.502
3.761
3.906
2
3.147
3.239
3.298
3.431
3.517
3.779
3.931
3
3.138
3.231
3.283
3.411
3.497
3.771
3.925
4
3.158
3.244
3.307
3.434
3.510
3.781
3.963
5
3.184
3.271
3.331
3.435
3.551
3.787
3.981
6
3.171
3.260
3.312
3.418
3.537
3.776
3.964
7
3.191
3.272
3.315
3.437
3.544
3.773
3.978
8
3.152
3.248
3.294
3.418
3.526
3.739
3.954
Std Dev
0.021
0.015
0.018
0,012
0.20
03015
0.027
%RSD
0.67%
0.46%
0.55%
0.36%
0.56%
0.40%
0.68%
Ave Migration Time Std Dev = 0.018 min =1.1 sec Ave %RSD = 0.53%
6500-13
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January 1998
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TABLE 3
COMPARISON OF CAPILLARY ION ELECTROPHORESIS WITH CHROMATE
ELECTROLYTE AND APPROVED METHODS USING A PERFORMANCE EVALUATION
STANDARD

Analyte
CI
O
z
so4
no3
F
po4
Performance
Evaluation
Standard1
True
Value
in mg/L
43.00
1.77
37.20
15,37
2.69
6.29
Official
Anion
Methods
Wet Chem & IC
Measured
Mean2
43.20
1.77
37.00
15.42
2.75
6.38
Measured
Std Dev
3.09
0.07
2 24
1.15
0.26
0.21
CIE Using
Chromate
Electrolyte3
Ave CIE
n=18
42.51
1.78
37.34
14.06
2.63
6.34
CIE/Mean
CIE/True Value
0.984
0.989
1.006
1.006
1.009
1.003
0.911
0.945
0.956
0.978
0.994
1.008
Source: Reference 2
The performance evaluation standard was purchased from APG Laboratories and diluted 1:100 with
Type I Dl water.
The measured result is the average from numerous laboratories using Approved Standard Methods
and EPA wet chemistry and ion chromatography methods
*The CIE results were determined using the proposed EPA and ASTM method, and are the average
from 4 laboratories using the Youden Pair Standards for quantitation.
6500 -14
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January 1998
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TABLE 4
CAPILLARY ION ELECTROPHORESIS WITH CHROMATE ELECTROLYTE
INTERLABORATORY REPRODUCIBILITY AND PRECISION1
Analyte2
CI
no2
CO
p
NO,
F
Lab 1
n = 5
43.22 ±
0.22
1.58 ±
0.09
36.39 ±
0.33
14.57 ±
0.12
2.54 ±
0.10
Lab 2
ns5
43.68 ±
0.61
1.58 ±
0.08
37.01 ±
0.37
13.94 ±
0.09
2.69 ±
0.02
Lab 3
n-5
43.93 ±
0.39
1.60 ±
0.06
37.68 ±
0.24
15.05 ±
0.11
2.691
0.03
Lab 4
n=3
42.51 ±
0.22
1.78 ±
0.06
37.34 ±
0.19
14.CK ±
0.07
2.69 ±
0.02
Average Mean
± Std Dev
43.34 ±
0.36
1.64 ±
0.07
37.11 ±
0.28
14.41 ±
0.10
2.64 ±
0.04
% RSD
0.83%
4.5%
0.77%
0.67%
1.61%
1ResuKs from 4 laboratories analyzing the performance evaluation standard using the Youden
Pair Standards for quantitation. Results expressed as mg/L.
2Only 1 lab reported results for PO> as 6.34 ± 0.02 mg/L on triplicate samplings yielding an
%RSD of 0.07%
6500 -15
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TABLE 5
CAPILLARY ION ELECTROPHORESIS WITH CHROMATE ELECTROLYTE KNOWN ADDITION
RECOVERY AND PRECISION USING PERFORMANCE EVALUATION STANDARD WITH
DRINKING WATER
Analyte
Ci
no2
SO4
NOj
F
po4
Milford
Drinking Water
n=3, as ppm
24.27 ±
0.15
Not
Detected
7.99 ±
0.07
0.361
0.05
Not
Detected
Not
Detected
%RSD
0.73%

0.91%
13.3%

Performance
Evaluation Std1
43.00
1.77
37.20
15.37
. 2.69
6.29
MDW+ PES
n=3, as ppm
66.57 ±
0.34
1.74 ±
0.03
45.19 ±
0.17
15.42 ±
0.12
2.62 ±
0.07
5.55 ±
0.31
%RSD
0.51
1.85
0.38
0.79
2.69
5.52
% Recovery
97.9%
98.3%
100.2%
98.1%
97.4%
88.2%
Source: Reference 2.
'The performance evaluation standard was diluted 1:100 with Drinking Water.
6500 -16
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January 1998
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TABLE 6
COMPARISON OF APPROVED METHOD AND CAPILLARY ION ELECTROPHORESIS
WITH CHROMATE ELECTROLYTE FOR THE DETERMINATION OF CHLORIDE
Data given as mg/L
Analyte
Sample #
Titration1
IC2
CIE
Effluent
1
—3
149
147

2
—
162
161

3
__
153
152

4
—
139
140

5
—
111
110

6
_
109
107

7
—
3.6
3.5
Drinking Water
1
5.5
5.1
5.0

2
5.5
5.0
4.9

3
5.3
5.2
5.1

4
5.5
5.1
5.1

5
5.3
5.0
5.0

s
5.3
4.9
4.9

7
5.5
4.9
4.9
Landfill
1
0.1
<0.1
ND
Leachate
2
230
245
240
Source: Reference 2.
1 Chloride determined using 4500 CI C, lodometric Method
2 Chloride determined using 4110 C, Single Column Ion Chromatography Using
Direct Conductivity Detection
3 A dash line indicates test not performed. ND indicates anion not detected
6500-17
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January 1998
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TABLE 7
COMPARISON OF APPROVED METHOD AND CAPILLARY ION ELECTROPHORESIS
WITH CHROMATE ELECTROLYTE FOR THE DETERMINATION OF FLUORIDE
Analyte
Sample #
Electrode1
IC2
CIE
Effluent
1
1.7
1.2
1.5

2
0.9
0.6
0.6

3
0.8
0.5
0.6

4
0.8
0.4
0.7

5
0.9
0.5
0.8

6
0.9
0.5
0.7

7
<0.1
ND
<0.1
Drinking Water
1
1,2
0.9
0.9

2
1.3
0.9
0.9

3
1.3
0.9
0.9

4
1.3
0.9
0.9

5
1.3
0.9
0.9

6
0.9
0.6
0.6

7
1.3
0.9
0.9
Landfill
1
<0.2
ND
ND
Leachate
2
16
10.6
10.9
Source: Reference 2,
1 Fluoride determined using 4500-F C, Ion Selecteive Electrode Method
2 Fluoride determined using 4110 C, Single Column Ion Chromatography Using Direct
Conductivity Detection
6500 -18
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January 1998
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TABLE 8
COMPARISON OF APPROVED METHOD AND CAPILLARY ION ELECTROPHORESIS
WITH CHROMATE ELECTROLYTE FOR THE DETERMINATION OF SULFATE
Data given as mg/L
Analyte
Sample#
Turbidimetric1
10s
CIE
Effluent
1
58
87.5
98.0 "

2
110
95.3
95.9

3
130
118
115

4
130
139
136

5
110
113
110

6
100
107
106

7
6
5.6
5.8
Drinking Water
1
6
5.8
6.0

2
6
5.8
6.0

3
6
5.9
6.1

4
i
5.9
6.1

5
5
5.8
6.2

6
4
3.0
3.4

7
5
5.8
6.1
Landfill
1
<1
ND
ND
Leachate
2
190
211
201
Source: Reference 2.
1 Sulfate determined using 4500 S04 E, Turbidimetric Method
2 Sulfate determined using 4110 C, Single Column ion Chromatography Using Direct
Conductivity Detection
6500 -19
Revision 0
January 1998
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TABLE 9
COMPARISON OF APPROVED METHOD AND CAPILLARY ION ELECTROPHORESIS
WITH CHROMATE ELECTROLYTE FOR THE DETERMINATION OF NITRITE + NITRATE3
Data given as mg/L
Analyte
Sample #
Cd Red'n1
IC*
CIE
Effluent
1
0.3
ND
ND

2
—
ND
ND

3
—
ND
ND

4
—
ND
0.5

5
—
2.1
2.4

6
2.4
1.9
2.2

7
0.7
0.3
- 0.4
Drinking Water
1
0.6
0.3
0.4

2
0.6
0.3
4.4

3
0.4
0.3
4.4

4
0.6
0.3
0.3

5
0.6
0.3
0.4

6
0.3
0.1
0.1

7
0.5
0.3
0.4
Landfill
1

ND
ND
Leachate
2

ND
ND
Source: Reference 2,
1 Total nitrite + nitrate determined using 4500-N03 F, Cadmium Reduction Method
2 Nitrite + nitrate determined using 4110 C, Single Column Ion chromatography Using Direct
Conductivity Detection
3 Each technique gave separte nitrate and nitrate values; because of their liability results were
added for comparison purposes
6500 - 20
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January 1998
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TABLE 10
COMPARISON OF APPROVED METHOD AND CAPILLARY ION ELECTROPHORESIS
WITH CHROMATE ELECTROLYTE FOR THE DETERMINATION OF ORTHO-PHOSPHATE
Data given as mg/L
Analyte
Sample #
Ascorbic Acid1
IC2
CIE
Effluent
1
3,4
ND
2.8 "

2
4,9
ND
4.4

3
4,7
ND
4.5

4
5.3
ND
4.2

5
3.0
ND
3.0

6
2.9
ND
2.3

7
<0.1
ND
<0.1
Drinking Water
1
<0.1
ND
ND

2
<0.1
ND
ND

3
„
ND
ND

4
<0.1
ND
ND

5
<0.1
ND
ND

6
—
ND
ND

7
_
ND
ND
Landfill
1
<0.1
ND
<0.1
Leachate
2
2.2
1.6
1.4
Source: Reference 2.
1 Phosphate determined using 4500 P04 E, Ascorbic Add Method
2 Phosphate determined using 4110 C, Single Column ion Chromatography Using Direct
Conductivity Detection
6500 - 21
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January 1998
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TABLE 11
CAPILLARY ION ELECTROPHORESIS ANION ANALYSIS ROUND ROBIN1
USING CHROMATE ELECTROLYTE (mg/L)
Sample
Chloride
Bromide
Nitrite
Sulfate
Nitrate
Fluoride
Phosphate
1. Bleachwaste
<0.046
<0.046
<0.072
0.30±0.37
<0.84
<0.020
<0.041
2. Creekwater
3.06±0.27
<0.046
<0.072
3.0010.30
0.37±0.19
0.11 ±0.09
<0.061
3. Wastewater
24.6±0.62
<0.046
<0.072
2.02±0.56
<0.084
0.08±0.08
3.74±0.75
4. Wastewater
59.7±2.9
0.85±0.52
<0.072
109±4.4
44.9±1.6
0.988±0.21
4.94±1.32
5. Wastewater
63.8±2.0
0.68±0.52
<0:072
115±3.9
44.3±1.06
1.04±0.17
4.78±1.55
6. Wastewater
72.0±5.4
0.05±0.01
<0:072
144±11.8
5.38±2.57
0.57±0.21
1.18±1.01
7. Wastewater
139±10.0
<0.046
4.0±1.3
584±35
353±25.5
3.01 ±0.80
9.34±5.17
8. Wastewater
51.4±7.7
<0.046
<0.072
40.2±6.1
39.9±7.9
1.17±0.24
6.99±1.31
9. Wastewater
29.9±4.3
<0.046
2.14±1.35
217±19
13.9±4.9
1.33±0.28
9.95±5.04
10. Wastewater
766±44
<0.046
<0.072
489±46
12.9±6.9
<0.020
41.3±8.5
11. Surfacewater
3.71 ±0.39
<0.046
<0.072
2.70±0.39
0.23±0.20
0.11±0.097
<0.041
12. Wastewater
22.1 ±0.62
8.47±0.30
<0.072
133±4.4
<0.084
0.76±0.11
<0.041
13. Drinking Water
5.15±0.35
<0.046
<0.072
2.64±0.26
0.50±0.27
0:59±0.097
<0.041
14. Drinking Water
4.95±0;24
<0.046
<0.072
2.62±0.21
0.54±0.25
0.56±0.09
<0.041
Source: Reference 2.
1 Five laboratory interlaboratory precision.
6500 - 22
Revision 0
January 1998
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FIGURE 1
HARDWARE SCHEMATIC OF A CAPILLARY
ION ELECTROPHORESIS SYSTEM
75 (im x 60 cm
Silica Capillary
Constant
Temperature
Compartment,
25-30 °C
FIGURE 2
PICTORIAL DIAGRAM OF ANION MOBILITY AND
ELECTROOSMATIC FLOW MODIFIER
SH + + + + + + + + + + + + + + +
High Mobility '
Low Mobility Anton
«—(, AUCattom* )
Catftode
Anode
Injection Skis
Dstaction Side
6500 - 23
Revision 0
January 1998
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FIGURE 3
SELECTIVITY DIAGRAM OF ANION MOBILITY
USING CAPILLARY ION ELECTROPHORESIS
Inorganic
Anions
Ci, Br,
NO*,SO<,
NO 3.
SOa, S2O3
DiValent
Org Acids,
Oxymetals
F. PO«,
002, CI03,
Formats
Monovalent
Organic Acids
C2 thru Cs
Migration Time
Water
and All
Neutral
Organics
MT=0
High Mobility
Anions
Low Mobility
Anions
MT >7 min
FIGURE 4
PICTORIAL DIAGRAM OF INDIRECT UV DETECTION
.-ff777W
eeeeeAAAAeeeeeee
eeeeeeAAAAeeeeeee
High UV Absorbing
Electrolyte
Analyle ion (A) displaces electraiye too (e)
cltaiga for charge or transfer ratio causing
a net decrease in background absoifianca.
The change in absorbance is directly
related to Analyte concentration.
6500 - 24
Revision 0
January 1998
-------
FIGURE 5
ELECTROPHEROGRAM OF THE INORGANIC ANIONS AND TYPICALLY FOUND ORGANIC
ACIDS USING CAPILLARY ION ELECTROPHORESIS WITH CHROMATE ELECTROLYTE
a
'
4.500
3.500
4.000
3.000
Minutes
Electrolyte: 4.7 mM Na^rO^.O mM TTAOH /10 mU CHES /0.1 mM Calcium Gluconate
Capiary: 75 |im (id) x 375 |im x 60 cm (length), Uncoated Sica
Vottage: 15 kV using a Negative Power Supply
Current 14 ± (lA, Constant Current
Sampling: Hydrostatic at 10 cm for 30 seconds
Detection: Indirect UV using a Hg Lamp and 254 nm Filter
Anion
Cone.
Mg/L
Migration
Time in
Mintuas
Migration
Time Ratio to
CI
Peak
Area
Time
Corrected
Peak Area
1. Chloride
2.0
3.200
1.000
1204
376.04
2. Bromide
4.0
3.296
1.030
1147
348.05
3. Nitrite
4.0
3.343
1.045
2012
601.72
4. Sulfate
4.0
3.465
1.083
1948
- 562.05
5. Nitrate
4.0
3.583
1.120
1805
503.69
6. Oxalate
5.0
3.684
1.151
3102
842.14
7. Fluoride
1.0
3.823
1.195
1708
446.65
8. Formate
5.0
3.873
1.210
1420
366,61
9. O-Phosphate
4.0
4.004
1.251
2924
730.25
10. Carbonate
-
4.281
1.338
-
-
11. Acetate
5.0
4.560
1.425
3958
868.01
6500 - 25
Revision 0
January 1998
-------
FIGURE 6
ELECTROPHEROGRAM OF 0.1 MG/L INORGANIC ANIONS
MINIMUM DETECTION LIMIT WITH CHROMATE ELECTROLYTE
Seven replicates of the 0.1 mg/L inorganic anion standard was used to calculate the minimum
detection limits, as mg/L, using analytical protocol described in Standard Methods 1030 E.
Chloride = 0.046 Bromide = 0.090 Nitrite ~ 0.072 Sulfate = 0,032
Nitrate = 0.084 Fluoride = 0.020 phosphate = 0.041
PO«
4.000
3.800
Minutes
FIGURE 7
ELECTROPHEROGRAM OF TYPICAL DRINKING WATER
USING CHROMATE ELECTROLYTE
Q = 24.72 mgfl.
SO. = 7.99
NO> = 0.36
F < 0.10
HCO»» Natural
3.S00
3.000
Minutes
6500 - 26
Revision 0
January 1998
-------
FIGURE 8
ELECTROPHEROGRAM OF TYPICAL MUNICIPAL WASTEWATER DISCHARGE
USING CHROMATE ELECTROLYTE
Anions in ma/l. No DMmion
1 CMorMe =93.3
2 Nitrite = 0.46
3 SuHate = 60.3
4 Nitrate = 40,8
5 Carbonate = Natural
3.000
3.500
Minutes
4.000
4,500
FIGURE 9
ELECTROPHEROGRAM OF TYPICAL INDUSTRIAL WASTEWATER DISCHARGE
USING CHROMATE ELECTROLYTE
3
"£
6
JLaJ
3.000
3.500 Minutes 4 000
Anions in ma/L, No Dilution
1 CMorMe = 2.0
= 1.6
=.34.7
¦» 16.5
< 0.05
12.3
2 Nitrite
3 Sulfate =
4 Nitrate *
5 Formate <
6 Phosphate =
Carbonate = Natural
4. £00
6500 - 27
Revision 0
January 1998
-------
FIGURE 10
LINEARTY CALIBRATION CURVE FOR CHLORIDE, BROMIDE, AND SULFATE
USING CHROMATE ELECTROLYTE
10
3 7.5
2.5
TCP* = 187-62 |CI] ~ 32,93
TCP* = 326.76fSO«l -16.12
TCPA= ?8.23[Br|* 11.76
RJ« 0.9996
R' ¦ 0.9996
R»=> 0 9995
3 Data Points per Concentration
Based upon Youden Pair Design
- 1- '
20 30
mg/L Anion
i
50
FIGURE 11
LINEARTY CALIBRATION CURVE FOR FLUORIDE AND O-PHOSPHATE
USING CHROMATE ELECTROLYTE
a. tj
3 Data Points per Concentration
Based upon Youden Pair Design
TCM = 376 67|F] +1005;
• 0.9985
TCW = IM.2S(PO..| -19.68; R'= 0.9996
_T_
20 30
mg/L Anion
40
50
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FIGURE 12
LINEARITY CALIBRATION CURVE FOR NITRITE AND NITRATE
USING CHROMATE ELECTROLYTE
10
«
S
<
7.5
m «
a. «
•a m
0> 01
o g
tB U
2: jc
5 *-
O
I ~ 28.93; «'= 0.9996
TCP* = 1Q6.B2[N0al * 126.81; H'= 0-9992
20 30
mg/L Anion
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METHOD 6500
DISSOLVED INORGANIC ANIONS IN AQUEOUS MATRICES
BY CAPILLARY ION ELECTROPHORESIS
Start
Stop
11.2,5 Monitor UV response
at 254 nm.
11.2.2 Program system for
hydrostatic sampling for
30 seconds.
11.1 Set-up Capillary
Electrophoresis system
according to manufacturers
instructions.
11.2.3 Program system for
constant current 14 u/i and
a run time of 5 min.
11.2.1 Condition capillary
with NaOH for 5 min.
followed by chromate
electrolyte soln. for 5 min.
11,3 Analyze all standards
and samples.
11.2.4 Program system for
data acquisition rate of
20 points per second.
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METHOD 6800
ELEMENTAL AND SPECIATED ISOTOPE DILUTION MASS SPECTROMETRY
1.0 SCOPE AND APPLICATION
1.1 This method consists of two approaches: isotope dilution mass spectrometry (IDMS)
for the determination of total metals and speciated isotope dilution mass spectrometry (SIDMS) for
the determination of elemental species. This method is applicable to the determination of total
metals and metal species at sub pg/L levels in water samples or in waste extracts or digests. In
general, elements that have more than one available stable isotope can be analyzed by IDMS.
SIDMS may require more isotopes of an element, depending on the number of interconvertable
species. The current method is applicable to the following elements.
Element CASRN3
Antimony
(Sb)
7440-36-0.
Boron
(B)
7440-42-8
Barium
(Ba)
7440-39-3
Cadmium
(Cd)
7440-43-9
Calcium
(Ca)
7440-70-2
Chromium
(Or)
7440-47-3
Copper
(Cu)
7440-50-8
Iron
(Fe)
7439-89-6
Lead
(Pb)
7439-92-1
Magnesium
(Mg)
7439-95-4
Mercury
(Hg)
7439-97-6
Molybdenum
(Mo)
7439-98-7
Nickel
(N«)
7440-02-0
Potassium
(K)
7440-09-7
Selenium
(Se)
7782-49-2
Silver
(Ag)
7440-22-4
Strontium
(Sr)
7440-24-6
Thallium
(Tl)
7440-28-0
Vanadium
(V)
7440-62-2
Zinc
(Zn)
7440-66-6
3 Chemical Abstracts Service Registry Number
Other elements and species may be analyzed by this method if appropriate performance is
demonstrated for the analyte of interest, in the matrices of interest, at the concentration levels of
interest (see Section 9.0).
1.2 Isotope dilution is based on the addition of a known amount of enriched isotope to a
sample. Equilibration of the spike isotope with the natural element/species in the sample alters the
isotope ratio that is measured. With the known isotopic abundance of both spike and sample, the
amount of the spike added to the sample, and the altered isotope ratio, the concentration of the
element/species in the sample can be calculated.
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1.3 IDMS has proven to be a technique of high accuracy for the determination of total
metals in various matrices (Reference 1). IDMS has several advantages over conventional
calibration methodologies. Partial loss of the analyte after equilibration of the spike and the sample
will not influence the accuracy of the determination. Fewer physical and chemical interferences
influence the determination as they have similar effects on each isotope of the same element. The
isotope ratio to be measured for quantification in IDMS can be measured with very high precision,
typically RSDsO.25%.
1.4 SIDMS takes a unique approach to speciated analysis that differs from traditional
methods. Traditional speciation methods attempt to hold each species static while making the
measurement. Unfortunately, speciation extraction and analysis methods inherently measure the
species after species conversions have occurred. SIDMS has been developed to address the
correction for the species conversions. In SIDMS, each species is "labeled" with a different isotope-
enriched spike in the corresponding species form. Thus, the interconversions that occur after spiking
are traceable and can be corrected. While SIDMS maintains the advantages of IDMS, it is capable
of correcting the degradation of the species or the interconversion between the species (Reference
2). SIDMS is also a diagnostic tool that permits the evaluation of species-altering procedures and
permits evaluation and validation of other more traditional speciation analysis methods.
1.5 Both IDMS and SIDMS require the equilibration of the spike isotope(s) and the natural
isotopes. For IDMS, the spike and sample can be in different chemical forms; only total elemental
concentrations will result. In general, equilibration of the spike and sample isotopes occurs as a
result of decomposition, which also destroys all species-specific information when the isotopes of
an element are all oxidized or reduced to the same oxidation state. For SIDMS, spikes and samples
must be in the same speciated form. This requires the chemical conversion of the elements in
spikes. For solution samples, spiking and equilibration procedures can be as simple as mixing the
known amount of the sample and the spikes. Efforts are taken to keep the species in their original
species forms after spiking. While drinking water, ground water, and other aqueous samples may
be directly spiked, soils, sludges, sediments, and other solid wastes require extraction or digestion
prior to analysis to solubilize the elemental species.
1.6 Detection limits, sensitivity, and optimum ranges of the elements will vary with the
matrix, separation method, and isotope ratio measurement methods. With the popularity of
chromatography and ICP-MS, it is convenient to separate elemental species and to measure the
isotope ratios. Although this method is not restricted to chromatography as the separation method
of the species and the ICP-MS as the isotope ratio measurement method, this method will use these
two techniques as examples in describing the procedures. Other species separation methods, such
as extraction, precipitation, and solid phase chelation, and other isotope ratio measurement
techniques, such as thermal ionization mass spectrometry (TIMS), can also be used.
2.0 SUMMARY OF METHOD
2.1 IDMS method:
2.1.1 Samples may require a variety of sample preparation procedures, depending
on sample matrices and the isotope ratio measurement methods. One primary purpose of
sample preparation is to solubilize the analyte of interest and equilibrate the spike isotopes
with sample isotopes. Solids, slurries, and suspended material must be subjected to digestion
after spiking using appropriate sample preparation methods (such as Method 3052). Water
samples may not require digestion when ICP-MS is used as a detection method because ICP
can destroy elemental species and thus many species are indistinguishable for ICP-MS.
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2.1.2 A representative measured sample is thoroughly mixed with a measured
amount of the isotopic spike. If a digestion procedure is required, the spiked sample is then
digested to equilibrate the spikes and samples. The sample solutions are then measured with
mass spectrometry such as ICP-MS to obtain the altered isotope ratios. Method 6020 can be
used as a reference method for ICP-MS detection. In addition to Method 6020, dead time
correction and mass bias correction must be included in the measurement protocol. The
equations described in Section 12.1 are used to calculate the concentrations. Figure 2 shows
an example of an IDMS determination of vanadium in crude oil (Reference 1).
2.2 SIDMS method:
2.2.1 Speciated samples generally require sample preparation specific to the
sample matrices, species, and the isotope ratio measurement method, The purpose of sample
preparation is to solubiiize the species of interest and to equilibrate the natural and spiked
species, creating a homogeneous solution. Solids, slurries, and suspended material must be
subjected to extraction before spiking, using appropriate sample preparation methods (such
as Method 3060 for the determination of Cr(VI) in soils). Water samples may not need
extraction. In contrast to total element analysis, efforts must be taken to avoid the destruction
of the species in SIDMS.
2.2.2 Although SIDMS is a general method applicable to many elements in various
species forms, environmental samples, such as water samples or soil extracts, containing
chromium species Cr(ill) and Cr(VI) will be used for demonstration purposes. Two isotopic
spites are prepared and characterized: ^Crflll) spike enriched in ^Cr and ^Cr^/I) enriched in
^Cr. The dominant natural isotope for Cr is 52Cr, at 83.79% f°Cr, 4.35%; ^Cr, 9.50%; ^Cr,
2.36%). A measured amount of a representative aqueous sample is mixed well with an
appropriate amount of MCr(lll) and ^Cr^VI) spike solutions. The spiked sample is then
separated into Cr(lll) and Cr(Vl) using chromatography or other separation method. Four
isotope ratios are measured: ^Crflll^CrCIII), ^CrOliy^Crflll), mCr^/l)FCr0/\), and
^CrCVIJ/^CrCVI). The concentrations of the species are determined from speciated isotope
dilution calculations. Figure 4 and Figure 5 show an example of the SIDMS for the
determination of chromium species in an aqueous sample. Any transformation from Cr(VI) to
Cr(lli) or from Cr(!ll) to Cr(VI) are mathematically corrected, as described in Section 12.2.
3.0 Definitions
3.1 Isotope dilution mass spectrometry (IDMS): A quantitative method for total
concentration of an analyte based on the measurement of the isotope ratio of a nuclide using mass
spectrometry after isotope dilution.
3.2 Isotope dilution: Mixing of a given nuclide with one or more of its isotopes. The isotope
usually has an enriched isotopic abundance different from that occurring naturally.
3.3 Speciated isotope dilution mass spectrometry (SIDMS). A quantitative method for
determining elemental species based on the measurement of isotope ratio(s) in each species of a
nuclide using mass spectrometry after speciated isotope dilution. Samples are mixed with one or
more isotopic spikes which have different isotopic abundance and are artificially converted to
chemical forms corresponding to the species to be analyzed. The spiked samples are then
subjected to the separation of the species and the measurement of the altered isotope ratios in each
species. Both species concentrations and species conversions can be mathematically deconvolved.
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3.4 Isotopic abundance: The relative number of atoms of a particular isotope in a mixture
of the isotopes of an element, expressed as a fraction of all the atoms of the element.
3.5 isotopes: Nuclides having the same atomic number but different mass numbers.
3.6 Species: Chemical forms in which an element exists.
3.7 Natural isotopic abundance: Isotopic abundance of elements from natural sources.
Most elements (except lithium, lead and uranium) found in nature have a constant isotope
abundance.
3.8 Isotope ratio: Ratio of the isotopic abundances of two isotopes.
3.9 Speciation (or speciated) analysis: Quantification of elements in specific chemical
forms.
3.10 Isotope-enriched material: Material containing elements artificially enriched in minor
isotopes.
3.11 Isotopic spike (Isotope-enriched spike): Standards prepared from isotope-enriched
materials.
3.12 Dead time: The interval during which the detector and its associated counting
electronics are unable to resolve successive pulses. The measured counts are lower than the true
counts ff no correction is performed.
3.13 Gain loss: The loss of gain in detector caused by the inability of the multiplier's dynode
string to supply enough current to maintain constant dynode voltage drops. The measured counts
are lower than the true counts, and cannot be mathematically corrected if gain loss occurs.
3.14 Mass Bias: The deviation of the measured isotope ratio from the true value caused by
the differential sensitivity of the instrument to mass. This effect may occur in the ionization process
or from differential transmission/detection by the mass spectrometer.
3.15 Mass bias factor A number used to correct the mass bias of the measured isotope
ratios. Mass bias factor is measured by employing an isotopically certified standard.
3.16 isotopic-abundance-certified standard (Isotopically certified standard): Standard
material with certified isotopic abundance.
3.17 Inverse isotope dilution: Analysis method to determine the concentrations of isotopic
spikes. A known quantity and isotopic abundance of an isotopic spike is mixed with a known amount
and isotopic abundance (usually tabulated natural isotopic abundance or certified isotopic
abundance) of standard(s), and the altered isotope ratio(s) is(are) measured and used in the
calculation to find the concentration of the isotopic spike. Usually, a natural material is used to
calibrate and determine the concentration of the separated isotopic spike solution using this method.
Only in the case of such elements as uranium, lead, and lithium are the natural isotopic abundances
not constant in terrestrial materials.
3.18 Single spiking: Addition of one isotopic spike to the sample.
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3.19 Double spiking: Addition of two isotopic spikes to the sample. The two isotopic spikes
are enriched in different isotopes, and are prepared in different chemical forms, .each of which
corresponds to a species form.
3.20 Unidirectional conversion: One directional transformation occurring between two
species. One species can convert to the other; the reverse transformation does not occur.
3.21 Interconversion: Bi-directional transformation occurring between two species. Species
convert back and forth between the two chemical forms.
3.22 Time resolved analysis (TRA): A data collection mode in which the data can be
acquired at specified intervals for a continuously aspirated sample, over a user-defined period of
time.
4.0 INTERFERENCES
4.1 Sample preparation
4.1.1 Because this method requires the equilibration of the spike isotope(s) and the
natural isotopes, the sample must be digested, dissolved or extracted into a solution. If the
analyte of interest does not completely dissolve, if the spike or sample isotopes are selectively
lost before equilibration, or if contamination occurs in the sample preparation process, the
measured isotope ratio will not reflect the accurate ratio of added spike atoms to sample atoms
for that element or species (Reference 1).
4.1.2 In general, SIDMS incorporates the assumption that all the converted species
can be found in other species that are monitored. As an example, in the interconversion
between Cr(lll) and Cr(VI), the lost Cr in one species must be found in the other species.
Thus, efforts should be made to keep all species in solution.
4.1.3 Preservation of the species is required in SIDMS since the interconversion
degrades the precision of the determination. The complete conversion of the species will
disable the deconvolution of the species concentration. Thus, digestion methods used for total
metals are inappropriate for SIDMS. However, the altered isotope ratios will indicate the
conversion that has occurred and will not lead to an incorrect answer, but to a situation where
the concentration cannot be determined. Approaches that have been developed to maintain
the species are applicable to SIDMS.
4.2 Isotope ratio measurement
4.2.1 Discussions about isobaric interference, doubly-charged ion interference, and
memory interference in Method 6020 are applicable to this method. The discussion about the
physical interference, suggesting the addition of an internal standard, does not apply. The
internal standard is unnecessary because the isotope ratio measurement is free from physical
interferences. (General considerations for isotope ratio measurement can be found in the
document of Section 13.3.1).
4.2.2 Dead time measurement must be performed daily. At high count rates, two
effects cause pulse counting systems to count less events than actually occur (Section 13.3.2).
The first is dead time (i), the interval during which the detector and its associated counting
electronics are unable to resolve successive pulses. If the true rate, n, is much less than 1/r,
then:
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m » «(1-nr)
where m is the observed rate. The second effect is the loss of gain at high rates caused by the
inability of the multiplier's dynode string to supply enough current to maintain constant dynode
voltage drops. This effect is indicated by a sharp increase in apparent dead time at high count rates.
Both effects cause the measured isotope ratios to diverge from the true isotope ratios with
increasing count rate. While the dead time can be mathematically corrected, the gain loss cannot.
A series of solutions with different concentrations can be prepared from isotopically certified
standards for the determination of dead time. The concentrations may not be accurate, but the
concentrations should spread out evenly, covering the blank to the highest count rate that may be
used in measurements. The isotope pairs that are monitored should have large differences between
their isotopic abundances, since the major isotopes suffer dead time effects much more seriously
than minor isotopes; this makes the dead time correction significant. The sum of the dead-time-
corrected counts is used for calculating the isotope ratios after background subtraction.
Isotopel Isotope 1_
^samplefctandard *~ ^background
tsotope2Q lsotope2«
"sample/standard ~ ^background
• R„, is the dead-time-corrected isotope ratio;
• and
sample/standard an(* are the integrated dead-time-corrected-
counts for the sample or standard of Isotopel and lsotope2, respectively;
• isotopeland teotope2Sbac(qjKJltf1d are the integrated dead-time-corrected-counts for the
background of Isotopel and Isotopel, respectively.
As shown in Figure 1, which displays the xCrPCr ratios for SRM 979 (Cr(N03)3«9H20) as a
function of the count rate, the isotope ratios are highly dependent on the number used for dead time
correction. When the dead time is set to 43.5 ns, the isotope ratios are approximately constant up
to the count rate of 5.8x10s. At higher count rates, gain loss will occur and cannot be mathematically
corrected. Therefore, the solutions must be diluted in the case where the count rate is higher than
this value.
NOTE: Dead time correction is performed before mass bias correction, so the dead-time-
corrected isotope ratios may be different from the certified isotope ratios. Although it is
unnecessary to use isotopically certified material for the determination of dead time, the
certified material is still required for the measurement of mass bias factors. Thus, it is
convenient to use the same certified materia! for both dead time and mass bias factor
measurement.
NOTE: It has been observed that using different isotope pairs for dead time measurement may
obtain different dead times. Thus, it is required to do the dead time measurement for each
isotope pair that will be used. The dead time must be determined daily.
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4.2.3 Instrumental discrimination/fraction effects are changes induced in the true"
isotope ratios from the ionization process or from differential transmission/detection by the
mass spectrometer. This effect can bias the ratios either positively or negatively. To correct
the mass bias, mass bias factors should be determined with isotopically certified materials.
mass bias factor = R, /Rm
where:
• R, and R„, are the certified isotope ratio and the measured dead-time-corrected-isotope-
ratios of the standard material.
The dead-time-corrected isotope ratios of the samples can be corrected using:
Rs = mass bias factor x Rn,
where:
• Rc and Rm are the corrected isotope ratio and the measured dead-time-eorrected-
isotope-ratios of the sample, respectively.
Mass discrimination is a time-dependent instrumental effect, so the mass bias factors must be
determined periodically during the measurement of the samples. Samples are run with the
assumption that mass bias factors remain constant. In general, the mass bias factors are stable
over several hours for ICP-MS measurements.
NOTE: Some previous work observed the following relationship between the measured and
the true isotope ratios for ICP-MS: Rm=R, (1+an), where a is the bias per mass unit, n is the
mass difference between isotopes. This enables the calculation of the mass bias factors of
other isotope pairs based on the measurement of one pair of isotopes. However, this must be
verified experimentally. Otherwise, the mass bias factor for each isotope pair must be
determined.
5.0 SAFETY
5.1 Refer to Chapter Three for a discussion on safety related references and issues.
5.2 Many chromium compounds are highly toxic if swallowed, inhaled, or absorbed through
the skin. Extreme care must be exercised in the handling of hexavalent chromium reagents.
Hexavalent chromium reagents should only be handled by analysts who are knowledgeable of their
risks and of safe handling procedures.
6.0 EQUIPMENT AND SUPPLIES
6.1 Inductively coupled plasma-mass spectrometer (ICP-MS) or other mass spectrometer
systems capable of base line (at least 1 amu) resolution are required. The data system should allow
for corrections of isobaric interferences, dead time and mass bias, or the raw data may be exported
to a computer for further processing. For quadrupole mass spectrometers, the dwell time should be
adjustable since proper settings of dwell time can significantly improve the precision of the isotope
ratio measurement. Both scan mode and peak jump mode can be used, depending on the
instrumentation. The use of a mass-flow controller for the nebulizer angon and a peristaltic pump
for the sample solution are recommended. When chromatography is coupled to ICP-MS for on-line
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detection, the ICP-MS data system must be capable of correcting interferences, dead time and mass
bias, and calculating the isotope ratios in time resolved analysis mode (TRA), or the raw data can
be exported for off-line processing. Other mass spectrometers may also be used, providing a
precision of 0.5% or better can be obtained for the isotope ratio measurement,
6.2 Chromatography or other separation methods are used to isolate species prior to
isotope ratio measurement. Chromatography, such as ion exchange chromatography, may be used
to separate the species on-line in SIDMS (Figure 3). Chromatography components should be
chemically inert based on the specific reagents and analytes. The eluent components and the flow
rate of the chromatography system must be compatible with ICP-MS. An interface between the
chromatography and ICP-MS may be required for compatibility reasons. Alternatively, any
appropriate separation methods, including extraction, chelation, and precipitation, can be used after
validation.
7.0 REAGENTS AND STANDARDS
7.1 All reagents should be of appropriate purity to minimize the blank levels due to -
contamination. Whenever possible, acids should be sub-boiling distilled. All references to water in
the method refer to high purity reagent water. Other reagent grades may be used if it is first
ascertained that the reagent is of sufficient purity to permit its use without lessening the accuracy
of the determination. If the purity of a reagent is questionable, analyze the reagent to determine the
level of impurity.
7.2 For higher precision, solutions may be prepared by weight. For IDMS, standard stock
solutions with natural isotopic abundance may be purchased or prepared from ultra-high purity grade
chemicals or metals. See Method 6010 for instruction on preparing standard solutions from solids.
Generally, the same procedures are applicable to isotope-enriched materials. However, when a
limited amount of the isotope-enriched material is used (usually due to cost considerations) to
prepare the stock solutions, the solutions require calibration with inverse isotope dilution (Section
7.4.1). Isotope-enriched materials with known enrichment can be purchased from several suppliers,
such as the Oak Ridge National Laboratory Electromagnetic isotope Enrichment Facility (ORNL-
EMIEF).
7.3 Currently, few standard stock solutions made for speciation analysis are commercially
available. Thus, in addition to the dissolution of the standard solid, the chemical conversion of the
element into the desired species is usually required for SIDMS. The preparation of Cr(VI) and Cr(lll)
stock standards for SIDMS will be illustrated as an example. For other elements and species,
procedures must be specifically developed.
7.3.1 There are five standards to be prepared for the simultaneous analysis of
Cr(Vl) and Cr(lll), including ratCr(VI) and rkatCr(lll) with natural abundance, ^^(Vl) enriched in
^Cr, ^Crflll) enriched in and isotopio-abundance-certified Cr standard solution.
7.3.2 1 mg/mL Cr(VI) and Cr(lll) standards are commercially available. ^CrO/l) and
natCr(lll) can also be prepared from f^Cr207 and Cr metal, respectively.
7.3.2.1 "trtVI) standard solution, stock, 1 g = 1 mg Cr: Dissolve 0.2829
grams of K2Cr207 in about 80 mL of reagent water and dilute to 100 g with reagent
water.
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7,3.2.2 "^Crflll) standard solution, stock, 1 g = 1 mg Cr Dissolve 0.1 g
Cr metal in a minimum amount of 6M HQ and dilute the solution with 1% HNOa to 100
grams.
7.3.3 53Cr
-------
SIDMS measurement is used to calibrate the isotope-enriched spike and to determine the species
distribution. The characterization of ^CrfVI) spike solution will be illustrated as an example.
7.4.1 Calibration of total concentration of spfce solution with natural material: Weigh
the proper amount (W*) of 10 jifl/fl {C^nda*) ""Cr standard and the proper amount (Ws) of the
^CrCVI) spike (nominal concentration is 10 pg/g ) into a polymeric container, and dilute the
mixture with 1% HN03 to a concentration suitable for isotope ratio measurement. Use direct
aspiration mode to determine the isotope ratio of ^CrPCr (Ra^a). The concentration of the
spike, 0^, can be calculated using the following equations.
'Spike
CSMS
cxwx
w*
53
i* _ p S2*
"X "S3/52 "X
^ *53*2 MS *S
CX = ^standard
where, Cs and C* are the concentrations of the isotope-enriched spike and the standard with
natural isotopic abundance in mmole/g, respectively. Ms and Mx are the average atomic
weights of the spike and the standard in g/mol, respectively. mAs and SAX are the atomic
fractions of sfcr for the spike and standard, respectively. ^Ag and 52Ax are the atomic fractions
of sCr for the spike and standard, respectively.
NOTE: The same procedure is applicable to the calibration of the isotope-enriched spike
solutions in IDMS. The same procedure is also applicable to the calibration of "Crflll) by
changing isotope MCr to ^Cr.
NOTE: Average atomic weight = I(atomic weight of the isotope x atomic fraction)
7.4.2 Calibration of the concentration of the Cr(Vl) in the ^CrfVI) spike with natCr(VI):
Weigh the proper amount (Wx) of 10 pg/g (C^aniaril)nalCr(V!) standard and the proper amount
(Wg) of the ^CrCVl) spike (nominal concentration is 10 pg/g) into a polymeric container, and
dilute the mixture with reagent water to a concentration suitable for measurement. Acidify the
solution to pH 1.7-2.0 with concentrated HN03. Separate the Cr(VI) with chromatography or
other separation methods and measure the isotope ratio of ^Cr/^Cr in Cr(VI) species (R^).
The concentration of Cr(Vi) in the spike, can be calculated using the following equations.
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<&. = Cs
rv> C?WX
Cs ""W
53A _ RVI 52* N
"X *5352 "X
D ^ ® A _ »
*53/52 MS *8 J
c? = C™ /Mx
where, <% and Gx are the concentrations of Cr(VI) in the isotope-enriched spike and standard
with natural isotopic abundance in [jmole/g, respectively. Msand Mx are the average atomic
weights of the spike and the standard in g/mol, respectively, "As and are the atomic
fractions of "Cr for the spike and standard, respectively. SAS and KAX are the atomic fractions
0f s2crfor the spike and standard, respectively.
NOTE: This set of equations is similar to those used in the determination of total Cr in
®Cr(Vl) standard (Section 7.4.1). The general equations for inverse SIDMS are not so
simple. However, for speo'ation of Cr(VI) and Cr(lll) in standard solutions, because the
matrix is so simplified, only the reduction of Cr(VI) to Cr(lll) is observed at low pH.
Thus, the existence of Cr(ll!) species will not influence the isotope ratio of Cr(VI), and
the complex equations can be simplified to the equations shown above (Reference 3).
7.4.3 The distribution of Cr(lll) and Cr(VI) in HCr(VI) spike can be calculated as:
percentage of Cr(VI) =
* VI
"'Spike
-'Spike
* 100%
percentage of Cr(lll)
1 -
% VI
"'Space
"'Spike
x 100%
NOTE: No determination of the species distribution in ^Citlll) spike is required
because only Cr(lll) is present in this solution.
7.5 Blanks: Three types of blanks are required for the analysis: background blank for
subtracting background in isotope ratio measurement, preparation blank for monitoring possible
contamination resulting from the sample preparation procedures, and rinse blank for flushing the
system between all samples and standards.
7.5.1 The background blank consists of the same concentration(s) of the acid(s)
used to prepare the final dilution of the sample solution (often 1% HNOa (v/v) in reagent water).
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7.5.2 The preparation (or reagent) blank must be carried through the complete
preparation procedure and contain the same volumes of reagents as the sample solutions.
7.5.3 The rinse blank consists of 1 to 2 % HN03 (v/v) in reagent water. Prepare a
sufficient quantity to flush the system between standards and samples. Refer to Method 6020
for interference check solution.
7.6 Refer to Method 6020 for preparing mass spectrometer tuning solution.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 All samples must have been collected using a sampling plan that addresses the
considerations discussed in Chapter Nine of this manual.
8.2 Due to the possible degradation or interconversion of the species, samples collected
for spedation analysis must be isotopically spiked as soon as possible. The measurement, however,
can be carried out later provided that less than 80% degradation or interconversion occurs. The
holding time prior to measurement depends on the preservation of the species.
8.3 Proper methods to retard the chemical activity of the species are applicable to SIDMS.
8.4 All sample containers must be prewashed with detergents, acids, and water. Polymeric
containers should be used. See Chapter Three of this manual for further information on clean
chemistry procedures to reduce blank effects in these measurements.
9.0 QUALITY CONTROL
9.1 All qualify control data must be available for reference or inspection. This method is
restricted to use by, or under supervision of, experienced analysts. Refer to the appropriate section
of Chapter One for additional quality control guidance.
9.2 Duplicate samples should be processed on a routine basis. A duplicate sample is a
sample processed through the entire sample preparation and analytical procedure. A duplicate
sample should be processed with each analytical batch or every 20 samples, whichever is the
greater number. A duplicate sample should be prepared for each matrix type (i.e., soil, sludge, etc.).
9.3 Spiked samples and/or standard reference materials should be included with each
group of samples processed or every 20 samples, whichever is the greater number. A spiked
sample should also be included whenever a new sample matrix is being analyzed. For SIDMS,
because the species may degrade or convert to other species when they are spiked into samples,
low recovery may be observed. Thus, the low recovery does not immediately invalidate this method.
For example, if Cr(III) is spiked into a basic solution, due to the hydrolysis of Cr(lll) and the limited
solubility of chromium hydroxide, low recovery of Cr(lll) will be obtained. Low recovery may indicate
an unfavorable matrix for preserving the corresponding species (Reference 4).
9.4 Blank samples should be prepared using the same reagents and quantities used in
sample preparation, placed in vessels of the same type, and processed with the samples.
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10,0 CALIBRATION AND STANDARDIZATION
10.1 IDMS calibration:
10.1.1 Follow the appropriate sections in Method 6020 to set up and tune the ICP-
MS. The determination is performed in direct aspiration mode. The following procedure is
illustrated with the measurement of ^Cr/^Cr and ^Cr/^Cr isotope ratios.
10.1.2 Determine the dead time (Section 4,2.2). Solutions prepared from reference
material SRM 979 (CrtN03)3*9H20) are used in this determination. A range of solutions of
different concentrations should be prepared, but do not need to be accurately known. Masses
50, 52 and 53, as well as masses which could affect data quality should be monitored. The
raw count rates for each solution are measured and integrated. Assume a dead time and use
the equation described in Section 4.2.2 to correct the integrated counts. The dead-time-
corrected counts are then used for calculating the isotope ratios after background subtraction.
By trial and error, the dead time is determined to bring the isotope ratios obtained from
solutions of different concentrations to a constant (the relative standard deviation of the
isotope ratios reach the minimum). The isotope ratios obtained from high counts may be
excluded as gain loss may occur.
NOTE: The concentration range of the solutions may be adjusted depending on the
sensitivity and dynamic range of the instrument.
NOTE: For direct aspiration mode, the dead time correction can be done either before
or after the integration of the raw data. However, it is simpler to do the dead time
correction after the integration.
10.1.3 Determine the mass bias factor (Section 4.2.3). The mean of isotope ratios
obtained in Section 10.1.2 is used for calculating the mass bias factor. The equation is
provided in Section 4.2.3. The measurement of the mass bias factor must be done periodically
between sample measurements. The interval between these measurements depends on the
mass bias stability of the instrument. The relative difference between two consecutive mass
bias factors should not exceed 1 %.
10.2 SIDMS calibration:
10.2.1 Follow the appropriate sections in Method 6020 to set up and tune the ICP-
MS. Follow Section 10.1.2 to measure the dead time. If the calibration of the isotope-enriched
spikes is required, the mass bias factors for direct aspiration mode and the altered isotope
ratios for the spiked standards are measured at this step. The measured isotope ratios
obtained at this step are used in the calibration of total concentrations.
10.2.2 Determine the mass bias factor (Section 4.2.3). Connect the chromatography
outlet to the nebulizer of the ICP-MS. Stabilize the entire system. Background blank and an
isotopic abundance certified standard are used for the measurement of the mass bias factors
for TRA mode. The raw data at each point are corrected for dead time using the equation
described in Section 4.2.2 and then integrated by summing the data across each peak. The
intervals between two consecutive injections must be long enough for the signal to return to
baseline. The integrated counts are then used to calculate the isotope ratios with the equation
shown in Section 4.2.2. Apply the equation in Section 4.2.3 to the calculation of the mass bias
factors for each isotope pair by comparing the measured isotope ratios to the certified isotope
ratios.
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NOTE: For the TRA mode, the dead time correction must be done at each data point
before the data integration.
11.0 PROCEDURE
11.1 IDMS
11.1.1 Closed-vessel microwave digestion is used as an example method to
decompose, solubilize and stabilize the elements of interest. The following procedure is
applicable to samples specified in Method 3052. Refer to Method 3052 for specification of the
microwave apparatus.
11.1.2 Prepare the isotope-enriched spike and calibrate it with the inverse Isotope
dilution mass spectrometry procedure described in Sections 7.2 and 7.4.1.
11.1.3 Weigh a representative sample to the nearest 0.001 g into an appropriate
microwave digestion vessel equipped with a pressure relief mechanism. Spike the sample with
the calibrated isotope-enriched spike. The concentration of the spike should be high enough
so that only a small volume of the solution is used. At least three significant figures should be
maintained for the mass of the spike.
11.1.4 Digest the sample according to the procedure described in Method 3052.
NOTE: For filtered and acidified aqueous samples, digestion may not be required.
Sample solutions can be directly analyzed with ICP-MS after spiking and equilibration.
11.1.5 Measurement of the isotope ratios can be carried out using ICP-MS or other
appropriate mass spectrometers.
11.1.5.1 Determine the mass bias factor periodically as described in
Section 10.1.3.
11.1.5.2 Measure the isotope ratio of each sample. Flush the system with
the rinse blank. The ideal isotope ratio is 1:1. Isotope ratios must be within the range
from 0.1:1 to 10:1, except for blanks and samples with extremely low concentrations.
Samples must be diluted if too high a count rate is observed to avoid gain loss of the
detector.
NOTE: For elements such as lithium, lead, and uranium, the unspiked
solution is used to measure the isotopic abundance of all the isotopes
because the isotopic abundances of these elements are not invariant in
nature.
11.2 SIDMS:
11.2.1 SIDMS is currently applicable to the quantification of elemental species in
various aqueous solutions. Solid samples require isolation and separation to solubilize the
elemental species before spiking. Procedures for such extraction of the species from different
matrices must be specifically designed. The following procedure is an illustration of the
simultaneous determination of Cr(lll) and Cr(VI) in water samples or soil or sediment extracts.
Solids are extracted for Cr(VI) using Method 3060.
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11.2.2 Prepare the isotope-enriched spikes in species forms arid calibrate them with
inverse isotope dilution mass spectrometry described in Section 7.4.
11.2.3 Extract the species from the samples such as soils and sludges. Proposed
Method 3060 can be used to extract Cr(VI) from soils.
NOTE: For aqueous samples, extraction may not be required.
11.2.4 Weigh a proper amount of water sample or extract to the nearest 0.0001 g
into a polymeric container. Spike the sample with 10 yg/g ^CrO/l) spike to a concentration so
that the isotope ratio of^PCr in Cr(Vt) will be approximately 1:1. Thoroughly mix the spike
and the sample. The isotope ratios ^CrPCr for samples must be within the range of 0.1:1 to
10:1, except for blanks or samples with extremely low concentrations.
11.2.5 Dilute the 53Cr(VI)-spiked sample with reagent water. If the solution is strongly
basic, neutralize the sample with concentrated HNO3 to avoid the hydrolysis of Cr(lll). Spike
the diluted sample with 10 pg/g ^CrP) spike to a concentration so that the isotope ratios of
^Cr/^Cr in Cr(lll) will also be approximately 1:1 and the species concentrations are suitable
for measurement. The measured isotope ratios ^Crf^Cr for samples must be within the range
of 0.1:1 to 10:1 except for blanks and samples with extremely low concentrations, or the
sample should be respiked and analyzed. (Sections 11.2.4 and 11.2.5 should be completed
as quickly as possible.)
NOTE: If only the Cr(VI) is of interest, the sample can be single spiked with ^rfVI)
instead of double-spiking with both ^Ci-flll) and ^CrO/l). However, this is based on the
assumption that only unidirectional conversion, the reduction Cr(VI) to Cr(lil), can occur
after spiking. This is usually true if the sample is acidified to low pH after spiking,
especially for matrices containing reducing agents.
11.2.6 Acidify the spiked samples to pH 1.7 to 2.0; under these conditions Cr is
usually retained in the solutions, (although there might be interconversion between Cr(lll) and
cm))- The spiked samples can be stored at 4°C to retard the interconversion of the species.
Other methods that can retard the transformation of the species are applicable as long as no
interference with the isotope ratio measurement is introduced. For example, some soil extracts
contain large concentrations of reducing agents that reduce Cr(VI) rapidly after acidification.
To slow down the reduction, stoichiometric amounts of KMn04 can be added to the sample to
compete with Cr(VI) in the oxidation of reducing matrices.
NOTE: Studies have shown that the lower the interconversion, the more precise the
determination (Reference 3). Thus, efforts should be made to prevent interconversion
between the species.
11.2.7 The measurement of the isotope ratios in each species can be carried out
using ICP-MS or other equivalent mass spectrometers following the separation of the species
using chromatography or other separation methods. An ion-exchange chromatograph coupled
with ICP-MS will be illustrated as an example in the measurement of ^Cr/^Cr and "Cr^Cr
isotope ratios in both Cr(lll) and Cr(VI) species in samples.
11.2.7.1 Determine the mass bias factors periodically as described in
Section 10.2.2.
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11,2.7.2 Measure the isotope ratios of each sample. Flush the system with
the eluent until the signal returns to the baseline. The ideal isotope ratios for ^Cr/^Cr
in Cr(Ili) and ^Cr/^Cr in Cr(VI) are 1:1. Ratios between 0.1:1 to 10:1 are also
acceptable. Samples may be respiked to achieve an isotope ratio close to 1:1.
Samples must be diluted if excessively high count rates are observed to avoid gain loss
of the detector.
NOTE: For elements such as lithium, lead, and uranium, the unspiked
solution is used to measure the isotopic abundance of all the isotopes
because the isotopic abundances are not invariant in nature.
12.0 DATA ANALYSIS AND CALCULATIONS
12.1 IDMS-Calcutations: The quantitative values shall be reported in appropriate units, such
as micrograms per liter (pg/L) for aqueous samples and milligrams per kilogram (mg/kg) for solid
samples. If dilutions are performed, the appropriate corrections must be applied to the sample
values.
12.1.1 Calculate the isotope ratios. Calculations should include appropriate
interference corrections (see Section 4.2 for data integration, dead time correction, and mass
bias correction).
12.1.2 The following equations are applied to the calculation of the concentration of
the element, Cga^^g/g), in the final sample solutions.
^Sarr**» =
= ^Spike /Ms
r _ csws
x WY
' % - wv
p _ S3.
^ 53/52 MX MX
where, Cs and Cx are the concentrations of the isotope-enriched spike and the sample in
mmole/g, respectively. Ms and Mx are the average atomic weights of the isotope-enriched
spike and the sample hi g/mole, respectively. MAS and "Ax are the atomic fractions of ^Cr for
the isotope-enriched spike and sample, respectively. 52AS and 52AX are the atomic fractions of
^Cr for the isotope-enriched spike and sample, respectively. Cspjke is the concentration of the
isotope-enriched spike in (xg/g.
NOTE: When isotope MCr is used, substitute 53 with 50 in the above equations.
12.1.3 If appropriate or required, calculate results for solids on a dry-weight basis as
follows:
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(1) A separate determination of percent solids must be performed.
(2) The concentrations determined in the digest are to be reported on the basis of the dry
weight of the sample.
r
Concentration (dry weight)(mg/kg) = ^sample
S
where, Cga^ = Concentration based on the wet sample (pg/g)
S - % Solids
100
12.2 SIDMS-Calculations: The quantitative values shall be reported in appropriate units,
such as micrograms per liter (pg/L) for aqueous samples and milligrams per kilogram (mg/kg) for
solid samples. If dilutions are performed, the appropriate corrections must be applied to the sample
values.
12.2.1 Calculate the isotope ratios. Calculations should include appropriate
interference corrections, dead time correction, and mass bias correction (Section 4.2).
12.2.2 The following equations are used to deconvolute the concentrations of the
species at the time of spiking, as well as the conversion of the species after spiking.
. (»AA*W.*aA?C.*vO0-°) * (^C^^CX1) P
" (HA,tfx*s2AKw;)(i-a) ~
M52 " (^t?Wx^A:c",W;)(1-o) - ("A^X^WWflS
Rv, a + pA^X^AycXHl-P)
"® " CV!wa * (HAxcX+K^?<)(1-|i)
„ a * (^cX^CXIa-B
« ~ (%CX^CX1 (1 -P)
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where,
R'sq/52 is the measured isotope ratio of ®Cr to ^Cr of Cr(lll) in the spiked sample
^Ax is the atomic fraction of ^Cr in the sample (usually a constant in nature)
c£' is the concentration of Cr(lll) in the sample (pmole/g, unknown)
Wx is the weight of the sample (g)
is the atomic fraction of ^Cr in the "Crflll) spike
Cg1 is the concentration of Cr(lll) in the "Crflll) spike (pmole/g)
Wg is the weight of the ^Crflll) spike (g)
C™ is the concentration of Cr(VI) in the sample (pmole/g, unknown)
a is the percentage of Cr(lll) oxidized to Cr(VI) after spiking (unknown)
p is the percentage of Cr(VI) reduced to Cr(lll) after spiking (unknown)
NOTE' Hie unit of the concentrations shown above is ^jmole/g. The conversion factor
from ymole/g to fjg^g is: M, where M is the average atomic weight of the element in
g/mole (Section 7.4.1). The following equation can be used to convert the unit of the
concentration. Be aware that samples with different isotopic abundance have different
average atomic weights.
Concentration (pmole/g)x M = Concentration (yg/g)
NOTE: Although the species distribution of the isotopic spike is determined (Section
7.4), the above equations assume that each isotope-enriched spike is only in one
species form to simplify the equations. This has been validated for Cr(lll) and MCr(\/l)
spikes prepared using the procedures described in Section 7.3. For other speciation
analysis, this assumption must be verified experimentally, or the distribution of the
species in the isotope-enriched spikes must be taken into account.
NOTE: For the quantification of the single-spiked samples, the following equations are
used:
^Sample " ^XMX
Cs -
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-M
cX
w.
53* V! pVt 52 .VI
"S "SSSZ AS
RVI 52* _ 53A
^ "53/52 nX
where, G| and are the concentrations of the isotope-enriched spike and the sample
in pmolefg, respectively. Ms and Mx are the average atomic weight of the isotope-
enriched spike and the sample in g/mole, respectively. ^Ag and ®AX are the atomic
fraction of^Cr for the isotope-enriched spike and sample, respectively. 52Aj. and82Ax
are the atomic fractions of ®Crfor the isotope-enriched spike and sample, respectively.
Ctpfly is the concentration of the isotope-enriched spike in pg/g.
NOTE: When isotope ^Cr is used, substitute 53 with 50 in the above equations.
12.2.3 A computer program such as a spreadsheet can be developed to solve this
set of second power, four variable equations. Solutions of the values for, C^, Cx, a and p are
required. The following mathematics is a way to solve the equations iteratively. To assist the
analyst a spreadsheet file with these preprogrammed equations has been placed on the
internet (Reference 10). Additional discussion and alternate equations are also available.
To make the expression simpler, assume
C™WX = n£ cX - Hi cX « N'l, Cs Ws = Ns
At the beginning of the iteration, arbitrary values can be assigned to and a. For example,
both of them are assigned as 0s. Now we need to know the expression of Nx and p. After careful
derivation, we can get the following equations:
o-«K«'V,"axK .[tfU (»axn; ~S2aXi) - (%>« ."AXl 0 • "ASKf1-")
(i-a^!V"A>K * KUSV« •"«) - ("M! .eAXl 0 = X)NS1-"|)
These equations can be rewritten as;
A^x + Bt{3 = Ct
+ = ^2
The solutions are

A,
0^
p2

and
P =
K
C2

A,
B,

**
B1
*2
B,

K
b2
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Use these two values in the following equations to solve N^1 and a
.*«) - ("v; -"'WJ <¦ - (-"i."*; •>
d-e)KU,Vs\K * WV! *XK) - pAX .1«|. - (-rSUX >
Repeating the calculation, the variables , N^1, a and P will converge to constant values, and
these values are the solution of the equations.
12.2.4 Results should be discarded when a + p > 80% because the interconversion
will be too extensive and cause inaccuracy and imprecision in the corrections. Samples should
be respiked with the isotope-enriched spikes and analyzed, and the preservation should be
improved to retard the conversion of the species.
12.2.5 If appropriate or required, calculate results for solids on a dry-weight basis as
described in Section 12.1.3.
13.0 METHOD PERFORMANCE
13.1 Performance and use of lDMS as a definitive method in standard reference material
certification has been well established in practice and in the literature. Review and discussion
articles are referenced for performance criteria of this highly accurate method (References 1, 8, 9).
13.2 Accuracy, precision, and capability of SIDMS in correcting species interconversion are
shown in Table 1. Table 2 and Table 3 compare data against Method 7196 analysis for Cr(VI) in
chromium ore process residues and soil extracts. Table 1 demonstrates the ability of Method 6800
to correct for transformations of both Cr(VI) and Cr(lll) in aqueous samples and also the magnitude
of errors that may be expected when using other methods that are unable to determine the
conversion of these species. Table 2 indicates a sample type where both the traditional 3060/7196
methods and 3060/6800 methods produced statistically similar data indicating confirmation between
these two analytical methods. Table 3 demonstrates the correction necessary in some soil samples
Rewrite the equation as:
A3N? + B3a - C3
A4Nx + B4cc - C4
again
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where the sample matrix would cause a bias in more traditional methods. These bias corrections
demonstrate the ability of Method 6800 to identify and correct for the degradation of a species during
the measurement process.
13.3 The following documents may provide additional guidance and insight on this method
and technique:
13.3.1 Javis, K. E.; Gray, A. L; Houk, R. S. Handbook of Inductively Coupled
Plasma Mass Spectrometry, Blackie: London, 1992.
13.3.2 Russ, G. P., Ill; Bazan, J. M. Spectrochim. Acta, Part B 1987,42B, 49- 62.
14.0 Pollution Prevention
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quality
and/or toxicity of waste at the point of generation. Numerous opportunities for pollution prevention
exist in laboratory operation. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of the first
choice. Whenever feasible, laboratory personnel should use pollution prevention techniques to
address their waste generation. When wastes cannot be feasibly reduced at the source, the Agency
recommends recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Less is Better. Laboratory Chemical Management for Waste Reduction
available from the American Chemical Society.
15.0 Waste Management
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with ail solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society.
16.0 References
1. Fasset, J. D.; Paulsen, P. J. Isotope Dilution Mass Spectrometry Anal. Chem. 1989, 61, 643A-
649A.
2. Kingston, H. M. Method of Speciated Isotope Dilution Mass Spectrometry, US Patent Number;
5,414,259, 1995.
3. Kingston, H. M.; Huo, D.; Lu, Y. "Speciated Isotope Dilution Mass Spectrometry (SIDMS): The
Accurate Determination of Reactive Species," (submitted Anal. Chem. 1998).
4. Janes, B. R.; Petura, J. C.; Vitale, R. J.; Mussoline, G. R. Environ. Sci. Tech. 1995,29,2377.
6800-21
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5. Kingston, H. M.; Huo, D.; Lu, Y.; Chalk, S. "Accuracy in Species Analysis. Speciated Isotope
Dilution Mass Spectrometry (SIDMS) Exemplified by Evaluation of Chromium," (Accepted)
Spectrochim Acta. 1998.
6. Kingston, H. M.; Huo, D.; Chalk, S.; Walter, P. The Accurate Determination of Species by
Speciated Isotope Dilution Mass Spectrometry: Exemplified by the Evaluation of Chromium (VI)
in Soil, The Twelfth Annual Waste Testing & Quality Assurance Symposium; Washington, DC,
July 23-36 1996; 112-119.
7. Lu, Y.; Huo, D.; Kingston, H. M. "Determination of Analytical Biases and Chemical Mechanisms
in the Analysis of Cr(VI) Using EPA Protocols, (submitted Environ. Sci. Tech., -1998).
8. Bowers, G. N, Jr.; Fassett, J. D.; White, D. V. Anal. Chem. 1993, 65,475R.
9. Moore, L. J.; Kingston, H. M.; Murphy, T. J. Environ. Intern. 1984,10,169.
10. Kingston, H. M.; Huo, D. (copywright 1998) SamplePrep Web™ [Homepage of SamplePrep
Web™], [Online], Available: http://www.sampieprep.duq.edu/sampleprep/ [1998, January 22].
17.0 Tables, Diagrams, Flowcharts, and Validation Data
The pages to follow contain Tables 1 through 3, Figures 1 through 5, and a method procedure
flow diagram.
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TABLE 1
ANALYSIS OF AN ARTIFICIALLY SYNTHESIZED WATER SAMPLE.
(Reference 6)
Concentration Conversion
(ng/g) (%)

Days after

Cr(lll) to
Cr(VI) to
Aliquot
spiking
CifllD
Cr(VI)
Cr(Vl)
Cr(HI)

1
69.8 ±0.3
66.8 ± 0.3
4.87 ±0.22
3.57 ±0.03
1
4
69.2 ± 0.6
69.4 ±0.3
3.47 ±0.11
11.9 ±0.5

13
70.5±0.9
68.5 ± 0.4
2.80 ±0.13
22.4 ±0.2

1
69.6 ±0.2
68.8 ±0.4
17.6 ±0.1
2.95 ±0.02
2
4
69.3 ±0.7
69.6 ±0.6
14.6 ±1.3 *
11.4 ±0.7

13
70.7 ±0.4
68.8 ±0.3
12.8 ±0.1
22.1 ±0.3

1
69.8 ±0.6
69.0 ±0.2
23.8 ±0.3
2.76 ±0.08
3
4
69.0 ± 0.8
69.6 ± 0.3
21.6 ±0.2
102 ±0.1

13
70.4 ±0.5
68.9 ±0.8
17.6 ±0.3
22.1 ±0.1
True

69.67
68.63

mean ± 95% confidence interval
Aliquots 1,2 and 3 were from the same isotopically-spiked synthesized sample. These aliquots were treated
in different ways to induce different degrees of interconversion between Cr(lll) and Cr{Vl), Measurement were
done on different days to check the stability of the species during storage. Despite the different degrees of
interconversion, the deconvolved concentrations for both Crflll) and Cr(VI) were always corrected successfully
within experimental error to the true concentrations.
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TABLE 2
CONCENTRATIONS OF CR(VI) IN COPR SAMPLES DETERMINED WITH METHOD 7196 AND SIDMS
(Reference 7)

Method 7196
SIDMS

sample
Cone, of Cr(Vl)
Average
Cone, of Cr(Vl)
Average

(Mg/g)
(mean ±std)
(pg/g)
(mean ±std)
COPR1
1330
1410 ±85
1373
1445 ±7Q

1410

1449

1500

1512

COPR3
91.2
85.3 ±5.2
93.9
88.8 ±6.1

81.5

82.1

83.1

90.4

COPR4
408.9
407.8 ±7.2
419.8
418.0 ±9.2

414.4

426.1

400.2

408.0

COPR: chromite ore processing residue.
Method 3060 was used for Cr(VI) extraction.
Results obtained from SIDMS and Method 7196 are comparable for COPR samples.
TABLE 3
RECOVERY OF CR(VI) SPIKED INTO SOIL EXTRACTS
(Reference 7)
Sample
Mass of Soil (g)
Spiked "^(Vl)
Recovery (%)

(Mg/g)
Method 7196
SIDMS
1
0
2.997
101 ±0.4
100 ±1.3
2
1.53
3.033
91.8 ±1.7
100 ±0.3
3
3.06
2.993
81.9 ± 1.1
101 ±0.3
4
3.12
1.587
71.6 ±2.5
99.3 ±0.3
Results obtained from SIDMS and Method 7196 are incomparable for soil extracts due to the serious matrix
effects resulting from the coexisting reducing agents in soil. Method 7196 is incapable of correcting conversion
of Cr(VI) leading to low recoveries. Results are based on N = 3 with uncertainties expressed in standard
deviation.
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FIGURE 1
THE INFLUENCE OF THE DEAD TIME CORRECTION ON THE ISOTOPE RATIOS MEASURED WITH
ICP-MS EQUIPPED WITH A CONTINUOUS DYNODE MULTIPLIER
Gain loss occurs when the count rate exceeds 5.8*1 G5.
0,054 -I dead time (ns)
0.053
0.052
O
IN
o
B
o 0.051
n
a 0.050
o
o
m
~ 0.049
0,048-
OE+0

0 '

20
-±-
43.5

80
Gain loss
2E+5
4E+5
SE+5
8E+5
1E+6
Counts per second torMCr
FIGURE 2
IDMS DETERMINATION OF VANADIUM IN CRUDE OIL. NUMBERS SHOWN ABOVE THE BARS ARE
THE ATOMIC FRACTION
(Revised from Reference 1}
»
Q.
o
0
01
K
Vanadium site
Spited sample
Natural vanadium
R„,_ *0.5647
50/51
Re„„ =0.09483
50/51
R50,S1=°.0025
0.9134
0.9975
0.G391
0.08662
0.3609
0.0025
50 51
50 51
isotope
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FIGURE 3
SEPARATION AND DETECTION OF CR(HI) AND CR(VI) WITH ION-EXCHANGE CHROMATOGRAPHY
COUPLED WITH AN ICP-MS
(Reference 5)
100
Cr(in): ] 00 ppb
Cr(VI): 100 ppb
Flow rate: 1.0 mL/min
Eluent: 0.06M N03", pH = 3
Column: CETAC ANX 4605 Cr
25.00

50.00
75.00
100.00
time (s)
FIGURE 4
SEPARATION OF THE UNSPIKED SAMPLE AND ISOTOPICALLY SPIKED SAMPLE
(Reference 3)
(a): Chromatograms of a solution containing Cr(lll) and Cr(VI) with natural isotope abundance,
(b): Chromatograms of the same solution spiked with isotope-enriched spikes *°Cr(Hl) and ^CifVl).
100%
ma-50
5 60%
S 40%
ntfZ"53
¦MHWrar
20 30 40 SO
70 80 90 100 110 120
100%
80%
60%
30 40 SO 60 70
40%
20%
90 100 110 120
6800-26
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FIGURE 5
GRAPHIC CALCULATED ILLUSTRATION OF THE APPLICATION OF SIDMS TO THE SIMULTANEOUS
DETERMINATION OF CR(III) AND CR(VI)
Cr(III) Cr(VI)
(a| !•<*<*>• msMbutton of Via un>plk«d ikw><

140 .

120 -
1
100 .
Is
80 -

60 -
I
40-
¦
20-

0 -
50
S3
¦ CrjVI)
spike
a cr(vi)
natural
a cm
spike
B Cr(IU>
natural
(c) Isnisfia dUtrlbutlon of 01 • tplk«d swnpit
120 -
100.
I Cr(V7)
spike
BCr{VI)
natiral
0 Cr(i8J
spite
B Cr(B)
natural
(•I Isotap* dlsMbstlon of m* iplktd tampla «Jth 20S
inlwconvwitan
1 Cr(VI)
0 CrtlB)
BCr(Hi)
natural
(b) l««op« dltirlBuilon af mi untpliwd >*mpl»
OCrpn
140-¦
mem
100 -¦
H Cf(VI}
(d) ! siXop* dMiifeutlgn of Bn tpikad tamp)*
~ CrJB)
140 . ¦
120 • ¦
BOOD
too..
HCrfVI)
(fl list opt distribution of tti> *plk*d umpl« with 20%
IMMtonvwalsn
OCrffl
140 ¦
120-
mcm
(a) and
-------
METHOD 6800
ELEMENTAL AND SPECIATED ISOTOPE DILUTION MASS SPECTROMETRY:
ISOTOPE DILUTION MASS SPECTROMETRY flDMS)
11.1,5 Measure the isotope
ratios of the samples. Measure
mass bias factors wrth an
interval of several hours.
Type of
analysis?
11.2 Speciated istotope
Dilution Mass
Spectrometry (SIDMS).
Speciation
Total Element
11.15
Are the isotopic
ratios oatside 0,1 to 10
for samples containing
significant amounts
of analyte?
11.1.2
Are the isotope
enriched standards
ready?
7.4 Prepare isotope
enriched standards
11.1.2 Spike the isotope
enriched standards with
standard solution that
has natural isotopic
abundance
11.1,5.2 Dilute
the samples
11.1.5,2
Is the
response so high
that the gain
depression might
occur?
11.1,3 Spike samples with
the isotope enriched
standards.
11.1.4
Is the sample
acidified and pre-
filtered water?
11.1.4 Digest the
spiked samples
11.1.5
Is the instrument
ready for
measuring isotopie
ratios?
10.1.1 Set-up and tune
instrument according to
Method 6020
10.1,2 Determine the
dead time of the detector.
12.1.1 Integrate the
counts, perform dead
time correction and mass
bias correction.

r
12.1.1 Calculate the
isotope ratios.

t
12.1-2 Ca
concentre
the measi.
ra
leulate the
lions using
red isotope
•OS.
12.1.3 If required,
calculate results for
solids on a dry-
weight basis.
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METHOD 6800
ELEMENTAL AND SPECIATED ISOTOPE DILUTION MASS SPECTROMETRY:
SPECIATED ISOTOPE DILUTION MASS SPECTROMETRY fSIDMSY
11.2 speclati'cn
11.1 Total Element
Type of
analysis?
11.1 isotope Dilution
Mass Spectrometry
(IQMS)
11.2.2
Are the
isotope enriched
standards
Cr(lll) & 53Cf{Vt)
ready?
7.4 Prepare isotope
ernlched standards
S0Cr(lll) & ^CrfVi).
7 A Spike tne Isotope
enrlctied standards with
the standard solutions
that hawe natural
isotopic abundance
11.2.3
Is the
sample an aqueous
solution?
11.2,3 Extract tie
species from the
sample
11,2.6 Dilute and

11.2.4 Spike
neutralize the
-
sample with *aCr(VI).
sample.

*
f

11.2.5 & 11.2,6 Spike the

sample witt- s0Cr;n:) and

acidify the solution to pH 2.

10.2.1 Set-up and tune
instrument according
1c Method 6020.
11.2.7
is the
instrument reedy
for measuring
isotopic ratios?
10.2.2 Measure the
dead time.
10-2.2 Connect the
chromatography to ICP-MS.
11,2.7 Measure ttie isotope
ratios of samples. Measure -
mass bias factors with an interval
of several hours.
11.2,7,2
Are the isotopic
ratios outside 0,1 to 10
for samples containing
significant 8mourns
of anafyte?
11.2.7.2 Dilute
the spiked
samples
1.2.7.2
is the
response so high
that the gain
depression might
occur?
12.2.1 Perform dead time
correction at each point
across peaks.
12.2.integrate me peaks
by summing up the counts
across peaks.
12.2,1 Calculate the isotope
ratios and perform mass
Was correction
12.2.2 Deconvolve tie
ecr.cer»tration of each
species and the
irrterconvensiofl of the
species^
12.2,3 If required,
calculate results tor
dry-weight basis.
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METHOD 7000B
FLAME ATOMIC ABSORPTION SPECTROPHOTOMETRY
1.0 SCOPE AND APPLICATION
1.1 Metals in solution may be readily determined by flame (direct aspiration) atomic
absorption spectrophotometry. The method is simple, rapid, and applicable to a large number of
environmental samples including, but not limited to, ground water, aqueous samples, extracts,
industrial wastes, soils, sludges, sediments, and similar wastes. With the exception of the analyses
for dissolved constituents, all samples require digestion prior to analysis (refer to Chapter Three).
Analysis for dissolved elements does not require digestion if the sample has been filtered and then
acidified.
Note: The analyst should be aware that organo-metallic species may not be detected if the
sample is not digested.
This method is applicable to the following elements:
ELEMENT

CASRN®
Aluminum
(Al)
7429-90-5
Antimony
(Sb)
7440-36-0
Barium
(Ba)
7440-39-3
Beryllium
(Be)
7440-41-7
Cadmium
(Cd)
7440-43-9
Calcium
(Ca)
7440-70-2
Chromium
(Cr)
7440-47-3
Cobalt
(Co)
7440-48-4
Copper
-------
downward with scale expansion and upward by using a less sensitive wavelength or by rotating the
burner head. Detection limits by direct aspiration may also be extended through concentration of the
sample and/or through solvent extraction techniques. Method detection limits (MDLs) must be
established empirically for each matrix type analyzed (refer to Chapter One for guidance) and would
be required for each preparatory/determinative method combination used. These MDLs must be
documented and kept on file and should be updated when a change in operation or instrument
conditions occurs. Refer to Chapter One for guidance.
1.3 Users of this method should state the data quality objectives prior to analysis and must
document and have on file the required initial demonstration performance data described in the
following sections prior to using the method for analysis.
1.4 Where direct-aspiration atomic absorption techniques do not provide adequate
sensitivity, refer to specialized procedures such as graphite furnace atomic absorption (Method
7010) or the gaseous-hydride methods.
1.5 Other elements and matrices may be analyzed by this method as long as the method
performance is demonstrated for these additional elements of interest, in the additional matrices of
interest, at the concentration levels of interest in the same manner as the listed elements and
matrices (see Sec. 9.0).
1.6 Use of this method is restricted to analysts who are knowledgeable in the chemical and
physical interferences as described in this method.
2.0 SUMMARY OF METHOD
2.1 Although methods have been reported for the analysis of solids by atomic absorption
spectrophotometry, the technique generally is limited to metals in solution or dissolved through some
form of sample processing.(refer to Chapter Three). Preliminary treatment of waste water, ground
water, extracts, and industrial waste is always necessary because of the complexity and variability
of sample matrix. Solids, slurries, and suspended material must be subjected to a solubilization
process before analysis. This process may vary because of the metals to be determined and the
nature of the sample being analyzed. Solubilization and digestion procedures are presented in
Chapter Three.
2.2 In direct-aspiration atomic absorption spectrophotometry, a sample is aspirated and
atomized in a flame. A light beam from a hollow cathode [amp or an electrodeless discharge lamp
is directed through the flame into a monochromator, and onto a detector that measures the amount
of absorbed light. Absorption depends upon the presence of free unexcited ground-state atoms in
the flame. Because the wavelength of the light beam is characteristic of only the metal being
determined, the light energy absorbed by the flame is a measure of the concentration of that metal
in the sample. This principle is the basis of atomic absorption spectrophotometry.
3.0 DEFINITIONS
Refer to Chapter One and Chapter Three for a listing of applicable definitions.
4.0 INTERFERENCES
4.1 The most troublesome type of interference in atomic absorption spectrophotometry is
usually termed "chemical" and is caused by lack of absorption of atoms bound in molecular
combination in the flame. This phenomenon can occur when the flame is not sufficiently hot to
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dissociate the molecule, as in the case of phosphate interference with magnesium, or when the
dissociated atom is immediately oxidized to a compound that will not dissociate further at the
temperature of the flame. The addition of lanthanum will overcome phosphate interference in
magnesium, calcium, and barium determinations. Similarly, silica interference in the determination
of manganese can be eliminated by the addition of calcium. A nitrous oxide/acetylene gas mixture
may be used to help prevent interferences from refractory compounds.
4.2 Chemical interferences may also be eliminated by separating the metal from the
interfering material. Although complexing agents are employed primarily to increase the sensitivity
of the analysis, they may also be used to eliminate or reduce interferences.
4.3 The presence of high dissolved solids in the sample may result in an interference from
non-atomic absorbance such as light scattering. In the absence of background correction, this can
result in false positives and/or falsely elevated values. If background correction is not available, a
non-absorbing wavelength should be checked. Signal contribution finom uncorrected background can
not be diagnosed through the analysis of spike recovery, nor is it compensated for by the application
of the method of standard additions (MSA). If background correction is not available and the non-
absorbing wavelength test indicates the presence of background interference, the sample digestates
must be extracted (liquid-liquid or solid phase) prior to analysis, or another analytical method must
be selected.
4.4 Ionization interferences occur when the flame temperature is sufficiently high to
generate the removal of an electron from a neutral atom, giving a positively charged ion. This type
of interference can generally be controlled by the addition, to both standard and sample solutions,
of a large excess (1,000 mg/L) of an easily ionized element such as K, Na, Li or Cs. Each sample
and standard should contain 2 mL KCI/100 mL of solution. Use 95 g of potassium chloride in 1 L
of reagent water for the KCI solution!
4.5 Spectral interference can occur when an absorbing wavelength of an element present
in the sample, but not being determined, falls within the width of the absorption line of the element
of interest. The results of the determination will then be erroneously high, due to the contribution of
the interfering element to the atomic absorption signal. Interference can also occur when resonant
energy from another element in a multielement lamp, or from a metal impurity in the lamp cathode,
falls within the bandpass of the si setting when that other metal is present in the sample. This type
of interference may sometimes be reduced by narrowing the slit width.
4.6 The analyst should be aware that viscosity differences and/or high dissolved or
suspended solids may alter the aspiration rate.
4.7 All metals are not equally stable in the digestate, especially if it only contains nitric acid
and not a combination of acids including hydrochloric acid. The addition of HCI helps stabilize Sn,
Sb, Mo, ia, and Ag in the digestate. The digestate should be analyzed as soon as possible, with
preference given to these analytes. Refer to Chapter Three for the suggested decomposition
methods.
4.8 Specific interference problems related to the individual analytes are located in this
section.
4.8.1 Aluminum: Aluminum may be as much as 15% ionized in a nitrous-
oxide/acetylene flame. Use of an ionization suppressor (1,000 ug/mL K as KCI) as described
in Sec. 4.4 will eliminate this interference.
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4.8.2 Antimony. In the presence of lead (1,000 mg/L), a spectral interference may
occur at the 217.6-nm resonance line. In this case, the 231.1-nm resonance line should be
used. Excess concentrations of copper and nickel (and potentially other elements), as well as
acids, can interfere with antimony analyses. If the sample contains these matrix types, either
matrices of the standards should be matched to those of the sample or the sample should be
analyzed using a nitrous oxide/acetylene flame.
4.8.3 Barium: Barium undergoes significant ionization in the nitrous oxide/acetylene
flame, resulting in a significant decrease in sensitivity. All samples and standards must contain
2 mL of the KG ionization suppressant per 100 mL of solution (refer to Sec. 4.4). In addition,
high hollow cathode current settings and a narrow spectral band pass must be used because
both barium and calcium emit strongly at barium's analytical wavelength.
4.8.4 Beryllium: Concentrations of Al greater than 500 ppm may suppress
beryllium absorbance. The addition of 0.1% fluoride has been found effective in eliminating
this interference. High concentrations of magnesium and silicon cause similar problems and
require the use of the method of standard additions.
4.8.5 Calcium: AH elements forming stable oxyanions will complex calcium and
interfere unless lanthanum is added. Addition of lanthanum to prepared samples rarely
presents a problem because virtually all environmental samples contain sufficient calcium to
require dilution to be within the linear range of the method.
4.8.6 Chromium: An ionization interference may occur if the samples have a
significantly higher alkali metal content than the standards. If this interference is encountered,
an ionization suppressant (KQ) should be added to both samples and standards (refer to Sec.
4.4).
4.8.7 Magnesium: All elements forming stable oxyanions (P, B, SI, Cr, S, V, Ti, Al,
etc.) wifl complex magnesium and interfere unless lanthanum is added. Addition of lanthanum
to prepared samples rarely presents a problem because virtually all environmental samples
contain sufficient magnesium to require dilution.
4.8.8 Molybdenum: Interference in an air/acetylene flame from Ca, Sr, S04, and
Fe are severe. These interferences are greatly reduced in the nitrous oxide flame and by the
addition of 1,000 mg/L aluminum to samples and standards (refer to Sec. 7.7).
4.8.9 Nickel: High concentrations of iron, cobalt, or chromium may interfere,
requiring either matrix matching or use of a nitrous-oxide/acetylene flame. A non-response line
of Ni at 232.14 nm causes non-linear calibration curves at moderate to high nickel
concentrations, requiring sample dilution or use of the 352.4 nm line.
4.8.10 Osmium: Due to the volatility of osmium, standards must be made on a daily
basis, and the applicability of sample preparation techniques must be verified for the sample
matrices of interest.
4.8.11 Potassium: In air/acetylene or other high temperature flames (>2800°C),
potassium can experience partial ionization, which indirectly affects absorption sensitivity. The
presence of other alkali salts in the sample can reduce ionization and thereby enhance
analytical results. The ionization-suppressive effect of sodium is small if the ratio of Na to K
is under 10. Any enhancement due to sodium can be stabilized by adding excess sodium
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(1,000 ug/mL) to both sample and standard solutions. If more stringent control of ionization
is required, the addition of cesium should be considered,
4.8.12 Silver Since silver nitrate solutions are light sensitive and have the tendency
to plate silver out on the container walls, they should be stored in dark-colored bottles. In
addition, it is recommended that the stock standard concentrations be kept below 2 ppm and
the chloride content increased to prevent precipitation. If precipitation is occurring, a 5%:2%
HCI:HN03 stock solution may prevent precipitation. Daily standard preparation may also be
needed to prevent precipitation of silver.
4.8.13 Strontium: Chemical interference caused by silicon, aluminum, and phosphate
are controlled by adding lanthanum chloride. Potassium chloride is added to suppress the
ionization of strontium. All samples and standards should contain 1 mL of lanthanum
chloride/potassium chloride solution per 10 mL of solution (refer to Sec. 7.8}.
4.8.14 Vanadium: High concentrations of aluminum or titanium, or the presence of
Bi, Cr, Fe, acetic add, phosphoric acid, surfactants, detergents, or alkali metals, may interfere.
The interference can be controlled by adding 1,000 mg/L aluminum to samples and standards
(refer to Sec, 7.7).
4.8.15 Zinc: High levels of silicon, copper, or phosphate may interfere. Addition of
strontium (1,500 mg/L) removes the copper and phosphate interference.
5.0 SAFETY
5.1 Refer to the guidance in Chapter Three.
5.2 Concentrated nitric and hydrochloric adds are moderately toxic and extremely irritating
to skin and mucus membranes. Use these reagents in a hood whenever possible and if eye or skin
contact occurs, flush with large volumes of water. Always wear safety glasses or a shield for eye
protection when working with these reagents.
5.3 Many metal salts, including those of osmium, are extremely toxic if inhaled or
swallowed. Extreme care must be taken to ensure that samples and standards are handled property
and that all exhaust gases are properly vented. Wash hands thoroughly after handling.
5.4 Protective eyeware and/or flame shields should be used when conducting analyses by
acetylene-nitrous oxide flame due to the emission of UV light.
6.0 EQUIPMENT AND SUPPLIES
6.1 Atomic absorption spectrophotometer - Single- or dual-channel, single- or double-beam
instrument having a grating monochromator, photomultiplier detector, adjustable slits, a wavelength
range of 190 to 800 nm, and provisions for a computer or graphical interface.
6.2 Burner - The burner recommended by the particular instrument manufacturer should
be used. For certain elements the nitrous oxide burner is required. Under no circumstance should
an acetylene-air burner head be used with an acetylene-nitrous oxide flame,
6.3 Hollow cathode lamps - Single-element lamps are preferred, but multielement lamps
may be used. Electrodeless discharge lamps may also be used when available. Other types of
lamps meeting the performance criteria of this method may be used.
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6.4 Graphical display and recorder - A recorder is recommended for flame work so that
there will be a permanent record and that any problems with the analysis such as drift, incomplete
atomization, losses during charring, changes in sensitivity, peak signal, etc., can be easily
recognized.
6.5 Pipets - Class A or microliter, with disposable tips. Sizes can range from 5 to 100 uL
as required. Pi pet tips should be checked as a possible source of contamination when
contamination is suspected or when a new source or batch of pipet tips is received by the laboratory.
. The accuracy of variable pipets must be verified daily. Class A pipets can be used for the
measurement of volumes equal to or larger than 1 mL.
6.6 Pressure-reducing valves - The supplies of fuel and oxidant should be maintained at
pressures somewhat higher than the controlled operating pressure of the instrument by suitable
valves.
6.7 Glassware - All glassware, polypropylene, or fluorocarbon (PFA or TFM) containers,
including sample bottles, flasks and pipets, should be washed in the following sequence: 1:1
hydrochloric acid, tap water, 1:1 nitric acid, tap water, detergent, tap water, and reagent water.
(Chromic add should not be used as a cleaning agent for glassware if chromium is to be included
in the analytical scheme.) If it can be documented through an active analytical quality control
program using spiked samples and method blanks that certain steps in the cleaning procedure are
not required for routine samples, those steps may be eliminated from the procedure. Alternative
cleaning procedures must also be documented.
6.8 Volumetric flasks of suitable precision and accuracy.
7.0 REAGENTS AND STANDARDS
7.1 Reagents: Analytical reagent grade or trace metals grade chemicals should be used
in all tests. Unless otherwise indicated, it is intended that all reagents shall conform to the
specifications of the Committee on Analytical Reagents of the American Chemical Society, where
such specifications are available. Other grades may be used, provided it is first ascertained that the
reagent is of sufficiently high purity to permit its use without lessening the accuracy of the
determination. All reagents should be analyzed to demonstrate that the reagents do not contain
target analytes at or above the MDL.
7.2 Reagent water: All references to water in this method refer to reagent water unless
otherwise specified. Reagent grade water is defined in Chapter One.
7.3 Nitric acid, HN03: Use a spectrograde acid certified for AA use. Prepare a 1:1 dilution
with water by adding the concentrated acid to an equal volume of water. If the method blank does
not contain target analytes at or above the MDL, then the acid may be used.
7.4 Hydrochloric add (1:1), HCI: Use a spectrograde add certified for AA use. Prepare a
1:1 dilution with water by adding the concentrated add to an equal volume of water. If the method
blank does not contain target analytes at or above the MDL, then the add may be used.
7.5 Fuel and oxidant: High purity acetylene is generally acceptable. Air may be supplied
from a compressed air line, a laboratory compressor, or a cylinder of compressed air and should be
dean and dry. Nitrous oxide is also required for certain determinations. A centrifuge filter on the
compressed air lines is also recommended to remove particulates.
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7.6 Stock standard metal solutions: Stock standard solutions are prepared from analytical
reagent grade High purity metals, oxides, or nonhygroscopic salts using reagent water and redistilled
nitric or hydrochloric acids. Sulfuric or phosphoric adds should be avoided as they produce an
adverse effect on many elements, The stock solutions are prepared at concentrations of 1.000 ma
of the metal oer liter. Commercially available standard solutions may also be used. When using pure
metals (especially wire) for standards preparation, cleaning procedures, as detailed in Chapter
Three, should be used to ensure that the solutions are not compromised. Stability of standards will
be verified through the use of the QC protocols as specified in this method. Comparison of the daily
ICVs and CCVs with the calibration curve enables the standards to be prepared as needed.
7.6.1 Aluminum: Dissolve 1.000 g of aluminum metal in dilute HCI with gentle
warning and dilute to 1 L with reagent water.
7.6.2 Antimony: Carefully weigh 2.743 g of antimony potassium tartrate,
K(Sb0)C4H406»1/2H20, and dissolve in reagent water. Dilute to 1 L with reagent water.
7.6.3 Barium: Dissolve 1.779 g barium chloride, BaCI2*2H20, analytical grade and
dilute to 1 L with reagent water.
7.6.4 Beryllium: Dissolve 11.659 g beryllium sulfate, BeS04, in reagent water
containing 2 mL nitric add (conc.) and dilute to 1 L with reagent water.
7.6.5 Cadmium: Dissolve 1.000 g cadmium metal in 20 mL of 1:1 HN03 and dilute
to 1 L with reagent water.
7.6.6 Calcium: Suspend 2.500 g of caldum carbonate, CaCOa, dried for 1 hour at
180°C in reagent water and dissolve by adding a minimum of dilute HCI. Dilute to 1 L with
reagent water.
7.6.7 Chromium: Dissolve 1.923 g of chromium trioxide, Cr03, in reagent water,
addify (to pM * 2) with redistilled HNO, (cone.), and dilute to 1 L with reagent water.
7.6.8 Cobalt: Dissolve 1.000 g of cobalt metal in 20 mL of 1:'1 HN03 and dilute to
1 L with reagent water. Chloride or nitrate salts of cobalt(ll) may be used. Although numerous
hydrated forms exist, they are not recommended unless the exact composition of the
compound is known.
7.6.9 Copper: Dissolve 1.000 g of electrolytic copper in 5 mL of redistilled HN03
(conc.) and dilute to 1 L with reagent water.
7.6.10 Iron: Dissolve 1.000 g iron wire in 10 mL redistilled HNO, (conc.) and
reagent water and dilute to 1 L with reagent water. Note that iron passivates in conc. HN03,
and therefore some water should be present.
7.6.11 Lead: Dissolve 1.599 g of lead nitrate, PbfNO^, in reagent water, addify with
10 mL redistilled HN03 (conc.), and dilute to 1 L with reagent water.
7.6.12 Lithium: Dissolve 5.324 g lithium carbonate, Li2C03, in a minimum volume
of 1:1 HCI and dilute to 1 L with reagent water.
7.6.13 Magnesium: Dissolve 1.000 g of magnesium metal in 20 mL 1:1 HN03 and
dilute to 1 L with reagent water.
7000B-7
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7,6.14 Manganese: Dissolve 1.000 g manganese metal in 10 mL redistilled HN03
(cone.) and dilute to 1 L with reagent water.
7.6.15 Molybdenum: Dissolve 1.840 g of ammonium molybdate, (NHJ6Mo7024*4H20,
and dilute to 1 L with reagent water,
7.6.16 Nickel: Dissolve 1.000 g nickel metal or 4.953 g nickel nitrate,
Ni(N03)2»6H20, in 10 mL HNOa (conc.) and dilute to 1 L with reagent water,
7.6.17 Osmium: Procure a certified aqueous standard from a supplier and verify by
comparison with a second standard. If necessary, standards can be made from osmium
compounds. However, due to the toxicity of these compounds, this approach is not advised,
7.6.18 Potassium: Dissolve 1.907 g of potassium chloride, KCI, dried at 110°C, in
reagent water and dilute to 1 L with reagent water
7.6.19 Silver: Dissolve 1.575 g of anhydrous silver nitrate, AgN03, in reagent water.
Add 10 mL of HN03 (conc.) and dilute to 1 L with reagent water. Store in a dark-colored glass
bottle in a refrigerator.
7.6.20 Sodium: Dissolve 2.542 g sodium chloride, NaCI, in reagent water, acidify
with 10 mL redistilled HN03 (conc.), and dilute to 1 L with reagent water.
7.6.21 Strontium: Dissolve 2.415 g of strontium nitrate, Sr(N03)2, in 10 mL of conc.
HQ and 700 mL of reagent water. Dilute to 1 L with reagent water.
7.6.22 Thallium: Dissolve 1.303 g thallium nitrate, TIN03, in reagent water, acidify
(to pH s 2) with 10 mL conc. HN03, and dilute to 1 L with reagent water.
I .
7.6.23 Tjo: Dissolve 1.000 g of tin metal in 100 mL conc. HCI and dilute to 1 L with
reagent water.
7.6.24 Vanadium: Dissolve 1.785 g of vanadium pentoxide, V2Os, in 10 mL of conc.
HN03 and dilute to 1 L with reagent water.
7.6.25 Zinc: Dissolve 1.000 g zinc metal in 10 mL of conc. HN03 and dilute to 1 L
with reagent water.
7.7 Aluminum nitrate solution: Dissolve 139 g aluminum nitrate, AI(N03)3«9H20, in 150 mL
reagent water and heat to effect solution. Allow to cool and make to 200 mL. Add 2 mL of this
solution to a 100 mL volume of standards and samples.
7.8 Lanthanum chloride/potassium chloride solution: Dissolve 11.73 g of lanthanum oxide,
LaA, in a minimum amount (approximately 50 mL) of conc. HCI. Add 1.91 g of potassium chloride,
KCI. Allow solution to cool to room temperature and dilute to 100 mL with reagent water. CAUTION
- REACTION IS VIOLENT! Add add slowly and in small portions to control the reaction rate upon
mixing.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 See the introductory material in Chapter Three, Inorganic Analytes.
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9.0 QUALITY CONTROL
9.1 All quality control data should be maintained and available for easy reference or
inspection.
9.2 For each batch of samples processed, at least one method blank must be carried
throughout the entire sample preparation and analytical process as described in Chapter One. A
method blank is prepared by using a volume or weight of reagent water at the volume or weight
specified in the preparation method and then earned through the appropriate steps of the analytical
process. These steps may include but are not limited to digestion, dilution, filtering, and analysis.
If the method blank does not contain target analytes at a level that interferes with the project-specific
DQOs then the method blank would be considered acceptable, in the absence of project-specific
DQOs, if the blank is less than the MDL or less than 10% of the lowest sample concentration for
each analyte, whichever is greater, then the method blank would be considered acceptable. If the
method blank cannot be considered acceptable, the method blank should be re-run once and if still
unacceptable then all samples after the last acceptable method blank must be reprepped and
reanalyzed along with the other appropriate batch QC samples. These blanks will be useful in
determining if samples are being contaminated. Refer to Chapter One for the proper protocol when
analyzing blanks.
9.3 For each batch of samples processed, at least one laboratory control samples must be
carried throughout the entire sample preparation and analytical process as described in Chapter
One. The laboratory control samples should be spiked with each analyte of interest at the project-
specific action level or when lacking project-specific action levels, between the low and midlevel
standards. Acceptance criteria should be set at a laboratory derived limit developed through the use
of historical analyses. In the absence of historical date this limit should be set at ± 20% of the spiked
value. After the determination of historical data, ±20% must still be the limit of maximum deviation
to express acceptability. If the laboratory control sample cannot be considered acceptable, the
laboratory control sample should be re-run once and if still unacceptable then ail samples after the
last acceptable laboratory control sample must be reprepped and reanalyzed. Refer to Chapter One
for more information.
9.4 Matrix Spike/Matrix Spike Duplicates (MS/MSDs): At the laboratory's discretion, a
separate spike sample and a separate duplicate sample may be analyzed in lieu of the MS/MSD. For
each batch of samples processed, at least one MS/MSD sample must be carried throughout the
entire sample preparation and analytical process as described in Chapter One. MS/MSDs are
intralaboratory split samples spiked with identical concentrations of each analyte of interest. The
spiking occurs prior to sample preparation and analysis. An MS/MSD is used to document the bias
and precision of a method in a given sample matrix.. Refer to the definitions of bias and precision,
in Chapter One, for the proper data reduction protocols. MS/MSD samples should be spiked at the
same level as the corresponding laboratory control sample that is at the project-specific action level
or, when lacking project-specific action levels, between the low and midlevel standards. Acceptance
criteria should be set at a laboratory derived limit developed through the use of historical analyses.
In the absence of historical data this limit should be set at ± 20% of the spiked value for precision
and £ 20 relative percent difference (RPD). After the determination of historical data, 20% must still
be the limit of maximum deviation for both percent recovery and relative percent difference to
express acceptability. Refer to of Chapter One for guidance. If the bias and precision indicators are
outside the laboratory control limits or if the percent recovery is less than 80% or greater than 120%
or if the relative percent difference is greater than 20%, the interference test as discussed in Sec.
9.5.2 and 9.7 should be conducted.
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9.5 Interference tests
9.5.1 Recovery test (post-digestion spike) - The recovery test must be done on all
samples within a batch that fails that batch's MS/MSD. To conduct this test, withdraw an
aliquot of the test sample and add a known amount of anafyte to bring the concentration of the
analyte to 2 to 5 times the original concentration. If spiking at 2-5 times would exceed the linear
range of the instrument, a lesser spike may be used, if all of the samples in the batch have
analyte concentrations below the detection limit, spike the selected sample at the project-
specific action level or when lacking project-specific action levels, between the low and
midlevel standards. Analyze the spiked sample and calculate the spike recovery. If the
recovery is less than 85% or greater than 115%, the method of standard additions should be
used for all samples in the batch.
9.5.2 Dilution test - The dilution test is to be conducted when interferences are
suspected and the sample concentration is high enough to allow for proper interpretation of
the results. To conduct this test, determine the apparent concentration in the undiluted
sample. Dilute the sample by a minimum of five fold (1+4) and reanalyze. Agreement within
a 10% difference ( RPD) between the concentration for the undiluted sample and five times
the concentration for the diluted sample indicates the absence of interferences, and such
samples may be analyzed without using the method of standard additions. If agreement
between the dilutions is greater than 10%, the MSA should be used for all samples in the
batch.
9.6 Where the sample matrix is so complex that viscosity, surface tension, and components
cannot be accurately matched with standards, the method of standard addition (MSA) is
recommended (see Section 9.7 below). Other options including, the use of different matrix
modifiers, different furnace conditions, different preparatory methods or different analytical methods
may also be attempted to properly characterize a samplel Section 9.5 provides tests to determine
the potential of an interference and evaluates the need for using the MSA.
9.7 Method of standard additions - The standard addition technique involves adding known
amounts of standard to one or more aliquots of the processed sample solution. This technique
attempts to compensates for a sample constituent that enhances or depresses the analyte signal,
thus producing a different slope from that of the calibration standards. It will not correct for additive
interferences which cause a baseline shift. The method of standard additions may be appropriate
for analysis of extracts, on analyses submitted as part of a delisting petition, whenever a new
sample matrix is being analyzed and on every batch that fails the recovery test.
9.7.1 The simplest version of this technique is the single-addition method, in which
two identical aliquots of the sample solution, each of volume V„ are taken. To the first (labeled
A) is added a known volume Vs of a standard analyte solution of concentration Cs. To the
second aliquot (labeled B) is added the same volume Vs of reagent water. The analytical
signals of A and B are measured and corrected for non-anajyte signals. The unknown sample
concentration C, is calculated:
C
"-(Sa-W,
where SA and S8 are the analytical signals (corrected for the blank) of solutions A and B,
respectively. Vs and C8 should be chosen so that SA is roughly twice S8 on the average,
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avoiding excess dilution of the sample. If a separation or concentration step is used, the additions
are best made first and carried through the entire procedure.
9.7.2 Improved results can be obtained by employing a series of standard additions.
To equal volumes of the sample are added a series of standard solutions containing different
known quantities of the analyte, and all solutions are diluted to the same final volume. For
example, addition 1 should be prepared so that the resulting concentration is approximately
50 percent of the expected absorbance from the indigenous analyte in the sample. Additions
2 and 3 should be prepared so that the concentrations are approximately 100 and 150 percent
of the expected endogenous sample absorbance. The absorbance of each solution is
determined and then plotted on the vertical axis of a graph, with the concentrations of the
known standards plotted on the horizontal axis. When the resulting line is extrapolated to zero
absorbance, the point of interception of the abscissa is the endogenous concentration of the
analyte in the sample. The abscissa on the left of the ordinate is scaled the same as on the
right side, but in the opposite direction from the ordinate. An example of a plot so obtained is
shown in Figure 1. A linear regression program may be used to obtain the intercept
concentration.
9.7.3 For the results of this MSA technique to be valid, the following limitations must
be taken into consideration:
1. The apparent concentrations from the calibration curve must be linear (0.995
or greater) over the concentration range of concern. For the best results, the
slope of the MSA plot should be nearly the same as the slope of the standard
curve.
2. The effect of the interference should not vary as the ratio of analyte
concentration to sample matrix changes, and the standard addition should
respond in a similar manner as the analyte.
3. The determination must be free of spectral interference and corrected for
nonspecific background interference.
9.8 All quality control measures described in Chapter One should be followed.
9.9 Independent source laboratory control samples or Standard Reference Materials
(SRMs) should be used to help assess the quality of the analysis scheme. Follow the directions
provided by the SRM's manufacturer for use and acceptance criteria.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Calibration standards - For those instruments which do not read out directly in
concentration, a calibration curve is prepared to cover the appropriate concentration range. Usually,
this means the preparation of a blank and standards which produce an absorbance of 0.0 to 0.7.
Calibration standards can prepared by diluting the stock metal solutions in the same acids and acid
concentrtions as the samples.
10.1.1 Calibration standards can be prepared fresh each time a batch of samples
is analyzed. If the ICV solution is prepared daily and the ICV is analyzed within the acceptance
criteria, calibration standards do not need to be prepared daily and may be prepared and
stored for as long as the calibration standard viability can be verified through the use of the
ICV. If the ICV is outside of the acceptance criteria, the calibration standards must be prepared
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fresh arid the instalment recalibrated. Prepare a blank and at least three calibration standards
in graduated amounts in the appropriate range of the linear part of the curve.
10.1.2 The calibration standards should be prepared using the same type of add or
combination of acids and at the same concentration as will result in the samples following
processing.
10.1.3 Beginning with the blank and working toward the highest standard, aspirate
the solutions and record the readings. Repeat the operation with both the calibration standards
and the samples a sufficient number of times to secure an average reading for each solution.
Calibration curves are always required.
10.2 A calibration curve must be prepared each day with a minimum of a calibration blank
and three standards. The curve must be linear and have a correlation coefficient of at least 0.995.
10.2.1 After initial calibration, the calibration curve must be verified by use of an
initial calibration blank (ICB) and an initial calibration verification (ICV) standard. The ICV
standard must be made from an independent (second source) material at or near mid-range.
The acceptance criteria for the ICV standard must be ±10% of its true value and the ICB must
not contain target analytes at or above the MDL for the curve to be considered valid. If the
calibration curve cannot be verified within the specified limits, the cause must be determined
and the instrument recalibrated before samples are analyzed. The analysis data for the ICV
must be kept on file with the sample analysis data.
10.2.2 The calibration curve must also be verified at the end of each analysis batch
and/or after every 10 samples by use of a continuing calibration blank (CCS) and a continuing
calibration verification (CCV) standard. The CCV standard should be made from the same
material as the initial calibration standards at or near midrange. The acceptance criteria for
the CCV standard must be ±10% of its true value and the CCB must not contain target
analytes at or above the MDL for the curve to be considered valid. If the calibration cannot be
verified within the specified limits, the sample analysis must be discontinued, the cause
determined and the instrument recalibrated. All samples following the last acceptable
CCV/CCB must be reanalyzed. The analysis data for the CCV/CCB must be kept on file with
the sample analysis data.
10.3 It is recommended that each standard should be analyzed (injected) twice and an
average value determined. Replicate standard values should be within ±10% RPD.
10.4 If conducting trace analysis, it is recommended that the lowest calibration standard be
set at the laboratory's quantitation level. The laboratory can use a reporting limit that is below the
quantitation level but all values reported below the low standard should be reported as estimated
values.
11.0 PROCEDURE
11.1 Preliminary treatment of aqueous and solid wastes is always necessary because of the
complexity and variability of sample matrices. Solids, slurries, and suspended material must be
subjected to a solubilization process before analysis. This process may vary because of the metals
to be determined and the nature of the sample being analyzed. Solubilization and digestion
procedures are presented in Chapter Three. Samples which are to be analyzed for dissolved
constituents need not be digested *rf they have been filtered and then acidified. See first note of
Section 1.0.
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11.2 All atomic absorption analyses must be performed using a suitable form of background
correction. Refer to Chapter Two for a detailed discussion on background correction.
11.3 Differences between the various makes and models of satisfactory atomic absorption
spectrophotometers prevent the formulation of detailed instructions applicable to every instrument.
The analyst should follow the manufacturer's operating Instructions for a particular instrument.
11.3.1 In general, after choosing the proper lamp for the analysis, allow the lamp to
warm up for a minimum of 15 minutes.
11.3.2 During this period, align the instrument, position the monochromator at the
correct wavelength, select the proper monochromator slit width, and adjust the current
according to the manufacturer's recommendation.
11.3.3 Light the flame and regulate the flow of fuel and oxidant. Adjust the burner
and nebulizer flow rate for maximum percent absorption and stability. Balance the photometer. *
11.3.4 Run a series of standards of the element under analysis. Construct a
calibration curve by plotting the concentrations of the standards against absorbances. Set the
curve corrector of a direct reading instrument to read out the proper concentration.
11.3.5 Aspirate the samples and determine the concentrations either directly or from
the calibration curve. Standards must be run each time a sample or series of samples is run.
12.0 DATA ANALYSIS AND CALCULATIONS
12.1 For determination of metal concentration, read the concentration from the calibration
curve or directly from the read-out system of the instrument.
12.1.1 If dilution of the sample was required:
yg/L metal in sample = —
.0
where:
A = pg/L of metal in diluted aliquot from calibration curve.
B = Starting sample volume, mL.
C = Final volume of sample, mL.
12.1.2 For solid samples, report all concentrations in consistent units based on
weight. Ensure that if the dry weight was used for the analysis, percent solids should be
reported to the client.
A x V
mg metal/kg sample*
W
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where:
A = mg/L of metal in processed sample from calibration curve.
V = Final volume of the processed sample, L.
W = Weight of sample, Kg.
12.1.3 Different integration times must not be used for samples and standards.
Instead, the sample should be diluted and the same integration time should be used for both
samples and standards. If dilution of the sample was required:
ji/L of metal sample = — ^^
c
where:
Z = MS"- of metal read from calibration curve or read-out system.
B = Starting sample volume, mL.
C = Final volume of sample, mL.
13.0 METHOD PERFORMANCE
13.1 Refer to the individual applicable methods from reference 1.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity
and/or toxicity of waste at the point of generation. Numerous opportunities for pollution prevention
exist in laboratory operation. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of first choice.
Whenever feasible, laboratory personnel should use pollution prevention techniques to address their
waste generation. When wastes cannot be feasibly reduced at the source, the Agency recommends
recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Less is Better Laboratory Chemical Management for Waste Reduction
available from the American Chemical Society, Department of Government Relations and Science
Policy, 1155 16th Street, NW, Washington, DC 20036, (202) 872-4477.
15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society at the address listed in Section 14.2.
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16.0 REFERENCES
1. Methods for Chemical Analysis of Water and Wastes: U.S. Environmental Protection Agency,
Office of Research and Development Environmental Monitoring and Support Laboratory. ORD
Publication Offices of Center for Environmental Research Information: Cincinnati, OH, 1983;
EPA-600/4-79-020.
2. Reagent Chemicals. American Chemical Society Specifications. Rohrtoough, W.Q.; et al. 7th
ed.; American Chemical Society: Washington, DC, 1986.
3. 1985 Annual Book of ASTM Standards. Vol. 11.01; "Standard Specification for Reagent
Water"; ASTM: Philadelphia, PA, 1985; D1193-77.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 and 2, Figure 1 and a flow diagram of the method procedures.
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TABLE 1
ATOMIC ABSORPTION DETECTION LIMITS AND SENSITIVITY FOR ANALYTES
IN REAGENT WATER
Direct Aspiration
Detection Limit Sensitivity
Metal (mg/L) (mg/L)
Aluminum
0.1
1
Antimony
0.2
0.5
Barium
0.1
0.4
Beryllium
0.005
0,025
Cadmium
0.005
0.025
Calcium
0.01
0.08
Chromium
0.05
0.25
Cobalt
0.05
0.2
Copper
0.02
0.1
Iron
0.03
0.12
Lead
0.1
0.5
Lithium
0.002
0.04
Magnesium
0.001
0.007
Manganese
0.01
0.05
Molybdenum
0.1
0.4
Nickel
0.04
0.15
Osmium
0.03
1
Potassium
0.01
0.04
Silver
0.01
0,06
Sodium
0.002
0.015
Strontium
0.03
0.15
Thallium
0.1
0.5
Tin
0.8
4
Vanadium
0.2
0.8
Zinc
0.005
0.02
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TABLE 2
INSTRUMENT PARAMETERS (Ref. 1)
ELEMENT
WAVELENGTH
(rimy
FUEL
OXIDANT
TYPE OF FLAME
Al
324.7
acetylene
nitrous oxide
fuel rich
Sb
217.6.
2311
acetylene
air
fuel lean
ia
553.6
acetylene
nitrous oxide
fuel rich
Be
234.9
acetylene
nitrous oxide
fuel rich
Cd
228.8
acetylene
air
fuel lean
Ca
422.7
acetylene
nitrous eodde.
stoichiometric
Cr
357.9
acetylene
nitrous oxide
fuel rich
Co
240.7
acetylene
air
fuel lean
Cu
324,7
acetylene
air
fuel lean
Fe
248.3.
248.8,271.8,
302.1.252.7
acetylene
air
fuel lean
Pto
283.3.
217.0
acetylene
air
fuel lean
U
670,8
acetylene
air
hid lean
Ma
285.2
acetylene
air
fuel lean
Mn
279 S
403.1
acetylene
air
fuel lean to
stoichiometric
Mo
313.3
acetylene
nitrous oxide
fuel rich
Ni
232.0.
352.4
acetylene
air
fuel lean
08
290.0
acetylene
nitrous oxide
fuel rich
K
766.5
acetylene
air
fuel lean
Aa
328.1
acetylene
air
fuel lean
Na
589.6
acetylene
air
fuel lean
Sr
480.7
acetylene
air
fuel lean
n
276.8
acetylene
air
fuel lean
Sn
266.3
acetylene
nitrous oxide
fuel rich
V
318,4
acetylene
nitrous oxide
fuel rich
Zn
213.9
acetylene
air
fuel lean
Note; If more than one wavelength is listed, the primary line is underlined.
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FIGURE 1
STANDARD ADDITION PLOT
Zero
Absorbance
Cone, of
Sample
AddnO
No Addn
Addn 1
Addn of 50%
Addn 2 Addn 3
Addn of 100% Addn of 150%
of Expected of Expected of Expected
Amount Amount Amount
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METHOD 7000B
FLAME ATOMIC ABSORPTION SPECTROPHOTOMETRY
Start
Stop
12.0 Determine concentrations.
11.3.5 Aspirate sample.
11.3.1 Choose and prepare
proper lamp for analysis.
11.3.2 Adjust and align
• equipment.
11.3.3 Light the flame and regulate flow
of fuel and oxidant.
11.1 Sotubilize or digest samples
(See Chapter Three, Section 3.2.)
11.2 Make Background
Correction.
11,3.4 Run standards. Construct
calibration curve and set curve
corrector, (See Section 10.0.)
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METHOD 7010
GRAPHITE FURNACE ATOMIC ABSORPTION SPECTROPHOTOMETRY
10 SCOPE AND APPLICATION
1.1 Metals in solution may be readily determined by graphite furnace atomic absorption
spectrophotometry (GFAA). The method is simple, quick, and applicable to a large number of metals
in environmental samples including, but not limited to, ground water, domestic and industrial wastes,
extracts, soils, sludges, sediments, and similar wastes. With the exception of the analyses for
dissolved constituents, all samples require digestion prior to analysis. Analysis for dissolved
elements does not require digestion if the sample has been filtered and then acidified.
NOTE: The analyst should be aware that organo-metallic species may not be detected if the
sample is not digested.
This method is applicable to the following elements:
Element

CASRN*
Antimony
(Sb)
7440-36-0
Arsenic
(As)
7440-38-2
Barium
(Ba)
7440-39-3
Beryllium
(Be)
7440-41-7
Cadmium
(Cd)
7440-43-9
Chromium
(Cr)
7440-47-3
Cobalt
(Co)
7440-48-4
Copper
(Cu)
7440-50-8
iron
(Fe)
7439-89-6
Lead
(Pb)
7439-92-1
Manganese
(Mn)
7439-96-5
Molybdenum
(Mo)
7439-98-7
Nickel
(Ni)
7440-02-0
Selenium
(Se)
7782-49-2
Silver
(Ag)
7440-22-4
Thallium
(Tl)
7440-28-0
Vanadium
(V)
7440-62-2
Zinc
(Zn)
7440-66-6
""Chemical Abstract Service Registry Number
1.2 Method detection limits, sensitivity, and optimum ranges of the metals will vary with
the matrices and models of atomic absorption spectrophotometers. The data shown in Table 1
provide some indication of the detection limits obtainable by the furnace technique. The detection
limits given in Table 1 are somewhat dependent on equipment (such as the type of
spectrophotometer and furnace accessory, the energy source, the degree of electrical expansion of
the output signal), and are greatly dependent on sample matrix. Method detection limits (MDLs)
must be established, empirically, for each matrix type analyzed (refer to Chapter One for guidance)
and are required for each preparatory/determinative method combination used.
1.3 Users of this method should state the data quality objectives prior to analysis and must
document and have on file the required initial demonstration performance data described in the
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following sections prior to using the method for analysis. When using furnace techniques, the
analyst should be cautioned as to possible chemical reactions occurring at elevated temperatures
which may result in either suppression or enhancement of the analysis element (see Sec. 4.0). To
ensure valid data with furnace techniques, the analyst must examine each sample for interference
effects (see Sec. 9.0) and, if detected, treat them accordingly, using either successive dilution,
matrix modification, or the method of standard additions (see Sec. i.7).
1.4 Other elements and matrices may be analyzed by this method as long as the method
performance is demonstrated for these additional elements of interest, in the additional matrices of
interest, at the concentration levels of interest in the same manner as the listed elements and
matrices (see Sec. 9.0).
1.5 Use of this method is restricted to analysts who are knowledgeable in the chemical and
physical interferences as described in this method.
2.0 SUMMARY OF THE METHOD
2.1 Although methods have been reported for the analysis of solids by atomic absorption
spectrophotometry, the technique generally is limited to metals in solution or solubilized through
some form of sample processing. Refer to Chapter Three for a description of appropriate digestion
methods.
2.2 Preliminary treatment of wastes, both solid and aqueous, is always necessary because
of the complexity and variability of sample matrix. Solids, slurries, and suspended material must be
subjected to a solubilization process before analysis. This process may vary because of the metals
to be determined and the nature of the sample being analyzed. Solubilization and digestion
procedures are presented in Chapter Three.
2.3 When using the furnace technique in conjunction with an atomic absorption
spectrophotometer, a representative aliquot of a sample is placed in. the graphite lube in the furnace,
evaporated to dryness, charred, and atomized. As a greater percentage of available analyte atoms
is vaporized and dissociated for absorption in the tube rather than the flame, the use of smaller
sample volumes or detection of lower concentrations of elements is possible. The principle is
essentially the same as with direct aspiration atomic absorption, except that a fumace, rather than
a flame, is used to atomize the sample. Radiation from a given excited element is passed through
the vapor containing ground-state atoms of that element. The intensity of the transmitted radiation
decreases in proportion to the amount of the ground-state element in the vapor. The metal atoms
to be measured are placed in the beam of radiation by increasing the temperature of the furnace,
tfiereby causing the injected specimen to be volatilized. A monochromator isolates the characteristic
radiation from the hollow cathode lamp or electrodeless discharge lamp, and a photosensitive device
measures the attenuated transmitted radiation.
3.0 DEFINITIONS
Refer to Chapter One and Chapter Three for a listing of applicable definitions.
4.0 INTERFERENCES
4.1 Although the problem of oxide formation is greatly reduced with fumace procedures
(because atomization occurs in an inert atmosphere), the technique is still subject to chemical
interferences. The composition of the sample matrix can have a major effect on the analysis. It is
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those effects which must be determined arid taken into consideration in the analysis of each different
matrix encountered. See Sec. 9.6 for additional guidance.
4.2 Background correction is important when using flameless atomization, ©specialty below
350 nm. Certain samples, when atomized, may absorb or scatter light from the lamp. This can be
caused by the presence of gaseous molecular species, salt particles, or smoke in the sample beam.
If no correction is made, sample absorbance will be greater than it should be, and the analytical
result will be erroneously high. Zeeman background correction is effective in overcoming composition
or structured background interferences. It is particularly useful when analyzing for As in the presence
of A! and when analyzing for Se in the presence of Fe.
4.3 Memory effects occur when the analyte is not totally volatilized during atomization. This
condition depends on several factors: volatility of the element and its chemical form, whether
pyrolytic graphite is used, the rate of atomization, and furnace design. This situation is detected
through blank bums. The tube should be cleaned by operating the furnace at full power for the
required time period, as needed, at regular intervals during the series of determinations.
4.4 Gases generated in the furnace during atomization may have molecular absorption
bands encompassing the analytical wavelength. When this occurs, use either background correction
or choose an alternate wavelength. Background correction may also compensate for nonspecific
broad-band absorption interference and light scattering.
4.5 Continuum background correction cannot correct for all types of background
interference. When the background interference cannot be compensated for, chemically remove the
analyte or use an alternate form of background correction, refer to Chapter Two. A single
background correction device to be used with this method is not specified; however, it must provide
an analytical condition that is not subject to the occurring interelement spectral interferences of
palladium on copper, iron on selenium and aluminum on arsenic.
4.6 Interference from a smoke-producing sample matrix can sometimes be reduced by
extending the charring time at a higher temperature or utilizing an ashing cycle in the presence of
air. Care must be taken, however, to prevent loss of the analyte.
4.7 Samples containing large amounts of organic materials should be oxidized by
conventional acid digestion before being placed in the furnace. In this way, broad-band absorption
will be minimized.
4.8 Anion interference studies in the graphite furnace indicate that, under conditions other
than isothermal, the nitrate anion is preferred. Therefore, nitric acid is preferable for any digestion
or solubilization step. When another acid in addition to nitric acid is required, a minimum amount
should be used. This applies particularly to hydrochloric and, to a lesser extent, to sulfuric and
phosphoric acids.
4.9 Carbide formation resulting from the chemical environment of the furnace has been
observed. Molybdenum may be cited as an example. When carbides form, the metal is released very
slowly from the resulting metal carbide as atomization continues. Molybdenum may require 30
seconds or more atomization time before the signal returns to baseline levels/Carbide formation is
greatly reduced and the sensitivity increased with the use of pyrolytically coated graphite. Elements
that readily form carbides are noted with the symbol (p) in Table 1.
4.10 Spectral interference can occur when an absorbing wavelength of an element present
in the sample, but not being determined, falls within the width of the absorption line of the element
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of interest. The results of the determination will then be erroneously high, due to the contribution of
the interfering element to the atomic absorption signal, interference can also occur when resonant
energy from another element in a multielement lamp, or from a metal impurity in the lamp cathode,
falls wrttiin the bandpass of the slit setting when that other metal is present in the sample. This type
of interference may sometimes be reduced by narrowing the slit width.
4.11 It is recommended that all graphite furnace analyses be carried out using an
appropriate matrix modifier. The choice of matrix modifier is dependent on analytes, conditions, and
instrumentation and should be chosen by the analyst as the situation dictates. Follow the instrument
manufacturers instructions for the preferred matrix modifier. If necessary, refer to Chapter Two for
additional guidance.
4.12 It is recommended that a stabilized temperature platform be used to maximize an
isothermal environment within the furnace cell to help reduce interferences. Refer to Chapter Two
for additional guidance.
4.13 Cross-contamination and contamination of the sample can be major sources of error
because of the extreme sensitivities achieved with the furnace. The sample preparation work area
should be kept scrupulously clean. All glassware should be cleaned as directed in Sec. 6.6. Pipet
tips are a frequent source of contamination. The analyst should be aware of the potential for the
yellow tips to contain cadmium. If suspected, they should be acid soaked with 1:5 nitric acid and
rinsed thoroughly with tap and reagent water. The use of a better grade of pipet tip can greatly
reduce this problem. Special attention should be given to assessing the contamination in method
blanks during the analysis. Pyrolytic graphite, because of the production process and handling, can
become contaminated. As many as five to ten high-temperature bums may be required to clean the
tube before use. In addition, auto sampler tips may also be a potential source of contamination.
Flushing the tip with a dilute solution of nitric acid between samples can help prevent cross-
contamination.
4.14 Specific interference problems related to individual analytes are located in this
section.
4.14.1 Antimony: High lead concentration may cause a measurable spectral
interference on the 217.6 nm line. Choosing the secondary wavelength or using background
correction may correct the problem.
4.14.2 Arsenic:
4.14.2.1 Elemental arsenic and many of its compounds are volatile;
therefore, samples may be subject to losses of arsenic during sample preparation
Likewise, caution must be employed during the selection of temperature and times
for the dry and char (ash) cycles. A matrix modifier such as nickel nitrate or
palladium nitrate should be added to all digestates prior to analysis to minimize
volatilization tosses during drying and ashing.
4.14.2.2 In addition to the normal interferences experienced during
graphite furnace analysis, arsenic analysis can suffer from severe nonspecific
absorption and light scattering caused by matrix components during atomization.
Arsenic analysis is particularly susceptible to these problems because of its low
analytical wavelength (133.7 nm). Simultaneous background correction must be
employed to avoid erroneously high results. Aluminum is a severe positive
interferant in the analysis of arsenic, especially using D2 arc background correction.
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Although Zeeman background correction is very useful in this situation, use of any
appropriate background correction technique is acceptable.
4.14.3 Barium: Barium can form barium carbide in the furnace, resulting in less
sensitivity and potential memory effects. Because of chemical interaction, nitrogen should not
be used as a purge gas and halide adds should not be used.
4.14.4 Beryllium: Concentrations of aluminum greater than 500 ppm may suppress
beryllium absorbance. The addition of 0.1% fluoride has been found effective in eliminating
this interference. High concentrations of magnesium and silicon cause similar problems and
require the use of the method of standard additions.
4.14.5 Cadmium: Cadmium analyses can suffer from severe non-specific absorption
and light scattering caused by matrix components during atomization. Simultaneous
background correction is required to avoid erroneously high results. Excess chloride may
cause premature volatilization of cadmium; an ammonium phosphate matrix modifier may
minimize this loss.
4.14.6 Chromium: Low concentrations of calcium and/or phosphate may cause
interferences; at concentrations above 200 mg/L, calcium's effect is constant and eliminates
the effect of phosphate. Therefore, add calcium nitrate (calcium nitrate solution: dissolve 11.8
g of calcium nitrate in 1 L reagent water) to ensure a constant effect. Nitrogen should not be
used as the purge gas because of a possible CN band interference.
4.14.7 Cobalt: Since excess chloride may interfere, it is necessary to verity by
standard additions that the interference is absent unless it can be shown that standard
additions are not necessary.
4.14.8 Lead: If poor recoveries are obtained, a matrix modifier may be necessary.
Add 10 uL of phosphoric acid to 1 mL of prepared sample.
4.14:9 Molybdenum: Molybdenum is prone to carbide formation; use a pyrolytically
coated graphite tube.
4.14.10 Nickel: Severe memory effects for nickel may occur in graphite furnace tubes
used for other GFAA analyses, due to the use of a nickel nitrate matrix modifier in those
methods. Use of graphite furnace tubes and contact rings for nickel analysis that are separate
from those used for arsenic and selenium analyses is strongly recommended.
4.14.11 Selenium:
4.14.11.1 Elemental selenium and many of its compounds are volatile;
therefore, samples may be subject to losses of selenium during sample preparation.
Likewise, caution must be employed during the selection of temperature and times for
the dry and char (ash) cycles. A matrix modifier such as nickel nitrate or palladium
nitrate should be added to all digestates prior to analysis to minimize volatilization
losses during drying and ashing.
4.14.11.2 In addition to the normal interferences experienced Airing graphite
furnace analysis, selenium analysis can suffer from severe nonspecific absorption and
light scattering caused by matrix components during atomization. Selenium analysis
is particularly susceptible to these problems because of its low analytical wavelength
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(196.0 rim). Simultaneous background correction must be employed to avoid
erroneously high results. High iron levels can give overcorrection with deuterium
background. Although Zeeman background correction is very useful in this situation,
use of any appropriate background correction technique is acceptable.
4.14.11.3 Selenium analysis suffers interference from chlorides (>800 mg/L)
and sulfate (>200 mg/L). The addition of nickel nitrate such that the final concentration
is 1% nickel will lessen this interference.
4.14.12 Silver: Silver chloride is insoluble, therefore HCI should be avoided unless
the silver is already in solution as a chloride complex. In addition, it is recommended that the
stock standard concentrations be kept below 2 ppm and the chloride content increased to
prevent precipitation. If precipitation is occurring, a 5%:2% HCI:HN03 stock solution may
prevent precipitation. Daily standard preparation may also be needed to prevent precipitation
of silver. Analysts should be aware that this technique may not be the best choice for this
analyte and that alternative techniques should be considered.
4.14.13 Thallium: HCI or excessive chloride will cause volatilization of thallium at low
temperatures. Verification that losses are not occurring must be made for each matrix type
(as detailed in 9.6.1).
4.14.14 Vanadium: Vanadium is refractory and prone to form carbides. Consequently,
memory effects are common, and care should be taken to clean the furnace before and after
analysis.
5.0 SAFETY
Refer to Chapter Three for a discussion on safety related references and issues.
6.0 EQUIPMENT AND SUPPLIES
6.1 Atomic absorption spectrophotometer - Single- or dual-channel, single- or double-beam
instrument having a grating monochromator, photomultiplier detector, adjustable slits, a wavelength
range of 190 to 800 nm, and provisions for interfacing with a graphical display. The instrument must
be equipped with an adequate correction device capable of removing undesirable nonspecific
absorbance over the spectral region of interest and provide an analytical condition not subject to the
occurrence of interelement spectral overlap interferences.
6.2 Hollow cathode lamps - Single-element lamps are preferred but multielement lamps
may be used. Eiectrodeless discharge lamps may also be used when available. Other types of
lamps meeting the performance criteria of this method may be used.
6.3 Graphite furnace - Any furnace device capable of reaching the specified temperatures
is satisfactory. For all instrument parameters (i.e., drying, ashing, atomizing, times and
temperatures) follow the specific instrument manufacturers instructions for each element.
6.4 Data systems recorder - A recorder is recommended for furnace work so that there will
be a permanent record and that any problems with the analysis such as drift, incomplete atomization,
losses during charring, changes in sensitivity, peak shape, etc., can be easily recognized.
6.5 Pipets - Microliter, with disposable tips. Sizes can range from 5 to 100 yL as required.
Pipet tips should be checked as a possible source of contamination when contamination is
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suspected or when a new source or batch of pipet tips is received by the laboratory. The accuracy
of variable pipets must be verified daily. Class A pipets can be used for the measurement of
volumes equal to or larger than 1 mL.
6.6 Glassware - AH glassware, polypropylene, or fluorocarbon (PFA or TFE) containers,
including sample bottles, flasks and pipets, should be washed in the following sequence: 1:1
hydrochloric acid, tap water, 1:1 nitric acid, tap water, detergent, tap water, and reagent water.
Chromic add should not be used as a cleaning agent for glassware if chromium is to be included
in the analytical scheme. If it can be documented through an active analytical quality control program
using spiked samples and method blanks that certain steps in the cleaning procedure are not
required for routine samples, those steps may be eliminated from the procedure. Leaching of
polypropylene for longer periods at lower acid concentrations is necessary to prevent degradation
of the polymer. Alternative cleaning procedures must also be documented. Cleaning for ultra-trace
analysis should be reviewed in Chapter Three.
6.7 Volumetric flasks of suitable precision and accuracy.
7.0 REAGENTS AND STANDARDS
7.1 Reagents: Analytical reagent grade or trace metals grade chemicals should be used
in all tests. Unless otherwise indicated, it is intended that all reagents shall conform to the
specifications of the Committee on Analytical Reagents of the American Chemical Society, where
such specifications are available. Other grades may be used, provided it is first ascertained that the
reagent is of sufficiently high purity to permit its use without lessening the accuracy of the
determination. All reagents should be analyzed to demonstrate that the reagents do not contain
target analytes at or above the MDL.
7.2 Reagent water: All references to water in this method refer to reagent water unless
otherwise specified. Refer to Chapter One for a definition of reagent water.
7.3 Nitric acid, HN03: Use a spectrograde add certified for AA use. Prepare a 1:1 dilution
with water by adding the concentrated acid to an equal volume of water. If the method blank does
not contain target analytes at or above the MDL, then the add may be used.
7.4 Hydrochloric acid (1:1), HCI: Use a spectrograde add certified for AA use. Prepare a
1:1 dilution with water by adding the concentrated add to an equal volume of water. If the method
blank does not contain target analytes at or above the MDL, then the add may be used.
7.5 Purge Gas: A mixture of H2 (5%) and argon (95%). The argon gas supply must be
high-purity grade, 99.99% or better. If performance can be documented, alternative gases may be
used. ~ .
7.6 Stock standard metal solutions: Stock standard solutions are prepared from analytical
reagent grade high purity metals, oxides, or nonhygroscopic salts using reagent water and redistilled
nitric or hydrochloric adds. (See individual methods for spedfic instructions.) Sulfuric or phosphoric
adds should be avoided as they produce an adverse effect on many elements. The stock solutions
are prepared at concentrations of 1.000 mo of the metal oer liter. Commercially available standard
solutions may also be used. When using pure metals (espedally wire) for standards preparation,
cleaning procedures, as detailed in Chapter Three, should be used to ensure that the solutions are
not compromised. Examples of appropriate standard preparations can be found in Sections 7.6.1
through 7.6.18.
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7.6.1 Antimony: Carefully weigh 2.743 g of antimony potassium tartrate,
K(Sb0)C4H406*1/2H20, and disserve in reagent water. Dilute to 1 L with reagent water;
7.6.2 Arsenic: Dissolve 1.320 g of arsenic tiioxide, ASjOa, or equivalent in 100 mL
of reagent water containing 4 g NaOM. Acidify the solution with 20 mL conc. HN03 and dilute
to 1 L with reagent water.
7.6.3 Barium: Dissolve 1.779 g barium chloride, BaC(2«2H20, in reagent water and
dilute to 1 L with reagent water.
7.6.4 Beryllium: Dissolve 11.659 g beryllium sulfate, BeS04, in reagent water
containing 2 mL nitric acid (conc.) and dilute to 1 L with reagent water.
7.6.5 Cadmium: Dissolve 1.000 g cadmium metal in 20 mL of 1:1 HNOa and dilute
to 1 L with reagent water.
7.6.6 Chromium: Dissolve 1.923 g of chromium tiioxide, Cr03, in reagent water,
acidify with redistilled HN03, and dilute to 1 L with reagent water.
7.6.7 Cobalt: Dissolve 1.000 g of cobalt metal in 20 mL of 1:1 HN03 and dilute to
1 Lwith reagent water. Chloride or nitrate salts of cobalt(li) may be used. Although numerous
hydrated forms exist, they are not recommended, unless the exact composition of the
compound is known.
7.6.8 Copper: Dissolve 1.000 g of electrolytic copper in 5 mL of redistilled HN03
and dilute to 1 L with reagent water.
7.6.9 Iron: Dissolve 1.000 g iron wire in 10 mL redistilled HN03 and reagent water
and dilute to 1 L with reagent water. Note that iron passivates in conc. HN03l and therefore
some water should be present.
7.6.10 Lead: Dissolve 1.599 g of lead nitrate, Pb(N03>2, in reagent water, acidify with
10 mL redistilled HN03, and dilute to 1 L with reagent water.
7.6.11 Manganese: Dissolve 1.000 g manganese metal In 10 mL redistilled HN03
and dilute to 1 L with reagent water.
7.6.12 Molybdenum: Dissolve 1.840 g of ammonium moJybdate, (NH^gMop^Hp,
and dilute to 1 L with reagent water.
7.6.13 Nickel: Dissolve 1.000 g nickel metal or 4.953 g nickel nitrate, Ni(N03)2*6H20
in 10 mL HN03 and dilute to 1 L with reagent water.
7.6.14 Selenium: Dissolve 0.345 g of selenious add (actual assay 94.6% H2SeOa)
or equivalent and dilute to 200 mL with reagent water.
NOTE: Due to the high toxicity of selenium, preparation of a smaller volume of reagent
has been described. Larger volumes may be prepared if required,
7.6.15 Silver Dissolve 1.575 g of anhydrous silver nitrate, AgNOa, in reagent water.
Add 10 mL of HNOa (conc.) and dilute to 1 L with reagent water. Because this standard is light
sensitive, store in a amber glass bottle in a refrigerator.
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7.6.16 Thallium: Dissolve 1.303 g thallium nitrate, TIN03, in reagent water, acidify
with 10 mL conc. HN03, and dilute to 1 L with reagent water.
7.6.17 Vanadium: Dissolve 1.785 g of vanadium pentoxide, V2Os, in 10 mL of conc.
HN03 and dilute to 1 L with reagent water.
7.6.18 Zinc: Dissolve 1.000 g zinc metal in 10 mL of conc. HNQj and dilute to 1 L
with reagent water.
7.7 Common matrix modifiers: The use of a palladium modifier is strongly recommended
for the determination of all analytes. This will correct for general chemical interferences as well as
allow for higher char and atomization temperatures without allowing the premature liberation of
anatyte. Other matrix modifiers may also be used as recommended by the instrument manufacturer
or when an interference is evident.
7.7.1 Palladium solution (Pd/Mg): Dissolve 300 mg of palladium powder in
concentrated HN03 (1 mL of HNOs, adding 0 .1 mL of conc. HCI, if necessary). Dissolve 200
mg of MgCNO^j in reagent water. Pour the two solutions together and dilute to 100 mL with
reagent water.
7.7.2 Nickel nitrate solution (5%): Dissolve 25g of NKNO^ei^O in reagent water
and dilute to 100 mL.
7.7.3 Nickel nitrate solution (1%): Dilute 20 mL of the 5% nickel nitrate solution to
100 mL with reagent water.
7.7.4 Ammonium phosphate solution (40%): Dissolve 40 g of ammonium
phosphate, (NH4)2HP04, in reagent water and dilute to 100 mL
7.7.5 Palladium chloride: Weigh 0.25 g of PdCIj to the nearest 0.0001 g and
dissolve in 10 mL of 1:1 HN03. Dilute to 1 L with reagent water
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 See Chapter Three.
9.0 QUALITY CONTROL
9.1 All quality control data should be maintained and available for easy reference or
inspection.
9.2 For each batch of samples processed, at least one method btank must be carried
throughout the entire sample preparation and analytical process as described in Chapter One. A
method blank is prepared by using a volume or weight of reagent water at the volume or weight
specified in the preparation method and then earned through the appropriate steps of the analytical
process. These steps may include but are not limited to digestion, dilution, filtering, and analysis.
If the method blank does not contain target analytes at a level that interferes with the project-specific
DQOs then the method blank would be considered acceptable, in the absence of project-specific
DQOs, if the blank is less than the MDL or less than 10% of the lowest sample concentration for
each anatyte, whichever is greater, then the method blank would be considered acceptable. If the
method blank cannot be considered acceptable, the method blank should be re-run once and if still
unacceptable, then all contaminated samples after the last acceptable method blank must be
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reprepped and reanalyzed along with the other appropriate batch QC samples. These blanks will be
useful in determining if samples are being contaminated. Refer to Chapter One for the proper
protocol when analyzing blanks.
9.3 For each batch of samples processed, at least one laboratory control sample must be
carried throughout the entire sample preparation and analytical process as described in Chapter
One. The laboratory control samples should be spiked with each anafyte of interest at the project-
specific action level or when lacking project-specific action levels, between the low and midlevel
standards. Acceptance criteria should be set at a laboratory derived limit developed through the use
of historical analyses. In the absence of historical data this limit should be set at ± 20% of the spiked
value. Alter the determination of historical data, ±20% must still be the limit of maximum deviation
to express acceptability. If the laboratory control sample cannot be considered acceptable* the
laboratory control sample should be re-run once and if still unacceptable then all samples after the
last acceptable laboratory control sample must be reprepped and reanalyzed. Refer to Chapter One
for more information.
9.4 Matrix Spike/Matrix Spike Duplicates (MS/MSDs): At the laboratory's discretion, a
separate spike sample and a separate duplicate sample may be analyzed in lieu of the MS/MSD. For
each batch of samples processed, at least one MS/MSD sample must be carried throughout the
entire sample preparation and analytical process as described in Chapter One. MS/MSDs are
intralaboratory split samples spiked with identical concentrations of each analyte of interest. The
spiking occurs prior to sample preparation and analysis. An MS/MSD is used to document the bias
and precision of a method in a given sample matrix. Refer to the definitions of bias and precision,
in Chapter One, forthe proper data reduction protocols. MS/MSD samples should be spiked at the
same bevel as the corresponding laboratory control sample that is at the project-specific action level
or, when lacking project-specific action levels, between the low and midlevel standards.
Acceptance criteria should be set at a laboratory derived limit developed through the use of historical
analyses. In the absence of historical data this limit should be set at ± 20% of the spiked value for
precision and <; 20 relative percent difference (RPD). After the determination of historical data, 20%
must still be the limit of maximum deviation for both percent recovery and relative percent difference
to expreiss acceptability. Refer to Chapter One for guidance. If the bias and precision indicators are
outside the laboratory control limits or if the percent recovery is less than 80% or greater than 120%
or if the relative percent difference is greater than 20%, the interference test as discussed in Sea
9.5.2 and 9.7 should be conducted.
9.5 Interference tests
9.5.1 Recovery test (post-digestion spike) - The recovery test must be done on
every sample. To conduct this test withdraw an aliquot of the test sample and add a known
amount of analyte to bring the concentration of the analyte to 2 to 5 times the original
concentration. If spiking at 2-5 times would exceed the linear range of the instrument, a lesser
spite may be used. If all of the samples in the batch have analyte concentrations below the
detection limit, spike the selected sample at the project-specific action level or when lacking
project-specific action levels, between the low and midlevel standards. Analyze the spiked
sample and calculate the spike recovery. If the recovery is <85% or >115%, MSA should be
used for the sample.
9.5.2 Dilution test - The dilution test is to be conducted when interferences are
suspected (Sec. 9.5.1) and the sample concentration is high enough to allow for proper
interpretation of the results. To conduct this test, determine the apparent concentration in the
undiluted sample. Dilute the sample by a minimum of five fold (1+4) and reanalyze.
Agreement within an RPD of 10 between the concentration forthe undiluted sample and five
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times the concentration for the diluted sample indicates the absence of interferences, and such
samples may be analyzed without using the method of standard additions. If agreement
between the dilutions is greater than 10%, the MSA should be used for all samples in the
batch.
9.6 Where the sample matrix is so complex that viscosity, surface tension, and components
cannot be accurately matched with standards, the method of standard addition (MSA) is
recommended (see Section 9.7 below). Other options including, the use of different matrix
modifiers, different furnace conditions, different preparatory methods or different analytical methods
may also be attempted to property characterize a sample. Section 9.5 provides tests to determine
the potential for an interference and evaluates the need for using the MSA.
9.7 Method of standard additions - The standard addition technique involves adding known
amounts of standard to one or more aliquots of the processed sample solution. This technique
attempts to compensate for a sample constituent that enhances or depresses the analyte signal,
thus producing a different slope from that of the calibration standards. It will not correct for additive
interferences which cause a baseline shift. The method of standard additions may be appropriate
for analysis of extracts, on analyses submitted as part of a delisting petition, whenever a new
sample matrix is being analyzed and on every batch that fails the recovery test
9.7.1 The simplest version of this technique is the single-addition method, in which
two identical aliquots of the sample solution, each of volume Vx, are taken. To the first (labeled
A) is added a known volume Vs of a standard analyte solution of concentration Gs. To the
second aliquot (labeled B) is added the same volume Vs of reagent water. The analytical
signals of A and i are measured and corrected for non-analyte signals. The unknown sample
concentration Cx is calculated:
c - S°VsC=
" (VS=)V
where SA and SB are the analytical signals (corrected for the blank) of solutions A and B,
respectively. V, and C» should be chosen so that SA is roughly twice SB on the average,
avoiding excess dilution of the sample. If a separation or concentration step is used, the
additions are best made first and carried through the entire procedure.
9.7.2 Improved results can be obtained by employing a series of standard additions.
To equal volumes of the sample are added a series of standard solutions containing different
known quantities of the analyte, and all solutions are diluted to the same final volume. For
example, addition 1 should be prepared so that the resulting concentration is approximately
50 percent of the expected absorbance from the indigenous analyte in the sample. Additions
2 and 3 should be prepared so that the concentrations are approximately 100 and 150 percent
of the expected endogenous sample absorbance. The absorbance of each solution is
determined and then plotted on the vertical axis of a graph, with the concentrations of the
known standards plotted on the horizontal axis. When the resulting line is extrapolated to zero
absorbance, the point of interception of the abscissa is the endogenous concentration of the
analyte in the sample. The abscissa on the left of the ordinate is scaled the same as on the
right side, but in the opposite direction from the ordinate. An example of a plot so obtained is
shown in Figure 1. A linear regression program may be used to obtain the intercept
concentration.
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9.7.3 For the results of this MSA technique to be valid, the following limitations must
be taken into consideration:
1. The apparent concentrations from the calibration curve must be linear (0.995
or greater) over the concentration range of concern. For the best results, the
slope of the MSA plot should be nearly the same as the slope of the standard
curve.
2. The effect of the interference should not vary as the ratio of analyte
concentration to sample matrix changes, and the standard addition should
respond in a similar manner as the analyte.
3. The determination must be free of spectral interference and corrected for
nonspecific background interference.
9.8 All qualify control measures described in Chapter One should be followed.
9.9 Independent source laboratory control sample or standard reference materials (SRMs)
should be used to help assess the quality of the analytical scheme.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Calibration standards - All analyses require that a calibration curve be prepared to
cover the appropriate concentration range. Usually, this means the preparation of a blank and
standards which produce an absorbance of 0.0 to 0.7. Calibration standards can prepared by
diluting the stock metal solutions in the same acids and acid concentrtions as the samples.
10.1.1 Calibration standards can be prepared fresh each time a batch of samples
is analyzed. If the ICV solution is prepared daily and the ICV is analyzed within the acceptance
criteria, calibration standards do not need to be prepared daily and may be prepared and
stored for as long as the calibration standard viability can be verified through the use of the
ICV. If the ICV is outside of the acceptance criteria, the caSbrat'on standards must be prepared
fresh and the instrument recalibrated. Prepare a blank and at least three calibration standards
in graduated amounts in the appropriate range of the linear part of the curve.
10.1.2 The calibration standards should be prepared using the same type of acid or
combination of acids and at the same concentration as will result in the samples following
processing. ^ ?
10.1.3 Beginning with the blank and working toward the highest standard, inject the
solutions and record the readings. Calibration curves are always required.
10.2 A calibration curve must be prepared each day with a minimum of a calibration blank
and three standards. The curve must be linear and have a correlation coefficient of at least 0.995.
10.2.1 After initial calibration, the calibration curve must be verified by use of an
initial calibration blank (ICB) and an initial calibration verification (ICV) standard. The ICV
standard must be made from an independent (second source) material at or near mid-range.
The acceptance criteria for the ICV standard must be ±10% of its true value and the ICB must
not contain target analytes at or above the MDL for the curve to be considered valid. If the
calibration curve cannot be verified within the specified limits, the cause must be determined
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and the instrument recalibrated before samples are analyzed. The analysis data for the ICV
must be kept on file with the sample analysis data.
10.2.2 The calibration curve must also be verified at the end of each analysis batch
and/or after every 10 samples by use of a continuing calibration blank (CCB) and a continuing
calibration verification (CCV) standard. The CCV standard should be made from the same
material as the initial calibration standards at or near midrange. The acceptance criteria for
the CCV standard must be ±10% of its true value and the CCB must not contain target
analytes at or above the MDL fort he curve to be considered valid. If the calibration cannot be
verified within the specified limits, the sample analysis must be discontinued, the cause
determined and the instrument recalibrated. All samples following the last acceptable
CCV/CCB must be reanalyzed. The analysis data for the CCV/CCB must be kept on file with
the sample analysis data.
10.3 It is recommended that each standard should be analyzed (injected) twice and an
average value determined. Replicate standard values should be within ±10% RPD.
10.4 Standards are run in part to monitor the life and performance of the graphite tube. Lack
of reproducibility or significant change in the signal for the standard indicates that the tube should
be replaced. Tube life depends on sample matrix and atomization temperature. A conservative
estimate would be that a tube will last at least 50 firings. A pyrolytic coating will extend that estimated
life by a factor of three.
10.5 If conducting trace analysis, it is recommended that the lowest calibration standard be
set at the laboratory's quantitation level. The laboratory can use a reporting limit that is below the
quantitation level but all values reported below the low standard should be reported as estimated
values.
11.0 PROCEDURE
11.1 Preliminary treatment of waste water, ground water, extracts, and industrial waste is
always necessary because of the complexity and variability of sample matrices. Solids, slurries, and
suspended material must be subjected to a solubilization process before analysis. This process may
vary because of the metals to be determined and the nature of the sample being analyzed.
Solubilization and digestion procedures are presented in Chapter Three. Samples which are to be
analyzed only for dissolved constituents need not be digested if they have been filtered and acidified.
11.2 Furnace devices (flameless atomization) are a most useful means of extending
detection limits. Because of differences between various makes and models of satisfactory
instruments, no detailed operating instructions can be given for each instrument. Instead, the analyst
should follow the instructions provided by the manufacturer of a particular instrument. A generalized
set of instructions follows:
11.2.1 Inject an aliquot of sample into the furnace and atomize. If the concentration
found is greater than the highest standard, the sample should be diluted in the same add
matrix and reanalyzed. The use of multiple injections can improve accuracy and help detect
furnace pipetting errors.
11.2.2 To verify the absence of interference, follow the interference procedure given
in Sec. 9.5.
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12.0 DATA ANALYSIS AND CALCULATIONS
12.1 For determination of metal concentration by GFAA: Read the metal value from the
calibration curve or directly from the read-out system of the instrument.
12.1.1 If dilution of sample was required:
|jg/L metal in sample = —
w
where:
A = \}g!L of metal in diluted aliquot from calibration curve.
B = Starting sample volume, mL.
C = Final volume of sample, mL.
12.1.2 For solid samples, report all concentrations in consistent units based on wet
weight. Ensure that if the dry weight was used for the analysis, percent solids should be
reported to the client. Hence:
A V V
mg metal/kg sample =
where:
A = mg/L of metal in processed sample from calibration curve.
V = Final volume of the processed sample, L.
W = Weight of sample, Kg.
12.1.3 Different injection volumes must not be used for samples and standards.
Instead, the sample should be diluted and the same size injection volume be used for both
samples and standards.
13.0 METHOD PERFORMANCE
13.1 See the individual methods from reference 1.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity
and/or toxicity of waste at the point of generation. Numerous opportunities for pollution prevention
exist in laboratory operation. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of first choice.
Whenever feasible, laboratory personnel should use pollution prevention techniques to address their
waste generation. When wastes cannot be feasibly reduced at the source, the Agency recommends
recycling as the next best option.
7010 -14
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14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Less is Better: Laboratory Chemical Management for Waste Reduction
available from the American Chemical Society, Department of Government Relations and Science
Policy, 1155 16th Street, NW, Washington, DC 20036, (202) 872-4477.
15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society at the address listed in Section 14.2.
16.0 REFERENCES
1- Methods for Chemical Analysis of Water and Wastes: U.S. Environmental Protection Agency.
Office of Research and Development. Environmental Monitoring and Support Laboratory. ORD
Publication Offices of Center for Environmental Research Information: Cincinnati, OH, 1983;
EPA-600/4-79-020.
2. Rohrbough, W.G.; et al. Reagent Chemicals. American Chemical Society Specifications. 7th
ed.; American Chemical Society: Washington, DC, 1986.
3. 1985 Annual Book of ASTM Standards. Vol. 11.01; "Standard Specification for Reagent
Water"; ASTM: Philadelphia, PA, 1985; D1193-77,
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 and 2, Figure 1, and a flow diagram of the method
procedures.
7010-15
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TABLE 1
FURNACE ATOMIC ABSORPTION DETECTION LIMITS FOR ANALYTES
IN REAGENT WATER
Furnace Procedure*''
Detection Limit
Metal (pg/L)
Antimony
3
Arsenic
1
iarium(p)
2
Beryllium
0.2
Cadmium
0.1
Chromium
1
Cobalt
1
Copper
1
Iron
1
Lead
1
Manganese
0.2
Molybdenum(p)
1
Nickel
1
Selenium
2
Silver
0.2
Thallium
1
Vanadium(p)
4
Zinc
0.05
NOTE: The symbol (p) indicates the use of pyrolytic graphite with the furnace
procedure.
•For furnace sensitivity values, consult instrument operating manual.
The listed furnace values are those expected when using a 20-pL injection and normal
gas flow, except in the cases of arsenic and selenium, where gas interrupt is used.
Source: Reference 1.
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TABLE 2
INSTRUMENT PARAMETERS
I ELEMENT
WAVELENGTH (nm)
PURGE GAS1
COMMENTS
Sb
217.5,
231.1
argon or nitrogen

As
193.7
argon

ia
553.6
argon
nitrogen should not
be used
Be
234.9
argon

Cd
228.8
argon

Cr
357.9
argon
nitrogen should not
be used
Co
240.7
anon

Cu
324.7
argon or nitrogen

Fe
?48.3,
248.8,271.8,
302.1,252.7
argon or nitrogen

Pb
283.3,
217.0
argon

Mn
279-5,
403.1
argon or nitrogen

Mo
313.3
argon
nitrogen should not
be used
Ni
232.0,
352.4
argon or nitrogen

Se
196.0
argon 4...

Ag
328.1
argon :

Tl
276.8
arpon or nitrogen

V
318.4
argon
nitrogen should not
be used
Zn
213.9
argon or nitrogen

Note: if more than one wavelength is listed, the primary line is underlined.
1The argon/H, purge gas is also applicable.
Source: Reference 1
7010-17
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FIGURE 1
STANDARD ADDITION PLOT
Cone, of
Sample
AddnO
No Addn
Addn 2
Addn of 100% Addn of 150%
Addn 3
Addn 1
Addn of 50%
of Expected of Expected of Expected
Amount Amount Amount
7010-18
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METHOD 7010
GRAPHITE FURNACE ATOMIC ABSORPTION SPECTROPHOTOMETRY
concentration >
highest
. standard? ,
Yes
No
Stop
12.0 Determine
concentrations.
11.1 Solubilize and
digest sample (See
Chapter 3.)
11.2.1 Inject and
atomize an aliquot
of sample.
11.2.1 Dilute
sample.
11.2 Follow operating
instructions from instrument
manufacturer.
11.2.2 Fallow the
interference procedure
of Sec. 9.5 to verify the
absence of interference
7010-19
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METHOD 7471B
MERCURY IN SOLID OR SEMISOLID WASTE (MANUAL COLD-VAPOR TECHNIQUES
1.0 SCOPE AND APPLICATION
1.1 Method 7471 is approved for measuring total mercury (organic and inorganic) in soils,
sediments, bottom deposits, and sludge-type materials. All samples must be subjected to an
appropriate dissolution step prior to analysis. If this dissolution procedure is not sufficient to dissolve
a specific matrix type or sample, then this method is not applicable for that matrix.
2.0 SUMMARY OF METHOD
2.1 Prior to analysis, the solid or semi-solid samples must be prepared according to the
procedures discussed in this method.
2.2 This method is a cold-vapor atomic absorption method and is based on the absorption
of radiation at the 253.7-nm wavelength by mercury vapor. The mercury is reduced to the elemental
state and aerated from solution in a closed system. The mercury vapor passes through a cell
positioned in the light path of an atomic absorption spectrophotometer. Absorbance (peak height)
is measured as a function of mercury concentration.
2.3 The typical instrument detection limit (IDL) for this method is 0.0002 mg/L.
3.0 DEFINITIONS
Refer to Chapter One and Chapter Three for a listing of applicable definitions.
4.0 INTERFERENCES
4.1 Potassium permanganate is added to eliminate possible interference from sulfide.
Concentrations as high as 20 mg/Kg of sulfide, as sodium sulfide, do not interfere with the recovery
of added inorganic mercury in reagent water.
4.2 Copper has also been reported to interfere; however, copper concentrations as high
as 10 mg/Kg had no effect on recovery of mercury from spiked samples.
4.3 Samples high in chlorides require additional permanganate (as much as 25 mL)
because, during the oxidation step, chlorides are converted to free chlorine, which also absorbs
radiation of 254 nm. Care must therefore be taken to ensure that free chlorine is absent before the
mercury is reduced and swept into the cell. This may be accomplished by using an excess of
hydroxylamine sulfate reagent (25 mL). In addition, the dead air space in the BOD bottle must be
purged before adding stannous sulfate.
4.4 Certain volatile organic materials that absorb at this wavelength may also cause
interference. A preliminary run without reagents should determine if this type of interference is
present.
5.0 SAFETY
Refer to Chapter Three for a discussion on safety related references and issues.
7471B -1
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6.0 EQUIPMENT AND SUPPLIES
6.1 Atomic absorption spectrophotometer or equivalent: Any atomic absorption unit with
an open sample presentation area in which to mount the absorption cell is suitable. Instrument
settings recommended by the particular manufacturer should be followed. Instruments designed
specifically for the measurement of mercury using the cold-vapor technique are commercially
available and may be substituted for the atomic absorption spectrophotometer.
6.2 Mercury hollow cathode lamp or electrodeless discharge lamp.
6.3 Recording device: Any multirange variable-speed recorder compatible with the UV
detection system or any other compatible data collection device.
6.4 Absorption cell: Standard spectrophotometer cells 10 cm long with quartz end windows
may be used. Suitable cells may be constructed from Plexlglas tubing, 1 in. O.D. x 4.5 in. The ends
are ground perpendicular to the longitudinal axis, and quartz windows (1 in. diameter x 1/16 in.
thickness) are cemented in place. The cell is strapped to a burner for support and aligned in the light
beam by use of two 2-in. x 2-in. cards. One inch diameter holes are cut in the middle of each card.
The cards are then placed over each end of the cell. The cell is then positioned and adjusted
vertically and horizontally to give the maximum transmittance.
6.5 Air pump: Any peristaltic pump capable of delivering 1 L/min air may be used. A
Masterflex pump with electronic speed control has been found to be satisfactory.
6.6 Flowmeter: Capable of measuring an air flow of 1 L/min.
6.7 Aeration tubing: A straight glass frit with a coarse porosity. Tygon tubing is used for
passage of the mercury vapor from the sample bottle to the absorption cell and return.
6.8 Drying tube: 6-in. x 3/4-in. diameter tube containing 20 g of magnesium perchiorate
or a small reading lamp with 60-W bulb which may be used to prevent condensation of moisture
inside the cell. The lamp should be positioned to shine on the absorption cell so that the air
temperature in the cell is about 10°C above ambient.
6.9 The cold-vapor generator is assembled as shown in Figure 1 of reference 1 or
according to the instrument manufacturers instructions. The apparatus shown in Figure 1 is a closed
system. An open system, where the mercury vapor is passed through the absorption cell only once,
may be used instead of the closed system. Because mercury vapor is toxic, precaution must be
taken to avoid its inhalation. Therefore, a bypass has been included in the system either to vent the
mercury vapor into an exhaust hood or to pass the vapor through some absorbing medium, such as:
1. Equal volumes of 0.1 M KMn04 and 10% H2S04, or
2. Iodine 0.25% in a 3% Kl solution.
A specially treated charcoal that will adsorb mercury vapor is also available from Bameby and
Cheney, East 8th Avenue and North Cassidy Street, Columbus, Ohio 43219, Cat. #580-13 or #580-
22.
6.10 Heating source - Adjustable and capable of maintaining a temperature of 95 ± 3°C. (e.g.,
hot plate, block digestor, microwave, etc.)
6.11 Graduated cylinder or equivalent volume measuring device.
7471B-2
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7.0 REAGENTS AND STANDARDS
7.1 Reagent Water. Reagent water will be interference free. All references to water in this
method refer to reagent water unless otherwise specified. Refer to Chapter One for a definition of
reagent water.
7.2 Aqua regia: Prepare immediately before use by carefully adding three volumes of
concentrated HCI to one volume of concentrated HN03.
7.3 Sulfuric add, 0.5 N: Dilute 14.0 mL of concentrated sulfuric acid to 1 liter.
7.4 Stannous sulfate: Add 25 g stannous sulfate to 250 mL of 0.5 N sulfuric add. This
mixture is a suspension and should be stirred continuously during use. A 10% solution of stannous
chloride can be substituted for stannous sulfate.
7.5 Sodium chloride-hydroxylamine sulfate solution: Dissolve 12 g of sodium chloride and
12 g of hydroxylamine sulfate in reagent water and dilute to 100 mL. Hydroxylamine hydrochloride
may be used in place of hydroxylamine sulfate.
7.6 Potassium permanganate, mercury-free, 5% solution (w/v): Dissolve 5 g of potassium
permanganate in 100 mL of reagent water.
7.7 Mercury stock solution: Dissolve 0.1354 g of mercuric chloride in 75 mL of reagent
water. Add 10 mL of concentrated nitric add and adjust the volume to 100.0 mL (1.0 mL = 1.0 rug
Hg).
7.8 Mercury working standard: Make successive dilutions of the stock mercury solution to
obtain a working standard containing 0.1 Mg/mL. This working standard and the dilution of the stock
mercury solutions should be prepared fresh daily. Acidity of the working standard should be
maintained at 0.15% nitric add. This add should be added to the flask, as needed, before adding
the aliquot.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 All samples must have been collected using a sampling plan that addresses the
considerations discussed in Chapter Nine of this manual.
8.2 All sample containers must be prewashed with detergents, adds, and reagent water.
Plastic and glass containers are both suitable.
8.3 Non-aqueous samples shall be refrigerated, when possible, and analyzed "as soon as
possible."
i.O QUALITY CONTROL
Refer to Method 7000.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Standard preparation: Transfer 0.0-, 0.5-, 1.0-, 2.0-, 5.0-, and 10-mL aliquots of the
mercury working standard, containing 0-1.0 ug of mercury, to a series of 300-mL BOD bottles or
equivalent. Add enough reagent water to each bottle to make a total volume of 10 mL. Add 5 mL
7471B- 3
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of aqua regia and heat 2 min at 95 ± 3°C. Allow the sample to cool; add 50 mL reagent water and
15 mL of KMn04 solution to each bottle and heat again at 95 ± 3°C for 30 min. Cod and add 6 mL
of sodium chloride-hydroxylamine sulfate solution to reduce the excess permanganate. Add 50 mL
of reagent water. Treating each bottle individually, add 5 mL of stannous sulfate solution,
immediately attach the bottle to the aeration apparatus, and continue as described in Section 11.3.
10.2 Construct a calibration curve by plotting the absortoances of standards versus
micrograms of mercury. Determine the peak height of the unknown from the chart or other recording
device and read the mercury value from the standard curve. Duplicates, spiked samples, and check
standards should be routinely analyzed.
11.0 PROCEDURE
11.1 Sample preparation: Weigh a 0.5 - 0.6g aliquot of a well homogenized sample and
place in the bottom of a BOD bottle or other appropriate analysis vessel. Add 5 mL of reagent water
and 5 mL of aqua regia. Heat two minutes at 95 ± 3°C. Cool; then add 50 mL reagent water and
15 mL potassium permanganate solution to each sampte. Mix thoroughly, then heat for 30 min at
95 ± 3°C. Cool and add 6 mL of sodium chloride-hydroxylamine sulfate to reduce the excess
permanganate.
CAUTION: Do this addition under a hood, as Cl2 could be evolved. Add 55 mL of reagent
water. Treating each bottle individually, add 5 mL of stannous sulfate and immediately attach
the bottle to the aeration apparatus. Continue as described under Section 11.3.
11.2 An alternate digestion procedure employing an autoclave may also be used. In this
method, 5 mL of concentrated H2S04 and 2 mL of concentrated HN03 are added to the 0.5 - 0.6 g
of sample. Add 5 mL of saturated KMn04 solution and cover the bottle with a piece of aluminum foil.
The samples are autoclaved at 121 ± 3°C and 15 lb for 15 min. Cool, dilute to a volume of 100 mL
with reagent water, and add 6 mL of sodium chloride-hydroxylamine sulfate solution to reduce the
excess permanganate. Purge the dead air space and continue as described under Section 11.3.
Refer to the caution statement in Section 11.1 for the proper protocol in reducing the excess
permanganate solution and adding stannous sulfate.
11.3 Analysis: At this point, the sample is allowed to stand quietly without manual agitation.
The circulating pump, which has previously been adjusted to a rate of 1 L/min, is allowed to run
continuously. The absorbance, as exhibited either on the spectrophotometer or the recorder, mil
increase and reach maximum within 30 seconds. As soon as the absorbance reading levels off
(approximately 1 minute), open the bypass valve and continue the aeration until the absorbance
returns to its minimum value. Close the bypass valve, remove the fritted tubing from the BOD bottle,
and continue the aeration.
12.0 DATA ANALYSIS AND CALCULATIONS
12.1 Calculate metal concentrations: (1) by the method of standard additions, (2) from a
calibration curve, or (3) directly from the instrument's concentration read-out. All dilution or
concentration factors must be taken into account. Concentrations reported for muitiphased or wet
samples must be appropriately qualified (e.g., 5 pg/g dry weight).
13.0 METHOD PERFORMANCE
13.1 Precision and accuracy data are available in Method 245.5 of Methods for Chemical
Analysis of Water and Wastes.
7471B - 4
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13.2 The data shown in Table 1 were obtained from records of state and contractor
laboratories. The data are intended to show the precision of the combined sample preparation and
analysis method.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity
and/or toxicity of waste at the point of generation. Numerous opportunities for pollution prevention
exist in laboratory operation. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of first choice.
Whenever feasible, laboratory personnel should use pollution prevention techniques to address their
waste generation. When wastes cannot be feasibly reduced at the source, the Agency recommends
recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Less is Better: Laboratory Chemical management for Waste Reduction
available from the American Chemical Society's Department of Government Relations and Science
Policy, 1155 16th St., N.W. Washington, D C. 20036, (202) 872-4477.
15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society at the address listed in Sec. 14.2.
16.0 REFERENCES
1. Methods for Chemical Analysis of Water and Wastes, EPA-600/4-82-055, December 1982,
Method 245.5.
2. Gaskill, A., Compilation and Evaluation of RCRA Method Performance Data, Work
Assignment No. 2, EPA Contract No. 68-01-7075, September 1986.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Table 1 and a flow diagram of the method procedure.
7471B - 5
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TABLE 1
METHOD PERFORMANCE DATA
Sample Preparation Laboratory
Matrix Method Replicates
Emission control dust Not known 12, 12 yg/g
Wastewater treatment sludge Not known 0.4, 0.28 pg/g
7471B - 6
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METHOD 7471B
MERCURY IN SOUP OR SEMISOLID WASTE fMANUAL COLD-VAPOR TECHNIQUES
Stilt
Standard Preparation
Sample Preparation

Type 1
7.6 Calculate
metal
concentration.
7.3 Add
stannous sulfate,
attach to aeration
apparatus.
7.3 Transfer alt quota
of He working
standards to
bottles.
7.1 Heat. cool,
add sodium
chloride-
hydro xyiomino
sulfate.
7.3 Add reagent
water and KMnQ
solution, heat
and cool.
7.3 Add sodium
chloride
hydroxyiamine
sulfate and
reagent water.
7.6 Construct
calibration curve,
determine peak
height and Hg value.
7.1 Add reagent
water, stannous
sulfate, attach to
aeration apparatus.
7.2 Add sodium
chloride-
hyd roxyiomrno
sulfate, purge
dead air space.
7.1 Heat, cool,
add reagent water
and KMnO^.
7.3 Add reagent
water to-volume,
and aqua regia
heat and cool.
7.1 Weigh cample
end reagent water
and aqua rag la.
7.2 Add
KMnQ) cover,
heat and cool,
dilute with
raagant water.
7471B - 7
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METHOD 7473
MERCURY IN SOLIDS AND SOLUTIONS BY THERMAL DECOMPOSITION. AMALGAMATION
AND ATOMIC ABSORPTION SPECTROPHOTOMETRY
1.0 SCOPE AND APPLICATION
1.1 Method 7473 is designated for the determination of mercury (CAS No. 7439-97-6)
in solids, aqueous samples, and digested solutions in both the laboratory arid field environments.
Integration of thermal decomposition sample preparation and atomic absorption detection reduces
the total analysis time of most samples to less than five minutes in either the laboratory or field
setting. Total mercury (organic and inorganic) in soils, sediments, bottom deposits, and sludge-
type materials as well as in aqueous wastes and ground waters can be determined without sample
chemical pretreatment using this method, except as noted. Alternatively, this method can be used
for the detection of total mercury from total decomposition sample preparation methods, such as
Method 3052, or for detection of extracted or leached mercury compounds or species from
methods such as the SW-846 3000 series methods (as detailed in Chapter Three).
NOTE: For unique circumstances when mercury could be bound in silicates or other
matrices that may not thermally decompose, validation of direct analysis of the solid should
be confirmed with total decomposition with an EPA approved method (such as Method
3052) and analysis with this method.
2.0 SUMMARY OF METHOD
2.1 Controlled heating in an oxygenated decomposition furnace is used to liberate
mercury from solid and aqueous samples in the instrument. The sample is dried and then
thermally and chemically decomposed within the decomposition furnace. The decomposition
products are earned by flowing oxygen to the catalytic section of the furnace. Here oxidation is
completed and halogens and nitrogen/sulfur oxides are trapped. The remaining decomposition
products are then carried to an amalgamator that selectively traps mercury. After the system is
flushed with oxygen to remove any remaining gases or decomposition products, the amalgamator
is rapidly heated, releasing mercury vapor. Flowing oxygen carries the mercury vapor through
absorbance cells positioned in the light path of a single wavelength atomic absorption
spectrophotometer. Absorbance (peak height or peak area) is measured at 253.7 nm as a function
of mercury concentration.
2.2 The typical working range for this method is 0.05 - 600 ng. The mercury vapor is
first carried through a long pathlength absorbance cell and then a short pathlength absorbance cell.
(The lengths of the first cell and the second cell are in a ratio of 10:1 or another appropriate ratio.)
The same quantity of mercury is measured twice, using two different sensitivities (see Figure 1),
resulting in a dynamic range that spans at least four orders of magnitude.
2.3 The instrument detection limit (IDL) for this method is 0.01 ng total mercury.
3.0 DEFINITIONS
3.1 Thermal Decomposition: Partial or complete degradation of sample components
using convection and conduction heating mechanisms resulting in the release of volatile
components such as water, carbon dioxide, organic substances, elements in the form of oxides
or complex compounds, and elemental gases.
7473 -1
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3.2 Amalgamation: The process by which mercury forms a metal alloy with gold.
3.3 Amalgamator: A system composed of gold particles at a high surface area to
volume ratio for the purpose of amalgamating mercury vapor.
3.4 Primary Calibration: A complete calibration of the instrument's working range. This
calibration is performed initially and when any significant instrumental parameters are changed.
For example, in this method a primary calibration should be performed after the decomposition
tube, amalgamator, or oxygen tank is replaced.
3.5 Daily Calibration: A calibration performed with minimal standards to ensure that the
primary calibration is valid. For example, when two standards within the range of interest are
analyzed and agree within 10% of their true value the primary calibration is assumed to be valid.
3.6 Memory Effects: Mercury vapor may remain in the decomposition tube,
amalgamator, or absorbance cells and be released in a subsequent analysis resulting in a positive
bias. For example, this may result when a low concentration sample is analyzed after a sample
of high mercury content.
3.7 Sample Boat: The non-amalgamating thermally stable vessel used for containment
and transport of the solid or liquid sample for thermal decomposition.
4.0 INTERFERENCES
4.1 In areas where mercury contamination is an existing problem, the background signal
may be significantly increased.
4.2 Memory effects between analyses may be encountered when analyzing a sample
of high mercury concentration (> 400 ng) prior to analyzing one of low concentration (s 25 ng).
Typically, to minimize memory effects, analyze the samples in batches of low and high
concentrations, always analyzing those of low concentration first. If this batching process cannot
be accomplished, , a blank analysis with an extended decomposition time may be required following
the analysis of a highly concentrated sample to limit memory effects.
4.3 Co-absorbing gases, such as free chlorine and certain organics (as indicated in
Methods 7470 and 7471), should not interfere due to the release of decomposition products by the
decomposition furnace, removal of some decomposition products by the decomposition catalyst,
and the selective entrapment of mercury vapor on the amalgamator.
5.0 SAFETY
5.1 Refer to Chapter Three for a discussion on safety related references and issues.
5.2 Many mercury compounds are highly toxic if swallowed, inhaled, or absorbed
through the skin. Extreme care must be exercised in the handling of concentrated mercury
reagents. Concentrated mercury reagents should only be handled by analysts knowledgeable of
their risks and of safe handling procedures.
6.0 EQUIPMENT AND SUPPLIES
6.1 The working scheme of the mercury analysis system is illustrated in Figure 2. The
sample introduction device consists of a motorized support with a metal or metal alloy sample boat
7473 - 2
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that is appropriate for solids and liquids. An example of an appropriate boat would be made of
nickel with a liquid capacity of 0.5 -1.0 mL. Once the sample is either manually or automatically
dispensed into the sample boat, the boat is mechanically introduced automatically into a quartz
decomposition tube. The decomposition tube is heated by two independently programmable ovens,
the decomposition and catalyst furnaces, each furnace is capable of maintaining a temperature of
at least 750°C. The sample is dried and thermally decomposed in an oxygen environment,
releasing mercury vapor. The mercury vapor is transported by oxygen over the amalgamator that
traps the mercury. Once the sample is completely decomposed the trapped mercury is desorbed
rapidly by heating the amalgamator with the mercury release furnace. The mercury vapor passes
through two absorbance cuvette, in series, that are separated by a collection flask outside the
optical axis. The flow path through the spectrometer and cuvettes is maintained at approximately
120°C, by a heating unit, to prevent condensation and minimize carry-over effects. A mercury
vapor lamp is used as the light source. The detector is connected to a computer for data
acquisition and analysis.
6.2 The DMA 80 automatic mercury analyzer (Milestone, Inc.) is the instrument used
for the scheme outlined above. It has been tested for use with this method. Other instruments
based on these principles may also be appropriate.
6.3 This mefriod is not limited to mercury vapor generation by thermal decomposition.
Alternatively, other mercury vapor introduction systems, such as mercury cold vapor generation,
may be appropriate. Alternative sample introduction apparatus may be applied after validation with
data similar to those in Tables 1 and 2.
6.4 This method is not limited to analyzing total mercury content. This detection
scheme can be used for analysis of individual species of mercury that have been separated by an
appropriate method or instrument system.
7.0 REAGENTS AND STANDARDS
7.1 Reagent water Reagent water will be interference free. All references to water in
this method refer to reagent water unless otherwise specified,
7.2 High purity oxygen gas: High purity oxygen should be interference and mercury free.
If the oxygen is possibly contaminated with mercury vapor, a gold mesh filter should be inserted
between the gas cylinder and the mercury analysis instrument to prevent any mercury from entering
the instrument.
7.3 Mercury stock solution: Dissolve 0.1354 g of mercuric chloride in 75 mL of reagent
water. Add 10 mL of concentrated nitric acid and adjust the volume to 100.0 mL (1.0 mL = 1.0 mg
Hg). Stock solutions may also be purchased. Verify the quality of the standard by checking it
against a second source standard (Sec. 9.2).
7.4 Mercury working standards: Make successive dilutions of the stock mercury solution
to obtain standards containing 100 ppm and 10 ppm. For calibration of the high range, standards
of 0,1, 2, 3,4, 5, and 6 ppm are recommended. These are prepared by dilution of the 100 ppm
standard. For calibration of the low range, standards of 0.00, 0,05, 0.1,0.2, 0.3, 0.4, and 0.5 ppm
are recommended. These are prepared by dilution of the 10 ppm standard. A blank calibration
solution is also used for a zero calibration. Acidity of the working standards should be maintained
at least 0.15% nitric acid, as also recommended in Methods 7470 and 7471.
7473 - 3
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NOTE: The concentrations listed above are only recommended concentrations. The
concentration of the working standards may need adjustment according to specific
instrumental working ranges and/or manufactures" recommendations.
NOTE: The stability of the mercury standards is limited to 24 - 48 hours. Fresh mercury
standards must be prepared daily.
7.5 Standard reference material: In place of aqueous mercury standards, solid
reference material with a certified value for mercury may by used for calibration.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 All samples should be collected using a sampling plan that addresses the
considerations discussed in Chapter Nine of this manual.
8.2 All sample containers must be prewashed with detergents, acids, and reagent
water. Glass, plastic, and PTFE containers are suitable in most cases. Polymers are not suitable
for samples containing metallic mercury.
8.3 Metallic mercury, some inorganic mercury compounds, and many organic mercury
compounds are volatile and unstable. It is advantageous to analyze the samples as soon as
possible to determine the total mercury in the sample but in no cases exceed the 28-day limit as
defined in Chapter Three of this manual. Non-aqueous samples shall be analyzed as soon as
possible. If solid samples are not analyzed immediately, refrigeration is necessary.
9.0 QUALITY CONTROL
9.1 All quality control data should be maintained and available for easy reference or
inspection.
9.2 If more than 10 samples per day are analyzed, the working standard curve must be
verified by measuring satisfactorily a mid-range standard or reference standard after every 10
samples. This sample value must be within 20% of the true value, or the previous 10 samples
must be reanalyzed.
9.3 Matrix Spike/Matrix Spike Duplicates (MS/MSDs): At the laboratory's discretion, a
separate spike sample and a separate duplicate sample may be analyzed in Jieu of the MS/MSD.
For each batch of samples processed, at least one MS/MSD sample must be carried throughout
the entire sample preparation and analytical process as described in Chapter One. MS/MSDs are
intralaboratory split samples spiked with identical concentrations of each .analyte of interest. The
spiking occurs prior to sample preparation and analysis. An MS/MSD is used to document the bias
and precision of a method in a given sample matrix. Refer to the definitions of bias and precision,
in Chapter One, for the proper data reduction protocols. MS/MSD samples should be spiked at the
same level as the corresponding laboratory control sample that is at the project-specific action level
or, when lacking project-specific action levels, between the low and midlevel standards.
Acceptance criteria should be set at a laboratory derived limit developed through the use of
historical analyses. In the absence of historical data this limit should be set at ± 20% of the spiked
value for precision and s 20 relative percent difference (RPD). After the determination of historical
data, 20% must still be the limit of maximum deviation for both percent recovery and relative
percent difference to express acceptability. Refer to Chapter One of this manual for guidance.
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9.4 For each batch of samples processed, at least one method blank must be carried
throughout the entire sample preparation and analytical process as described in Chapter One. A
method blank is prepared by using a volume or weight of reagent water at the volume or weight
specified in the preparation method and then carried through the appropriate steps of the analytical
process. These steps may include but are not limited to digestion, dilution, filtering, and analysis.
If the method blank does not contain target analytes at a level that interferes with the project-
specific DQOs then the method blank would be considered acceptable. In the absence of project-
specific DQOs, if the blank is less than the MDL or less than 10% of the lowest sample
concentration for each analyte, whichever is greater, then the method blank would be considered
acceptable. If the method blank cannot be considered acceptable, the method blank should be re-
run once and if still unacceptable then all samples after the last acceptable method blank must be
reprepped and reanalyzed along with the other appropriate batch QC samples. These blanks will
be useful in determining if samples are being contaminated. Refer to Chapter One for the proper
protocol when analyzing blanks.
9.5 For each batch of samples processed, at least one laboratory control samples must
be carried throughout the entire sample preparation and analytical process as described in
Chapter One. The laboratory control samples should be spiked with each analyte of interest at the
project-specific action level or when lacking project-specific action levels, between the low and
midlevel standards. Acceptance criteria should be set at a laboratory derived limit developed
through the use of historical analyses. In the absence of historical data this limit should be set at
± 20% of the spiked value. After the determination of historical data, ±20% must still be the limit
of maximum deviation to express acceptability. If the laboratory control sample cannot be
considered acceptable, the laboratory control sample should be re-ran once and if still
unacceptable then all samples after the last acceptable laboratory control sample must be
reprepped and reanalyzed. Refer to Chapter One for more information.
9.6 The method of standard additions can be used to verify linearity or if matrix
interference is suspected. Refer to Method 7000 for standard addition procedures.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Primary calibration: 100 jjL of a working standard is dosed onto the sample boat.
Analytical parameters for drying, decomposition, and wait times as recommended by the
manufacturer are chosen for the analysis (Section 11.1). Each standard solution is analyzed twice.
For the DMA 80, parameters of 70 seconds drying, 100 seconds decomposition, and 40 seconds
wait times (abbreviated 70/100/40) would be chosen for each standard analysis. Typical calibration
curves obtained in laboratory conditions are illustrated in Figures 3a and b and a calibration curve
obtained in field conditions is illustrated in Figure 4. Conduct curve using standards described in
Section 7.4.
10.2 Daily calibration: At least a high and low concentration standard for each working
range is analyzed using the analytical parameters as recommended by the manufacturer. The
woriting calibration standards must be measured within 10% of their true value for the curve to be
considered valid.
10.3 An alternative calibration using standard reference materials (SRMs) may be used.
In this method, an amount of the reference material is weighed (accurate to ± 0.001 g or better)
onto a tared sample boat. The analytical parameters chosen are based on the weight, moisture
content, and organic content of the soil and should be as similar to the matrix of interest as
possible (refer to Sec. 11.1). This procedure is repeated with several different weights of the
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standard reference material containing mercury concentrations in the desired working range (see
Figure 5).
NOTE: Do not dry the standard reference material as indicated on the certificate of
analysis unless the SRM was prepared and analyzed that way for mercury certification.
Drying may result in loss of mercury that is thermally unstable. Drying a separate sample
at the time of analysis and correcting for moisture content is appropriate.
11.0 PROCEDURE
11.1 General analytical parameters: the analytical parameters depend on the sample
see and matrix and are instrument specific. The following table shows the guidelines given for the
DMA 80. Consult the operating manual for manufactures' recommendations.
Analytical parameters as recommended by Milestone, Inc. for the DMA 80.
Sample
Maximum
Drying Time (s)1
Decomposition
Wait
Type
Capacity

Time (s)1
Time (s)
Aqueous
500 pL or
= [0.7 s * vol. (pL)]
100
40

1000 pL2

Solid (dry)
500 mg
10
= [0.4 s* wt.
40

(mg) + 100 s]

Solid (moist)
500 mg
= [0.7 s * wt (mg) *
= [0.4 s * wt.
40

% water content]
(mg) +100 s]

Solid (high
500 mg
= [0.7 s * wt (mg) *
100
40
organic % water content]
content)
1 The variability of some matrices requires calculating the drying and decomposition times.
2 Maximum sample size is dependent on the volume of the sample boat. Typical sample
boat sizes are either 0.5 or 1.0 mL.
11.2 Sample analysis: For solids, a homogenized amount of the sample is weighed (to ±
0.001 g or better) onto a tared sample boat. The sample boat is inserted into the instrument with
appropriate clean techniques. The analytical parameters chosen are based on the weight,
moisture content, and organic content of the soil (refer to Sec. 11.1). For example, for 200 mg of
sediment with a water content of 45%, the parameters for the DMA 80 would be: 63/180/40. For
aqueous samples and previous prepared samples (using appropriate SW-846 3000 series
methods), a known volume of the sample is dosed onto the sample boat (3, 4). The analytical
parameters chosen are based on the volume of the sample dosed (refer to Sec, 11.1). For
example, for 200 pL of prepared sample, the parameters for the DMA 80 would be: 140/100/40.
11.3 Field analysis: With a stable power supply, this method can be transported to the field
for direct sample analysis without acid digestions.
11.4 Construct a calibration curve by plotting the absorbances of the standards versus
nanograms of mercury. Determine the peak height or peak area of the sample from the chart and
calculate the mercury value from the standard curve. Duplicates, spiked samples, and check
standards should be routinely analyzed as detailed in Section 9.0 of this method. Samples
exceeding the calibration range should be diluted and reanalyzed. Refer to Section 10.0 for
additional guidance on calibrating the instalment.
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12.0 DATA ANALYSIS AND CALCULATIONS
Calculate metal concentrations: (1) by the method of standard addition, (2) from a calibration
curve, or (3) directly from the instrument's concentration read-out. All dilution or concentration
factors must be taken into account Concentrations reported for multiphased or wet samples must
be appropriately qualified (e.g., 5 pg/g dry weight).
13.0 METHOD PERFORMANCE
13.1 This method has been validated with both solid samples and digests of solid samples.
National Institute of Standards and Technology (NIST) Solid Standard Reference Materials (SRMs)
were selected for there homogeneity and availability. The selected SRMs encompass various
chemical forms of mercury, including biological forms, geological forms, and contaminated
environmental forms. The SRMs were analyzed directly as the solid and as the digested sample
as prepared by Method 3052. These results are summarized in Table 1.
Field capabilities of this instrumental method were tested. Direct analysis of various SRMs
were performed in a field setting. A summary of the results is given in Table 2. Field analysis with
this instrumental method resulted in the data in Table 2. Using this method randomly collected field
soil samples were tested. A sample was collected and homogenized in approximately ten minutes
and was analyzed in triplicate in an additional 15 minutes. Field data of randomly collected soil
samples indicate that typical % RSD of less than 10% can be achieved, however this is dependent
on many factors including concentration of mercury and homogeneity of the sample.
13.2 The following documents may provide additional guidance and insight on this method
and technique:
13.2.1 Salvato, N. and Pirola, C.; Analysis of Mercury Traces by Means of Solid
Sample Atomic Absorption Spectrometry. Mikrochimica Acta. Vol. 123,63 - 71,1996.
13.2.2 Walter, P.J., and Kingston, H.M.; The Fate of Mercury in Sample
Preparation", The Pittsburgh Conference, Atlanta, GA, March 1997, paper #1223.
13.2.3 Kingston, H.M., Walter, PJ., Chalk, S., Lorentzen E., and Link, D,; "Chapter
3: Environmental Microwave Sample Preparation: Fundamentals, Methods, and
Applications" in Microwave Enhanced Chemistry, Kingston, H.M. and Haswell, S., Eds.;
American Chemical Society, Washington DC, 1997.
13.2.4 Milestone, Inc., DMA 80 Operating Manual, 160B Shelton Rd., Monroe, CN
06468.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the
quantity and/or toxicity of waste at the point of generation. Numerous opportunities for pollution
prevention exist in laboratory operation. The EPA has established a preferred hierarchy of
environmental management techniques that places pollution prevention as the management option
of first choice. Whenever feasible, laboratory personnel should use pollution prevention techniques
to address their waste generation. When wastes cannot be feasibly reduced at the source, the
Agency recommends recycling as the next best option.
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14.2 For information about pollution prevention that may tie applicable to laboratories and
research institutions consult Less is Better: Laboratory Chemical Management for Waste Reduction
available from the American Chemical Society.
15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society.
16.0 REFERENCES
1. Boylan, H.M., Walter, P.J., and Kingston, H.M.; "Direct Mercury Analysis: Field and
Laboratory Validation for EPA Method 7473".
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 and 2, Figures 1 through 5, and a method procedure
flow diagram.
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TABLE 1
LABORATORY ANALYSIS RESULTS (MEAN ± 95% CONFIDENCE INTERVAL) OF DIRECT
AND DIGESTED (METHOD 3052) ANALYSES OF VARIOUS NIST SRMs
USING THE DMA 80 (MILESTONE, INC.)
(Ref. 1)
Standard
Direct
Digested
Certified
Reference
Analysis
Sample
Value
Material
(ng/g)
Analysis (ng/g)
(ng/g)
Apple Leaves NIST SRM 1515
48.3 ± 2.4
NA
44 ±4
Citrus Leaves NIST SRM 1572
100 ± 12
97 ±9
80 ±20
Estuarine Sediment NIST SRM 1646
75.2 ± 4.9
65.7 ± 8.7
63 ± 12
Oyster Tissue NIST SRM 1566a
67.1 ± 3.2
NA
64.2 ±6.7
Coal Fly Ash NIST SRM 1633b
139 ± 6
132 ± 12
141 ±19
Buffalo River SedimentNIST SRM 2704
1,450 ±24
1,450 ±26
1,440 ±70
Montana Highly Contaminated

Soil NIST SRM 2710
33,100 ±310
33,400 ±230
32,600 ± 1,800
NA: Not analyzed
mZ
TABLE 2
FIELD ANALYSIS RESULTS (MEAN ± 95% CONFIDENCE INTERVAL) OF DIRECT
ANALYSES OF VARIOUS NIST SRMs USING THE DMA 80 (MILESTONE, INC.)
(Ref. 1)
Standard Reference Material
Direct Analysis (ng/g)
Certified Value (ng/g)
EstuarineSediment NIST SRM 1646
74.7 ± 2.4
63 ±12
Oyster Tissue NIST SRM 1566a
68.0 ±2.0
64.2 ±6.7
Coal Fly Ash NIST SRM 1633b
139.2 ±2.2
141 ±19

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FIGURE 1
SPECTRAL OUTPUT OF DMA 80
The two individual peaks correspond to the two absorbance cells of different
sensitivities. The maximum intensity of the long pathlength cuvette (low range
cell) occurs at -8 seconds and the maximum intensity of the short pathlength
cuvette (high range ceil) occurs at ~20 seconds.
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Detector
Interference
en
ow
regulator
Short
ineatiurmg
Delay
vessel
cuvulte
Sample
boat
Amalgamator

Long
measuring
cuvette
Mercury
release
Catalyat
furnace
mposilion
furnace
Spectromete
Doting
equipment
tig lamp
Catalyst furnace
Dryil
decomposition furnacfe
Mercury release furnace
\
Sample boat
Amalgamator

o.
750°C
FIGURE 2
DIAGRAM OF THE MERCURY ANALYSIS SYSTEM
120 - 750°
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FIGURES 3a AND 3b
PRIMARY CALIBRATION CURVES USING THE DMA 80
The low range curve (3a) corresponds to the long pathlength cell. The high range curve (3b)
corresponds to the short pathlength ceil.
Primary Calibration Curve
Low Ranee
» | \ i i \ j i i i i | i i i i pt*i't i yi i r"T'| i >
y = 0.023725 + 0.023015x R= 0 J9782
0.8
0.6
0.4
0.2
0
0
ngHg
Primary Calibration Curve
y = 0,046033 + 0.0014433* R= 0.35c33j
0.8
0.6
0.4
0.2
0 100 200 300 400 500 600
ng Hg
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FIGURE 4
PRIMARY CALIBRATION CURVE USING THE DMA 80 IN FIELD ANALYSIS CONDITIONS
ri '"i i i i r [ i r i i [ i » i i | i i i i i i i [ i i i i | i i i t
41
U
C
Rt
•e
o
w
&
<
y = 0.023495 + 0.020552x R= 0.99766 _
t I « » ¦ i 1 «
I 1 1
0 »in.i ill 111111, i.ii 1.1 ii. 111111 111 1
0 5 10 15 20 25 30 35 40
ng Hg
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FIGURE 5
PRIMARY CALIBRATION CURVES USING THE DMA 80 -
COMPARISON OF THE CALIBRATION USING AQUEOUS STANDARD SOLUTIONS AND
SOLID NATIONAL INSTITUTE OF STANDARDS AND TECHNOLOGY STANDARD
REFERENCE MATERIAL 2704 (BUFFALO RIVER SEDIMENT)
-i 'i | i ¦ i""") "i i i i i i i i r ') 'i 'j i t i i | i i i i
y = 0.05268 + 0.0014197x R= 0.99765
- y = 0.072404 + 0.001448x R=0$p698
u
C
•fi
'O
513
<
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
*— Aqueous Standards
-*— Solid SRM Standards
L I
U-i.
X
XX,
100 200
300 400
rig Hg
500 600
7473 -14
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METHOD 7473
MERCURY IN SOLIDS AND SOLUTIONS BY THERMAL DECOMPOSITION
AMALGAMATION. AND ATOMIC ABSORPTION SPECTROPHOTOMETRY
Existing
primary
calibration?
Yes
No
Are results^
within 10% of
their true
values? .
No
Yes
Type of
sample?
Liquid
Solid
Stop
Set analytical parameters
according to volume.
7,2 Perform primary
calibration.
Proceed with
analysis.
.7.3 Perform daily
calibration.
Set analytical parameters
according to weight, %
moisture, and organic
content.
Calculate concentration of
sample based on calibration
curve, standard addition, or
instrument readout.
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METHOD 7474
MERCURY IN SEDIMENT AND TISSUE SAMPLES
BY ATOMIC FLUORESCENCE SPECTROMETRY
1.0 SCOPE AND APPLICATION
1.1 This procedure measures total mercury in sediment and tissue samples.
1.2 The range of this method is from approximately 1 part per billion to the part per million
range. Analysis of the entire range cannot be accomplished at once, but rather different portions of
this range can be analyzed depending upon the instrument gain settings.
1.3 This method should only be used by analysts experienced with the analysis of trace
elements at very low concentrations when analyzing samples in the ppb range.
2.0 SUMMARY OF THE METHOD
2.1 A representative portion of sample is digested in a microwave unit (a variation of
Method 3052) using nitric and hydrochloric adds in a closed fluorocarbon container. The sample
is digested under pressure to aid in the dissolution of organic compounds containing mercury.
2.2 An aliquot of the digested sample is diluted and subjected to cold digestion with an
acid/bromate/bromide mixture.
2.3 Stannous chloride is added to the digested sample as a reducing agent to produce
Hg°. The reduced mercury is separated from the sample/reagent mixture as a vapor that is carried
to the fluorescence detector by a stream of high purity argon.
3.0 DEFINITIONS
Refer to Chapter One and Chapter Three for a listing of applicable definitions.
4.0 INTERFERENCES
4.1 Method 3052 will overcome the problems associated with incomplete digestion.
Incomplete digestion may lead to the incomplete solubilization of sparingly soluble Hg compounds.
4.2 High purity argon (99.999%) must be used as the carrier gas. Nitrogen will reduce
the sensitivity by a factor eight-fold, while the use of air will reduce the sensitivity thirty-fold,
4.3 The presence of water vapor in the fluorescence detector may produce scattering
effects, positive interferences and degradation of the analytical signal. The use of a dfyer tube is
required to remove any water vapor from the flow before reaching the detector.
4.4 Contamination is always a potential problem in trace element determinations. See
Chapter Three for clean laboratory procedures.
5.0 SAFETY
5.1 Refer to Chapter Three for a discussion on safety related references and issues.
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5.2 Many mercury compounds are highly toxic if swallowed, inhaled, or absorbed through
the skin. Extreme care must be exercised in the handling of concentrated mercury reagents.
Concentrated mercury reagents should only be handled by analysts knowledgeable of their risks and
of safe handling procedures.
6.0 EQUIPMENT AND SUPPLIES
6.1 Atomic fluorescence system:
6.1.1 Autosampler (optional) - A multi-position computer controlled autosampler
may be used. However, it is essential that the autosampler have a wash cycle or "wash pot"
to rinse the probe between sampling positions. The autosampler wash water should closely
approximate the wash water (Section 7.13) in add strength.
6.1.2 Peristaltic pump - A three channel peristaltic pump that can deliver reagents
and sample at flow rates up to 10 mL/min by varying the pump speed, the pump tubing, or.
both is required. Silicone pump tubing is required for ppt determinations as PVC pump
tubing has been found to adsorb mercury.
6.1.3 Solenoid switching valve - A software controlled valve is required to switch
between wash and sample at the proper intervals in the analysis cycle.
6.1.4 Mass flow controllers - Mass flow controllers are required for the earner and
sheath gas flows when analyzing near the detection limit where maximum stability of
conditions is critical.
6.1.5 Gas liquid separator - A gas-liquid separator is required to sparge reduced
mercury from the liquid stream, direct the mercury vapor and argon carrier gas to the
fluorescence detector, arid direct the liquid reagents to waste.
6.1.6 Dryer tube - A dryer tube is to be placed in line between the gas-liquid
separator and the detector to remove water vapor from the carrier gas stream. Any dryer
tube which does not degrade the analysis or sensitivity is acceptable. (PermaPure MD-250-
12 or equivalent.)
6.1.7 Fluorescence detector - A fluorescence detector with a high intensity mercury
light source and a photomultiplier tube at a right angle to the source is required. Use of 254
nm filter coupled with the chemistry of the stannous chloride reduction in the vapor
generator/gas-liquid separator makes the detector highly specific for mercury.
6.1.8 Computer controller - A computer controller and software is required to
operate and coordinate the various components of the system and acquire the data as it is
produced.
6.1.9 Argon gas supply - High purity argon (99.999%) is required. A gas purifier
cartridge is also recommended.
6.1.10 Microwave apparatus. Refer to Method 3051 for a description of an
appropriate microwave digestion apparatus.
6.2 Data systems recorder - A recorder is recommended so that there will be a
permanent record and that any problems with the analysis can be easily recognized.
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6.3 Pipets - Microliter, with disposable tips. Pipet tips should be checked as a possible
source of mercury contamination prior to their use. Class A pipets can be used for the measurement
of volumes equal to or larger than 1 mL.
6.4 Glassware - All glassware, vessels, pipets, etc., must be very clean. Glass, plastic,
and fluorocarbon polymer (PFA or TFM) containers may be used but polymers are not suitable for
samples containing metallic mercury. Hie following is an example of a cleaning procedure
successfully used in a trace level laboratory. Soak glassware overnight in a cleaning solution (such
as Micro®). Rinse four times with Type I water (ASTM Type I water) and soak overnight in an
acid/bromate/bromide mixture. Hie acid/bromate/bromide mixture is made by adding the
bromate/bromide solution from Section 7.9 to dilute acid (approximately 5% v/v) until a yellow color
forms (the exact composition is not critical). The container should be covered or closed, as an open
container will pick up mercury from the atmosphere and permit bromine vapor to escape to the air.
After soaking overnight, add enough 5% hydroxylamine from Section 7.10 to eliminate the yellow
color. Rinse six times with Type I water. Cap the glassware tightly if it is not to be used immediately.
Store in a reduced mercury atmosphere.
When running samples on a daily basis, vessels require the rigorous cleaning procedure
described above every 5 to 7 uses. In between and after each use, the vessels should be soaked
in cleaning solution for 2 hours to loosen deposits. They are then cleaned thoroughly with cotton
swabs (tested for mercury contamination) and soaked again in cleaning solution overnight. Rinse
six times with reagent water. Repeat the soaking and rinsing steps if necessary.
6.5 Balance - A top-loader balance with an accuracy of ± 0.01 g is required.
6.6 Muffle furnace - A muffle furnace capable of reaching and maintaining a temperature
of 150°C is required for purifying the potassium bromate and potassium bromide reagents.
7.0 REAGENTS AND STANDARDS
7.1 Reagent grade Chemicals shall be used in air tests. Unless otherwise indicated, it is
intended that ail reagents shall conform to the specifications of the Committee on Analytical
Reagents of the American Chemical Society, where such specifications are available. Other grades
may be used, provided it is first ascertained that the reagent is of sufficiently high purity to permit its
use without lessening the accuracy of the determination. All reagents should be analyzed to provide
proof that all constituents are below the MDLs.
7.2 Reagent water: All references to water in this method refer to reagent water unless
otherwise specified. Refer to Chapter One for a definition of reagent water.
7.3 Nitric acid, HN03: Use a trace metal grade with negligible mercury content. If the
reagent blank is less than the MDL, the acid may be used.
7.4 Hydrochloric acid, HCI: Use a trace metal grade with negligible mercury content. If
the reagent blank is less than the MDL, the acid may be used.
7.6 Mercury stock standard solution: A mercury stock solution should be purchased from
a reputable source with a concentration of 1.0 mg Hg/mL.
7.7 Potassium bromate (CAS 7758-01-2): Volatilize trace mercury impurities by heating
in a muffle furnace at 150°C for at least S hours. This procedure is recommended every time the
compound is used but if it is stored in a desiccator, ensure that is not contaminated prior to use.
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7.8 Potassium bromide (CAS 7758-02-3): Volatilize trace mercury impurities by heating
in a muffle furnace at 150°C for at least 8 hours.
7.9 Bromate/Bromide solution: Dissolve 1.39 g potassium bromate and 5.95 g potassium
bromide in 500 ml_ reagent water. Prepare weekly.
7.10 Hydroxylamine hydrochloride (CAS 5470-11-1): Use a source that is specified as
suitable for mercury analysis.
7.11 Hydroxylamine hydrochloride solution (5% w/v): Dissolve 2.5 g hydroxylamine in 50
mL of reagent water. Prepare weekly.
7.12 Stannous chloride solution (CAS 10025-69-1), 2% in 10% HCI: Add approximately
500 mL of reagent water to a 1L volumetric flask followed by the addition of 100 mL conc. HCI. Add
20.0 g stannous chloride and stir to dissolve. Dilute to 1 L with reagent water. The solution should
be sparged with argon for 30 minutes prior to analysis to remove any traces of mercury. Prepare
daily.
7.13 Wash water (reagent blank), 5% HCI: Add approximately 1000 mL of reagent water
to a 2 L flask. Add 100 mL conc. HCI and 80 mL of the bromate/bromide solution (Section 7.9).
Bring to volume with reagent water. Prepare daily.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 See Chapter Three, Inorganic Analytes.
9.0 QUALITY CONTROL
9.1 All quality control data should be maintained and available for easy reference or
inspection.
9.2 For each batch of samples processed, method blanks must be carried throughout
the entire sample preparation and analytical process according to the frequency described in Chapter
One. A method blank is prepared by using a volume or weight of reagent water at the volume or
weight specified in the preparation method and then carried through the appropriate steps of the
analytical process. These steps may include but are not limited to digestion, dilution, filtering, and
analysis. These blanks will be useful in determining if samples are being contaminated. Refer to
Chapter One for the proper protocol when analyzing blanks.
9.3 Matrix Spike/Matrix Spike Duplicates (MS/MSDs): MS/MSDs are intralaboratory split
samples spiked with identical concentrations of target analytes. The spiking occurs prior to sample
preparation and analysis. An MS/MS D is used to document the bias and precision of a method in
a given sample matrix. MS/MSDs are to be analyzed at the frequency of one per analytical batch
as described in Chapter One. Refer to the definitions of bias and precision, in Chapter One, for the
proper data reduction protocols. MS/MSD samples should be spiked at the project-specific action
level or when lacking project-specific action levels, between the low and midievel standards.
Acceptance criteria should be set at a laboratory derived limit developed through the use of historical
analyses. In the absence of historical data this limit should be set at ± 20% of the spiked value for
precision and s 20 relative percent difference (RPD). After the determination of historical data, 20%
must still be the limit of maximum deviation for both percent recovery and relative percent difference
to express acceptability.
7474 -4
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9,4 For each batch of samples processed, laboratory control samples must be carried
throughout the entire sample preparation and analytical process according to the frequency of one
per analytical batch as described in Chapter One. The laboratory control samples should be spiked
at the project-specific action level or when lacking project-specific action levels, between the low and
midievel standards. Acceptance criteria should be set at a laboratory derived limit developed
through the use of historical analyses. In the absence of historical data this limit should be set at
± 20% of the spiked value. Refer to Chapter One for more information.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Calibration standards - All analyses require that a calibration curve be' prepared to
cover the appropriate concentration range. Calibration standards are prepared by diluting the stock
metal solutions at the time of analysis and digesting them using the same procedure used for actual
samples. If running more than one batch of samples during the same week, the microwave-digested
standards can be kept in clean dedicated glassware from which dilutions can be made daily which
are then prepared with add and bromate/bromine just as the samples are prepared after
microwaving.
10.1.1 Calibration standards must be prepared fresh (or from the weekly microwave
digest described in 10.1) each time a batch of samples is analyzed. Prepare a reagent blank
and at least three calibration standards in graduated amounts in the appropriate range of the
linear part of the curve.
10.1.2 The calibration standards should be prepared using the same type of bromine,
add or combination of adds and at the same concentration as will result in the samples
following processing.
10.2 A calibration curve must be prepared each day with a minimum of a reagent blank
and three standards. After calibration, the calibration curve must be verified by use of at least a
calibration blank and a check standard (made from a reference material or other independent
standard material) at or near the mid-range. The calibration curve must also be verified at the end
of each analysis batch and/or after every ten samples. The calibration check standard must be
measured within 20% of its true value for the curve to be considered valid.
10.3 The working standard curve must be verified by measuring satisfactorily a mid-range
standard or check standard arid a reagent blank at the end of each analysis batch and/or after every
10 samples. This sample value must be within 10% of the true value, or the previous ten samples
must be reanalyzed. The reagent blank must be less than the MDL If the aforementioned criteria
are not met, reanalyze the samples analyzed since the last passing calibration check and calibration
blank.
11.0 PROCEDURE
11.1 Prepare samples in a microwave unit using only nitric and hydrochloric adds. Follow
instrument manufacturers instructions.
11.1.2 Transfer approximately 1.0 gram of wet sample to a digestion vessel.
Add 2.0 mL of concentrated nitric and 6.0 mL of concentrated hydrochloric and cap.
11.1.3 Microwave the samples with a program appropriate for complete digestion.
Typically, the temperature should be ramped slowly to 190°C without overpressurization and
held at 190°C for 10 minutes or until the digestion is complete.
7474 -5
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11.1.4 Cool the samples. During the cooling period, vent and swirl the samples
occasionally to release dissolved gases. After the samples have cooled and the dissolved
gases have been dissipated, transfer 1.0 mL of the digested sample into a graduated 50 mL
centrifuge tube that contains reagent water, 2.0 mL bromate/bromide solution (Sec. 7.9) and
2.5 mL HO. Bring to volume with reagent water and cold digest for 15 minutes. Document
the accuracy of the centrifuge tubes through mass/volume records or use volumetric
glassware.
11.2 Set up a software controlled timing sequence for the analysis. Follow your instrument
manufacturer's instructions for all settings. Timing sequences that should be addressed are:
11.2.1 Delay: The delay step allows times for the sample line to fill with sample.
11.2.2 Rise: The rise step allows sample to enter the gas/liquid separator and react
with the stannous chloride. Mercury in the digested sample is reduced to Hg° and the argon
earner stream carries the Hg° as mercury vapor to the detector.
11.2.3* Analysis: The analysis time allows for the peak height to rise to its maximum
while the software measures the peak height or area.
11.2.4 Memory: The memory time allows the signal to return to the baseline level.
11.2.5 Set the gas flows at a level providing adequate sensitivity for the desired
analytical range. Flow rates for the following gases should be established: carrier gas, sheath
gas, and dryer tube gas.
11.2.6 When using a variable speed peristaltic pump, choose appropriate sized
tubing to obtain an approximate ratio of 2:1 between sample/wash (Sec. 7.13) flow rates and
the reductant (Sec. 7.12) flow rate,
11.2.7 Set the gain on the detector to the sensitivity range required for the analysis.
11.2.8 If an autosampler is used, set up a wash solution for the autosampler probes.
The autosampler wash solution should closely approximate the wash water (Sec. 7.13) in
add concentration or contain acid at a sufficient strength (typically 5%) to preclude any
sample carryover!
11.2.9 Allow 30 minutes for the system to equilibrate before initiating sample
analysis.
11.3 Sample Analysis
11.3.1 Add 0.10 mL of the hydroxylamine solution (Sec. 7.11) to remove excess
bromine and decolorize the sample.
11.3.2 Allow precipitate or sediment in diluted samples to settle to avoid fouling the
valves with solid material during analysis.
11.3.3 Any sample that gives a response greater than the highest standard must be
diluted and rerun. Add appropriate amounts of reagents to ensure the reagent concentration
of the diluted sample match that of the other samples and the wash (Sec. 7.13).
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11.3.4 Any samples that fall outside the laboratory calculated and derived QC range
must be re-digested and reanalyzed.
12.0 DATA ANALYSIS AND CALCULATIONS
12.1 Maintain an instalment log book that contains all information necessary to reproduce
the analytical conditions associated with a sample run.
12.2 Sample calculation:
result ng Hg 0.05 L final vol. 0.008 L dig soln.
Hg in yg/Kg = x x
L 0.001 L dig sample g sample (x %solids)
13.0 METHOD PERFORMANCE
13.1 Accuracy: Results for accuracy from the US EPA Region IV laboratory are given in
Table 1. Data for liquid reference materials and liquid calibration checks used with this procedure
are included along with the sediment and tissue materials.
13.2 Precision: Results for precision from the Region IV laboratory are provided in Table
2.
13.3 The laboratory for which accuracy and precision data are presented here also
participated in three intercomparison studies.
13.3,1 In the first study, two sediments and two tissues were analyzed for the
National Oceanic and Atmospheric Administration in the Seventh Round Intercomparison for
Trace Metals in Marine Sediments and Biological Tissues.
13.3.1.1 The accepted value for the sample identified as Sediment T
is 0.107 mg/Kg with an acceptable range of 0.087 to 0.127. The value reported from
this laboratory was 0.100 with a standard deviation of 0.003 and percent relative
standard deviation of 3.0. The number of labs reporting results for this sample was
32.
13.3.1.2 The accepted value for the sample identified as BCSS-1 is
0.163 mg/Kg with an acceptable range of 0.096 to 0.230. The value reported from
this laboratory was 0.199 with a standard deviation of 0.013 and percent relative
standard deviation of.3. The number of laboratories reporting results for this sample
was 28.
13.3.1.3 The accepted value for the tissue sample identified as Tissue
S is 0.0618 mg/Kg with an acceptable range of 0.0409 to 0,0827. The value reported
from this laboratory was 0.0574 with a standard deviation of 0.0047 and percent
relative standard deviation of 8.3. The number of laboratories reporting results for
this sample was 33.
13.3.1.4 The certified value for the tissue sample identified as NIST
1566a is 0.0642 mg/Kg with an acceptable range of 0,0575 to 0.0709. The value
reported from this laboratory was 0.0631 with a standard deviation of 0.0042 and
7474 -7
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percent relative standard deviation of 6,8. The number of laboratories reporting
results for this sample was 27.
13.3.2 Two sediments and two tissues were analyzed in the Eighth Round
Intercomparision for Trace Metais in Marine Sediments and Biological Tissues for the
National Oceanic and Atmospheric Administration.
13.3.2.1 The accepted value for the sediment sample identified as
Sediment U is 0.55 mg/Kg with an acceptable range of 0.42 to 0.68, The value
reported from this laboratory was 0.63 with a standard deviation of 2.7. The number
of laboratories reporting results for this sample was 28.
13.3.2.2 The accepted value for the sediment sample identified as
BCSS-1 is 0.180 mg/Kg with an acceptable range of 0.109 to 0.251. The value
reported from this laboratory was 0.23 with a standard deviation of 0.01 and percent
relative standard deviation of 5.7. The number of laboratories reporting results for this
sample was 26.
13.3.2.3 The accepted value for the tissue sample identified as Tissue
V is 0.0654 mg/Kg with an acceptable range of 0.0462 to 0.0846. The value reported
from this laboratory was 0.058 with a standard deviation of 0.006 and percent relative
standard deviation of 8.0. The number of laboratories reporting results for this sample
was 32,
13.3.2.4 The certified value for the tissue sample identified as NIST
1566a is 0.0654 mg/Kg with an acceptable range of 0.0587 to 0.0721. The value
reported from this laboratory was 0.066 with a standard deviation of 0.003 and
percent, relative standard deviation of 5.2. The number of laboratories reporting
results for this sample was 28.
13.3.3 Ten sediment samples from the Florida Everglades were analyzed and the
results from two laboratories are presented in Table 3.
13.4 Comparison data for this method (CVAF) versus cold vapor atomic absorption
generated within the Region IV laboratory are presented in Tables 4 and 5. Attention should be
drawn to the fact that the lowest results in Table 5 are near the limits of detection for the CVAA
method, but well within the CVAF range, while the higher results are obtained by diluting samples
for the CVAF method, but are well within the range for the CVAA method. Therefore, the CVAA
method may be more appropriate for the samples with higher levels of mercury and the CVAF
method is more appropriate for lower level samples.
13.5 The following documents may provide additional guidance and insight on this method
and technique:
13.5.1 W. Van Delft and G. Vos, Analytics Chimica Acta, 209 (1988) 147-156.
13.5.2 "Yorkshire Water Methods of Analysis", 5th Edition, 1988. (ISBN 090507236)
13.5.3 "Safety in Academic Chemistry Laboratories", American Chemical Society
Publication, Committee on Chemical Safety, 3rd Edition, 1979.
7474 - 8
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13.5.4 "OSHA Safety and Health Standards, General Industry", (29 CFR 1910),
Occupational Safety and Health Administration, OSHA 2206, revised January 1976.
13.5.5 "Proposed OSHA Safety and Health Standards, Laboratories", Occupational
Safety and Health Administration, FR July 24,1986.
13.5.6 1985 Annual Book of ASTM Standards. Vol. 11.01; "Standard Specification
for Reagent Water"; ASTM: Philadelphia, PA, 1985; D1193-77.
13.5.7 Standard Methods, 18th Edition, 1992.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the
quantity and/or toxicity of waste at the point of generation. Numerous opportunities for pollution
prevention exist in laboratory operation. The EPA has established a preferred hierarchy of
environmental management techniques that places pollution prevention as the management option
of first choice. Whenever feasible, laboratory personnel should use pollution prevention techniques
to address their waste generation. When wastes cannot be feasibly reduced at the source, the
Agency recommends recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Lass is Better: Laboratory Chemical Management for Waste Reduction
available from the American Chemical Society, Department of Government Relations and Science
Policy, 1155 16th Street, NW, Washington, D.C. 20036, (202) 872-4477.
15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society at the address listed in Section 14.2.
16.0 REFERENCES
1. "Method for Total Mercury in Drinking Water, Surface, Ground, Industrial and Domestic
Waste Waters and Saline Waters* P.S. Analytical Ltd., Sevenoaks, Kent, U.K.
2. "Method for the Determination of Ultra Trace Level Total Mercury in Sediment and Tissue
Samples by Atomic Fluorescence Spectrometry", EPA Region IV.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The following pages contain Tables 1 through 5 and a method procedure flow diagram.
7474 - 9
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TABLE 1
RECOVERY DATA FOR SEDIMENT AND TISSUE REFERENCE MATERIALS,
SPIKES, METHODS CHECKS (MC), AND CALIBRATION CHECKS
Matrix
True value1
Avg. % Rec.
# of samples
Std dev
NIST 8406 SED
60 M9/Kg
103.3
70
9.3
NRCC BEST1
SED
92.0 M0/Kg
100.9
21
7.4
Sediment spike
22.7 to 68,9
jjg/Kg
89.2
38
13.7
NIST 1575 Plant
tissue
150pg/Kg
95.6
13
7.5
NIST 1566 Oyster
tissue
84.2 pg/Kg
96.7
72
12.3
NRCC DORM-1
Fish tissue
798 pg/Kg
108
9
8.1
NBS 1641B inorg
water (MC)
38 to 60.8 ng/L
95.6
15
6.4
NBS 1641C Inorg
water (MC)
36.8 to 73.5
ng/L
105.6
6
4.3
EPA WS024
Inorg+org water
43.2 to 108 ng/L
88.2
16
7.0
EPA WS029
Inorg+org water
10.1 to 50.6
ng/L
97.3
45
7.3
EPA WS030
Inorg+org water
43.2 ng/L
100.6
24
8.6
EPAWS031
inorg+org water
9.08 to 45.4
ng/L
100.8
49
11.1
Calib checks
water
20.0 to 100 ng/L
101.5
139
4.2
1True values analyzed at various dilutions.
Source: Reference 2
7474-10
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TABLE 2
PRECISION DATA FOR SEDIMENT AND TISSUE SAMPLES1
Replicates
Avg %RSD
# Of samples
Std dev
Sediment original
10.2
280
8.2
Sediment duplicate
8.8
43
8.8
Sediment original vs
duplicate
7.5
43
7.0
Tissue original
11.4
61
10.5
Tissue duplicate
12.0
13
10.6
Tissue original vs
duplicate
5.7
13
4.7
^II samples analyzed twice.
Source: Reference 2.
7474-11
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TABLE 3
EVERGLADES SEDIMENT SAMPLE COMPARISON DATA (jig/Kg)
Sample
Reference Laboratory
Region IV Laboratory
Rep 1
Rep 2
Fluorescence
Rep 1
Fluorescence
Rep 2
Method 245.5
1
11
13
8.6
10.3

2
6

5.6
5.4
<25
3
76

76
73
68
4
58

63
64
59
5
410

458
444
424
6
296

501
463
453
7
42

48
46

8
40

47
58

9
36

37
37

10
34
40
42
39

Source: Reference 2
7474-12
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TABLE 4
COMPARISON OF CVAF METHOD VS. CVAA METHOD 245,5 RESULTS
ON EVERGLADES SEDIMENTS, PEAT AND MARL (pg/Kg)
SAMPLE
AVG
%RSD
CVAA
CVAA %RSD
1
169
2.35
148
11.8
2
197
0.35
248
20.3
3
89
8.04
46
56.6
4
64
6.21
48
26.0
5
28
3.09
23
16.4
6
56
3.76
47
15.4
7
97
2.88
75
22.6
a
172
0.11
161
5.7
9
79
7.89
71
10.0
10
75
4.52
68
8.3
11
79
0.70
76
28
12
99
1,54
81
18.1
13
67
1,97
58
12.8
14
146
19.25
130
10.6
15
63
0.52
59
6.3
16
157
0.10
140
10.7
17
144
2.37
144
0.2
18
450
2.55
424
5.5
19
482
6.99
463
5.6
20
247
2.89
226
7.7
21
161
1.97
195
17.8
22
108
4.76
118
7.6
23
278
15.96
356
22.0
24
278
13.71
175
40.0
25
273
2.68
209
23.5
26
456
7.79
285
41.1
27
136
6.83
128
5.42
28
263
3.95
215
17.9
29
89
0.75
80
9.3
30
108
4,79
119
8.4
Source: Reference 2
7474-13
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TABLE 5
COMPARISON OF CVAF METHOD VS. CVAA METHOD 245.5 RESULTS
ON TISSUE SAMPLES (yg/Kg)
Sample
Avg
% RSD
CVAA
%RSD CVAA
Alligator Liver 1
802
18.7
950
15.0
Alligator Liver 2
133
19.9
170
. 21.1
Alligator Liver 3
216
5.2
230
5.5
Alligator Liver 4
62.7
4.6
72
11.7
Gambusia 1
38.2
6.4
66
47.4
Gambusia 2
165
26.7
120'
28.2
Mixed Fish Comp.
70.1
10.6
74
3.4
Bass Filet 1
460
22.2
370
19.4
Bass Filet 2
987
5.5
910
7.2
Bass Filet 3
559
15.9
560
0.2
Bass Filet 4
274
52.8
202
26.9
Bass Filet 5
172
2.6
155
9.5
Catfish Filet 1
62.2
12.3
80
22.3
Catfish Filet 2
133
0.5
150
10.4
Catfish Filet 3
119
7.1
130
7.9
Bluegill Filet 1
36.4
5.1
37
1.6
Bluegill Filet 2
53.7
0.0
36
35.1
Clam Tissue 1
28.5
87.1
25
11.8
Clam Tissue 2
21.1
11.6
30
31.0
Clam Tissue 3
17.6
12.2
30
46.3
Source; Reference 2
7474 -14
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METHOD 7474
MERCURY IN SEDIMENT AND TISSUE SAMPLES
bv ataiii^ ci i lADCCACMrc eDC/rroAHCTDV
P ¦ A I UMIw r LUUIatOwtlivt Oi tw i KyiViC i r\ T
Start
\r
/ 11.3.3 \
IS sample
response
greater than
highest
V std? /
Yes
No
Stop
Dilute
Sample.
Analyze sample.
11.3.2 Allow precipitate to
settle.
11.2 Set-up software controlled
timing sequence for analysis.
11.3.1 Add 0.10 mL of
hydioxylarrine solution.
11,1.1 Transfer 1g of sample
to digestion vessel. Add 2 mL
cone. HMO3 and 6 mL cone. HCI.
11.1.2 Microwave samples with
a program appropriate for
coirfslete digestion.
11.1.3 Cool samples. Vent and swirl
samples. Transfer 1.0 mL of sample
into a centrifuge tube; add reagent
water, 2 mL brorrntc.'bromkle soln.
and 2.5 ml HCI; bring to volume
with water and cold digest for 15 min.
7474-15
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METHOD 9000
DETERMINATION OF WATER IN WASTE MATERIALS
BY KARL FISCHER TITRATION
1,0 SCOPE AND APPLICATION
1.1 The Karl Fischer titration technique is capable of quantifying the water content of
materials from 1 ppm to nearly 100%. Coulometric titration is used for direct analysis of samples
with water contents between 1 ppm and 5%, while volumetric titration is more suitable for direct
analysis of higher levels (100 ppm to 100%). With proper sample dilution, the range of the
coulometric technique can also be extended to 100% water. Both coulometric and volumetric
procedures are presented.
1.2 Multiphasic samples should be separated into physical phases (liquid, solid, etc,} prior
to analysis to assure representative aliquots are analyzed.
1.3 Establishing the water content in a sample may be useful for the reasons to follow.
1.3.1 It is useful in determining the total composition of a sample. In combination
with other analytical results, the mass balance of a sample can be determined.
1.3.2 It is useful in identifying which samples can be analyzed by Infrared
Spectroscopy using sodium chloride cells or which require zinc selenide cells.
1.3.3 It is useful in determining the amount of alcohol in an aqueous solution.
1.3.4 It is useful when distinguishing an aqueous from a nonaqueous solution.
1.3.5 It is useful when setting the proper mixture of feed materials in the incineration
of waste.
2.0 SUMMARY OF METHOD
2.1 In the volumetric procedure, tie sample or an extract of it, is added to a Karl Fischer
solvent consisting of sulfur dioxide and an amine dissolved in anhydrous methanol. This solution
is titrated with an anhydrous solvent containing iodine. The iodine titrant is first standardized by
titrating a known amount of water.
2.2 In the coulometric procedure, the sample or an extract of it, is injected into an
electrolytic cell containing the Karl Fischer solvent, where the iodine required for reaction with water
is produced by anodic oxidation of iodide. With this technique, no standardization of reagents is
required.
2.3 In both procedures, the endpoint is determined amperometrically with a platinum
electrode that senses a sharp change in cell resistance when the iodine has reacted with ail of the
water in the sample.
2.4 In the coulometric procedure, the coulombs of electricity required to generate the
necessary amount of iodine are converted to micrograms of water by the instrument microprocessor,
while in the volumetric procedure, the volume of iodine titrant required to reach the endpoint is
9000 -1
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converted to micrograms of water. Most instruments will also calculate concentration (ppm or
percent) if the sample weight is keyed in.
3.0 DEFINITIONS
Refer to Chapter One and Chapter Three for a listing of applicable definitions.
4.0 INTERFERENCES
4.1 Interfering side-neadions can occur between the various species in the Karl Fischer
reagent and the sample components, resulting in an overestimation of the water content in the
sample.
4.1.1 Hydroxide ions will titrate as water when injected directly Into the titration cell.
This is a significant problem with samples having a pH > 14. When this is suspected to be
a problem, a water vaporization module (furnace) should be used. The sample is heated in
this module and the water vapor carried to the titration cell, while the hydroxide remains In
the module.
4.1.2 Ketones can interfere with some Karl Fischer reagents by reacting with
alcoholic solvents like methanol to form ketals and acetals which can decompose to form
water. This problem can be avoided by substitution of a non-reactive alcohol or increasing
the pH.
4.1.3 The reduction of iodine by oxidizable species such as thiols, ammonia and
thiosulfate results in the consumption of iodine and an overestimation of the water content.
4.2 Undesired interfering side-reactions can also result in the underestimation of the
water content in the sample. These include:
4.2.1 Sulfur dioxide, base, carbonyl functional groups on aldehydes and ketones
and other substances that form bisulfite complexes.
4.2.2 Oxidation of iodide and bisulfite complexes.
5.0 SAFETY
5.1 The toxicity or carcinogenicity of each reagent used in this method has not been
precisely defined; however, each chemical compound should be treated as a potential health hazard.
From this viewpoint, exposure to these chemicals must be reduced to the lowest possible level by
whatever means available. The laboratory is responsible for maintaining a current awareness file
of OSHA regulations regarding the safe handling of the chemicals specified in this method. A
reference file of material data handling sheets should also be made available to all personnel
involved in the chemical analysis.
5.2 Care should be taken in avoiding the inhalation of the reagent vapors or skin contact
with the reagents. If any of the reagents comes in contact with the skin, wash thoroughly with
copious amounts of water. To avoid inhalation of vapors, fill and empty the cell or electrode
assembly in a working laboratory hood. Once the cell is assembled, solvent vapors are contained
so long as the system remains sealed.
5.3 Protective laboratory clothing, eyewear and gloves should be worn at all times.
9000 - 2
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6.0 EQUIPMENT AND SUPPLIES
6.1 Coulometric water titrator - An automatic Karl Fischer titration system with
amperometric, potentiometric or potential difference end point detection. It consists of an electrolytic
titration cell, dual platinum electrode, magnetic stirrer and control unit.
6.2 Volumetric water titrator - An automatic Karl Fischer titration system consisting of a
titration cell, dual platinum electrode, magnetic stirrer, dispensing buret and control unit.
6.3 Syringes - 5 pL, 10 pL and 100 pL.
6.4 Analytical balance - Minimum capacity of 160 g and capable of weighing to 0.0001
grams.
6.5 Screw cap vials, 20 mL.
6.6 Furnace module for determining water in the presence of high levels of hydroxide or
in samples not otherwise amenable to direct titration or extraction. This is interfaced with the titration
cell. An appropriate sample introduction apparatus will also be required.
7.0 REAGENTS AND STANDARDS
7.1 Coulometric cell solutions.
7.1.1 Anode reagent - Main ingredients consisting of methanol, organic base, sulfur
dioxide and a suitable iodine compound.
7.1.2 Cathode reagent - Main ingredients consisting of methanol, organic base,
sulfur dioxide and possibly carbon tetrachloride.
7.2 Volumetric reagents.
7.2.1 Volumetric titrant - A mixture of an organic amine, sulfur dioxide and iodine
dissolved in a non-hygroscopic solvent. Reagents with titers of 1, 2 and 5 mg H20/mL can
be commercially obtained.
7.2.2 Karl Fischer Solvent - Typically consisting of an organic amine and sulfur
dioxide dissolved in anhydrous methanol.
7.3 The reagents described in 7.1 and 7.2 are commercially available.
7.4 Methanol or other appropriate solvent for extracting samples, anhydrous, > 99.8%.
7.5 Water - Reagent water (as defined in Chapter One).
8.0 SAMPLE COLLECTION, PRESERVATION AND STORAGE
8.1 Samples should be collected and stored in containers which will protect them from
changes in volume or water content. Storage in glass with Teflon-lined caps is required if analytes
requiring such storage are to be determined.
9000 -3
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8.2 Samples should be refrigerated at 4°C ± 2°C and then brought to room temperature
prior to analysis if analytes requiring such storage are to be determined.
9.0 QUALITY CONTROL
9.1 For each batch of twenty samples processed, at least one duplicate sample must be
carried throughout the entire sample preparation and analytical process as described in Chapter
One. The relative standard deviation of the duplicate analyses should be <10%.
9.2 For each batch of twenty samples processed, at least one spiked sample must be
carried throughout the entire sample preparation and analytical process as described in Chapter
One. The spike recovery should be 90 to 110%. In the absence of other information, a spike of 50%
water is recommended. Spikes to some matrices, e.g., paints, may not be meaningful due to their
high water levels and problems with spiking emulsions. In these cases, a spike of their extract may
be the best option.
9.3 Certified reference materials should be analyzed where available.
9.4 To assess the accuracy of coulometric titrators, three 5 mg injections of reagent water
are to be performed daily with average recoveries of 90 to 110% and relative standard deviations
of < 5% to be achieved. If the recoveries fall outside of this range, the instrument problem must be
corrected before continuing with sample analysis.
9.5 Background levels of water in reagents are minimized by using anhydrous reagents
and by pre-titration of reagents prior to sample analysis.
9.6 To prevent the carryover of moisture into the syringe, the syringe should be rinsed
once with methanol between samples and twice with the sample prior to loading the volume to be
analyzed. Alternatively, use several syringes that have been oven dried, rotating the drying/use cycle
so that the syringe in use reaches room temperature prior to use.
9.7 Only small aliquots of samples should be handled near the titrator to prevent
contamination of the bulk sample by Karl Fischer reagent solvents.
9.8 When methanol or other solvent extractions are performed, three solvent blanks
should be analyzed with these extracts and the extract results corrected for the mean of these
blanks.
10.0 CALIBRATION AND STANDARDIZATION
10.1 Coulometric procedure - Since coulometric titrators generate iodine on demand by
the titration cell, standardization of titrant is not required.
10.2 Volumetric procedure - The titer of the titrant must be checked on a daily basis. Using
a 5 uL syringe, 3.0 uL of water is injected into the titration cell containing solvent that has been pre-
titrated to remove residual moisture. The titer is calculated as follows:
3.0 mg H20/mL of titrant consumed = mg/mL HaO equivalent of titrant.
9000 -4
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11.0 PROCEDURES
11.1 Sample introduction - The approach which should be used will depend on the
viscosity and solubility of the sample and is left to the discretion of the analyst.
11.2 Direct injection - A sample (5 |jL for coulometric, 100 |iL for volumetric) is weighed
by difference in a cleaned and dried syringe which has the needle inserted in a small piece of
silicone rubber to reduce sample evaporation. The sample is injected into the titration cell septum
and the syringe reweighed to determine the actual weight injected. The sample is titrated to the
endpoint and the results recorded.
11.3 Extraction - A 500 mg sample is extracted with 10 mL of an appropriate solvent by
shaking in a 20 mL vial for 2-3 minutes. The mixture is allowed to settle. Centrifugation may be
required. A100 pi aliquot of the supernatant is titrated. The volume of the aliquot to be titrated can
be varied to achieve results within the linear range of the titrator. Methanol is most commonly used,
but is not appropriate for all materials. Toluene, DMF, pyridine and diglyme are suitable for paints.
11.4 Water Vaporization - A 10 to 100 mg sample is weighed into a sampling tube and
introduced into the furnace or injected into hot mineral oil and the water vapor carried by a gas
stream into the titration cell. This approach is most commonly used with samples which cannot be
directly titrated or extracted or for samples containing high levels of hydroxide ion as described in
4.1.1. Consult the instrument instruction manual for proper use.
12.0 DATA ANALYSIS AND CALCULATIONS
12.1 The water content in ppm or percent in the sample is either calculated by the
instrument, if sample weight was keyed in, or the instrument readout in g HzO found is divided by
the g of sample injected with appropriate corrections for any dilutions or extractions performed.
12.2 Data analysis worksheets should be prepared for all samples analyzed. The
information to be included is the sample identification, sample weight, water content measured,
water content in the original sample and results of quality control tests performed as described in
Section 9.
13.0 METHOD PERFORMANCE
13.1 Coulometric Procedure: Crude oil analysis: In ASTM Method D4928-89, crude oils
containing 0.02 to 5% water were tested in an interlaboratory study. Within laboratory precision was
5 to 10% and between laboratory 7 to 20% relative standard deviation between 0.1 and 5% water.
13.2 Volumetric Procedure
13.2.1 Used oil analysis: A series of used oil standards was prepared by spiking
dried used oil with water over the range 0 to 20%. Additional standards were made by
spiking a hydrocarbon based cutting fluid at 25% and 50%. The results in w/w percent are
shown in Table 1. Over the range 1 to 50% water, a linear regression of the results by the
method vs. the spiked water content followed the relationship: y = 1.0137x + 0.0917 with R2
= 0.9997.
Certified reference materials covering the range 2 to 90% water were analyzed using the
direct injection procedure and volumetric titration. The results are shown in Table 2 and
9000 - 5
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agreed with those obtained using Method 9001 and the certified value. The relative standard
deviations ranged from 1 to 10% for 6 to 10 determinations.
13.2.2 Paint analysis: A certified reference material, ERM-19, Water and Volatiles
in Latex Paint, was analyzed 10 times by the extraction procedure and volumetric titration.
A 500 mg sample was extracted with diglyme, cenirifuged and 125 |jl_ of the supernatant
titrated. The results agreed with results determined by Method 9001:
In ASTM Method D4017-90, paints containing 25 to 75% water wire tested in an
interlaboratory study. Within laboratory precision was 1.7% and between laboratory 5.3%
relative standard deviation.
13.2.3 Otherwastes: In ASTM Method D5530, hazardous waste fuels containing 13
to 32% water yielded within laboratory precision of 1.3% and between laboratory of 4.3%
relative standard deviation.
13.2.4 Soil analysis: A marine sediment was dried and spiked with water over the
range 0 to 40% (w/w). The results are shown in Table 3 and followed the relationship y =
0.9972X + 0.1103 with R2 = 0.3391.
13.2.5 Alcohol analysis: Mixtures of ethanol and water covering the range 0 to 100%
water and three distilled spirits were analyzed by this method and Method 9001. The results
are given in Table 4. Because total dissolved solids like sugars and other carbohydrates
often present in beers, wines and distilled spirits will be counted as "alcohol" when water
content is used to estimate alcohol content, their contribution must be considered and if
necessary, determined and subtracted from the non-water content to determine the alcohol
content.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the
quantity and/or toxicity of waste at the point of generation. The EPA has established a preferred
hierarchy of environmental management techniques that places pollution prevention as the
management option of first choice. Whenever feasible, laboratory personnel should use pollution
prevention techniques to address their waste generation. When waste cannot be feasibly reduced
at the source, the Agency recommends recycling as the next best option.-
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions, consult Less is Better: Laboratory Chemical Management for Waste Reduction,
available from the American Chemical Society's Department of Government Relations and Science
Policy, 1155 16th St., N.W. Washington, D C. 20036, (202) 872-4477.
15.0 WASTE MANAGEMENT
15.1 Spent reagents and samples should be stored and disposed appropriately.
Method 9000
Method 9001
ERM-19. %w/w
42.34 ±1.25
44.91 ±0.31
9000 - 6
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15.2 The Environmental Protection Agency requires that laboratory waste management
practices be conducted consistent with all applicable rules and regulations. The Agency urges
laboratories to protect the air, water and land by minimizing and controlling ail releases from hoods
and bench operations, complying with the letter and spirit of any sewer discharge permits and
regulations, and by complying with all solid and hazardous waste regulations, particularly with
hazardous waste identification rules and land disposal restrictions. For further information on waste
management, consult the Management Manual for Laboratory Personnel, available from the
American Chemical Society at the address listed in Section 14.2.
16.0 REFERENCES
1. ASTM Method D3401-92, Standard Test Method for Water in Halogenated Organic Solvents
and Their Admixtures.
2. ASTM Method D4017-90, Standard Test Method for Water in Paints and Paint Materials by
Karl Fischer Method.
3. ASTM Method D5530-94, Standard Test Method for Total Moisture of Hazardous Waste Fuel
by Karl Fischer Titrimetry.
4. ASTM Method D4928-89 Standard Test Methods for Water in Crude Oils by Coulometric Karl
Fischer Titration.
5. ASTM Method D4377-93a Standard Test Method for Water in Crude Oils by Potentiometric
Karl Fischer Titration.
6. Validation Data for Draft Methods 9000 and 9001 for the Determination of Water Content in
Liquid and Solid Matrices, Dexsil Corp., Hamden, CT.
7. MacLeod, Steven K. "Moisture Determination Using Karl Fischer Titrations." Analytical
Chemistry. Volume 63. Pages 557-566. May 15, 1991.
8. U.S. EPA NEIC Method. "Water Content of Waste Material Samples by Coulometric Karl
Fischer Titration." Pages 1-12, August 1991.
17.0 TABLES, DIAGRAMS, FLOW CHARTS AND VALIDATION DATA
The following pages contain Tables 1 through 4 and a method procedure flow diagram.
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TABLE 1
DETERMINATION OF WATER IN USED OIL
(w/w %)
Expected
Method 9001
Method 9000
0
0.161
0.061
0.1
0.149
0.145
0.2
0.226
0.255
0.5
0.459
0.561
1.0
0.948
1.07
2.0
2.36
2.46
5.0
5.03
5.05
10.0
9.82
9.97
20.0
20.2
20.0
25.0
26.37
26.05
50.0
50.05
50.60
Source: Reference 6
TABLE 2
ANALYSIS OF USED OIL CERTIFIED REFERENCE MATERIALS3
CRM
Certified Value, wt %
Method 9001, wt %
Method 9000, wt %
ERM-34
1.95
1.92±0.02
1.86±0.09
ERM-35
5.86
5.91 ±0.61
6.13±0.55
ERM-36
10.3
10.30±0.85
10.3±0.81
ERM-41
87.4
88.4±6.7
86.4±6.6
"ERM-34 to 41 Water Content in Used Oil Mixtures from Environmental Reference Materials, Inc.
Source: Reference 6
9000 -8
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TABLE 3
DETERMINATION OF WATER IN MARINE SEDIMENT
(w/w%)
Expected
Method 9001
Method 9000
0
1.14
0.579
10
10.06
9.74
20
18.99
19.67
30
28.52
29.95
40
38.47
40.34
Source: Reference 6

TABLE 4
DETERMINATION OF ALCOHOL IN WATER/ALCOHOL MIXTURES
Expected % Alcohol, v/v
Method 9001 (% v/v)
Method 9000 (% v/v)
0
0
0
10
10.0
10.3
25
25.6
25.0
40
40.9
38.7
50
48,5
49.1
80
80.6
79.8
100
99.9
100.0
Vodka, 40
41.9
42.0
Whiskey, 40
40.0
41.9
Gin, 47
47.2
48.7
Source: Reference 6.
9000-9
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METHOD 9000
DETERMINATION OF WATER IN WASTE MATERIALS
BY KARL FISCHER TITRATION
Start
Karl Fischer
Titration
Volumetric
Procedure
Determine
Concentrations
11.3 Extraction
11.2 Direct
Injection
11.4 Water
Vaporization
Sample introduction
10.0 Calibrate titrator
if necessary.
9000 -10
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METHOD 9001
DETERMINATION OF WATER IN WASTE MATERIALS BY QUANTITATIVE
CALCIUM HYDRIDE REACTION
1.0 SCOPE AND APPLICATION
1.1 This quantitative calcium hydride reaction method is capable of determining water in
the concentration range from 0.1% to 100% in liquid and solid materials including oils, paints, soils
and water/alcohol mixtures. It is intended to be used as either a field or laboratory method.
1.2 Multiphasic samples should be separated into physical phases (liquid, solid, etc.) prior
to analysis to assure representative aliquots are analyzed.
1.3 Establishing the amount of water in a sample may be useful for the reasons to follow.
1.3.1 It is useful in determining the total composition of a sample. In combination
with other analytical results, the mass balance of a sample can be determined.
1.3.2 It is useful in the distinction of which samples can be analyzed by infrared
spectroscopy using sodium chloride cells or which require zinc selenide cells.
1.3.3 It is useful in determining the amount of alcohol in an aqueous alcohol
solution.
1.3.4 It is useful when distinguishing aqueous from nonaqueous solutions.
1.3.5 It is useful when setting the proper mixture of feed materials in the incineration
of waste.
2.0 SUMMARY OF METHOD
2.1 A sample of the material to be tested is treated with a specially formulated calcium
hydride reagent which reacts with water in the sample to liberate hydrogen gas as shown below:
CaHz + 2H20 - Ca(OH)2 + 2H21
2.2 The reaction is carried out in a sealed pressure vessel and the resulting pressure is
then measured using a specially designed meter. The results are displayed directly in weight or
volume percent, depending on the sampling method used.
2.3 The reaction is quantitative, measuring all water present in the sample over the range
0.1 to 100%.
3.0 DEFINITIONS
Refer to Chapter One and Chapter Three for a listing of applicable definitions.
9001-1
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4.0 INTERFERENCES
4.1 This method has no known positive Interferences. Tests conducted on 20% (w/w)
solutions of the compounds listed below using twice the normal sample size produced no response.
The following compounds are representative of substances either known to react with calcium
hydride and/or which are likely to be present in materials to be tested using this method:
Ethanol
Methanol
Acetone
Methyl ethyl ketone
Tetrahydrofuran
Diethylene glycol dimethyl ether
Ethylene glycol
Diethylene glycol
~propylene glycol
Stearic acid
2-Ethyl hexanoic add
Lead oxide (II and III)
Aluminum oxide (Brockman I)
4.2 Nitric add reacts with calcium hydroxide to form calcium nitrate tetra-hydrate crystals,
which trap water in the add before it can react with calcium hydride. This yields results as much as
80% lower than the actual water content. This interference is only significant when determining the
water content of concentrated nitric add mixtures.
5.0 SAFETY
5.1 The toxidty or cardnogenidty of each reagent used in this method has not been
predsely defined; however, eadi chemical compound should be treated as a potential health hazard.
From this viewpoint, exposure to these chemicals must be reduced to the lowest possible level by
whatever means available. The laboratory is responsible for maintaining a current awareness file
of OSHA regulations regarding the safe handling of the chemicals specified in this method. A
reference file of material data handling sheets should also be made available to all personnel
involved in the chemical analysis.
5.2 Protective laboratory clothing, eyewear and gloves should be worn at all times.
5.3 The amount of hydrogen gas generated is minimal and is nrot a hazard to the user.
6.0 EQUIPMENT AND SUPPLIES
6.1 Quantitative calcium hydride reaction test kit - Hydroscout test system (Dexsil
Corporation, One Hamden Park Drive, Hamden, CT), or equivalent. Each commercially available
test kit will supply or specify the apparatus and materials necessary for successful completion of the
test.
9001-2
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7.0 REAGENTS AND STANDARDS
7.1 Each commercially available test kit will supply or specify the reagents necessary for
successful completion of the test. Reagents should be labeled with appropriate expiration dates.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 Samples should be collected and stored in containers which will protect them from
changes in volume or water content. Storage in glass with PTFE-lined caps is required if analytes
requiring such storage are to be determined.
8.2 Samples should be refrigerated at 4°C ± 2°C and brought to room temperature prior
to analysis if analytes requiring such storage are to be determined.
9.0 QUALITY CONTROL
9.1 Follow the manufacturer's instructions for quality control procedures.
9.2 For each batch of twenty samples processed, at least one duplicate sample must be
carried throughout the entire sample preparation and analytical process as described in Chapter
One. The relative standard deviation of the duplicate determinations should be <10%.
9.3 For each batch of twenty samples processed, at least one spiked sample must be
carried throughout the entire sample preparation and analytical process as described in Chapter
One, The spike recovery should be 90 to 110%. In the absence of other information, a spike of 50%
water is recommended. Spikes to some matrices (e.g., oils and paints) may not be meaningful due
to their high water levels and problems with spiking emulsions. In these cases, a spike of their
extract may be the best option.
9.4 A test sample provided with the kit should be analyzed to verify proper performance of
the test and meter operation.
9.5 A blank correction for water is not required. Reagents are ampulized instead of bulk
packaged and thus are less likely to absorb water from the air.
9.6 Certified reference materials should be analyzed where available.
10.0 CALIBRATION AND STANDARDIZATION
10.1 The meter provided with the kit is factory calibrated to read directly in percent water.
Every time the meter is turned on, a new zero calibration point is determined.
-m n pcnrpni iop
11.1 Follow the directions provided by your kit manufacturer.
11.2 Oil samples are analyzed by directly reacting a measured 0.4 to 0.8 mL (for v/v
measurements) or 1 g (for w/w measurements) sample with the calcium hydride reagent. Samples
up to 5 mL can be used to determine water in the 0.1 to 1.0% range. The resulting pressure due
9001-3
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to hydrogen gas is converted by the meter to percent water. The meter has separate programs for
reporting results in v/v or w/w percent water.
11,3 Paint and soil samples are analyzed after extracting 1 g samples with a dilution solvent.
A 0.8 mL aliquot of the extract is reacted with the calcium hydride reagent. The meter results are
reported in w/w percent water.
12.0 DATA ANALYSIS AND CALCULATIONS
12.1 The meter provided with the kit is factory calibrated and the equations converting
pressure to percent water are stored in designated programs. The matrix and sample size determine
the appropriate program to use.
12.2 Data analysis worksheets should be prepared for all samples analyzed. The
information to be included is the sample identification, sample weight or volume, water content (as
read from the instrument readout), water content in the original sample (accounting for any dilutions
or extractions) and results of quality control tests performed as described in Section 9.0.
13.0 METHOD PERFORMANCE
13.1 Used oil analysis: A series of used oil standards were prepared by spiking dried used
oil with water over the range 0 to 20%. Additional standards were made by spiking a hydrocarbon
based cutting fluid at 25% and 50%. The results in w/w percent are shown in Table 1. Over the
range 1 to 50% water, a linear regression of the results by the method vs. the spiked water content
followed the relationship: y - 1,007x + 0.1024 with R2 = 0.9993.
Certified reference materials covering the range 2 to 90% water were analyzed using this
method and Method 9000. The results are shown in Table 2. The relative standard deviations
ranged from 1 to 10% for 6 to 10 determinations and the results agreed with the certified value and
those determined by Method 9000.
13.2 Paint analysis: A certified reference material, ERM-19, Water and Volatiles in Latex
Paint, was analyzed 10 times. The results in w/w% were 44.91 ± 0.31%. The RSD of the
measurements was 0.7%. The results by this method agreed with those obtained using Method
9000 (43.38 ± 1.29%).
13.3 Soil analysis: A marine sediment was dried and spiked with water over the range 0 to
40% (w/w). The results are shown in Table 3 and followed the relationship y = 0.9311x + 0.8149
with R2 = 0.9994.
13.4 Alcohol analysis: Mixtures of ethanol and water covering the range 0 to 100% water and
three distilled spirits were analyzed by this method and Method 9000. The results are given in Table
4. Because total dissolved solids like sugars and other carbohydrates often present in beers, wines
and distilled spirits will be counted as "alcohol" when water content is used to estimate alcohol
content, their contribution must be considered and if necessary, determined and subtracted from the
non-water content to determine the alcohol content.
13.5 Other wastes: Concentrated sulfuric and nitric acids and 10 N sodium hydroxide were
analyzed. The water content of the sulfuric acid was determined to be 4.33% vs. the bottle assay
value of 4.2%. The water content of 10 N NaOH was found to be greater than 20%, the upper limit
of the method for undiluted samples. This is expected for 10 N NaOH, which has a nominal water
9001-4
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content in excess of 50%. The water content of concentrated nitric acid was determined to be
around 6% vs. the assay value of 30%.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity
and/or toxicity of waste at the point of generation. Numerous opportunities for pollution prevention
exist in laboratory operation. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of first choice.
Whenever feasible, laboratory personnel should use pollution prevention techniques to address their
waste generation. When waste cannot be feasibly reduced at the source, the Agency recommends
recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions, consult Less is Better Laboratory Chemical Management for Waste Reduction,
available from the American Chemical Society's Department of Government Relations and Science
Policy, 1155 16th St., N.W. Washington, D.C., 20036, (202) 872-4477.
15.0 WASTE MANAGEMENT
15.1 On completion of a test, the reaction tube will contain water, the original sample matrix
and a solution of calcium hydroxide. Samples requiring dilution with an organic solvent will also
require disposal of the solvent Reacted samples and spent solvents should be stored and disposed
appropriately.
15.2 The Environmental Protection Agency requires that laboratory waste management
practices be conducted consistent with all applicable rules and regulations. The Agency urges
laboratories to protect the air, water and land by minimizing and controlling all releases from hoods
and bench operations, complying with the letter and spirit of any sewer discharge permits and
regulations, and by complying with all solid and hazardous waste regulations, particularly with
hazardous waste identification rules and land disposal restrictions. For further information on waste
management, consult the Management Manual for Laboratory Personnel, available from the
American Chemical Society at the address listed in Section 14 2.
16.0 REFERENCES
1. Operating Manual, Hydroscout System, Dexsil Corporation.
2. Lynn, Theodore B., Validation Data for Draft Methods 9000 and 9001 for the Determination of
Water Content in Liquid and Solid Matrices, Dexsil Corp., Hamden, CT. -
17.0 TABLES, DIAGRAMS, FLOW CHARTS AND VALIDATION DATA
The pages to follow contain Tables 1 through 4 and a method procedure flow diagram.
9001-5
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TABLE 1
DETERMINATION OF WATER IN USED OIL
(w/w %)
Expected
Method 9001
Method 9000
0
0.161
0.061
0.1
0.149
0.145
0.2
0.226
0.255
0.5
0.459
0.561
1.0
0.948
1.07
2.0
2.36
2.46
5.0
5.03
5.05
10.0
9.82
9.97
20.0
20.2
20.0
25.0
26.37
26.05
50.0
50.05
50.60
Source: Reference 2
TABLE 2
ANALYSIS OF USED OIL CERTIFIED REFERENCE MATERIALS3
CRM
Certified Value, wt %
Method 9001, wt %
Method 9000, wt %
ERM-34
1.95
1.92±0.02
1.86±0.09
ERM-35
5.86
5.91±0.61
6.13±0.55
ERM-36
10.3
10,30±0,85
10.3±0.81
ERM-41
87.4
88.416.7
86.416.6
aERM-34 to 41 Water Content in Used Oil Mixtures from Environmental Reference Materials, Inc.
Source: Reference 2
9001-6
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TABLE 3
DETERMINATION OF WATER IN MARINE SEDIMENT
(w/w %)
Expected
Method 9001
Method 9000
0
1.14
0.579
10
10.06
9.74
20
18.99
19.67
30
28.52
29.95
40
38.47
40.34
Source: Reference 2.
TABLE 4
DETERMINATION OF ALCOHOL IN WATER/ALCOHOL MIXTURES
Expected % Alcohol, v/v
Method 9001 (% v/v)
Method 9000 (% v/v)
0
0
0
10
10.0
10.3
25
25.6
25.0
40
40.9
38.7
50
48.5
49.1
80
80.6
79.8
100
99.9
100.0
Vodka, 40
41.9
42.0
Whiskey, 40
40.0
41.9
Gin, 47
47.2
48.7
Source: Reference 2
9001-7
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METHOD 9001
DETERMINATION OF WATER IN WASTE MATERIALS BY QUANTITATIVE
CALCIUM HYORIDE REACTION
Start
Type of
sample?
Paint and Soils
Oil
Stop
11.3 Record results
in w/w percent water.
11.3 Record results in
v/v or w/w percent water.
11.1 Follow directions
provided by kit
manufacturer.
11.3 Extract 1g sample
with dilution solvent. React
0.8mL aliquot of extract
with the calcium hydride
reagent.
11.2 React measured
0.4 - 0.8 mL or ig sample
with the calcium hydride
reagent.
9001-8
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CHAPTER FOUR
ORGANIC ANALYTES
Prior to employing the methods m this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information In Chapter Two for guidance on the allowed
flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified in a
regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in this chapter is provided by EPA as guidance to be used
by the analyst and the regulated community in making judgements necessary to meet the data
quality objectives or needs for the intended use of the data.
4.1 SAMPLING CONSIDERATIONS
4.1.1 Introduction
Following the initial and critical step of designing a sampling plan (Chapter Nine) is the
implementation of that plan such that a representative sample of the solid waste is collected. Once
the sample has been collected it must be stored and preserved to maintain the chemical and
physical properties that it possessed at the time of collection. The sample type, type of containers
and their preparation, possible forms of contamination, and preservation methods are all items which
must be thoroughly examined in order to maintain the integrity of the samples. This section
highlights considerations which must be addressed in order to maintain a sample's integrity and
representativeness. This section is, however, applicable only to trace analyses.
Quality Control (QC) requirements need not be met for ail compounds presented in the Table
of Analytes for the method in use, rather, they must be met for all compounds reported. A report of
non-detect is considered a quantitative report, and must meet all applicable QC requirements for that
compound and the method used.
4.1.2 Sample Handling and Preservation
This section deals separately with volatile and semivolatile organics. Refer to Chapter Two and
Table 4-1 of this section for sample containers, sample preservation, and sample holding time
information.
Volatile Organics
Standard 40 mL glass screw-cap VOA vials with Teflon lined silicone septa may be used for
liquid matrices. Special 40 mL VOA vials for purge-and-trap of solid samples are described in
Method 5035. VOA vials for headspace analysis of solid samples are described in Method 5021.
Standard 125 mL widemouth glass containers may be used for Methods 5031 and 5032. However,
the sampling procedures described in Method 5035 may minimize sample preparation analyte loss
better than the procedures described in Methods 5031 and 5032. The vials and septa should be
washed with soap and water and rinsed with distilled deionized water. After thoroughly cleaning the
vials and septa, they should be placed in an oven and dried at 100°C for approximately one hour.
NOTE: Do not heat the septa for extended periods of time (i.e., more than one hour,
because the silicone begins to slowly degrade at 105°C).
When collecting the samples, liquids and solids should be introduced into the vials gently to
reduce agitation which might drive off volatile compounds.
FOUR -1
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In general, liquid samples should be poured into the vial without introducing any air bubbles
within the vial as it is being filled. Should bubbling occur as a result of violent pouring, the sample
must be poured out and the vial refilled. The vials should be completely filled at the time of
sampling, so tiiat when the septum cap is fitted and sealed, and the vial inverted, no headspace is
visible. The sample should be hermetically sealed in the vial at the time of sampling, and must not
be opened prior to analysis to preserve their integrity.
Due to differing solubility and diffusion properties of gases in LIQUID matrices at
different temperatures, it is possible for the sample to generate some headspace
during storage. This headspace will appear in the form of micro bubbles, and should
not invalidate a sample for volatiles analysis.
The presence of a macro bubble in a sample vial generally indicates either improper
sampling technique or a source of gas evolution within the sample. The latter case
is usually accompanied by a buildup of pressure within the vial, (e.g. carbonate-
containing samples preserved with acid). Studies conducted by the USEPA
(EMSL-Ci, unpublished data) indicate that "pea-sized" bubbles (i.e., bubbles not
exceeding 1/4 inch or 6 mm in diameter) did not adversely affect volatiles data.
These bubbles were generally encountered in wastewater samples, which are more
susceptible to variations in gas solubility than are groundwater samples.
Immediately prior to analysis of liquid samples, the aliquot to be analyzed should be taken
from the vial using the instructions from the appropriate sample introduction technique:
For smaller analysis volumes, a gas-tight syringe may be inserted directly through the
septum of the vial to withdraw the sample.
For larger analysis volumes, (e.g. purge-and-trap analyses) the sample may be
carefully poured into the syringe barrel. Opening a volatile sample to pour a sample
into a syringe destroys the validity of the sample for future analysis. Therefore, if
there is only one VOA vial, it is strongly recommended that the analyst fill a second
syringe at this time to protect against possible loss of sample integrity. This second
sample is maintained only until such time as the analyst has determined that the first
sample has been analyzed properly.
If these guidelines are not followed, the validity of the data generated from the samples may be
suspect.
VOA vials for samples with solid or semi-solid matrices (e.g., sludges) should be filled
according to the guidance given in the appropriate 5000 series sample introduction method (see
Table 4-1) to be used. When 125-mL widemouth glass containers are used, the containers should
be filled as completely as possible. The 125-mL vials should be tapped slightly as they are filled to
try and eliminate as much free air space as possible. A minimum of two vials should also be filled
per sample location.
At least two VOA vials should be filled and labeled immediately at the point at which the
sample is collected. They should NOT be filled near a running motor or any type of exhaust system
because discharged fumes and vapors may contaminate the samples. The two vials from each
sampling location should then be sealed in separate plastic bags to prevent cross-contamination
between samples, particularly if the sampled waste is suspected of containing high levels of volatile
organics. (Activated carbon may also be included in the bags to prevent cross-contamination from
highly contaminated samples). VOA samples may also be contaminated by diffusion of volatile
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organics through the septum during shipment and storage. To monitor possible contamination, a
trip blank prepared from organic-free reagent water (as defined in Chapter One) should be carried
throughout the sampling, storage, and shipping process.
Semivolatile Organics (including Pesticides, PCis and Herbicides.)
Containers used to collect samples for the determination of semivolatile organic compounds
should be soap and water washed Mowed by methanol (or isopropanol) rinsing (see Sec. 4.1.4 for
specific instructions on glassware cleaning). The sample containers should be of glass or Teflon,
and have screw-caps with Teflon lined septa. In situations where Teflon Is not available, solvent-
rinsed aluminum foil may be used as a liner. However, acidic or basic samples may react with the
aluminum foil, causing eventual contamination of the sample. Plastic containers or lids mav NOT
be used few the storage of samples due to the possibility of sample contamination Iron the phthalate
esters and other hydrocarbons within the plastic. Sample containers should be filled with care so
as to prevent any portion of the collected sample coming in contact with the sampler's gloves, thus
causing contamination. Samples should not be collected or stored in the presence of exhaust
fumes. If the sample comes in contact with the sampler (e.g. if an automatic sampler is used), run
organic-free reagent water through the sampler and use as a field blank.
4.1.3 Safety
Safety should always be the primary consideration in the collection of samples. A thorough
understanding of the waste production process, as well as all of the potential hazards making up the
waste, should be investigated whenever possible. The site should be visually evaluated just prior
to sampling to determine additional safety measures. Minimum protection of gloves and safety
glasses should be worn to prevent sample contact with the skin and eyes. A respirator should be
worn even when working outdoors if organic vapors are present More hazardous sampling missions
may require the use of supplied air and special clothing.
4.1.4 Cleaning of Glassware
In the analysis of samples containing components in the parts per billion range, the
preparation of scrupulously clean glassware is necessary. Failure to do so can lead to a myriad of
problems in the interpretation of the final chromatograms due to the presence of extraneous peaks
resulting from contamination. Particular care must be taken with glassware such as Soxhlet
extractors, Kudema-Danish evaporative concentrators, sampling-train components, or any other
glassware coming in contact with an extract that will be evaporated to a smaller volume. The
process of concentrating the compounds of interest in this operation may similarly concentrate the
contaminating substance(s), which may seriously distort the results.
The basic cleaning steps are:
1. Removal of surface residuals immediately after use;
2. Hot soak to loosen and float most particulate material;
3. Hot water rinse to flush away floated particulates;
4. Soak with an oxidizing agent to destroy traces of organic compounds;
5. Hot water rinse to flush away materials loosened by the deep penetrant soak;
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6. Distilled water rinse to remove metallic deposits from the tap water;
7. Alcohol, e.g., isopropanol or methanol, rinse to flush off any final traces of organic
materials and remove the water, and
8. Flushing the item immediately before use with some of the same solvent that will be
used in the analysis.
Each of these eight fundamental steps are discussed here in the order in which they
appeared on the preceding page.
1. As soon possible after glassware (i.e., beakers, pipets, flasks, or bottles) has come in
contact with sample or standards, the glassware should be flushed with alcohol before
it is placed in the hot detergent soak. If this is not done, the soak bath may serve to
contaminate all other glassware placed therein.
2. The hot soak consists of a bath of a suitable detergent in water of 50°C or higher. The
detergent, powder or liquid, should be entirely synthetic and not a fatty add base.
There are very few areas of the country where the water hardness is sufficiently low to
avoid the formation of some hard-water scum resulting from the reaction between
calcium and magnesium salts with a fatty acid soap. This hard-water scum or curd
would have an affinity particularly for many chlorinated compounds and, being almost
wholly water-insoluble, would deposit on all glassware in the bath in a thin film.
There are many suitable detergents on the wholesale and retail market. Most of the
common liquid dishwashing detergents sold at retail are satisfactory but are more
expensive than other comparable products sold industrially. Alconox, in powder or
tablet form, is manufactured by Alconox, Inc., New York, and is marketed by a number
of laboratory supply firms. Sparkleen, another powdered product, is distributed by
Fisher Scientific Company.
3. No comments required.
4. The most common and highly effective oxidizing agent for removal of traces of organic
compounds is the traditional chromic acid solution made up of concentrated sulfuric
acid and potassium or sodium dichromate. For maximum efficiency, the soak solution
should be hot (40-50°C). Safety precautions must be rigidly observed in the handling
of this solution. Prescribed safety gear should include safety goggles, rubber gloves,
and apron. The bench area where this operation is conducted should be covered with
fluorocarbon sheeting because spattering will disintegrate any unprotected surfaces.
The potential hazards of using chromic-sulfuric acid mixture are great and have been
well publicized. There are now commercially available substitutes that possess the
advantage of safety in handling. These are biodegradable concentrates with a claimed
cleaning strength equal to the chromic add solution. They are alkaline, equivalent to
ca. 0.1 N NaOH upon dilution, and are daimed to remove dried blood, silicone greases,
distillation residues, insoluble organic residues, etc. They are further daimed to remove
radioactive traces and will not attack glass or exert a corrosive effect on skin or dothing.
One such product is "Chem Solv 2157," manufactured by Mallinckrodt and available
through laboratory supply firms. Another comparable product is "Detex," a product of
Borer-Chemie, Solothum, Switzerland.
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5,6, arid 7. No comments required,
8. There is always a possibility that between the time of washing and the next use, the
glassware could pick up some contamination from either the air or direct contact. To
ensure against this, it is good practice to flush the item immediately before use with
some of the same solvent that will be used in the analysis.
The drying and storage of the cleaned glassware is of critical importance to prevent the
beneficial effects of the scrupulous cleaning from being nullified. Pegboard drying is not
recommended. It is recommended that laboratory glassware and equipment be dried at 100°C.
Under no circumstances should such small items be left in the open without protective covering.
The dust cloud raised by the daily sweeping of the laboratory floor can most effectively
recontaminate the clean glassware.
As an alternate to solvent rinsing, the glassware can be heated to a minimum of 300°C to
vaporize any organics. Do not use this high temperature treatment on volumetric glassware,
glassware with ground glass joints, or sintered glassware.
4.1.5 High Concentration Samples
Cross contamination of trace concentration samples may occur when prepared in the
same laboratory with high concentration samples. Ideally, if both type samples are being
handled, a laboratory and glassware dedicated solely to the preparation of high concentration
samples would be available for this purpose. If this is not feasible, as a minimum when
preparing high concentration samples, disposable glassware should be used or, at least,
glassware dedicated entirely to the high concentration samples. Avoid cleaning glassware
used for both trace and high concentration samples in the same area.
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TABLE 4-1.
SAMPLE CONTAINERS, PRESERVATION, TECHNIQUES, AND HOLDING TIMES
VOLATILE ORGANICS
Sample Matrix
Container
Preservative
Holding Time
Concentrated Waste
Samples
Method 5035: 40-mL vials with stirring bar.
Method 5,021: See method.
Methods 5031 & 5032: 125-mL widemouth
glass container.
Use Teflon-lined lids for all procedures.
Cool to 4°C.
14 days
Aqueous Samples With
No Residual Chlorine
Present
Methods 5030, 5031, & 5032: 2 X 40-mL
vials with Teflon-lined septum caps
Cool to 4°C and adjust pH to less than 2 with
H2S04, HCI, or solid NaHS04.
14 days
Aqueous Samples WITH
Residual Chlorine Present
Methods 5030, 5031, & 5032: 2 X 40-mL
vials with Teflon-lined septum caps
Collect sample in a 125-mL container which
has been pre-preserved with 4 drops of 10%
sodium thiosulfate solution. Gently swirl to mix
sample and transfer to a 40-mL VOA vial.
Cool to 4°C and adjust pH to less than 2 with
H2S04l HCI, or solid NaHS04.
14 days
Acrolein and Acrylonitrile
in Aqueous Sample
Methods 5030, 5031, & 5032: 2 X 40-mL
vials with Teflon-lined septum caps
Adjust to pH 4-5. Cool to 4°C.
14 days
Solid Samples
(e.g. soils, sediments,
sludges, ash)
Method 5035: 40-mL vials with septum and
stirring bar.
Method 5021: See method.
Methods 5031 & 5032: 125-mL widemouth
glass container with Teflon-lined lids.
See the individual methods.
14 days
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TABLE 4-1 (Continued)

SEMIVOLATILE ORGANICS/ORGANOCHLORINE PESTICIDES/PCBs AND HERBICIDES

Sample Matrix
Container
Preservative
Holding Time
Concentrated Waste
Samples
125-mL widemouth glass with Teflon-lined
lid
None
Samples extracted
within 14 days and
extracts analyzed
within 40 days
following extraction.
Aqueous Samples With
No Residual Chlorine
Present
1-gal., 2 x 0.5-gal., or 4 x 1-L amber glass
container with Teflon-lined lid
Cool to 4°C
Samples extracted
within 7 days and
extracts analyzed
within 40 days
following extraction.
Aqueous Samples WITH
Residual Chlorine Present
1-gal., 2 x 0.5-gal., or 4 x 1-L, amber glass
container with Teflon-lined lid.
Add 3-mL 10% sodium thiosulfate solution per
gallon (or 0.008%). Addition of sodium
thiosulfate solution to sample container may be
performed in the laboratory prior to field use.
Cool to 4°C.
Samples extracted
within 7 days and
extracts analyzed
within 40 days
following extraction.
Solid Samples
(e.g. soils, sediments,
sludges, ash)
250-mL widemouth glass container with
Teflon-lined lid
Cool to 4°C
Samples extracted
within 14 days and
extracts analyzed
within 40 days
following extraction.
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4.2 SAMPLE PREPARATION METHODS
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the
allowed flexibility in the choice of apparatus, reagents, and supples. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. Hie information contained in each procedure is provided by EPA as guidance to
be used by the analyst and the regulated community in making judgements necessary to meet
the data quality objectives or needs for the intended use of the data.
4.2.1 EXTRACTIONS AND PREPARATIONS
The following methods are included in this section:
Method
3500B:
Method
3510C:
Method
3520C:
Method
3535A:
Method
3540C:
Method
3541:
Method
3542:
Method
3545A:
Method
3550B:
Method
3560:
Method
3561:
Method
3562:
Method
3580A:
Method
3585:
Method
5000:
Method
5021:
Method
5030B:
Method
5031:
Method
5032:
Method
5035:
Method
5041A:
Organic Extraction and Sample Preparation
Separatory Funnel Liquid-Liquid Extraction
Continuous Liquid-Liquid Extraction
Solid-Phase Extraction (SPE)
Soxhlet Extraction
Automated Soxhlet Extraction
Extraction of Semivolatile Analytes Collected Using Method
0010 (Modified Method 5 Sampling Train)
Pressurized Fluid Extraction (PFE)
Ultrasonic Extraction
Supercritical Fluid Extraction of Total Recoverable Petroleum
Hydrocarbons
Supercritical Fluid Extraction of Polynuclear Aromatic
Hydrocarbons
Supercritical Fluid Extraction of Polychlorinated Biphenyis
(PCBs) and Organochlorine Pesticides
Waste Dilution
Waste Dilution for Volatile Organics
Sample Preparation for Volatile Organic Compounds
Volatile Organic Compounds in Soils and Other Solid Matrices
Using Equilibrium Headspace Analysis
Purge-and-Trap for Aqueous Samples
Volatile, Nonpurgeable, Water-Soluble Compounds by
Azeotropic Distillation
Volatile Organic Compounds by Vacuum Distillation
Closed-System Purge-and-Trap and Extraction for Volatile
Organics in Soil and Waste Samples
Analysis for Desorption of Sorbent Cartridges from Volatile
Organic Sampling Train (VOST)
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METHOD 3535A
SOLID-PHASE EXTRACTION (SPE)
1.0 SCOPE AND APPLICATION
1.1 This method describes a procedure for isolating target organic analytes from aqueous
samples using solid-phase extraction (SPE) media. The method describes conditions for extracting
a variety of organic compounds from aqueous matrices that include: groundwater, wastewater, and
TCLP leachates. The method describes the use of disk extraction media for eight groups of analytes
and the use of cartridge extraction media for one group of analytes. Other solid-phase extraction
media may be employed as described in see Sec. 4.0. The extraction procedures are specific to the
analytes of interest and vary by group of analytes and type of extraction media. The groups of
analytes that have been evaluated thus far are listed below, along with the types of media that have
been evaluated, and the determinative methods in which the corresponding performance data can
be found.
Analyte
Extraction
Determinative
Group
Media Type
Method
Phthalate esters
Disks
8061
Organochlorine pesticides
Disks
8081
Polychlorinated biphenyls (PCBs)
Disks
8082
Organophosphorus pesticides
Disks
8141
Nitroaromatics and nitramines
Disks and Cartridges
8330
TCLP leachates containing organochlorine pesticides
Disks
8081
TCLP leachates containing semivolatiles
Disks
8270
TCLP leachates containing phenoxyacid herbicides
Disks
8321
1.2 The technique may also be applicable to other semivolatile or extractable compounds.
It may also be used for the extraction of additional target analytes or may employ other solid-phase
media, provided that the analyst demonstrates adequate performance (e.g., recovery of 70 -130%,
or project-specific recovery criteria) using spiked sample matrices and an appropriate determinative
method of the type included in Chapter Four (Sec, 4.3). The use of organic-free reagent water alone
is not considered sufficient for conducting such performance studies, and must be supported by data
from actual sample matrices.
1.3 This method also provides procedures for concentrating extracts and for solvent
exchange.
1.4 Solid-phase extraction is called liquid-solid extraction in some methods associated with
the Safe Drinking Water Act.
1.5 This method is restricted to use by or under the supervision of trained analysts. Each
analyst must demonstrate the ability to generate acceptable results with this method.
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2.0 SUMMARY OF METHOD
2.1 Sample preparation procedures vary by arialyte group. Extraction of some groups
requires that the pH of the sample be adjusted to a specified value prior to extraction (see Sec. 7.2).
Other groups do riot require a pH adjustment.
2.2 Following any necessary pH adjustment, a measured volume of sample is extracted
by passing it through the solid-phase extraction medium (disks or cartridges), which is held in an
extraction device designed for vacuum filtration of the sample.
2.3 Target analytes are eluted from the solid-phase media using an appropriate solvent
(see Sees. 7.8 and 7.9) which is collected in a receiving vessel. The resulting solvent extract is dried
using sodium sulfate and concentrated, as needed.
2.4 As necessary for the specific analysis, the concentrated extract may be exchanged into
a solvent compatible extract with subsequent cleanup procedures (Chapter Four, Sec. 4.2) or
determinative procedures (Chapter Four, Sec. 4.3) for the measurement of the target analytes.
3.0 INTERFERENCES
3.1 Refer to Method 3500.
3.2 The decomposition of some analytes has been demonstrated under basic extraction
conditions. Organochlorine pesticides may dechlorinate and phthalate esters may hydrolyze. The
rates of these reactions increase with increasing pH and reaction times.
3.3 Bonded-phase silica (e.g., C18) will hydrolyze on prolonged exposure to aqueous
samples with pH less than 2 or greater than 9. Hydrolysis will increase at the extremes of this pH
range and with longer contact times. Hydrolysis may reduce extraction efficiency or cause baseline
irregularities. Styrene divinytbenzene (SDB) extraction disks should be considered when hydrolysis
is a problem.
3.4 Phthalates are a ubiquitous laboratory contaminant. All glass extraction apparatus
should be used for this method because phthalates are used as release agents when molding rigid
plastic (e.g., PVC) and as plasticizers for flexible tubing. A method blank, as described in Chapter
One, should be analyzed, demonstrating that there is no phthalate contamination of the sodium
sulfate or other reagents listed in this method.
3.5 Sample particulates may clog the solid-phase media and result in extremely slow
sample extractions. Use of an appropriate filter aid will result in shorter extractions without loss of
method performance if dogging is a problem. Even when a filter aid is employed, this method may
not be appropriate for aqueous samples with high levels of suspended solids (>1%), as the
extraction efficiency may not be sufficient, given the small volumes of solvents employed and the
short contact time.
4.0 APPARATUS AND MATERIALS
The apparatus and materials described here are based on data provided to EPA for the
extraction of eight groups of analytes using disk-type materials and for the extraction of one group
of analytes using cartridge-type materials. Other solid-phase extraction media configurations may
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be employed, provided that the laboratory demonstrates adequate performance for the analytes of
interest. The use of other SPE configurations will require modifications to the procedures described
in Sec. 7.0. Consult the manufacturer's instructions regarding such modifications.
4.1 Solid-phase disk extraction system - Empore™ manifold that holds three 90-mm filter
standard apparatus or six 47-mm standard filter apparatus, or equivalent. Other manual, automatic,
or robotic sample preparation systems designed for solid-phase media may be utilized for this
method if adequate performance is achieved and all quality control requirements are satisfied.
4.1.1 Manifold station - (Fisher Scientific 14-378-1B [3-place], 14-378-1A [6-place],
or equivalent).
4.1.2 Standard filter apparatus - (Fisher Scientific 14-378-2A [47-mm], 14-378-2B
[90-mm], or equivalent), consisting of a sample reservoir, damp, fritted disk and filtration head
with drip tip.
4.1.3 Collection tube - 60-mL. The collection tube should be of appropriate ID and
length so that the drip tip of the standard filter apparatus can be positioned well into the neck
of the tube to prevent splattering.
4.1.4 Filter flask - 2-L with a ground-glass receiver joint (optional). May be used to
carry out individual disk extractions with the standard filter apparatus and collection vial in an
all-glass system.
4.2 Solid-phase cartridge extraction system - Visiprep solid-phase extraction manifold
(Supefco) or equivalent system suitable for use with the extraction cartridges (see Sec. 4.4). Consult
the manufacturer's recommendations for the associated glassware and hardware necessary to
perform sample extractions.
4.3 Solid-phase extraction disks - Empore™, 47-mm, 90-mm, or equivalent Disks are
available in 47-mm and 90-mm diameters, composed of a variety of solid-phase materials. Other
solid phases may be employed, provided that adequate performance is demonstrated for the
analytes of interest. Guidance for selecting the specific disk is provided in Table 1.
4.3.1 G18 disks - Empore™ disks, 47-mm diameter (3M product number 98-0503-
0015-5), 90-mm diameter (3M product number 98-0503-0019-7), or equivalent.
4.3.2 C18 fast flow disks - Empore™ disks, 47-mm diameter (3M product number
98-0503-0138-5), 90-mm diameter (3M product number 98-0503-0136-9), or equivalent. These
disks may be a better choice for samples that are difficult to filter even with the use of a filter
aid.
4.3.3 Styrene di vinyl benzene (SDB-XC) disks - Empore™ disks, 47-mm diameter
(3M product number 98-0503-0067-6), 90-mm diameter (3M product number 98-0503-0068-4),
or equivalent.
4.3.4 Styrene divinylbenzene reversed-phase sulfonated (SDB-RPS) disks -
Empore™ disks, 47-mm diameter (3M product number 98-0503-0110-4), 90-mm diameter (3M
product number 98-0503-0111-2), or equivalent.
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4.4 Solid-phase extraction cartridges - Porapak® R SPE device, Waters Corporation, or
equivalent. Other solid phases may be employed, provided that adequate performance is
demonstrated for the analytes of interest.
4.5 Filtration aid (optional)
4.5.1 Filter Aid 400 - (Fisher Scientific 14-378-3, or equivalent).
4.5.2 In-situ glass micro-fiber prefilter - (Whatman GMF 150, 1-miti pore size, or
equivalent).
4.6 Drying column - 22-mm ID glass chromatographic column with a PTFE stopcock
(Kontes K-420530-0242, or equivalent).
NOTE: Fritted glass discs used to retain sodium sulfate in some columns are difficult to
decontaminate after contact with highly contaminated or viscous extracts. Columns
suitable for this method use a small pad of glass wool to retain the drying agent.
4.7 Kudema-Danish (K-D) apparatus
4.7.1 Concentrator tube - 10-mL, graduated. A ground-glass stopper is used to
prevent evaporation of extracts during short-term storage.
4.7.2 Evaporation flask - 500-mL, or other size appropriate for the volumes of
solvents to be concentrated. Attach to concentrator tube using springs or clamps.
4.7.3 Three-ball macro-Snyder column.
4.7.4 Two-ball micro-Snyder column (optional).
4.7.5 Springs - 14-inch.
4.8 Solvent Vapor Recovery System - Kontes 545000-1006 or K-547300-0000, Ace Glass
6614-30, or equivalent.
NOTE: The glassware in Sec. 4.6 is recommended for the purpose of solvent recovery during the
concentration procedures (Sees. 7.10 and 7.11) requiring the use of Kudema-Danish
evaporative concentrators. Incorporation of this apparatus may be required by State or
local municipality regulations that govern air emissions of volatile organics. EPA
recommends the incorporation of this type of reclamation system as a method to
implement an emissions reduction program. Solvent recovery is a means to conform with
waste minimization and pollution prevention initiatives.
4.9 Boiling chips - Solvent extracted, approximately 10/40 mesh (silicon carbide, or
equivalent).
4.10 Water bath • Heated, with concentric ring cover, capable of temperature control to
within ± 5°C. The bath should be used in a hood.
4.11 Nitrogen evaporation apparatus (optional) - N-Evap, 12- or 24-position (Organomation
Model 112, or equivalent).
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4.12 Vials, glass - Sizes as appropriate, e.g., 2-mL or 10-mL, with PTFE-lined screw caps
or crimp tops for storage of extracts.
4.13 pH indicator paper - Wide pH range.
4.14 Vacuum system - Capable of maintaining a vacuum of approximately 66 cm (26 indies)
of mercury.
4.15 Graduated cylinders - Sizes as appropriate.
4.16 Pipets - disposable.
5.0 REAGENTS
5.1 Reagent grade chemicals shall be used in all tests. Unless otherwise indicated, it is
intended that all reagents shall conform to the specifications of the Committee on Analytical
Reagents of the American Chemical Society, where such specifications are available. Other grades
may be used, provided K is first ascertained that the reagent is of sufficiently high purity to permit its
use without decreasing the accuracy of the determination. Reagents should be stored in glass to
prevent the leaching of contaminants from plastic containers.
5.2 Organio-free reagent water - All references to water in this method refer to organic-free
reagent water, as defined in Chapter One.
5.3 Sodium sulfate (granular, anhydrous), Na2SO< - Purify by heating at 400°C for 4 hours
in a shallow tray, or by precleaning the sodium sulfate with methylene chloride.
5.4 Solutions for adjusting the pH of samples before extraction.
5.4.1 Sulfuric acid solution (1:1 v/v), H2S04 - Slowly add 50 mL of concentrated
H2S04 (sp. gr. 1.84) to 50 mL of organio-free reagent water.
5.4.2 Sodium hydroxide solution (10N), NaOH - Dissolve 40 g NaOH in organic-free
reagent water and dilute to 100 mL
5.5 Extraction, washing, and exchange solvents - At a minimum, all solvents must be
pesticide quality or equivalent.
5.5.1
Methylene chloride, CH2CI2.
5.5.2
Hexane, C6H14.
5.5.3
Ethyl acetate, CH3C(OH)OCH2CH3.
5.5.4
Acetonitrile, CH3CN.
5.5.5
Methanol, CH3OH.
5.5.6
Acetone, (CH^CO.
5.5.7
Methyl-ferf-butyl ether (MTBE), CjH^O.
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6.0 SAMPLE COLLECTION, PRESERVATION, AND HANDLING
See the introductory material to Chapter Four, Organic Analytes, Sec. 4.1, Method 3500, Sec.
7.1 of this method, and the specific determinative methods to be employed.
7.0 PROCEDURE
The procedures for solid-phase extraction are vety similar for most organic analytes.
Therefore, this section describes procedures for sample preparation, pH adjustment, preparation of
the extraction apparatus, and extract concentration that apply to al! target analytes. The procedures
for disk washing, disk conditioning, sample extraction, and sample etution vary among the groups
of analytes.
7.1 Sample preparation
Most of the specific procedures described in this method were developed for a nominal sample
size of 1 L, as this sample size is usually employed for other extraction methods such as separatory
funnel or continuous liquid-liquid extraction. This method also may be employed with smaller
samples when overall analytical sensitivity is not a concern or when high levels of the target analytes
are anticipated. However, such samples are best collected in a container of appropriate size. The
extraction of aqueous samples presents several challenges that must be considered during sample
preparation. First, the analytes of interest are often associated with the particulate matter in the
sample and sample preparation procedures must ensure that any particulates in the original sample
are included in the sample aliquot that is extracted. Secondly, the majority of the organic analytes
are hydrophobic and may preferentially adhere to the surfaces of the sample container. For this
reason, most extraction methods have traditionally specified that once the sample has been
transferred to the extraction apparatus, the sample container be rinsed with solvent which is added
to the apparatus. As a result, it is generally not appropriate to extract only part of the sample from
a sample container, e.g., 250 mL from a 1-L sample bottle.
The appropriate sample volume may vary with the intended use of the results and, in general,
is foe volume necessary to provide the analytical sensitivity necessary to meet the objectives of the
project (see Chapter Two). Under ideal conditions, the sample should be collected by completely
filling the container. The sample should generally be collected without additional volume and with
little or no headspace. Thus, a 1-L sample is collected in a 1-L container, a 250-mL sample is
collected in a 250-mL container, etc.
Any surrogates and matrix spiking compounds (if applicable) are added to the sample in the
original container. The container is then recapped and shaken to mix the spiked analytes into the
sample. The extraction of some groups of analytes also requires that the pH of the sample be
adjusted to a specified value (see Table 1). When pH adjustment is necessary, it should be
performed after the surrogates and matrix spiking compounds (if applicable) have been added and
mixed with the sample. Otherwise, the recoveries of these compounds will have little relevance to
those of the target analytes in the sample.
If this approach is not possible, then a sample aliquot may be transferred to a graduated
cylinder and spiked. However, in such instances, the analyst must take great care to mix the sample
well, by shaking, to ensure a homogeneous distribution of the particulate matter and must record the
fact that the container was not rinsed.
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NOTE: This method may riot be appropriate for aqueous samples with greater than 1 % solids, as
such samples can be difficult to filter and the extraction efficiency may be reduced as a
result of the small volumes of solvents employed and the short contact time. If the
particulate load significantly slows or prevents filtration, it may be more appropriate to
employ an alternative extraction procedure.
7.1.1 Mark the level of the sample on the outside of the sample container for later
determination of the sample volume used. Shake the container for several minutes, with the
cap tightly sealed, to ensure that any particulate matter is evenly distributed throughout the
sample.
7.1.2 Prepare a method Wank from a 1-L volume of organic-free reagent water, or
a volume similar to that of the samples (e.g., a 250-mL blank should be used when the sample
size is 250 mL, etc.). The blank may be prepared in a graduated cylinder, beaker, or other
suitable container. The frequency of method blank preparation is described in Chapter One.
7.1.3 Add any surrogate standards listed in the determinative method to the *
samples in their original containers and to the blank. For disk extractions, also add 5.0 mL of
methanol to each sample in the original container. All samples, blanks, and QC samples
should receive the same amount of methanol. (This step is not necessary for the cartridge
extraction of nitroaromatics and nitramines.) Shake the samples to mix the surrogates and
allow the sample to stand for at least several minutes. This will permit the surrogates to
dissolve in the sample and will also allow the particulate matter to settle after spiking, which
will speed the filtration process somewhat.
7.1.4 Prepare matrix spikes by adding listed matrix spike standards to
representative sample replicates in their original containers . The frequency with which matrix
spikes are prepared and analyzed is described in Chapter One or as part of the determinative
method. Mix the matrix spike samples as described in Sec.7.1.3 and allow to stand.
7.1.5 If cleanup procedures are to be employed that result in the loss of extract,
adjust the amount of surrogate and spiking cocktail(s) accordingly. In the case of Method
3640, Gel Permeation Cleanup, double the amount of standards to compensate for the loss
of one half of the extract concentrate when loading the GPC column.
7.2 pH adjustment
Check thepHof the sample with wide-range pH paper and, if necessary, adjust the pH to the
range listed below. If pH adjustment is required, this step should be performed in the original sample
container to ensure that analytes are not lost in precipitates or flocculated material. Any adjustment
of the sample pH should take place after the surrogates and matrix spiking compounds are added,
so that they are affected by the pH in the same manner as the target analytes.
NOTE: The efficiency of solid-phase extraction of acid herbicide compounds is greatly affected by
pH. If acid herbicides are to be extracted from TCLP leachates or other samples, adjust
the pH to 1.0 before extraction.
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Analvte Group
Extraction pH
Phthalate esters
5-7

Organochlorine pesticides
5-9

Polychlorinated biphenyls (PCBs)
5-9

Organophosphorus pesticides
as received

Nitroaromatics and nitramines
as received

TCLP leachates containing organochlorine pesticides
as produced
by TCLP
TCLP leachates containing semivolatiles
as produced
by TCLP
TCLP leachates containing phenoxyacid herbicides
1.0

7.3 Setting up the extraction apparatus
7.3.1 Assemble a manifold for multiple disk extractions (Figure 1) using 47-mm or
90-mm extraction disks. Use a filter flask with the standard filter apparatus for single
extractions. The solid-phase disks that are generally appropriate for each group of analytes
are listed below, and in Table 1.
Analvte Group Disk Medium
Phthalate esters C18
Organochlorine pesticides C18
Polychlorinated biphenyls (PCBs) C18
Organophosphorus pesticides SDB-RPS
Nitroaromatics and nitramines SDB-RPS
TCLP leachates containing organochlorine pesticides SDB-XC
TCLP leachates containing semivolatiles SDB-XC
TCLP leachates containing phenoxyacid herbicides SDB-XC
For nitroaromatics and nitramines, samples also may be extracted using an SPE
cartridge. Assemble the cartridge apparatus according to the manufacturer's instructions,
using Porapak R, or equivalent, SPE cartridges, and proceed to Sec. 7.6.
7.3.2 If samples contain significant quantities of particulates* the use of a filter aid
or prefitter is advisable for disk extractions. Empore™ Filter Aid 400, Whatman GMF 150, or
equivalent prefilters are recommended.
7.3.2.1 Pour about 40 g of Filter Aid 400 onto the surface of the disk after
assembling the standard filter apparatus.
7.3.2.2 Alternatively, place the Whatman GMF 150 on top of the extraction
disk prior to clamping the glass reservoir into the standard filter apparatus.
7.3.2.3 Do noj add the filter aid if using the cartridge extraction procedure
for nitroaromatics and nitramines.
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7.4 Washing the extraction apparatus
Prior to use, the extraction disks must undergo two separate washing steps, usually with
different solvents. The steps involved in washing the extraction apparatus before use depend on
the analytes of interest and the sample matrix.
7.4.1 First washing step
The following table illustrates the solvents recommended for the first washing step.
Analvte Group
Phthalate esters
Organochlorine pesticides
Polychlorinated biphenyls (PCBs)
Organophosphorus pesticides
NHroaromatics and nitramines
TCLP leachates containing organochlorine pesticides
TCLP leachates containing semivolatiles
TCLP leachates containing phenoxyacid herbicides
1st solvent wash volume
20 mL methylene chloride
20 mL methylene chloride
20 mL methylene chloride
5 mL acetone
5 mL acetonitrile
5 mL acetone
5 mL acetone
5 mL acetonitrile
Wash the extraction apparatus and disk with the volume of the solvent listed above by rinsing
the solvent down the sides of the glass reservoir. Pull a small amount of solvent through the
disk with a vacuum. Turn off the vacuum and allow the disk to soak for about one minute. Pull
the remaining solvent through the disk and allow the disk to dry.
7!4.1.1 When using a filtration aid, adjust the volume of ail wash solvents
so the entire filtration bed is submerged.
7.4.1.2 in subsequent conditioning steps, volumes should be adjusted so
that a level of solvent is always maintained above the entire filter bed.
7.4.2 Second washing step
The following table illustrates the solvents recommended for the second washing step.
Analvte Group
Phthalate esters
Organochlorine pesticides
Polychlorinated biphenyls (PCBs)
Organophosphorus pesticides
NHroaromatics and nitramines
TCLP leachates containing organochlorine pesticides
TCLP leachates containing semivolatiles
TCLP leachates containing phenoxyacid herbicides
2nd solvent wash volume
10 mL acetone
10 mL acetone
not required
5 mL methanol
15 mL acetonitrile
5 mL ethyl acetate
5 mL ethyl acetate
not required
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7.5
Disk conditioning
"Hie extraction disks are composed of hydrophobic materials which will not allow water to pass
unless they are pre-wetted with a water-miscible solvent before being used for sample extraction.
This step is referred to as conditioning, and the solvent used is dependent on the analytes of
interest. The following table illustrates the solvents recommended for specific groups of analytes.
NOTE: Beginning with the conditioning step, it is CRITICAL that the disk NOT go dry until after the
extraction steps are completed. Should a disk accidentally go dry during the conditioning
steps, the conditioning steps for that disk must be repeated prior to adding .the sample.
AnaMe Group
Fhthalate esters
Organochlorine pesticides
Polychlorinated biphenyls (PCBs)
Organophosphoms pesticides
NHroaromatics and nitramines
TCLP leachates containing organochlorine pesticides
TCLP leachates containing semivolatiles
TCLP leachates containing phenoxyacid herbicides
Conditioning steps
20 mL methanol, soak 1 min,
20 mL reagent water
20 mL methanol, soak 1 min,
20 mL reagent water
20 mL methanol, soak 1 min,
20 mL reagent water
5 mL methanol, soak 1 min,
20 mL reagent water
15 mL acetonitrile, soak 3 min
30 mL reagent water
5 mL methanol soak 1 min,
15 mL reagent water
5 mL methanol soak 1 min,
15 mL reagent water
5 mL methanol soak 1 min,
15 mL reagent water
7.5.1 Add the conditioning solvent to the extraction apparatus. Apply a vacuum
until a few drops of solvent pass through the disk, ensuring that the disk is soaked with the
solvent. Turn off the vacuum and allow the disk to soak in the solvent for the time specified
above.
7.5.2 When using a filtration aid, adjust the volume of conditioning solvents so that
the entire filtration bed remains submerged until the extraction is completed.
7.5.3 Once the soaking time is over, apply the vacuum again, drawing all but a thin
layer of solvent through the disk. Stop the vacuum iust before the disk goes dry.
7.5.4 Add the volume of organic-free reagent water listed above and apply vacuum
to draw the water through the disk. Stop the vacuum iust before the disk goes dry, leaving 2-3
mm of water above the surface of the disk.
7.5.5 Proceed to Sec. 7.7 for the sample extraction instructions.
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7.6 Cartridge procedure for nitroaromatics arid nitramines
Aqueous samples to be analyzed for nitroaromatics and nitramines may also be extracted
using the SPE cartridge technique described below. The same sample preparation considerations
discussed in Sec. 7.1 also apply to this procedure.
7.6.1 After assembling the SPE cartridge In the extraction apparatus (see Sec.
7.3.1), wash the cartridge with 10 mL of acetonitrile, using gravity flow. Do riot allow the
cartridge to go dry.
7.6.2 When only a thin layer of solvent remains above the sorberit bed in the
cartridge, add 30 mL of reagent water to the cartridge and allow it to flow through the sorbent
bed under gravity flow. Stop the flow just before the cartridge goes dry.
7.6.3 Attach a connector to the top of the cartridge. The other end of the connector
should be fitted with fle)dble PTFE tubing long enough to reach into the sample bottle or other
container (e.g., a beaker) holding the sample.
7.6.4 Turn on the vacuum, and draw the sample through the cartridge at a rate of
about 10 mUmin, until all of the sample has passed through the cartridge. As particulate
matter plugs the cartridge and slows the flow, increase the vacuum to maintain a reasonable
flow rate.
7.6.5 Once all of the sample has been pulled through the cartridge, shut off the
vacuum and add 5 mL of reagent water to the cartridge. Allow the reagent water to pass
through the cartridge under gravity flow, if practical, or apply a vacuum to complete the
process. Shut off the flow once the water has been drawn through the cartridge.
7.6.6 Method Wanks and matrix spike aliquots (Sec. 7.1) are handled in the same
manner as the samples.
7.6.7 Proceed with sample elution, as described in Sec. 7.9.
7.7 Sample extraction using SPE disks
7.7.1 Pour the sample into the reservoir and, under full vacuum, filter it as quickly
as the vacuum will allow (at least 10 minutes). Transfer as much of the measured volume of
water as possible.
NOTE: With heavily particle-laden samples, allow the sediment in the sample to settle and
decant as much liquid as is practical into the reservoir. After most of the aqueous
portion of the sample has passed through the disk, swirf the portion of the sample
containing sediment and add it to the reservoir. Use additional portions of organic-
free reagent water to transfer any remaining particulates to the reservoir.
Particulates must be transferred to the reservoir before all of the aqueous sample
has passed through the disk.
7.7.2 After the sample has passed through the solid-phase media, dry the disk by
maintaining vacuum for about 3 minutes. Method blanks and matrix spike aliquots (Sec. 7.1)
are handled in the same manner as the samples.
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7.8 Elution of the analytes from the disk
The choice of elution solvent is critical to the success of solid-phase extraction. The
recommended elution solvent for each group of analytes is listed below.
Analvte Group
Phthalate esters
Organochlorine pesticides
Polyehlorinated biphenyls (PCBs)
Organophosphorus pesticides
Nitroaromatics and nitramines
TCLP leachates containing organochlorine
pesticides
TCLP leachates containing semivolatiles
TCLP leachates containing phenoxyacid
herbicides
Sample elution steps
5 mL acetone, soak 15-20 sec. Rinse bottle
with 15 mL acetonitrile and add to disk.
5 mL acetone, soak 15-20 sec. Rinse bottle
with 15 mL methylene chloride and add to disk.
5 mL acetone, soak 15-20 sec. Rinse bottle
with 20 mL acetonitrile and add to disk.
0.6 mL acetone, soak 1 min. Rinse bottle with
5 mL MTBE and add to disk. Repeat bottle
rinse twee more.
5 mL acetonitrile, soak 3 min.
Rinse bottle with 4 mL acetone and add to disk.
Rinse glassware with 2 mL acetone and add to
disk. Soak 1 min. Rinse bottle twice with 5 mL
ethyl acetate and add to disk.
Rinse bottle with 4 mL acetone and add to disk.
Rinse glassware with 2 mL acetone and add to
disk. Soak 1 min. Rinse bottle twice with 5 mL
ethyl acetate and add to disk.
Rinse bottle with 5 mL acetonitrile and add to
disk. Soak 1 min. Rinse bottle twice more with
5 mL acetonitrile and add to disk.
7.8.1 Remove the entire standard filter assembly (do not disassemble) from the
manifold and insert a collection tube. The collection tube should have sufficient capacity to
hold all of the elution solvents. The drip tip of the filtration apparatus should be seated
sufficiently below the neck of the collection tube to prevent analyte loss due to splattering when
vacuum is applied. When using a filter flask for single extractions, empty the water from the
flask before inserting the collection tube.
7.8.2 An initial elution with a water-miscible solvent, i.e., acetone or acetonitrile,
improves the recovery of analytes trapped in water-filled pores of the sorbent. Use of a water-
miscible solvent is particularly critical when methylene chloride is used as the second elution
solvent. With the collection tube in place, add the volume of elution solvent listed above to the
extraction apparatus. Allow the solvent to spread out evenly across the disk (or inert filter)
then quickly turn the vacuum on and off to pull the first drops of sovlent through the disk. Allow
the disk to soak for 15 to 20 seconds before proceeding to Sec. 7.8.3
7.8.3 Rinse the sample bottle and/or glassware that held the sample with the
second solvent listed above and transfer the solvent rinse to the extraction apparatus. As
needed, use a disposable pipette to rinse the sides of the extraction apparatus with solvent
from the bottle.
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7.8.4 Draw about half of the solvent through the disk and then release the vacuum.
Allow the remaining elution solvent to soak the disk and particulates for about one minute
before drawing the remaining solvent through the disk under vacuum. When using a filtration
aid, adjust the volume of elution solvent so that the entire filtration bed is initially submerged.
7.8.5 Repeat the bottle rinsing step as listed in the table above, continuing to apply
vacuum and collecting the solvent in the tube.
7.9 Eluting the nitroaromatics and nitramines from the cartridge
Once the reagent water has passed through the column, place a collection tube under the
cartridge. Add 5 mL of acetonitrile to the top of the cartridge and allow it to pass through the
cartridge under gravity flow, collecting the solvent in the collection tube. Measure the volume of
acetonitrile recovered from the cartridge.
7.10 K-D concentration technique
Where necessary to meet the sensitivity requirements, sample extracts may be concentrated
to the final volume necessary for the determinative method and specific application, using the K-D
technique or nitrogen evaporation.
7.10.1 Assemble a Kudema-Danish (K-D) concentrator by attaching a 10-mL
concentrator tube to an appropriately sized evaporation flask.
7.10.2 Dry the combined extracts in the collection tube (Sees. 7.8 and 7.9) by
passing them through a drying column containing about 10 g of anhydrous sodium sulfate.
Collect the dried extract in the K-D concentrator. Use acidified sodium sulfate (see Method
8151) if acidic analytes are to be measured.
7.10.3 Rinse the collection tube and drying column into the K-D flask with an
additional 20-mL portion of solvent in order to achieve a quantitative transfer.
7.10.4 Add one or two clean boiling chips to the flask and attach a three-ball Snyder
column. Attach the solvent vapor recovery glassware (condenser and collection device, see
Sec. 4.6) to the Snyder column of the K-D apparatus, following the manufacturer's instructions.
Pre-wet the Snyder column by adding about 1 mL of methylene chloride (or other suitable
solvent) to the top of the column. Place the K-D apparatus on a hot wafer bath (15 - 20°C
above the boiling point of the solvent) so that the concentrator tube is partially immersed in the
hot water and the entire lower rounded surface of the flask is bathed with hot vapor. Adjust
the vertical position of the apparatus and the water temperature as required to complete the
concentration in 10 - 20 minutes. At the proper rate of distillation the balls of the column will
actively chatter, but the chambers will not flood. When the apparent volume of liquid readies
1 mL, remove the K-D apparatus from the water bath and allow it to drain and cool for at least
10 minutes.
7.10.4.1 If a solvent exchange is required (as indicated in Table 1),
momentarily remove the Snyder column, add 50 mL of the exchange solvent and a new
boiling chip.
7.10.4.2 Reattach the Snyder column. Concentrate the extract, raising the
temperature of the water bath, if necessary, to maintain a proper distillation rate.
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7.10.5 Remove the Snyder column. Rinse the K-D flask and the lower joints of the
Snyder column into the concentrator tube with 1 - 2 mL of solvent. The extract may be further
concentrated by using one of the techniques outlined in Sec. 7.11, or adjusted to a final volume
of 5.0 - 10.0 mL using an appropriate solvent (Table 1).
7.11 If further concentration is required, use either the micro-Snyder column technique
(7.11.1) or nitrogen evaporation technique (7.11.2).
7.11.1 Micro-Snyder column technique
7.11.1.1 Add a fresh dean boiling chip to the concentrator tube and attach
a two-ball micro-Snyder column directly to the concentrator tube. Attach the solvent
vapor recovery glassware (condenser and collection device) to the micro-Snyder
column of the K-D apparatus, following the manufacturer's instructions. Pre-wet the
Snyder column by adding 0.5 mL of methylene chloride or the exchange solvent to the
top of the column. Race the micro-concentration apparatus in a hot water bath so that
the concentrator tube is partially immersed in the hot water. Adjust the vertical position
of the apparatus and the water temperature, as necessary, to complete the
concentration in 5 • 10 minutes. At the proper rate of distillation the balls of the column
will actively chatter, but the chambers will not flood.
7.11.1.2 When the apparent volume of liquid reaches 0.5 mL, remove the
apparatus from the water bath and allow it to drain and cool for at least 10 minutes.
Remove the Snyder column and rinse its lower joints into the concentrator tube with
0.2 mL of solvent. Adjust the final extract volume to 1.0 - 2.0 mL.
7.11.2 Nitrogen evaporation technique
7.11.2.1 Place the concentrator tube in a warm bath (30 °C) and evaporate
the solvent volume to 0.5 mL using a gentle stream of clean, dry nitrogen (filtered
through a column of activated carbon).
CAUTION: New plastic tubing must not be used between the carbon trap and the
sample, since it may introduce phthalate interferences.
7.11.2.2 Rinse down the internal wall of the concentrator tube several times
with solvent during the concentration. During evaporation, position the concentrator
tube to avoid condensing water into the extract. Under normal procedures, the extract
must not be allowed to become dry.
CAUTION: When the volume of solvent is reduced below 1 mL, some semivolatife
analytes such as cresols may be lost.
7.12 The extract may now be subjected to cleanup procedures or analyzed for the target
analytes using the appropriate determinative technique(s). If further handling of the extract will not
be performed immediately, stopper the concentrator tube and store in a refrigerator. If the extract
will be stored longer than 2 days, it should be transferred to a vial with a PTFE-lined screw-cap, and
labeled appropriately.
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8.0 QUALITY CONTROL
8.1 Arty reagent blanks or matrix spike samples should be subjected to exactly the same
analytical procedures as those used for actual samples.
8.2 Refer to Chapter One for general quality control procedures and Method 3500 for
specific QC procedures for extraction and sample preparation.
9.0 METHOD PERFORMANCE
Refer to the determinative methods listed In Sec. 1.1 for performance data related to solid-
phase extraction.
10.0 REFERENCES
1. Lopez-Avila, V., Beckert, W., et. a!., "Single Laboratory Evaluation of Method 8060 - Phthalate
Esters", EPA/600/4-89/039.
2. Tomkins, B.A., Merriweather, R., et. al., "Determination of Eight Organochlorine Pesticides at
Low Nanogram/Liter Concentrations in Groundwater Using Filter Disk Extraction and Gas
Chromatography", JAOAC International, 75(6). pp. 1091-1099 (1992).
3. Markell, C., "3M Data Submission to EPA," letter to B. Lesnik, June 27,1995.
4. Jenkins, T. F., Thome, P. G.f Myers, K. F., McCormick, E. F., Parker, D. E., and B. L. Escalon
(1995). Evaluation of Clean Solid Phases for Extraction of Nitroaromatics and Nitramines from
Water. USACE Cold Regions Research and Engineering Laboratory, Special Report 95-22.
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TABLE 1
SPECIFIC EXTRACTION CONDITIONS FOR VARIOUS DETERMINATIVE METHODS
Determinative Method
Extraction pH
Disk Medium8
Elution Solvent
Exchange Solvent
Final Extract
Volume for
Analysis (mL)b
8061 (phthalate esters)
8081 (organochlorine
pesticides)
8082 (PCBs)
8141 (organophosphorus
pesticides)
8330 (nitroaromatics and
nitramines)
TCLP pesticides (8081)
TCLP semivolatiles (8270)
TCLP phenoxyacid
herbicides (8321)
5-7
5-9
5-9
as received
as received
as produced by TCLP
as produced by TCLP
1.0
Cis
Cis
Cis
SDB-RPS
SDB-RPS
SDB-XC
SDB-XC
SDB-XC
acetonitrile
methylene chloride
methylene chloride
MTBE
acetonitrile
ethyl acetate
ethyl acetate
acetonitrile
hexane 10.0
hexane 10.0
hexane 10.0
hexane 10.0
acetonitrile 10.0
hexane 10.0
methylene chloride 1.0
hexane 10.0
a SDB has a greater capacity than C18 and a greater affinity for more analytes but they may be more difficult to elute.
b For methods where the suggested final extract volume is 10.0 mL, the volume may be reduced to as low as 1.0 mL to achieve lower detection
limits.
3535A-16
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Reservoir
mnii'
•damp
,TM
Empore
Extraction Disk
Base
(Fritted or with Screen)
Dn'p Tube
Filter Flask or Manifold
FIGURE 1
DISK EXTRACTION APPARATUS
3535A - 17
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METHOD 3535A
SOLID-PHASE EXTRACTION (SPE)
Start
No
1 f
7.1 Pripir* tha aarnpta.
1,2 Adjust tha pH.
7,7 Parform disk a*tr»etton.
7,6 Prapara eartridga and
lead aampla on eartridga.
7,5 Condition tha dtak.
7.9 Eluta targat compounds
from eartridga.
7-10 Parlour? KD concentration.
7.8 Eluta analytaa from tha disk
7.4 Parfoim 2-stap withing.
7.3 Salact appropnat* disk
lor cartridge} and >at-up
apparatus
additional
concantration
naeaaaary?
Ya*
7.11 Parform Micro-Snydar or
Nj avaporation.
7.12 Go to appreprtata claanup
mathod or larnpia analyai*
taehniqua.
3535A - 18
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METHOD 3545A
PRESSURIZED FLUID EXTRACTION (PFB)
1.0 SCOPE AND APPLICATION
1.1 Method 3545 is a procedure for extracting water insoluble or slightly water soluble
organic compounds from soils, clays, sediments, sludges, and waste solids. The method uses
elevated temperature (100 - 180°C) and pressure (1500 - 2000 psi) to achieve analyte recoveries
equivalent to those from Soxhlet extraction, using less solvent and taking significantly less time than
the Soxhlet procedure. This procedure was developed and validated on a commercially-available,
automated extraction system.
1.2 This method is applicable to the extraction of semivolatile organic compounds,
organophosphorus pesticides, organochlorine pesticides, chlorinated herbicides, PCBs, and
PCDDs/PCDFs, which may then be analyzed by a variety of chromatographic procedures.
1.3 This method has been validated for solid matrices containing 250 to 12,500 pg/kg of
semivolatile organic compounds, 250 to 2500 |jg/kg of organophosphorus pesticides, 5 to 250 jjg/kg
of organochlorine pesticides, 50 to 5000 ytg/kg of chlorinated herbicides, 1 to 1400 pg/kg of PCBs,
and 1 to 2500 ng/kgof PCDDs/PCDFs. The method may be applicable to samples containing these
analytes at higher concentrations and may be employed after adequate performance has been
demonstrated for the concentrations of interest (see Method 3500, Sec. 8.0).
1.4 This method is applicable to solid samples only, and is most effective on dry materials
with small particle sizes. Therefore, waste samples must undergo phase separation, as described
in Chapter Two, and only the solid phase material is to be extracted by this procedure. If possible,
soil/sediment samples may be air-dried and ground to a fine powder prior to extraction. Alternatively,
if the loss of analytes or during drying is a concern, soil/sediment samples may be mixed with
anhydrous sodium sulfate or pelletized diatomaceous earth. (Drying and grinding samples containing
PCDDs/PCDFs is not recommended, due to safety concerns). The total mass of material to be
prepared depends on the specifications of the determinative method and the sensitivity required for
the analysis, but 10 - 30 g of material are usually necessary and can be accommodated by this
extraction procedure.
1.5 This method is restricted to use by or under the supervision of trained analysts. Each
analyst must demonstrate the ability to generate acceptable results with this method.
2.0 SUMMARY OF METHOD
2.1 Samples are prepared for extraction either by air drying the sample, or by mixing the
sample with anhydrous sodium sulfate or pelleteed diatomaceous earth. The sample is then ground
and loaded into the extraction cell. Drying and grinding samples containing PCDDs/PCDFs is not
recommended, due to safety concerns. Grinding may also be a concern for other more volatile
analytes. (See Sec. 7.1).
2.2 The extraction cell containing the sample is heated to the extraction temperature (see
Sec. 7.8), pressurized with the appropriate solvent system, and extracted for 5 minutes (or as
recommended by the instrument manufacturer). Multiple extractions are recommended for some
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groups of analytes. The solvent systems used for this procedure vary with the analytes of interest
and are described in Sec. 5.5.
2.3 The solvent is collected from the heated extraction vessel and allowed to cool.
2.4 The extract may be concentrated, if necessary, and, as needed, exchanged into a
solvent compatible with the cleanup or determinative step being employed.
3.0 INTERFERENCES
3.1 Refer to Method 3500.
3.2 If necessary, Florisil and/or sulfur cleanup procedures may be employed,
cases, proceed with Method 3620 and/or Method 3660,
3.3 Samples for PCDD/PCDF analysis should be subjected to the various
procedures described in the determinative methods (8280 and 8290).
4.0 APPARATUS AND MATERIALS
4.1 Pressurized fluid extraction device
4.1.1 Dionex Accelerated Solvent Extractor or Supelco SFE-400 with appropriately-
sized extraction cells. Currently, cells are available that will accommodate 10-g, 20-g and 30-g
samples. Cells should be made of stainless steel or other material capable of withstanding the
pressure requirements (2000+ psi) necessary for this procedure.
4.1.2 Other system designs may be employed, provided that adequate performance
can be demonstrated for the analytes and matrices of interest.
4.2 Apparatus for determining percent dry weight
4.2.1 Oven-drying
4.2.2 Desiccator
4.2.3 Crucibles - porcelain or disposable aluminum
4.3 Apparatus for grinding - capable of reducing particle size to < 1 mm.
4.4 Analytical balance - capable of weighing to 0.01 g.
4.5 Vials for collection of extracts - 40-mL or 60-mL, pre-cleaned, open top screw-cap with
PTFE-lined silicone septum (Dionex 049459, 049460, 049461, 049462 or equivalent).
4.6 Filter disk -1.91 cm, Type D28 (Whatman 10289356, or equivalent).
4.7 Cell cap sealing disk (Dionex 49454,49455, or equivalent).
In such
cleanup
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5.0 REAGENTS
5.1 Reagent grade chemicals shall be used in all tests. Unless otherwise indicated, it is
intended that all reagents shall conform to the specifications of the Committee on Analytical
Reagents of Vie American Chemical Society, where such specifications are available. Other grades
may be used, provided it Is first ascertained that the reagent is of sufficiently high purity to permit its
use without lessening the accuracy of the determination.
5.2 Organic-free reagent water. All references to water in this method refer to organic-free
reagent water, as defined in Chapter One.
5.3 Drying agents
5.3.1 Sodium sulfate (granular anhydrous), Na2S04.
5.3.2 Pelletized diatomaceous earth.
5.3.3 The drying agents should be purified by heating at 400°C for 4 hours in a
shallow tray, or by extraction with methylene chloride. If extraction with methylene chloride is
employed, then a reagent blank should be prepared to demonstrate that the drying agent is
free of interferences.
5.3.4 Quartz sand. Although not strictly a drying agent, dean sand may be used
to facilitate grinding of some sample matrices, to fill void volumes in the extraction cell, and to
increase the flow of solvent through the sample. It may be prepared as described in Sec.
5.3.3.
5.4 Acids
5.4.1 Phosphoric acid solution (see Sec. 5.5.5). Prepare a 1:1 (v/v) solution of 85%
phosphoric acid (H3PQ4) in organic-free reagent water.
5.4.2 Trifluoroaceticacid solution (see Sec. 5.5.5). Prepare a 1% (v/v) solution of
trifluoroacetic add in acetonitrile.
5.4.3 Glacial acetic acid (see Sec. 5.5.6).
5.5 Extraction solvents
The extraction solvent to be employed depends on the analytes to be extracted, as described
below. All solvents should be pesticide quality or equivalent. Solvents may be degassed prior to
use.
5.5.1 Organochlorine pesticides may be extracted with acetone/hexane (1:1, v/v),
CH3COCH3/C6H14 or acetone/methylene chloride (1:1,v/v), CH3COCH.j/CH2Cl2 .
5.5.2 Semivolatile organics may be extracted with acetone/methylene chloride (1:1,
v/v), CH3COCH3/CH2CI2 or acetone/hexane (1:1, v/v), CH3COChyc6H14.
5.5.3 PCBs may be extracted with acetone/hexane (1:1, v/v), CHjCOCI-yCgH^ or
acetone/methylene chloride (1:1, v/v), CHjCOChyCH^ or hexane, CgH,4.
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5.5.4 Organophosphoais pesticides may be extracted with methylene chloride,
CHjCIj or acetone/methylene chloride (1:1, v/v), CH3COCH3/CH2CI2.
5.5.5 Chlorinated herbicides may be extracted with an acetone/methylene
chloride/phosphoric acid solution (250:125:15, v/v/v), CHgCOCH^CHjCyHaPO,,, or an
acetone/methylene chloride/trifluoroacetic add solution (250:125:1, v/v/v),
CHsCCXJKVCH^JIj/CFaCOOH. (If the second option is used, the trifluoroacetic acid solution
should be prepared by mixing 1% trifluoroacetic acid in acetonitrile.) Make fresh solutions
before each batch of extractions.
5.5.6 PCDDs/PCDFs may be extracted with toluene, C6H5CH3. Fly ash samples to
be extracted for PCDDs/PCDFs may be extracted with a toluene/acetic acid solution (5% v/v
glacial acetic acid in toluene) in lieu of the HCI pretreatment described in Methods 8280 and
8290.
5.5.7 Other solvent systems may be employed, provided that the analyst can
demonstrate adequate performance for the analytes of interest in the sample matrix (see
Method 3500, Sec. 8.0).
CAUTION: For best results with very wet samples (e.g., *30% moisture), reduce or eliminate
the quantity of hydrophilic solvent used.
5.6 High-purity gases such as nitrogen, carbon dioxide, or helium are used to purge and/or
pressurize the extraction cell. Follow the instrument manufacturer's recommendation for the choice
of gases.
6.0 SAMPLE COLLECTION, PRESERVATION, AND HANDLING
See the introductory material to this chapter, Organic Analysis, Sec. 4.1, and Method 3500.
7.0 PROCEDURE
7.1 Sample preparation
7.1.1 Sediment/soil samples - Decant and discard any water layer on a sediment
sample. Mix the sample thoroughly, especially composited samples. Discard any foreign
objects such as sticks, leaves, and rocks. Air dry the sample at room temperature for 48 hours
in a glass tray or on hexane-rinsed aluminum foil. Alternatively, mix the sample with an equal
volume of anhydrous sodium sulfate or pelletized diatomaceous earth until a free-flowing
powder is obtained.
NOTE: Dry, finely-ground soil/sediment allows the best extraction efficiency for nonvolatile,
nonpolarorganics, e.g., 4,4'-DDT, PCBs, etc. Air-drying may not be appropriate for
the analysis of the more volatile organochkjrine pesticides (e.g., the BHCs) or the
more volatile of the semivolatile organics because of losses during the drying
process. Drying of samples for PCDDs/PCDFs is not generally recommended, due
to safety concerns with samples containing these analytes. The use of sodium
sulfate as a drying agent can lead to clogging of the frits in the cell with recrystallized
sodium sulfate. (See "Caution" following Sec. 5.5.6.)
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7.1.2 Waste samples - Multiphase waste samples must be prepared by the phase
separation method in Chapter Two before extraction. This extraction procedure is for solids
only.
7.1.3 Dry sediment/soil and dry waste samples amenable to grinding - Grind or
otherwise reduce the particle size of the waste so that it either passes through a 1-mm sieve
or can be extruded through a 1-mm hole. Disassemble grinder between samples, according
to manufacturer's instructions, and decontaminate with soap and water, followed by acetone
and hexane rinses. Grinding of samples for PCDDs/PCDFs is not generally recommended,
due to safety concerns with samples containing these analytes.
NOTE: The note in Sec. 7.1.1 also applies to the grinding process.
7.1.4 Gummy, fibrous, or oily materials not amenable to grinding should be cut,
shredded, or otherwise reduced in size to allow mixing and maximum exposure of the sample
surfaces for the extraction. The analyst may add anhydrous sodium sulfate, pelletized
diatomaceous earth, sand, or other clean, dry reagents to the sample to make it more
amenable to grinding.
7.1.5 Solid samples for PCDD/PCDF analysis are generally carefully mixed with
clean sand and a drying agent such as diatomaceous earth or sodium sulfate, breaking up
lumps with a spatula or other suitable tool.
7.1.6 Fly ash samples may be pretreated with an HCI solution prior to extraction
(See Sec. 7 of Method 8280 or 8290). Alternatively, they may be extracted with the
toluene/acetic acid solution described in Sec. 5.5.6.
7.2 Determination of percent dry weight - When sample results are to be calculated on a
dry weight basis, a second portion of sample should be weighed at the sarnie time as the portion
used for analytical determination.
WARNING: The drying oven should be contained in a hood or vented. Significant laboratory
contamination may result from drying a heavily contaminated sample.
7.2.1 Immediately after weighing the sample for extraction, weigh 5 -10 g of the
sample into a tared crucible. Dry this aliquot overnight at 105°C. Allow to cool in a desiccator
before weighing. Calculate the % dry weight as follows:
% dry weight = 9 of drY sample x 1Q0
g of sample
7.3 Grind a sufficient weight of the dried sample from Sec. 7.1 to yield the sample weight
needed for the determinative method (usually 10-30 g). Grind the sample until it passes through
a 10 mesh sieve. Grinding of samples for PCDDs/PCDFs is not generally recommended, due to
safety concerns with samples containing these analytes.
7.4 Transfer the ground sample to an extraction cell of the appropriate size for the aliquot.
Generally, an 11-mL cell will hold about 10 g of material, a 22-mL cell will hold about 20 g of
material, and a 33-mL cell will hold about 30 g of material. The weight of a specific sample that a
cell will contain depends on the bulk density of the sample and the amount of drying agent that must
be added to the sample in order to make it suitable for extraction. Analysts should ensure that the
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sample aliquot extracted is large enough to provide the necessary sensitivity and choose the
extraction cell size accordingly. Use disposable cellulose or glass fiber filters in the cell outlets.
Clean sand may be used to fill any void volume in the extraction cells.
7.5 Add the surrogates (or labeled Internal standards for PCDDs/PCDFs) listed in the
determinative method to each sample. Add the matrix spike/matrix spike duplicate compounds listed
in the determinative method to the two additional aliquots of the sample selected for spiking.
7.6 Place the extraction cell into the instrument or autosampler tray, as described by the
instrument manufacturer.
7.7 Place a precleaned collection vessel in the instrument for each sample, as described
by the instrument manufacturer. The total volume of the collected extract will depend on the specific
instrumentation and the extraction procedure recommended by the manufacturer and may range
from 0.5 to 1.4 times the volume of the extraction cell. Ensure that the collection vessel is
sufficiently large to hold the extract.
7.8 Recommended extraction conditions
7.8.1 Semivolatiles, organophosphorus pesticides, organochlorine pesticides,
herbicides, and PCBs
Oven temperature: 100°C
Pressure:
Static time:
Flush volume:
Nitrogen purge:
Static Cycles:
1500 - 2000 psi
5 min (after 5 min pre-heat equilibration)
60% of the cell volume
60 sec at 150 psi (purge time may be extended for larger cells)
1
7.8.2 PCDDs/PCDFs
Oven temperature: 150 - 175°C
Pressure: 1500 - 2000 psi
Static time: 5-10 min (after 5 min pre-heat equilibration)
Flush volume: 60 - 75% of the cell volume
Nitrogen purge: 60 sec at 150 psi (purge time may be extended for larger cells)
Static Cycles: 2 or 3
7.8.3 Optimize the conditions, as needed, according to the manufacturer's
instructions. In general, the pressure is not a critical parameter, as the purpose of pressurizing
the extraction cell is to prevent the solvent from boiling at the extraction temperature and to
ensure that the solvent remains in intimate contact with the sample. Any pressure in the range
of 1500 - 2000 psi should suffice.
7.8.4 Once established, the same pressure should be used for all samples
extracted for the same analysis type.
7.9 Begin the extraction according to the manufacturer's instructions. For PCDD/PCDF
extraction, 2 to 3 static extractions are recommended.
7.10 Collect each extract in a clean vial (see Sec. 7.7). Allow the extracts to cool after the
extractions are complete.
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7.11 The extract is now ready for concentration, deanup, or analysis, depending on the
extent of interferants and the determinative method to be employed. Refer to Method 3600 for
guidance on selecting appropriate cleanup methods. Excess water present in extracts may be
removed by filtering the extract through a bed of anhydrous sodium sulfate. Certain cleanup and/or
determinative methods may require a solvent exchange prior to cleanup and/or sample analysis.
7.12 If the phosphoric acid solution in Sec. 5.5.5 is used for the extraction of chlorinated
herbicides, then the extractor should be rinsed by pumping acetone through all the lines of the
system. The use of other solvents for these analytes may not require this rinse step.
8.0 QUALITY CONTROL
8.1 Refer to Chapter One and Method 8000 for guidance on quality control procedures.
Refer to Method 3500 for specific guidance on extraction and sample preparation procedures.
8.2 Before processing any samples, the analyst should demonstrate that all parts of the
equipment in contact with the sample and reagents are interference-free. This is accomplished
through the analysis of a solid matrix method blank (e.g., clean sand). Each time samples are
extracted, and when there is a change in reagents, a method blank needs to be extracted and
analyzed for the compounds of interest. The method blank should be carried through all stages of
the sample preparation and measurement.
8.3 Standard quality assurance practices should be used with this method. Field duplicates
should be collected to validate the precision of the sampling procedures. A matrix spike/matrix spike
duplicate, or matrix spike and duplicate sample analysis, and a laboratory control sample should be
prepared and analyzed with each batch of samples prepared by this procedure, unless the
determinative method provides other guidance.
8.4 When listed in the appropriate determinative method, surrogate standards should be
added to all samples prior to extraction. For PCDDs/PCDFs, the labeled internal standards listed
in the determinative methods should be added to all samples prior to extraction.
9.0 METHOD PERFORMANCE
9.1 Chlorinated pesticides and semivolatile organics
Single-laboratory accuracy data were obtained for chlorinated pesticides and semivolatile
organics at three different spiking concentrations in three different soil types. Spiking concentrations
ranged from 5 to 250 ^ig/kg for the chlorinated pesticides and from 250 to 12500 ^ig/kg for the
semivolatiles. Spiked samples were extracted both by the Dionex Accelerated Solvent Extraction
system and by a Perstorp Environmental Soxtec™ (automated Soxhlet). Extracts were analyzed
either by Method 8270 or Method 8081. Method blanks, spikes and spike duplicates were included
for the law concentration spikes; matrix spikes were included for all other concentrations. The data
are reported in detail in Reference 1, and represent seven replicate extractions and analyses for
each sample. Data summary tables are included in Methods 8270 and 8081.
9.2 Organophosphorus pesticides and chlorinated herbicides
Single-laboratory accuracy data were obtained for organophosphorus pesticides (OPPs) and
chlorinated herbicides at two different spiking concentrations in three different soil types. Spiking
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concentrations ranged from 250 to 2500 MS/kg for the OPPs and from 50 to 5000 |jg/kg for the
chlorinated herbicides. Chlorinated herbicides were spiked with a mixture of the free acid and the
ester (1:1). Spiked samples were extracted both by the Dionex Accelerated Solvent Extractor and
by Soxhlet for the OPPs. Extracts were analyzed by Method 8141. Spiked chlorinated herbicides
were extracted by the Oionex Accelerated Solvent Extractor and by the shaking method described
in Method 8151. Extracts were analyzed by Method 8151. Method blanks, spikes and spike
duplicates were included for the low concentration spikes; matrix spikes were included for all other
concentrations. The data are reported in detail in Reference 2, and represent seven replicate
extractions and analyses for each sample. Data summary tables are included in Methods 8141 and
8151.
9.3 PCBs
Single-laboratory accuracy data were obtained for PCBs from a soil sample with PCB content
certified by NIST (Standard Reference Material, SRM 1939, River Sediment). A PCB-contaminated
soil was purchased from a commercial source. Spiking or certified concentrations ranged from 1 to
1400 M9/kg- Samples were extracted by the Dionex Accelerated Solvent Extractor and by Soxtec™
(Perstorp Environmental). Extracts were analyzed using Method 8082. Method blanks, spikes and
spite duplicates were included. The data are reported in Reference 2, and represent seven replicate
extractions arid analyses for each sample. Data summary tables are included in Method 8082.
9.4 PCDDs/PCDFs
Single-laboratory data were obtained for PCDDs/PCDFs from ground chimney brick, urban
dust, fly ash, a relatively highly contaminated soil sample (EC-2, National Water Research Institute,
Burlington, Ontario, Canada), a low-level sediment sample (HS-2, National Research Council
Institute of Marine Biosciences, Halifax, Nova Scotia, Canada) and various field-contaminated soils
and sediments. Concentrations of PCDDs/PCDFs ranged from low ng/kg to mid pg/kg levels.
Samples were extracted by the Dionex Accelerated Solvent Extractor and by traditional Soxhlet
techniques. Extracts were analyzed by a high resolution mass spectrometry method employing
isotope dilution quantitation. The data are reported in Reference 3. Data summary tables are
included in Method 8290.
10.0 REFERENCES
1. B. Richter, Ezzell, J., and Felix, D., "Single Laboratory Method Validation Report. Extraction
of TCL/PPL (Target Compound List/Priority Pollutant List) BNAs and Pesticides using
Accelerated Solvent Extraction (ASE) with Analytical Validation by GC/MS and GC/ECD";
Document 116064.A, Dionex Corporation, June 16,1994.
2. B. Richter, Ezzell, J., and Felix, D., "Single Laboratory Method Validation Report. Extraction
of TCL/PPL (Target Compound List/Priority Pollutant List) OPPs, Chlorinated Herbicides and
PCBs using Accelerated Solvent Extraction (ASE)". Document 101124, Dionex Corporation,
December 2,1994).
3. B. E. Richter et a/., "Extraction of Polychlorinated Dibenzo-p-Dioxins and Polychlorinated
Dibenzofurans from Environmental Samples Using Accelerated Solvent Extraction (ASE)."
Chemosphem, 34(5-7), pp. 975-987,1997.
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11.0 SAFETY
The use of organic solvents, elevated temperatures, and high pressures in Method 3545
present potential safety concerns in the laboratory. Common sense laboratory practices can be
employed to minimize these concerns. However, the following sections describe additional steps
that should be taken.
11.1 Extraction cells in the oven are hot enough to bum unprotected skin. Allow the cells
to cool before removing them from the oven or use appropriate protective equipment (e.g., insulated
gloves or tongs), as recommended by the manufacturer.
11.2 During the gas purge step, some solvent vapors may exit through a vent port in the
instalment. Follow the manufacturer's directions regarding connecting this port to a fume hood or
other means to prevent release of solvent vapors to the laboratory atmosphere.
11.3 The instrument may contain flammable vapor sensors and should be operated with all
covers in place and doors closed to ensure proper operation of the sensors. If so equipped, follow
the manufacturer's directions regarding replacement of extraction cell seals when frequent vapor
leaks are detected.
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METHOD 3545A
PRESSURIZED FLUID EXTRACTION fPFE)
Start
7.11 Perform
cleanup or
determinative
method.
7.1 Prepare
sample.
7.6 Place extraction
cells into auto
sampling train.
7.2 Determine
sample % dry
weight.
7.7 Load
collection tray.
7.5 Add surrogates
and matrix spiking
standards.
7.4 Transfer ground
sample to an
extraction cell.
7.9 Begin
extraction.
7.3 Grind sufficient
weight of the dried
sample.
7.10 Collect
extracts and
allow to cool.
7.8 Optimize
conditions of
extractor.
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METHOD 3562
SUPERCRITICAL FLUID EXTRACTION OF POLYCHLORINATED BIPHENYLS fPCBs)
AND ORGANOCHLOR1NE PESTICIDES
1.0 SCOPE AND APPLICATION
1.1 Method 3562 describes the extraction with supercritical fluids of polychlorinated
btphenyls (PCBs) and organochlorine pesticides (OCPs) from soils, sediments, fly ash, solid-phase
extraction media, and other solid materials which are amenable to extraction with conventional
solvents. The method is suitable for use with any supercritical fluid extraction (SFE) system that
allows extraction conditions (e.g., pressure, temperature, flow rate) to be adjusted to achieve
separation of the PCBs and OCPs from the matrices of concern. The following compounds have
been extracted by this method during validation studies. Similar compounds not listed should also
be amenable to this extraction.
Compound
CAS Registry No.
IUPAC No.
2,4,4'-Trichlorobiphenyl
7012-37-5
28
2,2',5,5-T etrachlorobiphenyl
35693-99-3
52
2,2',4,5,5*-Pentachlorobiphenyl
37680-73-2
101
2,3,3',4,4-Pentachlorobiphenyl
32598-14-4
105
2,3',4,4',5-Pentachlorobiphenyl
31508-00-6
118
2,2,,3,3',4,4'-Hexachlorobiphenyl
38380-07-3
128
2,2',3,4,4',5-Hexachlorobiphenyl
35065-28-2
138
2,2',3,4',5',6-Hexachlorobiphenyl
38380-04-0
149
2,2,,4,4,,5I5'-Hexachlorobiphenyl
35065-27-1
153
2,3,3\4,4',5'-Hexachlorobiphenyl
38380-08-4
156
2,2',3,3',4,4',5-HeptachIorobiphenyf
35065-30-6
170
2,2')314,4',5,5,-Heptachlorobiphenyl
35065-29-3
180
Aidrin
309-00-2

P-Hexachlorocyclohexane (p-BHC)
319-85-7

5-Hexachlorocyciohexane (5-BHC)
319-86-8

Y-Hexachlorocyclohexane (y-BHC, or Lindane)
58-89-9

a-Chiordane
5103-71-9

4,4'-DDD
72-54-8

4,4'-DDE
72-55-9

4,4-DDT
50-29-3

Dieldrin
60-57-1

Endosulfan II
33213-65-9

Endrin
72-20-8

Endrin aldehyde
7421-93-4

Heptachlor
76-44-8

Heptachlor epoxide
1024-57-3

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1.2 Method 3562 is riot suitable for the extraction of PCBs or organochlorine pesticides
from liquid samples without some treatment of the liquid before introduction into the SFE to
"stabilize" the liquid. Otherwise, the sample may be extruded through the end pieces of the
extraction vessel without the benefit of SFE. The use of soiid-phase extraction (SPE) media is one
way to stabilize a liquid sample and it allows an easy coupling of two selective sample preparation
techniques. The use of large diameter (ca. 90 mm) SPE disks coupled with SFE allows large
volumes of aqueous samples to be prepared without the need for organic solvent elution.
Furthermore, SFE may allow an in-line cleanup to be performed, thus eliminating the need for
separate column cleanup and subsequent solvent concentration steps.
1.3 The extraction conditions listed in this procedure (Sec. 11.6) employed a variable
restrictor and solid trapping media. Other extraction conditions and equipment are acceptable once
appropriate method performance is demonstrated. The method applicability demonstration should
be based on the extraction of a certified reference sample or an environmentally-contaminated
sample, not on spiked soil/solids, whenever possible. It should be noted that there are currently no
"certified" samples for organochlorine pesticides. An authentic, weathered, environmental sample
which has been extracted by a traditional sample preparation technique should be used as the
reference for these compounds.
1.4 This method is restricted to use by or under the supervision of trained analysts. Each
analyst must demonstrate the ability to generate acceptable results with this method.
2.0 SUMMARY OF METHOD
2.1 In order to assure a homogeneous sample and minimum subsampling errors, at least
100 g of sample are homogenized with an equal volume of solid C02 "snow". A1 - 5 g aliquot of this
mixture is packed into a stainless steel SFE extraction vessel. Copper powder may be added to the
cell to remove sulfur from the sample extract. Surrogates and/or internal standards are added to the
portion of the sample in the cell and the cell is placed in the SFE extraction device.
2.2 The sample is extracted using supercritical carbon dioxide with no modifiers. Samples
to be analyzed for PCBs are subjected to a 10-minute static extraction, followed by a 40-minute
dynamic extraction. Samples for organochlorine pesticides are subjected to a 20-minute static
extraction, followed by a 30-minute dynamic extraction.
2.3 The sample extract is trapped on a solid-phase sorbent (Florisil for PCBs and octadecyl
silane for pesticides). The trapping material is then rinsed with solvent to collect the analytes of
interest and reactivate the trapping material for reuse.
2.4 The sample extracts may be subjected to additional cleanup steps (see Method 3600)
and then analyzed by the appropriate determinative methods.
3.0 DEFINITIONS
Dynamic extraction - An application of SFE in which the supercritical extraction fluid flows
through the sample and out of the extraction cell to a collection device during the extraction.
Dynamic extraction is contrasted with static extraction (see below).
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Modifier - A liquid or gaseous component added to the supercritical fluid to change its
extraction capabilities, often through changes in the solvation power of the extraction fluid.
Modifiers may be polar or non polar.
Supercritical fluid - A gas maintained above its critical temperature through the application of
pressure.
Supercritical fluid extraction (SFE) - The use of a gas maintained above its critical temperature
as an extraction fluid.
Static extraction - An application of SFE in which the supercritical extraction fluid is held in the
extraction vessel during the entire procedure, and is then released to a collection device.
Static extraction is contrasted with dynamic extraction (see above).
4.0 INTERFERENCES
4.1 The analyst must demonstrate through the analysis of method blanks that the
supercritical fluid extraction system is free from interferants. To do this, perform a simulated
extraction using an empty extraction vessel and a known amount of C02 under the same conditions
as those used for sample extraction, and determine the background contamination by analyzing the
extract by the determinative method that will be used for sample analysis.
4.2 The extraction vessel(s), the end-frits, the nozzle restrictor(s), and the multi-port
valve(s) may retain solutes whenever high-concentration samples are extracted. Therefore, it is
good practice to clean the extraction system after such extractions. Suspect parts of the system
should be replaced when reagent blanks indicate carryover. At least one reagent blank should be
prepared and analyzed daily when the instrument is in use. Furthermore, reagent blanks should be
prepared and analyzed after each extraction of a high-concentration sample (high part per million
range). If reagent blanks continue to indicate contamination, even after replacement of the
extraction vessel (and the restrictor, if a fixed restrictor system is used), then the multi-port valve
must be cleaned. The operator must be ever vigilant against impurities arising from liquid solvents
and COz itself. Avoid any apparatus, valves, solenoids, and other hardware that contain lubricants
or chlorofluorohydrocarbon materials that can serve as background contaminant sources.
4.3 No modifier was employed in the development of this method for either PCBs or
organochlorine pesticides. Use of a modifier may cause many other problems in these samples,
if the method is modified by the user to include an on-line modifier, or pre-mixed tanks of 0O2 and
modifier, considerable effort must be made to validate this change.
4.4 Refer to Method 3500, Section 3.0, for general extraction interference guidance.
5.0 SAFETY
5.1 SFE involves the use of high pressure gases. Typical SFE systems have maximum
operating pressures of approximately 400 atm (6000 psi). Great care must be taken to ensure that
8//components of the system are capable of withstanding such pressures.
5.2 SFE also involves heating portions of the system above ambient temperature, resulting
in further increases in pressure. The combined effects of the starting pressure and temperature
increase must be taken into account when evaluating the capabilities of system components.
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5.3 SFE devices typically employ gases at high pressure directly from a tank, with no
pressure regulator. In addition to making it difficult to monitor the level of gas in the tank, the lack
of a regulator means that system leaks may involve gases at 200Q psi or more.
5.4 When liquid C02 comes in contact with skin, K can cause "b'ums" because of its low
temperature (-70°C). lums are especially severe when C02 is modified with organic liquids.
5.5 The extraction fluid usually exhausts through an exhaust gas and liquid waste port on
the rear of the panel of the extractor. This port must be connected to a chemical fume hood to
prevent contamination of the laboratory atmosphere.
5.6 Combining modifiers with supercritical fluids requires an understanding and evaluation
of the potential chemical interaction between the modifier and the supercritical fluid, and between
the supercritical fluid and/or modifier and the analyte(s) or matrix.
5.7 When COj is used for cryogenic cooling, typical coolant consumption is 5 Umin, which
results hi a C02 level of 900 ppm for a room of 4.5 m x 3.0 m x 2.5 m, assuming 10 air exchanges
per hour.
6.0 EQUIPMENT AND SUPPLIES
6.1 Supercritical fluid extractor and associated hardware - Any supercritical fluid extraction
system that can achieve the extraction conditions and performance specifications detailed in this
procedure may be used.
WARNING: A safety feature to prevent over-pressurization is required on the extractor. This
feature should be designed to protect the laboratory personnel and the instrument
from possible injuries or damage resulting from equipment failure under high
pressure.
6.1.1 Extraction vessel - Stainless-steel vessel with end fittings with 2 pm frits. Use
the extraction vessel supplied by the manufacturer of the SFE system being used. Fittings
used for the extraction vessel must be capable of withstanding the required extraction
pressures. The maximum operating pressure for most extractors is 400 atm. Pressures above
400 atm, especially at elevated temperatures, are likely to exceed the ratings of standard
chromatography tubing and fittings. Check with the manufacturer of the particular extraction
system and especially the tubing manufacturer for the maximum operating pressure and
temperature for that system. Make sure that the extraction vessels are rated for such
pressures and temperatures.
6.1.2 Restrictor - This method was developed with continuously variable nozzle
restrictors that do not require that the operator take steps to remove water from the sample.
If a fixed restrictor is used, additional validation must be done to verify that moisture from the
sample does not adversely affect the chromatography of the determinative step.
6.1.3 Collection device - This method is based on a solid trap used at sub-ambient
and above ambient temperatures for the different classes of analytes (PCBs vs. OCPs).
However, a liquid (solvent) trap may also be used.
6.1.3.1 Use Florisil, 30-40 ym particle diameter (commonly used in SPE
cartridges), as a solid trap for the PCBs.
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6.1.3.2 For organochiorine pesticides, octadecyi silane (ODS) may be
used as a solid trap, although the use of Florisil is also possible.
6.1.3.3 Analytes may be collected in a small volume of solvent in a
suitable vial, however, great care must be taken to recover the most volatile
compounds. The use of a glass wool plug in the inner tube of the collection vial
improves recoveries. Gas flow must not be so high as to evaporate the collection
solvent to dryness. A 15-mL collection solvent volume is recommended,
6.2 CO2 cylinder balance (optional) - Balances from Scott Specialty Gases, Model 5588D,
equivalent, may be used to monitor the fluid usage. Such a device is useful because C02 tanks
used forSFE are not equipped with regulators, and it is difficult to determine when the tank needs
to be replaced.
6.3 Glass microfiber filter paper disks - Cored out of Whitman QF/F filter paper (Whatman
No. 1825021), or equivalent A disk is placed at both ends of the sample. This ultra-fine filter paper
has good retentive properties for particulate matter down to 0.7 pm and is easy to core. The normal
background is insignificant, but blanks must be run on each batch.
7.0 REAGENTS AND STANDARDS
7.1 CO2 - SFE-grade C02 is absolutely necessary for use in SFE. Aluminum cylinders are
preferred to steel cylinders. The cylinders must be fitted with eductor tubes.
7.2 C02 for cryogenic cooling - Certain parts of some models of extractors (i.e., the
high-pressure pump head and the analyte trap) must be cooled during use. The C02 used for this
purpose must be supplied in tanks with a full-length eductor tube, but need not be SFE-grade. A
low-cost industrial grade is acceptable.
7.3 Reconstitution solvents - Hie reconstitution solvents dispensed by the SFE instruments
that use solid-phase trapping may be the same solvent that is used for liquid trapping. This method
was developed with only sub-ambierit solid trapping. Liquid trapping will work for this method as
well, however the trapping volume is typically ten times larger than that with a solid trap. Further,
the use of liquid trapping will likely require the use of manual Florisil or silica cleanup. These manual
cleanup steps will also require the concentration of the solvent after the cleanup, a step that can be
avoided through use of solid-phase trapping.
7.4 Internal Standards - Refer to the appropriate determinative method for information of
tfje choice of internal standards, where applicable. However, note that for RCBs, certain ethers work
well as internal standards, but do not survive the SFE extraction particularly well.
7.4.1 Internal standards for PCBs - Internal standards that have been evaluated
using this method include PCB 35, PCB 36, PCB 169, 2,4-dichlorobenzyl hexyt ether, 2,4-
dichlorobenzyl heptyl ether, 1,2,3,4-tetrachloronaphthalene, hexabromobenzene, and
octachloronaphthalene.
7.4.2 Internal standard for organochiorine pesticides - PentachloronHrobenzene
7.5. Surrogate standards - Refer to the appropriate determinative method for information
of the choice of surrogates. Surrogates that have been evaluated using this method include
hexabromobenzene, PCB 35, PCB 36, PCB 169, 1,2,3,4-tetrachloronaphthalene, and
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octachioronaphthalene. Prepare a stock solution of 10 mg/mL Apply 150-pL aliquots to the soil
samples within the extraction vessels at the exit end of the flow-through vessels. It has been
observed that a veiy small volume (10 jjL) of a concentrated surrogate mixture often gives poor
recoveries, while adding a larger volume of more dilute surrogate standard to the sample matrix
achieved the expected recoveries.
7.6 Copper powder - Electrolytic grade. Added to samples that contain elemental sulfur.
The powder is pretreated by rinsing 20 g with 150 mL organic-free reagent water, 150 mL acetone,
150 mL of hexane, and drying in a rotary evaporator. The powder is kept under argon or helium until
used. Copper powder must have a shiny bright appearance to be effective. If it has oxidized and
turned dark, it should not be used.
7.7 Sodium Sulfate - Anhydrous (12-60 mesh), Baker Analyzed grade, or equivalent.
7.8 Celtte 545 - 60/80 mesh, J. T. Baker, or equivalent Prepare a reagent blank to assure
that no background contaminants are present.
7.9 Solvents - Used for eluting the analytes of interest from the solid trapping material aid
rinsing the trapping material prior to reuse. All solvents should be pesticide-grade or equivalent.
7.9.1 n-Heptane, C,H16
7.9.2 Methylene chloride, CH2CI2
7.9.3 Acetone, CH3COCH3
7.10 Florisil - Pesticide residue grade.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 See the introductory material to this chapter, Organic Analytes, Sec. 4.1.
8.2 Solid samples to be extracted by this procedure should be collected and stored as any
other solid samples containing semivolatile organics.
9.0 QUALITY CONTROL
9.1 Refer to Chapter One and Method 8000 for specific Quality Control procedures and to
Method 3500 for sample preparation quality control procedures.
9.2 Each time samples are extracted, and when there is a change in reagents, a method
blank should be prepared and analyzed for the compounds of interest as a safeguard against chronic
laboratory contamination. Any method blanks, matrix spike samples, or replicate samples should
be subjected to the same analytical procedures (Sec. 11) as those used on actual samples.
9.3 All instrument operating conditions should be recorded.
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10.0 CALIBRATION AND STANDARDIZATION
There are no calibration or standardization steps associated with this sample extraction
procedure other than establishing the extraction conditions in Sec. 11.6.
11.0 PROCEDURE
11.1 Sample handling - Decant and discard any water layer on a sediment sample. Discard
any foreign objects such as pieces of wood, glass, leaves and rocks.
11.2 Determination of sample % dry weight - In certain cases, sample results are desired
based on dry-weight basis. When such data are desired, a separate portion of sample for this
determination should be weighed out at the same time as the portion used for analytical
determination.
WARNING: The drying oven should be contained in a hood or vented. Significant laboratory
contamination may result from a heavily contaminated hazardous waste sample.
11.2.1 Immediately after weighing the sample aliquot to be extracted, weigh an
additional 5 -10 g aliquot of the sample into a tared crucible. Determine the % dry weight of
the sample by drying overnight at 1Q5°C. Allow to cool in a desiccator before weighing.
11.2.2 Calculate the % dry weight as follows:
% dry weight = 9 of drY sample x 100
g of sample
11.3 Sample grinding and homogenization
NOTE: Sample grinding is a critical step in the SFE process. The soil/solid must be a fine particle
to ensure efficient extraction.
11.3.1 Mix at least 100 grams of sample with an equal volume of C02 solid "snow"
prepared from the extraction grade C02. Place this mixture in a small food-type chopper, and
grind for two minutes. Place the chopped sample on a clean surface and allow the CO^ to
sublime away. As soon as the sample appears free-flowing and solid C02 is no longer visible,
weigh the sample and place it in the extraction vessel. This procedure will ensure the
homogeneity of the sample without loss of the volatile analytes and also retains the original
moisture content of the sample.
11.3.2 Weigh 1.0 to 5.0 g of the homogenized sample from Sec. 11.3.1 into a
pre-cleaned aluminum dish. For samples in the mg/kg (ppm) concentration range, use a 0.1-
gram sample after carefully homogenizing (Sec. 11.3.1) the bulk sample, to avoid sub-sampling
errors.
11.4 For samples known to contain elemental sulfur, use copper powder (Sec. 7.6) to
remove the dissolved sulfur from tire sample and C02 eluant. The copper powder (1 to 2 grams per
sample) can be mixed with the sample in the extraction vessel itself, or packed in a separate vessel
between the extraction vessel and the nozzle (restrictor). The copper addition to samples is a useful
precaution, whether or not one suspects the presence of elemental sulfur. In tests, no adverse
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effect from the addition of copper was observed and it appears that finely divided copper may
enhance the dispersion of C02. If copper powder is added to the samples, it must also be added
to the method blank.
11.5 Packing the extraction cell
The procedure used for a 7.0-mL SFE extraction vessel with sample and copper powder is as
11.5.1 Place a small disk of fiber glass filter paper at the bottom of the extraction
vessel to protect the end frits from particulate matter (this makes the cleanup very easy
between samples and lessens any chance of plugging of the frits).
11.5.2 Place approximately two grams of anhydrous sodium sulfate on top of this
disk in the extraction vessel. Weigh 1.0 gram of solid waste sample into a weighing dish. Add
two grams of electrolytic grade copper powder to the same weighing dish, followed by 7 grams
of anhydrous sodium sulfate. Mix the weighed materia!. Transfer the entire homogeneous
mixture to the extraction vessel on top of the existing small layer of sodium sulfate. Finally,
place a top layer (2 grams) of sodium sulfate on top of the mixture. The densities of the
respective materials are such that this still leaves a small volume at the top of a 7-mL vessel.
These ratios may be adjusted for different sample sizes and vessel sizes, but should be kept
consistent among samples and blanks.
11.5.3 If a surrogate is being added, transfer half the weighed sample to the
extraction vessel. Add 150 jjL of surrogate standard to the sample in the vessel and then add
the remainder of the sample material.
11.5.4 To ensure efficient extraction, fill the extraction vessel completely, avoiding
any dead volume. If any dead volume remains, fill the space with an inert, porous material,
e.g., pre-deaned Pyrex glass wool, Celite, etc.
11.6 Sample extraction conditions
11.6.1 Recommended conditions for PCBs
follows:
11.6.1.1 Extraction conditions
Pressure:
Extraction chamber temperature:
Density:
Extraction fluid composition:
Static equilibration time:
Dynamic extraction time:
Extraction fluid flow rate:
10 minutes
40 minutes
2.5 mL/minute
4417 psi (305 bar)
SOX
0.75 g/mL
CO,
'2
The resultant thimble volume swept is 17.6 times the volume of the cell at
1 bar (this is equivalent to 100 mL of liquid C02 at a reference temperature
of 4.0°C and a density 0.92 g/mL, or 92 g of C02).
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11.6.1.2 Collection conditions (during extraction)
Trap packing: Florisil
Trap temperature: 15-20X
Nozzle temperature: 45-55X (variable restrictor)
11.6.1.3 Reconstitution conditions for coliected extracts
The reconstitution process consists of four rinse steps. The first
rinse is used to elute the analytes of interest from the trapping material. All
four rinse steps are performed with a recommended trap temperature of
38°C, a nozzle temperature of 30°C, and a flow rate of 1.0 mL/min.
Rinse Substep 1:
Rinse solvent n-Heptane
Collected rinse volume: 1.6 mL
Rinse Substep 2:
Rinse solvent n-Heptane
Collected rinse volume: 1.6 mL
This second rinse step is an "insurance rinse". The vial is usually not
analyzed unless there is a need or desire to assure that the entire sample
rinsed in substep 1.
Rinse Substep 3:
Rinse solvent Methylene chloride:Acetone (1:1)
Collected rinse volume: 4.0 mL (to waste)
This third rinse step provides a means of rinsing the solid Florisil trap to
remove interfering compounds such as lipids, hydrocarbons, and PAHs. The
rinse solvent is then discarded.
Rinse Substep 4:
Rinse solvent n-Heptane
Collected rinse volume: 3.0 mL (to waste)
This fourth rinse step provides a means of regenerating the solid Florisil trap
to prepare it (reactivate) for reuse.
11.6.2 Recommended conditions for organochlorine pesticides
11.6.2.1 Extraction conditions
Pressure:
Extraction chamber temperature:
Density:
Extraction fluid composition:
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4330 psi (299 bar)
50 °C
0.87 g/mL
C02
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Static equilibration time:
Dynamic extraction time:
Extraction fluid flow rate:
20 minutes
30 minutes
1.0 mliminute
The resultant thimble volume swept is 4.6 times the volume of the cell at 1
bar (this is equivalent to 30 ml. of liquid C02 at a reference temperature of
4.0°C and a density 0.92 g/mL, or 28 g of COj).
11.6.2.2 Collection conditions (during extraction)
Trap packing:
Trap temperature:
Nozzle temperature:
ODS
20 °C
50 °C (variable restrictor)
11.6.2.3 Reconstitution conditions for collected extracts
The extraction of organochlorine pesticides requires only a single rinse step.
Rinse solvent:
Collected fraction volume:
Trap temperature:
Nozzle temperature:
Rinse solvent flow rate:
n-Hexane
1.3 ml
50° C
30°C (variable restrictor)
2 mL/minute
NOTE: If a fixed restrictor and liquid trapping are used, a restrictor temperature
between 100 and 150°C is recommended.
11.7 Label the extract with the fraction designation and vial number.
11.8 If the rapper powder was not added to the sample prior to loading the cell, additional
sulfur cleanup of the extracts may be required prior to analysis.
11.9 SFE System Maintenance
11.9.1 Depressurize the system following the manufacturer's instructions.
11.9.2 After extraction of an especially "tarry" sample, the end-frits of the extraction
vessel may require extensive cleanup or replacement to ensure adequate flow of extraction
fluid without an excessive pressure drop. In addition, very fine particles may clog the exit frit,
necessitating its replacement. By placing a layer of inert material such as Ceiite or sand
between the sample and the exit frit (and placing disks of filter paper or glass fiber filter on top
of the inert material), this maintenance may be delayed.
11.9.3 Clean the extraction vessel after each sample extraction. The cleaning
procedure depends upon the type of sample. After removing the bulk of the extracted sample
from the extraction vessel, the cell and end-frits should be scrubbed with a solution of
detergent and water using a stiff brush. Placing the parts in an ultrasonic bath with a warm
detergent solution may help. Rinse the parts with organic-free reagent water. Repeat the
ultrasonic bath treatment with either methyl alcohol, or acetone, or both, followed by air drying.
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12.0 DATA ANALYSIS AND CALCULATIONS
There are no calculations explicitly associated with this extraction procedure. See the
appropriate determinative method for calculation of final sample results.
13.0 METHOD PERFORMANCE
13.1 Tables in Method 8081 contain single laboratory performance data for the
organochioiine pesticides using supercritical fluid extraction Method 3562 on an HP 7680. Samples
were analyzed using GC/ELCD. The method was performed using a variable restrictor and solid
trapping material. Three different soil samples were spiked at 5 and 250 ug/kg. Soil 1 (Delphi) is
described as loamy sand, with 2.4% clay, 94% sand, 0.9% organic matter, 3.4% silt, and 0.1%
moisture. Soil 2 (McCarthy) is described as sandy-loam, with 11% clay, 56% sand, 22% organic
matter, 33% silt, and 8.7% moisture. Soil 3 (Auburn) is described as clay loam, with 32% day, 21%
sand, 5.4% organic matter, 46% silt, and 2.2% moisture. Seven replicat# extractions were made
of each soil at the 2 concentrations.
13.2 Tables in Method 8082 contain laboratory performance data for several PCB congeners
using supercritical fluid extraction Method 3562 on an HP 7680. Seven replicate extractions on each
sample were performed. The method was performed using a variable restrictor and solid trapping
material (Florisil). Sample analysis was performed by GC/ECD. The following soil samples were
used for this study:
13.2.1 Two field-contaminated certified reference materials were extracted by a
single laboratory. One of the materials was a lake sediment from Environment Canada (EC-5).
The other material was soil from a dump site and was provided by the National Science and
Engineering Research Council of Canada (EC-1). The average recoveries for EC-5 are based
on the certified value for that sample. The average recoveries for EC-1 are based on the
certified value of the samples or a Soxhlet value, if a certified value was unavailable for a
specific analyte.
13.2.2 Four certified reference materials were extracted by two independent
laboratories. The materials were: a marine sediment from NEST (SRM 1941), a fish tissue
from NIST (SRM 2974), a sewage sludge from BCR European Union(CRM 392), and a soil
sample from BCR European Union (CRM 481). The average recoveries are based on the
certified value of the samples or a Soxhlet value, if a certified value was unavailable for a
specific analyte.
13.2.3 A weathered sediment sample from Michigan (Saginaw Bay) was extracted
by a single laboratory. Soxhlet extractions were carried out on this sample and the SFE
recovery is relative to that for each congener. The average recoveries are based on the
certified value of the samples. Additional data is shown in the tables for some congeners that
were not certified.
14.0 POLLUTION PREVENTION
Extraction of organic compounds using supercritical fluid extraction conforms with EPA's
pollution prevention goals. The volumes of solvent employed, if any, are significantly smaller than
with other extraction procedures. Minimal waste is generated.
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15.0 WASTE MANAGEMENT
Laboratory waste management procedures must be consistent with federal, state, and local
regulations.
16.0 REFERENCES
Gere, D, "Final Deliverables for PCB/OCP SFE Draft Method,9 letter to B. Lesnik, April 15,
1995.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
A flow diagram for the method procedure follows.
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METHOD 3562
SUPERCRITICAL FLUID EXTRACTION OF POLYCHLORINATED BIPHENYLS (PCBs)
AND ORGANOCHLORINE PESTICIDES.

I
11.1 Use appropriate
sample handling
I

11.2 Determine
sample % dry weight.

11.3.1 Grind & homogenize
sample with dry ice
11.3.2 Weigh 1-5 g of
sample
11.4 Is
elemental sulfur
in sample?

11.4 Add
S
copper powder

to sample
11.5 Transfer weighed sample to
extraction vessel and add surrogates
O
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METHOD 3562
(Continued)
Analyte
interest
Stop
11.7 Label extract
11.6 Sample
Extraction
11.9 Perform SFE
system maintenance
11.6.1 Collection
of PCBs
11.6.2 Collection
of organochlorlde
pesticides
11.8 Additional sulfur cleanup
of extract if necessary
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4.2 SAMPLE PREPARATION METHODS
Prior to employing the methods In this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the
allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to
be used by the analyst and the regulated community in making judgements necessary to meet
the data quality objectives or needs for the intended use of the data.
4.2.2 CLEANUP
The following methods are included in this section:
Method 3600C:
Method 3610B:
Method 3611B:
Method 3620B:
Method 3630C:
Method 3640A:
Method 3650B:
Method 3660B:
Method 3665A:
Cleanup
Alumina Cleanup
Alumina Column Cleanup and Separation of
Petroleum Wastes
Florist! Cleanup
Silica Gel Cleanup
Gel-Permeation Cleanup
Acid-Base Partition Cleanup
Sulfur Cleanup
Sulfuric Acid/Permanganate Cleanup
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4.3 DETERMINATION OF ORGANIC ANALYTES
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the
allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to
be used by the analyst and the regulated community in making judgements necessary to meet
the data quality objectives or needs for the intended use of the data.
4.3,1 GAS CHROMATOGRAPHIC METHODS
The following methods are included in this section:
Method 8000B:
Method 8011;
Method 801 SB:
Method 8021B:
Method 8031:
Method 8032A:
Method 8033:
Method 8041:
Method 8061A:
Method 8070A:
Method 8081B:
Method 8082A:
Method 8091:
Method 8100:
Method 8111:
Method 8121:
Method 8131:
Method 8141B:
Method 8151A;
Determinative Chromatographic Separations
1,2-Dibromoethane and 1,2-Dibromo-3-chloropropane by
Microextraction and Gas Chromatography
Nonhalogenated Organics Using GC/FID
Aromatic and Halogenated Voiatiles by Gas Chromatography
Using Photoionization and/or Electrolytic Conductivity
Detectors
Acrylonitrile by Gas Chromatography
Acrylamide by Gas Chromatography
Acetonitrile by Gas Chromatography with Nitrogen-
Phosphorus Detection
Phenols by Gas Chromatography
Phthalate Esters by Gas Chromatography with Electron
Capture Detection (GC/ECD)
Nitrosamines by Gas Chromatography
Organochiorine Pesticides by Gas Chromatography
Polychlorinated Biphenyls (PCBs) by Gas Chromatography
Nitroaromatics and Cyclic Ketones by Gas Chromatography
Polynuclear Aromatic Hydrocarbons
Haloethers by Gas Chromatography
Chlorinated Hydrocarbons by Gas Chromatography: Capillary
Column Technique
Aniline and Selected Derivatives by Gas Chromatography
Organophosphorus Compounds by Gas Chromatography
Chlorinated Herbicides by GC Using Methylation or
Pentafluorobenzylation Derivatization
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METHOD 80818
ORGANOCH LORIN E PESTICIDES BY GAS CHROMATOGRAPHY
1.0 SCOPE AND APPLICATION
1.1 Method 8081 may be used to determine the concentrations of various organochiorine
pesticides in extracts from solid and liquid matrices, using fused-silica, open-tubular, capillary
columns with electron capture detectors (ECD), When compared to the packed columns, these
columns offer improved resolution, better selectivity, increased sensitivity, and faster analysis. The
compounds listed below may be determined by either a single- or dual-column analysis system.
Compound
CAS Registry No.
Aldrin
309-00-2
a-BHC
319-84-6
P-BHC
319-85-7
Y-BHC (Lindane)
58-89-9
5-BHC
319-86-8
Chlorobenzilate
510-15-6
a-Chlordane
5103-71-9
Y-Chlordane
5103-74-2
Chlordane - not otherwise specified
57-74-9
DBCP
96-12-8
4,4-DDD
72-54-8
4,4-DDE
72-55-9
4,4'-DDT
50-29-3
Diallate
2303-16-4
Dteldrin
60-57-1
Endosulfan I
959-98-8
Endosulfan II
33213-65-9
Endosulfan sulfate
1031-07-8
Endrin
72-20-8
Endrin aldehyde
7421-93-4
Endrin ketone
53494-70-5
Heptachlor
76-44-8
Heptachlor epoxide
1024-57-3
Hexachlorobenzene
118-74-1
Hexachlorocyclopentadiene
77-47-4
Isodrin
465-73-6
Methoxychlor
72-43-5
Toxaphene
8001-35-2
8081B -1
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1.2 This revision of Method 8081 no longer includes the PCBs as Aroclors in the list of
target analytes. The analysis of PCBs should be undertaken using Method 8082, which includes
specific cleanup and quantitation procedures designed for PCB analysis. This change was made
to obtain PCB data of better quality and to eliminate the complications inherent in a combined
organochlorine pesticide and PCB method. Therefore, if the presence of PCBs is expected, use
Method 8082 for PCB analyses, and this method (8081) for the organochlorine pesticides. If there
is no information of the likely presence of PCBs, either employ a PCB-specific screening procedure
such as an immunoassay (e.g., Method 4020), or split the sample extract prior to any cleanup steps,
and process part of the extract for organochlorine pesticide analysis and the other portion for PCB
analysis using Method 8082.
1.3 The analyst must select columns, detectors and calibration procedures most
appropriate for the specific analytes of interest in a study. Matrix-specific performance data must
be established and the stability of the analytical system and instrument calibration must be
established for each analytical matrix (e.g., hexane solutions from sample extractions, diluted oil
samples, etc.).
1.4 Although performance data are presented for many of the target analytes, it is unlikely
that all of them could be determined in a single analysis. The chemical and chromatographic
behaviors of many of these chemicals can result in co-elution of some target analytes. Several
cleanup/fractionation schemes are provided in this method and in Method 3600.
1.5 Several multi-component mixtures (i.e., Chlordane and Toxaphene) are listed as target
analytes. When samples contain more than one multi-component anaiyte, a higher level of analyst
expertise is required to attain acceptable levels of qualitative and quantitative analysis. The same
is true of multi-component analytes that have been subjected to environmental degradation or
degradation by treatment technologies. These result in "weathered" multi-component mixtures that
may have significant differences in peak patterns than those of standards.
1.6 Compound identification based on single-column analysis should be confirmed on a
second column, or should be supported by at least one other qualitative technique. This method
describes analytical conditions for a second gas chromatographic column that can be used to
confirm the measurements made with the primary column. GC/MS Method 8270 is also
recommended as a confirmation technique, if sensitivity permits (Sec. 8.0).
1.7 This method includes a dual-column option. The option allows a hardware
configuration of two analytical columns joined to a single injection port. The option allows one
injection to be used for dual-column analysis. Analysts are cautioned that the dual-column option
may not be appropriate when the instrument is subject to mechanical stress, many samples are to
be run in a short period, or when contaminated samples are analyzed.
1.8 This method is restricted to use by, or under the supervision of, analysts experienced
in the use of gas chromatographs (GC) and skilled in the interpretation of gas chromatograms. Each
analyst must demonstrate the ability to generate acceptable results with this method.
1.9 Extracts suitable for analysis by this method may also be analyzed for
organophosphorus pesticides (Method 8141). Some extracts may also be suitable for triazine
herbicide analysis, if low recoveries (normally samples taken for triazine analysis must be preserved)
are not a problem.
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1.10 The following compounds may also be determined using this method:
Compound
CAS Registry No.
Aiachlor
15972-60-8
Captafol
2425-06-1
Chloroneb
2675-77-6
Chloropropyiate
5836-10-2
Chlorothaionil
1897-45-6
DCPA
1861-32-1
Dichlone
117-80-6
Dicofol
115-32-2
Etridiazole
2593-15-9
Halowax-1000
58718-66-4
Halowax-1001
58718-67-5
Halowax-1013
12616-35-2
Halowax-1014
12616-36-3
Halowax-1051
2234-13-1
Halowax-1099
39450-05-0
Mirex
2385-85-5
Nitrofen
1836-75-5
PCNB
82-68-8
Permethrin (c/s + trans)
52645-53-1
Perthane
72-56-0
Propachlor
1918-16-7
Strobane
8001-50-1
frans-Nonachlor
39765-80-5
Trifluralin
1582-09-8
1.11 Kepone extracted from samples or standards exposed to water or methanol may
produce peaks with broad tails that elute later than the standard by up to 1 minute. This shift is
presumably the result of the formation of a hemi-acetal from the ketone functionality. As a result,
Method 8081 is not recommended for determining Kepone. Method 8270 may be more appropriate
for the analysis of Kepone.
2.0 SUMMARY OF METHOD
2.1 A measured volume or weight of sample (approximately 1 L for liquids, 2 g to 30 g for
solids) is extracted using the appropriate matrix-specific sample extraction technique.
2.2 Liquid samples are extracted at neutral pH with methylene chloride using either Method
3510 (separatory funnel), Method 3520 (continuous liquid-liquid extractor), Method 3535 (solid-phase
extraction), or other appropriate technique.
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2.3 Solid samples are extracted with hexane-acetone (1:1) or methylene chloride-acetone
(1:1) using Method 3540 (Soxhlet), Method 3541 (automated Soxhiet), Method 3545 (pressurized
fluid extraction), Method 3550 (ultrasonic extraction), or other appropriate technique.
2.4 A variety of cleanup steps may be applied to the extract, depending on the nature of
the matrix interferences and the target analytes. Suggested cleanups include alumina (Method
3610), Florisil (Method 3620), silica gel (Method 3630), gel permeation chromatography (Method
3640), and sulfur (Method 3660).
2.5 After cleanup, the extract is analyzed by injecting a 1-|jL sample into a gas
chromatograph with a narrow- or wide-bore fused-siiica capillary column and electron capture
detector (GC/ECD) or an electrolytic conductivity detector (GC/ELCD).
3.0 INTERFERENCES
3.1 Refer to Methods 3500 (Sec. 3.0, in particular), 3600, and 8000, for a discussion of
interferences.
3.2 Sources of interference in this method can be grouped into three broad categories.
3.2.1 Contaminated solvents, reagents, or sample processing hardware.
3.2.2 Contaminated GC carrier gas, parts, column surfaces, or detector surfaces.
3.2.3 Compounds extracted from the sample matrix to which the detector will
respond.
3.2.4 Interferences co-extracted from the samples will vary considerably from waste
to waste. While general cleanup techniques are referenced or provided as part of this method,
unique samples may require additional cleanup approaches to achieve desired degrees of
discrimination and quantitation.
3.3 Interferences by phthalate esters introduced during sample preparation can pose a
major problem in pesticide determinations.
3.3.1 These materials may be removed prior to analysis using Method 3640 (Gel
Permeation Cleanup) or Method 3630 (Silica Gel Cleanup).
3.3.2 Common flexible plastics contain varying amounts of phthalate esters which
are easily extracted or leached from such materials during laboratory operations,
3.3.3 Cross-contamination of clean glassware routinely occurs when plastics are
handled during extraction steps, especially when solvent-wetted surfaces are handled.
3.3.4 Interferences from phthalate esters can best be minimized by avoiding contact
with any plastic materials and checking all solvents and reagents for phthalate contamination.
Exhaustive cleanup of solvents, reagents and glassware may be required to eliminate
background phthalate ester contamination.
3.4 Glassware must be scrupulously cleaned. Clean all glassware as soon as possible
after use by rinsing with the last solvent used. This should be followed by detergent washing with
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hot water, and rinses with tap water and organic-free reagent water. Drain the glassware and dry
it in an oven at 130°C for several hours, or rinse with methanol and drain. Store dry glassware in
a clean environment.
3.5 The presence of elemental sulfur will result in broad peaks that iinterfere with the
detection of earty-eluting organochlorine pesticides. Sulfur contamination should be expected with
sediment samples. Method 3660 is suggested for removal of sulfur. Since the recovery of Endrin
aldehyde (using the TBA procedure) is drastically reduced, this compound must be determined prior
to sulfur cleanup.
3.6 Waxes, lipids, and other high molecular weight materials can be removed by Method
3640 (gel-permeation cleanup).
3.7 Other halogenated pesticides or industrial chemicals may interfere with the analysis of
pesticides. Certain co-eluting organophosphorus pesticides are eliminated by Method 3640 (gel
permeation cleanup - pesticide option). Co-eluting chlorophenols may be eliminated by using Method
3630 (silica gel), Method 3620 (Florisil), or Method 3610 (alumina). Polychlorinated biphenyls
(PCBs) also may interfere with the analysis of the organochlorine pesticides. The problem may be
most severe for the analysis of multicomponent analytes such as Chlordane, Toxaphene, and
Strobane. If PCBs are known or expected to occur in samples, the analyst should consult Methods
3620 and 3630 for techniques that may be used to separate the pesticides from the PCBs.
3.8 Co-elution among the many target analytes in this method can cause interference
problems. The following target analytes may coelute on the GC columns listed, when using the
single-column analysis scheme:
DB 608 Trifluralin/Diallate isomers
PCNP/Dichlone/I sodrin
DB 1701 Captafol/Mirex
Methoxychlor/Endosulfan sulfate
3.9 The following compounds may coelute using the dual-column analysis scheme. In
general, the DB-5 column resolves fewer compounds that the DB-1701.
DB-5 Permethrin/Heptachlor epoxide
Endosulfan l/a-Chlordane
Perthane/Endrin
Endosulfan ll/Chloropropylate/Chlorobenzilate
4,4-DDT/Endosulfan sulfate
Methoxychlor/Dicofol
DB-1701 Chlorothalonil/p-BHC
5-BHC/DCPA/Permethrin
a-Chlorda ne/fra ns-Nonachlor
Nitrofen, Dichlone, Carbophenothion, Dichloran exhibit extensive peak tailing on both columns.
Simazine and Atrazine give poor responses on the ECD detector. Triazine compounds should
be analyzed using Method 8141 (NPD option).
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4.0 APPARATUS AND MATERIALS
4.1 Gas chromatograph - An analytical system complete with gas chromatograph suitable
for on-column and split-splitless injection and all required accessories including syringes, analytical
columns, gases, electron capture detectors (ECD), and recorder/integrator or data system.
4.2 GC columns
This method describes procedures for both single-column and dual-column analyses. The
single-column approach involves one analysis to determine that a compound is present, followed
by a second analysis to confirm the identity of the compound (Sec. 8.4 describes how GC/MS
confirmation techniques may be employed). The single-column approach may employ either narrow-
bore (sO.32 mm ID) columns or wide-bore (0.53 mm ID) columns. The dual-column approach
involves a single injection that is split between two columns that are mounted in a single gas
chromatograph. The dual-column approach employs only wide-bore (0.53 mm ID) columns.
The columns listed m this section were the columns used to develop the method performance
data. The mention of these columns in this method is not intended to exclude the use of other
columns that may be developed. Laboratories may use other capillary columns provided that they
document method performance data (e.g., chromatographic resolution, analyte breakdown, and
MDLs) that equals or exceeds the performance described in this method, or as appropriate for the
intended application.
4.2.1 Narrow-bore columns for single-column analysis (use both columns to confirm
compound identifications unless another confirmation technique such as GC/MS is employed).
4.2.1.1 30-m x 0.25 or 0.32 mm ID fused-silica capillary column chemically
bonded with SE-54 (DB-5 or equivalent), 1 Mm film thickness.
4.2.1.2 30-m x 0.25 mm ID fused-silica capillary column chemically
bonded with 35 percent phenyl methylpolysiloxane (DB-608, SPB-608, or equivalent),
2.5 pm coating thickness, 1 pm film thickness.
4.2.1.3 Narrow-bore columns should be Installed in split/splitless (Grob-
type) injectors.
4.2.2 Wide-bore columns for single-column analysis (use two of the three columns
listed to confirm compound identifications unless another confirmation technique such as
GC/MS is employed).
4.2.2.1 30-m x 0.53 mm ID fused silica capillary column chemically
bonded with 35 percent phenyl methylpolysiloxane (DB-608, SPB-608, RTx-35, or
equivalent), 0.5 Mm or 0.83 Mm film thickness.
4.2.2.2 30-m x 0.53 mm ID fused silica capillary column chemically
bonded with 50 percent phenyl methylpolysiloxane (DB-1701, or equivalent), 1.0 pm
film thickness.
4.2.2.3 30-m x 0.53 mm ID fused silica capillary column chemically
bonded with 95 percent dimethyl - 5 percent diphenyl polysiioxane (DB-5, SPB-5, RTx-
5, or equivalent), 1.5 Mm film thickness.
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4.2.2.4 Wide-bore columns should be installed in 1/4 inch injectors, with
deactivated liners designed specifically for use with these columns.
4.2.3 Wide-bore columns for dual-column analysis (choose one of the two pairs of
columns listed below).
4.2.3.1 Column pair 1
30-m x 0.53 mm ID fused silica capillary column chemically bonded with SE-
54 (DB-5, SPB-5, RTx-5, or equivalent), 1.5 ym film thickness.
30-m x 0.53 mm ID fused silica capillary column chemically bonded with 50
percent phenyl methylpolysiloxane (DB-1701, or equivalent), 1.0 (jnfi film thickness.
Column pair 1 is mounted in a press-fit Y-shaped glass 3-way union splitter
(J&W Scientific, Catalog No. 705-0733) or a Y-shaped fused-silica connector (Restek,
Catalog No. 20405), or equivalent.
4.2.3.2 Column pair 2
30-m x 0.53 mm ID fused silica capillary column chemically bonded with SE-
54 (DB-5, SPB-5, RTx-5, or equivalent), 0.83 Mm film thickness.
30-m x 0.53 mm ID fused silica capillary column chemically bonded with 50
percent phenyl methylpolysiloxane (DB-1701, or equivalent), 1.0 Mm film thickness.
Column pair 2 is mounted in an 8 in. deactivated glass injection tee (Supelco,
Catalog No. 2-3665M), or equivalent.
4.3 Column rinsing kit - Bonded-phase column rinse kit (J&W Scientific, Catalog No. 430-
3000), or equivalent.
4.4 Volumetric flasks, 10-mL and 25-mL, for preparation of standards.
5.0 REAGENTS
5.1 Reagent grade or pesticide grade chemicals shall be used in all tests. Unless
otherwise indicated, it is intended that all reagents shall conform to specifications of the Committee
on Analytical Reagents of the American Chemical Society, where such specifications are available.
Other grades may be used, provided it is first ascertained that the reagent is of sufficiently high purity
to permit its use without lessening the accuracy of the determination.
NOTE: Store the standard solutions (stock, composite, calibration, internal, and surrogate) at 4°C
in polytetrafluoroethylene (PTFE)-sealed containers in the dark. When a lot of standards
is prepared, it is recommended that aliquots of that lot be stored in individual small vials.
All stock standard solutions must be replaced after one year or sooner if routine QC tests
(Sec. 8.0) indicate a problem. All other standard solutions must be replaced after six
months or sooner if routine QC (Sec. 8.0) indicates a problem.
5.2 Solvents used in the extraction and cleanup procedures (appropriate 3500 and 3600
series methods) include n-hexane, diethyl ether, methylene chloride, acetone, ethyl acetate, and
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isooctane (2,2,4-trimethyIpentane) and must be exchanged to n-hexane or isooctane prior to
analysis.
Therefore, n-hexane and isooctane will be required in this procedure. Acetone or toluene may
be required for the preparation of some standard solutions (see Sec. 5.4.2). All solvents should be
pesticide quality or equivalent, and each lot of solvent should be determined to be phthalate free.
5.3 Organic-free reagent water - All references to water in this method refer to organic-free
reagent water as defined in Chapter One.
5.4 Stock standard solutions (1000 mg/L) - May be prepared from pure standard materials
or can be purchased as certified solutions.
5.4.1 Prepare stock standard solutions by accurately weighing about 0.0100 g of
pure compound. Dissolve the compound in isooctane or hexane and dilute to volume in a 10-
mL volumetric flask. If compound purity is 96 percent or greater, the wight can be used
without correction to calculate the concentration of the stock standard solution. Commercially
prepared stock standard solutions can be used at any concentration if they are certified by the
manufacturer or by an independent source.
5.4.2 (J-BHC, Dieldrin, and some other standards may not be adequately soluble
in isooctane. A small amount of acetone or toluene should be used to dissolve these
compounds during the preparation of the stock standard solutions.
5.5 Composite stock standard - May be prepared from individual stock solutions.
5.5.1 For composite stock standards containing less than 25 components, take
exactly 1 mL of each individual stock solution at a concentration of 1000 mg/L, add solvent,
and mix the solutions in a 25-mL volumetric flask. For example, for a composite containing 20
individual standards, the resulting concentration of each component in the mixture, after the
volume is adjusted to 25 mL, will be 1 mg/25 mL. This composite solution can be further
diluted to obtain the desired concentrations.
5.5.2 For composite stock standards containing more than 25 components, use
volumetric flasks of the appropriate volume (e.g., 50 mL, 100 mL), and follow the procedure
described above.
5.6 Calibration standards should be prepared at a minimum of five different concentrations
by dilution of the composite stock standard with isooctane or hexane. The concentrations should
correspond to the expected range of concentrations found in real samples and should bracket the
linear range of the detector.
5.6.1 Although all single component analytes can be resolved on a new 35 percent
phenyl methyl silicone column (e.g., DB-608), two calibration mixtures should be prepared for
the single component analytes of this method. This procedure is established to minimize
potential resolution and quantitation problems on confirmation columns or on older 35 percent
phenyl methyl silicone (e.g. DB-608) columns and to allow determination of Endrin and DDT
breakdown for method QC (Sec. 8.0).
5.6.2 Separate calibration standards are required for each multi-component target
analyte (e.g., Toxaphene and Chlordane). Analysts should evaluate the specific Toxaphene
standard carefully. Some Toxaphene components, particularly the more heavily chlorinated
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components, are subject to dechlorination reactions. As a result, standards from different
vendors may exhibit marked differences which could lead to possible false negative results or
to large differences in quantitative results.
5.7 Internal standard (optional)
5.7.1 Pentachloronitrobenzene is suggested as an internal standard for the single-
column analysis, when it is not considered to be a target analyte. 1 -bromo-2-nitrobenzene may
also be used. Prepare a solution of 5000 mg/L (5000 ng/pL) of pentachloronitrobenzene or 1-
bromo-2-nitrobenzene. Spike 10 jjL of this solution into each 1 mL sample extract.
5.7.2 1 -bromo-2-nitrobenzene is suggested as an internal standard for the dual-
column analysis. Prepare a solution of 5000 mg/L (5000 ng/pL) of. 1 -bromo-2-nitrobenzene.
Spike 10 |iL of this solution into each 1 mL of sample extract.
5.8 Surrogate standards
The performance of the method should be monitored using surrogate compounds.
Surrogate standards are added to all samples, method blanks, matrix spikes, and calibration
standards. The following compounds are recommended as possible surrogates.
5.8.1 Decachlorobiphenyl and tetrachloro-m-xylene have been found to be a useful
pair of sun-ogates for both the single-column and dual-column configurations. Method 3500,
Sec. 5.0, describes the procedures for preparing these surrogates.
5.8.2 4-Chloro-3-nitrobenzotrifluoride may also be useful as a surrogate if the
chromatographic conditions of the dual-column configuration cannot be adjusted to preclude
co-el ution of a target analyte with either of the surrogates in Sec. 5.8.1. However, this
compound elutes early in the chromatographic run and may be subject to other interference
problems. A recommended concentration for this surrogate is 500 ng/pL. Use a spiking
volume of 100 pL for a 1-L aqueous sample.
5.8.3 Store surrogate spiking solutions at 4°C in PTFE-sealed containers In the
dark.
6.0 SAMPLE COLLECTION, PRESERVATION, AND HANDLING
6.1 See Chapter Four, Organic Analytes, Sec. 4.0, for sample collection and preservation
instructions.
6.2 Extracts must be stored under refrigeration in the dark and analyzed within 40 days of
extraction.
7.0 PROCEDURE
7.1 Sample extraction
Refer to Chapter Two and Method 3500 for guidance in choosing the appropriate
extraction procedure. In general, water samples are extracted at a neutral pH with methylene
chloride using a separatory funnel (Method 3510), a continuous liquid-liquid extractor (Method
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3520), solid-phase extraction (Method 3535), or other appropriate technique. Solid samples
are extracted with hexane-acetone (1:1) or methylene chloride-acetone (1:1) using one of the
Soxhlet extraction (Method 3540 or 3541), pressurized fluid extraction (Method 3545),
ultrasonic extraction (Method 3550), supercritical fluid extraction (Method 3562), or other
appropriate technique.
NOTE: Hexane-acetone (1:1) may be more effective as an extraction solvent for
organochlorine pesticides in some environmental and waste matrices than is
methylene chloride-acetone (1:1). Relative to the methylene chloride-acetone
mixture, use of hexane-acetone generally reduces the amount of interferences that
are extracted and improves signal-to-noise.
Spiked samples are used to verify the applicability of the chosen extraction technique
to each new sample type. Each sample type must be spiked with the compounds of interest
to determine the percent recovery and the limit of detection for that sample (see Chapter One).
See Method 8000 for guidance on demonstration of initial method proficiency as well as
guidance on matrix spikes for routine sample analysis.
7.2 Extract cleanup
Cleanup procedures may not be necessary for a relatively clean sample matrix, but
most extracts from environmental and waste samples will require additional preparation before
analysis. The specific cleanup procedure used will depend on the nature of the sample to be
analyzed and the data quality objectives for the measurements. General guidance for sample
extract cleanup is provided in this section and in Method 3600.
7.2.1 If a sample is of biological origin, or contains high molecular weight materials,
the use of Method 3640 (GPC cleanup - pesticide option) is recommended. Frequently, one
of the adsorption chromatographic cleanups (alumina, silica gel, or Florisil) may also be
required following the GPC cleanup.
7.2.2 Method 3610 (alumina) may be used to remove phthaiate esters.
7.2.3 Method 3620 (Florisil) may be used to separate organochlorine pesticides
from aliphatic compounds, aromatics, and nitrogen-containing compounds.
7.2.4 Method 3630 (silica gel) may be used to separate single component
organochlorine pesticides from some interferants.
7.2.5 Elemental sulfur, which may be present in certain sediments and industrial
wastes, interferes with the electron capture gas chromatography of certain pesticides. Sulfur
should be removed by the technique described in Method 3660.
7.3 GC conditions
This method allows the analyst to choose between a single-column or a dual-column
configuration in the injector port. Either wide- or narrow-bore columns may be used. Identifications
based on retention times from a single-column must be confirmed on a second column or with an
alternative qualitative technique.
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7.3.1 Single-column analysis
This capillary GC/ECD method allows the analyst the option of using 0.25-0.32 mm ID
capillary columns (narrow-bore) or 0.53 mm ID capillary columns (wide-bore). Performance
data are provided for both options. Figures 1-6 provide example chromatograms.
7.3.1.1 The use of narrow-bore (sO.32 mm ID) columns is recommended
when the analyst requires greater chromatographic resolution. Use of narrow-bore
columns is suitable for relatively clean samples or for extracts that have been prepared
with one or more of the clean-up options referenced in the method.- Wide-bore
columns (0.53 mm ID) are suitable for more complex environmental and waste
matrices.
7.3.1.2 Table 1 lists average retention times for the target analytes using
wide-bore capillary columns. Table 2 lists average retention times for the target
analytes using narrow-bore capillary columns.
7.3.1.3 Table 4 lists the GC operating conditions for the single-column
method of analysis,
7.3.2 Dual-column analysts
The dual-column/dual-detector approach involves the use of two 30-m x 0.53 mm ID
fused-silica open-tubular columns of different polarities, thus, different selectivities towards the
target analytes. The columns are connected to an injection tee and separate electron capture
detectors.
7.3.2.1 Retention times for the organochlorine analytes on dual-columns
are in Table 6. The GC operating conditions for the compounds in Table 6 are given
in Table 7.
7.3.2.2 Multi-component mixtures of Toxaphene and Strobane were
analyzed separately (Figures 5 and 6) using the operating conditions found in Table 7.
7.3.2.3 Figure 6 is a sample chnomatogram for a mixture of organochlorine
pesticides. The retention times of the individual components detected in these
mixtures are given in Tables 6 and 7.
7.3.2.4 Operating conditions for a more heavily loaded DB-5/DB-1701 pair
are given in Table 8. This column pair was used for the detection of multi-component
organochlorine compounds.
7.3.2.5 Operating conditions for a DB-5/DB-1701 column pair with thinner
films, a different type of splitter, and a slower temperature programming rate are
provided in Table 7. These conditions gave better peak shapes for Nitrofen and
Dicofol. Table 6 lists the retention times for the compounds on this column pair.
7.4 Calibration
7.4.1 Prepare calibration standards using the procedures in Sec. 5.0. Refer to
Method 8000 (Sec. 7.0) for proper calibration techniques for both initial calibration and
calibration verification. The procedure for either internal or external calibration may be used.
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In most cases, external standard calibration is used with Method 8081 because of the
sensitivity of the electron capture detector and the probability of the internal standard being
affected by interferences. Because several of the pesticides may co-elute on any single-
column, analysts should use two calibration mixtures (see Sec. 3.8). The specific mixture
should be selected to minimize the problem of peak overlap.
NOTE: Because of the sensitivity of tiie electron capture detector, the injection port and
column should always be cleaned prior to performing the initial calibration.
7.4.1.1 Unless otherwise necessary for a specific project, the analysis of
the multi-component arialytes employs a single-point calibration. A single calibration
standard near the mid-point of the ejected calibration range of each multi-component
analyte is included with the initial calibration of the single component anaiytes for
pattern recognition, so that the analyst is familiar with the patterns and retention times
on each column.
7.4.1.2 For calibration verification (each 12-hour shift) all target anaiytes
required in the project plan must be injected.
7.4.2 Establish the GC operating conditions appropriate for the configuration
(single-column or dual column, Sec. 7.3) using Tables 4, 5,7, or 8 as guidance. Optimize the
instrumental conditions for resolution of the target anaiytes and sensitivity. An initial oven
temperature £ 140 -150°C is required to resolve the four BHC isomers. A final temperature of
240 - 270°C is required to elute decachlorobiphenyl. Use of injector pressure programming
will improve the chromatography of late eluting peaks.
NOTE: Once established, the same operating conditions must be used for both calibrations
and sample analyses.
7.4.3 A 2-jjL injection volume of each calibration standard is recommended. Other
injection volumes may be employed, provided that the analyst can demonstrate adequate
sensitivity for the compounds of interest.
7.4.4 Because of the low concentration of pesticide standards injected on a
GC/ECD, column adsorption may be a problem when the GC has not been used for a day or
more. Therefore, the GC column should be primed (or deactivated) by injecting a pesticide
standard mixture approximately 20 times more concentrated than the mid-concentration
standard. Inject this standard mixture prior to beginning the initial calibration or calibration
verification.
CAUTION: Several anaiytes, including Aldrin, may be observed in the injection just following
this system priming. Always run an acceptable blank prior to running any
standards or samples.
7.4.5 Calibration factors
When external standard calibration is employed, calculate the calibration factor for each
analyte at each concentration, the mean calibration factor, and the relative standard deviation
(RSD) of the calibration factors, using the formulae below. If internal standard calibration is
employed, refer to Method 8000 for the calculation of response factors.
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7.4.5.1 Calculate the calibration factor for each analyte at each
concentration as:
QP „ Pea^ Area (or Height) of the Compound in the Standard
Mass of the Compound Injected (in nanograms)
7.4.5.2 Calculate the mean calibration factor for each analyte as:
_ ECFi
mean CF = CF = —
n
where n is the number of standards analyzed.
7.4.5.3 Calculate the standard deviation (SDj and the RSD of the
calibration factors for each analyte as:
SD -
£(CFrCF)2
M
n-1
RSD
SD
CF
x 100
If the RSD for each analyte is <; 20%, then the response of the instrument is considered
linear and the mean calibration factor can be used to quantitate sample results. If the
RSD is greater than 20%, then linearity through the origin cannot be assumed. The
analyst must use a calibration curve or a non-linear calibration model (e.g., a
polynomial equation) for quantitation. See Method 8000 for information on non-linear
calibrations.
7.4.6 Retention time windows
Absolute retention times are used for compound identification. Retention time windows
are crucial to the identification of target compounds, and should be established by one of the
approaches described in Method 8000.
7.4.6.1 Before establishing the retention time windows, make sure the gas
chromatographic system is operating within optimum conditions.
7.4.6.2 The widths of the retention time windows are defined as described
in Method 8000. However, the experience of the analyst should weigh heavily in the
interpretation of the chromatograms.
7.5 Gas chromatographic analysis of sample extracts
7.5.1 The same GC operating conditions used for the initial calibration must be
employed for samples analyses.
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7.5.2 Verity calibration each 12-hour shift by injecting calibration verification
standards prior to conducting any sample analyses. Analysts should alternate the use of high
and low concentration mixtures of single-component analytes and multi-component anatytes
for calibration verification. A calibration standard must also be injected at intervals of not less
than once every twenty samples (after every 10 samples is recommended to minimize the
number of samples requiring re-injection when QC limits are exceeded) and at the end of the
analysis sequence. See Sec. 8.4.4 for additional guidance on the frequency of the standard
injections.
7.5.2.1 The calibration factor for each analyte should not exceed a ± 15
percent difference from the mean calibration factor calculated for the initial calibration.
If a non-linear calibration model or a linear model not through the origin has been
employed for the initial calibration, consult Sec. 7 of Method 8000 for the specifics of
calibration verification.
Qp _ Qp
% Difference = —=—- * 100
CF
7.5.2.2 If this criterion is exceeded for any analyte, use the approach
described in Sec. 7 of Method 8000 to calculate the average percent difference across
a}! analytes. If the average of the responses for all analytes is within ±15%, then the
calibration has been verified. However, the conditions in Sec. 7 of Method 8000 also
apply, e.g., the average must include all analytes in the calibration, regardless of
whether they are target analytes for a specific project, and the data user must be
provided with the calibration verification data or a list of those analytes that exceeded
the ±15% limit,
7.5.2.3 If the calibration does not meet the ±15% limit (either on the basis
of each compound or the average across all compounds), check the instrument
operating conditions, and if necessary, restore them to the original settings, and inject
another aliquot of the calibration verification standard. If the response for the analyte
is still not within ±15%, then a new initial calibration must be prepared. The effects of
a failing calibration verification standard on sample results are discussed in Sec. 7.5.7.
7.5.3 Compare the retention time of each analyte in the calibration standard with
the absolute retention time windows established in Sec. 7.4.6. As described in Method 8000,
the center of the absolute retention time window for each analyte is its retention time in the
mid-concentration standard analyzed during the initial calibration. Each analyte in each
standard must fall within its respective retention time window. If not, the gas chromatographic
system must either be adjusted so that a second analysis of the standard does result in all
analytes falling within their retention time windows, or a new initial calibration must be
performed and new retention time windows established.
7.5.4 Inject a 2-|jL aliquot of the concentrated sample extract. Record the volume
injected to the nearest 0.05 |j|_ and the resulting peak size in area units.
7.5.5 Tentative identification of an analyte occurs when a peak from a sample
extract falls within the absolute retention time window. Each tentative identification must be
confirmed using either a second GC column of dissimilar stationary phase or using another
technique such as GC/MS (see Sec. 7.7).
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When results are confirmed using a second GC column of dissimilar stationary phase,
the analyst should check the agreement between the quantitative results on both columns once
the identification has been confirmed. See Sec. 7 of Method 8000 for a discussion of such a
comparison. Unless otherwise specified in an approved project plan, the higher result should
be reported, as this is a conservative approach relative to protection of the environment. If the
relative percent difference of the results exceeds 40%, consult Method 8000 for steps that may
be taken to address the discrepancy.
7.5.6 When using the external calibration procedure (Method 8000), determine the
quantity of each component peak in the sample chromatogram which corresponds to the
compounds used for calibration purposes, as follows. Proper quantitation requires the
appropriate selection of a baseline from which the peak area or height can be determined.
7.5.6.1 For aqueous samples
Concentration (pg/L) -
(CFXV.XVg
where:
A* = Area (or height) of the peak for the analyte in the sample.
Vt = Total volume of the concentrated extract (pL).
D = Dilution factor, if the sample or extract was diluted prior to analysis. If no
dilution was made, D - 1. The dilution factor is always dimensionless.
£F = Mean calibration factor from the initial calibration (area/ng).
Vj = Volume of the extract injected (pL). The injection volume for samples and
calibration standards must be the same. For purge-and-trap analysis, V, is not
applicable and therefore is set at 1.
V, = Volume of the aqueous sample extracted in mL. If units of liters are used for
this term, multiply the results by 1000.
Using the units specified here for these terms will result in a concentration in units of
ng/mL, which is equivalent to pg/L.
7.5.6.2 For non-aqueous samples
Concentration (pg/kg) - (A*)(V')(D)
(CFXV^OAQ
where A*, Vt, D, CF, and Vj are the same as for aqueous samples, and
W, = Weight of sample extracted (g). The wet weight or dry weight may be used,
depending upon the specific application of the data. If units of kilograms are
used for this term, multiply the results by 1000.
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Using the units specified here for these terms will result in a concentration in units of
ng/g, which is equivalent to pg/kg.
7.5.6.3 See Method 8000 for the equation used for internal standard
quantitation.
7.5.6.4 If the responses exceed the calibration range of the system, dilute
the extract and reanalyze. Peak height measurements are recommended over peak
area integration when overlapping peaks cause errors in area integration.
7.5.6.5 If partially overlapping or coefuting peaks are found, change GC
columns or try GC/MS quantitation (see Sec. 8.0 and Method 8270).
7.5.7 Each sample analysis must be bracketed with an acceptable initial calibration,
calibration verification standard(s) (each 12-hour analytical shift), or calibration standards
interspersed within the samples.
Although analysis of a single mid-concentration standard (standard mixture or multi-
component anaiyte) will satisfy the minimum requirements, analysts are urged to use different
calibration verification standards during organochlorine pesticide analyses. Also, multi-level
standards (mixtures or multi-component analytes) are highly recommended to ensure that the
detector response remains stable for all the analytes over the calibration range.
The results from these bracketing standards must meet the calibration verification
criteria in Sec. 7.5.2. When a calibration verification standard fails to meet the QC criteria, all
samples that were injected after the last standard that last met the QC criteria must be
evaluated to prevent mis-quantitations and possible false negative results, and re-injection of
the sample extracts may be required. More frequent analyses of standards will minimize the
number of sample extracts that would have to be reinjected if the QC limits are violated for the
standard analysis.
However, if the standard analyzed after a group of samples exhibits a response for an
anaiyte that is above the acceptance limit, i.e., >15%, and the anaiyte was not detected in the
specific samples analyzed during the analytical shift, then the extracts for those samples do
not need to be reanalyzed, as the verification standard has demonstrated that the anaiyte
would have been detected were it present. In contrast, if an anaiyte above the QC limits was
detected in a sample extract, then re-injection is necessary to ensure accurate quantitation.
If an anaiyte was not detected in the sample and the standard response is more than 15%
below the initial calibration response, then re-injection is necessary to ensure that the detector
response has not deteriorated to the point that the anaiyte would not have been detected even
though it was present (i.e., a false negative result).
7.5.8 Sample injections may continue for as long as the calibration verification
standards and standards interspersed with the samples meet instrument QC requirements.
It is recommended that standards be analyzed after every 10 samples (required after every 20
samples and at the end of a set) to minimize the number of samples that must be re-injected
when the standards fail the QC limits. The sequence ends when the set of samples has been
injected or when qualitative and/or quantitative QC criteria are exceeded.
7.5.9 If the peak response is less than 2.5 times the baseline noise level, the
validity of the quantitative result may be questionable. The analyst should consult with the
source of the sample to determine whether further concentration of the sample is warranted.
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7.5.10 Validation of GC system qualitative performance
7.5.10.1 Use the calibration standards analyzed during the sequence to
evaluate retention time stability. The retention time windows are established using the
absolute retention time of each analyte as described in Method B000.
7.5.10.2 Each subsequent injection of a standard during the 12-hour
analytical shift (i.e., those standards injected every 20 samples, or more frequently)
must be checked against tie retention time windows. If any of these subsequent
standards fall outside their absolute retention time windows, the QC system is out of
control. Determine the cause of the problem and correct it. if the problem cannot be
corrected, a new initial calibration must be performed.
7.5.11 Identification of mixtures (i.e. Chlordane and Toxaphene) is based on the
characteristic "fingerprint" retention time and shape of the indicator peak(s); and quantitation
is based on the area under the characteristic peaks as compared to the area under the
corresponding calibration peak(s) of the same retention time and shape generated using either
internal or external calibration procedures.
7.5.12 If compound identification or quantitation is precluded due to interference
(e.g., broad, rounded peaks or ill-defined baselines are present) cleanup of the extract or
replacement of the capillary column or detector is warranted. Rerun the sample on another
instrument to determine if the problem results from analytical hardware or the sample matrix.
Refer to Method 3600 for the procedures to be followed in sample cleanup.
7.6 Quantitation of multi-component analytes - Multi-component analytes present problems
in measurement. Suggestions are offered in the following sections for handling Toxaphene,
Strobane, Chlordane, BHC, and DDT.
7.6.1 Toxaphene and Strobane - Toxaphene is manufactured by the chlorination
of camphenes, whereas Strobane results from the chlorination of a mixture of camphenes and
pinenes. Quantitation of Toxaphene or Strobane is difficult, but reasonable accuracy can be
obtained. To calculate Toxaphene from GC/ECD results:
7.6.1.1. Adjust the sample size so thatthe majorToxaphene peaks are 10-
70% of full-scale deflection (FSD).
7.6.1.2 Inject a Toxaphene standard that is estimated to be within ± 10 ng
of the sample amount.
7.6.1.3 Quantitate Toxaphene using the total area of the Toxaphene
pattern or using 4 to 6 major peaks.
7.6.1.3.1 While Toxaphene contains a large number of
compounds that will produce well resolved peaks in a GC/ECD
chromatogram, it also contains many other components that are not
chromatographically resolved. This unresolved complex mixture results in
the "hump" in the chromatogram that is characteristic of this mixture.
Although the resolved peaks are important for the identification of the
mixture, the area of the unresolved complex mixture contributes a significant
portion of the area of the total response.
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7.6.1.3.2 To measure total area, construct the baseline of
Toxaphene in the sample chromatogram between the retention times of the
first and last eluting Toxaphene components in the standard. In order to use
the total area approach, the pattern in the sample chromatogram must be
compared to that of the standard to ensure that all of the major components
in the standard are present in the sample. Otherwise, the sample
concentration may be significantly underestimated.
7.6.1.3.3 Toxaphene may also be quantitated on the basis of
4 to 6 major peaks. A collaborative stutfy of a series of Toxaphene residues
evaluated several approaches to quantitation of this compound, including the
use of the total area of the peaks in the Toxaphene chromatogram and the
use of a subset of 4 to 6 peaks. That study indicated that the use of 4 to 6
peaks provides results that agree well with the total peak area approach and
may avoid difficulties when interferences with Toxaphene peaks are present
in the early portion of the chromatogram from compounds such as DDT.
Whichever approach is employed should be documented and available to the
data user, if necessary.
7.6.1.3.4 When Toxaphene is determined using the 4 to 6
peaks approach, the analyst must take care to evaluate the relative areas of
the peaks chosen in the sample and standard chromatograms. It is highly
unlikely that the peaks will match exactly, but the analyst should not employ
peaks from the sample chromatogram whose relative sizes or areas appear
to be disproportionate larger or smaller in the sample compared to the
standard.
7.6.1.3.5 The heights or areas of tiie 4 to 6 peaks should be
summed together and used to determine the Toxaphene concentration.
Alternatively, use each peak in the standard to calculate a calibration factor
for that peak, using the total mass of Toxaphene in the standard. These
calibration factors are then used to calculate the concentration of each
corresponding peak in the sample chromatogram and the 4 to 6 resulting
concentrations are averaged to provide the final result for the sample.
7.6.2 Chlordane - Technical Chloidane is a mixture of at least 11 major components
and 30 or more minor components that is used to prepare specific pesticide formulations. The
CAS Registiy number for Technical Chlordane is property given as 12789-03-6. Trans-
Chlordane (or a-Chlordane, CAS RN 5103-71-9) and c/s-Chlordane (y-Chlordane, CAS RN
5103-74-2), are the two most prevalent major components of Technical Chlordane. However,
the exact percentage of each in the technical material is not completely defined, and is not
consistent from batch to batch. Moreover, changes may occur when the technical material is
used to prepare specific pesticide formulations. The approach used for evaluating and
reporting Chlordane results will often depend on the end use of the results and the analyst's
skill in interpreting this multi-component pesticide residue. The following sections discuss
three specific options: reporting Technical Chlordane (12789-03-6), reporting Chlordane (not
ottterwise specified, 57-74-9), and reporting the individual Chlordane components that can be
identified under their individual CAS numbers.
7.6.2.1 When the GC pattern of the residue resembles that of Technical
Chlordane, the analyst may quantitate Chlordane residues by comparing the total area
of the Chlordane chromatogram using three to five major peaks or the total area. If the
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Heptachlor epoxide peak is relatively small, include it as part of the total Chlordane
area for calculation of the residue. If Heptachlor and/or Heptachlor epoxide are much
out of proportion, calculate these separately and subfcact their areas from tfie total area
to give a corrected Chlordane area.
NOTE: Octachloro epoxide, a metabolite of Chlordane, can easily be mistaken for
Heptachlor epoxide on a nonpolar GC column.
-i
To measure the total area of the Chlordane chromatogram, inject an amount
of a Technical Chlordane standard which will produce a chromatogram in which the
major peals are approximately the same size as those in the sample chromatograms.
Construct the baseline of Technical Chlordane in the standard chromatogram between
tie retention times of the first and last eluting toxaphene components. Use this area
and the mass of Technical Chlordane in the standard to calculate a calibration factor.
Construct a similar baseline In the sample chromatogram, measure the area, and use
the calibration factor to calculate the concentration in the sample.
7.6.2.2 The GC pattern of a Chlordane residue in a sample may differ
considerably from that of the Technical Chlordane standard. In such instances, it may
not be practical to relate a sample chromatogram back to the pesticide active
ingredient Technical Chlordane. Therefore, depending on the objectives of the
analysis, the analyst may choose to report the sum of all the identifiable Chlordane
components as "Chlordane (n.o.s.)* under the CAS number §7-74-9.
7.6.2.3 The third option is to quantitate the peaks of a-Chlordane, y-
Chlordane, and Heptachlor separately against the appropriate reference materials, and
report these individual components under their respective CAS numbers.
7.6.2.4 To measure the total area of the Chlordane chromatogram, inject
an amount of a Technical Chlordane standard which will produce a chromatogram in
which the major peaks are approximately the same size as those in the sample
chromatograms.
7.6.3 Hexachlorocyclohexane - Hexachlorocydohexane is also known as BHC, from
the former name, benzene hexachioride. Technical grade BHC is a cream-colored amorphous
solid with a very characteristic musty odor. It consists of a mixture of six chemically distinct
isomers and one or more heptachlorocyclohexanes and octachlorocyclohexanes. Commercial
BHC preparations may show a wide variance in the percentage of individual isomers present.
Quantitate each isomer (a, (3, y. and 0) separately against a standard of the respective pure
isomer.
7.6.4 DDT - Technical DDT consists primarily of a mixture of 4,4-DDT
(approximately 75%) and 2,4-DDT (approximately 25%). As DDT weathers, 4,4-DDE, 2,4-
DDE, 4,4'-DDD, and 2,4-DDD are formed. Since the 4,4'-isomers of DDT, DDE, and DDD
predominate in the environment, these are the isomers normally regulated by EPA. Therefore,
sample extracts should be quantitated against standards of the respective pure isomers of 4,4-
DDT, 4,4-DDE, and 4,4-DDD.
7.7 GC/MS confirmation may be used in conjunction with either single-column or dual-
column analysis if the concentration is sufficient for detection by GC/MS.
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7.7.1 Full-scan GC/MS will normally require a concentration of approximately 10
ng/|iL in the final extract for each single-component compound. Ion trap or selected ion
monitoring will normally require a concentration of approximately 1 ng/nL.
7.7.2 The GC/MS must be calibrated for the specific target pesticides when H is
used for quantitative analysis. If GC/MS is used only for confirmation of the identification of
the target anaiytes, then the analyst must demonstrate that those pesticides identified by
GC/ECD can be confirmed by GC/MS. This demonstration may be accomplished by analyzing
a single-point standard containing the anaiytes of interest at or below the concentrations
reported in the GC/EC analysis.
7.7.3 GC/MS is not recommended for confirmation when concentrations are below
1 ng/yl_ in the extract, unless a more sensitive masss spectrometer is employed.
7.7.4 GC/MS confirmation should be accomplished by analyzing the same extract
that is used for GC/ECD analysis and the extract of the associated method blank.
7.7.5 The base/neutral/acid extract and the associated blank may be used for
GC/MS confirmation if the surrogates and internal standards do not interfere and if it is
demonstrated that the analyte is stable during acid/base partitioning. However, if the
compounds are not detected in the base/neutral/acid extract, then GC/MS analysis of the
pesticide extract should be performed.
7.8 Suggested chromatographic system maintenance - When system performance does
not meet the established QC requirements, corrective action is required, and may include one or
more of the following.
7.8.1 Splitter connections - For dual-columns which are connected using a press-fit
Y-shaped glass splitter or a Y-shaped fused-silica connector, clean and deactivate the splitter
port insert or replace with a cleaned and deactivated splitter. Break off the first few
centimeters (up to 30 cm) of the injection port side of the column. Remove the columns and
solvent backflush according to the manufacturer's instructions. If these procedures fail to
eliminate the degradation problem, it may be necessary to deactivate the metal injector body
and/or replace the columns.
7.8.2 GC injector ports can be of critical concern, especially in the analysis of DDT
and Endrin. Injectors that are contaminated, chemically active, or too hot can cause the
degradation ("breakdown") of the anaiytes. Endrin and DDT breakdown to endrin aldehyde,
endrin ketone, DDD, or DDE. When such breakdown is observed, clean and deactivate the
injector port, break off at least 30 cm of the column and remount it. Check the injector
temperature and lower it to 205°C, if required. Endrin and DDT breakdown are less of a
problem when ambient on-column injectors are used.
7.8.3 Metal injector body - Turn off the oven and remove the analytical columns
when the oven has cooled. Remove the glass injection port insert (instruments with on-column
injection). Lower the injection port temperature to room temperature. Inspect the injection port
and remove any noticeable foreign material.
7.8.3.1 Place a beaker beneath the injector port inside the oven. Using
a wash bottle, serially rinse the entire inside of the injector port with acetone and then
toluene, catching the rinsate in the beaker.
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7.8.3,2 Prepare a solution of a deactivating agent (Sylon-CT or equivalent)
following manufacturer's directions. After all metal surfaces inside the injector body
have been thoroughly coated with the deactivation solution, rinse the injector body with
toluene, methanol, acetone, then hexane. Reassemble the injector and replace the
columns.
7.8.4 Column rinsing - The column should be rinsed with several column volumes
of an appropriate solvent. Both polar and nonpolar solvents are recommended. Depending
on the nature of the sample residues expected, the first rinse might be water, followed by
methanol and acetone. Methylene chloride is a good final rinse and in some cases may be the
only solvent required. The column should then be filled with methylene chloride and allowed
to stand flooded overnight to allow materials within the stationary phase to migrate into the
solvent. The column is then flushed with fresh methylene chloride, drained, and dried at room
temperature with a stream of ultrapure nitrogen.
8.0 QUALITY CONTROL
8.1 Refer to Chapter One and Method 8000 for specific quality control (QC) procedures.
Quality control procedures to ensure the proper operation of the various sample preparation
techniques can be found in Method 3500. If an extract cleanup procedure was performed, refer to
Method 3600 for the appropriate quality control procedures. Each laboratory should maintain a
formal quality assurance program. The laboratory should also maintain records to document the
quality of the data generated.
8.2 Quality control procedures necessary to evaluate the GC system operation are found
in Method 8000, Sec. 7.0 and include evaluation of retention time windows, calibration verification,
and chromatographic analysis of samples.
8.3 Initial Demonstration of Proficiency
8.3.1 Each laboratory must demonstrate initial proficiency with each sample
preparation and determinative method combination it utilizes, by generating data of acceptable
accuracy and precision for target analytes in a clean matrix. The laboratory must also repeat
the following operations whenever new staff are trained or significant changes in
instrumentation are made.
8.3.2 K is suggested that the quality control (QC) reference sample concentrate (as
discussed in Sec. 8.0 of Methods 8000 and 3500) contain each analyte of interest at 10 mg/L.
If this method is to be used for analysis of Chlordane or Toxaphene only, the QC reference
sample concentrate should contain the most representative multi-component mixture at a
suggested concentration of 50 mg/L in acetone. See Method 8000, Sec. 8.0 for additional
information on how to accomplish this demonstration.
8.3.3 Calculate the average recovery and the standard deviation of the recoveries
of the analytes in each of the four QC reference samples. Refer to Sec. 8.0 of Method 8000
for procedures for evaluating method performance.
8.4 Sample Quality Control for Preparation and Analysis - The laboratory must also have
procedures for documenting the effect of the matrix on method performance (precision, accuracy,
and detection limit). At a minimum, this includes the analysis of QC samples, including a method
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blank, a matrix spike, a duplicate, and a laboratory control sample (LCS), and the addition of
surrogates to each field sample and QC sample.
8.4.1 Documenting the effect of the matrix should include the analysis of at least
one matrix spike and one duplicate unspiked sample or one matrix spike/matrix spike duplicate
pair. The decision on whether to prepare and analyze duplicate samples or a matrix
spike/matrix spike duplicate must be based on a knowledge of the samples in the sample
batch. If samples are expected to contain target analytes, then laboratories may use one
matrix spike and a duplicate analysis of an unspiked field sample. If samples are not expected
to contain target analytes, the laboratories should use a matrix spike and matrix spike duplicate
pair,
8.4.2 In-house method performance criteria should be developed using the
guidance found in Sec. 8.0 of Method 8000 for procedures for evaluating method performance.
8.4.3 A Laboratory Control Sample (LCS) should be included with each analytical
batch. The LCS consists of an aliquot of a clean (control) matrix simitar to the sample matrix
and of the same weight or volume. The LCS is spiked with the same analytes at the same
concentrations as the matrix spike. When the results of the matrix spike analysis indicates a
potential problem due to the sample matrix itself, the LCS results are used to verify that the
laboratory can perform the analysis in a clean matrix.
8.4.4 Include a calibration standard after each group of 20 samples (it is
mcommended that a calibration standard be included after every 10 samples to minimize the
number of repeat injections) in the analysis sequence as a calibration check. Thus, injections
of method blank extracts, matrix spike samples, and other non-standards are counted in the
total. Solvent blanks, injected as a check on cross-contamination, need not be counted in the
total. The response factors for the calibration should be within ±15% of the initial calibration
(see Sec. 7.5.2). When this calibration verification standard falls out of this acceptance
window, the laboratory should stop analyses and take corrective action.
8.4.5 Whenever quantitation is accomplished using an internal standard, internal
standards must be evaluated for acceptance. The measured area of the internal standard
must be no more than 50 percent different from the average area calculated during calibration.
When the internal standard peak area is outside the limit, all samples that fall outside the QC
criteria must be reanalyzed.
8.4.6 DDT and endrin are easily degraded in the injection port. Breakdown occurs
when the injection port liner is contaminated with high boiling residue from sample injection or
when the injector contains metal fittings. Check for degradation problems by injecting a
standard containing only 4,4-DDT and endrin. Presence of 4,4-DDi, 4,4-DDD, endrin ketone
or endrin indicates breakdown. If degradation of either DDT or endrin exceeds 15%, take
corrective action before proceeding with calibration.
8.4.6.1 Calculate percent breakdown as follows:
% breakdown of DDT - "" °f de9radrt°" P63" areas
-------
8.4.6.2 The breakdown of PPT and endrin should be measured before
samples are analyzed and at the beginning of each 12-hour shift. Injector maintenance
and recalibration should be completed if the breakdown is greater than 15% for either
compound (Sec. 7.8.2).
8.4.7 Whenever silica gel (Method 3630} or Florisii (Method 3620) cleanups are
used, the analyst must demonstrate that the fractionation scheme is reproducible. Batch to
batch variation in the composition of the silica gel or Florisii or overloading the column may
cause a change in the distribution patterns of the organochlorine pesticides. When
compounds are found in two fractions, add the concentrations found in the fractions, and
correct for any additional dilution.
8.4.8 See Method 8000, Sec. 8.0 for the details on carrying out sample quality
control procedures for preparation and analysis.
8.5 Surrogate recoveries
The laboratory must evaluate surrogate recovery data from individual samples versus the
surrogate control limits developed by the laboratory. See Method 8000, Sec. 8.0, for information on
evaluating surrogate data and developing and updating surrogate limits.
8.6 It is recommended that the laboratory adopt additional quality assurance practices for
use with this method. The specific practices that are most productive depend upon the needs of the
laboratory and the nature of the samples. Whenever possible, the laboratory should analyze
standard reference materials and participate in relevant performance evaluation studies.
9.0 METHOD PERFORMANCE
9.1 The method detection limit (MDL) is defined in Chapter One. Each laboratory should
develop its own matrix-specific MDLs using the guidance found in Chapter One. Estimated
quantitation limits (EQLs) may be determined using the factors in Table 3.
9.2 The chromatographic separations in this mettiod have been tested in a single laboratory
by using clean hexane and liquid and solid waste extracts that were spiked with the test compounds
at three concentrations. Single-operator precision, overall precision, and method accuracy were
found to be related to the concentration of the compound and the type of matrix
9.3 The accuracy and precision that can be acheived with this method depend on the
sample matrix, sample preparation technique, optional cleanup techniques, and calibration
procedures used.
9.4 Tables 9 and 10 contain precision and recovery data generated for sewage sludge and
dichloroethane stillbottoms. Table 11 contains recovery data for a clay soil, taken from Reference
10. The spiking concentration was for the clay soil was 500 pg/kg. The spiking solution was mixed
into the soil and then immediately transferred to the extraction device and immersed in the extraction
solvent. The spiked sample was then extracted by Method 3541 (Automated Soxhlet). The data
represent a single determination. Analysis was by capillary column gas chromatography/electron
capture detector following Method 8081 for the organochlorine pesticides.
9.5 Table 12 contains single-laboratory precision and accuracy data for solid-phase
extraction of TCLP buffer solutions spiked at two levels and extracted using Method 3535.
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9.6 Table 13 contains multiple-laboratory data for solid-phase extraction of spiked TCLP
soil leachates extracted using Method 3535.
9.7 Table 14 contains single-laboratory data on groundwater and wastewater samples
extracted by solid-phase extraction, using Method 3535.
9.8 Tables 15 and 16 contain single-laboratory performance data using supercritical fluid
extraction Method 3562. Samples were analyzed by GC/ELCD. The method was performed using
a variable restrictor and solid trapping material (octadecyl silane [ODS]). Three different soil
samples were spiked at 5 and 250 pgftg. Soil 1 (Delphi) is described as loamy sand, with 2.4% clay,
94% sand, 0.9% organic matter, 3.4% silt, and 0.1% moisture. Soil 2 (McCarthy) is described as
sandy-loam, with 11% day, 56% sand, 22% organic matter, 33% silt, and 8.7% moisture. Soil 3
(Aubum) is described as clay loam, with 32% day, 21% sand, 5.4% organic matter, 46% silt, and
2.2% moisture. Seven replicate extractions were made of each soil at the two concentrations.
10.0 REFERENCES
1. Lopez-Avila, V., Baldin, E., Benedicto, J, Milanes, J., Beckert. W.F., "Application of Open-
Tubular Columns to SW-846 GC Methods", report to the U.S. Environmental Protection Agency
on Contract 68-03-3511; Mid-Pacific Environmental Laboratory, Mountain View, CA, 1990.
2. Development and Application of Test Procedures for Specific Organic Toxic Substances in
Wastewaters. Category 10 - Pestiddes and PCB Report for the U.S. Environmental Protection
Agency on Contract 68-03-2606.
3. Goerlitz, D.F., Law, L.M., "Removal of Elemental Sulfur Interferences from Sediment Extracts
for Pestidde Analysis", Bull. Environ. Contam. Toxicol., 1971,6, 9.
4. Jensen, S., Renberg, L, Reutergandth, L., "Residue Analysis of Sediment and Sewage Sludge
for Organochlorines in the Presence of Elemental Sulfur", Anal. Chem., 1977, 49, 316-318.
5. Wise, R.H., Bishop, D.F., Williams, R.T., Austem, B.M., "Gel Permeation Chromatography in
the GC/MS Analysis of Organics in Sludges", U.S. Environmental Protection Agency,
Cindnnati, OH, 45268.
6. Pionke, H.B., Chesters, G., Armstrong, D.E., "Extraction of Chlorinated Hydrocarbon
Insectiddes from Soil", Agron. J., 1968,60, 289.
7. Burke, J.A., Mills, P.A., Sostwick, D.C., "Experiments with Evaporation of Solutions of
Chlorinated Pestiddes", J. Assoc. Off. Anal. Chem., 1966, 49, 999.
8. Glazer, JA, et al., "Trace Analyses for Wastewaters", Environ. Sci. and Techno!., 1981,15,
1426.
9. Mars den, P. J., "Performance Data for SW-846 Methods 8270, 8081, and 8141", U.S.
Environmental Protection Agency, EMSL-Las Vegas, EPA/600/4-90/015.
10. Lopez-Avila, V. (Beckert, W., Project Officer), "Development of a Soxtec Extraction Procedure
for Extracting Organic Compounds from Soils and Sediments", EPA 600/X-91/140, US
Environmental Protection Agency, Environmental Monitoring Systems Laboratory, Las Vegas,
NV, October 1991.
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11. Marked, C., "Solid-Phase Extraction of TCLP Leachates," Proceedings of the Tenth Annual
Waste Testing and Quality Assurance Symposium, Arlington, VA, July, 1994.
12. Bennett, D., Lesnik, B., Lee, S.M., "Supercritical Fluid Extraction of Onganochlorine Pesticide
Residues from Soils," Proceedings of the Tenth Annual Waste Testing and Quality Assurance
Symposium, Arlington, VA, July, 1994.
13. Marked, C., "3M Data Submission to EPA," letter to B. Lesnik, June 27,1995.
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TABLE 1
GAS CHROMATOGRAPHIC RETENTION TIMES FOR THE ORGANOCHLORINE PESTICIDES
USING WIDE-BORE CAPILLARY COLUMNS, SINGLE-COLUMN METHOD OF ANALYSIS

Retention Time (min)
Compound
DB-6089
DB-1701"
Aldrin
11.84
12.50
a-BHC
8.14
9.46
p-BHC
9.%
13.58
5-BHC
11.20
14.39
Y-BHC (Lindane)
9.52
10.84
a-Chlordane
15.24
16.48
Y-Chlordane
14.63
16.20
4,4-DDD
18.43
19.56
4,4*-DDE
16.34
16.76
4,4'-DDT
19.48
20.10
Dieldrin
16.41
17.32
Endosulfan 1
15.25
15.96
Endosulfan II
18.45
19.72
Endosulfan sulfate
20.21
22.36
Endrin
17.80
18.06
Endrin aldehyde
19.72
21.18
Heptachlor
10.66
11.56
Heptachlor epoxide
13.97
15,03
Methoxychlor
22.80
22.34
Toxaphene
MR
MR
MR = Multiple response compound,
8 See Table 4 for GC operating conditions.
8081B -26
Revision 2
January 1998
-------
TABLE 2
GAS CHROMATOGRAPHIC RETENTION TIMES FOR THE ORGANOCHLORINE PESTICIDES
USING NARROW-BORE CAPILLARY COLUMNS, SINGLE-COLUMN METHOD OF ANALYSIS

Retention Time (min)
Compound
DB-608®
DB-5"
Aldrin
14.51
14.70
ct-BHC
11.43
10.94
P-BHC
12.59
11.51
5-BHC
13.69
12.20
Y-BHC (b'ndane)
12.46
11.71
a-Chlordane
NA
NA
y-Chlordane
17.34
17.02
4,4-DDD
21.67
20.11
4,4-DDE
19.09
18.30
4,4-DDT
23.13
21.84
Dieldrin
19.67
18.74
Endosulfan I
18.27
17.62
Endosulfan II
22.17
20.11
Endosulfan sulfate
24.45
21.84
Endrin
21.37
19.73
Endrin aldehyde
23.78
20.85
Heptachlor
13.41
13.59
Heptachlor epoxide
16.62
16.05
Methoxychlor
28.65
24.43
Toxaphene
MR
MR
NA = Data not available.
MR = Multiple response compound.
a See Table 4 for GC operating conditions.
8081B-27
Revision 2
January 1998
-------
TABLE 3
FACTORS FOR DETERMINATION OF ESTIMATED QUANTITATION LIMITS" (EQLs)
FOR VARIOUS MATRICES
Matrix • Factor
Ground water 10
Low-concentration soil by sonication with GPC cleanup 670"
High-concentration soil and sludges by sonication 10,000
Non-water miscible waste 100,000
• Laboratories may estimate the quantitation limits of the target analytes in environmental and
waste media by generating MDLs in organic-free reagent water and using the following equation
(see Sec. S.O of Chapter One for information on generating MDL data):
EQL = [MDL in water] * [factor in this table]
For nonaqueous samples, the factor is on a wet-weight basis. Sample EQLs are highly matrix-
dependent. EQLs determined using these factors are provided as guidance and may not always
be achievable.
8081B - 28
Revision 2
January 1998
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TABLE 4
GC OPERATING CONDITIONS FOR ORGANOCHLOR1NE COMPOUNDS
SINGLE-COLUMN ANALYSIS USING NARROW-BORE COLUMNS
Column 1 - 30 m x 0.25 or 0.32 mm ID fused-silica capillary column chemically bonded with
SE-54 (DB-5 or equivalent), 1 Mm film thickness.
Cairier gas
Carrier gas pressure
Injector temperature
Detector temperature
Initial temperature
Temperature program
Helium
16 psi
225°C
300°C
100°C, hold 2 minutes
100°C to 160°C at 15°C/min, followed by 160°C to 270°C at
5°C/min
Final temperature
270°C
Column 2 - 30 m x 0.25 mm ID fused-silica capillary column chemically bonded with 35
percent phenyl methyipolysiioxane (DB-608, SPB-608, or equivalent), 25 jjm coating
thickness, 1 Mm film thickness.
Carrier gas
Nitrogen
Carrier gas pressure
20 psi
Injector temperature
225°C
Detector temperature
300°C
Initial temperature
160°C, hold 2 minutes
Temperature program
160°C to 290°C at 5°C/min
Final temperature
290°C, hold 1 min
8081B-29 Revision 2
January 1998
-------
TABLE 5
GC OPERATING CONDITIONS FOR ORGANOCHLORINE COMPOUNDS
SINGLE-COLUMN ANALYSIS USING WIDE-BORE COLUMNS
Column 1 - 30 m x 0.53 mm ID fused-siliea capillary column chemically bonded with 35
percent phenyl methylpolysiloxane (DB-608, SPB-608, RTx-35, or equivalent), 0.5 Mm or
0.83 Mm Aim thickness.
Column 2 - 30 m x 0.53 mm ID fused-silica capillary column chemically bonded with 50
percent phenyl methylpolysiloxane (DB-1701, or equivalent), 1.0 pm film thickness.
Both Column 1 and Column 2 use the same GC operating conditions.
Carrier gas
Carrier gas flow rate
Makeup gas
Makeup gas flow rate
Injector temperature
Detector temperature
Initial temperature
Temperature program
Final temperature
Helium
5-7 mL/minute
argon/methane (P-5 or P-10) or nitrogen
30 mL/min
250°C
290°C
150°C, hold 0.5 minute
150°C to 270 °C at 5°C/min
270°C, hold 10 min
Column 3 - 30 m x 0.53 mm ID fused-silica capillary column chemically bonded with SE-54
(DB-5, SPB-5, RTx-5, or equivalent), 1.5 Mm film thickness.
Carrier gas
Carrier gas flow rate
Makeup gas
Makeup gas flow rate
Injector temperature
Detector temperature
Initial temperature
Temperature program
Helium
6 mL/minute
argon/methane (P-5 or P-10) or nitrogen
30 mL/min
205°C
290°C
140°C, hold 2 min
140°C to 240°C at 10°C/min, hold 5 minutes at 240°C, 240°C
to 265°C at 5°C/min
Final temperature
265 °C, hold 18 min
8081B - 30
Revision 2
January 1998
-------
TABLE 6
RETENTION TIMES OF THE ORGANOCHLOR1NE PESTICIDES9
DUAL-COLUMN METHOD OF ANALYSIS
Compound
DB-5 RT (min)
DB-1701 RT (min)
DBCP
2.14
2.84
Hexachlorocyclopentadiene
4.49
4.88
Etridiazole
6.38
8.42
Chloroneb
7.46
10.60
Hexachlorobenzene
12.79
14.58
Diallate
12.35
15.07
Propachlor
9.96
15.43
Trifluralin
11.87
16.26
a-BHC
12.35
17.42
PCNB
14.47
18.20
y-bhc
14.14
20.00
Heptachlor
18.34
21.16
Aldrin
20.37
22.78
Aiachlor
18.58
24.18
Chlorothalonil
15.81
24.42
Aiachlor
18.58
24.18
P-BHC
13.80
25.04
Isodrin
22.08
25.29
DCPA
21.38
26.11
6-BHC
15.49
26.37
Heptachlor epoxide
22.83
27.31
Endosulfan-I
25.00
28.88
y-Chlordane
24.29
29.32
a-Chlordane
25.25
29.82
frans-Nonachlor
25.58
30.01
44-.DDE
26.80
30.40
Dieldrin
26.60
31.20
Perthane
28.45
32.18
Endrin
27.86
32.44
Chloropropylate
28.92
34.14
Chloroberizilate
28.92
34.42
Nitrofen
27.86
34.42
4,4-DDD
29.32
35.32
Endosulfan II
28.45
35.51
4,4-DDT
31.62
36.30
Endrin aldehyde
29.63
38.08
8081B - 31
Revision 2
January 1998
-------
TABLE 6
(continued)
Compound
DB-5 RT (min)
DB-1701 RT (min)
Mirex
37.15
38.79
Endosulfan sulfate
31.62
40.05
Methoxychlor
35.33
40.31
Captafoi
32.65
41,42
Endrin ketone
33.79
42.26
Permethrin
41.50
45.81
Kepone
31.10

Dicofol
35.33

Dichlorie
15.17

a,a -Dibromo-m-xylene
9.17
11.51
2-Bromobiphenyi
8.54
12.49
* See Table 7 for GC operating conditions.
b Not detected at 2 ng per injection.
8081B - 32
Revision 2
January 1998
-------
TABLE 7
GC OPERATING CONDITIONS FOR ORGANOCHLORINE PESTICIDES
FOR DUAL-COLUMN METHOD OF ANALYSIS, LOW TEMPERATURE, THIN FILM
Column 1: DB-1701 or equivalent
30-m x 0.53 mm ID
1.0 film thickness
Column 2: DB-5 or equivalent
30-m x 0.53 mm ID
0.83 tim film thickness
Helium
6 mL/minute
Nitrogen
20 mUmin
250 °C
320°C
140°C, hold 2 minutes
140°C to 270°C at 2.8°C/min
270°C, hold 1 minute
Carrier 93s
Carrier gas flow rate
Makeup gas
Makeup gas flow rate
Injector temperature
Detector temperature
Initial temperature
Temperature program
Final temperature
8081B - 33
Revision 2
January 1998
-------
TABLE 8
GC OPERATING CONDITIONS FOR ORGANOCHLORINE PESTICIDES
FOR THE DUAL COLUMN METHOD OF ANALYSIS
HIGH TEMPERATURE, THICK FILM
Column 1:
DB-1701 (J&W) or equivalent

30 mx 0.53 mm ID

1.0 (inn film thickness
Column 2:
DB-5 (J&W) or equivalent

30 mx0,53mm ID

1.5 |jm film thickness
Carrier gas:
Helium
Carrier gas flow rate:
6 mL/minute
Makeup gas:
Nitrogen
Makeup gas flow rate:
20 (mL/min)
Injector temperature:
250°C
Detector temperature:
320°C
Initial temperature:
150°C, hold 0.5 min
Temperature program:
150°C to 190°C at 12°C/min, hold 2 min190°C to 275°C

at 4°C/min
Final temperature
275°C, hold 10 min
8081B - 34
Revision 2
January 1998
-------
TABLE 9
AIMALYTE RECOVERY FROM SEWAGE SLUDGE
Ultrasonic Extraction Soxhlet
Compound %Recovery %RSD %Recovery %RSD
Hexachloroethane
80
7
79
1
2-Chloronapthalene
50
56
67
8
4-Bromodiphenyl ether
118
4
nd
nd
a-BHC
88
25
265
18
y-bhc
55
9
155
29
Heptachlor
60
13
469
294
Aldrin
92
33
875
734
P-BHC
351
71
150
260
5-BHC
51
11
57
2
Heptachlor epoxide
54
11
70
3
Endosulfan I
52
11
70
4
y-Chlordane
50
9
65
1
a-Chiordane
49
8
66
0
DDE
52
11
74
1
Dieldrin
89
19
327
7
Endrin
56
10
92
15
Endosulfan II
52
10
88
11
DDT
57
10
95
17
Endrin aldehyde
45
6
42
10
DDD
57
11
99
8
Tetrachloro-m-xylene
71
19
82
1
Decachlorobiphenyl
26
23
28
48
Concentration spiked in the sample: 500-1000 ng/g, three replicates analyses.
Soxhlet extraction by Method 3540 with methylene chloride.
Ultrasonic extraction by Method 3550 with methylene chloride/acetone (1:1).
Cleanup by Method 3640.
GC column: DB-608, 30 mx 0.53 mm ID.
8081B - 35
Revision 2
January 1998
-------
TABLE 10
ANALYTE RECOVERY FROM DICHLOROETHANE STILLBOTTOMS
Compound
Ultrasonic Extraction
%Recovery %RSD
Soxhlet
%Recovery
%RSD
Hexachloroethane
70
2
50
30
2-Chloronapthalene
59
3
35
35
4-Bromodiphenyl ether
159
14
128
137
a-BHC
55
7
47
25
P-BHC
43
6
30
30
Heptachlor
48
6
55
18
Aldrin
48
5
200
258
P-BHC
51
7
75
42
5-BHC
43
4
113
129
Heptachlor epoxide
47
6
66
34
Endosulfan I
47
4
41
18
Y-Chlordane
48
5
47
13
a-Chlondane
45
5
37
21
DDE
45
4
70
40
Dieldrin
45
5
58
24
Endrin
50
6
41
23
Endosulfan II
49
5
46
17
DDT
49
4
40
29
Endrin aldehyde
40
4
29
20
DDD
48
5
35
21
Tetrachloro-m-xylene
49
2
178
211
Decachlorobiphenyt
17
29
104
93
Concentration spiked in the sample: 500-1000 ng/g, three replicates analyses.
Soxhlet extraction by Method 3540 with methylene chloride.
Ultrasonic extraction by Method 3550 with methylene chloride/acetone (1:1).
Cleanup by Method 3640.
GC column: DB-608, 30 m x 0.53 mm ID.
80815 - 36
Revision 2
January 1998
-------
TABLE 11
SINGLE LABORATORY ACCURACY DATA FOR THE EXTRACTION OF
ORGANOCHLORINE PESTICIDES FROM SPIKED CLAY SOIL BY METHOD 3541
(AUTOMATED SOXHLET)®
% Recovery
Compound Name DB-5 DB-1701
a-BHC
89
94
p-BHC
86
ND
Heptachlor
94
95
Aldrin
ND
92
Heptachlor epoxide
97
97
trans-Chlordane
94
95
Endosulfan I
92
92
Dieldrin
ND
113
Endrin
111
104
Endosulfan II
104
104
4,4-DDT
ND
ND
Mirex
108
102
' The operating conditions for the automated Soxhlet were:
Immersion time 45 min; extraction time 45 min; 10 g sample size; extraction solvent, 1:1
acetone/hexane. No equilibration time following spiking.
ND = Not able to determine because of interference.
AH compounds were spiked at 500 pg/kg.
Data taken from Reference 10.
8081B - 37
Revision 2
January 1998
-------
TABLE 12
SINGLE LABORATORY RECOVERY DATA FOR SOUD-PHASE EXTRACTION OF
ORGANOCHLORINE PESTICIDES FROM TCLP BUFFERS SPIKED AT TWO LEVELS

Spike Level
(Mg/L)
Buffer 1 (pH =
2.886)
Buffer 2 (pH =
4.937)
Compound
Recovery (%)
RSD
Recovery (%)
, RSD
Low Level Spike
Toxaphene
250
86
13
77
17
Chlordane
15
88
7
95
6
Y-BHC (Lindane)
200
115
7
98
5
Heptachior
4
95
11
77
23
Heptaehlor
epoxide
4
107
9
104
12
Endrin
10
89
5
100
6
Methoxychlor
5000
97
8
95
6
High Level Spike
Toxaphene
1000
106
7
85
15
Chlordane
60
116
12
107
12
Y-BHC (Lindane)
800
109
19
112
5
Heptachior
16
113*
18*
93
3
Heptachior
epoxide
16
82
17
91
7
Endrin
40
84
19
82
4
Methoxychlor
20,000
100
4
87
8
Results were from seven replicate spiked buffer samples, except where noted with *, which indciates
that only three replicates were analyzed.
8081B - 38
Revision 2
January 1998
-------
TABLE 13
RECOVERY DATA FROM THREE LABORATORIES FOR SOLID-PHASE EXTRACTION OF ORGANOCHLORINE PESTICIDES
FROM SPIKED TCLP LEACHATES FROM SOIL SAMPLES

Spike Level
(pg/L)*

Lab 1

Lab 2

Lab 3

Compound
%R
RSD
n
%R
RSD
n
%R
RSD
n
Buffer 1 pH = 2.886
Toxaphene
500
75
25
7
95.4
2.4
3
86.0
4.3
3
Chlordane
30
80
15
7
57.8
12.0
3
73.8
0.9
3
Y-BHC (Lindane)
400
104
11
7
99.3
0.6
3
86.6
6.4
3
Heptachlor
8
88
13
7
70.8
20.4
3
88.0
9.1
3
Heptachlor epoxide
8
92
13
7
108.7
6.9
3
75.0
2.8
3
Endrin
20
106
12
7
110
0
3
78.3
4.6
3
Methoxychlor
10,000
107
12
7
86.7
2.2
3
84.8
8.5
3
Buffer 2 pH = 4.937
Toxaphene
500
87
9
7
98
4.1
3
88.8
4.1
3
Chlordane
30
91
8
7
66.7
5.0
3
73.7
11.5
3
Y-BHC (Lindane)
400
74
20
7
102.7
2.2
3
89.3
3.1
3
Heptachlor
8
71
21
7
62.5
20
3
85.0
1.5
3
Heptachlor epoxide
8
118
1
3
113
0
3
81.3
2.7
3
Endrin
20
124
7
3
111.7
2.6
3
83.0 ,
3.4
3
Methoxychlor
10,000
73
22
7
88.8
2.7
3
89.6
2.7
3
* 250-mL aliquots of leachate were spiked by Labs 2 and 3 at the levels show. Lab 1 spiked at one-half these levels.
8081B- 39
Revision 2
January 1998
-------
TABLE 14
SINGLE LABORATORY ACCURACY AND PRECISION DATA FOR SOLID-PHASE EXTRACTION BY METHOD 35351
Bias (%) Precision (%) MDL
Compound
Ground
water
(low)
Ground
water
(high)
Waste
water
(low)
Waste
water
(high)
Ground
water
(low)
Ground
water
(high)
Waste
water
(low)
Waste
water
(high)
Ground
water
(Mfl/L)
Waste
water
(pg/L)
Aldrin
37.3
93.5
79.3
94.0
23.7
5.5
6.7
3.4
1.4
0.83
P-BHC
89.2
107.8
79.7
82.3
6.5
2.5
1.6
4.2
0.91
0.20
5-BHC
106.2
86.0
88.9
83.4
5.6
2.4
2.5
4.2
0.93
0.35
a-Chlordane
75.4
112.3
78.9
89.5
12.8
2.7
4.7
2.4
1.5
0.58
y-Chlordane
70.7
98.9
79.9
93.9
15.8
2.7
4.6
2.9
1.8
0.58
Dieldrin
83.4
96.1
81.2
93.3
7.1
2.3
3.8
3.6
0.9
0.49
Endosulfan I
79.6
99.1
79.6
87.9
10.6
2.3
4.1
3.8
1.3
0.51
Endosulfan II
94.5
101.6
82.7
93.5
5.8
2.8
4.2
4.1
0.9
0.54
Endrin
88.3
98.4
85.1
89.6
6.2
2.3
3.1
2.9
1.7
0.82
Endrin aldehyde
87.5
99.9
69.0
80.2
6.0
4.0
3.3
5.9
0.8
0.36
Heptachlor
43.1
95.4
71.8
78.6
19.2
3.9
5.0
2.8
1.3
0.56
Heptachlor
epoxide
76.4
97.6
75.3
83.4
12.1
2.4
2.9
3.3
1.5
0.34
Lindane
81.3
115.2
82.1
85.3
11.1
3.2
2.4
3.1
1.4
0.32
p,p'-DDE
80.3
96.0
85.1
97.9
8.3
2.5
4.4
2.4
1.0
0.59
P.P'-DDT
86.6
105.4
105
111
4.4
2.7
4.3
4.7
0.6
0.71
p,p'-TDE (DDD)
90.5
101.1
74.9
79.6
4.8
2.4
4.6
2.9
1.4
0.85
1AII results determined from seven replicates of each sample type. Two spiking levels were used. "Low" samples were spiked at 5-10 )jg/L for
each analyte, while "high" samples were spiked at 250 - 500 fjg/L. MDL values were determined from the "low" samples without further
consideration of the spiking level.
8081B- 40
Revision 2
January 1998
-------
TABLE 15
RECOVERY (BIAS) OF ORGANOCHLORINE PESTICIDES USING SFE METHOD 3562
(Seven replicates)
Compounds
Delphi8
250 ug/kg
Delphi-5a
5 ug/kg
McCarthy"
250 ug/kg
McCarthy"
5 ug/kg
Auburn®
250 ug/kg
Auburn0
5 ug/kg
Mean
Recovery
y-bhc
102.6
66.4
80.7
82.7
86.0
86.1
84.1
P-BHC
101.9
73.0
86.1
85.1
87.4
86.3
86.6
Heptachlor
101.3
61.6
78.0
79.1
83.3
80.4
80.6
5-BHC
120.9
82.3
90.4
89.6
92.9
89.4
94.2
Aldrin
56.7
28.7
52.1
77.1
42.1
74.6
55.2
Heptachlor epoxide
102.3
71.9
87.1 .
87.4
89.6
91.1
88.2
a-Chlordane
106.4
87.1
88.1
105.9
91.7
97.1
96.1
4,4'DDE
110.9
75.7
88.4
118.7
83.6
110.9
98.0
Dieldrin
106.9
80.4
88.1
140.8
90.6
80.1
97.8
Endrin
211.0
87.0
111.7
98.7
90.5
87.6
114.4
4,4-DDD
93.0
80.4
85.0
88.1
83.7
90.4
86.8
Endosulfan II
105.6
89.9
92.1
88.6
87.7
92.9
92.5
4,4-DDT
126.7 ,
81.3
110.9
199.7
83.6
124.3
121.1
Endrin aldehyde
64.3
74.0
63.0
86.7
21.0
38.3
37.9
Matrix Mean Recovery
107.9
74.3
85.9
102.0
79.8
87.8
89.5
a Delphi: Loamy sand soil
b McCarthy: Sandy loamy-organic rich soil
0 Auburn: Clay-loamy soil
8081B - 41
Revision 2
January 1998
-------
TABLE 16
RELATIVE STANDARD DEVIATION (PRECISION) OF ORGANOCHLORINE PESTICIDES USING SFE METHOD 3562
(Seven replicates)
Compounds
Delphi8
250 ug/kg
Delphi0
5 ug/kg
McCarthy"
250 ug/kg
McCarthy"
5 ug/kg
Auburn0
250 ug/kg
Auburn®
5 ug/kg
Mean
y-bhc
3.9
3.3
3.3
6.5
4.0
1.6
3.7
P-BHC
6.5
3.0
3.0
4.3
4.6
2.0
3.9
Heptachlor
4.4
2.1
4.3
5.0
4.4
2.6
3.8
6-BHC
5.3
3.1
3.3
7.1
4V
3.5
4.4
Aldrin
2.9
5.5
2.8
4.6
1.6
1.9
3.2
Heptachlor epoxide
3.0
2.7
3.6
4.3
4.7
4.2
3.8
a-Chlordane
3.6
5.7
4.8
13.8
4.2
2.5
5.8
4,4'DDE
5.2
15.3
4.8
4.2
7.7
3.4
6.8
Dieldrin
4.3
4.5
2.9
23.9
5.0
3.1
7.3
Endrin
7.2
6.0
4.5
6.0
4.3
10.5
6.4
4,4-DDD
6.9
3.1
3.7
3.5
4.3
7.4
4.8
Endosulfan II
5.1
4.7
3.2
3.3
5.5
4.6
4.4
4,4'-DDT
12.5
6.2
6.6
5.9
4.9
3.4
6.6
Endrin aldehyde
3.9
7.5
4.7
11.6
1.9
26.0
9.3
Matrix Mean Recovery
5.3
5.2
4.0
7.4
4.4 ,
5.5
5.3
a Delphi: Loamy sand soil
" McCarthy: Sandy loamy-organic rich soil
0 Auburn: Clay-loamy soil
8081B - 42
Revision 2
January 1998
-------
FIGURE 1
GAS CHROMATOGRAM OF THE MIXED ORGANOCHLORINE PESTICIDE STANDARD
Start Tin* : 0.00 m'tn
Scale Factor: Q
End Mm : 33.60 min
Plot Offset: 20 mto
Low p&int : 20,00 to/
Plot Scale: 409 WV
Nigfi Point : 420.00 i
Response [mV]
o-
L?l-
33
ft
fD
~
6' -i.
H
n>
K3
O"
K>
U»"
Ut
O"
to
OiDOlOmOCJiQ
TO Q O O O O Q
%MHf HMlT %pr %bmp VrmT
' ' I i I i i m I i i i i h i i i I i i i i I i i i i I i i i i I i
K>
U!
CK
o
CJ1
i.i i I n
MM
4.68
5wl>7
-0.95
-7.99
-8.60
67
3-10.47
-9.93
-10.78
-11. 05
-17.63
18.56
11.81
13.65
14.34
14.92
16.32
17.17
15=23
^207j.6
21.20
21.93
22.77
23.80
26.23
28.64
23.18
30.19
Column:
Temperature program:
30 m x 0.25 mm ID, DB-5
100°c (hold 2 minutes) to 160°C at 15cC/min, then at 5aC/min to 270°C;
carrier He at 16 psi
8081B-43
Revision 2
January 1998
-------
FIGURE 2
GAS CHROMATOGRAM OF INDIVIDUAL ORGANOCHLORINE PESTICIDE STANDARD MIX A
Start Time : 0.00 min
Scale Factor: 0
End Time t 33.00 min
Plot Offset: 20 ttN
Lay Point : 20.00 mV
Plot Scale: 250 mv
High Point : 275.00 mv
Response [mV]
Oi
o
j_i_L
o
o
) I I. I
-» K>
m o
o o
I 1 I I I 1 1
to
en
O '
'ill
Ul-
71
03
nT
o
~ v>
H
3
tD
I—I KJ_
3 °
m
LJ
or
5.13
•7.93

<*¦60
-12.33
-14.27
-17.08
.22
77
I
"2^—22. 68
-23.73
28.52
-3.3$
-S.54
-9.86
-10.98
-13.58
-17.54
-19.24
-19.78
-21.13
-23.OS
-30.05
Column:
Temperature program:
30 m x 0.2S mm ID, DB-5
10Q°C (hold 2 minutes) to 160°C at 15°C/min, then at 5°C/min to 270°C;
carrier He at 16 psi.
8081B - 44
Revision 2
January 1998
-------
FIGURE 3
GAS CHROMATOGRAM OF INDIVIDUAL ORGANOCHLORINE PESTICIDE STANDARD MIX B
Start Time .; 0,03 mm End Time : 33.OD rein Low Point : 2C.QQ mV High Pairs : 270.30.rr.v
Scale Factor; 0 Plot Offset: 20 mv Plot Scale: 250 mv
Response [mV]
-i -» ro K5
oi q m o (_rt
¦2.74
CJ1'
=-5.13
i-lvli
s—6.97
•8.54
•9. 60
o
-10.71
¦11.73
14.27
¦14.84
•16.23
-—17.08
—17.63
¦18. 31
¦19.11
19.54
¦20.19
¦=£20.69
4-22.00
-21.03
-22.68
30,04

Column: 30 mx 0.25 mm ID, DB-5
Temperature program: 100°C (hold 2 minutes) to 160°C at 15°C/min, then at 5°C/min to 270°C;
carrier He at 16 psi.
8081B - 45
Revision 2
January 1998
-------
FIGURE 4
GAS CHROMATOGRAM OF TOXAPHENE
7.00-
5.00—
4.00—
3.00—
0.00
2.00
4.00
B.00
12.00
14.00
Retention time In minutes
Toxaphene analyzed on an SPB-608 fused-silica open-tubular column. The GC operating conditions were
as follows: 30 m x 0,53 mm ID SPB-608. Temperature program: 200°C (2 mln hold) to 290°C at 6°C/min.
8081B-46
Revision 2
January 1998
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FIGURE 5
GAS CHROMATOGRAM OF STROBANE
V
m
*
r-
* ©
til im

DB-1701
0"
O
T

U4ji 41

DB-5
i> O-u
uil'
fWI A) .® d
, •
-------
FIGURES
GAS CHROMATOGRAM OF ORGANOCHLORINE PESTICIDES
t *
IS 5t>

L
u
OB-5
13 IS
^ -»A
24 S
Ittti
25
3* 3} *2
}i
1?
JS

U
u
M
13
41
«
20
12 i I ill IS I
~V*Jl

1AJ
DB-1701
10 11 12 IS

Jl It 4; 41
»
»
40
y
all
20,
JL
Organochlorine pesticides analyzed on a DB-5/DB-1701 fused-silica open-tubular column pair. The GC
operating conditions were as follows: 30 m x 0.53 mm ID DB-5 (0.83-ym film thickness) and 30 m x 0.53
mm ID DB-1701 (1.0-pm film thickness) connected to an 8 in. injection tee (Supelco Inc.). Temperature
program: 140°C (2 min hold) to 270°C (1 min hold) at 2.8°C/min.
8081B-48
Revision 2
January 1998
-------
METHOD 8081B
ORGANOCHLORINE PESTICIDES BY GAS CHROMATOGRAPHY
' 7.5.91s N
peak at least
2.5 times
noise?
No
Yes
/ 7.6 Any \
multicomponent
analytes
v present?
Yes
Stop
7.2 Routine cleanup/
fractionation.
7,4.3 Inject each
calibration standard.
7.4.2 Establish GC
operating conditions.
7.4.1 Refer to Method 8000
for proper calibration
techniques.
7.4.6 Calculate determination
time windows for
each analyte.
7.4.5 Calculate
calibration factors
for each analyte.
7.3 Add specified
matrix spike to sample.
7.3 Choose single-column
or dual-column GC
configuration.
7.5.8 Additional
cleanup or
concentration.
7.6.1 - 7.6.4 Calculate
concentration of
Toxaphene, Strobane,
Chlordane, BHC, or DDT.
7.5.4 Inject an aliquot
of sample extract.
7.1 Choose
appropriate extraction
technique (see Chapter 2
and Method 3500).
7.5.7 Bracket the
sample analysis with
calibration standards
(every 10 samples).
7.5.5 - 7.5.6 Identify
and quantify the peaks
observed in the
ehromatogram.
8081B- 49 Revision 2
January 1998
-------
METHOD 8082A
POLYCHLORINATED BIPHENYLS fPCBs> BY GAS CHROMATOGRAPHY
1.0 SCOPE AND APPLICATION
1.1 Method 8082 is used to determine the concentrations of polychlorinated biphenyls
(PCBs) as Aroclors or as individual PCB congeners in extracts from solid and aqueous matrices.
Open-tubular, capillary columns are employed with electron capture detectors (ECD) or electrolytic
conductivity detectors (ELCD). When compared to packed columns, these fused-silica, open-tubular
columns offer improved resolution, better selectivity, increased sensitivity, and faster analysis. The
target compounds listed below may be determined by either a single- or dual-column analysis
system. The PCB congeners listed below have been tested by this method, and the method may
be appropriate for additional congeners.
Compound
CAS Registry No."
IUPAC#
Aroclor 1016
12674-11-2
-
Aroclor 1221
11104-28-2
-
Aroclor 1232
11141-16-5
-
Aroclor 1242
53469-21-9
-
Aroclor 1248
12672-29-6
-
Aroclor 1254
11097-69-1
-
Aroclor 1260
11096-82-5
-
2-Chlorobiphenyl
2051-60-7
1
2,3- Dichlorobi phenyl
16605-91-7
5
2,2',5-Trichlorobiphenyl
37680-65-2
18
2,4',5-T richlorobiphenyl
16606-02-3
31
2,2,,3,5'-Tetrachlorobiphenyl
41464-39-5
44
2,2', 5,5'-Tetrachlorobiphenyl
35693-99-3
52
2,3" ,4,4'-Tetrachlorobiphenyl
32598-10-0
66
2,2', 3,4,5-Pentachlorobiphenyl
38380-02-8
87
2,2',4,5,5-Pentachlorobiphenyl
37680-73-2
101
2,3,3' ,4',6-Pentachlorobi phenyl
38380-03-9
110
2,2,,3,4,4'15'-Hexachlorobiphenyl
35065-28-2
138
2,2',3,4,5,5-HexachlorobiphenyI
52712-04-6 "
141
2,2',3,5,5',6-Hexachlorobipheny!
52663-63-5
151
2,2,,4,4\5,5'-Hexachlorobiphenyl
35065-27-1
153
2,2',3, S'^', 5-Heptachlorobiphenyl
35065-30-6
170
2,2',3,4,4,,5,5'-Heptachlorobiphenyl
35065-29-3
180
2,2',3,4,4',5',6-Heptachlorobiphenyl
52663-69-1
183
2,2',3,4',5,5',6-Heptachlorobiphenyl
52663-68-0
187
2,2',3,3,,4,4,!5,5,,6-Nonachlorobiphenyl
40186-72-9
206
8082A -1
Revision 1
January 1998
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1.2 Aroclors are multi-component mixtures. When samples contain more than one Aroclor,
a higher level of analyst expertise Is required to attain acceptable levels of qualitative and
quantitative analysis. The same is true of Arodors that have been subjected to environmental
degradation ("weathering") or degradation by treatment technologies. Such weathered multi-
component mixtures may have significant differences in peak patterns than those of Aroclor
standards.
1.3 Quantitation of PCBs as Aroctors is appropriate for many regulatory compliance
determinations, but is particularly difficult when the Aroclors have been weathered by long exposure
in the environment. Therefore, this method provides procedures for the determination of selected
individual PCB congeners. The 19 PCB congeners listed above have been tested by this method.
1.4 The PCB congener approach potentially affords greater quantitative accuracy when
PCBs are known to be present. As a result, this method may be used to determine Aroclors, some
PCB congeners, or "total PCBs," depending on regulatory requirements and project needs. The
congener method is of particular value in determining weathered Aroclors. However, analysts should
use caution when using the congener method when regulatory requirements are based on Aroclor
concentrations.
1.5 Compound identification based on single-column analysis should be confirmed on a
second column, or should be supported by at least one other qualitative technique. This method
describes analytical conditions for a second gas chromatographic column that can be used to
confirm the measurements made with the primary column. GC/MS Method 8270 is also
recommended as a confirmation technique when sensitivity permits (Sec. 8.0).
1.6 This method also describes a dual-column option. The option allows a hardware
configuration of two analytical columns joined to a single injection port. The option allows one
injection to be used for dual-column analysis. Analysts are cautioned that the dual-column option
may not be appropriate when the instrument is subject to mechanical stress, many samples are to
be run in a short period, or when highly contaminated samples are analyzed.
1.7 The analyst must select columns, detectors and calibration procedures most
appropriate for the specific analytes of interest in a study. Matrix-specific performance data must
be established and the stability of the analytical system and instrument calibration must be
established for each analytical matrix (e.g., hexane solutions from sample extractions, diluted oil
samples, etc.). Example chromatograms and GC conditions are provided as guidance.
1.8 The MDLs for Aroclors vary in the range of 0.054 to 0.90 |jg/L in water and 57 to 70
pg/kg in soils. Estimated quantitation limits may be determined using the data in Table 1.
1.9 This method is restricted to use by, or under the supervision of, analysts experienced
in the use of gas chromatographs (GC) and skilled in the interpretation of gas chromatograms. Each
analyst must demonstrate the ability to generate acceptable results with this method.
2.0 SUMMARY OF METHOD
2.1 A measured volume or weight of sample (approximately 1 L for liquids, 2 g to 30 g for
solids) is extracted using the appropriate matrix-specific sample extraction technique.
8082A - 2
Revision 1
January 1998
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2.2 Aqueous samples are extracted at neutral pH with methylene chloride using a
separatory funnel (Method 3S10), a continuous liquid-liquid extractor (Method 3520), solid-phase
extraction (Method 3535), or other appropriate technique.
2.3 Solid samples are extracted with hexane-acetone (1:1) or methylene chloride-acetone
(1:1) using a Soxhlet extractor (Method 3540), an automated Soxhlet (Method 3541), supercritical
fluid extraction (Method 3562), or other appropriate technique.
2.4 Extracts for PCB analysis may be subjected to a sulfuric add/potassium permanganate
cleanup (Method 3665) designed specifically for these analytes. This cleanup technique will remove
(destroy) many single component organochlorine or organophosphorus pesticides. Therefore,
Method 8082 is not applicable to the analysis of those compounds. Instead, use Method 8081.
2.5 After cleanup, the extract is analyzed by injecting a 2-mL aliquot into a gas
chromatograph with a narrow- or wide-bore fused-silica capillary column and electron capture
detector (GC/ECD).
2.6 The chromatographic data may be used to determine the seven Aroclors in Sec. 1.1,
individual PCB congeners, or total PCBs.
3.0 INTERFERENCES
3.1 Refer to Methods 3500 (Sec. 3.0, in particular), 3600, and 8000 for a discussion of
interferences.
3.2 Interferences co-extracted from the samples will vary considerably from matrix to
matrix. While general cleanup techniques are referenced or provided as part of this method, unique
samples may require additional cleanup approaches to achieve desired degrees of discrimination
and quantitation. Sources of interference in this method can be grouped into three broad categories.
3.2.1 Contaminated solvents, reagents, or sample processing hardware.
3.2.2 Contaminated GC earner gas, parts, column surfaces, or detector surfaces.
3.2.3 Compounds extracted from the sample matrix to which the detector will
respond.
3.3 Interferences by phthalate esters introduced during sample preparation can pose a
major problem in PCB determinations.
3.3.1 Common flexible plastics contain varying amounts of phthalate esters which
are easily extracted or leached from such materials during laboratory operations. Interferences
from phthalate esters can best be minimized by avoiding contact with any plastic materials and
checking all solvents and reagents for phthalate contamination.
3.3.2 Exhaustive cleanup of solvents, reagents and glassware may be required to
eliminate background phthalate ester contamination.
3.3.3 These materials can be removed through the use of Method 3665 (sulfuric
acid/permanganate cleanup).
8082A - 3
Revision 1
January 1998
-------
3.4 Cross-contamination of clean glassware routinely occurs when plastics are handled
during extraction steps, especially when solvent-wetted surfaces are handled. Glassware must be
scrupulously cleaned.
Clean all glassware as soon as possible after use by rinsing with the last solvent used. This
should be followed by detergent washing with hot water, and rinses with tap water and organic-free
reagent water. Drain the glassware, and dry it in an oven at 130°C for several hours, or rinse with
methanol and drain. Store dry glassware in a dean environment.
NOTE: Oven-drying of glassware used for PCB analysis can increase contamination because
PCBs are readily volatilized in the oven and spread to other glassware. Therefore,
exercise caution, and do not dry glassware from samples containing high concentrations
of PCBs with glassware that may be used for trace analyses.
3.5 Elemental sulfur (S8) is readily extracted from soil samples and may cause
chromatographic interferences In the determination of PCBs. Sulfur can be removed through the
use of Method 3660.
4.0 APPARATUS AND MATERIALS
4.1 Gas chromatograph - An analytical system complete with gas chromatograph suitable
for on-column and split-splitless injection and all required accessories including syringes, analytical
columns, gases, electron capture detectors (ECD), and recorder/integrator or data system.
4.2 GC columns
This method describes procedures for both single-column and dual-column analyses. The
single-column approach involves one analysis to determine that a compound is present, followed
by a second analysis to confirm the identity of the compound (Sec. 8.4 describes how GC/MS
confirmation techniques may be employed). The single-column approach may employ either narrow-
bore (s 0.32 mm ID) columns or wide-bore (0.53 mm ID) columns. The dual-column approach
involves a single injection that is split between two columns that are mounted in a single gas
chromatograph. The dual-column approach employs only wide-bore (0.53 mm ID) columns. A third
alternative is to employ dual columns mounted in a single GC, but with each column connected to
a separate injector and a separate detector.
The columns listed in this section were the columns used to develop the method performance
data, listing these columns in this method is not intended to exclude the use of other columns that
may be developed. Laboratories may use other capillary columns provided that they document
method performance (e.g., chromatographic resolution, analyte breakdown, and MDLs) that equals
or exceeds the performance specified in this method.
4.2.1 Narrow-bore columns for single-column analysis (use both columns to confirm
compound identifications unless another confirmation technique such as GC/MS is employed).
Narrow bore columns should be installed in split/splitless (Grob-type) injectors.
4.2. ¦1.11 30 m x 0.25 or 0.32 mm ID fused-sica capillary column chemically
bonded with SE-54 (DB-5 or equivalent), 1 Mm film thickness.
8082A-4
Revision 1
January 1998
-------
4.2.1.2 30 m x 0.25 mm ID fused-silica capillary column chemically
bonded wfth 35 percent phenyl methylpolysiloxane (DB-608, SPB-608, or equivalent),
2.5 pm coating thickness, 1 pm film thickness.
4.2.2 Wide-bore columns for single-column analysis (use two of the three columns
listed to confirm compound identifications unless another confirmation technique such as
GC/MS is employed). Wide-bore columns should be installed in 1/4 inch injectors, with
deactivated liners designed specifically for use with these columns.
4.2.2.1 30 m x 0.53 mm ID fused-silica capillary column chemically
bonded with 35 percent phenyl methylpolysiloxane (DB-608, SPB-608, RTx-35, or
equivalent), 0.5 pm or 0.83 pm film thickness.
4.2.2.2 30 m x 0.53 mm ID fused-silica capillary column chemically
bonded with 14% cyanopropylmethylpoiysiloxane (DB-1701, or equivalent), 1.0 pm Aim
thickness.
4.2.2.3 30 m x 0.53 mm ID fused-silica capillary column chemically
bonded with SE-54 (DB-5, SPB-5, RTx-5, or equivalent), 1.5 film thickness.
4.2.3 Wide-bore columns for dual-column analysis (choose one of the two pairs of
columns listed below).
4.2.3.1 Column pair 1
30 m x 0.53 mm ID fused-silica capillary column chemically bonded with SE-
54 (DB-5, SPB-5, RTx-5, or equivalent), 1.5 pm film thickness.
30 m x 0.53 mm ID fused-silica capillary column chemically bonded with 14%
cyanopropylmethylpoiysiloxane (DB-1701, or equivalent), 1.0 pm film thickness.
Column pair 1 is mounted in a press-fit Y-shaped glass 3-way union splitter
(J&.W Scientific, Catalog Nq . 7Q5-Q733\ ac a Y-ska-qecL
Catalog No. 20405), or equivalent
4.2.3.2 Column pair 2
30 m x 0.53 mm ID fused-silica capillary column chemically bonded with SE-
54 (DB-5, SPB-5, RTx-5, or equivalent), 0.83 pm film thickness.
30 m x 0.53 mm ID fused-siiica capillary column chemically bonded with 14%
cyanopropylmethylpoiysiloxane (DB-1701, or equivalent), 1.0 pm film thickness.
Column pair 2 is mounted in an 8 in. deactivated glass injection tee (Supelco,
Catalog No. 2-3665M), or equivalent.
4.3 Column rinsing kit - Bonded-phase column rinse kit (J&W Scientific, Catalog No. 430-
3000), or equivalent.
4.4 Volumetric flasks - 10-mL and 25-mL, for preparation of standards.
8082A-5 , Revision 1
January 1998
-------
5.0 REAGENTS
5.1 Reagent grade or pesticide grade chemicals shall be used in all tests. Unless
otherwise indicated, K is intended that all reagents shaft conform to specifications of the Committee
on Analytical Reagents of the American Chemical Society, where such specifications are available.
Other grades may be used, provided K is first ascertained that the reagent is of sufficiently high purity
to permit its use without lessening the accuracy of the determination.
NOTE: Store the standard solutions (stock, composite, calibration, internal, and surrogate
standards) at 4°C in polytetrafluoroethylene (PTFE)-sealed containers in the dark. When
a lot of standards is prepared, it is recommended that aliquots of that lof be stored in
individual small vials. All stock standard solutions must be replaced after one year or
sooner if routine QC (Sec. 8.0) indicates a problem. All other standard solutions must be
replaced after six months or sooner if routine QC (Sec. 8.0) indicates a problem.
5.2 Sample extracts prepared by Methods 3510, 3520, 3540, 3541, 3545, or 3550 need to
undergo a solvent exchange step prior to analysis. The following solvents are necessary for dilution
of sample extracts. AH solvent lots should be pesticide quality or equivalent and should be
determined to be phthalate-free.
5.2.1 n-Hexane, C6H14
5.2.2 Isooctane, (CH3)3CCH2CH(CH3)2
5.3 The following solvents may be necessary for the preparation of standards. All solvent
lots must be pesticide quality or equivalent and should be determined to be phthalate-free.
5.3.1 Acetone, (CH^CO
5.3.2 Toluene, CeHsCH3
5.4 Organic-free reagent water- All references to water in this method refer to organic-free
reagent water as defined in Chapter One.
5.5 Stock standard solutions (1000 mg/L) - May be prepared from pure standard materials
or can be purchased as certified solutions.
5.5.1 Prepare stock standard solutions by accurately weighing about 0.0100 g of
pure compound. Dissolve the compound in isooctane or hexane and dilute to volume in a 10-
mL volumetric flask. If compound purity is 96 percent or greater, the weight may be used
without correction to calculate the concentration of the stock standard solution.
5.5.2 Commercially-prepared stock standard solutions may be used at any
concentration if they are certified by the manufacturer or by an independent source.
5.6 Calibration standards for Aroclors
5.6.1 A standard containing a mixture of Aroclor 1016 and Aroclor 1260 will include
many of the peaks represented in the other five Aroclor mixtures. As a result, a multi-point
initial calibration employing a mixture of Aroclors 1016 and 1260 at five concentrations should
be sufficient to demonstrate the linearity of the detector response without the necessity of
performing initial calibrations for each of the seven Aroclors. In addition, such a mixture can
8082A - 6
Revision 1
January 1998
-------
be used as a standard to demonstrate that a sample does not contain peaks that represent any
one of the Aroclors. This standard can also be used to determine the concentrations of either
Arodor 1016 or Arodor 1260, should they be present in a sample. Prepare a minimum of five
calibration standards containing equal concentrations of both Aroclor 1016 and Arodor 1260
by dilution of the stock standard with isooctane or hexane. The concentrations should
correspond to the expected range of concentrations found in real samples and should bracket
the linear range of the detector,
5.6.2 Single standards of each of the other five Aroclors are required to aid the
analyst in pattern recognition. Assuming that the Arodor 1016/1260 standards described in
Sec. 5.6.1 have been used to demonstrate the linearity of the detector, these single standards
of the remaining five Arodors are also used to determine the calibration factor for each Aroclor.
Prepare a standard for each of the other Aroclors. The concentrations should correspond to
the mid-point of the linear range of the detector.
5.7 Calibration standards for PCB congeners
5.7.1 If results are to be determined for individual PCB congeners, then standards
for the pure congeners must be prepared. The table in Sec. 1.1 lists 19 PCB congeners that
have been tested by this method along with the IUPAC numbers designating these congeners.
This procedure may be appropriate for other congeners as well.
5.7.2 Stock standards may be prepared in a fashion similar to that described for the
Arodor standards, or may be purchased as eommerdaSy-prepared solutions. Stock standards
should be used to prepare a minimum of five concentrations by dilution of the stock standard
with isooctane or hexane. The concentrations should correspond to the expected range of
concentrations found in real samples and should bracket the linear range of the detector.
5.8 Internal standard
5.8.1 When POT congeners are to be determined, the use of an internal standard
is highly recommended. Pecachlorobiphenyl may be used as an internal standard, added to
each sample extract prior to analysis, and induded in each of the initial calibration standards.
5.8.2 When PCBs are to be determined as Aroclors, an internal standard is not
used, and decachlorobiphenyl is employed as a surrogate (see Sec. 5.8).
5.9 Surrogate standards
5.9.1 When PCBs are to be determined as Aroclors, decachlorobiphenyl is used
as a surrogate, and is added to each sample prior to extraction. Prepare a solution of
decachlorobiphenyl at a concentration of 5 mg/L in acetone.
5.9.2 When PCB congeners are to be determined, decachlorobiphenyl is
recommended for use as an internal standard, and therefore, cannot also be used as a
surrogate. Therefore, tetrachloro-meta-xylene may be used as a surrogate for PCB congener
analysis. Prepare a solution of tetrachloro-meta-xylene at a concentration of 5 mg/L in
acetone.
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6.0 SAMPLE COLLECTION, PRESERVATION, AND HANDLING
6.1 See Chapter Four, Organic Analytes for sample collection and preservation instructions.
6.2 Extracts should be stored under refrigeration in the dark and analyzed within 40 days
of extraction.
NOTE; The holding times listed in Sec. 6.2 are recommendations. PCBs are very stable in a
variety of matrices, and holding times under the conditions listed in Sec. 6.2 may be as
high as a year for some matrices.
7.0 PROCEDURE
7.1 Sample extraction
7.1.1 Refer to Chapter Two and Method 3500 for guidance in choosing the
appropriate extraction procedure. In general, water samples are extracted at a neutral pH with
methylene chloride using a separatory funnel (Method 3510) or a continuous liquid-liquid
extractor (Method 3520) or other appropriate procedure. Solid samples are extracted with
hexane-acetone (1:1) or methylene chloride-acetone (1:1) using one of the Soxhlet extraction
(Method 3540 or 3541) procedures, ultrasonic extraction (Method 3550), supercritical fluid
extraction (3562), or other appropriate procedure. Oils and other organic liquids may be
amenable to the waste dilution procedure in Method 3580.
NOTE: Use of hexane-acetone generally reduces the amount of interferences that are
extracted and improves signal-to-noise.
7.1.2 Reference materials, field-contaminated samples, or spiked samples should
be used to verify the applicability of the selected extraction technique to each new sample type.
Such samples should contain or be spiked with the compounds of interest in order to
determine the percent recovery and the limit of detection for that sample type (see Chapter
One). When other materials are not available and spiked samples are used, they should be
spiked with the analytes of interest, either specific Aroclors or PCB congeners. When the
presence of specific Aroclors is not anticipated, the Aroclor 1016/1260 mixture may be an
appropriate choice for spiking. See Methods 3500 and 8000 for guidance on demonstration
of initial method proficiency as well as guidance on matrix spikes for routine sample analysis.
7.2 Extract cleanup
Refer to Methods 3660 and 3665 for information on extract cleanup.
7.3 GC conditions
This method allows the analyst to choose between a single-column or a dual-column
configuration in the injector port. Either wide- or narrow-bore columns may be used. See Sec.
7.7 for information on techniques for making positive identifications of multi-component
analytes.
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7.3.1 Single-column analysis
This capillary GC/ECD method allows the analyst the option of using 0.25-0.32 mm ID
capillary columns (narrow-bore) or 0.53 mm ID capillary columns (wide-bore). The use of
narrow-bore (0.25-0.32 mm ID) columns is recommended when the analyst requires greater
chromatographic resolution. Use of narrow-bore columns is suitable for relatively dean
samples or for extracts that have been prepared with one or more of the clean-up options
referenced in the method. Wide-bore columns (0.53 mm ID) are suitable for more complex
environmental and waste matrices.
7.3.2 Dual-column analysis
The dual-column/dual-detector approach involves the use of two 30 m x 0.53 mm ID
fused-silica open-tubular columns of different polarities, thus different selectivities towards the
target compounds. The columns are connected to an injection tee and ECD detectors.
7.3.3 GC temperature programs and flow rates
7.3.3.1 Table 2 lists GC operating conditions for the analysis of PCBs as
Aroclors for single-column analysis, using either narrcrw^ore or wide-bore capillary
columns. Table 3 lists GC operating conditions for the dual-column analysis. Use the
conditions in these tables as guidance and establish the GC temperature program and
flow rate necessary to separate the analytes of interest.
7.3.3.2 When determining PCBs as congeners, difficulties may be
encountered with coelution of congener 153 and other sample components. When
determining PCBs as Aroclors, chromatographic conditions should be adjusted to give
adequate separation of the characteristic peaks in each Aroclor (see Sec. 7.4.6).
7.3.3.3 Tables 4 and 5 summarize the retention times of up to 73 Aroclor
peaks determined during dual-column analysis using the operating conditions listed in
Table 2. These retention times are provided as guidance as to what may be achieved
using the GC columns, temperature programs, and flow rates described in this method.
Note that the peak numbers used in these tables are not the IUPAC congener
numbers, but represent the elution order of the peaks on these GC columns.
7.3.3.4 Once established, the same operating conditions must be used
for the analysis of samples and standards.
7.4 Calibration
7.4.1 Prepare calibration standards as described in Sec. 5.0. Refer to Method 8000
(Sec. 7.0) for proper calibration techniques for both initial calibration and calibration verification.
When PCBs are to be determined as congeners, the use of internal standard calibration is
highly recommended. Therefore, the calibration standards must contain the internal standard
(see Sec. 5.7) at the same concentration as the sample extracts. When PCBs are to be
determined as Aroclors, external standard calibration is generally used.
NOTE: Because of the sensitivity of the electron capture detector, the injection port and
column should always be cleaned prior to performing the initial calibration.
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7.4.2 When PCBs are to be quantitatively determined as congeners, an initial multi-
point calibration must be performed that includes standards for ail the target analytes
(congeners).
7.4.3 When PCBs are to be quantitatively determined as Aroclors, the initial
calibration consists of two parts, described below.
7.4.3.1 As noted in Sec. 5.6.1, a standard containing a mixture of ArocJor
1016 and Aroclor 1260 will include many of the peaks represented in the other five
Aroclor mixtures. Thus, such a standard may be used to demonstrate the linearity of
the detector and that a sample does not contain peaks that represent any one of the
Aroclors. This standard can also be used to determine the concentrations of either
Aroclor 1016 or Aroclor 1260, should they be present in a sample. Therefore, an initial
multi-point calibration is performed using the mixture of Aroclors 1016 and 1260
described in Sec. 5.6.1. See Sec. 7.0 of Method 8000 for guidance on the use of linear
and non-linear calibrations.
7.4.3.2 Standards of the other five Aroclors are necessary for pattern
recognition. These standards are also used to determine a single-point calibration
factor for each Aroclor, assuming that the Aroclor 1016/1260 mixture in Sec. 7.4.3.1
has been used to describe the detector response. The standards for these five
Aroclors should be analyzed before the analysis of any samples, and may be analyzed
before or after the analysis of the five 1016/1260 standards in Sec. 7.4.3.1.
7.4.3.3 In situations where only a few Aroclors are of interest for a specific
project, the analyst may employ a multi-point initial calibration of each of the Aroclors
of interest (e.g., five standards of Aroclor 1232 if this Aroclor is of concern and linear
calibration is employed) and not use the 1016/1260 mixture described in Sec. 7.4.3.1
or the pattern recognition standards described in 7.4.3.2.
7.4.4 Establish the GC operating conditions appropriate for the configuration
(single-column or dual column, Sec. 7.3). Optimize the instrumental conditions for resolution
of the target compounds and sensitivity. A final temperature of 240-270°C may be required
to elute decachlorobiphenyl. Use of injector pressure programming will improve the
chromatography of late eluting peaks.
NOTE: Once established, the same operating conditions must be used for both calibrations
and sample analyses.
7.4.5 A 2-|jL injection of each calibration standard is recommended. Other injection
volumes may be employed, provided that the analyst can demonstrate adequate sensitivity for
the compounds of interest.
7.4.6 Record the peak area (or height) for each congener or each characteristic
Aroclor peak to be used for quantitation.
7.4.6.1 A minimum of 3 peaks must be chosen for each Aroclor, and
preferably 5 peaks. The peaks must be characteristic of the Aroclor in question.
Choose peaks in the Aroclor standards that are at least 25% of the height of the largest
Aroclor peak. For each Aroclor, the set of 3 to 5 peaks should include at ieast one
peak that is unique to that Aroclor. Use at ieast five peaks for the Aroclor 1016/1260
mixture, none of which should be found in both of these Aroclors.
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7.4.6.2 Late-eluting Aroclor peaks are generally the most stable in the
environment Table 6 lists diagnostic peaks in each Aroclor, along with their retention
times on two GC columns suitable for single-column analysis. Table 7 lists 13 specific
PCB congeners found in Aroclor mixtures. Table 8 lists PCB congeners with
corresponding retention times on a DB-5 wide-bore GC column. Use these tables as
guidance in choosing the appropriate peaks.
7.4.7 When determining PCB congeners by the internal standard procedure,
calculate the response factor (RF) for each congener in the calibration standards relative to
the internal standard, decachlorobiphenyl, using the equation that follows.
RF - A° "
K * c.
where:
As = Peak area (or height) of the analyte or surrogate.
Afe = Peak area (or height) of the internal standard.
Cs = Concentration of the analyte or surrogate, in yigll.
Cis = Concentration of the internal standard, in jig/L.
7.4.8 When determining PCBs as Aroclors by the external standard technique,
calculate the calibration factor (CF) for each characteristic Aroclor peak in each of the initial
calibration standards (from either Sec. 7.4.3.1 or 7.4.3.2) using the equation below.
QP Peak Area (or Height) in the Standard
Total Mass of the Standard Injected (in nanograms)
Using the equation above, a calibration factor will be determined for each characteristic peak.
using the total mass of the Aroclor injected. These individual calibration factors are used to
quantitate sample results by applying the factor for each individual peak to the area of that
peak, as described in Sec. 7.9.
For a five-point calibration, five sets of calibration factors will be generated for the Aroclor
1016/1260 mixture, each set consisting of the calibration factors for each of the five (or more)
peaks chosen for this mixture, e.g., there will be at least 25 separate calibration factors for the
mixture. The single standard for each of the other Aroclors (see Sec. 7.4.3.1) will generate at
least three calibration factors, one for each selected peak. If a non-linear calibration model is
employed, as described in Method 8000, then additional standards containing the Aroclor
1016/1260 mixture will be employed, with a corresponding increase in the total number of
calibration factors (e.g., at least 30 for a 6-point curve and 35 for a 7-point curve).
7.4.9 The response factors or calibration factors from the initial calibration are used to
evaluate the linearity of the initial calibration, if a linear calibration model is used. This involves
the calculation of the mean response or calibration factor, the standard deviation, and the
relative standard deviation (RSD) for each congener or Aroclor peak. See Method 8000 for the
specifics of, the evaluation of the linearity of the calibration and guidance on performing non-
linear calibrations. In general, non-linear calibrations also will consider each characteristic
Aroclor peak separately. Thus, for the 1016/1260 mixture, non-linear calibration will result in
at least five calibration models, one for each selected peak. Each model is then applied to
the determination of the concentration of that specific peak in the sample chromatogram, as
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describe din Sec. 7.9. When the Aroclor 1016/1260 mixture is used to demonstrate the
detector response, the calibration models (see Method 8000) chosen for this mixture must be
applied to the other five Aroclors for which only single standards are analyzed. If multi-point
calibration is performed for individual Aroclors (see Sec. 7.4.3.3), use the calibration factors
from those standards to evaluate linearity.
7.5 Retention time windows
Retention time windows are crucial to the identification of target compounds. Absolute
retention times are used for the identification of PCBs as Aroclors. When PCBs are determined as
congeners by an internal standard technique, absolute retention times may be used in conjunction
with relative retention times (relative to the internal standard). Retention time windows are
established to compensate for minor shifts in absolute retention times as a result of sample loadings
and normal chromatographic variability. The width of the retention time window should be carefully
established to minimize the occurrence of both false positive and false negative results. Tight
retention time windows may result in false negatives and/or may cause unnecessary reanalysis of
samples when surrogates or spiked compounds are erroneously not identified. Overly wide retention
time windows may result in false positive results that cannot be confirmed upon further analysis.
Analysts should consult Method 8000 for the details of establishing retention time windows.
7.6 Gas chromatographic analysis of sample extracts
7.6.1 The same GC operating conditions used for the initial calibration must be
employed for samples analyses.
7.6.2 Verily calibration each 12-hour shift by injecting calibration verification standards
prior to conducting any sample analyses. A calibration standard must also be injected at
intervals of not less than once every twenty samples (after every 10 samples is recommended
to minimize the number of samples requiring re-injection when QC limits are exceeded) and
at the end of the analysis sequence. For Aroclor analyses, the calibration verification standard
should be a mixture of Aroclor 1016 and Aroclor 1260. The calibration verification process
does not require analysis of the other Aroclor standards used for pattern recognition, but the
analyst may wish to include a standard for one of these Aroclors after the 1016/1260 mixture
used for calibration verification throughout the analytical sequence.
7.6.2.1 The calibration factor for each anaiyte calculated from the calibration
verification standard (CFV) must not exceed a difference of more than ± 15 percent when
compared to the mean calibration factor from the initial calibration curve.
CF - CF„
% Difference = ———- * 100 ~ .
CF
7.6.2.2 When internal standard calibration is used for PCB congeners, the
response factor calculated from the calibration verification standard (RFV) must not
exceed a ± 15 percent difference when compared to the mean response factor from the
initial calibration.
pp _ pp
% Difference = * 100
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7.6.2.3 If this criterion is exceeded for any calibration factor or response factor,
inspect the gas chromatographic system to determine the cause and perform whatever
maintenance is necessary before verifying calibration and proceeding with sample
analysis.
7.6.2.4 If routine maintenance does not return the instrument performance to
meet the QC requirements (Sec. B.2) based on the last initial calibration, then a new
initial calibration must be performed.
7.6.3 inject a 2-jjL aliquot of the concentrated sample extract. Record the volume
injected to the nearest 0.05 >jL and the areas (or heights) of the resulting peaks.
7.6.4 Qualitative identifications of target analytes are made by examination of the
sample chromatograms, as described in Sec. 7.7.
7.6.5 Quantitative results are determined for each identified analyte (Aroclors or
congeners), using the procedures described in Sees. 7.8 and 7.9 for either the internal or the
external calibration procedure (Method 8000). If the responses in the sample chromatogram
exceed the calibration range of the system, dilute the extract and reanalyze. Peak height
measurements are recommended over peak area when overlapping peaks cause errors in
area integration.
7.6.6 Each sample analysis must be bracketed with an acceptable initial calibration,
calibration verification standard(s) (each 12-hour shift), or calibration standards interspersed
within the samples. When a calibration verification standard fails to meet the QC criteria, all
samples that were injected after the last standard that last met the QC criteria must be re-
injected.
Multi-level standards (mixtures or multi-component analytes) are highly recommended
to ensure that detector response remains stable for all analytes over the calibration range.
7.6.7 Sample injections may continue for as long as the calibration verification
standards and standards interspersed with the samples meet instrument QC requirements.
It is recommended that standards be analyzed after every 10 samples (required after every 20
samples and at the end of a set) to minimize the number of samples that must be re-injected
when the standards fail the QC limits. The sequence ends when the set of samples has been
injected or when qualitative or quantitative QC criteria are exceeded.
7.6.8 If the peak response is less than 2.5 times the baseline noise level, the validity
of the quantitative result may be questionable. The analyst should consult with the source of
the sample to determine whether further concentration of the sample is warranted.
7.6.9 Use the calibration standards analyzed during the sequence to evaluate retention
time stability. If any of the standards fall outside their daily retention time windows, the system
is out of control. Determine the cause of the problem and correct it.
7.6.10 If compound identification or quantitation is precluded due to interferences (e.g.,
broad, rounded peaks or ill-defined baselines are present), corrective action is warranted.
Cleanup of the extract or replacement of the capillary column or detector may be necessary.
The analyst may begin by rerunning the sample on another instrument to determine if the
problem results from analytical hardware or the sample matrix. Refer to Method 3600 for the
procedures to be followed in sample cleanup.
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7.7 Qualitative identification
The identification of PCBs as either Aroclors or congeners using this method with an
electron capture detector Is based on agreement between the retention times of peaks in the
sample chromatogram with the retention time windows established through the analysis of
standards of the target analytes. See Method 8000 for information on the establishment of
retention time windows.
Tentative identification of an analyte occurs when a peak from a sample extract falls
within the established retention time window for a specific target analyte. Each tentative
identification must be confirmed: using a second GC column of dissimilar stationary phase (as
in the dual-column analysis), based on a clearly identifiable Aroclor pattern, or using another
technique such as GC/MS (see Sec. 7.10).
7.7.1 When simultaneous analyses are performed from a single injection (the dual-
column GC configuration described in Sec. 7.3), It is not practical to designate one column as
the analytical (primary) column and the other as the confirmation column. Since the calibration
standards are analysed on both columns, the results for both columns must meet the
calibration acceptance criteria. If the retention times of the peaks on both columns fall within
the retention time windows on the respective columns, then the target analyte identification has
been confirmed.
7.7.2 The results of a single column/single injection analysis may be confirmed on a
second, dissimilar, GC column. In order to be used for confirmation, retention time windows
must have been established for the second GC column. In addition, the analyst must
demonstrate the sensitivity of the second column analysis. This demonstration must include
the analysis of a standard of the target analyte at a concentration at least as low as the
concentration estimated from the primary analysis. That standard may be either the individual
congeners, individual Aroclor or the Aroclor 1016/1260 mixture.
7.7.3 When samples are analyzed from a source known to contain specific Aroclors,
the results from a single-column analysis may be confirmed on the basis of a clearly
recognizable Aroclor pattern. This approach should not be attempted for samples from
unknown or unfamiliar sources or for samples that appear to contain mixtures of Aroclors. In
order to employ this approach, the analyst must document:
• The peaks that were evaluated when comparing the sample chromatogram and the
Aroclor standard.
• The absence of major peaks representing any other Aroclor.
• The source-specific information indicating that Aroclors are anticipated in the sample
(e.g., historical data, generator knowledge, etc.).
This information should either be provided to the data user or maintained by the laboratory.
7.7.4 See Sec. 7.10 for information on GC/MS confirmation.
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7.8 Quantitation of PCBs as congeners
7.8.1 The quantitation of PCB congeners is accomplished by the comparison of the
sample chromatogram to those of the PCB congener standards, using the internal standard
technique (see Method 8000). Calculate the concentration of each congener.
7.8.2 Depending on project requirements, the PCB congener results may be
reported as congeners, or may be summed and reported as total PCBs. The analyst should
use caution when using the congener method for quantitation when regulatory requirements
are based on Aroclor concentrations. See Sec. 7.9.3,
7.9 Quantitation of PCBs as Aroclors
The quantitation of PCB residues as Aroclors is accomplished by comparison of the sample
chromatogram to that of the most similar Aroclor standard. A choice must be made as to which
Aroclor is most similar to that of the residue and whether that standard is truly representative of the
PCBs in the sample.
7.9.1 Use the individual Aroclor standards (not the 1016/1260 mixtures) to
determine the pattern of peaks on Aroclors 1221,1232,1242,1248, and 1254. The patterns
for Aroclors 1016 and 1260 will be evident in the mixed calibration standards.
7.9.2 Once the Aroclor pattern has been identified, compare the responses of 3 to
5 major peaks in the single-point calibration standard for that Aroclor with the peaks observed
in the sample extract. The amount of Arocior is calculated using the individual calibration
factor for each of the 3 to 5 characteristic peaks chosen in Sec. 7.4.6.1. and the calibration
model (linear or non-linear) established from the multi-point calibration of the 1016/1260
mixture. Non-linear calibration may result in different models for each selected peak. A
concentration is determined using each of the characteristic peaks, using the individual
calibration factor calculated for that peak in Sec. 7.4.8, and then those 3 to 5 concentrations
are averaged to determine the concentration of that Aroclor.
7.9.3 Weathering of PCBs in the environment and changes resulting from waste
treatment processes may after the PCBs to the point that the pattern of a specific Arocior is
no longer recognizable. Samples containing more than one Aroclor present similar problems.
If the purpose of the analysis is not regulatory compliance monitoring on the basis of Aroclor
concentrations, then it may be more appropriate to perform the analyses using the PCB
congener approach described in this method. If results in terms of Aroclors are required, then
the quantitation as Aroclors may be performed by measuring the total area of the PCB pattern
and quantitating on the basis of the Aroclor standard that is most similar to the sample. Any
peaks that are not identifiable as PCBs on the basis of retention times should be subtracted
from the total area. When quantitation is performed in this manner, the problems should be
fully described for the data user and the specific procedures employed by the analyst should
be thoroughly documented.
7.10 GC/MS confirmation may be used in conjunction with either single-or dual-column
analysis if the concentration is sufficient for detection by GC/MS.
7.10,1 Full-scan quadrupole GC/MS will normally require a higher concentration of
the analyte of interest than full-scan ion trap or selected ion monitoring techniques. The
concentrations will be instrument-dependent, but values for full-scan quadrupole GC/MS may
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be as high as 10 ng/pL in the final extract, while ion trap or SIM may only require a
concentration of 1 ng/ML.
7.10.2 The GC/MS must be calibrated for the target analytes when it is used for
Quantitative analysis. If GC/MS is used only for confirmation of the identification of the target
analytes, then the analyst must demonstrate that those pesticides identified by GC/ECD can
be confirmed by GC/MS. This demonstration may be accomplished by analyzing a single-point
standard containing the analytes of interest at or below the concentrations reported in the
GC/EC analysis. When using SIM techniques, the ions and retention times should be
characteristic of the Aroclors to be confirmed.
7.10.3 GC/MS confirmation should be accomplished by analyzing the same extract
used for GC/ECD analysis and the extract of the associated blank.
7.11 Chromatographic system maintenance as corrective action
When system performance does not meet the established QC requirements, corrective action
is required, and may include one or more of the following.
7.11.1 Splitter connections
For dual columns which are connected using a press-fit Y-shaped glass splitter or a Y-
shaped fused-silica connector, dean and deactivate the splitter port insert or replace with a
cleaned and deactivated splitter. Break off the first few inches (up to one foot) of the injection
port side of the column. Remove the columns and solvent backflush according to the
manufacturer's instructions. If these procedures fail to eliminate the degradation problem, it
may be necessary to deactivate the metal injector body and/or replace the columns.
7.11.2 Metal injector body
Turn off the oven and remove the analytical columns when the oven has cooled.
Remove the glass injection port insert (instruments with on-column injection). Lower the
injection port temperature to room temperature. Inspect the injection port and remove any
noticeable foreign material.
7.11.2.1 Place a beaker beneath the injector port inside the oven. Using
a wash bottle, rinse the entire inside of the injector port with acetone and then rinse it
with toluene, catching the rinsate in the beaker.
7.11.2.2 Consult the manufacturer's instructions regarding deactivating the
injector port body. Glass injection port liners may require deactivation with a silanizing
solution containing dimethyidichlorosilane.
7.11.3 Column rinsing
The column should be rinsed with several column volumes of an appropriate solvent.
Both polar and nonpolar solvents are recommended. Depending on the nature of the sample
residues expected, the first rinse might be water, followed by methanol and acetone.
Methylene chloride is a good final rinse and in some cases may be the only solvent required.
The column should then be filled with methylene chloride and allowed to stand flooded
overnight to allow materials within the stationary phase to migrate into the solvent. The column
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is then flushed with fresh methylene chloride, drained, and dried at room temperature with a
stream of ultrapure nitrogen.
8.0 QUALITY CONTROL
8.1 Refer to Chapter One and Method 8000 for specific quality control (GC) procedures.
Quality control procedures to ensure the proper operation of the various sample preparation
techniques can be found in Method 3500. If an extract cleanup procedure was performed, refer to
Method 3600 for the appropriate quality control procedures. Each laboratory should maintain a
formal quality assurance program. The laboratory should also maintain records to document the
qualify of the data generated.
8.2 Quality control procedures necessary to evaluate the GC system operation are found
in Method 8000, Sec. 7.0 and include evaluation of retention time windows, calibration verification
and chromatographic analysis of samples.
8.2.1 Include a calibration standard after each group of 20 samples (it is
recommended that a calibration standard be included after every 10 samples to minimize the
number of repeat injections) in the analysis sequence as a calibration check. The response
factors for the calibration should be within 15 percent of the initial calibration. When this
continuing calibration is out of this acceptance window, the laboratory should stop analyses
and take corrective action.
8.2.2 Whenever quantitation is accomplished using an internal standard, internal
standard responses must be evaluated for acceptance. The measured area of the internal
standard must be no more than 50 percent different from the average area calculated during
calibration. When the internal standard peak area is outside the limit, all samples that fall
outside the QC criteria must be reanalyzed.
8.3 Initial Demonstration of Proficiency - Each laboratory must demonstrate initial
proficiency with each sample preparation and determinative method combination it utilizes, by
generating data of acceptable accuracy and precision for target analytes in a clean matrix. The
laboratory must also repeat the following operations whenever new staff are trained or significant
changes in instrumentation are made. See Method 8000, Sec. 8.0, for information on how to
accomplish this demonstration.
8.3.1 The QC Reference Sample concentrate (Method 3500) should contain PCBs
as Aroclors at 10-50 mg/L for water samples, or PCBs as congeners at the same
concentrations. A 1-mL volume of this concentrate spiked into 1 L of organic-free reagent
water will result in a sample concentration of 10-50 gg/L. If Aroclors are not expected in
samples from a particular source, then prepare the QC reference samples with a mixture of
Aroclors 1016 and 1260. However, when specific Aroclors are known to be present or
expected in samples, the specific Aroclors should be used for the QC reference sample. See
Method 8000, Sec. 8.0, for additional information on how to accomplish this demonstration.
8.3.2 Calculate the average recovery and the standard deviation of the recoveries
of the analytes in each of the four QC reference samples. Refer to Sec. 8.0 of Method 8000
for procedures for evaluating method performance.
8.4 Sample Quality Control for Preparation and Analysis - The laboratory must also have
procedures for documenting the effect of the matrix on method performance (precision, accuracy,
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and detection limit). At a minimum, this includes the analysis of QC samples including a method
blank, a matrix spike, a duplicate, and a laboratory control sample (LCS) in each analytical batch and
the addition of surrogates to each field sample and QC sample.
8.4.1 Documenting the effect of the matrix should Include the analysis of at least
one matrix spike and one duplicate unspiked sample or one matrix spike/matrix spike duplicate
pair. The decision on whether to prepare and analyze duplicate samples or a matrix
spike/matrix spike duplicate must be based on a knowledge of the samples in the sample
batch. If samples are not expected to contain target analytes, laboratories should use a matrix
spike and matrix spike duplicate pair, spiked with the Aroclor 1016/1260 mixture. However,
when specific Aroclors are known to be present or expected in samples, the specific Aroclors
should be used for spiking. If samples are expected to contain target analytes, then
laboratories may use one matrix spike and a duplicate analysis of an unspiked field sample.
8.4.2 A Laboratory Control Sample (LCS) should be included with each analytical
batch. The LCS consists of an aliquot of a clean (control) matrix similar to the sample matrix
and of the same weight or volume. The LCS is spiked with the same analytes at the same
concentrations as the matrix spike. When the results of the matrix spike analysis indicate a
potential problem due to the sample matrix itself, the LCS results are used to verify that the
laboratory can perform the analysis in a clean matrix.
8.4.3 See Method 8000, Sec. 8.0, for the details on carrying out sample quality
control procedures for preparation and analysis.
8.5 Surrogate recoveries - The laboratory must evaluate surrogate recovery data from
individual samples versus the surrogate control limits developed by the laboratory. See Method
8000, Sec. 8.0, for information on evaluating surrogate data and developing and updating surrogate
limits.
8.6 It is recommended that the laboratory adopt additional quality assurance practices for
use with this method. The specific practices that are most productive depend upon the needs of the
laboratory and the nature of the samples. Whenever possible, the laboratory should analyze
standard reference materials and participate in relevant performance evaluation studies.
9.0 METHOD PERFORMANCE
9.1 The MDL is defined in Chapter One. The MDLs for Aroclors vary in the range of 0.054
to 0.90 jjg/L in water and 57 to 70 pg/kg in soils, with the higher MDLs for the more heavily
chlorinated Aroclors. Estimated quantitation limits may be determined using the factors in Table 1.
9.2 Estimated quantitation limits for PCSs as congeners vary by congener, in the range of
5-25 ng/L in water and 160 - 800 ng/kg in soils, with the higher values for the more heavily
chlorinated congeners.
9.3 The accuracy and precision obtainable with this method depend on the sample matrix,
sample preparation technique, optional cleanup techniques, and calibration procedures used. Table
9 provides single laboratory recovery data for Aroclors spiked into clay and soil and extracted with
automated Soxhlet. Table 10 provides multiple laboratory data on the precision and accuracy for
Aroclors spiked into soil and extracted by automated Soxhlet.
8082A -18
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January 1998
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9.4 During method performance studies, the concentrations determined as Arodors were
larger than those obtained using the congener method. In certain soils, interference prevented the
measurement of congener 66. Recoveries of congeners from soils spiked with Arodor 1254 and
Arodor 1260 were between 80% and 90%. Recoveries of congeners from environmental reference
materials ranged from 51 - 66% of the certified Arodor values.
9.5 Tables 11 through 13 contain laboratory performance data for several PCB congeners
using supercritical fluid extraction Method 3562 on an HP 7680. Seven replicate extractions were
performed on each sample. The method mis performed using a variable restrictor and solid trapping
material (Florisil). Sample analysis was performed by GC/ECD. The following soil samples were
used for this study:
9.5.1 Two field-contaminated certified reference materials were extracted by a
single laboratory. One of the materials was a lake sediment from Environment Canada (EC-5).
The other material was soil from a dump site and was provided by the National Science and
Engineering Research Council of Canada (EC-1). The average recoveries for EC-5 are based
on the certified value for that sample. The average recoveries for EC-1 are based on the
certified value of the samples or a Soxhlet value, if a certified value was unavailable for a
specific analyte.
9.5.2 Four certified reference materials were extracted by two independent
laboratories. The materials were: a marine sediment from NIST (SRM 1941), a fish tissue
from NIST (SRM 2974), a sewage sludge from BCR European Union (CRM 392), and a soil
sample from BCR European Union (CRM 481). The average recoveries are based on the
certified value of the samples or a Soxhlet value, if a certified value was unavailable for a
specific analyte.
9.5.3 A weathered sediment sample from Michigan (Saginaw Bay) was extracted
by a single laboratory. Soxhlet extractions were carried out on this sample and the SFE
recovery is relative to that for each congener. The average recoveries are based on the
certified value of the samples. Additional data are shown in the tables for some congeners for
which no certified values were available.
9.6 Tables 14 through 16 contain single laboratory recovery data for Arodor 1254 using
SPE Method 3535. Recovery data at 2,10, and 100 ug/L are presented. Results represent three
replicate solid-phase extractions of spiked wastewaters. Two different wastewaters from each
wastewater type were spiked. All of the extractions were performed using 90-mm C18 disks.
10.0 REFERENCES
1. Lopez-Avila, V., Baldin, E., Benedicto, J, Milanes, J., Beckert. W.F., Application of Open-
Tubular Columns to SW-846 GC Methods", final report to the U.S. Environmental Protection
Agency on Contract 68-03-3511, Mid-Pacific Environmental Laboratory, Mountain View, CA,
1990.
2. Development and Application of Test Procedures for Spedfic Organic Toxic Substances in
Wastewaters. Category 10 - Pestiddes and PCB Report for the U.S. Environmental Protection
Agency on Contract 68-03-2606.
3. Ahnoff, M„ Josefsson, B„ "Cleanup Procedures for PCB Analysis on River Water Extracts",
Bull. Environ. Contam. Toxicol., 1975, 13, 159.
8082A- 19
Revision 1
January 1998
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4. Marsden, P.J., "Performance Data for SW-846 Methods 8270, 8081, and 8141", U.S.
Environmental Protection Agency, EMSL-Las Vegas, EPA/600/4-90/015.
5. Marsden, P.J., "Analysis of PCBs", U.S. Environmental Protection Agency, EMSL-Las Vegas,
NV, EPA/600/8-90/004.
6. Erickson, M., Analytical Chemistry of PCBs. Butterworth Publishers, Ann Arbor Science Book,
(1986).
7. Stewart, J., "EPA Verification Experiment for Validation of the SOXTEC® PCB Extraction
Procedure", Oak Ridge National Laboratory, Oak Ridge, TN, 37831-6138, October 1988.
8. Lopez-Avila, V. (Beckert, W., Project Officer), "Development of a Soxtec Extraction Procedure
for Extracting Organic Compounds from Soils and Sediments", EPA 6Q0/X-91/140, U.S.
Environmental Protection Agency, Environmental Monitoring Systems Laboratory, Las Vegas,
NV, October 1991.
9. Stewart, J.H., Bayne, C.K., Holmes, R.L., Rogers, W.F., and Maskarinec, M.P., "Evaluation
of a Rapid Quantitative Organic Extraction System for Determining the Concentration of PCB
in Soils", Proceedings of the U.S. EPA Symposium on Waste Testing and Quality Assurance,
Oak Ridge National Laboratory, Oak Ridge, TN, 37831, July 11-15,1988.
10. Tsang, S.F., Marsden, P.J., and Lesnik, B,, "Quantitation of Polychlorinated Biphenyls Using
19 Specific Congeners", Proceedings of the 9th Annual Waste Testing and Quality Assurance
Symposium, Office of Solid Waste and Emergency Response, U.S. Environmental Protection
Agency, Washington, DC, July 1993.
11. S. Bewadt, B. Johansson, S. Wunderli, M. Zennegg, L. F. de Alencastro and D. Grandjean,
"Independent Comparison of Soxhlet and Supercritical Fluid Extraction for the Determination
of PCBs in an Industrial Soil," Ana!. Chem., 1995,67(14) 2424-2430.
12. Markell, C„ "3M Data Submission to EPA," letter to B. Lesnik, June 27,1995.
13. Richter, B, Ezzell, J., and Felix D, "Single Laboratory Method Validation Report - Extraction of
Organophosphorus Pesticides, Herbicides and Polychlorinated Biphenyls using Accelerated
Solvent Extraction (ASE) with Analytical Validation by GC/NPD and GC/ECD," Dionex, Salt
Lake City, UT, Document 101124, December 2,1994.
8082A - 20
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TABLE 1
FACTORS FOR DETERMINATION OF ESTIMATED QUANTITATION LIMITS8 (EQLs)
FOR VARIOUS MATRICES
Matrix Factor
Ground water 10
Low-concentration soil by sonication with GPC cleanup 670
High-concentration soil and sludges by sonication 10,000
Non-water miscible waste 100,000
aEQL ~ [MDL for water (see Sec. 1.8)] times [Factor in this table]
For nonaqueous samples, the factor is on a wet-weight basis. Sample EQLs are highly matrix-
dependent. EQLs determined using these factors are provided as guidance and may not always be
achievable.
8082A-21
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TABLE 2
GC OPERATING CONDITIONS FOR PGBs AS AROCLORS
SINGLE COLUMN ANALYSIS
Narrow-bone columns
Narrow-bore Column 1 - 30 m x 0.25 or 0.32 mm ID fused-siiica capillary column chemically bonded
with SE-54 (DB-5 or equivalent), 1 Mm Aim thickness.
Carrier gas (He) 16 psi
Injector temperature 225 °C
Detector temperature 3000C
Initial temperature 100°C, hold 2 minutes
Temperature program 100°C to 160°C at 15°C/min, followed
by 160°C to 270°C at 5cC/min
Final temperature 270°C
Narrow-bore Column 2 - 30 m x 0.25 mm ID fused-silica capillary column chemically bonded with 35
percent phenyl methylpofysfloxane (DB-608, SPB-608, or equivalent) 25 Mm coating thickness, 1 Mm
film thickness
Cam'er gas (N^ 20 psi
Injector temperature 225°C
Detector temperature 300°C
Initial temperature 160°C, hold 2 minutes
Temperature program 160°C to 290°C at 5°C/min
Final temperature 290°C, hold 1 min
Wide-bore columns
Wide-bore Column 1 - 30 m x 0.53 mm ID fused-silica capillary column chemically bonded with 35
percent phenyl methylpolysiloxane (DB-608, SPB-608, RTx-35, or equivalent), 0.5 or 0.83 Mm
film thickness.
Wide-bore Column 2 - 30 m x 0.53 mm ID fused-silica capillary column chemically bonded with 14%
cyanopropylmethylpolysiloxane (DB-1701, or equivalent), 1.0 fjm film thickness.
Cam'er gas (He) 5-7 mL/minute
Makeup gas (argon/methane
[P-5 or P-10] or N2) 30 mLimin
Injector temperature 2506C
Detector temperature 290°C
Initial temperature 1 SOX, hold 0.5 minute
Temperature program 150°C to 270°C at 5°C/min
Final temperature 270°C, hold 10 min
8082A - 22
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TABLE 2
(continued)
GC OPERATING CONDITIONS FOR PCBs AS AROCLORS
SINGLE COLUMN ANALYSIS
Wide-bore Columns (continued)
Wide-bore Column 3 - 30 m x 0.53 mm ID fused-silica capillary column chemically bonded with SE-
54 (DB-5, SPB-5, RTx-5, or equivalent), 1.5 vim film thickness.
Carrier gas (He)
Makeup gas (argon/methane
[P-5 or P-10] or N2)
Injector temperature
Detector temperature
Initial temperature
Temperature program
Final temperature
6 mL/minute
30 mL/min
205°C
290°C
140°C, hold 2 min
140 °C to 240°C at 10°C/min,
hold 5 minutes at 240°C,
240°C to 265°C at 5°C/min
265°C, hold 18 min
8082A - 23
Revision 1
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TABLE 3
GC OPERATING CONDITIONS FOR PCBs AS AROCLORS
FOR THE DUAL COLUMN METHOD OF ANALYSIS HIGH TEMPERATURE, THICK FILM
Column 1 - DB-1701 or equivalent, 30 mx 0.53 mm ID, 1.0 Mm film thickness.
Column 2 - DB-5 or equivalent, 30 m x 0.53 mm ID, 1.5 |jm film thickness.
Carrier gas (He) flow rate
Makeup gas (N2) flow rate
Temperature program
Injector temperature
Detector temperature
Injection volume
Solvent
Type of injector
Detector type
Range
Attenuation
Type of splitter
6 mL/min
20 mL/min
0.5 min hold
150°C to 190X, at 12°C/min, 2 mln hold
190°C to 275aC, at 4eC/min, 10 min hold
250°C
320 °C
2 mL
Hexane
Flash vaporization
Dual ECD
10
64 (DB-1701 )/64 (DB-5)
J&W Scientific press-fit Y-shaped inlet splitter
8082A - 24
Revision 1
January 1998
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TABLE 4
SUMMARY OF RETENTION TIMES OF AROCLORS
ON THE DB-5 COLUMN", DUAL-COLUMN ANALYSIS
Peak Aroclor Aroclor Aroclor Aroclor Aroclor doctor Aroclor
No." 1016 1221 1232 1242 1248 1254 1260
1

5.85
5.85

7.63
7.64
7.57

3
8.41
8.43
8.43
8.37

4
8.77
8.77
8.78
8.73

5
8.98
8.99
9.00
8.94
8.95

6
9.71

9.66

7
10.49
10.50
10.50
10.44
10.45

8
10.58
10.59
10.59
10.53

9
10.90

10.91
10.86
10.85

10
11.23
11.24
11.24
11.18
11 18

11
11.88

11.90
11.84
11.85

12
11.99

12.00
11.95

13
12.27
12.29
12.29
12.24
12.24

14
12.66
12.68
12.69
12.64
12.64

15
12.98
12.99
13.00
12.95
12.95

16
13,18

13.19
13.14
13.15

17
13.61

13.63
13.58
13.58
13.59
13.59
18
13.80

13.82
13.77
13.77
13.78

19
13.96

13.97
13.93
13.93
13.90

20
14.48

14.50
14.46
14.45
14.46

21
14.63

14.64
14.60
14.60

22
14.99

15.02
14.98
14.97
14.98

23
15.35

15.36
15.32
15.31
15.32

24
16.01

15.96

16.14
16.08
16.08
16.10

26
16.27

16.29
16.26
16.24
16.25
16.26
27

16.53

17.04

16.99
16.96
16.97
29

17.22
17.19
17.19
17.19
17.21
30

17.46
17.43
17.43
17.44

17.69
17.69

17.92
17.91
17.91

18.16
18.14
18.14

18.41
18.37
18.36
18.36
18.37
35

18.58
18.56
18.55
18.55

18.68
a GO operating conditions are given in Table 3. All retention times in minutes.
b The peaks listed in this table are sequentially numbered in elution order for illustrative purposes
only and are not isomer numbers.
8082A-25
Revision 1
January 1998
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TABLE 4
(continued)
Peak Aroclor Aroclor Aroclor Aroclor Aroclor Aroclor Aroclor
No." 1016 1221 1232 1242 1248 1254 1260
37
18.83
18.80
18.78
18.78
18.79
38
19.33
19.30
19.29
19.29
19.29
39

19.48
19.48
40

19.81
19.80
41
20.03
19.97
19.92
19.92

20.28
20.28
43

20.46
20.45

20.57
20.57
45

20.85
20.83
20.83
20.83
46
21.18
21.14
21.12
20.98

21.36
21.38
21.38
48

21.78
21.78
49

22.08
22.05
22.04
22.03
50

22.38
22.37
51

22.74
22.73
52

22.96
22.95
53

23.23
23.23
54

23.42
55

23.75
23.73
56

23.99
23.97
57

24.16
58

24.27

24.45
60

24.61
24.62
61

24.93
24.91
62

25.44
63

26.22
26.19
64

26.52
65

26.75
66

27.41
67

28.07
68

28.35
69

29.00
• GC operating conditions are given in Table 3. AH retention times in minutes.
b The peaks listed in this table are sequentially numbered in etution order for illustrative purposes
only and are not isomer numbers.
8082A - 26
Revision 1
January 1998
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TABLE 5
SUMMARY OF RETENTION TIMES OF AROCLORS
ON THE DB-1701 COLUMN", DUAL COLUMN ANALYSIS
Peak Aroclor Aroclor Aroclor Aroclor Aroclor Aroclor Aroclor
No.6 1016 1221 1232 1242 1248 1254 1260
1

4.45
4.45

5.38

5.78

5,86
5.86

5
6.33
6.34
6.34
6.28

6
6.78
6.78
6.79
6.72

7
6.96
6.96
6.96
6.90
6.91

8
7.64

7.59

9
8.23
8.23
8.23
8.15
8.16

10
8.62
8.63
8.63
8.57

11
8.88

8.89
8.83
8.83

12
9.05
9.06
9.06
8.99
8.99

13
9.46

9.47
9.40
9.41

14
9.77
9.79
9.78
9.71
9.71

15
10.27
10.29
10.29
10.21
10.21

16
10.64
10.65
10.66
10.59
10.59

10.96
10.95
10.95

18
11.01

11.02
11.02
11.03

19
11.09

11.10

20
11.98

11.99
11.94
11.93
11.93

21
12.39

12.39
12.33
12.33
12.33

12.77
12.71
12.69

23
12.92

12.94
12.93

24
12.99

13.00
13.09
13.09
13.10

25
13.14

13.16

13.24

27
13.49

13.49
13.44
13.44

28
13.58

13.61
13.54
13.54
13.51
13.52
29

13.67

13.68

14.08
14.03
14.03
14.03
14.02
31

14.30
14.26
14.24
14.24
14.25
32

14.39
14.36

14.49
14.46
14.46

14.56
14.56
35

15.10
15.10

15.38
15.33
15.32
15.32

3GC operating conditions are given in Table 3. All retention times in minutes.
The peaks listed in this table are sequentially numbered in elution order for illustrative purposes only
arid are not isomer numbers.
8082A - 27
Revision 1
January 1998
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TABLE 5
(continued)
Peak Aroclor ArocJor Aroclor Aroclor Aroclor ArocJor Aroclor
No.b 1016 1221 1232 1242 1248 1254 1260
37
15.65
15.62
15.62
15.61
16.61
38
15.78
15.74
15.74
15.74
. 15.79
39
16.13
16.10
16.10
16.08

16.19
41

16.34
16.34
42

16.44
16.45
43

16.55

44
16.77
16.73
16.74
16.77
16.77
45
17.13
17.09
17.07
17.07
17.08
46

17.29
17.31
47

17.46
17.44
17.43
17.43
48

17.69
17.69
17.68
17.68
49

18.19
18.17
18.18
50

18.48
18.49
18.42
18.40
51

18.59

18.86
18.86
53

19.13
19.13
19.10
19.09
54

19.42
19.43
55

19.55
19.59
56

20.20
20.21
57

20.34

20.43
59

20.57
20.55

20.62
20.66
61

20.88
20.87
62

21.03
63

21.53
21.53
64

21.83
21.81
65

23.31
23.27
66

23.85
67

24.11
68

24.46
69

24.59
70

24.87
71

25.85
72

27.05
73

27.72
a GC operating conditions are given in Table 3. All retention times in minutes.
b The peaks listed in this table are sequentially numbered in elution order for illustrative purposes
only and are not isomer numbers.
8082A-28
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January 1998
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TABLE 6
PEAKS DIAGNOSTIC OF PCBs OBSERVED ON 0.53 mm ID COLUMN
DURING SINGLE COLUMN ANALYSIS
Peak
RT on
RT on

No,"
DB-eoe*
DB-1701"
Aroclor®
I
4.90
4.66
1221
II
7.15
6.96
1221,1232,1248
III
7.89
7.65
1061.1221.1232, 1242
IV
9.38
9.00
1016,1232,1242, 1248
V
10.69
10.54
1016. 1232. 1242
VI
14.24
14.12
1248, 1254
VII
14.81
14.77
1254
Vill
16.71
16.38
1254
IX
19.27
18.95
1254,1260
X
21.22
21.23
1260
XI
22.89
22.46
1260
B Peaks are sequentially numbered in elution order and are not isomer numbers
b Temperature program: T, = 150°C, hold 30 seconds; 5°C/minute to 275°C.
c Underline indicates largest peak in the pattern for that Aroclor
8082A-29 , Revision 1
January 1998
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TABLE 7
SPECIFIC PCB CONGENERS IN AROCLORS
Congener
IUPAC number
1016 1221
Aroclor
1232 1242 1248 1254 1260
Biphenyl
_

2-CB
1
X
X
X
X

23-DCB
5
X
X
X
X
X

34-DCB
12
X

X
X
X

244-TCB
28*
X

X
X
X
X

22'35-TCB
44

X
X
X
X
X
23'44-TCB
66*

X
X
X
233'4'6-PCB
110

23'44'5-PCB
118*

X
X
22'44'55'-HCB
153

X
22'344'5'-HCB
138

X
22'344'55'-HpCB
180

X
22'33'44'5-HpCB
170

X
"Apparent co-elution of:
28 with 31 (2,4',5-trichlorobiphenyl)
66 with 95 (2,2l,3,5',6-pentachlorobipheny1)
118 with 149 (2,2',3,4',5',6-hexachlorobiphenyl)
8082A - 30
Revision 1
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TABLE 8
RETENTION TIMES OF PCS CONGENERS
ON THE DB-5 WIDE-BORE COLUMN
IUPAC # Retention Time (min)
1
6.52
5
10.07
18
11.62
31
13.43
52
14.75
44
15.51
66
17.20
101
18.08
87
19.11
110
19.45
151
19.87
153
21.30
138
21.79
141
22.34
187
22.89
183
23.09
180
24.87
170
25.93
206
30.70
209
32.63
(internal standard)
8082A- 31
Revision 1
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TABLE 9
SINGLE LABORATORY RECOVERY DATA FOR EXTRACTION OF
PCBs FROM CLAY AND SOIL BY METHOD 3541a (AUTOMATED SOXHLET)
Spike Level Percent
Matrix Aroclor (ppm) Trial Recovery*
Clay 1254 5 1 87.0
2 92.7
3 93.8
4 98.6
5 79.4
6 28.3
Clay 1254 50 1 65.3
2 72.6
3 97.2
4 79.6
5 49.8
6 59.1
Clay 1260 5 1 87.3
2 74.6
3 60.8
4 93.8
5 96.9
6 113.1
Clay 1260 50 1 73.5
2 70.1
3 92.4
4 88.9
5 90.2
6 67.3
8082A - 32
Revision 1
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TABLE 9
(continued)
Spike Level Percent
Matrix Aroclor (ppm) Trial Recovery1
Soil 1254 5 1 69.7
2 89.1
3 91.8
4 83.2
5 62.5
Soil 1254 50 1 84.0
2 77.5
3 91.8
4 66.5
5 82.3
6 61.6
Soil 1260 5 1 83.9
2 82.8
3 81.6
4 96.2
5 93.7
6 93.8
7 97.5
Soil 1260 50 1 76.9
2 69.4
3 92.6
4 81.6
5 83.1
6 76.0
a The operating conditions for the automated Soxhlet
Immersion time: 60 min
Reflux time: 60 min
b Multiple results from two different extractors
Data from Reference 9
8082A - 33
Revision 1
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TABLE 10
MULTIPLE LABORATORY PRECISION AND ACCURACY DATA FOR THE EXTRACTION
OF PCBs FROM SPIKED SOIL BY METHOD 3541 (AUTOMATED SOXHLET)
Percent Recovery at Percent Recovery at
Aroclor 1254 Spike Aroclor 1260 Spike Mean
Concentration (pg/kg) Concentration (pg/kg) Recovery

5
50
500
5
50
500
All Levels

n
3
3

3
3

12
Lab 1
Mean
101.2
74.0

83.9
78.5

84.4

S. D.
34.9
41.8

7.4
7.4

26.0

6
6

6
6
24
Lab 2
Mean

56.5
66.9

70.1
74.5
67.0

S. D.

7.0
15.4

14.5
10.3
13.3

n
3
3

3
3

12
Lab 3
Mean
72.8
63.3

70.6
57.2

66.0

S. D.
10.8
8.3

2.5
5.6

9.1

n
6
6

6
6

24
Lab 4
Mean
112.6
144.3

100.3
84.8

110.5

S. D.
18.2
30.4

13.3
3.8

28.5

3
3

3
3
12
Lab 5
Mean

97.1
80.1

79.5
77.0
83.5

S. D,

8.7
5.1

3.1
9.4
10.3

n
2
3

3
4

12
Lab 6
Mean
140.9
127.7

138.7
105.9

125.4

S.D.
4.3
15.5

15.5
7.9

18.4

n
3
3

3
3

12
Lab 7
Mean
100.1
123.4

82.1
94.1

99.9

S. D.
17.9
14.6

7.9
5.2

19.0

n
3
3

3
3

12
I ah 8
Mean
65.0
38.3

92.8
51.9

62.0

S. D.
16.0
21.9

36.5
12.8 -

29.1

n
20
30
9
21
31
9
120
All
Mean
98.8
92.5
71.3
95.5
78.6
75.3
87.6
Labs
S.D.
28.7
42.9
14.1
25.3
18.0
9.5
29.7
Data from Reference 7
8082A - 34
Revision 1
January 1998
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TABLE 11
PERCENT RECOVERY (BIAS) OF PCBs IN VARIOUS SOILS USING SFE METHOD 3562
PCB No.8
EC-1C Dump
Site Soil
Low#1
SRM 1941
Marine
Sediment
Low #2
EC-5C Lake
Sediment
Low #3
CRM 481"
European
Soil
High #1
Mich Bay
Saginaw
Sediment
High #2
CRM 392
Sewage
Sludge
High #3
SRM 2974
Fish Tissue
Mussel
Low #4
Congener Mean
28
148.4
63.3
147.7
67.3
114.7
89.2
101.7
104.6
52
88.5
106.6
115.8
84.5
111.1
96.2
131.4
104.9
101
93.3
91.2
100.2
84.5
111.5
93.9
133.2
101.1
149
92.6
105.1
101.5
73.2
111.2

69.4
92.2
118
89.9
66.1
108.9
82.1
110.8
73.5
82.7
87.7
153
90.8
65.1
95.1
82.8
118.6
97.3
107.5
94.0
105°
89.1
72.6
96.6
83.4
111.8

79.4
88.8
138
90.1
57.4
97.9
76.9
126.9

73.1
87.1
128
90.8
69.9
101.2
65.9
87.6

62.5
79.7
O
CO
m
90.6
88.9
94.3
85.2
101.1

59.3
86.6
180
92.4
142.4
93.3
82.2
109.2
100.5
65.7
98.0
170
91.3
101.1
95.2
80.5

33.0
81.8
Matrix
Mean
95.7
85.8 '
104.0
79.0
108.7
91.8 .
83.2
92.2
a Congeners which are either certified or have had Soxhlet confirmation
b Parts per million (|jg/g)
0 Congener 105 was not resolved from congener 132 and congener 156 was not resolved from congener 171 by the GC method used for samples
EC-1 and EC-5
8082A-35
Revision 1
January 1998
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TABLE 12
% RELATIVE STANDARD DEVIATION (PRECISION) OF PCBs USING SFE METHOD 3562
EC-1° Dump
Site Soil
PCBNo.a Low#1
SRM 1941
Marine
Sediment
Low #2
EC-5° Lake
Sediment
Low #3
CRM 481b
European
Soil
High #1
Mich Bay
Saginaw
Sediment
High #2
CRM 392
Sewage
Sludge
High #3
SRM 2974
Fish Tissue
Mussel
Low #4
Congener Mean
28 11.5
1.5
3.8
5.6
2.4
1.9
2.7
4.2
52 9.1
3.3
3.9
5.4
2.2
2.9
3.1
4.3
101 9.1
2.9
2.8
4.9
1.4
5.2
2.9
4.2
149 7.1
0.7
3.8
3.9
3.4

2.2
3.0
118 9.8
1.9
4.5
5.4
2.0
3.3
2.4
4.2
153 8.4
1.5
3.0
4.3
4.3
9.5
3.0
4.9
105c 6.6
3.7
2.7
4.3
2.7

2.5
3.2
138 9.2
1.8
3.1
4.7
2.3

2.9
3.4
128 6.0
5.3
3.3
4.9
2.8

3.3
3.7
156c 8.3
0.0
5.1
4.5
1.9

3.8
3.4
180 8.0
1.3
3.6
4.3
3.1
9.6
2.7
4.7
170 5.7
2.3
3.6
3.9
2.3

4.0
3.1
Matrix 8.2
Mean
2.2
I
3.6
4.7
2.6
2.7
3.0
3.8
a Congeners which are either certified or have had Soxhlet confirmation.
b Parts per million (mg/kg)
0 Congener 105 was not resolved from congener 132 and congener 156 was not resolved from congener 171 by the GC method used for samples
EC-1 and EC-5.
8082A - 36
Revision 1
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TABLE 13
METHOD DETECTION LIMITS (MDLs)a OF PCBs USING SFE METHOD 3562
PCB No. a
EC-1° Dump
Site Soil
Low#1
SRM 1941
Marine
Sediment
Low #2
EC-5C Lake
Sediment
Low #3
CRM 481b
European
Soil
High #1
Mich Bay
Saginaw
Sediment
High #2
CRM 392
Sewage
Sludge
High #3
SRM 2974
Fish Tissue
Mussel
Low #4
Congener Mean
28
13.2
0.5
0.6
n/a
15.4
5.3
5.0
6.6
52
22.3
0.6
1.9
n/a
29.9
6.9
9.1
11.8
101
23.9
0.9
1.9
n/a
3.3
20.6
9.7
10.1
149
7.1
0.7
3.8
n/a
3.7

4.1
3.2
118
9.8
1.9
4.5
n/a
3.3
7.5
6.9
5.7
153
8.4
1.5
3.0
n/a
3.5
97.3
9.4
20.5
105 d
6.6
3.7
2.7
n/a
2.6

3.1
3.1
138
9.2
1.8
3.1
n/a
1.7

7,2
3.8
128
6.0
5.3
3.3
n/a
0.5

0.6
2.6
156 d
8.3
0.0
5.1
n/a
0.2

0.6
2.4
180
8.0
1.3
3.6
n/a
1.9
94.5
0.9
18.4
170
5.7
2.3
3.6
n/a
0.6

3.1
2.6
Matrix
Mean
10.7
1.7 '
3.1
n/a
5.6
19.3
5.0
7.6
a MDLs are highly matrix-dependant. MDLs provided in SW-846 are for guidance purposes and may not always be achievable. Labs should
establish their own in-house MDLs to document method performance.
b Congeners which are either certified or have had Soxhlet confirmation.
0 Parts per million (mg/kg), THEREFORE LOW MDL IS NOT APPROPRIATE - Use mean M9/k9 value.
d Congener 105 was not resolved from congener 132 and congener 156 was not resolved from congener 171 by the GC method used for samples
EC-1 and EC-5.
8082A - 37
Revision 1
January 1998
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TABLE 14
SINGLE LABORATORY RECOVERY DATA FOR SPE (METHOD 3535)
OF AROCLOR 1254 FROM WASTEWATER MATRICES SPIKED AT 2 yg/L

Mean Cone.
Percent
Std. Dev.
RSD
Wastewater Type
(m'l)
Recovery
(MQ/L)
(%)
Chemical Industry
2.39
120
0.41
- 17.2
Chemical Industry
0.56
28
0.03
5.4
Paper Industry
3.00
150
0.56
18.5
Paper Industry
2.30
115
0.08
3.7
Pharmaceutical Industry
1.52
76
0,03
1.7
Pharmaceutical Industry
1.02
51
0.03
2.9
Refuse
0.54
27
0.04
6.7
Refuse
0.63
31
0.10
16.0
POTW
1.92
96
0.15
7.8
POTW
2.10
105
0.04
1.8
Results represent three replicate solid-phase extractions of spiked wastewaters. Two different
wastewaters from each wastewater type were spiked. All extractions were performed using 90-mm
C18 extraction disks.
8082A - 38
Revision 1
January 1998
-------
TABLE 15
SINGLE LABORATORY RECOVERY DATA FOR SPE (METHOD 3535)
OF AROCLOR 1254 FROM WASTEWATER MATRICES SPIKED AT 10 jjg/L

Mean Cone,
Percent
Std. Dev.
RSD
Wastewater Type
(mq/l)
Recovery
(M9/L)
<%>
Chemical Industry
8.75
88
1.07
12.2
Chemical Industry
8.08
81
0.06
0.7
Paper Industry
8.88
889
0.71
7.9
Paper Industry
10.14
101
0.15
1.4
Pharmaceutical Industry
9.19
92
0.24
2.6
Pharmaceutical Industry
8.42
84
0.17
2.0
Refuse
8.80
88
0.49
5.6
Refuse
8.00
80
1.44
18.0
POTW
9.52
82
0.17
2.1
POTW
8.18
82
0.17
2.1
Results represent three replicate solid-phase extractions of spiked wastewaters. Two different
wastewaters from each wastewater type were spiked. All extractions were performed using 90-mm
C18 extraction disks.
S082A - 39
Revision 1
January 1998
-------
TABLE 16
SINGLE LABORATORY RECOVERY DATA SPE (METHOD 3S3S)
OF AROCLOR 1254 FROM WASTEWATER MATRICES SPIKED AT 100 |ig/L

Mean Cone.
Percent
Std. Dev.
RSD
Wastewater Type
(MQ/L)
Recovery
(MQ/L)
(%)
Chemical Industry
81.72
82
1.46
- 1.8
Chemical Industry
89.71
90
0.66
0.7
Paper Industry
73.73
74
3.94
5.3
Paper Industry
95.26
95
1.89
2.0
Pharmaceutical industry
86.41
86
1.95
2.3
Pharmaceutical Industry
79.16
79
3.92
4.9
Refuse
85.70
86
1.59
1.9
Refuse
71.50
72
1.61
2.2
POTW
87,76
88
1.76
2.0
POTW
80.59
81
0.40
0.5
Results represent three replicate solid-phase extractions of spiked wastewaters. Two different
wastewaters from each wastewater type were spiked. All extractions were performed using 90-mm
C1S extraction disks.
8082A - 40
Revision 1
January 1998
-------
DB-170
m
m
CM
DB-5
FIGURE 1. GC/ECD chromatogram of Arodor 1016 analyzed on a DB-5/DB-1701 fused-silica
open-tubular column pair. The GC operating conditions were as follows: 30 m x
0,53 mm ID DB-5 (1,5-pm film thickness) and 30 m x 0.53 mm ID DB-1701 (1 .(Him
film thickness) connected to a J&W Scientific press-fit Y-shaped inlet splitter.
Temperature program: 150°C (0.5 min hold) to 190°C (2 min hold) at 12°C/min then
to 275°C (10 min hold) at 4°C/min.
8082A - 41
Revision 1
January 1998
-------
crti'd 4*
OB-S
FIGURE 2. GC/ECD chromatogram of Aroclor 1221 analyzed on a DB-5/DB-1701 fused-silica
open-tubular column pair. The GC operating conditions were as follows: 30 m x
0.53 mm ID DB-5 (1.5-pm film thickness) and 30 m x 0.53 mm ID DB-1701 (1.0-pm
film thickness) connected to a J&W Scientific press-fit Y-shaped inlet splitter.
Temperature program: 150°C (0.5 min hold) to 190°C (2 min hold) at 12°C/min then
to 275CC (10 min hold) at 4°C/min.
8082A-42
Revision 1
January 1998
-------
DB-1701
ir>
r-*
DB-5
FIGURE 3. GC/ECD chromatogram of ArocJor 1232 analyzed on a DB-5/DB-1701 fused-sfllca
open-tubular column pair. The GC operating conditions were as follows: 30 m x
0.53 mm ID DB-5 (1.5-|jm film thickness) and 30 m x 0.53 mm ID DB-1701 (1,0-ym
film thickness) connected to a J&W Scientific press-fit Y-shaped inlet splitter.
Temperature program: 150°C (0.5 min hold) to 190°C (2 min hold) at 12°C/min then
to 275°C (10 min hold) at 4°C/min.
8082A-43
Revision 1
January 1998
-------
r» —
* to
9 -
OB-1701
m
tn r«) M N
• —tti KV «
II
C4 l-Hil*# k<
OB-5
f" »'
fit
I* W

FIGURE 4. GC/ECD chromatogram of Arodor 1242 analyzed on a DB-5/DB-1701 fused-sllica
open-tubular column pair. The GC operating conditions were as follows: 30 m x
0.53 mm ID DB-5 (1.5-|jm film thickness) and 30 m x 0.53 mm ID DB-1701 (1,0-pm
film thickness) connected to a J&W Scientific press-fit Y-shaped inlet splitter.
Temperature program: 150°C (0.5 min hold) to 190°C (2 mm told) at 12°C/min then
to 275°C (10 min hold) at 4°C/min.
8082A- 44
Revision 1
January 1998
-------
m
OB-1701
P '

DB-5
(4
-------
*i « DB-1701
m
«0U» Q N
— ffi ft
r«
fcil
m
0 dm 0K> —
ii 6 m w
r« «"»* aw* —

FIGURE 6. GC/ECD chromatogram of Aroclor 1254 analyzed on a DB-5/DB-1701 fused-silica
open-tubular column pair. The GC operating conditions were as follows: 30 m x
0.53 mm ID DB-5 (1.5-pm film thickness) and 30 m x 0.53 mm ID DB-1701 (1,.0-pm
film thickness) connected to a J&W Scientific press-fit Y-shaped inlet splitter.
Temperature program: 150°C (0.5 min hold) to 190°C (2 min hold) at 12°C/min then
to 275°C (10 min hold) at 4°C/min.
8082A - 46
Revision 1
January 1998
-------
DB-1701

¦ *—•*
il ift ¦ r An i ' '

1A1L
= t
DB-5
Jul
FIGURE 7. GC/ECD chromatogram of Aroclor 1260 analyzed on a DB-5/DB-1701 fused-silica
open-tubular column pair. The GC operating conditions were as follows: 30 m x
0.53 mm ID DB-5 (1.6-fim film thickness) and 30 m x 0.53 mm ID DB-1701 (1.0-fim
film thickness) connected to a J&W Scientific press-fit Y-shaped inlet splitter.
Temperature program: 150°C (0 J min hold) to 190°C (2 min hold) at 12°C/min then
to 275°C (10 min hold) at 4°C/min.
8082A-47
Revision 1
January 1998
-------
METHOD 8082A
POLYCHLORINATED BIPHENYLS (PCBs) BY GAS CHROMATOGRAPHY
V
7.1 Choose
appropriate extraction
technique.

r
7.1 Add specified
matrix spike to sample.

r
7.2 Perform
extract cleanup.

f
7.3 Set
chromatographic
conditions.

f
7.4 Perform
initial calibration.

7.5 Establish retention
time windows.

' Does N
response fall
within
.calibration
\range? >/
No
Yes
' 7.6.10 N
Any sample
peak inter-
ferences?
Yes
No
Stop
7.6.5 Dilute
extract.
7.6.3 Inject sample
extract.
7.6 Perform GC
analysis of sample
extracts.
7.7 - 7.9 Qualitative
and Quantitative peak
identification.
Take corrective
action. (Cleanup or
system adjustment
may be necessary.)
8082A - 48
Revision 1
January 1998
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METHOD 8141B
ORGANOPHOSPHORUS COMPOUNDS BY GAS CHROMATOGRAPHY
1.0 SCOPE AND APPLICATION
1.1 This method provides procedures for the gas chromatographic (GC) determination of
organophosphorus (OP) compounds. The compounds listed in the table below can be determined
by GC using capillary columns with a flame photometric detector (FPD) or a nitrogen-phosphorus
detector (NPD). Triazine herbicides can also be determined with this method when the NPD is used.
Although performance data are presented for each of the listed chemicals, it is unlikely that all of
them could be determined in a single analysis. This limitation results because the chemical and
chromatographic behavior of many of these chemicals can result in co-el ution. The analyst must
select columns, detectors, and calibration procedures for the specific analytes of interest. Any listed
chemical is a potential method interference when it is not a target analyte.
Analyte
CAS Registry No.
Orga nophosphorus Pesticides
Aspon"
Azinphos-methyl
Azinphos-ethyP
Bolstar (Sulprofos)
Carbophenothion8
Chlorfenvinphos"
Chlorpyrifos
Chlorpyrifos methyl"
Coumaphos
Crotoxyphos"
Demeton-Oc
Demeton-S®
Diazinon
Dichlorofenthion0
Dichlorvos (DDVP)
Dicrotophos3
Dimethoate
Dioxathionac
Disulfoton
EPN
Ethion®
Ethoprop
Famphur3
2104-64-5
563-12-2
13194-48-4
52-85-7
35400-43-2
786-19-6
470-90-6
2921-88-2
5598-13-0
56-72-4
7700-17-6
8065-48-3
8065-48-3
333-41-5
97-17-6
62-73-7
141-66-2
60-51-5
78-34-2
298-04-4
3244-90-4
86-50-0
2642-71-9
8141B- 1
Revision 2
January 1998
-------
Analyte
CAS Registry No.
Fenitrothion3
122-14-5
Fensulfothion
115-90-2
Fenthion
55-38-9
Fonophos"
944-22-9
Leptophosa,d
21609-90-5
Malathion
121-75-5.
Merphos0
150-50-5
Mevinphos*
7786-34-7
Monocrotophos
6923-22-4
Naled
300-76-5
Parathion, ethyl
56-38-2
Parathion, methyl
298-00-0
Phorate
298-02-2
Phosmef
732-11-6
Phosphamidorf
13171-21-6
Ronnel
299-84-3
Stirophos (Tetrachlorvinphos)
22248-79-9
Suifotepp
3689-24-5
Tetraethyl pyrophosphate (TEPP)d
107-49-3
Tertoufos*
13071-79-9
Thionazinab (Zinophos)
297-97-2
Tokuthion" (Prothiofos)
34643-46-4
Trichtorfon8
52-68-6
Trichloronateb
327-98-0
Industrial Chemicals

Hexamethyf phosphoramide8 (HMPA)
680-31-9
Tri-o-cresyl phosphate8,15 (TOCP)
78-30-8
Triazine Herbicides (NPD only)
....
Atrazine®
1912-24-9
Simazine3
122-34-9
a This analyte has been evaluated using a 30-m column only (see Sec. 1.5).
b Production discontinued in the U.S., standard not readily available.
0 Standards may have multiple components because of oxidation.
" Compound is extremely toxic or neurotoxic.
• Adjacent major/minor peaks can be observed due to cis/trans isomers.
8141B-2
Revision 2
January 1998
-------
1.2 A dual-column/dual-detector approach may be used for the analysis of relatively dean
extracts. Two 15- or 30-m x 0,53-mm ID fused-silica, open-tubular columns of different polarities
are connected to an injection tee and each is connected to a detector. Analysts are cautioned
regarding the use of a dual column configuration when their instrument is subject to mechanical
stress, when many samples are analyzed over a short time, or when extracts of contaminated
samples are analyzed.
1.3 Two detectors can be used for the listed organophosphorus chemicals. The FPD works
by measuring the emission of phosphorus- or sulfur-containing species. Detector performance is
optimized by selecting the proper optical filter and adjusting the hydrogen and air flows to the flame.
The NPD is a flame ionization detector with a rubidium ceramic flame tip which enhances the
response of phosphorus- and nitrogen-containing analytes. The FPD is more sensitive and more
selective, but is a less common detector in environmental laboratories.
1.4 Table 1 lists method detection limits (MDLs) for the target analytes, using 15-m
columns and FPD, for water and soil matrices. Table 2 lists the estimated quantitation limits (EQLs)
for other matrices. MDLs and EQLs using 30-m columns will be very similar to those obtained from
15-m columns, however, laboratories should determine in-house MDLs for the analytes of Interest
using the specific instrumentation employed for sample analysis.
1.5 The use of a 15-m column system has not been fully validated for the determination of
all of the compounds listed in Sec. 1.1. The analyst must demonstrate chromatographic resolution
of all analytes, recoveries of greater than 70 percent, with precision of no more than 15 percent RSD,
before data generated on the 15-m column system can be reported for the following analytes, or any
additional analytes:
1.6 When Method 8141 is used to analyze unfamiliar samples, compound identifications
should be supported by confirmatory analysis. Sec. 8.0 provides gas ehromatograph/mass
spectrometer (GC/MS) criteria appropriate for the qualitative confirmation of compound
identifications.
1.7 This method is restricted to use by, or under the supervision of, analysts experienced
in the use of capillary gas chromatography and in the interpretation of chromatograms.
2.0 SUMMARY OF METHOD
2.1 Method 8141 provides gas chromatographic conditions for the determination of part per
billion concentrations of organophosphorus compounds. Prior to the use of this method, appropriate
sample preparation techniques must be used. Water samples are extracted at a neutral pH with
methylene chloride by using a separatory funnel (Method 3510), a continuous liquid-liquid extractor
(Method 3520), solid-phase extraction (Method 3535), or other appropriate technique. Solid samples
are extracted using Soxhlet extraction (Method 3540) or automated Soxhlet extraction (Method
3541), using methylene chloride/acetone (1:1), pressurized fluid extraction (Method 3545), or other
Azinphos-ethyl
Ethion
Carbophenothion
Famphur
Phosphamidon
Chlorfenvinphos
HMPA
Terbufos
Leptophos
TOCP
Phosmet
Dioxathion
8141B-3
Revision 2
January 1998
-------
appropriate technique. Both neat and diluted organic liquids (Method 3580, Waste Dilution) may be
analyzed by direct injection.
2.2 Analysis is performed on a gas chromatograph with a flame photometric or nitrogen-
phosphorus detector.
2.3 Organophosphorus esters and thioesters can hydrolyze under both add and base
conditions. Therefore, sample preparation procedures employing acid and base partitioning
procedures are not appropriate for extracts to be analyzed by Method 8141.
2.4 Ultrasonic Extraction (Method 3550) is not an appropriate sample preparation method
for Method 8141 because of the potential for destruction of target analytes during the ultrasonic
extraction process and should not be used.
3.0 INTERFERENCES
3.1 Refer to Methods 3500, 3600, and 8000, as well as to Sec. 1.1.
3.2 The use of Florisil Cleanup (Method 3620) for some of the compounds in this method
has been demonstrated to yield recoveries less than 85 percent and is therefore not recommended
for all compounds. Refer to Table 2 of Method 3620 for recoveries of organophosphorus
compounds. Use of an FPD often eliminates the need for sample cleanup. If particular
circumstances demand the use of an alternative cleanup procedure, the analyst must determine the
elution profile and demonstrate that the recovery of each analyte is not less than 85 percent.
3.3 The use of gel permeation cleanup (GPC) (Method 3640) for extract cleanup has been
demonstrated to yield recoveries of less than 85 percent for many method anatytes because they
elute before bis-(2-ethylhexyl) phthalate. Therefore Method 3640 is not recommended for use with
this method, unless analytes of interest are listed in Method 3640 or are demonstrated to give
greater than 85 percent recovery.
3.4 Use of a flame photometric detector in the phosphorus mode will minimize interferences
from materials that do not contain phosphorus or sulfur. Elemental sulfur will interfere with the
determination of certain organophosphorus compounds by flame photometric gas chromatography.
If Method 3660 is used for sulfur cleanup, only the tetrabutylammonium (TBA)-suIfite option should
be employed, since rapper may destroy OP pesticides. The stability of each analyte must be tested
to ensure that the recovery from the TBA-sulfite sulfur cleanup step is not less than 85 percent.
3.5 A halogen-specific detector (i.e., electrolytic conductivity or microcoulometry) is very
selective for the halogen-containing compounds and may be used for the determination of
Chlorpyrifos, Ronnel, Coumaphos, Tokuthion, Trichloronate, Dichlorvos, EPN, Naled, and Stirophos
only. Many of the OP pesticides may also be detected by the electron capture detector (ECD),
however, the ECD is not as specific as the NPD or FPD. The ECD should only be used when
previous analyses have demonstrated that interferences will not adversely effect quantitation, and
that the detector sensitivity is sufficient to meet project requirements..
3.6 Certain analytes will coeiute, particularly on 15-m columns (Table 3). If coelution is
observed, analysts should (1) select a second column of different polarity for confirmation, (2) use
30-m x 0.53-mm columns, or (3) use 0.25- or 0.32-mm ID columns. See Figures 1 through 4 for
combinations of compounds that do not coeiute on 15-m columns.
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3.7 The following pairs coeluted on the DB-5/DB-210 30-m column pair:
GC Column Coeluting pair
DB-5 T erbufos/tri-o-cresyl phosphate
Naled/Simazine/Atrazine
Dichlorofenthion/Demeton-0
T richloronate/Aspon
Bolstar/Stirophos/Carbophenothion
Phosphamidon/Crotoxyphos
Fensulfothion/EPN
DB-210 Terbufos/tri-o-cresyl phosphate
Dichlorofenthion/P hosphamidori
Chlorpyrifos, methyl/Parathion, methyl
Chlorpyrifos/Parathion, ethyl
Aspon/Fenthion
Demeton-O/Di m ethoate
Leptophos/Azinphos-methyl
EPN/Phosmet
Famphur/Carbophenothion
See Table 4 for the retention times of these compounds on 30-m columns.
3.8 Analytical difficulties encountered for target analytes
3.8.1 Tetraethyl pyrophosphate (TEPP) is an unstable diphosphate which is readily
hydrolyzed in water and is thermally labile (decomposes at 170°C). Care must be taken to
minimize loss during GC analysis and during sample preparation. Identification of bad
standard Jots is difficult since the electron impact (El) mass spectrum of TEPP is nearly
identical to its major breakdown product, triethyl phosphate.
3.8.2 The water solubility of Dichlorvos (DDVP) is 10 g/L at 20°C, and recovery is
poor from aqueous solution.
3.8.3 Naled is converted to Dichlorvos (DDVP) on column by denomination. This
reaction may also occur during sample preparation. The extent of debromination will depend
on the nature of the matrix being analyzed. The analyst must consider the potential for
debromination when Naled is to be determined.
3.8.4 Trichlorfon rearranges and is dehydrochlorinated in acidic, neutral, or basic
media to form Dichlorvos (DDVP) and hydrochloric acid. If this method is to be used for the
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determination of organophosphates in the presence ofTrichlorfon, the analyst should be aware
of the possibility of its rearrangement to Dichlorvos and the possibility of misidentification.
' 3.8.5 Demeton (Systox) is a mixture of two compounds; O.O-diethyl
0-[2-(ethyfthio)ethyl]phosphorothioate (Demeton-O) and O.O-diethyl S-[2-
(ethylthio)ethyl]phosphorothioate (Demeton-S). Two peaks are observed in all the
chromatograms corresponding to these two isomers, it is recommended that the early eluting
compound (Demeton-S) be used for quantitation.
3.8.6 Dioxathion is a single-component pesticide. However, several extra peaks
are observed in the chromatograms of standards. These peaks appear to be the result of
spontaneous oxygen-sulfur isomerization. Because of this, Dioxathion is not included in
composite standard mixtures.
3.8.7 Merphos (tributyl phosphorotrithioite) is a single-component pesticide that is
readily oxidized to its phosphorotrithioate (Merphos oxone). Chromatographic analysis of
Merphos almost always results two peaks (unoxidized Merphos ehites first). As the relative
amounts of oxidation of the sample and the standard are probably different, quantitation based
on the sum of both peaks may be most appropriate.
3.8.8 Retention tines of some analytes, particularly Monocrotophos, may increase
with increasing concentrations in the injector. Analysts should check for retention time shifts
in highly-contaminated samples.
3.8.9 Many analytes will degrade on reactive sites in the chromatographic system.
Analysts must ensure that injectors and splitters are free from contamination and are silanized.
Columns should be installed and maintained properly.
3.8.10 Performance of chromatographic systems will degrade with time. Column
resolution, analyte breakdown and baselines may be improved by column washing (Sec. 7).
Oxidation of columns is not reversible.
3.9 Metfiod interferences may be caused by contaminants in solvents, reagents, glassware,
and other sample processing hardware that lead to discrete artifacts or elevated baselines in gas
chromatograms. All these materials must be routinely demonstrated to be free from interferences
under the conditions of the analysis by analyzing reagent blanks (Sec. 8.0).
3.10 NP Detector interferences - Triazine herbicides, such as atrazine and simazine, and
other nitrogen-containing compounds may interfere.
4.0 APPARATUS AND MATERIALS
4.1 Gas chromatograph
An analytical system complete with a gas chromatograph suitable for on-column or
spJit/splitless injection, and all required accessories, including syringes, analytical columns, gases,
suitable detectors), and a recording device. The analyst should select the detector for the specific
measurement application, either the flame photometric detector or the nitrogen-phosphorus detector.
A data system for measuring peak areas and dual display of chromatograms is highly recommended.
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4.2 GC columns
This method employs capillary columns (0.53-mm, 0.32-mm, or 0.25-mm ID and 15-m or 30-m
length, depending on the resolution required). Columns of 0.53-mm ID are recommended for most
environmental and waste analysis applications. Dual-coiumn, single-injector analysis requires
columns of equal length and bore. See Sec. 3.0 and Figures 1 through 4 for guidance on selecting
the proper length and diameter for the column(s). Four columns are recommended.
4.2.1 Column 1-15-m or 30-m x 0.53-mm wide-bore capillary column, 1.0-jjm film
thickness, chemically bonded with 50% trifluoropropyl polysiloxane, 50% methyl .polysiloxane
(DB-210), or equivalent.
4.2.2 Column 2 - 15-m or 30-m x 0.53-mm wide-bore capillary column, 0.83-jjm film
thickness, chemically bonded with 35% phenyl methyl polysiloxane (DB-608, SPB-608, RTx-
35), or equivalent.
4.2.3 Column 3 -15-m or 30-m x 0.53-mm wide-bore capillary column, 1.0 pm film
thickness, chemically bonded with 5% phenyl polysiloxane, 95% methyl polysiloxane (DB-5,
SPB-5, RTx-5), or equivalent.
4.2.4 Column 4 -15- or 30-m x 0.53-mm ID wide-bore capillary column, chemically
bonded with methyl polysiloxane (DB-1, SPB-1, or equivalent), 1.0-pm or 1.5-|Jm film
thickness.
4.2.5 Column rinsing kit (optional) - Bonded-phase column rinse kit (J&W Scientific,
catalog no. 430-3000, or equivalent).
4.3 Splitter - If a dual-column, single-injector configuration is used, the open tubular
columns should be connected to one of the following splitters, or equivalent:
4.3.1 Splitter 1 - J&W Scientific press-fit Y-shaped glass 3-way union splitter (J&W
Scientific, catalog no. 705-0733).
4.3.2 Splitter 2 - Supelco 8-in glass injection tee, deactivated (Supelco, catalog no.
2-3668M).
4.3.3 Splitter 3 - Restek Y-shaped fused-siiica connector (Restek, catalog no.
20405).
4.4 Injectors
4.4.1 Packed column, 1/4-in injector port with hourglass liner are recommended for
0.53-mm column. These injector ports can be fitted with splitters (Sec. 4.3) for dual-column
analysis.
4.4.2 Split/splitless capillary injectors operated in the split mode are required for
0.25-mm and 0.32-mm columns.
4.5 Detectors
4.5.1 Flame Photometric Detector (FPD) operated in the phosphorus-specific mode
is recommended.
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4.5.2 Nitrogen-Phosphorus Detector (NPD) operated in the phosphorus-specific
mode is less selective but can detect triazine herbicides.
4.5.3 Halogen-Specific Detectors (electrolytic conductivity or microcoulometry) may
be used only for a limited number of halogenated or sulfur-containing analytes (Sec. 3.5).
4.5.4 Electron-capture detectors may be used for a limited number of analytes (Sec.
3.5).
4.6 Data system
4.6.1 A data system capable of presenting chromatograms, retention time, and
peak integration data is strongly recommended.
4.6.2 Use of a data system that allows storage of raw chromatographic data is
strongly recommended.
REAGENTS
5.1 Solvents - All solvents must be pesticide quality or equivalent.
5.1.1 Isooctane, (CH3)3CCH2CH(CH3)2
5.1.2 Hexane, C6H„
5.1.3 Acetone, CH3COCH3
5.1.4 Tetrahydrofuran (THF), C4HeO - for triazine standards only.
5.1.5 Methyl ferf-butyl-ether (MTBE), CH3Of-C4H9 -for triazine standards only.
5.2 Stock standard solutions (1000 mg/L) - May be prepared from pure standard materials
can be purchased as certified solutions.
5.2.1 Prepare stock standard solutions by accurately weighing about 0.0100 g of
pure compounds. Dissolve the compounds in suitable mixtures of acetone and hexane and
dilute to volume in a 10-mL volumetric flask. If compound purity is 96 percent or greater, the
weight may be used without correction to calculate the concentration of the stock standard
solution. Commercially-prepared stock standard solutions may be used at any concentration
if they are certified by the manufacturer or by an independent source.
5.2.2 Both Simazine and Atrazine have low solubilities in hexane. If standards of
these compounds are required, Atrazine should be dissolved in MTBE, and Simazine should
be dissolved in acetone/MTBE/THF (1:3:1).
5.2.3 Composite stock standard - This standard may be prepared from individual
stock solutions. The analyst must demonstrate that the individual analytes and common
oxidation products are resolved by the chromatographic system. For composite stock
standards containing less than 25 components, take exactly 1 mL of each individual stock
solution at 1000 mg/L, add solvent, and mix the solutions in a 25-mL volumetric flask. For
example, for a composite containing 20 individual standards, the resulting concentration of
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each component in the mixture, after the volume is adjusted to 25 mL, will be 40 mg/L. This
composite solution can be further diluted to obtain the desired concentrations. Composite
stock standards containing more than 25 components are not recommended.
5.2.4 Store the standard solutions (stock, composite, calibration, internal, and
surrogate) at 4°C in PTFE-sealed containers in the dark. All standard solutions should be
replaced after two months, or sooner if routine QC (Sec. 8.0) indicates a problem. Standards
for easily hydrolyzed chemicals including TEPP, Methyl Parathion, and Merphos should be
checked every 30 days.
5.2.5 It is recommended that lots of standards be subdivided and stored in small
vials. Individual vials should be used as working standards to minimize the potential for
contamination or hydrolysis of the entire lot.
5.3 Calibration standards should be prepared at a minimum of five concentrations by
dilution of the composite stock standard with isooctane or hexane. The concentrations should
correspond to the expected range of concentrations found in real samples and should bracket the
linear range of the detector. Organophosphorus calibration standards should be replaced after one
or two months, or sooner if comparison with check samples or historical data indicates that there is
a problem. Laboratories may wish to prepare separate calibration solutions for the easily hydrolyzed
standards identified above.
5.4 internal standard
Internal standards should only be used on well-characterized samples by analysts experienced
in the technique. Use of internal standards is complicated by coelution of some OP pesticides and
by the differences in detector response to dissimilar chemicals, if internal standards are to be used,
the analyst must select one or more internal standards that are similar in analytical behavior to the
compounds of interest. The analyst must further demonstrate that the measurement of the internal
standard is not affected by method or matrix interferences.
5.4.1 FPD response for organophosphorus compounds is enhanced by the
presence of sulfur atoms bonded to the phosphorus atom. It has not been established that a
thiophosphate can be used as an internal standard for an OP with a different numbers of sulfur
atoms (e.g., phosphorothioates [P=S] as an internal standard for phosphates [POJ) or
phosphorodithioates [P=SJ).
5.4.2 When 15-m columns are used, it may be difficult to fully resolve internal
standards from target analytes and interferences. The analyst must demonstrate that the
measurement of the internal standard is not affected by method or matrix interferences.
5.4.3 1 -bromo-2-nitrobenzene has been used as an NPD internal standard for a 30-
m column pair. Prepare a solution of 1000 mg/L of 1-bromo-2-nitrobenzene. For spiking, dilute
this solution to 5 mg/L Use a spiking volume of 10 jjL/mL of extract The spiking concentration
of the internal standards should be kept constant for all samples and calibration standards.
Since its FPD response is small, 1 -bromo-2-nitrobenzene is not an appropriate internal
standard for that detector. No FPD internal standard is suggested.
5.5 Surrogates
The analyst should monitor the performance of the extraction, cleanup (when used), and
analytical system, and the effectiveness of the method in dealing with each sample matrix, by spiking
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each sample, standard, and blank with one or two surrogates (e.g., organophosphorus compounds
not expected to be present in the sample). If multiple anatytes are to be measured, two surrogates
(an early and a late eluter) are recommended, Deuterated analogs of analytes are not appropriate
surrogates for gas chromatographic/FPD/NPD analysis.
5.5.1 If surrogates are to be used, the analyst must select one or more compounds
that are similar in analytical behavior to the compounds of interest. The analyst must further
demonstrate that the measurement of a surrogate is not affected by method or matrix
interferences. General guidance on Ihe selection and use of surrogates is provided in Sec. 5.0
of Method 3500.
5.5.2 Tributyl phosphate and triphenyl phosphate are recommended as surrogates
for either FPD and NPD analyses. A volume of 1.0 mL of a 1-pg/L spiking solution (containing
1 ng of surrogate) is added to each sample. If there is a co-elution problem with either of these
compounds, 4-chloro-3-nitrobenzo-trifluoride has also been used as a surrogate for NPD
analysis.
6.0 SAMPLE COLLECTION, PRESERVATION, AND HANDLING
6.1 See the introductory material to Chapter Four, "Organic Analytes," Sec. 4.0.
6.2 Even with storage at 4°C and use of mercuric chloride as a preservative, most
organophosphorus pesticides in groundwater samples collected for a national pesticide survey
degraded within a 14-day period. Therefore, begin sample extraction within 7 days of collection.
6.3 Store extracts at 4°C and perform analyses within 40 days of extraction.
6.4 Organophosphorus esters will hydrolyze under acidic or basic conditions. Adjust
samples to a pH of 5 to 8 using sodium hydroxide or sulfuric acid solution as soon as possible after
sample collection. Record the volume used.
7.0 PROCEDURE
7.1 Extraction and cleanup
Refer to Chapter Two and Method 3500 for guidance on choosing the appropriate extraction
procedure. In general, water samples are extracted at a neutral pH with methylene chloride, using
Method 3510,3520,3535, or other appropriate technique. Solid samples are extracted using either
Method 3540 or 3541 with methylene chloride/acetone (1:1 v/v) or hexane/acetone (1:1 v/v) as the
extraction solvent, Method 3545, or other appropriate technique.
Method 3550 is not an appropriate extraction technique for the target analytes of this method
(See Sec. 2.4).
Extraction and cleanup procedures that use solutions below pH 4 or above pH 8 are not
appropriate for this method.
7,1.1 If required, the sample extracts may be cleaned up using Florisii column
cleanup (Method 3620) and sulfur cleanup (Method 3660, TBA-sulfite option), which may have
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particular application for organophosphorus pesticides. Gel permeation cleanup (Method
3640) should not generally be used for organophosphorus pesticides.
7.1.2 If sulfur cleanup by Method 3660 is required, do not use the copper technique,
as the target anatytes may be degraded in the presence of copper.
7.1.3 GPC may only be employed if all the target organophosphorus pesticides of
interest are listed as anatytes of Method 3640, or if the laboratory has demonstrated a recovery
of greater than 85 percent for target organophosphorus pesticides at a concentration not
greater than 5 times the levels of interest (e.g., the regulatory limit). Laboratories must retain
data demonstrating acceptable recovery.
7.1.4 Prior to gas chromatographic analysis, the extract solvent may be exchanged
to hexane. The analyst must ensure quantitative transfer of the extract concentrate. Single-
laboratory data indicate that samples should not be transferred with 100-percent hexane during
sample workup, as the more polar organophosphorus compounds may be lost. Transfer of
oiganophosphorus esters is best accomplished using methylene chloride or a hexane/acetone
solvent mixture.
7.1.5 Methylene chloride may be used as an injection solvent with both the FPD and
the NPD.
NOTE: Follow manufacturer's instructions as to suitability of using methylene chloride with
any specific detector.
7.2 Gas chromatographic conditions
Four different 0.53-mm ID capillary columns are suggested for the determination of
organophosphates by this method. Column 1 (DB-210,or equivalent) and Column 2 (SPB-608,or
equivalent) of 30-m lengths are recommended if a large number of organophosphorus analytes are
to be determined. If superior chromatographic resolution is not required, 15-m columns may be
appropriate.
7.2.1 Suggested operating conditions for 15-m columns are listed in Table 8.
Suggested operating conditions for 30-m columns are listed in Table 9.
7.2.2 Retention times for analytes on each set of columns are presented in Tables
3 and 4. These data were developed using the operating conditions in Tables 8 and 9.
7.2.3 Establish the GC operating conditions appropriate for the column employed,
using Tables 8 and 9 as guidance. Optimize the instrumental conditions for resolution of the
target analytes and sensitivity.
NOTE: Once established, the same operating conditions must be used for both calibrations
and sample analyses.
7.3 Calibration
Refer to Method 8000 for proper calibration techniques. Use Table 8 and Table 9 for
establishing the proper operating parameters for the set of columns being employed in the analyses.
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7.4 Gas chromatographic analysis
Method 8000 provides instructions on the analysis sequence, appropriate dilutions, establishing
daily retention time windows and identification criteria.
7.4.1 Automated 1-jjL injections are recommended. Manual injections of no more
than 2 pL may be used if the analyst demonstrates quantitation precision of s 10 percent
relative standard deviation. The solvent flush technique may be used if the amount of solvent
is kept at a minimum. If the internal standard calibration technique is used, add 10 pL of
internal standard to each 1 mL of sample, prior to injection. Chromatograms of the target
organophosphorus compounds are shown in Figures 1 through 4.
7.4.2 Figures 5 and 6 show chromatograms with and without Simazine, Atrazine,
and Carbophenothion on 30-m columns.
7.5 Record the sample volume injected to the nearest 0.05 yL and the resulting peak sizes
(in area units or peak heights). Using either the internal or external calibration procedure (Method
8000), determine the identity and quantity of each component peak in the sample chromatogram
which corresponds to the compounds used for calibration purposes. See Method 8000 for
calculations.
7.5.1 If peak detection and identification are prevented by the presence of
interferences, the use of an FPD or further sample cleanup is required. Before using any
cleanup procedure, the analyst must process a series of calibration standards through the
procedure to establish elution patterns and to determine recovery of target compounds.
7.5.2 If the responses exceed the linear range of the system, dilute the extract and
reanalyze. It is recommended that extracts be diluted so that all peaks are on scale.
Overlapping peaks are not always evident when peaks are off-scale. Computer reproduction
of chromatograms, manipulated to ensure all peaks are on scale over a 100-fold range, are
acceptable if linearity is demonstrated. Peak height measurements are recommended over
peak area integration when overlapping peaks cause errors in area integration.
7.5.3 If the peak response is less than 2.5 times the baseline noise level, the
validity of the quantitative result may be questionable. The analyst should consult with the
source of the sample to determine whether further concentration of the sample extract is
warranted.
7.5.4 If partially overlapping or coeluting peaks are found, change columns or try
a GC/MS technique. Refer to Sec. 8.0 and Method 8270.
7.6 Suggested chromatograph maintenance
Connective measures may require any one or more of the following remedial actions. Refer to
Method 8000 for general information on the maintenance of capillary columns and injectors.
7.6.1 Splitter connections - For dual columns which are connected using a press-fit
Y-shaped glass splitter or a Y-shaped fused-silica connector, clean and deactivate the splitter.
Reattach the columns after cleanly cutting off at least one foot from the injection port side of
the column using a capillary cutting tool or scribe. The accumulation of high boiling residues
can change split ratios between dual columns and thereby change calibration factors.
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7.6.2 Columns wifl be damaged permanently and irreversibly by contact with oxygen
at elevated temperature. Oxygen can enter the column during a septum change, when oxygen
traps are exhausted, through neoprene diaphragms of regulators, and through leaks in the gas
manifold. Polar columns including the DB-210 and DB-608 are more prone to oxidation.
Oxidized columns will exhibit baselines that rise rapidly during temperature programming.
7.6.3 Peak tailing for all components will be exacerbated by dirty injectors, pre-
columns, and glass "Vs. Additionally, cleaning of this equipment (or replacement/clipping, as
appropriate) will greatly reduce the peak tailing. Components such as Fensulfothion, Naled,
Azinphos-methyl, and Dimethoate are very good indicators of system performance.
7.7 Detector maintenance
7.7.1 Older FPDs may be susceptible to stray light in the photomultiplier tube
compartment. This stray light will decrease the sensitivity and the linearity of the detector.
Analysts can check for leaks by initiating an analysis in a dark room and turning on the lights.
A shift in the baseline indicates that light may be leaking into the photomultiplier tube
compartment. Additional shielding should be applied to eliminate light leaks and minimize stray
light interference.
7.7.2 The bead of the NPD will become exhausted with time, which will decrease
the sensitivity and the selectivity of the detector. The collector may become contaminated
which decreased detector sensitivity.
7.7.3 Both types of detectors use a flame to generate a response. Flow rates of
air and hydrogen should be optimized to give the most sensitive, linear detector response for
target analytes.
7.8 GC/MS confirmation
7.8.1 GC/MS techniques should be judiciously employed to support qualitative
identifications made with this method. Follow the GC/MS operating requirements described
in Method 8270. GC/MS confirmation may be used in conjunction with either single-column
or dual-column analysis if the concentration is sufficient for detection by GC/MS.
7.8.2 The GC/MS must be calibrated for the specific target pesticides when it is
used for quantitative analysis. If GC/MS is used only for confirmation of the identification of
the target analytes, then the analyst must demonstrate that those pesticides identified by
GC/ECD can be confirmed by GC/MS. This demonstration may be accomplished by analyzing
a single-point standard containing the analytes of interest at or below the concentrations
reported in the GC/EC analysis.
7.8.3 GC/MS confirmation should be accomplished by analyzing the same extract
that is used for GC analysis and the extract of the associated method blank.
7.8.4 Where available, chemical ionization mass spectra may be employed to aid
in the qualitative identification process because of the extensive fragmentation of
organophosphorus pesticides during electron impact MS processes.
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8.0 QUALITY CONTROL
8.1 Refer to Chapter One and Method 8000 for specific quality control (QC) procedures.
Quality control procedures to ensure the proper performance of the various sample preparation
techniques can be found in Method 3500. If an extract cleanup procedure was performed, refer to
Method 3600 for the appropriate quality control procedures. Each laboratory should maintain a
formal quality assurance program. The laboratory should also maintain records to document the
quality of the data generated.
8.2 Quality control procedures necessary to evaluate the GC system operation are found
in Method 8000, Sec. 7.0 and include evaluation of retention time windows, calibration verification,
and chromatographic analysis of samples.
8.3 Initial Demonstration of Proficiency
8.3.1 Each laboratory must demonstrate initial proficiency with each sample,
preparation and determinative method combination it utilizes, by generating data of acceptable
accuracy and precision for target analytes in a clean matrix. The laboratory must also repeat
the following operations whenever new staff are trained or significant changes in
instrumentation are made.
8.3.2 It is suggested that the quality control (QC) reference sample concentrate (as
discussed in Section 8.0 of Methods 8000 and 3500} contain each analyte of interest at 10
mg/L. See Method 8000, Sec. 8.0 for additional information on how to accomplish this
demonstration.
8.3.3 Calculate the average recovery and the standard deviation of the recoveries
of the analytes in each of the four QC reference samples. Refer to Sec. 8.0 of Method 8000
for procedures for evaluating method performance.
8.4 Sample Quality Control for Preparation and Analysis - The laboratory must also have
procedures for documenting the effect of the matrix on method performance (precision, accuracy,
and detection limit). At a minimum, this includes the analysis of QC samples including a method
blank, a matrix spike, a duplicate, a laboratory control sample (LCS), and the addition of surrogates
to each field sample and QC sample.
8.4.1 Documenting the effect of the matrix should include the analysis of at least
one matrix spike and one duplicate unspiked sample or one matrix spike/matrix spike duplicate
pair. The decision on whether to prepare and analyze duplicate samples or a matrix
spike/matrix spike duplicate must be based on a knowledge of the samples in the sample
batch, if samples are expected to contain target analytes, then laboratories may use one
matrix spike and a duplicate analysis of an unspiked field sample. If samples are not expected
to contain target analytes, the laboratories should use a matrix spike and matrix spike duplicate
pair.
8.4.2 In-house method performance criteria should be developed using the
guidance found in Sec. 8.0 of Method 8000 for procedures for evaluating method performance.
8.4.3 A Laboratory Control Sample (LCS) should be included with each analytical
batch. The LCS consists of an aliquot of a clean (control) matrix similar to the sample matrix
and of the same weight or volume. The LCS is spiked with the same analytes at the same
concentrations as the matrix spike. When the results of the matrix spike analysis indicates a
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potential problem due to the sample matrix itself, the LCS results are used to verify that the
laboratory can perform the analysis in a clean matrix.
8.4.4 Include a calibration standard after each group of 20 samples (it is
mcornmndmithat a calibration standard be included after every 10 samples to minimize the
number of repeat injections) in the analysis sequence as a calibration check. Thus, injections
of method blank extracts, matrix spike samples, and other non-standards are counted in the
total. Solvent blanks, injected as a check on cross-contamination, need not be counted in the
total. The calibration factors for the calibration should be within ±15% of the initial calibration.
When this calibration verification standard falls out of this acceptance window, the laboratory
should stop analyses and take corrective action.
8.4.5 Whenever quantitation is accomplished using an internal standard, internal
standards must be evaluated for acceptance. The measured area of the internal standard
must be no more than 50 percent different torn the average area calculated during calibration.
When the internal standard peak area is outside the limit, all samples that fall outside the QC
criteria must be reanalyzed.
8.4.6 See Method 8000, Sec. 8.0 for the details on carrying out sample quality
control procedures for preparation and analysis.
8.5 Surrogate recoveries
The laboratory must evaluate surrogate recovery data from individual samples versus the
surrogate control limits developed by the laboratory. See Method 8000, Sec. 8.0 for information on
evaluating surrogate data and developing and updating surrogate limits.
8.6 It is recommended that the laboratory adopt additional quality assurance practices for
use with this method. The specific practices that are most productive depend upon the needs of the
laboratory and the nature of the samples. Whenever possible, the laboratory should analyze
standard reference materials and participate in relevant performance evaluation studies.
9.0 METHOD PERFORMANCE
9.1 Estimated MDLs and associated chromatographic conditions for water and clean soil
(uncontaminated with synthetic organics) are listed in Table 1. As detection limits will vary with the
particular matrix to be analyzed, guidance for determining EQLs is given in Table 2.
9.2 Recoveries for some method analytes are provided in Tables 5,6, and 7.
9.3 Tables 11 and 12 present data for solid-phase extraction of ground water and waste
water samples. Forty four organophosphorus compounds were divided into three sets of analytes.
Each set was spiked into seven 250-ml replicate samples of ground water and a waste water at
10 ppb and at 250 ppb. Ground water was obtained from the Stroh Brewery in St. Paul, MN, while
the waste water was obtained from a chemical manufacturing plant. The water samples were
extracted using a 47-mm Empore™ Extraction Disk with SDB-RPS, a reverse-phase, sulfonated,
poly(styrenedivinylbenzene) copolymer adsorbent. The samples were analyzed using gas
chromatography with a nitrogen-phosphorous detector.
9.4 Single-laboratory accuracy data were obtained for organophosphorus pesticides at two
different spiking concentrations in three different soil types. Spiking concentrations ranged from 250
8141B-15
Revision 2
January 1998
-------
to 2500 (jgfcg for the OPPs. Spiked samples were extracted both by the Dionex Accelerated Solvent
Extractor and by Soxhlet. Table 13 contains the data for the recoveries of the analytes from
pressurized fluid extraction as a percentage of the amount recovered by Soxhlet for all three soils.
Table 14 contains the bias, calculated as a percentage of the spiked concentration and the precision
of those results, calculated as the relative standard deviation (RSD). All data are taken from
Reference 15.
10.0 REFERENCES
1. Taylor, V.; Hickey, D.M.; Marsden, P.J. "Single Laboratory Validation of EPA Method 8140,"
U.S. Environmental Protection Agency, Environmental Monitoring Systems Laboratory, Office
of Research and Development, Las Vegas, NV, 1987; EPA-600/4-87-009.
2. Pressley, T.A; Longbottom, J.E. 'The Determination of Organophosphorus Pesticides in
Industrial and Municipal Wastewater. Method 614," U.S. Environmental Protection Agency,
Environmental Monitoring and Support Laboratory, Cincinnati, OH, 1982; EPA-600/4-82-004.
3. "Analysis of Volatile Hazardous Substances by GC/MS: Pesticide Methods Evaluation," Letter
Reports 6,12A, and 14 to the U.S. Environmental Protection Agency on Contract 68-03-2697,
1982.
4. "Method 622, Organophosphorus Pesticides," U.S. Environmental Protection Agency,
Environmental Monitoring and Support Laboratory, Cincinnati, OH 45268.
5. Lopez-Avila, V.; Baldin, E.; Benedicto, J; Milanes, J.; Beckert, W. F. "Application of Open-
Tubular Columns to SW-846 GC Methods," final report to the U.S. Environmental Protection
Agency on Contract 68-03-3511, Mid-Pacific Environmental Laboratory, Mountain View, CA,
1990.
6. Hatcher, M.D.; Hickey, D.M.; Marsden, P.J.; and Betowski, L.D.; "Development of a GC/MS
Module for RCRA Method 8141," final report to the U.S. EPA Environmental Protection Agency
on Contract 68-03-1958, S-Cubed, San Diego, CA, 1988.
7. Chau, A.S.Y.; Afghan, B.K. Analysis of Pesticides in Water, "Chlorine and Phosphorus-
Containing Pesticides," CRC Press, Boca Raton, FL, 1982, Vol. 2, pp 91-113, 238.
8. Hild, J.; Schulte, E; Thier, H.P. "Separation of Organophosphorus Pesticides and Their
Metabolites on Glass-Capillary Columns," Chromatographia, 1978,11-17.
9. Luke, M.A.; Froberg, J.E.; Doose, G.M.; Masumoto, H.T. "Improved Multiresidue Gas
Chromatographic Determination of Organophosphorus, Organonitrogen, and Organohalogen
Pesticides in Produce, Using Flame Photometric and Electrolytic Conductivity Detectors," J.
Assoc. Off. Anal. Chem. 1981,1187,64.
10. Sherma, J.; Berzoa, M. "Analysis of Pesticide Residues in Human and Environmental
Samples," U.S. Environmental Protection Agency, Research Triangle Park, NC, EPA-600/8-80-
038.
11. Desmarchelier, J.M.; Wustner, D.A.; Fukuto, T.R, "Mass Spectra of Organophosphorus Esters
and Their Alteration Products," Residue Reviews, 1974, pp 63, 77.
8141B-16
Revision 2
January 1998
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12. Munch, D.J. and Frebis, C.P., "Analyte Stability Studies Conducted during the National
Pesticide Survey," ES & T, 1992, vol 26,921-925.
13. T.L. Jones, "Organophosphorus Pesticide Standards: Stability Study," EMSL-LV Research
Report, EPA 600/X-92/040, April, 1992
14. Kotronarou, A., et al., "Decomposition of Parathion in Aqueous Solution by Ultrasonic
Irradiation," ES&T, 1992, Vol. 26,1460-1462.
15. Richter, B, Ezzell, J., and Felix D, "Single Laboratory Method Validation Report - Extraction of
Organophosphorous Pesticides, Herbicides and Polychlorinated Biphenyls using Accelerated
Solvent Extraction (ASE) with Analytical Validation by GC/NPD and GC/ECD," Dionex, Salt
Lake City, UT, Document 101124, December 2,1994.
8141B -17
Revision 2
January 1998
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TABLE 1
METHOD DETECTION LIMITS IN A WATER AND A SOIL MATRIX
USING 15-m COLUMNS AND A FLAME PHOTOMETRIC DETECTOR
Compound
Reagent Water1 (pg/L)
Soil5 (jig/kg)
Azinphos-methyl
0.10
5.0
Bolstar (Sulprofos)
0.07
3.5
Chlorpyrifos
0.07
5.0
Coumaphos
0.20
10.0
Demeton, -O, -S
0.12
6.0
Diazinon
0.20
10.0
Dichlorvos (DDVP)
0.80
40.0
Dimethoate
0.26
13.0
Disulfoton
0.07
3.5
EPN
0.04
2.0
Ethoprop
0.20
10.0
Fensulfothion
0.08
4.0
Fenthion
0.08
5.0
Malathion
0.11
5.5
Merphos
0.20
10.0
Mevinphos
0.50
25.0
Naled
0.50
25.0
Parathion, ethyl
0.06
3.0
Parathion, methyl
0.12
6.0
Phorate
0.04
2.0
Ronnel
0.07
3.5
Sulfotepp
0.07
" ... 3.5
TEPP®
0.80
40.0
T etrachlorovinphos
0.80
40.0
Tokuthion (Protothiofos)0
0.07
5.5
Trichloronate0
0.80
40.0
"Sample extracted using Method 3510, Separatory Funnel Liquid-Liquid Extraction.
"Sample extracted using Method 3540, Soxhlet Extraction.
"Purity of these standards not established by the EPA Pesticides and Industrial Chemicals
Repository, Research Triangle Park, NC.
8141B-18
Revision 2
January 1998
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TABLE 2
DETERMINATION OF ESTIMATED QUANTITATION LIMITS (EQLs)
FOR VARIOUS MATRICES*
Matrix
Factor
Ground water (Methods 3510 or 3520)
Low-concentration soil by Soxhlet and no cleanup
Non-water miscible waste (Method 3580)
10b
10°
10006
" EQL = [Method detection limit (see Table 1)] X [Factor found in Ms table]. For non-aqueous
samples, the factor is cm a wet-weight basis. Sample EQLs are higl% matrix dependent. The
EQLs to be determined herein are for guidance and may not always be achievable.
b Multiply this factor times the reagent water MDL in Table 1.
c Multiply this factor times the soil MDL in Table 1.
8141B-19
Revision 2
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TABLE 3
RETENTION TIMES ON 15-m COLUMNS
Retention Time (min)
Analyte
DB-5
SPB-608
DB-210
TEPP

6.44
5.12
Dichlorvos (DDVP)
9.63
7.91
12.79
Mevinphos
14.18
12.88
18.44
Demeton, -0 and -S
18.31
15.90
17.24
Ethoprop
18.62
16.48
18.67
Naled

19.01
17.40
Phorate
19.94
17.52
18.19
Monochrotophos
20.04
20.11
31.42
Sulfotepp
20.11
18.02
19.58
Dimethoate
20.64
20.18
27.96
Disutfoton
23.71
19.96
20.66
Diazinon
24.27
20.02
19.68
Merphos
26.82
21.73
32.44
Ronnel
29.23
22.98
23.19
Chlorpyrifos
31.17
26.88
25.18
Maiathion
31.72
28.78
32.58
Parathion, methyl
31.84
23.71
32.17
Parathion, ethyl
31.85
27.62
33.39
Trichloronate
32.19
28.41
29.95
Tetrachlorovinphos
34.65
32.99
33.68
Tokuthion (Protothiofos)
34.67
24.58
39.91
Fensulfothion
35.85
35.20
36.80
Bolstar (Sulprofos)
36.34
35.08
37.55
Famphur*
36.40
36.93
37.86
EPN

37.80
36.71
Azinphos-methyl
38.34
38.04
37.24
Fenthion
38.83
29.45
28.86
Coumaphos
39.83
38.87
39.47
DB-1
10.66
19.35
36.74
®GC operating conditions are shown in Table 8.
"Method 8141 has not been fully validated for Famphur.
8141B - 20
Revision
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TABLE 4
RETENTION TIMES ON 30-m COLUMNS8
Retention Time (min)
Analyte
DB-5
DB-210
DB-608
DB-1
Trimethylphosphate
b
2.36

Dichiorvos (DDVP)
7.45
6.99
6.56
10.43
Hexamethylphosphoramide
b
7.97

Trichlorfon
11.22
11.63
12.69

TEPP
b
13.82

Thionazin
12.32
24.71

Mevinphos
12.20
10.82
11.85
14.45
Ethoprop
12.57
15.29
18.69
18.52
Diazinon
13.23
18.60
24.03
21.87
Sulfotepp
13.39
16.32
20.04
19.60
Terbufos
13.69
18.23
22.97

T ri-o-cresyl phosphate
13.69
18.23

Naled
14.18
15.85
18.92
18.78
Phorate
12.27
16.57
20.12
19.65
Fonophos
14.44
18.38

Disulfoton
14.74
18.84
23.89
21.73
Merphos
14.89
23.22

26.23
Oxidized Merphos
20.25
24.87
35.16

Dichtorofenthion
15.55
20.09
26.11

Chlorpyrifos, methyl
15.94
20.45
26.29

Ronnei
16.30
21.01
27.33
23.67
Chlorpyrifos
17.06
22.22
29.48
24.85
Trichloronate
17.29
22.73
30.44

Aspon
17.29
21.98
~

Fenthion
17.87
22.11
29.14
24.63
Demeton-S
11.10
14.86
21.40
20.18
Demeton-O
15.57
17.21
17.70

Monocrotophosc
19.08
15.98
19.62
19.3
Dimethoate
18.11
17.21
20.59
19.87
Tokuthion
19.29
24.77
33.30
27.63
Maiathion
19.83
21.75
28.87
24.57
Parathion, methyl
20.15
20.45
25.98
22.97
8141B
-21
Revision 2
January 1998
-------
TABLE 4
(continued)
Retention Time (min)
Arialyte
DB-5
DB-210
DB-608
DB-1
Fenithnothion
20.63
21.42

Chlorfenvinphos
21.07
23.66
32.05

Parathion, ethyl
21.38
22.22
29.29
24.82
Bolstar
22.09
27.57
38.10
29.53
Stirophos
22.06
24.63
33.40
26.90
Ethion
22.65
27.12
37.61

Phosphamidon
22.77
20.09
25.88

Crotoxyphos
22.77
23.85
32.65

Leptophos
24.62
31.32
44.32

Fensutfbthion
27.54
26.76
36.58
28.58
EPN
27.58
29.99
41.94
31.60
Phosmet
27.89
29.89
41.24

Azinphos-methyl
28.70
31.25
43.33
32.33
Azinphos-ethyl
29.27
32.36
45.55

Famphur
29.41
27.79
38.24

Coumaphos
33.22
33.64
48.02
34.82
Atrazine
13.98
17.63

Simazine
13.85
17.41

Carbophenothion
22.14
27.92

Dioxathion
d
d
22.24

Trithion methyl

36.62

Dicrotophos

19.33

Internal Standard

1 -Bromo-2-nitro benzene
8.11
9.07

Surrogates

Tributyf phosphate

11.1

Triphenyl phosphate

33.4

4-Chloro-3-nitrobenzotrffluoride
5.73
5.40

33S8MBI
*GC operating conditions are shown in Table 8.
b Not detected at 20 ng per injection.
c Retention times may shift to longer times with larger amounts ir^ected (shifts of over 30
seconds have been observed, see Reference 6).
" Shows multiple peaks; therefore, not included in the composite.
8141B - 22
Revision 2
January 1998
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TABLE 5
RECOVERY OF 27 ORGANOPHOSPHATES BY SEPARATORY FUNNEL EXTRACTION
Anatyte
Percent Recovery at Three Spiking Levels
Low
Medium
High
Aziriphos methyl
126
143 ±8
101
Bolstar
134
141 ±8
101
Chlorpyrifos
7
89 ±6
86
Coumaphos
103
90 ±6
96
Demeton
33
67 ±11
74
Diazinon
136
121 ±9.5
82
Dichlorvos
80
79 ±11
72
Dimethoate
NR
47 ± 3
101
Disuifoton
48
92 ±7
84
EPN
113
125 ±9
97
Ethoprop
82
90 ±6
80
Fensulfonthion
84
82 ± 12
96
Fenthion
NR
48 ±10
89
Malathion
127
92 ±6
86
Merphos
NR
79
81
Mevinphos
NR
NR
55
Monocrotophos
NR
18±4
NR
Naled
NR
NR
NR
Parathion, ethyl
101
94 ± 5
86
Parathion, methyl
NR
46±4
44
Phorate
94
77 ±6
73
Ronnel
67
97 ±5
87
Sulfotep
87
85 ±4
83
TEPP
96
55 ±72
63
Tetrachlorvinphos
79
90 ±7
80
Tokuthion
NR
45 ±3
90
Trichloroate
NR
35
4
NR = Not recovered
8141B - 23
Revision 2
January 1998
-------
TABLE 6
RECOVERY OF 27 ORGANOPHOSPHATES BY CONTINUOUS LIQUID-LIQUID EXTRACTION
Arialyte
Percent Recovery at Three Spiking Levels
Low
Medium
High
Azinphos methyl
NR
129
122
Bolstar
NR
126
128
Chlorpyrifos
13
82 ±4
88
Coumaphos
94
79 ±1
89
Demeton
38
23 ±3
41
Diazinon
NR
128 ±37
118
DicWorvos
81
32 ± 1
74
Dimethoate
NR
10 ±8
102
Disulfotort
94
69 ±5
81
EPN
NR
104 ± 18
119
Ethoprop
39
76 ±2
83
Famphur
—
63 ±15
—
Fensulforrthion
90
67 ±26
90
Fenthion
8
32 ±2
86
Malathion
105
87 ±4
86
Merphos
NR
80
79
Mevinphos
NR
87
49
Monocrotophos
NR
30
1
Naled
NR
NR
74
Paraihion, ethyl
106
81 ±1
87
Parathion, methyl
NR
50 ±30
43
Ph orate
84
63±3
74
Ronnel
82
83 ±7
89
Sulfotep
40
77 ± 1
85
XHPP
39
18±7
70
Tetrachlorvinphos
56
70 ±14
83
Tokuthion
132
32 ±14
90
Trichloroate
NR
NR
21
NR = No! recovered
8141B-24
Revision 2
January 1998
-------
TABLE 7
RECOVERY OF 27 ORGANOPHOSPHATES BY SOXHLET EXTRACTION
Analyte
Percent Recovery at Three Spiking Levels
Low
Medium
High
Azinphos methyl
156
110 ±6
87
Botstar
102
103 ± 15
79
Chlorpyrifos
NR
66 ± 17
79
Coumaphos
93
89 ± 11
90
Demeton
169
64 ± 6
75
Diazinon
87
96 ± 3
75
Dichlorvos
84
39 ±21
71
Dimethoate
NR
48 ±7
98
Disulfoton
78
78 ±6
76
EPN
114
93 ±8
82
Ethoprop
65
70 ± 7
75
Fensulfonthion
72
81 ±18
111
Fenthion
NR
43 ±7
89
Malathion
100
81 ±8
81
Merphos
62
53
60
Mevinphos
NR
71
63
Monocrotophos
NR
NR
NR
Naled
NR
48
NR
Parathion, ethyl
75
80 ±8
80
Parathion, methyl
NR
41 ±3
28
Phorate
75
77 ±6
78
Ronnel
NR
83 ±12
79
Sulfotep
67
72 ±8
78
TEPP
36
34 ±33
63
Tetrachlorvinphos
50
81 ±7
83
Tokuthion
NR
40 ±6
89
Trichloroate
56
53
53
NR - Not recovered
8141B - 25
Revision 2
January 1998
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TABLE 8
SUGGESTED OPERATING CONDITIONS FOR 15-m COLUMNS
Columns 1 and 2 fDB-210 and SPB-608 or their equivalents)
Carrier gas (He) flow rate SmL/min
Initial temperature 50°C, hold for 1 minute
Temperature program 50°C to 140°C at 5°C/min, hold for 10 minutes,
followed by 140°C to 240°C at 10°C/min, hold
for 10 minutes (or a sufficient amount of time
for last compound to elute).
Column 3 fPB-5 or equivalent)
Carrier gas (He) flow rate SmL/min
Initial temperature 130°C, hold for 3 minutes
Temperature program 130"C, to 180°C at 5'C/min, hold for 10
minutes, followed by 180X to 250'C at
2-C/min, hold for 15 minutes (or a sufficient
amount of time for last compound to elute).
8141B - 26
Revision 2
January 1998
-------
TABLE 9
SUGGESTED OPERATING CONDITIONS FOR 30-m COLUMNS
Column 1
Column 2
Carrier gas flow rate
Makeup gas flow rate
Temperature program
Injector temperature
Detector temperature
Injection volume
Solvent
Type of injector
Detector type
Range
Attenuation
Type of splitter
Data system
Hydrogen gas pressure
Bead temperature
Bias voltage
DB-210
Dimensions: 30-m x 0.53-mm ID
Film Thickness (Mm): 1.0
DB-5
Dimensions: 30-m x 0.53-mm ID
Film Thickness Qim): 1.5
6 (mL/min) Helium
20 (mL/min) Helium
120 °C (3-min hold) to 270 °C (10-min hold) at
5°C/min
250°C
300°C
2 (JL
Hexane
Flash vaporization
Dual NPD
1
64
Y-shaped or Tee
Integrator
20 psi
400°C
4
8141B - 27
Revision 2
January 1998
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TABLE 10
QUANTITATION AND CHARACTERISTIC IONS FOR GC/MS ANALYSIS
OF ORGANOPHOSPHORUS PESTICIDES
Analyte
Quantitation ion
Characteristic ions
Azlnphos-methyl
160
77,132
Bolstar (Sulprofos)
156
140,143,113,33
Chlorpyrifos
197
97,199,125.314
Coumaphos
109
97,226,362,21
Demeton-S
88
60,114,170
Diazinon
137
179,152,93,199,304
Dichlorvos (DDVP)
109
79,185,145
Dimethoate
87
93,125,58,143
Disulfoton
88
89,60,61,97,142
EPN
157
169,141,63,185
Ethoprop
158
43,97,41,126
Fensulfothion
293
97,125,141,109,308
Fenthion
278
125,109,93,169
Malathion
173
125,127,93,158
Merphos
209
57,153,41,298
Mevinphos
127
109,67,192
Monocrotophos
127
67,97,192,109
Naled
109
145,147,79
Parathion, ethyl
291
97,109,139,155
Parathion, methyl
109
125,263,79
Phorate
75
"121,97,47,260
Ronnel
285
125,287,79,109
Stirophos
109
329,331,79
Sulfotepp
322
97,65,93,121,202
TEPP
99
155,127,81,109
Tokuthion
113
43,162,267,309
8141B - 28
Revision 2
January 1998
-------
TABLE 11
PERFORMANCE DATA FOR ORGANOPHOSPHORUS PESTICIDES
IN GROUND WATER USING METHOD 3535
Ground Water spiked at Ground Water spiked at
250 ppb 10 ppb
Analyte
% Recovery
RSD
% Recovery
RSD
MDL*
Aspon
85.6
11.5
77.7
6.8
1.7
Azinphos-methyl
83.0
13.4
109.7
7.0
2.4
Azinphos-ethyi
88.3
10.8
92.8
8.1
2.4
Bolstar
96.1
4.2
78.2
4.3
1.1
Carfaophenothion
85.6
11.0
81.7
7.2
1.9
Chlorfenvinphos
87.8
10.2
90.1
6.0
1.7
CWorpyrifos
98.8
5.7
77.5
4.2
1.0
Chlorpyrifos methyl
82.5
12.0
59.4
7.5
1.4
Coumaphos
84.3
8.7
100.8
13.5
4.3
Crotoxyphos
86.3 -
10.5
89.4
5.9
1.7
Demeton
93.6
4.5
73.8
5.1
1.2
Diazinon
91.7
4.7
70.0
5.0
1.1
Dichlorofenthion
85.2
10.9
75.6
6.0
1.4
Dichlorvos (DDVP)
88.1
6.7
90.1
7.9
2.2
Dicrotophos
88.6
10.8
75.7
5.7
1.3
Dimethoate
99.3
1.8
76.7
9.5
2.3
Dioxathion
81.6
14.1
92.7
11.0
3.2
Disulfoton
93.2
7.6
79.5
6.1
1.5
EPN
73.8
10.6
67.9
7.9
1.7
Ethion
85.5
10.6
79.2
6.5
1.6
Ethoprop
95.6
4.1
81.4
3.7
0.9
Famphur
85.2
10.2
75.6
8.3
2.0
Fenitrothion
91.2
8.8
85.0
5.0
1.3
Fensulfothion
86.2
6.4
97.2
6.0
1.8
8141B - 20
Revision 2
January 1998
-------
TABLE I! (cont.)
Ground Water spiked at Ground Water spiked at
250 ppb 10 ppb
Analyte
% Recovery
RSD
% Recovery
RSD
MDLa
Fenthion
91.2
5.4
79.5
4.3
1.7
Fonophos
91.0
8.0
81.6
3.6
0.9
Leptophos
81.3
12.2
73.6
8.8
2.0
Malathion
79.5
6.9
78.0
8.7
2.1
Merphos
113.1
9.3
84.6
4.5
1.2
Mevinphos
57.9
6.9
96.8
6.7
2.0
Naled
90.1
6.7
88.1
7.9
2.2
Parathion, ethyl
76.7
9.6
69.6
8.1
1.8
Parathion, methyl
93.9
5.8
83.6
4.7
1.2
Phorate
92.3
7.1
70.8
6.7
1.5
Phosmet
66.1
17.7
90.3
10.7
3.0
Phosphamidon
86.2
11.2
80.6
5.7
1.4
Ronnel
94.7
5.2
77.8
4.7
1.2
Stirophos
78.6
13.1
106.3
5.9
2.0
Sulfotepp
7S.3
9.3
68.9
8.6
1.9
Terbufos
87.1
10.5
78.0
3.7
0.9
Thionazin
95.1
8.0
88.6
3.4
1.0
Tokuthion
94.4
4.1
77.8
5.6
1.4
Trichlorfon
72.7
13.5
45.6
6.9
1.0
Trichloronate
95.3
4.5
75.7
3.9
0.9
3 All MDL values are in pg/L, and are highly matrix dependant MDLs provided in SW-846 are for
guidance purposes and may not always be achievable. Laboratories should establish their own in-
house MDLs to document method performance.
8141B - 30
Revision 2
January 1998
-------
TABLE 12
PERFORMANCE DATA FOR ORGANOPHOSPHORUS PESTICIDES
IN WASTEWATER USING METHOD 3535
Wastewater spiked at Wastewater spited at
250 ppb 10 ppb
Analyte
% Recovery
RSD
% Recovery
RSD
MDL1
Aspon
83.7
1.8
76.3
6.7
1.6
Azinphos-methyi
102.6
18.0
129.9
12.4
5.1
Azinphos-ethyl
79.8
6.8
96.0
6.7
2.0
Bolstar
94.4
8.3
84.9
* 1.4
0.4
Cartoophenothion
82.4
2.9
82.1
6.7
1.7
Chlorfenvinphos
81.7
6.5
88.0
7.2
2.0
Chlorpyrifos
91.0
8.3
86.5
1.7
0.5
Chlorpyrifbs methyl
77.6
2.2
56.7
7.1
1.3
Coumaphos
100.2
17.2
111.0
8.5
3.0
Crotoxyphos
81.3
5.7
87.5
7.0
1.9
Demeton
95.8
5.3
88.5
5.0
1.4
Diazinon
91.8
6.5
82.4
3.2
0.8
Dichlorfenthion
82.5
1.4
76.2
5.5
1.3
Dichlorvos (DDVP)
60.6
11.1
99.7
6.1
1.9
Dicrotophos
82.0
1.6
73.4
6.1
1.4
Dimethoate
93.5
4.1.
115.7
6.7
2.4
Dioxathion
84.6
5.6
100.4
9.4
3.0
Disulfoton
92.5
5.3
90.4 "
2.6
0.7
EPN
78.1
9.6
80.1
8.6
2.2
Ethion
83.5
2.0
78.4
6.4
1.6
Ethoprop
96.3
4.7
92.9
3.1
0.9
Famphur
85.9
2.5
78.6
7.9
1.9
Fenitrothion
83.5
4.8
82.3
5.9
1.5
Ferisulfothion
101.7
11.4
110.5
6.5
2.3
8141B-31
Revision 2
January 1998
-------
TABLE 12
PERFORMANCE DATA FOR ORGANOPHOSPHORUS PESTICIDES
IN WASTEWATER USING METHOD 3535
(continued)
Wastewater spiked at Wastewater spiked at
250 ppb 10 ppb
Analyte
% Recovery
RSD
% Recovery
RSD
- MDL*
Fenthion
91.7
7.3
88.2
2.7
0.7
Fonophos
83.4
2.6
81.3
5.0
1.3
Leptophos
81.9
3.3
73.2
7.5
1.7
Malathion
94.8
6.7
94.7
5.5
1.6
Merphos
94.5
12.7
90.7
1.4
0.4
Mevinphos
62.6
11.2
109.0
4.8
1.6
Naled
60.6
11.1
99.7
6.1
1.9
Parathion ethyl
80.2
8.1
83.6
8.6
2.3
Parathion methyl
92.9
6.5
93.8
4.4
1.3
Phorate
92.4 -
6.4
85.6
2.4
0.6
Phosmet
63.5
8.2
101.3
9.1
2.9
Phosphamidon
81.1
3.1
78.0
5.7
1.4
Ronnel
91.4
8.4
88.3
2.2
0.6
Stirophos
101.4
14.3
126.5
6.5
2.6
Sulfotepp
78.7
10.7
87.9
8.8
2.4
Terbufos
83.0
1.5
80.1
6.4
1.6
Thionazin
85.1
5.8
84.8
4.9
1.3
Tokuthion
91.8
8.4
83.6
1.8
0.5
Trichlorfon
66.8
4.6
52.2
8.7
1.4
Trichloronate
91.3
8.1
84.3
1.6
0.4
8 All MDL values are in }jg/L, and are highly matrix dependant. MDLs provided in SW-846 are for
guidance purposes and may not always be achievable. Labs should establish their own in-house
MDLs to document method performance.
8141B - 32
Revision 2
January 1998
-------
TABLE 13
RECOVERIES OF ANALYTES FROM SPIKED SOIL SAMPLES
USING PRESSURIZED FLUID EXTRACTION (METHOD 3545)
Clay Loam Sand
Analyte
Low
High
Low
High
Low
High
Dichlorvos
100.0
280.0
135.1
158.5
103.0
230.2
Mevinphos
100.6
98.0
104.0
99.8
, 91.5
107.8
Demeton O&S
103.7
106.2
124.3
108.4
103.4
106.0
Ethoprop
97.4
95.8
101.2
97.2
90.0
98.4
TEPP
100.0
100.0
100.0
100.0
100.0
100.0
Phorate
98.8
96.6
104.6
98.5
92.6
100.0
Sulfotep
102.6
99.3
113.2
119.1
129.4
104.2
Naled
100.0
100.0
100.0
100.0
100.0
100.0
Diazinon
97.6
96.2
104.2
101.7
89.3
100.4
Disuifoton
121.8
86.8
112.0
92.5
76.9
90.7
Monocrotophos
100.0
100.0
100.0
100.0
100.0
100.0
Dimethoate
92.5
90.7
94.3
89.0
88.5
101.8
Ronnel
96.4
95.3
102.4
85.0
94.1
98.7
Chlorpyrifos
98.3
96.3
98.0
97.0
90.2
100.2
Parathion methyl
94.9
97.7
98.9
98.5
91.3
98.3
Parathion ethyl
95.4
97.4
99.1
99.5
87.9
98.2
Fenthion
95.9
96.5
104.0
100.4
83.1
99.2
Tokuthion
97.1
95.8
102.4
96.5
94.2
98.2
Tetrachlorvinphos
93.8
93.8
144.0
92.3
95.2
101.9
Bolstar
99.1
98.4
105.1
97.5
96.6
102.5
Fensulfothion
100.0
89.9
81.2
76.8
90.1
102.1
EPN
85.8
97.3
88.8
97.1
92.8
104.8
Azinphos-methyl
100.0
92.4
85.0
81.4
96.3
103.4
Coumaphos
100.0
94.0
85.1
90.6
102.6
109.8
Results are expressed as the percentage of amount determined by a Soxhlet extraction. Data from
Reference 15.
8141B - 33
Revision 2
January 1998
-------
TABLE 14
BIAS AND PRECISION OF PRESSURIZED FLUID EXTRACTION
(METHOD 3545) OF THREE SPIKED SOIL SAMPLES
Clay Loam Sand
Low High Low High Low High
Analyie
Bias
Pre
Bias
Pre
Bias
Pre
Bias
Pre
Bias
Pre
Bias
Pre
Dichlorvos
0.0 ^
NA
5.6
19.0
10.4
11.4
6.5
222
13.9
13.4
9.9
22.2
Mevinphos
66.1
3.8
67.2
4.8
57.3
11.2
63.1
6.5
61.6
14.3
64.7
12.1
Demeton O&S
79.0
3.4
80.2
42
73.7
10.0
77.6
6.4
60.0
12.5
77.6
12.7
Ethoprop
83.0
4.7
84.8
4.8
76.1
10.7
77.0
4.9
-75.5
12.8
79.0
10.6
TEPP
0.0
NA
0.0
NA
0.0
NA
0.0
NA
0.0
NA
0.0
NA
Phorate
67.5
3.2
79.4
5.1
63.4
11.8
73.5
5.4
62.9
13.6
762
10.8
Sulfotep
66.6
3.7
69.4
4.7
62.6
11.0
66.8
7.3
62.1
13.8
67.7
132
Naled
0.0
NA
0.0
NA
0.0
NA
0.0
NA
0.0
NA
0.0
NA
Diazinon
80.2
4.7
80.3
4.8
74.4
12.0
75.9
6.0
73.9
14.0
77.4
112
Disulfoton
55.9
3.6
93.9
4.7
58.9
11.8
89.4
6.2
522
15.3
88.5
12.3
Moriocrotophos
0.0
NA
0.0
NA
0.0
NA
0.0
NA
0.0
NA
0.0
NA
Dimethoate
87.0
5.0
86.7
5.3
70.7
12.1
71.7
18.8
75.0
13.1
80.6
12.5
Ronriel
81.3
3.7
81.1
5.0
73.1
11.1
64.7
6.5
69.0
13.6
73.8
11.6
Chlorpyrifos
99.5
3.1
99.0
5.1
81.7
14.1
87.7
16.8
84.1
13.1
91.6
12.7
Parathion methyl
82.5
3.9
84.5
5.2
74.4
11.5
79.6
5.8
74.9
132
80.3
11.3
Parathion ethyl
85.0
3.8
83.5
5.2
77.3
11.9
79.6
6.1
78.0
12.7
80.3
11.5
Fenthion
56.4
3.8
71.4
5.0
44.1
10.8
50.9
6.6
44.3
12.5
51.9
12.6
Tokuthion
96.1
4.7
97.0
5.7
932
12.2
93.8
6.1
812
12.5
85.4
11.9
Tetrachlorvinphos
72.1
3.3
69.7
5.6
101.4
12.6
64.7
6.5
69.3
11.9
69.6
13.0
Bolstar
89.0
3.4
109.5
6.8
822
9.9
89.2
6.2
77.3
11.7
94.2
12.8
Fensulfothion
0.0
NA
69.7
4.3
70.4
9.3
522
7.1
63.0
9.2
62.0
13.1
EPN
72.6
44.3
76.9
8.0
92.9
10.1
70.4
7.1
68.6
11.2
71.9
11.6
Azinphos-methyl
0.0
NA
90.6
5.3
69.7
13.9
70.5
8.7
94.5
12.5
82.5
11.4
Coumaphos
0.0
NA
79.6
4.8
62.8
13.4
6.5
10.2
74.8
16.1
72.9
92
NA = not applicable
Bias was calculated as the percent recoveiy of the certified spiking value. Precision was calculated as the
relative standard deviation (RSD). Total number of analyses of each sample was 7.
Data from Reference 15.
8141B - 34
Revision 2
January 1998
-------
FIGURE 1
Chromatogram of target organophosphorus compounds from a 15-m DB-210 column with FPD detector.
More compounds are shown in Figure 2. See Table 3 for retention times.
300.00
250.00
200.00
150.00
100.00
50.00
0.00
*
a
1
W.
o
%
E
0)
\.k
e&c
£ o
III
*s
c
c
o
a.
c
o
c
£
1

IM
CO
la.
S«
V... j
'A*
> « I I II I I M f I « I | I M | M I |l I I | < If pi I ]l M | Mt ) M I | t
1 3 5 7 9 tl 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45
8141B - 35
Revision 2
January 1998
-------
FIGURE 2
Chromatogram of target organophosphoms compounds from a 15-m DB-210 column with FPD
detector. More compounds are shown in Figure 1. See Table 3 for retention times.
300.00
250.00
200.00
150.00
100.00
50.00
0.00
ft

Jl
M
iuli
r<
s.
£
3
E
(A
O
n
Q.
c
II
1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 ^
8141B - 36
Revision 2
January 1998
-------
FIGURE 3
Chromatogram of target organophosphoais compounds from a 15-m DB-210 column with NPD
detector. More compounds are shown in Figure 4. See Table 3 for retention times.
tL
%
c
300.00
c.
250.00
j=
u.
100.00 -
50.00-
I 11 II | i rrjy11 (I I »p » ;
8141B-37
Revision 2
January 1998
-------
FIGURE 4
Chromatogram of taiget organophosphorus compounds from a 15-m DB-210 column with NPD detector.
More compounds are shown in Figure 3. See Table 3 for retention times.
moo
o
200.00
150.00
CL
DC
en6"
100.00
50.00
0.00
35 37 39 41 43 45
1 3 5 7 9 11
31
8141B - 38
Revision 2
January 1998
-------
FIGURE 5
Chromatogram of target organophosphorus compounds on a 30-m DB-5/DB-210 column pair with NPD
detector, without Simazine, Atrazine and Carbophenothion. See Table 4 for retention times and for GC
operating conditions.
J
DB-210
m
v__
8141B - 39
Revision 2
January 1998
-------
FIGURE 6
Chromatogram of target organophosphorus compounds on a 30-m DB-5/DB-21Q column pair with NPD
detector, with Simazine, Atrazine and Carfcophenothion. See Table 4 for retention times and for GC
operating conditions.
DB-210
8141B- 40
Revision 2
January 1998
-------
METHOD 8141B
ORGANOPHOSPHORUS COMPOUNDS BY GAS CHROMATOGRAPHY
<=>
7.1 Select appropriate
extraction and,
if necessary, cleanup
technique.
I
7.1.4 Perform
solvent exchange
during K-D
procedures in all
extraction methods.
1
r
72 Select GC
conditions.
1
r
7.3 Refer to Method
8000 for
calibration techniques.

f
7.4 Refer to Method 8000,
Sec. 7.6 for instructions
on analysis sequence,
dilutions, retention times,
and identification
criteria.

f
7.4.1 Inject sample.

1
7.5 Record sample
volume injected and
resulting peak sizes.
Determine identity and
quantity of each
component peak; refer
to Method 8000, Sec. 7.8
for calculation equations.
7.5.1 Reanalyze with an
FPD or perform
appropriate cleanup and
reanalyze.
7.5.1
Is peak
detection and
identification
prevented by
interfer-
ences?
7.5.2
Is sample
dilution
necessary?
7.5.2 Dilute and
reanalyze
8141B-41
Revision 2
January 1998
-------
4.3 DETERMINATION OF ORGANIC ANALYTES
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the
allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to
be used by the analyst and the regulated community in making judgements necessary to meet
the data quality objectives or needs for the intended use of the data.
4.3.2 GAS CHROMATOGRAPHIC/MASS SPECTROMETRIC METHODS
The following methods are included in this section:
Method 8260B:
Method 8270D:
Method 8275A:
Method 8280B:
Method 8290A:
Appendix A:
Volatile Organic Compounds by Gas Chromatography/Mass
Spectrometry (GC/MS)
Semivolatile Organic Compounds by Gas
Chromatography/Mass Spectrometry (GC/MS)
Semivolatile Organic Compounds (PAHs and PCBs) in
Soils/Sludges and Solid Wastes Using Thermal
Extraction/Gas Chromatography/Mass Spectrometry
(TE/GC/MS)
Polychlorinated Dibenzo-p-Dioxins and Polychlorinated
Dibenzofurans by High Resolution Gas Chromatography/Low
Resolution Mass Spectrometry (HRGC/LRMS)
Polychlorinated Dibenzodioxins (PCDDs) and Polychlorinated
Dibenzofurans (PCDFs) by High-Resolution Gas
Chromatography/High-Resolution Mass Spectrometry
(HRGC/HRMS)
Procedures for the Collection, Handling, Analysis, and
Reporting of Wipe Tests Performed within the
Laboratory
FOUR-11
Revision 4
January 1998
-------
METHOD 8270D
SEMI VOLATILE ORGANIC COMPOUNDS
BY GAS CHROMATOGRAPHY/MASS SPECTROMETRY fGC/MS)
1.0 SCOPE AND APPLICATION
1.1 Method 8270 is used to determine the concentration of semivolatile organic compounds
in extracts prepared from many types of solid waste matrices, soils, air sampling media and water
samples. Direct injection of a sample may be used in limited applications. The following compounds
can be determined by this method:
Appropriate Preparation Techniques"
3540/
Compounds
CAS No"
3510
3520 "
3541
3550
3580
Acenaphthene
83-32-9
X
X
X
X
X
Acenaphthylene
208-96-8
X
X
X
X
X
Acetophenone
98-86-2
X
ND
ND
ND
X
2-Acetyiaminofluorene
53-96-3
X
ND
ND
ND
X
1-Acetyl-2-thiourea
591-08-2
LR
ND
ND
ND
LR
Aldrin
309-00-2
X
X
X
X
X
2-Aminoanthraquinone
117-79-3
X
ND
ND
ND
X
Aminoazobenzene
60-09-3
X
ND
ND
ND
X
4-Aminobiphenyl
92-67-1
X
ND
ND
ND
X
3-Amino-9-ethyicarbazole
132-32-1
X
X
ND
ND
ND
Anilazine
101-05-3
X
ND
ND
ND
X
Aniline
62-53-3
X
X
ND
X
X
o-Anisidine
90-04-0
X
ND
ND
ND
X
Anthracene
120-12-7
X
X
X
X
X
Aramite
140-57-8
HS(43)
ND
ND
ND
X
Arocior 1016
12674-11-2
X
X
X
X
X
Aroclor 1221
11104-28-2
X
X
X
X
X
Arocior 1232
11141-16-5
X
X
X
X
X
Aroclor 1242
53469-21-9
X
X -
X
X
X
Arocior 1248
12672-29-6
X
X
X
X
X
Aroclor 1254
11097-69-1
X
X
X
X
X
Aroclor 1260
11096-82-5
X
X
X
X
X
Azinphos-methyl
86-50-0
HS{62)
ND
ND
ND
X
Barban
101-27-9
LR
ND
ND
ND
LR
Benzidine
92-87-5
CP
CP
CP
CP
CP
Benzoic acid
65-85-0
X
X
ND
X
X
ienz(a) anthracene
56-55-3
X
X
X
X
X
Benzo(b)fluoranthene
205-99-2
X
X
X
X
X
8270D -
1
Revision 4
January 1998
-------
Appropriate Preparation Techniques'1
3540/
Compounds
CAS No'
3510
3520
3541
3550
3580
Benzo(k)fluoranthene
207-08-9
X
X
X
X
X
Benzo(g,h,i)perylene
191-24-2
X
X
X
X
X
Benzo(a)pyrene
50-32-8
X
X
X
X
X
p-Benzoquinone
106-51-4
OE
ND
ND
ND
X
Benzyl alcohol
100-51-6
X
X
ND
X
X
a-BHC
319-84-6
X
X
X
X
X
p-BHC
319-85-7
X
X
X
X
X
5-BHC
319-86-8
X
X
X
X
X
V-BHC (Lindane)
58-89-9
X
X
X
X
X
Bis(2-chIoroethoxy)methane
111-91-1
X
X
X
X
X
Bis(2-chloroethyl) ether
111-44-4
X
X
X
X
X
Bis(2-chloroisopropyl) ether
108-60-1
X
X
X
X
X
Bis(2-ethythexyl) phthaiate
117-81-7
X
X
X
X
X
4-Bromophenyl phenyl ether
101-55-3
X
X
X
X
X
Bromoxynil
1689-84-5
X
ND
ND
ND
X
Butyl benzyl phthaiate
85-68-7
X
X
X
X
X
Captafol
2425-06-1
HS(55)
ND
ND
ND
X
Captart
133-06-2
HS(40)
ND
ND
ND
X
Carbaryi
63-25-2
X
ND
ND
ND
X
Carbofuran
1563-66-2
X
ND
ND
ND
X
Carbophenothion
786-19-6
X
ND
ND
ND
X
Chlordane (NOS)
57-74-9
X
X
X
X
X
Chlorfenvinphos
470-90-6
X
ND
ND
ND
X
4-Chloroaniline
106-47-8
X
ND
ND
ND
X
Chlorobenzilate
510-15-6
X
ND
ND
ND
X
5-Chloro2-methylaniline
95-79-4
X
ND
ND
ND
X
4-Chloro-3-methylphenol
59-50-7
X
X
X
X
X
3-(Chloromethyl)pyridine
6959-48-4
X
ND
ND
ND
X
hydrochloride

1 -Chloronaphthalene
90-13-1
X
X
" X
X
X
2-Chloronaphthalene
91-58-7
X
X
X
X
X
2-Chlorophenol
95-57-8
X
X
X
X
X
4-Chloro-1,2-phenylenediamine
95-83-0
X
X
ND
ND
ND
4-Chloro-1,3-phenylenediamjne
5131-60-2
X
X
ND
ND
ND
4-Chlorophenyl phenyl ether
7005-72-3
X
X
X
X
X
Chrysene
218-01-9
X
X
X
X
X
Coumaphos
56-72-4
X
ND
ND
ND
X
p-Cresidine
120-71-8
X
ND
ND
ND
X
Crotoxyphos
7700-17-6
X
ND
ND
ND
X
8270D-2
Revision 4
January 1998
-------
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-------
Appropriate Preparation Techniques'*
3540/
Compounds
CAS No8
3510
3520
3541
3550
3580
2,4-DMtrophenol
51-28-5
X
X
X
X
X
2,4-Dinitrotoluene
121-14-2
X
X
X
X
X
2,6-Dinitrotoluene
606-20-2
X
X
X
X
X
Dinocap
39300-45-3
CP,HS(28)
ND
ND
ND
CP
Dinoseb
88-85-7
X
ND
ND
ND
X
Diphenylamine
122-39-4
X
X
X
X
X
5,5- Diphenyl hyd a ntoin
57-41-0
X
ND
ND
ND
X
1,2-Diphenyihydrazine
122-66-7
X
X
X
X
X
Di-rvoctyl phthalate
117-84-0
X
X
X
X
X
Disuifoton
298-04-4
X
ND -
ND
ND
X
Endosulfan I
959-98-8
X
X
X
X
X
Endosulfan II
33213-65-9
X
X
X
X
X
Endosulfan sulfate
1031-07-8
X
X
X
X
X
Endrin
72-20-8
X
X
X
X
X
Endrin aldehyde
7421-93-4
X
X
X
X
X
Endrin ketone
53494-70-5
X
X
ND
X
X
EPN
**4t\A *A g
Mm 1 1W
X
ND
ND
ND
X
Ethion
563-12-2
X
ND
ND
ND
X
Ethyl carbamate
51-79-6
DC(28)
ND
ND
ND
X
Ethyl methanesulfonate
62-50-0
X
ND
ND
ND
X
Famphur
52-85-7
X
ND
ND
ND
X
Fensulfothion
115-90-2
X
ND
ND
ND
X
Fenthion
55-38-9
X
ND
ND
ND
X
Fluchloralin
33245-39-5
X
ND
ND
ND
X
Fluoranthene
206-44-0
X
X
X
X
X
Fluorene
86-73-7
X
X
X
X
X
2-Fluorobiphenyt (surr)
321-60-8
X
X
X
X
X
2-Fluorophenol (surr)
367-12-4
X
X
X
X
X
Heptachlor
76-44-8
X
X
X
X
X
Heptachlor epoxide
1024-57-3
X
X
X
X
X
Hexachlorobenzene
118-74-1
X
X
X
X
X
Hexachlorobutadiene
87-68-3
X
X
X
X
X
Hexachlorocyclopentadiene
77-47-4
X
X
X
X
X
Hexachloroethane
67-72-1
X
X
X
X
X
Hexachlorophene
70-30-4
AW,CP(62)
ND
ND
ND
CP
Hexachloropropene
1888-71-7
X
ND
ND
ND
X
Hexamethylphosphoramide
680-31-9
X
ND
ND
ND
X
Hydroquinone
123-31-9
ND
ND
ND
ND
X
lndeno(1,2,3-cd)pyrene
193-39-5
X
X
X
X
X
8270D-
4
Revision 4
January 1998
-------
Appropriate Preparation Techniques'8
3540/
Compounds
CAS No"
3510
3520
3541
3550
3580
Isodrin
465-73-6
X
ND
ND
ND
X
Isophorone
78-59-1
X
X
X
X
X
Isosafrole
120-58-1
DC(46)
ND
ND
ND
X
Kepone
143-50-0
X
ND
ND
ND
X
Leptophos
21609-90-5
X
ND
ND
ND
X
Malathion
121-75-5
HS{5)
ND
ND
ND
X
Maleic anhydride
108-31-6
HE
ND
ND
ND
X
Mestranot
72-33-3
X
ND
ND
ND
X
Methapyrilene
91-80-5
X
ND
ND
ND
X
Methoxychlor
72-43-5
X
ND -
ND
ND
X
3-Methyfcholanthrene
56-49-5
X
ND
ND
ND
X
4,4'-Methylenebis (2-chloroaniline)
101-14-4
OE,OS(0)
ND
ND
ND
LR
4,4'-Methyienebis(N,N-dimethyl-
101-61-1
X
X
ND
ND
ND
anifine)

Methyl methanesulfonate
66-27-3
X
ND
ND
ND
X
2-Methylnaphthaiene
91-57-6
X
X
ND
X
X
Methyl parathion
298-00-0
X
ND
ND
ND
X
2-Methyiphenol
95-48-7
X
ND
ND
ND
X
3-Methylphenol
108-39-4
X
ND
ND
ND
X
4-Methylphenol
106-44-5
X
ND
ND
ND
X
Mevinphos
7786-34-7
X
ND
ND
ND
X
Mexacarbate
315-18-4
HE,HS(68)
ND
ND
ND
X
Mirex
2385-85-5
X
ND
ND
ND
X
Monocrotophos
6923-22-4
HE
ND
ND
ND
X
Naled
300-76-5
X
ND
ND
ND
X
Naphthalene
91-20-3
X
X
X
X
X
1,4-Naphthoquinone
130-15-4
X
ND
ND
ND
X
1-Naphthylamine
134-32-7
OS(44)
ND
ND
ND
X
2-Naphthyiamine
91-59-8
X
ND
ND
ND
X
Nicotine
54-11-5
DE(67)
ND
ND
ND
X
5-Nitroacenaphthene
602-87-9
X
ND
ND
ND
X
2-Nitroaniline
88-74-4
X
X
ND
X
X
3-Nitroaniline
99-09-2
X
X
ND
X
X
4-Nitroaniline
100-01-6
X
X
ND
X
X
5-Nitro-o-anisidine
99-59-2
X
ND
ND
ND
X
Nitrobenzene
98-95-3
X
X
X
X
X
4-Nitrobiphenyl
92-93-3
X
ND
ND
ND
X
Nitrofen
1836-75-5
X
ND
ND
ND
X
2-Nitro phenol
88-75-5
X
X
X
X
X
8270D -
5
Revision 4
January 1998
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Appropriate Preparation Techniques6
3540/
Compounds
CAS No"
3510
3520
3541
3550
3580
4-Nitrophenol
100-02-7
X
X
X
X
X
5-Nitro-o-toluidine
99-55-8
X
X
ND
ND
X
Nitroquinoline-1-oxide
56-57-5
X
ND
ND
ND
X
N-Nitrosodi-n-butyiamine
924-16-3
X
ND
ND
ND
X
N-Nitrosodiethylamine
55-18-5
X
ND
ND
ND
X
N-Nitrosodimethylamine
62-75-9
X
X
X
X
X
N-Nitrosomethylethylamine
10595-95-6
X
ND
ND
ND
X
N-Nitrosodiphenylamine
86-30-6
X
X
X
X
X
N-Nitrosodi-n-propyiami ne
621-64-7
X
X
X
X
X
N-Nitrosomorpholine
59-89-2
ND
ND -
ND
ND
X
N- Nitrosopiperidi ne
100-75-4
X
ND
ND
ND
X
N-Nitrosopyrrolidine
930-55-2
X
ND
ND
ND
X
Octamethyi pyrophosphoramide
152-16-9
LR
ND
ND
ND
LR
4,4'-OxydianiIine
101-80-4
X
ND
ND
ND
X
Parathion
56-38-2
X
X
ND
ND
X
Pentachlorobenzene
608-93-5
X
ND
ND
ND
X
Pentach loronitrobenzene
82-68-8
X
ND
ND
ND
X
Pentachlorophenol
87-86-5
X
X
X
X
X
Phenacetin
62-44-2
X
ND
ND
ND
X
Phenanthrene
85-01-8
X
X
X
X
X
Phenobarbital
50-06-6
X
ND
ND
ND
X
Phenol
108-95-2
DC<28)
X
X
X
X
1,4-Phenylenediamine
106-50-3
X
ND
ND
ND
X
Phorate
298-02-2
X
ND
ND
ND
X
Phosalone
2310-17-0
HS(65)
ND
ND
ND
X
Phosmet
732-11-6
HS(15)
ND
ND
ND
X
Phosphamidon
13171-21-6
HE(63)
ND
ND
ND
X
Phthalic anhydride
85-44-9
CP,HE(1)
ND
ND
ND
CP
2-Picoline (2-Methylpyridine)
109-06-8
X
X
ND
ND
ND
Piperonyl sulfoxide
120-62-7
X
ND
ND
ND
X
Pronamide
23950-58-5
X
ND
ND
ND
X
Propylthiouracil
51-52-5
LR
ND
ND
ND
LR
Pyrene
129-00-0
X
X
X
X
X
Resorcinol
108-46-3
DC,OE(10)
ND
ND
ND
X
Safrofe
94-59-7
X
ND
ND
ND
X
Strychnine
57-24-9
AW,0S(55)
ND
ND
ND
X
Sulfallate
95-06-7
X
ND
ND
ND
X
Terbufos
13071-79-9
X
ND
ND
ND
X
1,2,4,5-T etrachlorobenzene
95-94-3
X
ND
ND
ND
X
8270D-6
Revision 4
January 1998
-------
Appropriate Preparation Techniques'1
3540/
Compounds
CAS No®
3510
3520
3541
3550
3580
2,3,4,6-Tetrachlorophenol
58-90-2
X
ND
ND
ND
X
Tetrachlorvinphos
961-11-5
X
ND
ND
ND
X
Tetraethyl dithiopyrophosphate
3689-24-5
X
X
ND
ND
ND
Tetraethyi pyrophosphate
107-49-3
X
ND
ND
ND
X
Thionazine
297-97-2
X
ND
ND
ND
X
Thiophenol (Benzenethiol)
108-98-5
X
ND
ND
ND
X
Toluene diisocyanate
584-84-9
HE(6)
ND
ND
ND
X
o-Toiuidine
95-53-4
X
ND
ND
ND
X
Toxaphene
8001-35-2
X
X
X
X
X
1,2,4-T richlorobenzene
120-82-1
X
X
- X
X
X
2,4,5-T richlorophenol
95-95-4
X
X
ND
X
X
2,4,6-T richlorophenol
88-06-2
X
X
X
X
X
Trifluralin
1582-09-8
X
ND
ND
ND
X
2,4,5-Trimethylaniline
137-17-7
X
ND
ND
ND
X
Trim ethyl phosphate
512-56-1
HE(60)
ND
ND
ND
X
1,3,5-T rinitrobenzene
99-35-4
X
ND
ND
ND
X
Tris(2,3-dibromopropyI) phosphate
126-72-7
X
ND
ND
ND
LR
Tri-p-tolyl phosphate
78-32-0
X
ND
ND
ND
X
0,0,0-Triethyl phosphorothioate
126-68-1
X
ND
ND
ND
X
3 Chemical Abstract Service Registry Number
b See Sec. 1.2 for other acceptable preparation methods.
KEY TO ANALYTE LIST
AW = Adsorption to walls of glassware during extraction and storage.
CP = Nonreproducible chromatographic performance.
DC = Unfavorable distribution coefficient (number in parenthesis is percent recovery).
HE = Hydrolysis during extraction accelerated by acidic or basic conditions (number in
parenthesis is percent recovery).
HS = Hydrolysis during storage (number in parenthesis is percent stability).
LR = Low response.
ND = Not determined.
OE = Oxidation during extraction accelerated by basic conditions (number in parenthesis is
percent recovery).
OS = Oxidation during storage (number in parenthesis is percent stability).
X = Greater than TO percent recovery by this technique.
1.2 In addition to the sample preparation methods listed in the above anaiyte list, Method
3535 describes a solid-phase extraction procedure that may be applied to the extraction of
semivolatiles from TCLP leachates (Tables 16 and 17 contain performance data). Method 3542
8270D - 7
Revision 4
January 1998
-------
describes sample preparation for semivolatile organic compounds in air sampled by Method 0010
(Table 11 contains surrogate performance data), Method 3545 describes an automated solvent
extraction device for semivolatiles in solids (Table 12 contains performance data), and Method 3561
describes a supercritical fluid device for the extraction of PAHs from solids (see Tables 13,14, and
15 for performance data).
1.3 Method 8270 can be used to quantitate most neutral, acidic, and basic organic
compounds that are soluble in methylene chloride and capable of being eluted, without
derivatization, as shaip peaks from a gas chromatographic fused-silica capillary column coated with
a slightly polar silicone. Such compounds include poiynuclear aromatic hydrocarbons, chlorinated
hydrocarbons and pesticides, phthalate esters, organophosphate esters, nitrosamines, haloethers,
aldehydes, ethers, ketones, anilines, pyridines, quinoiines, aromatic nitro compounds, and phenols,
including nitrophenols. See Table 1 for a list of compounds and their characteristic ions that have
been evaluated.
In most cases, Method 8270 is not appropriate for the quantitation of multicomponent analytes,
e.g., Aroclors, Toxaphene, Chlordane, etc., because of limited sensitivity for those analytes. When
these analytes have been identified by another technique, Method 8270 may be appropriate for
confirmation of the identification of these analytes when concentration in the extract permits. Refer
to Sec. 7.0 of Methods 8081 and 8082 for guidance on calibration and quantitation of
multicomponent analytes such as the Aroclors, Toxaphene, and Chlordane.
1.4 The following compounds may require special treatment when being determined by this
method:
1.4.1 Benzidine may be subject to oxidative losses during solvent concentration and
its chromatographic behavior is poor.
1.4.2 Under the alkaline conditions of the extraction step from aqueous matrices,
a-BHC, y-BHC, Endosulfan I and If, and Endrin are subject to decomposition. Neutral
extraction should be performed if these compounds are expected.
1.4.3 Hexachlorocyclopentadiene is subject to thermal decomposition in the inlet
of the gas chromatograph, chemical reaction in acetone solution, and photochemical
decomposition.
1.4.4 N-nitrosodimethylamine is difficult to separate from the solvent under the
chromatographic conditions described.
1.4.5 N-nitrosodiphenylamine decomposes in the gas chromatographic inlet and
cannot be separated from diphenylamine.
1.4.6 Pentachlorophenol, 2,4-dinitrophenol, 4-nitrophenol, benzoic acid,
4,6-dinitro-2-methylphenoI, 4-chloro-3-methylphenol, 2-nitroaniline, 3-nitroaniline,
4-chloroaniline, and benzyl alcohol are subject to erratic chromatographic behavior, especially
if the GC system is contaminated with high boiling material.
1.4.7 Pyridine may perform poorly at the GC injection port temperatures listed in the
method. Lowering the injection port temperature may reduce the amount of degradation.
However, the analyst must use caution in modifying the injection port temperature, as the
performance of other analytes may be adversely affected. Therefore, if pyridine is to be
determined in addition to other target analytes, it may be necessary to perform separate
8270D-8
Revision 4
January 1SS8
-------
analyses. In addition, pyridine may be lost during the evaporative concentration of the sample
extract. As a result, many of the extraction methods listed above may yield low recoveries
unless great care is exercised during the concentration steps. For this reason, analysts may
wish to consider the use of extraction techniques such as pressurized fluid extraction (Method
3545) or supercritical fluid extraction, which involve smaller extract volumes, thereby reducing
or eliminating the need for evaporative concentration techniques for many applications.
1.4.8 Toluene diisocyanate rapidly hydrolyses in water (half-life of less then 30
min.). Therefore, recoveries of this compound from aqueous matrices should not be expected.
In addition, in solid matrices, toluene diisocyanate often reacts with alcohols and amines to
produce urethane and ureas and consequently cannot usually coexist in a solution containing
these materials.
1.4.9 In addition, analytes in the list provided above are flagged when there are
limitations caused by sample preparation and/or chromatographic problems.
1.5 The estimated quantitation limit (EQL) of Method 8270 for determining an individual
compound is approximately 660 pg/kg (wet weight) for soil/sediment samples, 1-200 mg/kg for
wastes (dependent on matrix and method of preparation), and 10 }jg/L for ground water samples
(see Table 2). EQLs will be proportionately higher for sample extracts that require dilution to avoid
saturation of the detector.
1.6 This method is restricted to use by or under the supervision of analysts experienced
in the use of gas chromatograph/mass spectrometers and skilled in the interpretation of mass
spectra. Each analyst must demonstrate the ability to generate acceptable results with this method.
2.0 SUMMARY OF METHOD
2.1 The samples are prepared for analysis by gas chromatography/mass spectrometry
(GC/MS) using the appropriate sample preparation (refer to Method 3500) and, if necessary, sample
cleanup procedures (refer to Method 3600).
2.2 The semivolatile compounds are introduced into the GC/MS by injecting the sample
extract into a gas chromatograph (GC) with a narrow-bore fused-silica capillary column. The GC
column is temperature-programmed to separate the analytes, which are then detected with a mass
spectrometer (MS) connected to the gas chromatograph.
2.3 Analytes eluted from the capillary column are introduced into the mass spectrometer
via a jet separator or a direct connection. Identification of target analytes is accomplished by
comparing their mass spectre with the electron impact (or electron impact-like) spectra of authentic
standards. Quantitation is accomplished by comparing the response of a major (quantitation) ton
relative to an internal standard using a five-point calibration curve.
2.4 The method includes specific calibration and quality control steps that supersede the
general requirements provided in Method 8000.
8270D-9
Revision 4
January 1998
-------
3.0 INTERFERENCES
3.1 Raw GC/MS data from all blanks, samples, and spikes must be evaluated for
interferences. Determine if the source of interference is in the preparation and/or cleanup of the
samples and take connective action to eliminate the problem.
3.2 Contamination by carryover can occur whenever high-concentration and
low-concentration samples are sequentially analyzed. To reduce carryover, the sample syringe must
be rinsed with solvent between sample injections. Whenever an unusually concentrated sample is
encountered, it should be followed by the analysis of solvent to check for cross-contamination.
4.0 APPARATUS AND MATERIALS
4.1 Gas chromatograph/mass spectrometer system
4.1.1 Gas chromatograph - An analytical system complete with a
temperature-programmable gas chromatograph suitable for splitless injection and all required
accessories, including syringes, analytical columns, and gases. The capillary column should
be directly coupled to the source.
4.1.2 Column - 30-m x 0.25-mm ID (or 0.32-mm ID) 1-pm film thickness
silicone-coated fused-silica capillary column (J&W Scientific DB-5 or equivalent).
4.1.3 Mass spectrometer
4.1.3.1 Capable of scanning from 35 to 500 amu every 1 sec or less,
using 70 volts (nominal) electron energy in the electron impact ionization mode. The
mass spectrometer must be capable of producing a mass spectrum for
decafluorotriphenylphosphine (DFTPP) which meets the criteria in Table 3 when 1 pL
of the GC/MS tuning standard is injected through the GC (50 ng of DFTPP).
4.1.3.2 An ion trap mass spectrometer may be used if it is capable of axial
modulation to reduce ion-molecule reactions and can produce electron impact-like
spectra that match those in the EPA/NIST Library. The mass spectrometer must be
capable of producing a mass spectrum for DFTPP which meets the criteria in Table 3
when 5 or 50 ng are introduced.
4.1.4 GC/MS interface - Any GOto-MS interface may be used that gives acceptable
calibration points at 50 ng per injection for each compound of interest and achieves acceptable
tuning performance criteria. For a narrow-bore capillary column, the interface is usually
capillary-direct into the mass spectrometer source.
4.1.5 Data system - A computer system should be interfaced to the mass
spectrometer. The system must allow the continuous acquisition and storage on
machine-readable media of all mass spectra obtained throughout the duration of the
chromatographic program. The computer should have software that can search any GC/MS
date file for ions of a specific mass and that can plot such ion abundances versus time or scan
number. This type of plot is defined as an Extracted Ion Current Profile (EICP). Software
should also be available that allows integrating the abundances in any EICP between specified
time or scan-number limits. The most recent version of the EPA/NIST Mass Spectra! Library
should also be available.
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4,1.6 Guard column (optional) - (J&W deactivated fused-silica, 0.25-mm ID x 6-m,
or equivalent) between the injection port and the analytical column joined with column joiners
(Hewlett-Packard Catalog No. 5062-3556, or equivalent).
4.2 Syringe - 10-pL.
4.3 Volumetric flasks, Class A - Appropriate sizes with ground-glass stoppers.
4.4 Balance - Analytical, capable of weighing 0.0001 g.
4.5 Bottles - glass with polytetrafluoroethylene (PTFE)-lined screw caps or crimp tops.
5.0 REAGENTS
5.1 Reagent grade inorganic chemicals shall be used in all tests. Unless otherwise
indicated, it is intended that all reagents shall conform to the specifications of the Committee on
Analytical Reagents of the American Chemical Society, where such specifications are available.
Other grades may be used, provided it is first ascertained that the reagent is of sufficiently high purity
to permit its use without lessening the accuracy of the determination.
5.2 Organic-free reagent water - All references to water in this method refer to organic-free
reagent water, as defined in Chapter One.
5.3 Stock standard solutions (1000 mg/L) - Standard solutions can be prepared from pure
standard materials or purchased as certified solutions.
5.3.1 Prepare stock standard solutions by accurately weighing about 0.0100 g of
pure material. Dissolve the material in pesticide quality acetone or other suitable solvent and
dilute to volume in a 10-mL volumetric flask. Larger volumes can be used at the convenience
of the analyst. When compound purity is assayed to be 96% or greater, the weight may be
used without correction to calculate the concentration of the stock standard. Commercially-
prepared stock standards may be used at any concentration if they are certified by the
manufacturer or by an independent source.
5.3.2 Transfer the stock standard solutions into bottles with PTFE-lined screw-caps.
Store, protected from light, at -10°C or less or as recommended by the standard manufacturer.
Stock standard solutions should be checked frequently for signs of degradation or evaporation,
especially just prior to preparing calibration standards from them.
5.3.3 Stock standard solutions must be replaced after 1 year or sooner if
comparison with quality control check samples indicates a problem.
5.3.4 It is recommended that nitrosamine compounds be placed together in a
separate calibration mix and not combined with other calibration mixes. When using a
premixed certified standard, consult the manufacturer's instructions for additional guidance.
5.3.5 Mixes with hydrochloride salts may contain hydrochloric add, which can cause
analytical difficulties. When using a premixed certified standard, consult the manufacturer's
instructions for additional guidance.
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5.4 Internal standard solutions - The internal standards recommended are 1,4-dichloro-
benzene-d* naphthalene-dgl acenaphthene-d 10l phenanthrene-d 10, chrysene-d 12l and pery1ene-d12
(see Table 5). Other compounds may be used as internal standards as long as the specifications
in Sec. 7.3.2 are met.
5.4.1 Dissolve 0.200 g of each compound with a small volume of carbon disulfide.
Transfer to a 50 mL volumetric flask and dilute to volume with methylene chloride so that the
final solvent is approximately 20% carbon disulfide. Most of the compounds are also soluble
in small volumes of methanol, acetone, or toluene, except for perylene-d12. The resulting
solution will contain each standard at a concentration of 4,000 ng/^L. Each 1-mL sample
extract undergoing analysis should be spiked with 10 pL of the internal standard solution,
resulting in a concentration of 40 ng/^L of each internal standard. Store at -10°C or less when
not in use. When using premixed certified solutions, store according to the manufacturer's
documented holding time and storage temperature recommendations.
5.4.2 If a more sensitive mass spectrometer is employed to achieve lower detection
levels, a more dilute internal standard solution may be required. Area counts of the internal
standard peaks should be between 50-200% of the area of the target analytes in the mid-point
calibration analysis.
5.5 GC/MS tuning standard - A methylene chloride solution containing 50 ng/pL of
decafluorotriphenylphosphine (DFTPP) should be prepared. The standard should also contain 50
ng/pL each of 4,4-DOT, pentachlorophenol, and benzidine to verify injection port inertness and GC
column performance. Store at -10°C or less when not in use. If a more sensitive mass
spectrometer is employed to achieve lower detection levels, a more dilute tuning solution may be
necessary. When using premixed certified solutions, store according to the manufacturer's
documented holding time and storage temperature recommendations.
5.6 Caltoration standards - A minimum of five calibration standards should be prepared at
five different concentrations. At least one of ttie calibration standards should correspond to a sample
concentration at or below that necessary to meet the data quality objectives of the project. The
remaining standards should correspond to the range of concentrations found in actual samples but
should not exceed the working range of the GC/MS system. Each standard should contain each
analyte for detection by this method.
5.6.1 It is the intent of EPA that all target analytes for a particular analysis be
included in the calibration standard(s). These target analytes may not include the entire list
of analytes (Sec. 1.1) for which the method has been demonstrated. However, ttie laboratory
shall not report a quantitative result for a target analyte that was not included in the calibration
standard(s).
5.6.2 Each 1-mL alquot of calibration standard should be spiked with 10 yL of the
internal standard solution prior to analysis. All standards should be stored at -10°C or less,
and should be freshly prepared once a year, or sooner if check standards indicate a problem.
The calibration verification standard should be prepared weekly and stored at 4°C. When
using premixed certified solutions, store according to the manufacturer's documented holding
time and storage temperature recommendations.
5.7 Surrogate standards - The recommended surrogates are phenoI-d6, 2-ffuorophenol,
2,4,6-tribromophenol, nitrobenzene-d5, 2-fluorobiphenyl, and p-terphenyl-d14. See Method 3500 for
instructions on preparing the surrogate solutions.
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5.7.1 Surrogate standard check - Determine what the appropriate concentration
should be for the blank extracts after all extraction, cleanup, and concentration steps. Inject
this concentration into the GC/MS to determine recovery of surrogate standards. It is
recommended that this check be done whenever a new surrogate spiking solution is prepared.
NOTE: Method 3561 (SFE Extraction of PAHs) recommends the use of bromobenzene and
p-quaterphenyl to better cover the range of PAHs listed in the method.
5.7.2 If a more sensitive mass spectrometer is employed to achieve lower detection
levels, a more dilute surrogate solution may be necessary.
5.8 Matrix spike and laboratory control standards - See Method 3500 for instructions on
preparing the matrix spike standard. The same standard may be used as the laboratory control
standard (LCS).
5.8.1 Matrix spike check - Determine what concentration should be in the blank
extracts after all extraction, cleanup, and concentration steps. Inject this concentration into the
GC/MS to determine recovery, it is recommended that this check be done whenever a new
matrix spiking solution is prepared.
5.8.2 If a more sensitive mass spectrometer is employed to achieve lower detection
levels, a more dilute matrix and LCS spiking solution may be necessary.
5.8.3 Some projects may require the spiking of the specific compounds of interest,
since the spiking compounds listed in Method 3500 would not be representative of the
compounds of interest required for the project. When this occurs, the matrix and LCS spiking
standards should be prepared in methanol, with each compound present at a concentration
appropriate for the project.
5.9 Solvents - Acetone, hexane, methylene chloride, isooctane, carton disulfide, toluene,
and other appropriate solvents. All solvents should be pesticide quality or equivalent.
6.0 SAMPLE COLLECTION, PRESERVATION, AND HANDLING
6.1 See the introductory material to this chapter, Organic Analytes, Sec. 4.1.
6.2 Store the sample extracts at -10°C, protected from light, in sealed vials (e.g., screw-cap
vials or crimp-capped vials) equipped with unpierced PTFE-lined septa.
7.0 PROCEDURE
7.1 Sample preparation
7.1.1 Samples are normally prepared by one of the following methods prior to
GC/MS analysis.
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Matrix
Methods
Air (particulates arid sorbent resin} 3542
Water (including TCLP leachates) 3510,3520,3535
Soil/sediment 3540, 3541, 3545, 3550,3560, 3561
Waste 3540, 3541, 3545, 3550, 3560, 3561, 3580
7.1.2 In very limited applications, dined injection of the sample into the GC/MS
system with a 10-jjL syringe may be appropriate. The detection limit is very high
(approximately 10,000 Mg/L). Therefore, it is only permitted where concentrations in excess
of 10,000 tig/i- are expected.
7.2 Extract cleanup - Extracts may be cleaned up by any of the following methods prior to
GC/MS analysis.
Analvtes of interest
Aniline & aniline derivatives
Phenols
Phthalate esters
Nitrosamines
Organochlorine pesticides & PCBs
Nitroaromatics and cyclic ketones
Polynuclear aromatic hydrocarbons
Haloethers
Chlorinated hydrocarbons
Organophosphorus pesticides
Petroleum waste
All base, neutral, and add
priority pollutants
a Method 8041 includes a derivatization
are encountered on GC/FID.
Methods
3620
3630, 3640, 8041"
3610, 3620, 3640
3610, 3620, 3640
3610, 3620, 3630, 3660, 3665
3620, 3640
3611,3630,3640
3620, 3640
3620,3640
3620
3611, 3650
3640
lique and a GC/ECD analysis, if interferences
7.3 Initial calibration
Establish the GC/MS operating conditions, using the following recommendations as guidance.
Mass range:
Scan time:
Initial temperature:
Temperature program:
Final temperature:
Injector temperature:
Transfer line temperature:
Source temperature:
Injector
Injection volume:
Carrier gas:
Ion trap only:
35-500 amu
1 sec/scan
40°C, hold for 4 minutes
40-270°C at 10°C/min
270°C, hold until benzo[g,h,i]perylene elutes
250-300®C
250-300°C
According to manufacturer's specifications
Grob-type, splitless
1-2 |iL
Hydrogen at 50 cm/sec or helium at 30 cm/sec
Set axial modulation, manifold temperature, and emission
current to manufacturer's recommendations
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Split injection is allowed if the sensitivity of the mass spectrometer is sufficient.
7.3.1 The GC/MS system must be hardware-tuned using a 50-ng injection of
DFTPP. Analyses must not begin until the tuning criteria are met.
7.3.1.1 In the absence of specific recommendations on how to acquire the
mass spectrum of DFTPP from the instrument manufacturer, the following approach
has been shown to be useful: Three scans (the peak apex scan and the scans
immediately preceding and following the apex) are acquired and averaged.
Background subtraction is required, and must be accomplished using a single scan
acquired no more than 20 scans prior to the elution of DFTPP. The background
subtraction should be designed only to eliminate column bleed or instrument
background ions. Do not subtract part of the DFTPP peak,
7.3.1.2 Use the DFTPP mass intensity criteria in Table 3 as tuning
acceptance criteria. Alternatively, other documented tuning criteria may be used (e.g.
CLP, Method 525, or manufacturer's instructions), provided that method performance
is not adversely affected.
NOTE: All subsequent standards, samples, MS/MSDs, and blanks associated with
a DFTPP analysis must use the identical mass spectrometer instrument
conditions.
7.3.1.3 The GC/MS tuning standard solution should also be used to
assess GC column performance and injection port inertness. Degradation of DDT to
DDE and DDD should not exceed 20%. (See Sec. 8.0 of Method 8081 for the percent
breakdown calculation). Benzidine and pentachlorophenol should be present at their
normal responses, and no peak tailing should be visible.
7.3.1.4 If degradation is excessive and/or poor chromatography is noted,
the injection port may require cleaning. It may also be necessary to break off the first
6-12 in. of the capillary column. The use of a guard column (Sec. 4.1.6) between the
injection port and the analytical column may help prolong analytical column
performance.
7.3.2 The internal standards selected in Sec. 5.4 should permit most of the
components of interest in a chromatogram to have retention times of 0.80-1.20 relative to one
of the internal standards. Use the base peak ion from the specific internal standard as the
primary ion for quantitation (see Table 1). If interferences are noted, use the next most intense
ion as the quantitation ion (i.e. for 1,4-dichlorobenzene-d4, use 152 m/z for quantitation).
7.3.3 Analyze 1-2 v>L of each calibration standard (containing internal standards)
and tabulate the area of the primary characteristic ion against concentration for each target
analyte (as indicated in Table 1). A set of at least five calibration standards is necessary (see
Sec. 5.6 and Method 8000). The injection volume must be the same for all standards and
sample extracts. Figure 1 shows a chromatogram of a calibration standard containing
base/neutral and acid analytes.
Calculate response factors (RFs) for each target analyte relative to one of the internal
standards as follows:
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where:
^ = Peak area (or height) of the analyte or surrogate.
A* = Peak area (or height) of the internal standard.
Cs = Concentration of the analyte or surrogate, in pg/L.
= Concentration of the internal standard, in pg/L.
7.3.4 System performance check compounds (SPCCs)
7.3.4.1 A system performance check must be performed to ensure that
minimum average RFs are met before the calibration curve is used. For semivolatiles,
the System performance check compounds (SPCCs) are: N-nitroso-di-n-propyiamine;
hexachlorocyclopentadiene; 2,4-dinitrophenol; and 4-nitrophenol.
7.3.4.2 The minimum acceptable average RF for these compounds is
0.050. These SPCCs typically have very low RFs (0.1-0.2) and tend to decrease in
response as the chromatographic system begins to deteriorate or the standard material
begins to deteriorate. They are usually the first to show poor performance. Therefore,
they must meet the minimum requirement when the system is calibrated.
7.3.4.3 If the minimum response factors are not met, the system must be
evaluated, and corrective action must be taken before sample analysts begins.
Possible problems include standard mixture degradation, injection port inlet
contamination, contamination at the front end of the analytical column, and active sites
in the column or chromatographic system. This check must be met before sample
analysis begins.
7.3.5 Calibration check compounds (CCCs)
7.3.5.1 The purpose of the CCCs are to evaluate the calibration from the
standpoint of the integrity of the system. High variability for these compounds may be
indicative of system leaks or reactive sites on the column. Meeting the CCC criteria
is not a substitute for successful calibration of the target analytes using one of the
approaches described in Sec. 7.0 of Method 8000.
7.3.5.2 Calculate the mean response factor and the relative standard
deviation (RSD) of the response factors for each target analyte. The RSD should be
less than or equal to 15% for each target analyte. However, the RSD for each
individual CCC (see Table 4) must be less than or equal to 30%.
ERF.
mean RF - RF -
i«1
SD =
E(RFrRF)2
_M
n-1
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RSD = 52 X 100
RF
7.3.5.3 if the RSD of any CCC is greater than 30%, then the
chromatographic system is too reactive for analysis to begin. Clean or replace the
injector liner and/or capillary column, then repeat the calibration procedure beginning
with Sec. 7.3.
7.3.5.4 If the CCCs are not included in the list of analytes for a project,
and therefore not included in the calibration standards, then refer to Sec, 7,0 of Method
8000.
7.3.6 Evaluation of retention times - The relative retention time (RRT) of each target
analyfe in each calibration standard should agree within 0.06 RRT units. Late-eluting target
analytes usually have much better agreement.
rrt - Retention time of the analyte
Retention time of the internal standard
7.3.7 Linearity of target analytes - If the RSD of any target analytes is 15% or less,
then the relative response factor is assumed to be constant over the calibration range, and the
average relative response factor may be used for quantitation (Sec. 7.6.2).
7.3.7.1 If the RSD of any target analyte is greater than 15%, refer to Sec.
7.0 in Method 8000 for additional calibration options. One of the options must be
applied to GC/MS calibration in this situation, or a new initial calibration must be
performed.
NOTE: Method 8000 designates a linearity criterion of 20% RSD. That criterion
pertains to GC and HPLC methods other than GC/MS. Method 8270 requires
15% RSD as evidence of sufficient linearity to employ an average response
factor.
7.3.7.2 When the RSD exceeds 15%, the plotting and visual inspection
of a calibration curve can be a useful diagnostic tool. The inspection may indicate
analytical problems, including errors in standard preparation, the presence of active
sites in the chromatographic system, analytes that exhibit poor chromatographic
behavior, etc.
7.4 GC/MS calibration verification - Calibration verification consists of three steps that are
performed at the beginning of each 12-hour analytical shift.
7.4.1 Prior to the analysis of samples or calibration standards, inject 50 ng of the
DFTPP standard into the GC/MS system. The resultant mass spectrum for DFTPP must meet
the criteria given in Table 3 before sample analysis begins. These criteria must be
demonstrated each 12-hour shift during which samples are analyzed.
7.4.2 The initial calibration (Sec. 7.3) for each compound of interest should be
verified once every 12 hours prior to sample analysis, using the introduction technique and
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conditions used for samples. This is accomplished by analyzing a calibration standard at a
concentration near the midpoint concentration for the calibrating range of the GC/MS. The
results from the calibration standard analysis should meet the verification acceptance criteria
provided in Sees. 7.4.4 through 7.4.7.
NOTE: The DFTPP and calibration verification standard may be combined into a single
standard as long as both tuning and calibration verification acceptance criteria for the
project can be met without interferences.
7.4.3 A method blank should be analyzed either after the calibration
standard, or at any other time during the analytical shift, to ensure that the total system
(introduction device, transfer lines and GC/MS system) is free of contaminants. If the method
blank indicates contamination, then it may be appropriate to analyze a solvent blank to
demonstrate that the contamination is not a result of carryover from standards or samples.
See Sec. 8.0 of Method 8000B for method blank performance criteria.
7.4.4 System performance check compounds (SPCCs)
7.4.4.1 A system performance check must be made during every 12-hour
analytical shift. Each SPCC in the calibration verification standard must meet a
minimum response factor of 0.050. This is the same check that is applied during the
initial calibration.
7.4.4.2 If the minimum response factors are not met, the system must be
evaluated, and corrective action must be taken before sample analysis begins.
Possible problems include standard mixture degradation, injection port inlet
contamination, contamination at the front end of the analytical column, and active sites
In the column or chromatographic system. This check must be met before sample
analysis begins.
7.4.5 Calibration check compounds (CCCs)
7.4.5.1 After the system performance check is met, the CCCs listed in
Table 4 are used to check the validity of the initial calibration. Use percent difference
when performing the average response factor model calibration. Use percent drift
when calibrating using a regression fit model. Refer to Sec. 7.0 of Method 8000 for
guidance on calculating percent difference and drift.
7.4.5.2 If the percent difference for each CCC is less than or equal to
20%, then the initial calibration Is assumed to be valid. If the criterion is not met (i.e.,
greater than 20% difference or drift) for any one CCC, then-corrective action must be
taken prior to the analysis of samples. If the CCCs are not included in the list of
analytes for a project, and therefore not included in the calibration standards, then all
analytes must meet the 20% difference or drift criterion.
7.4.5.3 Problems similar to those listed under SPCCs could affect the
CCCs. If the problem cannot be corrected by other measures, a new initial calibration
must be generated. The CCC criteria must be met before sample analysis begins.
7.4.6 Internal standard retention time - The retention times of the internal standards
in the calibration verification standard must be evaluated immediately after or during data
acquisition. If the retention time for any internal standard changes by more than 30 seconds
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from that in the mid-point standard level of the most recent initial calibration sequence, then
the chromatographic system must be inspected for malfunctions and corrections must be
made, as required. When corrections are made, reanalysis of samples analyzed while the
system was malfunctioning is required.
7.4.7 Internal standard response - If the E1CP area for any of the internal standards
in the calibration verification standard changes by a factor of two (-50% to +100%) from that
in the mid-point standard level of the most recent initial calibration sequence, the mass
spectrometer must be inspected for malfunctions and corrections must be made, as
appropriate. When corrections are made, reanalysis of samples analyzed while the system
was malfunctioning is required.
7.5 GC/MS analysis of samples
7.5.1 It is highly recommended that sample extracts be screened on a GC/FID or
GC/PID using the same type of capillary column used in the GC/MS system. This will minimize
contamination of the GC/MS system from unexpectedly high concentrations of organic
compounds.
7.5.2 Allow the sample extract to warm to room temperature. Just prior to analysis,
add 10 |iL of the internal standard solution to the 1-mL concentrated sample extract obtained
from sample preparation.
7.5.3 Inject a 1-2 |iL aliquot of the sample extract into the GC/MS system, using the
same operating conditions that were used for the calibration (Sec. 7.3). The volume to be
injected should contain 100 ng of base/neutral and 200 ng of acid surrogates (assuming 100%
recovery), unless a more sensitive GC/MS system is being used and the surrogate solution is
less concentrated then that listed in Sec. 5.7. The injection volume must be the same volume
used for the calibration standards.
7.5.4 If the response for any quantitation ion exceeds the initial calibration range
of the GC/MS system, the sample extract must be diluted and reanalyzed. Additional internal
standard solution must be added to the diluted extract to maintain the same concentration as
in the calibration standards (40 ng/pL, unless a more sensitive GC/MS system is being used).
Secondary ion quantitation should be used only when there are sample interferences with the
primary ion.
NOTE: It may be a useful diagnostic tool to monitor internal standard retention times and
responses (area counts) in all samples, spikes, blanks, and standards to effectively
check drifting method performance, poor injection execution, and anticipate the need
for system inspection and/or maintenance.
7.5.4.1 When ions from a compound in the sample saturate the detector,
this analysis must be followed by the analysis of an instrument blank consisting of
clean solvent. If the blank analysis is not free of interferences, then the system must
be decontaminated. Sample analysis may not resume until the blank analysis is
demonstrated to be free of interferences.
7.5.4.2 All dilutions should keep the response of the major constituents
(previously saturated peaks) in the upper half of the linear range of the curve.
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7.5.5 The use of selected ion monitoring (SIM) is acceptable for applications
requiring detection limits below the normal range of electron impact mass spectrometry.
However, SIM may provide a lesser degree of confidence in the compound identification unless
multiple ions are monitored for each compound.
7.6 Qualitative analysis
7.6.1 The qualitative identification of compounds determined by this method is
based on retention time and on comparison of the sample mass spectrum, after background
correction, with characteristic ions in a reference mass spectrum. The reference mass
spectrum must be generated by the laboratory using the conditions of this method. The
characteristic ions from the reference mass spectrum are defined as the three ions of greatest
relative intensity, or any ions over 30% relative intensity, if less than three such ions occur in
the reference spectrum. Compounds are identified when the following criteria are met.
7.6.1.1 The intensities of the characteristic ions of a compound must
maximize in the same scan or within one scan of each other. Selection of a peak by'
a data system target compound search routine where the search is based on the
presence of a target chromatographic peak containing ions specific for the target
compound at a compound-specific retention time will be accepted as meeting this
criterion.
7.6.1.2 The RRT of the sample component is within ± 0.06 RRT units of
the RRT of the standard component.
7.6.1.3 The relative intensities of the characteristic ions agree within 30%
of the relative intensities of these ions in the reference spectrum. (Example: For an
ion with an abundance of 50% in the reference spectrum, the corresponding
abundance in a sample spectrum can range between 20% and 80%,)
7.6.1.4 Structural isomers that produce very similar mass spectra should
be identified as individual isomers if they have sufficiently different GC retention times.
Sufficient GC resolution is achieved if the height of the valley between two isomer
peaks is less than 25% of the sum of the two peak heights. Otherwise, structural
isomers are identified as isomeric pairs. Diastereomeric pairs (e.g., Aramite and
Isosafirol) that may be separable by the GC should be identified, quantitated and
reported as the sum of both compounds by the GC.
7.6.1.5 Identification is hampered when sample components are not
resolved chromatographicaliy and produce mass spectra containing ions contributed
by more than one analyte. When gas chromatographic peaks obviously represent
more than one sample component (i.e., a broadened peak with shoulders) or a valley
between two or more maxima), appropriate selection of analyte spectra and
background spectra is important.
7.6.1.6 Examination of extracted ion current profiles of appropriate ions
can aid in the selection of spectra and in qualitative identification of compounds. When
analytes coelute (i.e., only one chromatographic peak is apparent), the identification
criteria may be met, but each analyte spectrum will contain extraneous ions contributed
by the coeluting compound.
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7.6.2 For samples containing components not associated with the calibration
standards, a library search may be made for the purpose of tentative identification. The
necessity to perform this type of identification will be determined by the purpose of the
analyses being conducted. Data system library search routines should not use normalization
routines that would misrepresent the library or unknown spectra when compared to each other.
For example, the RCRA permit or waste delisting requirements may require the
reporting of non-target analytes. Only after visual comparison of sample spectra with the
nearest library searches may the analyst assign a tentative identification. Guidelines for
tentative identification are:
(1) Relative intensities of major ions in the reference spectrum (ions > 10% of the
most abundant ion) should be present in the sample spectrum.
(2) The relative intensities of the major ions should agree within ± 20%. (Example;
For an ion with an abundance of 50% in the standard spectrum, the corresponding
sample ion abundance must be between 30 and 70%.) -
(3) Molecular ions present in the reference spectrum should be present in the sample
spectrum.
(4) Ions present in the sample spectrum but not in the reference spectrum should be
reviewed for possible background contamination or presence of coeluting
compounds.
(5) Ions present in the reference spectrum but not in the sample spectrum should be
reviewed for possible subtraction from the sample spectrum because of
background contamination or coeluting peaks. Data system library reduction
programs can sometimes create these discrepancies.
7.7 Quantitative analysis
7.7.1 Once a compound has been identified, the quantitation of that compound will
be based on the integrated abundance of the primary characteristic ion from the EICP.
7.7.2 If the RSD of a compound's response factor is 15% or less, then the
concentration in the extract may be determined using the average response factor (HF) from
initial calibration data (Sec. 7.3.5). See Method 8000, Sec. 7.0, for the equations describing
internal standard calibration and either linear or non-linear calibrations.
7.7.3 Where applicable, the concentration of any non-target analytes identified in
the sample (Sec. 7.6.2) should be estimated. The same formulae should be used with the
following modifications: The areas A* and A*, should be from the total ion chromatograms, and
the RF for the compound should be assumed to be 1.
7.7.4 The resulting concentration should be reported indicating: (1) that the value
is an estimate, and (2) which internal standard was used to determine concentration. Use the
nearest internal standard free of interferences.
7.7.5 Quantitation of multicomponent compounds (e.g., Toxaphene, Anoclors, etc.)
is beyond the scope of Method 8270. Normally, quantitation is performed using a GC/ECD,
by Methods 8081 or 8082. However, Method 8270 may be used to confirm the identification
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of these compounds, when the concentrations are at least 10 ng/^iL in the concentrated
sample extract.
7.7.6 Structural isomers that produce vety similar mass spectra should be
quantitated as individual isomers if they have sufficiently different GC retention times.
Sufficient GC resolution is achieved if the height of the valley between two isomer peaks is less
than 25% of the sum of the two peak heights. Otherwise, structural isomers are quantitated
as isomeric pairs. Diastereomeric pairs (e.g., Aramite and Isosafro!) that may be separable
by the GC should be summed and reported as the sum of both compounds.
8.0 QUALITY CONTROL
8.1 Refer to Chapter One and Method 8000 for specific quality control (QC) procedures.
Quality control procedures to ensure the proper operation of the various sample preparation and/or
sample introduction techniques can be found in Method 3500. Each laboratory should maintain a
formal qualify assurance program. The laboratory should also maintain records to document the
quality of the data generated.
8.2 Quality control procedures necessary to evaluate the GC system operation are found
in Sec. 7.0 of Method 8000 and include calibration verification and chromatographic analysis of
samples, in addition, instrument QC requirements may be found in the following sections of Method
8270:
8.2.1 The GG/MS system must be tuned to meet the DFTPP criteria discussed in
Sees. 7.3.1 and 7.4.1.
8.2.2 There must be an initial calibration of the GC/MS system as described in Sec.
7.3.
8.2.3 The GC/MS system must meet the calibration verification acceptance criteria
in Sec. 7.4, each 12 hours.
8.2.4 The RRT of the sample component must fall within the RRT window of the
standard component provided in Sec. 7.6.1.
8.3 Initial demonstration of proficiency - Each laboratory must demonstrate initial proficiency
with each sample preparation and determinative method combination it utilizes, by generating data
of acceptable accuracy and precision for target anaiytes in a clean matrix. The laboratory must also
repeat the following operations whenever new staff are trained or significant changes in
instrumentation are made. See Method 8000, Sec. 8.0 for information on bow to accomplish this
demonstration.
8.4 Sample quality control for preparation and analysis - The laboratory must also have
procedures for documenting the effect of the matrix on method performance (precision, accuracy,
and detection limit). At a minimum, this includes the analysis of QC samples including a method
blank, matrix spike, a duplicate, and a laboratory control sample (LCS) in each analytical batch and
the addition of surrogates to each field sample and QC sample.
8.4.1 Before processing any samples, the analyst should demonstrate, through the
analysis of a method blank, that interferences from the analytical system, glassware, and
reagents are under control. Each time a set of samples is analyzed or there is a change in
8270D - 22
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reagents, a method blank should be analyzed as a safeguard against chronic laboratory
contamination. The blanks should be carried through all stages of sample preparation and
measurement.
8.4.2 Documenting the effect of the malrix should include the analysis of at least
one matrix spike and one duplicate unspiked sample or one matrix spike/matrix spike duplicate
pair. The decision on whether to prepare and analyze duplicate samples or a matrix
spike/matrix spike duplicate must be based on a knowledge of the samples in the sample
batch, if samples are expected to contain target analytes, then laboratories may use one
matrix spike and a duplicate analysis of an unspiked field sample. If samples are not expected
to contain target analytes, laboratories should use a matrix spike and matrix spike duplicate
pair.
8.4.3 A laboratory control sample (LCS) should be included with each analytical
batch. The LCS consists of an aliquot of a clean (control) matrix similar to the sample matrix
and of the same weight or volume. The LCS is spiked with the same analytes at the same
concentrations as the matrix spike. When the results of the matrix spike analysis indicate a
potential problem due to the sample matrix itself, the LCS results are used to verify that the
laboratory can perform the analysis in a clean matrix.
8.4.4 See Method 8000, Sec. 8.0 for the details on carrying out sample quality
control procedures for preparation and analysis.
8.5 Surrogate recoveries - The laboratory must evaluate surrogate recovery data from
individual samples versus the surrogate control limits developed by the laboratory. See Method
8000, Sec. 8.0 for information on evaluating surrogate data and developing and updating surrogate
limits.
8.6 The experience of the analyst performing GC/MS analyses is invaluable to the success
of the methods. Each day that analysis is performed, the calibration verification standard should be
evaluated to determine if the chromatographic system is operating properly. Questions that should
be asked are: Do the peaks look normal? Is the response obtained comparable to the response
from previous calibrations? Careful examination of the standard chromatogram can indicate whether
the column is still performing acceptably, the injector is leaking, the injector septum needs replacing,
etc. If any changes are made to the system (e.g., the column changed, a septum is changed), see
the guidance in Sec 8.2 of Method 8000 regarding whether recalibration of the system must take
place.
8.7 It is recommended that the laboratory adopt additional quality assurance practices for
use with this method. The specific practices that are most productive depend upon the needs of the
laboratory and the nature of the samples. Whenever possible, the laboratory should analyze
standard reference materials and participate in relevant performance evaluation studies.
9.0 METHOD PERFORMANCE
9.1 Method 8250 (the packed column version of Method 8270) was tested by 15
laboratories using organic-free reagent water, drinking water, surface water, and industrial
wastewaters spiked at six concentrations ranging from 5 to 1,300 (jg/L. Single operator accuracy
and precision, and method accuracy were found to be directly related to the concentration of the
analyte and essentially independent of the sample matrix. Linear equations to describe these
relationships are presented in Table 7. These values are presented as guidance only and are not
8270D-23
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intended as absolute acceptance criteria. Laboratories should generate their own acceptance
criteria for capillary column method performance. (See Method 8000.)
9.2 Chromatograms from calibration standards analyzed with Day 0 and Day 7 samples
were compared to detect possible deterioration of GC performance. These recoveries (using
Method 3510 extraction) are presented in Table 8.
9.3 Method performance data using Method 3541 (automated Sodhlet extraction) are
presented in Table 9. Single laboratory accuracy and precision data were obtained for semivolatiie
organics in a clay soil by spiking at a concentration of 6 mg/kg for each compound. The spiking
solution was mixed into the soil during addition and then allowed to equilibrate for approximately 1
hour prior to extraction. The spiked samples were then extracted by Method 3541 (Automated
Soxhlet). Three extractions were performed and each extract. was analyzed by gas
chromatography/ma ss spectrometry following Method 8270. The low recovery of the more volatile
compounds is probably due to volatilization losses during equilibration. These data are listed in
Table 10 and were taken from Reference 7.
9.4 Surrogate precision and accuracy data are presented in Table 11 from a field dynamic
spiking study based on air sampling by Method 0010. The trapping media were prepared for analysis
by Method 3542 arid subsequently analyzed by Method 8270.
9.5 Single laboratory precision and bias data using Method 3545 (pressurized fluid
extraction) for semivolatiie organic compounds are presented in Table 12. The samples were
conditioned spiked samples prepared and certified by a commercial supplier that contained 57
semivolatiie organics at three concentrations (250, 2500, and 12,500 ygfkg) on three types of soil
(clay, loam and sand). Spiked samples were extracted both by the Dionex Accelerated Solvent
Extraction system and by the Perstorp Environmental Soxtec1** (automated Soxhlet). The data in
Table 12 represent seven replicate extractions and analyses for each individual sample and were
taken from reference 9. The average recoveries from the three matrices for all analytes and all
replicates relative to the automated Soxhlet data are as follows: clay 96.8%, loam 98.7% and sand
102.1%. The average recoveries from the three concentrations also relative to the automated
Soxhlet data are as follows: low-101.2%, mid-97.2% and high-99.2%.
9.6 Single laboratory precision and bias data using Method 3561 (SFE extraction of PAHs
with a variable restrictor and solid trapping material) were obtained for the method analytes by the
extraction of two certified reference materials (EC-1, a lake sediment from Environment Canada and
HS-3, a marine sediment from the National Science and Engineering Research Council of Canada,
both naturally-contaminated with PAHs). The SFE instrument used for these extractions was a
Hewlett-Packard Model 7680. Analysis was by GC/MS. Average recoveries from six replicate
extractions range from 85 to 148% (overall average of 100%) based on the certified value (or a
Soxhlet value if a certified value was unavailable for a specific analyte) for the lake sediment.
Average recoveries from three replicate extractions range from 73 to 133% (overall average of 92%)
based on the certified value for the marine sediment. The data are found in Tables 13 and 14 and
were taken from Reference 10.
9.7 Single laboratory precision and accuracy data using Method 3561 (SFE extraction of
PAHs with a fixed restrictor and liquid trapping) were obtained for twelve of the method analytes by
the extraction of a certified reference material (a soil naturally contaminated with PAHs). The SFE
instrument used for these extractions was a Dionex Model 703-M. Analysis was by GC/MS.
Average recoveries from four replicate extractions range from 60 to 122% (overall average of 89%)
based on the certified value. Following are the instrument conditions that were utilized to extract a
3.4 g sample: Pressure - 300 atm; Time - 60 min.; Extraction fluid - C02; Modifier -10% 1:1 (v/v)
8270D - 24
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methanol/methylene chloride; Oven temperature - 80°C; Restrictor temperature - 120°C; and,
Trapping fluid - chloroform (methylene chloride has also been used). The data are found in Table
15 and were taken from Reference 11,
9.8 Tables 16 and 17 contain single-laboratory precision and accuracy data for solid-phase
extraction of TCLP buffer solutions spiked at two levels and extracted using Method 3535,
9.9 Table 18 contains multiple-laboratory data for solid-phase extraction of spiked TCLP
soil leachates extracted using Method 3535.
10.0 REFERENCES
1. Eichelberger, J.W., Harris, L.E., and Budde, W.L., "Reference Compound to Calibrate Ion
Abundance Measurement In Gas Chromatography-Mass Spectrometry Systems", Analytical
Chemistry, 47,995-1000,1975.
2. "Method Detection Limit for Methods 624 and 625", Olynyk, P., Budde, W.L, and Eichelberger,
J.W., unpublished report, October 1980.
3. "tnteriaboratory Method Study for EPA Method 625-Base/Neutrals, Acids, and Pesticides",
Final Report for EPA Contract 68-03-3102.
4. Burke, J.A., "Gas Chromatography for Pesticide Residue Analysis: Some Practical Aspects",
Journal of the Association of Official Analytical Chemists (AOAC), 48,1037,1965.
5. Lucas, S.V., Komfeld, R.A., "GC-MS Suitability Testing of RCRA Appendix VIII and Michigan
List AnalytesU.S. Environmental Protection Agency, Environmental Monitoring and Support
Laboratory, Cincinnati, OH 45268, February 20,1987, Contract No. 68-03-3224.
6. Engel, T.M., Komfeld, R.A., Warner, J.S., and Andrews, K.D., "Screening of Semivolatile
Organic Compounds for Extractabiiity and Aqueous Stability by SW-846, Method 3510", U.S.
Environmental Protection Agency, Environmental Monitoring and Support Laboratory,
Cincinnati, OH 45268, June 5,1987, Contract 68-03-3224.
7. Lopez-Avila, V, (W, Beckert, Project Officer); "Development of a Soxtec Extraction Procedure
for Extraction of Organic Compounds from Soils and Sediments"; U.S. Environmental
Protection Agency, Environmental Monitoring and Support Laboratory, Las Vegas, NV,
October 1991; EPA 600/X-91/140.
8. Bursey, J., Merrill, R., McAllister, R., and McGaughey, J., "Laboratory Validation of VOST and
SemiVOST for Halogenated Hydrocarbons from the Clean Air Act Amendments List", Vol. 1
and 2, U.S. Environmental Protection Agency, EPA 600/R-93/123a and b, (NTIS PB 93-227163
and 93-27171), Research Triangle Park, NC, July 1993.
9. Richter, B., Ezzell, J., and Felix, D., "Single Laboratory Method Validation Report: Extraction
of Target Compound List/Priority Pollutant List BNAs and Pesticides using Accelerated Solvent
Extraction (ASE) with Analytical Validation by GC/MS and GC/ECD", Document 101124,
Dionex Corporation, Salt Lake City, UT, June 16,1994.
8270D - 25
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10. Lee, H.B., Peart, T.E., Hong-You, R.L., and Gere, D.R., "Supercritical Carbon Dioxide
Extraction of Polycyclic Aromatic Hydrocarbons from Sediments", J. Chromatography, A 653
83-91 (1993).
11. Warner, S,, "SFE Extraction of PNAs from Solid Matrices Using the Dionex 703M SFE
Extractor and a Liquid Trap," EPA Region lit, Central Regional Laboratory, 839 Bestgate Road,
Annapolis, MD 21401, December 12,1994.
12. Marked, C., "3M Data Submission to EPA," letter to B. Lesnik, June 27,1995.
8270D - 26
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TABLE 1
CHARACTERISTIC IONS FOR SEMI VOLATILE COMPOUNDS

Retention
Primary
Secondary lon(s)
Compound
Time (min)
Ion

2-Picoline
3.75s
93
66,92
Aniline
5.68
93
66,65
Phenol
5.77
94
65,66
Bis(2-chloroethy1) ether
5.82
93
63,95
2-ChlorophenoI
5.97
128
64,130
1,3-Dichlorobenzene
6.27
146
148,111
1,4-Dichtorobenzene-d4 (IS)
6.35
152
150,115
1,4- Dichlorobenzene
6.40
146
148,111
Benzyl alcohol
6.78
108
79,77
1,2-Dichlorobenzene
6.85
146
148,111
N-Nitrosornethylethylamine
6.97
88
42,43,56
Bis(2-chloroisopropyl) ether
7.22
45
77,121
Ethyl carbamate
7.27
62
44,45,74
Thiophenol (Benzenethiol)
7.42
110
66,109,84
Methyl methanesulfonate
7.48
80
79,65,95
N-Nitrosodi-n-propyiamine
7.55
70
42,101,130
Hexachloroethane
7.65
117
201,199
Maleic anhydride
7.65
54
98,53,44
Nitrobenzene
7.87
77
123,65
Isophorone
8.53
82
95,138
N-Nitrosodiethylamine
8.70
102
42,57,44,56
2-Nitrophenol
8.75
139
109,65
2,4-Dimethylphenol
9.03
122
107,121
p-Benzoquinone
9.13
108
54,82,80
Bis(2-chloroethoxy)methane
9.23
93
95,123
Benzoic acid
9.38
122
105,77
2,4-Dichlorophenol
9.48
162
164,98
Trimethyf phosphate
9.53
110
79,95,109,140
Ethyl methanesulfonate
9.62
79
109,97,45,65
1,2,4-Trichlorobenzene
9.67
180
182,145
Naphthalene-dg (IS)
9.75
136
68
Naphthalene
9.82
128
129,127
Hexachlorobutadiene
10.43
225
223,227
Tetraethyl pyrophosphate
11.07
99
155,127,81,109
Diethyl sulfate
11.37
139
45,59,99,111,125
4-Ch(oro-3-methylphenol
11.68
107
144,142
2-Methylnaphthalene
11.87
142
141
2-Methylphenol
12.40
107
108,77,79,90
Hexachloropropene
12.45
213
211,215,117,106,141
Hexachlorocyclopentad iene
12.60
237
235,272
N-Nitrosopyrrolidine
12.65
100
41,42,68,69
Acetophenone
12.67
105
71,51,120
8270D - 27
Revision 4
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TABLE 1
(continued)
Retention Primary Secondary lon(s)
Compound
Time (min)
ion

4-Methylphenol
12.82
107
108,77,79,90
2,4,6-T richlorophenoi
12.85
196
198,200
o-ToIuidine
12.87
106
107,77,51,79
3-Methylphenol
12.93
107
108,77,79,90
2-Chloronaphthalene
13.30
162
127,164
N-Nitrosopiperidirie
13.55
114
42,55,56,41
1,4-PhenyJenediamine
13.62
108
80,53,54,52
1 -Chloronaphthalene
13.65*
162
127,164
2-Nitroanitine
13.75
65
92,138
5-Chloro-2-methy1aniiine
14.28
106
.141,140,77,89
Dimethyl phthalate
14.48
163
194,164
Acenaphthylene
14.57
152
151,153
2,6-DinNrotoluene
14.62
165
63,89
Phthalic anhydride
14.62
104
76,50,148
o-Anisidine
15.00
108
80,123,52
3-Nitroaniline
15.02
138
108,92
Acenaphthene-d10 (IS)
15.05
164
162,160
Acenaphthene
15.13
154
153,152
2,4-Dinitrophenol
15.35
184
63,154
2,6-Dinitrophenol
15.47
162
164,126,98,63
4-Chloroaniiine
15.50
127
129,65,92
Isosafrole
15.60
162
131,104,77,51
Dibenzofuran
15.63
168
139
2,4-Diaminotoiuene
15.78
121
122,94,77,104
2,4-Oinitrotoluene
15.80
165
63,89
4-Nitrophenol
15.80
139
109,65
2-NaphthyIamine
16.00®
143
115,116
1,4-Naphthoquinone
16.23
158
104,102,76,50,130
p-Cresidine
16.45
122
94,137,77,93
Dichlorovos
16.48
109
185,79,145
Diethyl phthalate
16.70
149
177,150
Fluorene
16.70
166
165,167
2,4,5-T rimethylaniline
16.70
120
135,134,91,77
N-Nitrosodi-n-butylamine
16.73
84
57,41,116,158
4-Chlorophenyl phenyl ether
16.78
204
206,141
Hydroquinone
16.93
110
81,53,55
4,6-Dinitro-2-methyl phenol
17.05
198
51,105
Resorcinol
17.13
110
81,82,53,69
N-Nitrosodiphenylamirte
17.17
169
168,167
Safrole
17.23
162
104,77,103,135
Hexamethyl phosphoramide
17.33
135
44,179,92,42
3-(Chloromethy1) pyridine hydrochloride
17.50
92
127,129,65,39
Diphenylamine
17.54"
169
168,167
8270D - 28
Revision 4
January 1998
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TABLE 1
(continued)
Retention Primary Secondary lon(s)
Compound
Time (min)
Ion

1,2,4,5-Tetrachlorobenzene
17.97
216
214,179,108,143,218
1-Naphthytamine
18.20
143
115,89,63
1-Ace tyi-2-thio urea
18.22
118
43,42,76
4-Bromophenyl phenyl ether
18.27
248
250,141
Toluene diisocyanate
18.42
174
145,173,146,132,91
2,4,5-Trichlorophenol
18.47
196
198,97,132,99
Hexachlorobenzene
18.65
284
142,249
Nicotine
18.70
84
133,161,162
Pentachlorophenol
19.25
266
264,268
5-Nitro-o-toluidine
19.27
152
77,79,106,94
Thionazine
19.35
107
96,97,143,79,68
4-Nitroaniline
19.37
138
65,108,92,80,39
Phenanthrene-d10 (IS)
19.55
188
94,80
Phenanthrene
19.62
178
179,176
Anthracene
19.77
178
176,179
1,4-Dinitrobenzene
19.83
168
75,50,76,92,122
Mevinphos
19.90
127
192,109,67,164
Naled
20.03
109
145,147,301,79,189
1,3-Dinitnobenzene
20.18
168
76,50,75,92,122
Diallate (cis or trans)
20.57
86
234,43,70
1,2-Dinitrobenzene
20.58
168
50,63,74
Diallate (trans or cis)
20.78
86
234,43,70
Pentachlorobenzene
21.35
250
252,108,248,215,254
5-Nitro-o-anisidine
21.50
168
79,52,138,153,77
Pentachloronrtrabenzene
21.72
237
142,214,249,295,265
4-Nitroquinoline-1-oxide
21.73
174
101,128,75,116
Di-n-butyi phthalate
21.78
149
150,104
2,3,4,6-T etrachloraphenol
21.88
232
131,230,166,234,168
Dihydrosaffrole
22.42
135
64,77
Demeton-O
22.72
88
89,60,61,115,171
Fluoranthene
23.33
202
101,203
1,3,5-Trinitrobenzene
23.68
75
.74,213,120,91,63
Dicrotophos
23.82
127
67,72,109,193,237
Benzidine
23.87
184
92,185
Trifluralin
23.88
306
43,264,41,290
Bromoxynil
23.90
277
279,88,275,168
Pyrene
24.02
202
200,203
Monocrotophos
24.08
127
192,67,97,109
Phorate
24.10
75
121,97,93,260
Sulfallate
24.23
188
88,72,60,44
Demeton-S
24.30
88
60,81,89,114,115
Phenacetin
24.33
108
180,179,109,137,80
Dimethoate
24.70
87
93,125,143,229
8270D - 29
Revision 4
January 1998
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TABLE 1
(continued)
Compound
Retention
Time (min)
Primary
Ion
Secondary lon(s)
Phenobarbital
24.70
204
117,232,146,161
Carbofuran
24.90
164
149,131,122
Octamethyl pyrophosphoramide
24.95
135
44,199,286,153,243
4-Aminobiphenyl
25.08
169
168,170,115
Dioxathion
25.25
97
125,270,153
Terbufos
25.35
231
57,97,153,103
a,a-Dimethylphenylamine
25.43
58
91,65,134,42
Pronamide
25.48
173
175,145,109,147
Aminoazobenzene
25.72
197
92,120,65,77
Dichlone
25.77
191
163,226,228,135,193
Dinoseb
25.83
211
163,147,117,240
Disulfoton
25.83
88
97,89,142,186
Fluchloraiin
25.88
306
63,326,328,264,65
Mexacarbate
26.02
165
150,134,164,222
4,4-Oxydianiline
26.08
200
108,171,80,65
Butyl benzyl phthalate
26.43
149
91,206
4-Nitrobiphenyl
26.55
199
152,141,169,151
Phosphamidon
26.85
127
264,72,109,138
2-CydohexyJ-4,6-Dinitrophenol
26.87
231
185,41,193,266
Methyl parathion
27.03
109
125,263,79,93
Carbaryl
27.17
144
115,116,201
Dimethylaminoazobenzene
27.50
225
120,77,105,148,42
Propylthiouracil
27.68
170
142,114,83
Benz(a)anthracene
27.83
228
229,226
Chrysene-d12 (IS)
27.88
240
120,236
3,3-Dichlorobenzidine
27.88
252
254,126
Chrysene
27.97
228
226,229
Malathion
28.08
173
125,127,93,158
Kepone
28.18
272
274,237,178,143,270
Fenthion
28.37
278
125,109,169,153
Parathion
28.40
109
97,291,139,155
Anilazine
28.47
239
241,143,178,89
Bis(2-ethyfhexyl) phthalate
28.47
149
167,279
3,3'-Dimethy!benzidine
28.55
212
106,196,180
Carbophenothion
28.58
157
97,121,342,159,199
5-Nitroacenaphthene
28.73
199
152,169,141,115
Methapyrilene
28.77
97
50,191,71
Isodrin
28.95
193
66,195,263,265,147
Captan
29.47
79
149,77,119,117
Chlorfenvinphos
29.53
267
269,323,325,295
Crotoxyphos
29.73
127
105,193,166
Phosmet
30.03
160
77,93,317,76
EPN
30.11
157
169,185,141,323
8270D - 30
Revision 4
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TABLE 1
(continued)
Retention Primary Secondary ion(s)
Compound
Time (min)
Ion

Tetrachlorvinphos
30.27
329
109,331,79,333
Di-n-octyl phthatate
30.48
149
167,43
2-Aminoanthraquinone
30.63
223
167,195
Barban
30.83
222
51,87,224,257,153
Aramite
30.92
185
191,319,334,197,321
Benzo(b)fluoranthene
31.45
252
253,125
Nitrofen
31.48
283
285,202,139,253
Benzo(k)fluoranthene
31.55
252
253,125
Chlorobenzilate
31.77
251
139,253,111,141
Fensuifothion
31.87
293
97,308,125,292
Ethion
32.08
231
97,153,125,121
Diethylstilbestroi
32.15
268
145,107,239,121,159
Famphur
32.67
218
125,93,109,217
Tri-p-tolyi phosphate"
32.75
368
367,107,165,198
Benzo(a)pyrene
32.80
252
253,125
Perylene-d12 (IS)
33.05
264
260,265
7,12-Dimethyibenz(a)anth racene
33.25
256
241,239,120
5,5-Diphenylhydantoin
33.40
180
104,252,223,209
Captafol
33.47
79
77,80,107
Dinocap
33.47
69
41,39
Methoxychlor
33.55
227
228,152,114,274,212
2-Acetyiaminofluorene
33.58
181
180,223,152
4,4'-Methylenebis(2-ch!oroaniline)
34.38
231
266,268,140,195
3,3'-Dimethoxybenzidine
34.47
244
201,229
3-Methyteholanthrene
35.07
268
252,253,126,134,113
Phosalone
35.23
182
184,367,121,379
Azinphos-methyl
35.25
160
132,93,104,105
Leptophos
35.28
171
377,375,77,155,379
Mirex
35.43
2^2
237,274,270,239,235
Tris(2,3-dibromopropyl) phosphate
35.68
201
137,119,217,219,199
Dibenz(a,j)acridine
36.40
279
280,277,250
Mestranol
36.48
277
.310,174,147,242
Coumaphos
37.08
362
226,210,364,97,109
lndeno(1,2,3-cd)pyrene
39.52
276
138,227
Dibenz(a,h)anthracene
39.82
278
139,279
ienzo(g,h,i)perylene
41.43
276
138,277
1,2:4,5-Dibenzopyrene
41.60
302
151,150,300
Strychnine
45.15
334
334,335,333
Piperonyi sulfoxide
46.43
162
135,105,77
Hexachiorophene
47.98
196
198,209,211,406,408
Aidrin
—
66
263,220
Aroclor 1016
—
222
260,292
Arocior 1221
—
190
224,260
8270D-31
Revision 4
January 1998
-------
TABLE 1
(continued)
Compound
Retention
Time (min)
Primary
Ion
Secondary lon(s)
Aroclor 1232
_
190
224,260
Aroclor 1242
_
222
256,292
Aroclor 1248
—
292
362,326
Aroclor 1254
—
292
362,326 -
Aroclor 1260
—
360
362,394
a-BHC
—
183
181,109
p-BHC
—
181
183,109
6-BHC
—
183
181,109
Y-BHC (lindane)
—
183
181,109
4,4'-DDD
_
235
237,165
4,4-DDE
—
246
248,176
4,4-DDT

235
237,165
Dieldrin
—
79
263,279
1,2-Diphenylhydrazine
—
77
105,182
Endosulfan 1
—
195
339,341
Endosuifan II
—
337
339,341
Endosulfan sulfate
—
272
387,422
Endrin

263
82,81
Endrin aldehyde
—
67
345,250
Endrin ketone
—
317
67,319
2-Fluorobiphenyt (surr)
—
172
171
2-Fluorophenol (surr)
—
112
64
Heptachlor
—
100
272,274
Heptachlor epoxide
_
353
355,351
Nitrobenzene-d5 (surr)
_
82
128,54
N-Nitrosodimethylamine
—
42
74,44
Phenol-d6 (surr)
_
99
42,71
Terphenyi-d„ (surr)
—
244
122,212
2,4,6-Tribromophenol (surr)
—
330
332,141
Toxaphene

159
231,233
IS = internal standard
suit = surrogate
Estimated retention times
"Substitute for the non-specific mixture, tricresol phosphate
8270D - 32
Revision 4
January 1998
-------
TABLE 2
ESTIMATED QUANTITATION LIMITS (EQLs) FOR SEMI VOLATILE ORGANICS
Estimated Quantitation Limits8
Ground water Low Soil/Sediment"
Compound
(M9/L)
(M9^g)
Acenaphthene
10
660
Acenaphthyiene
10
660
Acetophenone
10
ND
2-Acetylaminofluorene
20
ND
1-Acetyl-2-thiourea
1000
ND
2-Am inoanthraquinone
20
ND
Aminoazobenzene
10
ND
4-Aminobiphenyl
20
ND
Anilazine
100
ND
o-Anisidine
10
ND
Anthracene
10
660
Aramite
20
ND
Azlnphos-methyl
100
ND
Barban
200
ND
Benz(a)anthracene
10
660
Benzo(b)fIuoranthene
10
660
Benzo(k)fluoranthene
10
660
Benzoic acid
50
3300
Benzo(g,h,i)perylene
10
660
Benzo(a)pyrene
10
660
p-Benzoquinone
10
ND
Benzyl alcohol
20
1300
Bis(2-chloroethoxy)methane
10
660
Bis(2-chioroethyl) ether
10
660
Bis(2-chloroisopropyl) ether
10
660
4-Bromophenyl phenyl ether
10
660
Bromoxynil
10
ND
Butyl benzyl phthalate
10
660
Captafol
20
ND
Captan
50
ND
Carbaryl
10
ND
Carbofuran
10
ND
Carbophenothion
10
ND
Chlorfenvinphos
20
ND
4-Chloroaniline
20
1300
Chlorobenzilate
10
ND
5-Chloro-2-methylaniline
10
ND
8270D - 33
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits"
Ground water Low Soil/Sediment"
Compound (pg/L) (yjg/kg)
4-Chloro-3-methylphenol
20
1300
3-(Chloromethyl)pyridine hydrochloride
100
ND
2-Chloronaphthalene
10
660
2-Chlorophenol
10
660
4-Chlorophenyl phenyl ether
10
660
Chrysene
10
660
Coumaphos
40
ND
p-Cresidine
10
ND
Crotoj^phos
20
ND
2-CycIohexyM,6-dinitrophenol
100
ND
Demeton-O
10
ND
Demeton-S
10
ND
Vallate (ds or trans)
10
ND
Vallate (trans or ds)
10
ND
2,4-Diaminotofuene
20
ND
Dibenz(aj)acridine
10
ND
Dibenz(a,h)anthracene
10
660
Dibenzofuran
10
660
Dibenzo(a,e)pyrene
10
ND
Di-n-butyl phthalate
10
ND
Dichlone
NA
ND
1,2-Dichlorobenzene
10
660
1,3-Dichlorobenzene
10
660
1,4-Dichlorobenzene
10
660
3,3'-Dichlorobenzidine
20
1300
2,4-Dichlorophenol
10
660
2,6-Dichlorophenol
10
ND
Dichlorovos
10 "
ND
Dicrotophos
10
ND
Diethyl phthalate
10
660
Diethyistilbestrol
20
ND
Diethyl sulfate
100
ND
Dimethoate
20
ND
3,3-Dimethoxybenzidine
100
ND
Dimeth^amino^obenzene
10
ND
7,12-Dimethylbenz(a)anthracene
10
ND
3,3'-Dimethylbenzidine
10
ND
2,4-Dimethylphenol
10
660
827QD - 34
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits8
Ground water Low Soil/Sediment"
Compound
(Mg/L)
(ug/kg)
Dimethyl phthaiate
10
660
1,2-Dinitrobenzene
40
ND
1,3-Dinitrobenzene
20
ND
1,4-Dinitrobenzene
40
ND
4,6-Dinitro-2-methylphenol
50
3300
2,4-Dinitrophenol
50
3300
2,4-DWtrotoluene
10
660
2,6-Dinitrotoluene
10
660
Dinocap
100
ND
Dinoseb
20
ND
5,5-Diphenylhydantoin
20
ND
K-n-octyl phthaiate
10
660
Disulfoton
10
ND
EPN
10
ND
Ethion
10
ND
Ethyl carbamate
50
ND
Bis(2-ethyihexy0 phthaiate
10
660
Ethyl methanesulfonate
20
ND
Famphur
20
ND
Fensulfothion
40
ND
Fenthion
10
ND
Fluchloralin
20
ND
Fluoranthene
10
660
Fluorene
10
660
Hexachlorobenzene
10
660
Hexachlonobutadiene
10
660
Hexachlorocydopentadiene
10
660
Hexachloroethane
10
660
Hexachlorophene
50
ND
Hexachloropropene
10
ND
Hexamethylphosphoramide
20
ND
lndeno(1,2,3-cd)pyrene
10
660
isodrin
20
ND
fsophorone
10
660
Isosafrole
10
ND
Kepone
20
ND
Leptophos
10
ND
Mafathion
50
ND
8270D - 35
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits"
Ground water Low Soil/Sediment*
Compound
(pg/L)
(pg/kg)
Mestranol
20
ND
Methapyrilene
100
ND
Methoxychlor
10
ND
3-Methylcholanthrene
10
ND
Methyl methanesulfonate
10
ND
2-Wethyinaphthalene
10
660
Methyi parathion
10
ND
2-Methylphenol
10
660
3-Methyiphenol
10
ND
4-Methyiphenol
10
660
Mevinphos
10
ND
Mexacarbate
20
ND
Mirex
10
ND
Monocrotophos
40
ND
Naled
20
ND
Naphthalene
10
660
1,4-Naphthoquinone
10
ND
1-Naphthylamine
10
ND
2-Naphthylamine
10
ND
Nicotine
20
ND
5-Nitroacenaphthene
10
ND
2-Nitroaniline
50
3300
3-Nitroaniline
50
3300
4-Nitroaniline
20
ND
5-Nitro-o-anisidine
10
ND
Nitrobenzene
10
660
4-Nitrobiphenyi
10
ND
Nitrofen
20
ND
2-Nitrophenol
10
660
4-Nitrophenol
50
3300
5-Nitro-o-toluidine
10
ND
4-Nitroquinoline-1 -oxide
40
ND
N-Nitrosodi-n-butylamine
10
ND
N-Nitrosodiethylamine
20
ND
N-Nitrosodiphenylamine
10
660
N-Nitroso-di-n-propylamine
10
660
N-Nitrosopiperidine
20
ND
N-Nitrosopyrrolidine
40
ND
8270D-36
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits'
Ground water Lew Soil/Sediment"
Compound
(m'L)
(Ma/kg)
Octamethyl pyrophosphoramide
200
ND
4,4-Oxydianiline
20
ND
Parathion
10
ND
Pentachlorobenzene
10
ND
Pentachloronitrobenzene
20
ND
Pentachlorophenol
50
3300
Phenacetin
20
ND
Phenanthrene
10
660
Phenobarbital
10
ND
Phenol
10
660
1,4-Phenylenediamine
10
ND
Phorate
10
ND
Phosaione
100
ND
Phosmet
40
ND
Phosphamidon
100
ND
Phthalic anhydride
100
ND
2-Picoline
ND
ND
Piperonyl sulfoxide
100
ND
Pronamide
10
ND
Propylthiouracil
100
ND
Pyrene
10
660
Resorcinol
100
ND
Safrole
10
ND
Strychnine
40
ND
Sulfallate
10
ND
Terbufos
20
ND
1,2,4,5-Tetrachlorobenzene
10
ND
2,3,4,6-Tetrachlorophenol
10
ND
Tetrachtorvinphos
20
ND
Tetraethyl pyrophosphate
40
ND
Thionazine
20
ND
Thiophenol (Benzenethiol)
20
ND
o-Toluidine
10
ND
1,2,4-T richlorobenzene
10
660
2,4,5-Trichlorophenol
10
660
2,4,6-Trichlorophenol
10
660
Trifluraiin
10
ND
2,4,5-T rimethylaniline
10
ND
8270D - 37
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits®
Compound
Ground water
(M9/L)
Low Soil/Sediment"
W*g)
Trimethyl phosphate
1,3,5-Tririrtro benzene
T ris(2,3-dibromopropyl) phosphate
Tri-p-tolyl phosphate(h)
10
10
200
10
ND
ND
ND
ND
* Sample EQLs are highly matrix-dependent. The EQLs listed here are provided for guidance and
may not always be achievable.
b EQLs listed for soil/sediment are based on wet weight Normally, data are reported on a dry
weight basis, therefore, EQLs will be higher based on the % dry weight of each sample. These
EQLs are based on a 30-g sample and gel permeation chromatography cleanup.
ND = Not Determined
NA = Not Applicable
Other Matrices Factor*
High-concentration soil and sludges by ultrasonic extractor 7.5
Non-water miscible waste 75
®EQL »(EQL for Low Soil/Sediment given above in Table 2) x (Factor)
8270D-38
Revision 4
January 1998
-------
TABLE 3
DFTPP KEY IONS AND ION ABUNDANCE CRITERIA4 "
Mass
Ion Abundance Criteria
51
30-60% of mass 198
68
< 2% of mass 69
70
<2% of mass 69
127
40-60% of mass 198
197
< 1% of mass 198
198
Base peak, 100% relative abundance
199
5-9% of mass 198
275
10-30% of mass 198
365
> 1% of mass 198
441
Present but less than mass 443
442
>40% of mass 198
443
17-23% of mass 442
a Data taken from Reference 3.
6 Alternate tuning criteria may be employed,
(e.g., CLP, Method 525, or manufacturers'
instructions), provided that method
performance is not adversely affected.
TABLE 4
CALIBRATION CHECK COMPOUNDS (CCC)
Base/Neutral Fraction
Acid Fraction
Acenaphthene
1,4-Diehloro benzene
Hexachlorobutadiene
Diphenytamine
Di-n-octyl phthalate
Fluoranthene
Benzo
-------
TABLE 5
SEM!VOLATILE INTERNAL STANDARDS WITH CORRESPONDING ANALYTES
ASSIGNED FOR QUANTITATION
1,4-Dichlorobenzene-d4
Naphthalene-d8
Acenaphthene-d10
Aniline
Acetophenone
Acenaphthene
Benzyl alcohol
Benzoic acid
Acenaphthylene
Bis(2-chloroethyi) ether
Bis(2-chloroethoxy)methane
1 -Chioronaphthalene
Bis(2-chloroisopropyf) ether
4-Chloroaniline
2-ChloronaphthaIene
2-Chlorophenol
4-Chloro-3-methylphenol
4-ChIorophenyl phenyl ether
1,3-Dichlorobenzene
2,4-Dichbrophenol
Dibenzofuran
1,4-Dichlorobenzene
2,6-Dichiorophenol
Diethyl phthalate
1,2-Dichloro benzene
ata-Dimethyl-
Dimethyl phthalate
Ethyl methanesulfonate
phenethylamine
2,4-Dinrtrophenol
2-Fluorophenol (surr)
2,4-Dimethylphenol
2,4-Dinitrotoluene
Hexachloroethane
Hexachlorobutadiene
2,6-Dinitrotoluene
Methyl methanesulfonate
Isophorone
Fluorene
2-Methylphenol
2-Methyinaphthalene
2-Fluorobipheriyi (surr)
4-Methylphenol
Naphthalene
Hexachiorocyclopentadiene
N-Nitrosodimethylamine
Nitrobenzene
1-Naphthylamine
N-Nitroso-di-n-propylamine
Nitrobenzene-da (surr)
2-Naphthylamine
Phenol
2-Nitrophenol
2-Nitroaniline
Phenol-d6 (surr)
N-Nitrosodi-n-butylamine
3-Nitroaniline
2-Picofine
N-Nitrosopiperidine
4-Nitroaniline

1,2,4-Trichlorobenzene
4-Nitrophenol
Pentachlorobenzene
1.2.4.5-Tetrachlorobenzene
2.3.4.6-Tetrachloropheno!
2.4.5-Tribromophenol (suit)
2.4.6-Trichlorophenol
2,4.5-Trichlorophenol
(suit) = surrogate
8270D - 40
Revision 4
January 1998
-------
TABLE 5
(continued)
Phenanthrene-d
10
Chrysene-d12
Perylene-d12
4-Aminobiphenyl
Anthracene
4-Bromophenyl phenyl ether
Di-n-butyl phthalate
4,6-Dinitro-2-methylphenol
Diphenylamine
Fluoranthene
Hexachlorobenzene
N-Nitrosodiphenylamine
Pentachlorophenol
Pentachloronitrobenzene
Phenacetin
Phenanthrene
Pronamide
Benzidine
Benzo(a)anthracene
Bis(2-ethylhexyl) phthalate
Butyl benzyl phthalate
Chrysene
3,3'-DichIorobenzidine
p-Dimethyl aminoazobenzene
Pyrene
Terphenyl-d14 (suit)
7,12-Oimethylbenz(a)
anthracene
Di-rv-octyl phthalate
lndeno(1,2,3-cd) pyrene
3-Methylcholanthrene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(g,h,i)perylene
Benzo(a)pyrene
Dibenz(a,j)acridine
Dibenz(a,h)anthracene
(suit) = surrogate
8270D-41
Revision 4
January 1998
-------
TABLE 6
MULTILABORATORY PERFORMANCE DATA"
L-I.ll ——-_1
Compound
Test conc.
(pg/L)
Limit for
s (pg/L)
Range for x
(pg/L)
Range
P« Ps C%)
Acenaphthene
100
27.6
60.1-132.3
47-145
Acenaphthylene
100
40.2
53.5-126.0
33-145
Aldrin
100
39.0
7.2-152.2
D-166
Anthracene
100
32.0
43.4-118.0
27-133
Benz(a)anihracene
100
27.6
41.8-133.0
33-143
Benzo(b)fluoranthene
100
38.8
42.0-140,4
24-159
Benzo(k)fluoranthene
100
32.3
25.2-145.7
11-162
Benzo(a)pyrene
100
39.0
31.7-148.0
17-163
Benzo(g,h,i)perylene
100
58.9
D-195.0
D-219
Benzyl butyl phthaiate
100
23.4
D-139.9
D-152
P-BHC
100
31.5
41.5-130.6
24-149
5-BHC
100
21.6
D-100.0
D-110
Bis(2-chloroethyl) ether
100
55.0
42.9-126.0
12-158
Bis(2-ehioroethoxy)methane
100
34.5
49.2-164.7
33-184
Bis(2-chloroisopropyl) ether
100
46.3
62.8-138.6
36-166
Bis(2-ethylhexyl) phthaiate
100
41.1
28.9-136.8
8-158
4-Bromophenyl phenyl ether
100
23.0
64.9-114.4
53-127
2-Chloronaphfhaiene
100
13.0
64.5-113.5
60-118
4-Chlorophenyl phenyl ether
100
33.4
38.4-144.7
25-158
Chrysene
100
48.3
44.1-139.9
17-168
4,4-DDD
100
31.0
D-134,5
D-145
4,4'-DDE
100
32.0
19.2-119.7
4-136
4,4-DDT
100
61.6
D-170.6
D-203
Dibenzo(a, h)anthracene
100
70.0
D-199.7
D-227
Di-n-butyl phthaiate
100
16.7
8.4-111.0
1-118
1,2-Dichloro benzene
100
30.9
48.6-112.0
32-129
1,3-Dichlorobenzene
100
41.7
16.7-153.9
D-172
1,4-Dichlorobenzene
100
32.1
37.3-105.7
20-124
3,3'-Dichlorobenzidine
100
71.4
8.2-212.5
D-262
Dieldrin
100
30.7
44.3-119.3
29-136
Diethyl phthaiate
100
26.5
D-100.0
D-114
Dimethyl phthaiate
100
23.2
D-100.0
D-112
2,4-Dinitrotoluene
100
21.8
47.5-126,0
39-139
2,6-Dinitrotoluene
100
29.6
68.1-136.7
50-158
Di-n-octyl phthaiate
100
31.4
18,6-131.8
4-146
Endosulfan sulfate
100
16.7
D-103.5
D-107
Endrin aldehyde
100
32.5
D-188.8
D-209
Fluoranthene
100
32.8
42.9-121.3
26-137
8270D - 42
Revision 4
January 1998
-------
TABLE 6
(continued)
Compound
Test conc.
(M0/L)
Limit for
s (M0/L)
Range for x
(pg/L)
Range
P> P«t%)
Fluorene
100
20.7
71.6-108.4
59-121
Heptachlor
100
37.2
D-172,2
D-192
Heptachior epoxide
100
54.7
70.9-109.4
26.155
Hexachlorobenzene
100
24.9
7.8-141.5
- D-152
Hexachlorobutadiene
100
26.3
37.8-102.2
24-116
Hexachloroethane
100
24.5
55.2-100,0
40-113
indeno(1,2,3~cd)pyrene
100
44.6
D-150.9
D-171
Isophorone
100
63.3
46.6-180.2
21-196
Naphthalene
100
30.1
35.6-119.6
21-133
Nitrobenzene
100
39.3
54.3-157.6
35-180
N-Nitrosodi-n-propylamine
100
55.4
13.6-197.9
D-230
Aroclor 1260
100
54.2
19.3-121.0
D-164
Phenanthrene
100
20.6
65.2-108.7
54-120
Pyrene
100
25.2
69.6-100.0
52-115
1,2,4-Trichlorobenzene
100
28.1
57.3-129.2
44-142
4-Chloro-3-methylphenol
100
37.2
40.8-127.9
22-147
2-Chlorophenol
100
28.7
36.2-120.4
23-134
2,4-Chlorophenoi
100
26.4
52.5-121.7
39-135
2,4-Dimethyiphenol
100
26.1
41.8-109.0
32-119
2,4-Dinitrophenol
100
49.8
D-172.9
CM 91
2-MethyM,6-dinitrophenol
100
93.2
53.0-100.0
D-181
2-Nitrophenol
100
35.2
45.0-166.7
29-182
4-Nitrophenol
100
47.2
13.0-106.5
D-132
Pentachlorophenol
100
48.9
38.1-151.8
14-176
Phenol
100
22.6
16.6-100.0
5-112
2,4,6-T richlorophenol
100
31.7
52.4-129.2
37-144
s = Standard deviation of four recovery measurements, in pg/L
x = Average recovery for four recovery measurements, in pg/L
p, p, = Measured percent recovery
D = Detected; result must be greater than zero
3 Criteria from 40 CFR Part 136 for Method 625, using a packed GC column. These criteria are
based directly on the method performance data in Table 7. Where necessary, the limits for
recovery have been broadened to assure applicability of the limits to concentrations below those
used to develop Table 7. These values are for guidance only. Appropriate derivation of
acceptance criteria for capillar/ columns should result in much narrower ranges. See Method 8000
for information on developing and updating acceptance criteria for method performance.
8270D - 43
Revision 4
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TABLE 7
METHOD ACCURACY AND PRECISION AS FUNCTIONS OF CONCENTRATION*

Accuracy, as
Single analyst
Overall precision,
Compound
recovery, x* (ng/L)
precision, sr' (pg/L)
S* (pg/L)
Acenaphthene
0.96C+0.19
0.15x-0.12
0.21X-0.67
Acenaphthyfene
0.89C+0.74
0.24X-1.06
0.26x-0.54
Aldrin
0.78C+1.66
0.27X-1.28
0.43x+1.13
Anthracene
0.80C+0.68
0.21x-0.32
0.27x-0.64
Benz(a)anthracene
0.88C-0.60
0.1SX+0.93
0.26X-0.21
Benzo(b)fluoranthene
0.93C-1.80
0.22X+0.43
0.29X+0.96
Benzo(k)fluoranth0ne
0.87C-1.56
0.19X+1.03
0.35X+0.40
Benzo(a)pyrene
0.90C-0.13
0.22X+0.48
0.32x+1.35
Benzo(g,h,i)perylene
0.98C-0.86
0.29x+2.40
0.51X-0.44
Benzyl butyl phthalate
0.66C-1.68
0.18X+0.94
0.53x+0.92
P-BHC
0.87C-0.94
0.20X-0.58
0.30X+1.94
6-BHC
0.29C-1.09
0.34x+0.86
0.93X-0.17
Bis(2-chloroethyl) ether
0.86C-1.54
0.35X-0.99
0.35x+0.10
Bis(2-ch!oroetho)ty)methane
1.12C-5.04
0.16X+1.34
0.26X+2.01
Bis(2-chloroisopropyl) ether
1.0302.31
0.24X+0.28
0.25 x+1.04
Bis(2-ethylhexyl) phthalate
0.84C-1.18
0.26X+0.73
0.36X+0.67
4-Bromophenyl phenyl ether
0.91C-1.34
0.13X+0.66
0.16X+0.66
2-Chloronaphthatene
0.89C+0.01
O.OTx+O.52
0.13X+0.34
4-Chlorophenyl phenyl ether
0.91C+0.53
0.20X-0.94
0.30X-0.46
Chrysene
0.93C-1.00
0.28x+0.13
0.33x-0.09
4,4'-DDD
0.56C-0.40
0.29X-0.32
0.66X-0.96
4,4-DDE
0.70C-0.54
0.26x-1.17
0.39X-1.04
4,4'-DDT
0.79C-3.28
0.42x+0,19
0.65x-0.58
Dibenzo(a, h)anthracene
0.88C+4.72
0.30x+8.51
0.59x+0.25
Di-n-butyl phthalate
0.59C+0.71
0.13X+1.16
0.39X+0.60
1,2-Dichlorobenzene
0.80C+0.28
0.20x+0.47
0.24x+0.39
1,3-Dichlorobenzene
0.86C-0.70
0.25x+0.68
0.41x+0.11
1,4-Dichlorobenzene
0.73C-1.47
0.24X+0.23
0.29X+0.36
3,3'-Dichloro benzidine
1.23C-12.65
G.28x+7.33 -
0.47X+3.45
Dieldrin
0.82C-0.16
0.20X-0.16
0.26x-0.07
Diethyl phthalate
0.43C+1.00
0.28 x+1.44
0.52x+0.22
Dimethyl phthalate
0.20C+1.03
0.54x+0.19
1.05X-0.92
2,4-Dinitrotoluene
0.92C-4.81
0.12x+1.06
0.21X+1.50
2,6-Dinitrotoluene
1.06C-3.60
0.14X+1.26
0.19X+0.35
Di-n-octyl phthalate
0.76C-0.79
0.21X+1.19
0.37X+1.19
Endosulfan sulfate
0.39C+0.41
0.12X+2.47
0.63x-1.03
Endrln aldehyde
0.76C-3.86
0.18X+3.91
0.73X-0.62
Fluoranthene
0.81C+1.10
0.22x-0.73
0.28x-0.60
Fluorene
0.90C-0.00
0.12X+0.26
0.13X+0.61
8270D-44
Revision 4
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TABLE 7
(continued)
Compound
Accuracy, as
recovery, x" (pg/L)
Single analyst
precision, sr' (pg/L)
Overall precision,
S'(Mg/L)
Heptachlor
0.87C-2.97
0.24x-0.56
0.50X-0.23
Heptachlor epoxide
0.92C-1.87
0.33x-0.46
0.28X+0.64
Hexachlorobenzene
0.74C+0.66
0.18x-0.10
0.43X-0.52
Hexachlorobutadiene
0.71C-1.01
0.19x+0.92
0.26X+0.49
Hexachtoroethane
0.73C-0.83
0.17X+0.67
0.17X+0.80
lndeno(1,2,3-cd)pyrene
0.78C-3.10
0.29X+1.46
0.50x-0.44
Isophorone
1.12C+1.41
0.27X+0.77
0.33X+0.26
Naphthalene
0.76C+1.58
0.21X-0.41
0.30x-0.68
Nitrobenzene
1.0903.05
0.19X+0.92
0.27X+0.21
N-Nitrosod»-n-propylamine
1.12C-6.22
0.27X+0.68
0.44X+0.47
Aroclor 1260
0.81C-10.86
0.35x+3.61
0.43X+1.82
Phenanthrene
0.87C+0.06
0.12X+0.57
0.15X+0.25
Pyrene
0.84C-0.16
0.16x+0.06
0.15X+0.31
1,2,4-Trichlorobenzene
0.94C-0.79
0.15x+0.85
0.21X+0.39
4-Chloro-3-methylphenol
0.84C+0.35
0.23X+0.75
0.29X+1.31
2-Chterophenol
0.78C+0.29
0.18X+1.46
0.28X+0.97
2,4-Dichlorophenol
0.87C-0.13
0.15X+1.25
0.21X+1.28
2,4-Dimethylphenol
0.71C+4.41
0.16X+1.21
0.22X+1.31
2,4-Dinitrophenol
0.81C-18.04
0.38X+2.36
0.42x+26.29
2-MethyM,6-dinitrophenol
1.04C-28.04
0.10X+42.29
0.26X+23.10
2-Nitrophenol
0.07C-1.15
0.16x+1.94
0.27x+2.60
4-Nitrophenol
0.61C-1.22
0.38X+2.57
0.44X+3.24
Pentachlorophenol
0.93C+1.99
0.24X+3.03
0.30X+4.33
Phenol
0.430+1.26
0.26X+0.73
0.35X+0.58
2,4,6-T richlorophenol
0.91C-0.18
0,16x+2.22
0.22 x+1.81
x' = Expected recovery for one or more measurements of a sample containing a concentration of
C, in pg/L.
sr' = Expected single analyst standard deviation of measurements at an average concentration of
x, in pg/L
S' = Expectedjnteriaboratory standard deviation of measurements at an average concentration
found of x, in pg/L.
C = True value for the concentration, in mq^I--
x = Average recovery found for measurements of samples containing a concentration of C, in
pg/L
• Criteria from 40 CFR Part 136 for Method 625, using a packed GC column. These criteria are
based directly on the method performance data in Table 7. These values are for guidance
only. Appropriate derivation of acceptance criteria for capillary columns should result in much
narrower ranges. See Method 8000 for information on developing and updating acceptance
criteria for method performance.
8270D - 45
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TABLE 8
EXTRACTION EFFICIENCY AND AQUEOUS STABILITY RESULTS
Percent Recovery, Day 0 Percent Recovery, Day 7
Compound
Mean
RSD
Mean
RSD
3-Amino-9-ethylcart)azole
80
8
73
3
4-Chloro-1,2-phenylenediamine
91
1
108
4
4-Chloro-1,3-phenylenediamine
84
3
70
3
1,2-Dibromo-3-chloropropane
97
2
98
5
Dinoseb
99
3
97
6
Parathion
100
2
103
4
4,4'-MethyIenebis(N, N-dimethylaniline)
108
4
90
4
t
5-Nitro-o-toluidine
99
10
93
4
2-Picoline
80
4
83
4
Tetraethyl d'rthiopyrophosphate
92
7
70
1
Data taken from Reference 6.
8270D - 46
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TABLE 9
MEAN PERCENT RECOVERIES AND PERCENT RSD VALUES FOR SEMI VOLATILE ORGANICS
FROM SPIKED CLAY SOIL AND TOPSOIL BY AUTOMATED SOXHLET EXTRACTION
(METHOD 3541) WITH HEXANE-ACETONE (1:1)a

Clay Soil

Topsoil

Mean
RSD
Mean
RSD
Compound
Recovery

Recovery

1,3-Dichlorobenzene
0
—
0
—
1,2-Dichlorobenzene
0
—
0
—
Nitrobenzene
0
—
0
--
ienzal chloride
0
—
0

Benzotrichloride
0
—
0
—
4-Chloro-2-nitrotoluene
0

0
—
Hexachlorocyclopentadie ne
4.1
15
7.8
23
2,4-Dichloronitrobenzene
35.2
7.6
21.2
15
3,4-Dichioronitrobenzene
34.9
15
20.4
11
Pentachlorobenzene
13.7
7.3
14.8
13
2,3,4,5-Tetrachloronitrobenzene
55.9
6.7
50.4
6.0
Benefin
62.6
4.8
62.7
2.9
alpha-BHC
58.2
7.3
54.8
4.8
Hexachlorobenzene
26.9
13
25.1
5.7
delta-BHC
95.8
4.6
99.2
1.3
Heptachlor
46.9
9.2
49.1
6.3
Aldrin
97.7
12
102
7.4
Isopropalin
102
4.3
105
2.3
Heptachlor epoxide
90.4
4.4
93.6
2.4
trans-Chlordane
90.1
4.5
95.0
2.3
Endosulfan!
96.3
4.4
101
2.2
Dieldrin
129
4.7
104
1.9
2,5-Dichlorophenyl-4-nitrophenyl ether
110
4.1
112
2.1
Endrin
102
4.5
106
3.7
Endosulfan II
104
4.1
105
0.4
p.p'-DDT
134
2.1
111
2.0
2,3,6-T richlorophenyl-4'-nitrophenyl ether
110
4.8
110
2.8
2,3,4-Trichlorophenyl-4'-nitrophenyl ether
112
4.4
112
3.3
Mirex
104
5.3
108
2.2
a The operating conditions for the Soxtec apparatus were as follows: immersion time 45 min;
extraction time 45 min; the sample size was 10 g; the spiking concentration was 500 ng/g, except
for the surrogate compounds at 1000 ng/g, 2,5-Dichlorophenyl-4-nitrophenyl ether, 2,3,6-
TrichiorophenyI-4-nitrophenyI ether, and 2,3,4-TrichlorophenyI-4-nitrophenyI ether at 1500 ng/g,
Nitrobenzene at 2000 ng/g, and 1,3-Dichlorobenzene and 1,2-Dichlorobenzene at 5000 ng/g.
8270D - 47
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TABLE 10
SINGLE LABORATORY ACCURACY AND PRECISION DATA FOR THE EXTRACTION
OF SEMIVOLATILE ORGANICS FROM SPIKED CLAY BY
AUTOMATED SOXHLET (METHOD 3541)a
Compound
Mean Recovery
RSD
Phenol
47.8
5.6
Bis(2-chloroethyl)ether
25.4
13
2-ChIorophenoI
42.7
4.3
Benzyl alcohol
55.9
7.2
2-Methyiphenol
17.6
6.6
Bis(2-chloroisopropyl)ether
15.0
15
4-MethyiphenoI
23.4
6.7
N-Nitroso-di-n-propy!ami ne
41.4
6.2
Nitrobenzene
28.2
7.7
Isophorone
56.1
4.2
2-Nitrophenol
36.0
6.5
2,4-Dimethylphenol
50.1
5.7
Benzoic acid
40.6
7.7
Sls(2-chloroethoxy)methane
44.1
3.0
2,4-DichIorophenol
55.6
4.6
1,2,4-T richlorobenzene
18.1
31
Naphthalene
26.2
15
4-Chloroaniline
55.7
12
4-Ch loro-3-m ethylph enol
65.1
5.1
2-Methyfnaphthalene
47.0
8.6
Hexachlorocyclopentadiene
19.3
19
2,4,6-T richlorophenol
70.2
6.3
2,4,5-Trichlorophenol
26.8
2.9
2-Chloronaphthalene
61.2
6.0
2-NilroaniIine
73.8
6.0
Dimethyl phthalate
74.6
5.2
Acenaphthylene
71.6
5.7
3-NitroaniIine
77.6
5.3
Acenaphthene
79.2
4.0
2,4-Dinitrophenol
91.9
8.9
4-Nitrophenol
62.9
16
Dibenzofuran
82.1
5.9
2,4-Dinitrotoluene
84.2
5.4
2,6-Dinitrotoluene
68.3
5.8
Diethyl phthalate
74.9
5.4
4-ChIorophenyl-phenyl ether
67.2
3.2
Fluorene
82.1
3.4
8270D - 48
Revision 4
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TABLE 10
(continued)
Compound
Mean Recovery
RSD
4-Nitroaniline
79.0
7.9
4,6-Dinitro-2-methylphenoI
63.4
6.8
N-Nitrosodiphenylamine
77.0
3.4
4-Bromophenyl-phenyl ether
62.4
3.0
Hexachlorobenzene
72.6
3.7
Pentachlorophenol
62.7
6.1
Phenanthrene
83.9
5.4
Anthracene
96.3
3.9
Di-n-butyl phthalate
78.3
40
Fluoranthene
87.7
6.9
Pyrene
102
0.8
Butyl benzyl phthalate
66.3
5.2
3,3-Dichlorobenzidine
25.2
11
Benzo{a)anthracene
73.4
3.8
Bis(2-ethy1hexyl) phthalate
77.2
4.8
Chrysene
76.2
4.4
Di-n-octyl phthalate
83.1
4.8
Benzo(b)fluoranthene
82.7
5.0
Benzo(k)fluoranthene
71.7
4.1
Benzo(a)pyrene
71.7
4.1
lndeno(1,2,3-cd)pyrene
72.2
4.3
Dibenzo(a,h)anthracene
66.7
6.3
Benzo(g,h,i)perylene
63.9
8.0
1,2-Dichlorobenzene
0
—
1,3-Dichlorobenzene
0
—
1,4-Dichlorobenzene
0
-
Hexachloroethane
0
_
Hexachlorobutadiene
0
—
8 Number of determinations was three. The operating conditions for the Soxtec apparatus were
as follows: immersion time 45 min; extraction time 45 min; the sample size was 10 g clay soil; the
spike concentration was 6 mg/kg per compound. The sample was allowed to equilibrate 1 hour
after spiking.
Data taken from Reference 7.
8270D-49
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TABLE 11
PRECISION AND BIAS VALUES FOR METHOD 35421
Compound
Mean Recovery
Standard Deviation
% RSD
2-Fluorophenol
74.6
28.6
38.3
Phenol-d5
77,8
27.7
35.6
N itrob enzene-d 5
65.6
32.5
49.6
2-Fluorobiphenyl
75.9
30.3
39.9
2,4,6-Tribromophenol
67.0
34.0
50.7
Terphenyl-d14
78.6
32.4
41.3
1 The surrogate values shown in Table 11 represent mean recoveries for surrogates In all Method
0010 matrices in a field dynamic spiking study.
8270D - 50
Revision 4
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TABLE 12
ACCELERATED SOLVENT EXTRACTION (METHOD 3545) RECOVERY VALUES
AS PERCENT OF SOXTEC™

Clav

Loam

Sand

Mean
Compound
Low
Mid
High
Low
Mid
High
Low .
Mid
High
Roc.
Phenol
93.3
78.7
135.9
73.9
82.8
124,6
108.8
130.6
89.7
102.0
Bis(2-ch!oroethyl) ether
102.1
85.1
109.1
96.0
88.0
103.6
122.3
119.9
-90.8
101.9
2-Chlorophenoi
100.8
82.6
115.0
93.8
88.9
111.1
115.0
115.3
91.9
101.6
1,3-Dichlorobenzene
127.7
129.7
110.0
*364.2
129.9
119,0
*241.3
*163.7
107.1
120.6
1,4-Dichlorobenzene
127.9
127.0
110.5
•365.9
127.8
116.4
*309.6
*164.1
105.8
119.2
1,2-Dichlorobenzene
116.8
115.8
101.3
•159.2
113.4
105.5
*189.3
134.0
100.4
112.5
2-Methytphenol
98.9
82.1
119.7
87.6
89.4
111.0
133.2
128.0
92.1
104.7
Bfe(2-chloro«opropyt)ether
109.4
71.5
108.0
81.8
81.0
88.6
118.1
148.3
94.8
100.2
o-Toluidine
100.0
89.7
117.2
100.0
*1525
120.3
100.0
*199.5
102.7
110.3
N-Nitroso-di-n-propylamine
103.0
79.1
107.7
B3.9
88.1
96.2
109.9
123.3
91.4
98.1
Hexachloroethane
97.1
125.1
111.0
*245.4
117.1
128.1
*566,7
147.9
103,7
118.6
Nitrobenzene
104.8
82.4
106,6
86.8
84.6
101.7
119.7
122.1
93.3
100.2
Isophorone
100.0
86.4
98.2
87.1
87.5
109.7
135.5
118,4
92.7
101.7
2,4-Dimethylphenol
100.0
104.5
140.0
100.0
114.4
123.1
100.0
*180.6
96.3
109.8
2-ffitrophenol
80.7
80.5
107.9
91.4
86.7
103.2
122.1
107.1
87.0
96.3
B's(ch[oroethoxy)methane
94.4
80.6
94.7
86.5
84.4
99.6
130.6
110.7
93.2
97.2
2,4-Dichloropheno)
88.9
87.8
111.4
85.9
87.6
103.5
123.3
107.0
92.1
98.6
1,2,4-T richlorobenzena
98.0
97.8
98.8
123.0
93.7
94.5
137.0
99.4
95.3
104.2
Naphthalene
101.7
97.2
123.6
113.2
102.9
129.5
*174.5
114.0
89.8
106.1
4-Chloroanine
100.0
"150.2
*162.4
100.0
125.5
*263.6
100.0
*250.8
114.9
108.1
Hexachlorobutadiene
101.1
98.7
102.2
124.1
90.3
98.0
134.9
96.1
96.8
104.7
4-Chtoro-3-methylphenoI
90.4
80.2
114.7
79.0
85.2
109.8
131.6
116.2
90.1
99.7
2-Methylnaphthalene
93.2
89.9
94.6
104.1
92.2
105.9
146.2
99.1
93.3
102.1
H exachiorocyclopentadiene
100.0
100.0
0.0
100.0
100.0
6.8
100.0
100.0
*238.3
75.8
2,4,6-T richloropheriol
94.6
90.0
112.0
84.2
91.2
103.6
101.6
95.9
89.8
95.9
2,4,5-T richlorophenoi
84.4
91.9
109.6
96.1
80.7
103.6
108.9
83.9
87.9
94.1
2-Chloronaphtha!ene
100.0
91.3
93.6
97.6
93.4
98.3
106.8
93.0
92.0
96.2
2-Nitroanitine
90.0
83.4
97.4
71.3
88.4
89.9
112.1
113.3
87.7
92.6
2,6-Dinitrotoluene
83.1
90.6
91.6
86.4
90.6
90.3
104.3
84.7
90.9
90.3
Acenaphthylene
104.9
95.9
100.5
99.0
97,9
108.8
118.5
97,8
92.0
101.7
3-Nitroaniline
*224.0
115.6
97.6
100.0
111.8
107.8
0.0
111.7
99.0
92.9
Acenaphthene
1(2.1
92.6
97.6
97.2
96.9
104,4
114.2
92.0
89.0
98.4
4-Nitrophend
0.0
93.2
121.5
18.1
87.1
116.6
69.1
90.5
84.5
75.6
2,4-Dinitrotoluene
73.9
91.9
100.2
84.7
93.8
98.9
100.9
84.3
67.3
90.7
S270D - 51
Revision 4
January 199B
-------
TABLE 12
(continued)

Clav

Loam

Sand

Mean
Compound
Low
Mid
High
Low
Mid
High
Low
Mid
High
Rec.
Dibenzofuran
89.5
91.7
109.3
98.5
92.2
111.4
113,8
92.7
90.4
98.8
4-Chloropheny) phenyl
83,0
94.5
88.7
95.7
94.3
94.2
111.4
87.7
90.3
94.4
ether

Fluorene
85.2
94.9
89.2
102.0
95.5
93.8
121.3
85.7
90.9
95.4
4-Nftroaniline
77.8
114.8
94.5
129.6
103.6
95.4
*154.1
89,3
87.5
99.1
N-Nitrosodiphenylamine
82.6
96.7
93.8
92.9
93.4
116.4
97.5
110.9
86.7
96.8
4-Bromophenyl phenyl
85.6
92.9
92.8
91.1
107.6
89.4
118.0
97.5
87.1
95.8
ether

Hexachlarobenzene
95.4
91.7
92.3
95.4
93.6
83.7
106.8
94.3
90.0
93.7
Pentachlorophenol
68.2
85.9
107.7
53.2
89.8
88.1
96.6
59.8
81.3
81.2
Phenanthrene
92.1
93,7
93.3
100.0
97.8
113.3
124.4
101.0
89.9
100.6
Anthracene
101.6
95.0
93.5
92.5
101.8
118.4
123.0
94.5
90.6
101.2
Carbazole
94.4
99.3
96.6
105.5
96.7
111.4
115.7
83.2
88.9
99.1
Fluoranthene
109,9
101.4
94.3
111.6
96.6
109.6
123.2
85.4
92.7
102.7
Pyrene
106.5
105.8
107.6
116.7
90.7
127.5
103.4
95.5
93.2
105.2
3,3'-Dichlorobenzidine
100.0
*492.3
131.4
100.0
*217.6
*167.6
100,0
*748.8
100.0
116.5
Benzo(a)anthracene
98.1
107.0
98.4
119.3
98.6
104.0
105.0
93.4
89.3
101.5
Chrysene
100.0
108.5
100.2
116.8
93.0
117.0
106.7
93.6
90.2
102.9
Benzo(b)fluoranthene
106.6
109.9
75.6
121.7
100.7
93.9
106.9
81.9
93.6
99.0
Benzo(k)tluoranthene
102.4
105,2
88.4
125.5
99.4
95.1
144.7
89.2
78.1
103.1
Benzo(a)pyrene
107.9
105.5
80.8
122.3
97.7
104.6
101.7
86.2
92.0
99.9
lndeno(1,2,3-cd)pyrene
95.1
105.7
93.8
126.0
105.2
90.4
133.6
82.6
91.9
102,7
Dibenz(a,h)anthracene
85.0
102.6
82.0
118.8
100.7
91.9
142.3
71.0
93.1
98.6
Benzofg, h,i)perylene
98.0
0.0
81.2
0.0
33.6
78.6
128.7
83.0
94.2
66.4
Mean
95.1
94.3
101.0
95.5
96.5
104.1
113.0
100.9
92.5

* Values greater than 150% were not used to determine the averages, but the 0% values were used.
827QD - 52
Revision 4
January 1998
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TABLE 13
SINGLE LABORATORY ACCURACY AND PRECISION FOR THE EXTRACTION OF PAHs
FROM A CERTIFIED REFERENCE SEDIMENT EC-1, USING METHOD 3561 (SFE - SOLID TRAP)
Compound
Certified Value
(mg/kg)
SFE Value"
(mg/kg)
Percent of
Certified Value
SFE
RSD
Naphthalene
(27.9)"
41.3 ±3.6
(148)
8.7
Acenaphthyiene
(0.8)
0.9 ±0.1
(112)
11.1
Acenaphthene
(0.2)
0.2 ± 0.01
(100)
0.05
Fluorene
(15.3)
15.6 ±1.8
(102)
11.5
Phenanthrene
15.8 ±1.2
16.1 ±1.8
- 102
11.2
Anthracene
(1.3)
1.1 ±0.2
(88)
18.2
Fluoranthene
23.2 12.0
24.1 ±2.1
104
8.7
Pyrene
16,7 ± 2.0
17.2 ±1.9
103
11.0
Benz(a)anthracene
8.7 ± 0.8
8.8 ± 1.0
101
11.4
Chrysene
(9.2)
7.9 ± 0.9
(86)
11.4
Benzo(b)fluoranthene
7.9 ± 0.9
8.5 ±1.1
108
12.9
Benzo(k)fluoranthene
4.4 ± 0.5
4.1 ± 0.5
91
12.2
Benzo(a)pyrene
5.3 ± 0.7
5.1 ± 0.6
96
11.8
lndeno(1,2,3-cd)pyrene
5.7 ± 0.6
5.2 ± 0.6
91
11.5
Benzo(g,h,i)peryiene
4.9 ± 0.7
4.3 ± 0.5
88
11.6
Dibenz(a,h)anthracene
(1.3)
1.1 ±0.2
(85)
18.2
a Relative standard deviations for the SFE values are based on six replicate extractions.
b Values in parentheses were obtained from, or compared to, Soxhlet extraction results which were
not certified.
Data are taken from Reference 10.
82T0D - 53
Revision 4
January 1998
-------
TABLE 14
SINGLE LABORATORY ACCURACY AND PRECISION FOR THE EXTRACTION OF PAHs
FROM A CERTIFIED REFERENCE SEDIMENT HS-3, USING METHOD 3561 (SFE - SOLID TRAP)
Compound
Certified Value
(mg/kg)
SFE Value3
(mg/kg)
Percent of
Certified Value
SFE
RSD
Naphthalene
9.0 ± 0.7
7.4 ± 0.6
82
8.1
Acenaphthylene
0.3 ±0.1
0.4 ± 0.1
133
25.0
Acenaphthene
4.5 ±1.5
3.3 ± 0.3
73
9.0
Fluorene
13.6 ±3.1
10.4 ± 1.3
77
12.5
Phenanthrene
85.0 ±20.0
86.2 ±9.5
" 101
11.0
Anthracene
13.4 ±0.5
12.1 ±1.5
90
12.4
Fluoranthene
60.0 ± 9.0
54.0 ±6.1
90
11.3
Pyrene
39.0 ± 9.0
32.7 ±3.7
84
11.3
Benz(a)anthracene
14.6 ±2.0
12.1 ±1.3
83
10.7
Chrysene
14.1 ±2.0
12.0 ±1.3
85
10.8
Benzo(b)fluoranthene
7.7 ±1.2
8.4 ± 0.9
109
10.7
Benzo(k)fl uoranthene
2.8 ±2.0
3.2 ±0.5
114
15.6
Benzo(a)pyrene
7.4 ± 3.6
6.6 ±0.8
89
12.1
lndeno(1,2,3-cd)pyrene
5.0 ±2.0
4.5 ± 0.6
90
13.3
Benzo(g ,h ,i) perylene
5.4 ±1.3
4.4 ± 0.6
82
13.6
Dibenz(a,h)anthracene
1.3 ±0.5
1.1 ±0.3
85
27.3
a Relative standard deviations for the SFE values are based on three replicate extractions.
Data are taken from Reference 10.
8270D - 54
Revision 4
January 1998
-------
TABLE 15
SINGLE LABORATORY ACCURACY AND PRECISION FOR THE EXTRACTION OF PAHs
FROM A CERTIFIED REFERENCE SOIL SRS103-100, USING METHOD 3561
(SFE- LIQUID TRAP)
Compound
Certified Value
(mg/kg)
SFE Value3
(mg/kg)
Percent of
Certified Value
SFE
RSD
Naphthalene
32.4 ± 8.2
29.55
91
10.5
2-Methylnaphthalene
62.1 ±11.5
76.13
122
2.0
Acenaphthene
632 ±105
577.28
91
2.9
Dibenzofuran
307 ± 49
302.25
98
4.1
Fluorene
492 ± 78
427.15
87
3.0
Phenanthrene
1618 ±340
1278.03
79
3.4
Anthracene
422 ± 49
400.80
95
2.6
Fluoranthene
1280 ±220
1019.13
80
4.5
Pyrene
1033± 285
911.82
88
3.1
Benz(a)anthracene
252 ±8
225.50
89
4.8
Chrysene
297 ± 26
283.00
95
3.8
Benzo(a)pyrene
97.2 ±17.1
58.28
60
6.5
Benzo(b)fluoranthene +
Benzo(k)fluoranthene
153 ±22
130.88
86
10.7
3 Relative standard deviations for the SFE values are based on four replicate extractions.
Data are taken from Reference 11,
8270D - 55
Revision 4
January 1998
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TABLE 16
SINGLE LABORATORY RECOVERY DATA FOR SOUD-PHASE EXTRACTION OF
BASBNEUTRAL/ACiD EXTRACTABLES FROM SPIKED TCLP BUFFERS
LOW SPIKE LEVEL
Analyte

Buffer 1 (pH
= 2.886)
Buffer 2 (pH ¦
4.937)
Spike Level *~
(P9/L)
Recovery (%)
RSD
Recovery (%),
RSD
1,4-Dichlorobenzene
3,750
63
10
63
9
Hexaehloroethane
1,500
55
6
77
4
Nitrobenzene
1,000
82
10
100
5
Hexachlorobutadiene
250
65
3
56
4
2,4-Dinitrotoluene
65
89
4
101
5
Hexachlorobenzene
65
98
5
95
6
o-Cresol
100,000
83
10
85
5
m-Cresol*
100,000
86
8
85
3
p-Cresol*
100,000
*
*
*
*
2,4,6-T richlorophenol
1,000
84
12
95
12
2,4,5-Trichlorophenol
200,000
83
11
88
3
Pentachlorophenol
50,000
82
9
78
9
Results from seven replicate spiked buffer samples.
* In this study, m-cresol and p-cresol co-eluted and were quantitated as a mixture of both isomers.
Data from Reference 12.
8270D - 56 Revision 4
January 1998
-------
TABLE 17
SINGLE LABORATORY RECOVERY DATA FOR SOLID-PHASE EXTRACTION OF
BASE/NEUTRAL/ACID EXTRACTABLES FROM SPIKED TCLP BUFFERS
HIGH SPIKE LEVEL
Analyte

Buffer 1 (pH
= 2.886}
Buffer 2 (pH
= 4.937)
Spike Level
(ng/L)
Recovery (%)
RSD
Recovery (%)
RSD
1,4-Dichlorobenzene
15,000
63
10
63
9
Hexachloroethane
6,000
54
7
46
7
Nitrobenzene
4,000
81
4
81
13
Hexachlorobutadiene
1,000
81
5
- 70
11
2,4-Dinitrotoluene
260
99
8
98
3
Hexachlorobenzene
260
89
8
91
9
o-Cresol*
400,000
92
15
90
4
m-Cresor
400,000
95
8
82
6
p-Cresol*
400,000
82
14
84
7
2,4,6-T richlorophenol
4,000
93
12
104
12
2,4,5-T richlorophenol
800,000
93
14
97
23
Pentachloropheno!
200,000
84
9
73
8
Results from seven replicate spiked buffer samples.
* In this study, recoveries of these compounds were determined from triplicate spikes of the
individual compounds into separate buffer solutions.
Data from Reference 12.
8270D - 57
Revision 4
January 1998
-------
TABLE 18
RECOVERY DATA FROM THREE LABORATORIES FOR SOLID-PHASE EXTRACTION
OF BASE/NEUTRAL/ACID EXTRACTABLES FROM SPIKED TCLP LEACHATES FROM SOIL SAMPLES
Buffer 1 dH = 2.886

Lab 1

Lab 2

Lab 3

Analvte
Spike Level
(ml D*
%R
RSD
n
%R
RSD
n
%R
RSD
n
o-Cresol
200,000
86
8
7
35.3
0.7
3
7.6
6
3
m-Cresol**
-
77
8
7
—
-
-
—
-
-
p-Cresol**
-
-
-

-
-
-
7.7
11
3
2,4,6-Trichlorophenol
2,000
106
6
7
96.3
3.9
3
44.8
5
3
2,4,5-T richlorophenol
400,000
93
3
7
80.5
4.5
3
63.3
11
3
Pentachlorophenol
100,000
79
2
7
33.8
12.2
3
29.2
13
3
1,4-Dichlorobenzene
7,500
51
5
7
81.3
5.3
3
19.2
7
3
Hexachloroethane
3,000
50
5
7
66.2
2.1
3
12.6
11
3
Nitrobenzene
2,000
80
8
7
76.3
5.3
3
63.9
12
3
Hexachlorobutadiene
500
53
8
7
63.3
4.8
3
9.6
9
3
2,4-Dinitrotoluene
130
89
8
7
35.7
2.6
3
58.2
17
3
Hexachlorobenzene
1,30
84
21
7
92.3
1.6
3
71.7
9
3
(continued)
8270D - 58
Revision 4
January 1998
-------
TABLE 18
(continued)
Buffer 2 dH = 4.937

Lab 1

Lab 2

Lab 3

Analvte
Spike Level
fua ID*
%R
RSD
n
%R
RSD
n
%R
RSD
n
o-Cresol
200,00
97
13
7
37.8
4.5
3
6.1
24
3
m-Cresol**
-
83
4
7
-
—
—
6.0
25
3
p-Cresol**
-
—
--

-
-
—
-
-
-
2,4,6-T richlorophenol
2,000
104
4
7
91.7
8.0
3
37.7
25
3
2,4,5-Trichlorophenol
400,000
94
4
7
85.2
0.4
3
64.4
10
3
Pentachlorophenol
100,000
109
11
7
41.9
28.2
3
36.6
32
3
1,4-Dichlorobenzene
7,500
50
5
7
79.7
1.0
3
26.5
68
3
Hexachloroethane
3,000
51
3
7
64.9
2.0
3
20.3
90
3
Nitrobenzene
2,000
80
4
7
79.0
2.3
3
59.4
6
3
Hexachlorobutadiene
500
57
5
7
60
3.3
3
16.6
107
3
2,4-Dinitrotoluene
130
86
6
7
38.5
5.2
3
62.2
6
3
Hexachlorobenzene
130
86
7
7
91.3
0.9
3
75.5
5
3
* 250-mL aliquots of leachate were spiked. Lab 1 spiked at one-half these levels.
** m-Cresol and p-Cresol coelute. Lab 1 and Lab 3 reported o-Cresol and the sum of m- and p-Cresol. Lab 2 reported the sum of all three
isomers of Cresol.
Data from Reference 12.
8270D - 59
Revision 4
January 1998
-------
FIGURE 1
GAS CHROMATOGRAM OF BASE/NEUTRAL AND ACID CALIBRATION STANDARD
RIC
Cti,'07/oto 8;2S:68
S*«Js BASE ACID STD.2U./2M0- UL
COGS.:
RnNCEi C 1.2780 UtfCLt N 6, 4.6
CbilM: 51Bh
-------
METHOD 8270D
SEMI VOLATILE ORGANIC COMPOUNDS BY GAS CHROMATOGRAPHY/MASS
SPECTROMETRY (GC/MS)
Start
/ 7.5.4 \
/ Does any
response exceed\ Yes
initial calibration/
v curve X
n. range? /
No
7.6 Identify analyte
by comparing the
sample and standard
mass spectra.
Stop
7.5,4 Dilute
extract.
7.5.3 Analyze
extract by GC/MS.
7.1 Prepare sample
using appropriate
3500 series method.
7.2 If necessary,
cleanup extract using
appropriate 3600
series method.
7.3 Establish GC/MS
operating conditions.
Tune to DFTPP.
Perform initial
calibration.
7.4 Perform daily
calibration verification
with SPCCs and CCCs
prior to analysis of
samples.
7.7 Calculate
concentration of
each individual analyte
confirmed present.
Report results.
7.5.1 Screen extract
on GC/FID or GC/PID
to identify highly
contaminated samples.
Dilute those samples
as needed.
8270D - 61
Revision 4
January 1998
-------
METHOD 8280S
POLYCHLORINATED DIBEKIZO-p-DIOXINS AND POLYCHLORINATED DIBENZQFURANS BY
HIGH RESOLUTION GAS CHROMATOGRAPHY/LOW RESOLUTION MASS SPECTROMETRY
(HRGC/LRMS)
1.0 SCOPE AND APPLICATION
1.1 This method is appropriate for the detection and quantitative measurement of 2,3,7,8-
tetrachlorinated dibenzo-p-dioxin (2,3,7,8-TCDD), 2,3,7,8-tetrachlorinated dibenzofuran (2,3,7,8-
TCDF), and the 2,3,7,8-substituted penta-, hexa-, hepta-, and octachlorinated dibenzo-p-dioxins
(PCDDs) and dibenzofurans (PCDFs) (Figure 1) in water (at part-per-triliion concentrations), soil, fly
ash, and chemical waste samples, including stilibottoms, fuel oil, and sludge matrices (at part-per-
billion concentrations). The following compounds can be determined by this method (see Sec. 1.4
for a discussion of "total" concentrations).
Compound
CAS Registry No.
2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)
1746-01-6
1,2,3,7,8-Pentachlorodibenzo-p-dioxin (PeCDD)
40321-76-4
1,2,3,4,7,8-HexachIorodibenzo-p-dioxin (HxCDD)
39227-28-6
1,2,3,6,7,8-HexachIorodibenzo-p-dioxin (HxCDD)
57653-85-7
1,2,3,7,8,9-Hexachlorodibenzo-p-dioxin (HxCDD)
19408-74-3
1,2,3,4,6,7,8-Heptachlorodibenzo-p-dioxin (HpCDD)
35822-46-9
1,2,3,4,6,7,8,9-Octachlorodibenzo-p-dioxin (OCDD)
3268-87-9
2,3,7,8-Tetrachlorodibenzofuran (TCDF)
51207-31-9
1,2,3,7,8-PentachIorodibenzofuran (PeCDF)
57117-41-6
2,3,4,7,8-Pentachlonodibenzofuran (PeCDF)
57117-31-4
1,2,3,4,7,8-Hexachlorodibenzofuran (HxCDF)
70648-26-9
1,2,3,6,7,8-Hexachlorodibenzofuran (HxCDF)
57117-44-9
1,2,3,7,8,9-Hexachlorodibenzofuran (HxCDF)
72918-21-9
2,3,4,6,7,8-Hexachlorodibenzofuran (HxCDF)
60851-34-5
1,2,3,4,6,7,8-Heptachlorodibenzofuran (HpCDF)
67562-39-4
1,2,3,4,7,8,9-Heptachlorodibenzofuran (HpCDF)
55673-89-7
1,2,3,4,5,6,7,8-Octachlorodibenzofuran (OCDF)
39001-02-0
Total Tetrachlorodibenzo-p-dioxin (TCDD)
41903-57-5
Total Pentachlocodibenzo-p-dioxin (PeCDD)
36088-22-9
Total Hexachlorodibenzo-p-dioxin (HxCDD)
34465-46-8
Total Heptachlorodibenzo-p-dioxin (HpCDD)
37871-00-4
Total Tetrachlorodibenzofuran (TCDF)
55722-27-5
Total Pentachlorodibenzofu ran (PeCDF)
30402-15-4
Total Hexachlorodibenzofuran (HxCDF)
55684-94-1
Total Heptachlorodibenzofuran (HpCDF)
38998-75-3
8280B -1
Revision 2
January 1998
-------
1.2 The analytical method requires the use of high resolution gas chromatography and low
resolution mass spectrometry (HRGC/LRMS) on sample extracts that have been subjected to
specified cleanup procedures. The calibration range is dependent on the compound and the sample
size. The sample size varies by sample matrix. Table 2 lists the quantitation limits for the various
matrices.
1.3 This method requires the calculation of the 2,3,7,8-TCDD toxicity equivalence according
to the procedures given in the U.S. Environmental Protection Agency "Update of Toxicity Equivalency
Factors (TEFs) for Estimating Risks Associated with Exposures to Mixtures of Chlorinated Dibenzo-
p-Dioxins and Dibenzofurans (CDDs/CDFs)" February 1989 (EPA 625/3-89/016). If the toxicity
equivalence is greater than or equal to 0.7 ppb (soil or fly ash), 7 ppt (aqueous), or 7 ppb (chemical
waste), analysis on a column capable of resolving all 2,3,7,8-substituted PCDDs/PCDFs may be
necessary. If the expected concentrations of the PCDDs and PCDFs are below the quantitation
limits in Table 2, use of Method 8290 may be more appropriate.
1.4 This method contains procedures for reporting the total concentration of all
PCDDs/PCDFs in a given level of chlorination (i.e., Total TCDD, Total PeCDF, etc.), although
complete chromatographic separation of all 210 possible PCDDs/PCDFs is not possible under the
instrumental conditions described here.
1.5 This method is restricted for use only by analysts experienced with residue analysis and
skilled in HRGC/LRMS. Each analyst must demonstrate the ability to generate acceptable results
with this method.
1.6 Because of the extreme toxicity of these compounds, the analyst must take necessary
precautions to prevent the exposure of laboratory personnel or others to materials known or believed
to contain PCDDs or PCDFs. Typical infectious waste incinerators are not satisfactory devices for
disposal of materials highly contaminated with PCDDs or PCDFs. A laboratory planning to use these
compounds should prepare a disposal plan. Additional safety instructions are outlined in Sec. 11.0.
2.0 SUMMARY OF THE METHOD
2.1 This procedure uses a matrix-specific extraction, analyte-specific cleanup, and high-
resolution capillary column gas chromatography/low resolution mass spectrometry (HRGC/LRMS)
techniques.
2.2 If interferants are encountered, the method provides selected deanup procedures to
aid the analyst in their elimination. The analysis flow chart is shown at the end of this procedure.
2.3 A specified amount of water, soil, fly ash, or chemical waste samples is spiked with
internal standards and extracted according to a matrix-specific extraction procedure. Aqueous
samples are filtered, and solid samples that show an aqueous phase are centrifuged before
extraction. The extraction procedures and solvents are:
2.3.1 Soil, fly ash, or chemical waste samples are extracted with the combination
of a Dean-Stark water trap and a Soxhlet extractor using toluene. Soil, fly ash, and other solids
may also be extracted using pressurized fluid extraction (PFE) by Method 3545.
2.3.2 Water samples are extracted with a separately funnel or liquid-liquid extractor
using methylene chloride, and the particulate fraction that results from filtering the water
samples are extracted separately in a Soxhlet extractor using toluene.
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2.4 The extracts are spiked with ^CI^.SJ.S-TCDD arid submitted to an acid-base washing
treatment, dried and concentrated. The extracts are cleaned up by column chromatography on
alumina, silica gel, and activated carbon on Celite 545s and concentrated again.
2.5 An aliquot of fte concentrated extract is injected into an HRGC/LRMS system capable
of performing the selected ion monitoring.
2.6 The identification of the target compounds is based on their ordered elution and
comparison to standard solutions (Table 1) from an appropriate GC column and MS identification.
Isomer specificity for all 2,3,7,8-substituted PCDDs/PCDFs cannot be achieved on a single column.
The use of both DB-5 and SP2331 (or equivalent) columns is advised. No analyses can proceed
unless all the criteria for retention times, peak identification, signal-to-noise and ion abundance ratios
are met by the GC/MS system after the initial calibration and calibration verification.
2.7 A calculation of the toxicity equivalent concentration (TEQ) of each sample is made
using international consensus toxicity equivalence factors (TEFs), and the TEQ is used to determine
if the concentrations of target compounds in the sample are high enough to'warrant confirmation of
the results on a second GC column.
3.0 INTERFERENCES
3.1 Solvents, reagents, glassware, and other sample processing hardware may yield
discrete artifacts and/or elevated baselines which may cause misinterpretation of chromatographic
data. All of these materials must be demonstrated to be free from interferents under the conditions
of analysis by running laboratory method blanks.
3.2 The use of high purity reagents and pesticide grade solvents helps to minimize
interference problems. Purification of solvents by distillation, in all glass systems, may be required.
3.3 interferants co-extracted from the sample will vary considerably from source to source,
depending upon the industrial process being sampled. PCDDs and PCDFs are often associated with
other interfering chlorinated compounds such as PCBs and polychiorinated diphenyl ethers
(PCDPEs) which may be found at concentrations several orders of magnitude higher than that of the
analytes of interest. Retention times of target analytes must be verified using reference standards.
While certain deanup techniques are provided as part of this method, unique samples may require
additional cleanup techniques to achieve the sensitivity specified in this method.
3.4 High resolution capillary columns are used to resolve as many isomers as possible;
however, no single column is known to resolve all of the 210 isomers. The columns employed by
the laboratory in these analyses must be capable of resolving all 17 of the 2,3,7,8-substituted
PCDDs/PCDFs sufficiently to meet the method specifications.
4.0 APPARATUS AND MATERIALS
4.1 Gas chromatograph/mass spectrometer system;
4.1.1 Gas chromatograph - An analytical system with a temperature-programmable
gas chromatograph and all necessary accessories including syringes, analytical columns, and
gases. The GC injection port shall be designed for capillary columns; a splitless or an on-
column injection technique is recommended. A 2-pL injection volume is assumed throughout
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this method; however, with some GC injection ports, other volumes may be more appropriate.
A 1-pL injection volume may be used if adequate sensitivity and precision can be
demonstrated.
4.1.2 GC column - Fused silica capillary columns are needed. The columns shall
demonstrate the required separation of all 2,3,7,8-specific isomers whether a dual column or
a single column analysis is chosen. Column operating conditions shall be evaluated at the
beginning and end of each 12-hour period during which samples or concentration calibration
solutions are analyzed.
4.1.2.1 Isomer specificity for all 2,3,7,8-subsfituted PCDDs/PCDFs cannot
be achieved on the 60 m DB-5 column. Problems have been associated with the
separation of 2,3,7,8-TCDD from 1,2,3,7-TCDD and 1,2,6,8-TCDD, and separation of
2,3,7,8-TCDF from 1,2,4,9-, 1,2,7,9-, 2,3,4,6-, 2,3,4,7-, and 2,3,4,8-TCDF. Because
of the toxicologic concern associated with 2,3,7,8-TCDD and 2,3,7,8-TCDF, additional
analyses may be necessary for some samples, as described in Sec. 7.15.8. In
instances where the toxicity equivalent concentration (TEQ) is greater than 0.7 ppb
(solids), 7 ppt (aqueous), or 7 ppb (chemical waste), the reanalysis of the sample
extract on a 60 m SP-2330 or SP-2331 GC, or DB-225 column (or equivalent column)
may be required in order to determine the concentrations of the individual 2,3,7,8-
substituted isomers. For the DB-225 column, problems are associated with the
separation of 2,3,7,8-TCDF from 2,3,4,7-TCDF and a combination of 1,2,3,9- and
2,3,4,8-TCDF.
4.1.2.2 For any sample analyzed on a DB-5 or equivalent column in which
2,3,7,8-TCDF Is reported as an Estimated Maximum Possible Concentration (Sec.
7.15.7) that is above the quantitation limit for the matrix, analysis of the extract is
recommended on a second GC column which provides better specificity for 2,3,7,8-
TCDF.
4.1.2.3 Analysis on a single column is acceptable if the required
separation of all the 2,3,7,8-specific isomers is demonstrated, and the minimum
acceptance criteria outlined in Sec. 7.12 are met. See Sec. 7.14.5 for the
specifications for the analysis of the 2,3,7,8-specific isomers using both dual columns
and single columns.
4.2 Mass spectrometer - A low resolution instrument is employed, utilizing 70 volts
(nominal) electron energy in the electron impact ionization mode. The system must be capable of
selected ion monitoring (SIM). The recommended configuration is for at least 18 ions per cycle, with
a cycle time of 1 sec or less, and a minimum integration time of 25 msec per m/z. Other cycle times
and integration times may be employed, provided that the analyst can demonstrate acceptable
performance for the calibration standards and window defining mixes. The integration time used to
analyze samples shall be identical to the time used to analyze the initial and continuing calibration
solutions and quality control samples.
4.2.1 Interfaces - GC/MS interfaces constructed of an glass or glass-lined materials
are necessary. Glass can be deactivated by silanizing with dichlorodimethylsilane. Inserting
a fused silica column directly into the MS source is recommended. Care must be taken not
to expose the end of the column to the electron beam.
4.2.2 Data system - An interfaced data system is necessary to acquire, store,
reduce and output mass spectral data.
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4.3 Miscellaneous equipment
4.3.1 Nitrogen evaporation apparatus (N-Evap* Analytical Evaporator Model 111,
Organomation Association inc., Northborough, MA, or equivalent).
4.3.2 Balance capable of accurately weighing ±0.01 g.
4.3.3 Water bath - Equipped with concentric ring cover and temperature controlled
within ± 2°C.
4.3.4 Stainless steel (or glass) pan large enough to hold contents of 1 pint sample
containers.
4.3.5 Glove box - For use in preparing standards from neat materials and in
handling soil/sediment samples containing fine particulates that may pose a risk of exposure.
4.3.6 Rotary evaporator, R-110, Buchi/Brinkman - American-Scientific No. E5045-10
or equivalent.
4.3.7 Centrifuge - Capable of operating at 400 x G with a 250-300 mL capacity.
4.3.8 Drying oven.
4.3.9 Vacuum oven - Capable of drying solvent-washed solid reagents at 110°C.
4.3.10 Mechanical shaker - A magnetic stirrer, wrist-action or platform-type shaker
that produces vigorous agitation. Used for pre-treatment of fly ash samples.
4.4 Miscellaneous laboratory glassware
4.4.1 Extraction Mrs - Amber glass with pdytetrafluoroethylene (PTFE)-lined screw
cap; minimum capacity of approximately 200 iiiL.j must be coi^ ipatible ^A^ith mechanical shaker
to be used.
4.4.2 Kudema-Danish (K-D) Apparatus - 500-mL evaporating flask, 10-mL
graduated concentrator tubes with ground glass stoppers, three-ball macro-Snyder column.
NOTE: The use of a solvent vapor recovery system (Kontes K-545000-1006 or K-547300-
0000, Ace Glass 6614-30, or equivalent) is recommended for the purpose of solvent
recovery during the concentration procedures requiring the use of Kudema-Danish
evaporative concentrators. Incorporation of this apparatus may be required by State
or local municipality regulations that govern air emissions of volatile organics. EPA
recommends the incorporation of this type of reclamation system as a method to
implement an emissions reduction program. Solvent recovery is a means to conform
with waste minimization and pollution prevention initiatives.
4.4.3 Disposable Pasteur pipets, 150 mm long x 5 mm ID.
4.4.4 Disposable serological pipets, 10-mL for preparation of the carbon column
described in Sec. 7.10.
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4.4.5 Vials - 0.3-mL arid 2-mL amber borosilicate glass with conical shaped
reservoir and screw caps lined with PTFE-faced silicone disks.
4.4.6 Funnels - Glass; appropriate size to accommodate filter paper (12.5 cm).
4.4.7 Chromatography columns - 300 mm x 10.5 mm glass chromatographic
column fitted with PTFE stopcock.
4.4.8 Soxhlet apparatus, 500-mL flask, all glass - Complete with glass extractor
body, condenser, glass extraction thimbies, heating mantle, and variable transformer for heat
control.
NOTE: Extraction thimbles must be of sufficient size to hokj 100 g of sand, 5 g of silica gel,
and at least 10 g of solid sample, with room to mix the sand and sample in the
thimble.
4.4.9 Dean-Stark water separator apparatus, with a PTFE stopcock. Must fit
between Soxhlet extractor body and condenser.
4.4.10 Concentrator tubes - 15-mL conical centrifuge tubes.
4.4.11 Separatory funnels - 125-mL and 2-L separatory funnels with a PTFE
stopcock.
4.4.12 Continuous liquid-liquid extractor - 1-L sample capacity, suitable for use with
heavier than water solvents.
4.4.13 PTFE boiling chips - wash with hexane prior to use.
4.4.14 Buchner funnel -15 cm.
4.4.15 Filtration flask - For use with Buchner funnel, 1-L capacity.
4.5 Filters
4.5.1 Filter paper - Whatman No. 1 or equivalent.
4.5.2 Glass fiber filter -15-cm, for use with Buchner funnel.
4.5.3 0.7 jim, Whatman GFF, or equivalent material compatible with toluene. Rinse
with toluene.
4.6 Glass wool, silanized - Extract with methylene chloride and hexane before use.
4.7 Laboratory glassware deaning procedures - Reuse of glassware should be minimized
to avoid the risk of using contaminated glassware. All glassware that is reused shall be scrupulously
cleaned as soon as possible after use, applying the following procedure.
4.7.1 Rinse glassware with the last solvent used in it
4.7.2 Wash with hot water containing detergent.
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4.7.3 Rinse with copious amounts of tap water and several portions of organic-free
reagent water. Drain dry.
4.7.4 Rinse with pesticide grade acetone and hexane.
4.7.5 After glassware is dry, store inverted or capped with aluminum foil in a dean
environment.
4.7.6 Do not bake reusable glassware as a routine part of cleaning. Baking may
be warranted after particularly dirty samples are encountered, but should be minimized, as
repeated baking may cause active sites on the glass surface that will irreversibly adsorb
PCDDs/PCDFs.
CAUTION: The analysis for PCDDs/PCDFs in water samples is for much lower
concentrations than in soil/sediment, fly ash, or chemical waste samples.
Extreme care must be taken to prevent cross-contamination between
soil/sediment, fly ash, chemical waste and water samples. Therefore, it is
strongly recommended that separate glassware be reserved for analyzing water
samples.
4.8 Pre-extraction of glassware - All glassware should be rinsed or pre-extracted with
solvent immediately before use. Soxhlet-Dean-Stark (SDS) apparatus and continuous liquid-liquid
extractors should be pre-extracted for approximately three hours immediately prior to use, using the
same solvent and extraction conditions that will be employed for sample extractions. The pooled
waste solvent for a set of extractions may be concentrated and analyzed as a method of
demonstrating that the glassware was free of contamination.
It is recommended that each piece of reusable glassware be numbered in such a fashion that
the laboratory can associate all reusable glassware with the processing of a partiadar sample. This
will assist the laboratory in:
1) Tracking down possible sources of contamination for individual samples,
2) Identifying glassware associated with highly contaminated samples that may require extra
deaning, and
3) Determining when glassware should be discarded.
5.0 REAGENTS
5.1 Solvents - all solvents must be pesticide grade, distilled-in-glass.
5.1.1
Hexane, C6H,4
5.1.2
Methanol, CH3OH
5.1.3
Methylene chloride, CH2CI2
5.1.4
Toluene, C6H5CH3
5.1.5
Isooctane, (CH^CCHjChKCI"^
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5.1.6 Cyclohexane, C6H12
5.1.7 Acetone, CH3COCH3
5.1.8 Tridecane, CH3(CH2)11CH3
5.1.9 Nonane, CgH^
5.2 White quartz sand - 60/70 mesh, for use in the Soxhlet-Dean-Stark
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5.8.5 Silica gal impregnated with 2% (w/w) sodium hydroxide * Add one part by
weight of 1 M NaOH solution to two parts silica gel (extracted and activated) in a screw-cap
bottle and mix with a glass rod until free of lumps.
5.8.6 Silica gel impregnated with 40% (w/w) sulfuric add. Add two parts by weight
concentrated sulfuric acid to three parts silica gel (extracted and activated), mix with a glass
rod until free of lumps, and store in a screw-cap glass bottle.
5.9 Calibration solutions (Table 1) - Prepare five tridecane (or nonane) solutions (CC1-CC5)
containing 10 unlabeled and 7 carbon-labeled PCDDs/PCDFs at known concentrations for use in
instrument calibration. One of these five solutions (CC3) is used as the calibration verification
solution and contains 7 additional unlabeled 2,3,7,8-isomers. The concentration ranges are
homologue-dependent, with the lowest concentrations associated with tetra- and pentachlorinated
dioxins and furans (0.1 to 2.0 ng/pL), and the higher concentrations associated with the hexa-
through octachlorinated homologues (0.5 to 10.0 ng/(jL). Commercially-available standards
containing all 17 unlabeled analytes in each solution may also be utilized.
5.10 Internal standard solution (Table 3) - Prepare a solution containing the five internal
standards in tridecane (or nonane) at the nominal concentrations listed in Table 3. Mix 10 pL with
1.0 mL of acetone before adding to each sample and blank.
5.11 Recovery standard solution (Table 3) - Prepare a solution in hexane containing the
recovery standards, 1^C12-1,2,3,4-TCDD and 13C12-1,2,3,7,8,9-HxCDD, at concentrations of 5.0 ng/ML,
in a solvent other than tridecane or nonane.
5.12 Calibration verification solution - Prepare a solution containing standards to be used
for identification and quantitation of target analytes (Table 4).
5.13 Cleanup standard - Prepare a solution containing 37d4-2,3,718-TCDD at a concentration
of 5 ng/yL (5 MQ/niL) in tridecane (or nonane). Add this solution to all sample extracts prior to
cleanup. The solution may be added at this concentration, or diluted into a larger volume of solvent.
The recovery of this compound is used to judge the efficiency of the cleanup procedures.
5.14 Matrix spiking standard - Prepare a solution containing ten of the 2,3,7,8-substituted
isomers, at the concentrations listed in Table 5 in tridecane (or nonane). Use this solution to prepare
the spiked sample aliquot. Dilute 10 piL of this standard to 1.0 mL with acetone and add to the
aliquot chosen for spiking.
5.15 Window defining mix - Prepare a solution containing the first and last eluting isomer of
each homologue (Table 6). Use this solution to verify that the switching times between the
descriptors have been appropriately set.
5.16 Column performance sol utions
Chromatographic resolution is verified using a test mixture of PCDDs/PCDFs specific
to each column and shown below.
DB-5 test mix: 1,2,3,7-TCDm,2,3,8-TCDD
2.3.7.8-TCDD
1.2.3.9-TCDD
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DB-225 test mix: 2,3,4,7-TCDF
2.3.7.8-TCDF
1.2.3.9-TCDF
SP-2331 test mix: 2,3,7,8-TCDD
1,4,7,8-TCDD
1.2.3.7-TCDD
1.2.3.8-TCDD
The concentrations of these isomers should be approximately 0.5 ng/jjL in tridecane (or nonane).
If the laboratory employs a column that has a different elution order than those specified here,
the laboratory must ensure that the isomers eluting closest to 2,3,7,8-TCDD are represented in the
column performance solution.
6.0 SAMPLE COLLECTION, HANDLING, AND PRESERVATION
6.1 See the introductory material to this chapter, Organic Analytes,
6.2 Sample collection
6.2.1 Sample collection personnel should, to the extent possible, homogenize
samples in the field before filling the sample containers. This should minimize or eliminate the
necessity for sample homogenization in the laboratory. The analyst should make a judgment,
based on the appearance of the sample, regarding the necessity for additional mixing. If the
sample is cleariy not homogeneous, the entire contents should be transferred to a glass or
stainless steel pan for mixing with a stainless steel spoon or spatula before removal of a
sample portion for analysis.
6.2.2 Grab and composite samples must be collected in glass containers.
Conventional sampling practices must be followed. The bottle must not be prewashed with
sample before collection. Sampling equipment must be free of potential sources of
contamination.
6.2.3 If residual chlorine is present in aqueous samples, add 80 mg sodium
thiosulfate per liter of sample. If sample pH is greater than 9, adjust to pH 7-9 with sulfuric
acid.
6.3 Storage and holding times-All samples should be stored at 4°C in the dark, extracted
within 30 days and completely analyzed within 45 days of extraction. Whenever samples are
analyzed after the holding time expiration date, the results should be considered to be minimum
concentrations and should be identified as such.
NOTE: The holding times listed in Sec. 6.3 are recommendations. PCDDs and PCDFs are very
stable in a variety of matrices, and holding times under the conditions listed in Sec. 6.3
may be as high as a year for certain matrices.
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7.0 PROCEDURE
Five types of extraction procedures are employed in these analyses, depending on the sample
matrix and the available equipment.
1) Chemical waste samples are extracted by refluxing with a Dean-Stark water separator.
2) Fly ash samples and soil/sediment samples may be extracted in a combination of a Soxhlet
extractor and a Dean-Stark water separator.
3) Water samples are filtered and then the filtrate is extracted using either a separatory funnel
procedure or a continuous liquid-liquid extraction procedure.
4) The filtered particulates are extracted in a combination of a Soxhlet extractor and a Dean-
Stark water separator.
5) Fly ash, soil/sediment, and other solid samples may also be extracted using pressurized
fluid extraction (PFE), employing Method 3545. (See Method 3545 for the equipment,
supplies, reagents, and procedures associated with PFE.)
Sec. 7.1 provides general information on the use of the Soxhlet-Dean-Stark apparatus. The first four
matrix-specific extraction procedures are described in Sees. 7.2 - 7.5. Pressurized fluid extraction
is described in Method 3545.
NOTE: EPA has not performed a formal evaluation of pressurized fluid extraction (PFE) with
respect to Method 8280. However, EPA has received and evaluated data regarding this
technique in conjunction with Method 8290, the high resolution mass spectrometry method
for PCDDs/PCDFs, and has incorporated those data into that method. Given that Method
8280 addresses a higher concentration range of the target analytes than Method 8290
(rather than a lower range), EPA believes that PFE will also be applicable to analyses
employing Method 8280. Analysts wishing to employ PFE are advised to proceed with
caution. Consult Method 3545 and the manufacturer of the PFE equipment for additional
information regarding PCDD/PCDF extraction. Laboratories wishing to employ PFE in
conjunction with Method 8280 should initially demonstrate the applicability of the technique
to typical range of concentrations and matrices addressed in Method 8280, focusing on the
use of reference materials rather than spiked samples whenever possible. "Hie results of
such a demonstration should be maintained on file at the laboratory.
7.1 General considerations for use of the Soxhlet-Dean-Stark (SDS) apparatus
The following procedures apply to use of the SDS apparatus for extracting matrices covered
by this protocol.
The combination of a Soxhlet extractor and a Dean-Stark trap is used for the removal of water
and extraction of PCDDs/PCDFs from samples of fly ash, soil/sediment, and the particulate fraction
of water samples.
For soil/sediment samples, the results of these analyses are reported based on the wet weight
of the sample. However, use of the SDS allows the water content of a sample to be determined
from the same aliquot of sample that is also extracted for analysis. The amount of water evolved
from the sample during extraction is used to approximate the percent solids content of the sample.
The percent solids data may be employed by the data user to approximate the dry weight
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concentrations. The percent solids determination does not apply to the extraction of particulates
from the filtration of water samples or to the extraction of fly ash samples which are treated with an
HCI solution prior to extraction.
7.1.1 The extraction of soil/sediment, fly ash, and particulates from water samples
will require the use of a Soxhlet thimble. See Sec. 4.6 for a discussion of pre-extradion of
glassware such as the SOS. Prior to pre-extradion, prepare the thimble by adding 5 g of
70/230 mesh silica ge! to the thimble to produce a thin layer in the bottom of the thimble. This
layer will hap fine particles in the thimble. Add 80-100 g of quartz sand on top of the silica gel,
and place the thimble in the extractor.
7.1.2 Pre-extrad the SIX for three hours with toluene, then allow the apparatus to
cool and remove the thimble. Mix the appropriate weight of sample with the sand in the
thimble, being careful not to disturb the silica gel layer.
7.1.3 If the sample aliquot to be extracted contains large lumps, or is otherwise not
easily mixed in the thimble, the sand and sample may be mixed in another container. Transfer
approximately 2/3offte sand from the thimble to a dean container, being careful not to disturb
the silica gel layer when transferring the sand. Thoroughly mix the sand with the sample with
a dean spatula, and transfer the sand/sample mixture to the thimble.
7.1.4 If a sample with particularly high moisture content is to be extracted, it may
be helpful to leave a small conical depression in the material in the thimble. This will allow the
water to drain through the thimble more quickly during the early hours of the extraction. As the
moisture is removed during the first few hours of extradion, the depression mil collapse, and
the sample will be uniformly extraded.
7.2 Chemical waste extradion (induding oily sludge/wet fuel oil and stillbottom/oil).
7.2.1 Assemble a flask, a Dean-Stark trap, and a condenser, and pre-extrad with
toluene for three hours (see Sec. 4.6). After pre-extradion, allow the apparatus to cool, and
discard the used toluene, or pod it for later analysis to verify the cleanliness of the glassware.
7.2.2 Weigh about 1 g of the waste sample to two decimal places into a tared pre-
extraded 125-mL flask. Add 1 mL of the acetone-diluted internal standard solution (Sec. 5.10)
to the sample in the flask. Attach the pre-extraded Dean-Stark water separator and condenser
to the flask, and extrad the sample by reftedng it with 50 mL of toluene for at least three hours.
Continue refluxing the sample until all the water has been removed. Cool the sample,
filter the toluene extrad through a rinsed glass fiber filter into a 100-mL round-bottom flask.
Rinse the filter with 10 mL of toluene; combine the extract andlinsate. Concentrate the
combined solution to approximately 10 mL using a K-D or rotary evaporator as described in
Sees. 7.6.1 and 7.6.2. Transfer the concentrated extrad to a 125-mL separatory funnel. Rinse
the flask with toluene and add the rinse to the separatory funnel. Proceed with acid-base
washing treatment per Sec. 7.8, the micro-concentration per Sec. 7.7, the chromatographic
procedures per Sees. 7.9 and 7.10, and a final concentration per Sec. 7.11.
7.2.3 Prepare an additional two 1-g aliquots of the sample chosen for spiking. After
weighing the sample in a tared pre-extraded flask (Sec. 7.2.2), add 1.0 mL of the acetone-
diluted matrix spiking standard solution (Sec. 5.14) to each of the two aliquots. After allowing
the matrix spiking solution to equilibrate to approximately 1 hour, add the internal standard
solution and extrad the aliquots as described in Sec. 7.2.2.
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7.3 Fly ash sample extraction
7.3.1 Weigh about 10 g of the fly ash to two decimal places, and transfer to an
extraction jar. Add 1 mL of the acetone-diluted internal standard solution to the sample.
7.3.2 Add 150 mL of 1 N HCI to the fly ash sample in the jar. Seal the jar with the
PTFE-lined screw cap, place on a mechanical shaker, and shake for 3 hours at room
temperature.
7.3.3 Rinse a Whatman #1 (or equivalent) filter paper with toluene, and then filter
the sample through the filter paper in a Buchner funnel into a 1 L receiving flask. Wash the
fly ash with approximately 500 mL of organic-free reagent water.
7.3.4 Mix the fly ash with the sand in the pre-extracted thimble (Sec. 7.1.2). Place
the filter paper from Sec. 7.3.3 on top of the sand. Place the thimble in a SDS extractor, add
200 mL toluene, and extract for 16 hours. The solvent should cycle completely through the
system 5-10 times per hour. Cod and filter the toluene extract through a rinsed glass fiber
filter into a 500-mL round-bottom flask. Rinse the Her with 10 mL of toluene. Concentrate the
extract as described in Sees. 7.6.1 or 7.6.2. Transfer the concentrated extract to a 125-mL
separatory funnel. Rinse the flask with toluene and add the rinse to the separatory funnel.
Proceed with acid-base washing treatment per Sec. 7.8, the micro-concentration per Sec. 7.7,
the chromatographic procedures per Sees. 7.9 and 7.10 and a final concentration per Sec.
7.11.
NOTE: A blank should be analyzed using a piece of filter paper handled in the same manner
as the fly ash sample.
7.3.5 Prepare an additional two 10-g aliquots of the sample chosen for spiking for
use as the matrix spike and matrix spike duplicate. Transfer each aliquot to a separate
extraction jar and add 1.0 mL of the acetone-diluted matrix spiking standard solution (Sec.
5.14) to each of the two aliquots. After allowing the matrix spiking solution to equilibrate for
approximately 1 hour, add the internal standard solution and extract the aliquots as described
in Sec. 7.3.1.
7.3.6 If pressurized fluid extraction is employed, consult Method 3545.
7.4 Soil/sediment sample extraction
NOTE: Extremely wet samples may require centrifugation to remove standing water before
extraction.
7.4.1 Weigh about 10 grams of the soil to two decimal places and transfer to a pre-
extracted thimble (Sec. 7.1.2). Mix the sample with the quartz sand, and add 1 mL of the
acetone-diluted internal standard solution (Sec. 5.10) to the sample/sand mixture. Add small
portions of the solution at several sites on the surface of the sample/sand mixture.
7.4.2 Place the thimble in the SDS apparatus, add 200 to 250 mL toluene, and
reflux for 16 hours. The solvent should cycle completely through the system 5-10 times per
hour.
7.4.3 Estimate the percent solids content of the soil/sediment sample by measuring
the volume of water evolved during the SDS extraction procedure. For extremely wet samples,
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the DeaoStark trap may need to be drained one or more times during the 16-hour extraction.
Collect the water from the trap, measure its volume to the nearest 0.1 mL. Assume a density
of 1.0 g/mL, and calculate the percent solids content according to the formula below;
Percent solids - Wet weight of sample - Weight of water ^
Wet weight of sample
7.4.4 Concentrate this extract as described in Sees. 7.6.1 or 7.6.2. Transfer the
concentrated extract to a 125 mL separatory funnel. Rinse the flask with toluene and add the
rinse to the separatory funnel. Proceed with acid-base washing treatment per Sec. 7.8, the
micro concentration per Sec. 7.7, the chromatographic procedures per Sees. 7.9 and 7.10 and
a final concentration per Sec. 7.11.
7.4.5 Prepare an additional two 10-g aiiquots of the sample chosen for spiking for
use as the matrix spike and matrix spike duplicate. After transferring each aliquot to a
separate pre-extracted Soxhlet thimble, add 1.0 mL of the acetone-diluted matrix spiking
standard solution (Sec. 5.14) to each of the two aiiquots. After allowing the matrix spiking
solution to equilibrate to approximately 1 hour, add the internal standard solution (Sec. 5.10)
and extract the aiiquots as described in Sec. 7.4.1.
7.4.6 If pressurized fluid extraction is employed, consult Method 3545.
7.5 Aqueous sample extraction
7.5.1 Allow the sample to come to ambient temperature, then marie the water
meniscus on the side of the 1-L sample bottle for determination of the exact sample volume.
7.5.2 Add 1 mLofthe acetone-diluted internal standard solution (Sec. 5.10) to the
sample bottle. Cap the bottle, and mix the sample by gently shaking for 30 seconds.
7.5.3 Filter the sample through a 0.7-Mm filter that has been rinsed with toluene.
Collect the aqueous filtrate in a clean flask. If the total dissolved and suspended solids
contents are too much to filter through the 0.7-Mm filter, centrifuge the sample, decant, and
then filter the aqueous phase. Alternatively, other filter configurations, including stacked filters
of decreasing pore sizes, may be employed. Procedures for extraction of the particulate
fraction are given in Sec. 7.5.4. The aqueous portion may be extracted using either the
separatory funnel technique (Sec. 7.5.5.1) or a pre-extracted continuous liquid-liquid extractor
(Sec. 7.5.5.2).
NOTE: Ofganio-free reagent water used as a blank must also be filtered in a similar fashion,
and subjected to the same cleanup and analysis as the water samples.
7.5.4 Particulate fraction
7.5.4.1 Combine the particulate on the filter and the filter itself, and if
centrifugation was used, the solids from the centrifuge bottie(s), with the quartz sand in the
pre-extracted Soxhlet thimble. Place the filter on top of the particulate/sand mixture, and place
the thimble into a pre-extracted SDS apparatus.
7.5.4.2 Add 200 to 250 mL of toluene to the SDS apparatus and reflux for
16 hours. The solvent should cycle completely through the system 5-10 times per hour.
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7.5.4.3 Allow the Soxhlet to cool, remove the toluene and concentrate this
extract as described in Sees. 7.6.1. or 7.6.2.
7.5.4.4 Pressurized fluid extraction has not been evaluated for the
extraction of the particulate fraction.
7.5.5 Aqueous filtrate
The aqueous filtrate may be extracted by either a separatory funnel procedure (Sec.
7.5.5.1) or a continuous liquid-liquid extraction procedure (Sec. 7.5.5.2).
7.5.5.1 Separatory funnel extraction - Pour the filtered aqueous sample
into a 2-L separatory funnel. Add 60 mL methylene chloride to the sample bottle, seal, and
shake 60 seconds to rinse the inner surface. Transfer the solvent to the separatory funnel and
extract the sample by shaking the funnel for 2 minutes with periodic venting. Allow the organic
layer to separate from the water phase for a minimum of 10 minutes. Drain the methylene
chloride extract into a 500-mL K-D concentrator (mounted with a 10-mL concentrator tube) by
passing the extract through a funnel packed with a glass wool plug and half-filled with
anhydrous sodium sulfate. Extract the water sample two more times using 60 mL of fresh
methylene chloride each time. Drain each extract through the funnel No the K-D concentrator.
After the third extraction, rinse the sodium sulfate with at least 30 mL of fresh methylene
chloride. Concentrate this extract as described in Sees. 7.6.1 or 7.6.2.
7.5.5.2 Continuous liquid-liquid extraction - A continuous liquid-liquid
extractor may be used in place of a separatory funnel when experience with a sample from a
given source indicates that a serious emulsion problem will result or an emulsion is
encountered using a separatory funnel. The following procedure is used for a continuous
liquid-liquid extractor.
7.5.5.2.1 Pre-extract the continuous liquid-liquid extractor for
three hours with methylene chloride and reagent water. Allow the extractor
to cool, discard the methylene chloride and the reagent water, and add the
filtered aqueous sample to the continuous liquid-liquid extractor. Add 60 mL
of methylene chloride to the sample bottle, seal and shake for 30 seconds.
7.5.5.2.2 Transfer the solvent to the extractor. Repeat the
sample bottle rinse with an additional 50 to 100 mL portion of methylene
chloride and add the rinse to the extractor. Add 200 to 500 mL methylene
chloride to the distilling flask and sufficient reagent water to ensure proper
operation. Extract for 16 hours. Allow to cool, then detach the flask and dry
the sample by running it through a rinsed funnel packed with a glass wool
plug and 5 g of anhydrous sodium sulfate into a 500-mL K-D flask.
Concentrate the extract according to Sees. 7.6.1 or 7.6.2.
7.5.6 Combination of extracts - The extracts from both the particulate fraction (Sec.
7.5.4) and the aqueous filtrate (Sec. 7.5.5) must be concentrated using the procedures in Sec.
7.6.1 and then combined together prior to the acid-base washing treatment in Sec. 7.8.
7.5.7 Determine the original aqueous sample volume by refilling the sample bottle
to the mark and transferring the liquid to a 1-L graduated cylinder. Record the sample volume
to the nearest 5 mL.
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7.5.8 Prepare an additional two 1-L aliquots of the sample chosen for spiking for
use as the matrix spike and matrix spike duplicate. Add 1.0 mL of the acetone-diluted matrix
spiking standard solution (Sec. 5.14) to each of the two aliquots in the original sample bottles.
After allowing the matrix spiking solution to equilibrate for approximately 1 hour, add the
internal standard solution and fitter and extract the aliquots as described in Sec. 7.5.2.
7.6 Macro-concentration procedures (all matrices)
Prior to cleanup, extracts from all matrices must be concentrated to approximately 10 mL. In
addition, as noted above, the concentrated extracts from the aqueous filtrate and the filtered
particulates must be combined prior to cleanup. Two procedures may be used for macro-
concentration: rotary evaporator, or Kudema-Danish (K-D). Concentration of toluene by K-D
involves the use of a heating mantle, as toluene boils above the temperature of a water bath. The
two procedures are described below.
7.6.1 Concentration by K-D
7.6.1.1 Add one or two clean boiling chips to the flask and attach a three-
ball Snyder column. Pre-wet the column by adding approximately 1 mL of toluene
through the top.
7.6.1.2 Attach the solvent recovery system condenser, place the round-
bottom flask in a heating mantle and apply heat as required to complete the
concentration in 15-20 minutes. At the proper rate of distillation, the balls of the
column will actively chatter but the chambers will not flood.
7.6.1.3 When the apparent volume of liquid readies 10 mL, remove the
K-D apparatus from the water bath and allow it to drain and cool for at least 10
minutes.
7.6.2 Concentration by rotary evaporator
7.6.2.1 Assemble the rotary evaporator according to manufacturer's
instructions, and warm the water bath to 45°C. On a daily basis, preclean the rotary
evaporator by concentrating 100 mL of clean extraction solvent through the system.
Archive both the concentrated solvent and the solvent in the catch flask for
contamination check if necessary. Between samples, three 2-3 mL aliquots of toluene
should be rinsed down the feed tube into a waste beaker.
7.6.2.2 Attach the round-bottom flask containing the sample extract to the
rotary evaporator. Slowly apply vacuum to the system and begin rotating the sample
flask. Lower the sample flask into the water bath and adjust the speed of rotation to
complete the concentration in 15-20 minutes. At the proper rate of concentration, the
flow of condensed solvent into the receiving flask will be steady, but no bumping or
visible boiling will occur.
7.6.2.3 When the apparent volume of the liquid reaches 10 mL, shut off
the vacuum and the rotation. Slowly admit air into the system, taking care not to
splash the extract out of the sample flask.
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7.7 Micro-concentration procedures (all matrices)
When further concentration is required, either a micro-Snyder column technique or a
nitrogen evaporation technique is used to adjust the extract to the final volume required.
7.7.1 Micro-Snyder column technique
7.7.1.1 Add another one or two clean boiling chips to the concentrator
tube and attach a two-ball micro-Snyder column. Prewet the column by adding about
0.5 mL of toluene to the top of the column.
7.7.1.2 Place the round-bottom flask in a heating mantle and apply heat
as required to complete the concentration in 5-10 minutes. At the proper rate of
distillation the balls of the column will actively chatter, but the chambers will not flood.
7.7.1.3 When the apparent volume of liquid reaches 0.5 mL, remove the
K-D apparatus from the water bath and allow it to drain ahd cool for at least 10
minutes. Remove the Snyder column and rinse the flask and its lower joints with about
0.2 mL of solvent and add to the concentrator tube. Adjust the final volume to 1.0 mL
with solvent.
7.7.2 Nitrogen evaporation technique
7.7.2.1 Place the concentrator tube in a warm water bath (approximately
35°C) and evaporate the solvent volume to the required level using a gentle stream of
clean, dry nitrogen (filtered through a column of activated carbon).
CAUTION: Do not use plasticized tubing between the carbon trap and the sample.
7.7.2.2 The internal wall of the tube must be rinsed down several times
with the appropriate solvent during the operation. During evaporation, the solvent level
in the tube must be positioned to prevent water from condensing into the sample (i.e.,
the solvent level should be below the level of the water bath). Under normal operating
conditions, the extract should not be allowed to become dry.
7.7.2.3 When the apparent volume of liquid readies 0.5 mL, remove the
concentrator tube from the water bath. Adjust the final volume to 1.0 mL with solvent.
7.8 Acid-base cleanup procedure (all matrices)
7.8.1 The concentrated extracts from all matrices are subjected to a series of
cleanup procedures generally beginning with an acid-base wash, aid continuing on with silica
gel chromatography, alumina chromatography, and carbon chromatography. The acid-base
wash may not be necessary for uncolored extracts, but all the other cleanup procedures should
be employed, regardless of the color of the extract. Begin the cleanup procedures by
quantitatively transferring each concentrated extract to a separate 125-mL separatory funnel.
7.8.2 Prior to cleanup, all extracts are spiked with the 37CI4-2,3,7,6-TCDD cleanup
standard (Sec. 5.13). The recovery of this standard is used to monitor the efficiency of the
cleanup procedures. Spike 5 |jL of the cleanup standard {or a larger volume of diluted solution
containing 25 ng of 37CI4-2,3,7,8-TCDD) into each separatory funnel containing an extract,
resulting in a concentration of 0.25 ng/jjL in the final extract analyzed by GC/MS.
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CAUTION: Concentrated acid and base produce heat when mixed with aqueous solutions,
and may cause solutions to boil or splatter. Perform the following extractions
carefully, allowing the heat and pressure in the separatory funnel to dissipate
before shaking the stoppered funnel.
7.8.3 Partition the concentrated extract against 40 mL of concentrated sulfuric add.
Shake for 2 minutes. Remove and discard the acid layer (bottom). Repeat the add washing
until no color is visible in the add layer. (Perform add washing a maximum of 4 times.)
7.8.4 Partition the concentrated extract against 40 mL of 5 percent (w/v) sodium
chloride. (Caution: Add entrained in the extract may produce heat when mixed with the
sodium chloride solution). Shake for two minutes. Remove and discard the aqueous layer
(bottom).
7.8.5 Partition the concentrated extract against 40 mL of 20 percent (w/v)
potassium hydroxide (KOH). (Caution: Allow heat to dissipate before shaking). Shake for 2
minutes. Remove and discard the base layer (bottom). Repeat the base washing until color
is not visible in the bottom layer (perform base washing a maximum of four times). Strong
base (KOH) is known to degrade certain PCDDs/PCDFs; therefore, contact time should be
minimized.
7.8.6 Partition the concentrated extract against 40 mL of 5 percent (w/v) sodium
chloride. (Caution: Base entrained in the extract may produce heat when mixed with the
sodium chloride solution). Shake for 2 minutes. Remove and discard the aqueous layer
(bottom). Dry the organic layer by pouring it through a funnel containing a rinsed filter half-filled
with anhydrous sodium sulfate. Collect the extract in an appropriate size (100- to 250-mL)
round-bottom flask. Wash the separatory funnel with two 15-mL portions of hexane, pour
through the funnel and combine the extracts.
7.8.7 Concentrate the extracts of all matrices to 1.0 mL of hexane using the
procedures described in Sec. 7.7. Solvent exchange is accomplished by concentrating the
extract to approximately 100 pL, adding 2-3 mL of hexane to the concentrator tube and
continuing concentration to a final volume of 1.0 mL.
7.9 Silica gel and alumina column chromatographic procedures
7.9.1 Silica gel column - Insert a glass wool plug into the bottom of a gravity column
(1 cm x 30 cm glass column) fitted with a PTFE stopcock. Add 1 g silica gel and tap the
column gently to settle the silica gel. Add 2 g sodium hydroxide-impregnated silica gel, 1 g
silica gel, 4 g sulfuric add-impregnated silica gel, and 2 g silica gel (Sec. 5.8). Tap the column
gently after each addition. A small positive pressure (5 psi) of dean nitrogen may be used if
needed.
7.9.2 Alumina column - Insert a glass wool plug onto the bottom of a gravity column
(1 cm x 30 cm glass column) fitted with a PTFE stopcock. Add 6 g of the activated add
alumina (Sec. 5.8.1). Tap the top of the column gently.
NOTE: Check each new batch of silica gel and alumina by combining 50 pL of the continuing
calibration solution (CC3) with 950 pL of hexane. Process this solution through both
columns in the same manner as a sample extract (Sees. 7.9.5 through 7.9.9).
Concentrate the continuing calibration solution to a final volume of 50 pL. Proceed
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to Sec. 7.14. If the recovery of any of the anaiytes is less than 80%, the batch of
alumina or silica gel may not be appropriate for use.
7.9.3 Add hexane to each column until the packing is free of air bubbles. A small
positive pressure (5 psi) of clean dry nitrogen may be used if needed. Check the columns for
channeling. If channeling is present, discard the column. Do not tap a wetted column.
7.9.4 Assemble the two columns such that the eluate from the silica gel column
drains directly into the alumina column. Alternatively, the two columns may be eluted
separately.
7.9.5 Apply the concentrated extract (in hexane) from Sec. 7.8.7 to the top of the
silica gel column. Rinse the vial with enough hexane (1-2 ml.) to complete the quantitative
transfer of the sample to the surface of the silica.
7.9.6 Using 90 mL of hexane, elute the extract from Column 1 directly onto Column
2 which contains the alumina. Do not allow the alumina column to run dry.
7.9.7 Add 20 mL of hexane to Column 2, and elute until the hexane level is just
below the top of the alumina. Do not discard the eluted hexane, but collect in a separate flask
and store it for later use, as it may be useful in determining where the labeled anaiytes are
being lost if recoveries are less than 50%.
7.9.8 Add 20 mL of 20% methylene chloride/80% hexane (v/v) to Column 2 and
collect the eluate.
7.9.9 Concentrate the extract to 2 to 3 mL using the procedures in Sec. 7.7.
CAUTION: Do not concentrate the eluate to dryness. The sample is now ready to be
transferred to the carbon column.
7.10 Carbon column chromatographic procedure
7.10.1 Thoroughly mix 9,0 g activated carbon (Carbopak C, Sec. 5.8.2) and 41.0 g
Celite 545® to produce a 18% w/w mixture. Activate the mixture at 130°C for 6 hours, and
store in a desiccator.
NOTE: Check each new batch of the carbon/Celite mixture by adding 50 mL of the calibration
verification solution to 950 yL of hexane. Process the spiked solution in the same
manner as a sample extract (Sees. 7.10.3 through 7.10.5). Concentrate the
calibration verification solution to 50 pL and proceed with Sec, 7.14. If the recovery
of any of the anaiytes is less than 80%, this batch of carbon/Celite mixture may not
be used.
7.10.2 Prepare a 4-inch long glass column by cutting off each end of a 10-mL
disposable serological pipe! Fire polish both ends and flare if desired. Insert a glass wool
plug at one end of the column, and pack it with 1 g of the Carbon/Celite mixture. Insert an
additional glass wool plug in the other end.
CAUTION: It is very important that the column be packed properly to ensure that carbon
fines are not earned into the eluate. PCDDs/PCDFs will adhere to the carbon
fines and greatly reduce recovery. If carbon fines are carried into the eluate in
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Sec. 7.10.5, fitter the eluate, using a 0.7-pm filter (pre-rinsed with toluene), then
proceed to Sec. 7.11.
7.10.3 Rinse the column with:
• 4 mL toluene
• 2 mL of methylene chloride/methanol/toluene (75:20:5 v/v)
• 4 mL of cyclohexane/methylene chloride (50:50 v/v)
Discard all the column ririsates.
7.10.4 While the column is still wet, transfer the concentrated eluate from Sec.
7.9.10 to the prepared cartoon column. Rinse the eluate container with two 0.5-mL portions
of hexane and transfer the rinses to the carbon column. Elute the column with the following
sequence of solvents.
• 10 mL of cyclohexane/methylene chloride (50:50 v/v).
• 5 mL of methylene chloride/methanol/toluene (75:20:5 v/v).
NOTE: The above two eluates may be collected and combined, and used as a check on
column efficiency.
7.10.5 Once the solvents have eluted through the column, turn the column over, and
elute the PCDD/PCDF fraction with 20 mL of toluene, and collect the eluate.
7.11 Final concentration
7.11.1 Evaporate the toluene fraction from Sec. 7.10.5 to approximately 1.0 mL,
using the procedures in Sees. 7.6 and 7.7. Transfer the extract to a 2.0-mL conical vial using
a toluene rinse.
CAUTION: Do not evaporate the sample extract to dryness.
7.11.2 Add 100 |jL tridecane (or nonane) to the extract and reduce the volume to 100
pL using a gentle stream of dean dry nitrogen (Sec. 7.7). The final extract volume should be
100 |jL of tridecane (or nonane). Seal the vial and store the sample extract in the dark at
ambient temperature until just prior to GC/MS analysis.
7.12 Chromatographic conditions (recommended)
7.12.1 Establish tfie GC operating conditions necessary to achieve the resolution and
sensitivity required for the analyses, using the following conditions as guidance for the DB-5
(or equivalent) column:
Helium Linear Velocity:
Initial Temperature:
Initial Time:
Temperature Program:
Hold Time:
Total Time:
35 - 40 cm/sec at 240°C
170°C
10 minutes
increase to 320 °C at 8°C/minute
until OCDF elutes
40-45 minutes
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On the DB-5 column, the chromatographic resolution is evaluated using the CC3 calibration
standard during both the initial calibration and the calibration verification. The chromatographic
peak separation between the %r2,3,7,8-TCDD peak and the 13C12-1,2,3,4-TCDD peak must
be resolved with a valley of s 25 percent, where:
Valley = (-) * 10
y
y = the peak height of any TCDD isomer
x = measured as shown in Figure 2
The resolution criteria must be evaluated using measurements made on the selected
ion current profile (SICP) for the appropriate ions for each isomer. Measurements are not
made from total ion current profiles.
Optimize the operating conditions for sensitivity and resolution, and employ the same
conditions for both calibration and sample analyses.
7.12.2 When an SP-2331 (or equivalent) GC column is used to confirm the results
for 2,3,7,8-TCDF, the chromatographic resolution is evaluated before the analysis of any
calibration standards by the analysis of a commercially-available column performance mixture
(Sea 5.16) that contains the TCDD isomers that elute most closely with 2,3,7,8-TCDD on this
GC column (1,4,7,8-TCDD and the 1,2,3,7/1,2,3,8-TCDD pair). Analyze a 2-mL aliquot of this
solution, using the column operating conditions and descriptor switching times previously
established. The GC operating conditions for this column should be modified from those for
the DB-5 (or equivalent) column, focusing on resolution of the closely-eluting TCDD and TCDF
isomers.
NOTE: The column performance mixture may be combined with the window defining mix into
a single analysis, provided that the combined solution contains the isomers needed
to determine that criteria for both analyses can be met.
The chromatographic peak separata between unlabeled 2,3,7,8-TCDD and the peaks
representing all other unlabeled TCDD isomers should be resolved with a valley of <; 25
percent, where:
% Valley = (-) * 100
y
y = the peak height of any TCDD isomer
x = measured as shown in Figure 2
The resolution criteria must be evaluated using measurements made on the selected
ion current profile (SICP) for the appropriate ions for each isomer. Measurements are not
made from total ion current profiles.
Further analyses may not proceed until the GC resolution criteria have been met.
7.13 GC/MS Calibration
Calibration of the GC/MS system involves three separate procedures, mass calibration of the
MS, establishment of GC retention time windows, and calibration of the target analytes. These three
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procedures are described in Sees. 7.13.1 to 7.13.3. Samples should not be analyzed until
acceptable descriptor switching times, chromatographic resolution, and calibrations are achieved
and documented. The sequence of analyses is shown in Figure 3.
NOTE: The injection volume for all sample extracts, blanks, quality control samples and calibration
solutions must be the same.
7.13.1 Mass calibration - Mass calibration of the MS is recommended prior to
analyzing the calibration solutions, blanks, samples and QC samples. It is recommended that
the instalment be tuned to greater sensitivity in the high mass range in order to achieve better
response for the later eluting compounds. Optimum results using FC-43 for mass calibration
may be achieved by scanning from 222-510 amu every 1 second or less, utilizing 70 volts
(nominal) electron energy in the electron ionization mode. Under these conditions, m/z 414
and m/z 502 should be 30-50% of mlz 264 (base peak).
7.13.2 Retention time windows - Prior to the calibration of the target anatytes, it is
necessary to establish the appropriate switching times for the SIM descriptors (Table 7). The
switching times are determined by the analysis of the Window Defining Mix, containing the first
and last eluting isomers in each homologue (Table 8). Mixes are available for various
columns.
The ions in each of the four recommended descriptors are arranged so that there is
overlap between the descriptors. The ions for the TCDD, TCDF, PeCDD, and PeCDF isomers
are in the first descriptor, the ions for the PeCDD, PeCDF, HxCDD and HxCDF isomers are in
the second descriptor, the ions for the HxCDD, HxCDF, HpCDD and HpCDF isomers are in the
third, and the ions for the HpCDD, HpCDF, OCDD and OCDF isomers are in the fourth
descriptor. The descriptor switching times are set such that the isomers that elute from the
GC during a given retention time window will also be those isomers for which the ions are
monitored. For the homologues that overlap between descriptors, the laboratory may use
discretion in setting the switching times. However, do not set descriptor switching times such
that a change in descriptors occurs at or near the expected retention time of any of the 2,3,7,8-
substituted isomers.
7.13.3 Calibration of target analytes - Two types of calibration procedures, initial
calibration and calibration verification, are necessary (Sees. 7.13.3.1 and 7.13.3.2). The initial
calibration is needed before any samples are analyzed for PCDDs/PCDFs, and intermittently
throughout sample analysis, as dictated by the results of the calibration verification. The
calibration verification is necessary at the beginning of each 12-hour time period during which
sample are analyzed.
7.13.3.1 Initial Calibration - Once the Window Defining Mix has been
analyzed and the descriptor switching times have been verified (and after the analysis
of the column performance solution, if using a GC column other than DB-5), analyze
the five concentration calibration solutions (CC1-CC5), described in Table 1, prior to
any sample analysis.
7.13.3.1.1 The relative ion abundance criteria for
PCDDs/PCDFs presented in Table 9 should be met for all PCDD/PCDF
peaks, including the labeled internal and recovery standards, in all solutions.
The lower and upper limits of the ion abundance ratios represent a ±15%
window around the theoretical abundance ratio for each pair of selected ions.
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The 37CI4-2,3,7,8-TCDD cleanup standard contains no ®CI, thus the ion
abundance ratio criterion does not apply to this compound.
7.13.3.1.2 if the laboratoiy uses a GC column other than those
described here, the laboratory must ensure that the isomers eluting closest
to 2,3,7,8-TCDD on that column are used to evaluate QC column resolution
7.13.3.2 Calculate the relative response factors (RFs) for the seventeen
unlabeled target analytes relative to their appropriate Internal standards (RF„) (Table
10), according to the formulae below. For the seven unlabeled analytes and the 37CL,-
2,3,7,8-TCDD cleanup standard that are found only in the CC3 solution, only one RF
is calculated for each analyte. For the other 10 unlabeled analytes, calculate the RF
of each analyte in each calibration standard.
Calculate the RFs for the five labeled internal standards and the cleanup
standard relative to the appropriate recovery standard (RF^ (Table 10), in each
calibration standard, according to the following formulae:
RF . * A»2> *Q*
(Ajs* * Aj) * Q„
(AJ + A*) * Q„
RF
' (Ara1 + Aj x Qjs
where:
A,,1 and fi^2 - integrated areas of the two quantitation ions of the isomer of
interest (Table 8)
A*1 and A*2 = integrated areas of the two quantitation ions of the appropriate
internal standard (Table 8)
AfSt and A„2 = integrated areas of the two quantitation ions of the appropriate
recovery standard (Table 8)
Qn = nanograms of unlabeled target analyte injected
0* - nanograms of appropriate internal standard injected
Qre = nanograms of appropriate recovery standard injected.
There is only one quantitation ion for the ^Cl cleanup standard. Calculate the
relative response factor as described for RF*, using one area for the cleanup standard,
and the sum of the areas of the ions from the recovery standard.
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The RFn and xRFte are dimensioniess quantities; therefore, the units used to
express the Qn, Qa, and Qre must be the same.
7.13.3.3 Calculate the relative response factors for the unlabeled
PCDDs/PCDFs relative to the recovery standards (RF„), where:
F_ = RF_ x RF
i* n
litis relative response factor is necessary when the sample is diluted to the extent that
the S/N ratio for the internal standard is less than 10.0.
7.13.3.4 Relative Response Factor Criteria - Calculate the mean RF and
percent relative standard deviation (%RSD) of the five RFs (CC1 to CCS) for each
unlabeled PCDD/PCDF and labeled internal standards present in all five concentration
calibration solutions. No mean RF or %RSD calculations are possible for the 2,3,7,8-
substituted isomers or the cleanup standard found only in the CC3 solution.
%RSD = Standi deviation x
RF
The %RSD of the five RFs (CC1-CC5) for the unlabeled PCDDs/PCDFs and the
internal standards should not exceed 15.0%.
7.13.3.5 The response factors to be used for determining the total
homologue concentrations are described in Sec. 7.15.2.
7.13.3.6 Calibration Verification - The calibration verification consists of two
parts: evaluation of the chromatographic resolution, and verification of the RF values
to be used for quantitation. At the beginning of each 12-hour period, the
chromatographic resolution is verified in the same fashion as in the initial calibration,
through the analysis of the CC3 solution on the D&-5 (or equivalent) column, or through
the analysis of the column performance solution on the SP-2331 (or equivalent)
column.
Prepare the CC3 solution by combining the volumes of the solutions listed in
Table 4 to yield a final volume of 1.0 mL at the concentrations listed for the CC3
solution in Table 1. Alternatively, use a commercially-prepared solution that contains
the target analytes at the CC3 concentrations listed in Table 1.
For the DB-5 (or equivalent) column, begin the 12-hour period by analyzing
the CC3 solution. Inject a 2-mL aliquot of the calibration verification solution (CC3) into
the GC/MS. The identical GC/MS/DS conditions used for the analysis of the initial
calibration solutions must be used for the calibration verification solution. Evaluate the
chromatographic resolution using the QC criteria in Sec. 7.12.1.
For the SP-2331 (or equivalent) column, or other columns with different
elution orders, begin the 12-hour period with the analysis of a 2-pL aliquot of the
appropriate column performance solution. Evaluate the chromatographic resolution
using the QC criteria in Sec. 7.12.2. If this solution meets the QC criteria, proceed with
the analysis of a 2-jjL aliquot of the CC3 solution. The identical GC/MS/DS conditions
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used for the analysis of the initial calibration solutions must be used for the calibration
verification solution.
Calculate the RFs for the seventeen unlabeled target anaiytes relative to their
appropriate internal standards (RF„) and the response factors for the five labeled
internal standards and the cleanup standard relative to the appropriate recovery
standard (RFte), according to the formulae in Sec. 7.13.3.2.
Calculate the RFs for the unlabeled PCDDs/PCDFs relative to the recovery
standards (RFJ, using the formula in Sec. 7.13.3.3.
Do not proceed with sample analyses until the calibration verification criteria
have been met for
1) GC Column Resolution Criteria - The chromatographic resolution on the DB-5 (or
equivalent) and /or the SP-2331 (or equivalent) column must meet the QC criteria
in Sec. 7.12. In addition, the chromatographic peak separation between the
1,2,3,4,7,8-HxCDD and the 1,2,3,6,7,8-HxCDD in the CC3 solution shall be
resolved with a valley of s 50 percent (Figure 2).
2) Ion Abundance Criteria - The relative ion abundances listed in Table 9 must be met
for all PCDD/PCDF peaks, including the labeled internal and recovery standards.
3) Instrument Sensitivity Criteria - For the CC3 solution, the signal-to-noise (S/N) ratio
shall be greater than 2.5 for the unlabeled PCDD/PCDF ions, and greater than 10.0
for the labeled internal and recovery standards.
4) Response Factor Criteria - The measured RFs of each analyte and internal
standard in the CC3 solution must be within ±30.0% of the mean RFs established
during initial calibration for the anaiytes in all five calibration standards, and within
± 30.0% of the single-point RFs established during initial calibration for those
anaiytes present in only the CC3 standard (see Sec. 7.13.3.2).
(RF - RFJ
% Difference = -—~—± x 100
RF
where:
RF = Mean Relative response factor established during initial calibration.
RFC - Relative response factor established during calibration verification.
7.13.3.7 In order to demonstrate that the GC/MS system has retained
adequate sensitivity during the course of sample analyses, the lowest standard from
the initial calibration is analyzed at the end of each 12-hour time period during which
samples are analyzed. This analysis must utilize the same injection volume and
instrument operating conditions as were used for the preceding sample analyses.
The results of this analysis must meet the acceptance criteria for retention
times, ion abundances, and S/N ratio that are listed in Sec 7.13.3.6 for the continuing
calibration standard. Response factors do no| need to be evaluated in this end-of-shift
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standard. If this analysis fails either the ion abundance or S/N ratio criteria, then any
samples analyzed during that 12-hour period that indicated the presence of any
PCDDs/PCDFs below the method quantitation limit or where estimated maximum
possible concentrations were reported must be reanalyzed. Samples with positive
results above the method quantitation limit need not be reanalyzed.
7,14 GC/MS analysis of samples
7.14.1 Remove the extract of the sample or blank from storage. Gently swirl the
solvent on the lower portion of the vial to ensure complete dissolution of the PCDDs/PCDFs.
7.14.2 Transfer a 50-jjL aliquot of the extract to a 0.3-mL vial, and add sufficient
recovery standard solution to yield a concentration of 0.5 ng/^L. Reduce the volume of the
extract back down to 50 jjL using a gentle stream of dry nitrogen.
7.14.3 Inject a 2-pL aliquot of the extract into the GC/MS instrument. Reseal the vial
containing the original concentrated extract. Analyze the extract by GC/MS, and monitor all
of the ions listed in Table 7. The same MS parameters used to analyze the calibration
solutions must be used for the sample extracts.
7.14.4 Dilution of the sample extract is necessary if the concentration of any
PCDD/PCDF in the sample has exceeded the calibration range, or the detector has been
saturated. An appropriate dilution will result in the largest peak in the diluted sample falling
between the mid-point and high-point of the calibration range.
7.14.4.1 Dilutions are performed using an aliquot of the original extract, of
which approximately 50 pL remain from Sec. 7.14.2. Remove an appropriate size
aliquot from the vial ami add it to a sufficient volume of tridecane (or nonane) in a clean
0.3-mL conical vial. Add sufficient recovery standard solution to yield a concentration
of 0.5 ng/pL. Reduce the volume of the extract back down to 50 jjL using a gentle
stream of dry nitrogen.
7.14.4.2 The dilution factor is defined as the total volume of the sample
aliquot and dean solvent divided by the volume of the sample aliquot that was diluted.
7.14.4.3 Inject 2 jjL of the diluted sample extract into the GC/MS, and
analyze according to Sees. 7.14.1 through 7.14.3.
7.14.4.4 Diluted samples in which the MS response of any internal standard
is greater than or equal to 10% of the MS response of that internal standard in the most
recent calibration verification standard are quantitated using the internal standards.
Diluted samples in which the MS response of any internal standard
is less than 10% of the MS response of that internal standard in the most
recent calibration verification standard are quantitated using the recovery
standards (see Sec. 7.15.3).
7.14.5 Identification Criteria - For a gas chromatographic peak to be unambiguously
identified as a PCDD or PCDF, it must meet all of the following criteria.
7.14.5.1 Retention times - In order to make a positive identification of the
2,3,7,8-substituted isomers for which an isotopically labeled internal or recovery
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standard is present in the sample extract, the absolute retention tame (RT) at the
maximum peak height of the analyte must be within -1 to +3 seconds of the retention
time of the corresponding labeled standard.
in order to make a positive identification of Ihe 2,3,7,8-substituted isomers for
which a labeled standard is not available, the relative retention time (RRT) of the
analyte must be within 0.05 RRT units of the RRT established by the calibration
verification. The RRT is calculated as follows:
= retention time of the analyte
retention time of the corresponding internal standard
For non-2,3,7,8~substituted compounds (tetra through hepta), the retention
time must be within the retention time windows established by the window defining mix
for the corresponding homologue (Sec. 7.13.2).
In order to assure that retention time shifts do not adversely affect the
identification of PCDDs/PCDFs, the absolute retention times of the two recovery
standards added to every sample extract immediately prior to analysis may not shift by
more than ± 10 seconds from their retention times in the calibration verification
standard.
7.14.5.2 Peak identification - All of the ions listed in Table 8 for each
PCDD/PCDF homologue and labeled standards must be present in the SICP. The ion
current response for the two quantitation ions and the M-[C0CLJ* ions for the analytes
must maximize simultaneously (± 2 seconds). This requirement also applies to the
internal standards and recovery standards. For the cleanup standard, only one ion is
monitored.
7.14.5.3 Signal-to-noise ratio - The integrated ion current for each analyte
ion listed in Table 8 must be at least 2.5 times background noise and must not have
saturated the detector (Figure 4). The internal standard ions must be at least 10.0
times background noise and must not have saturated the detector. However, if the M-
[COCLf ion does not meet the 2.5 times S/N requirement but meets all the other
criteria listed in Sec. 7.14.5 and, in the judgement of the GC/MS Interpretation
Specialist the peak is a PCDD/PCDF, the peak may be reported as positive and the
data flagged on the report form.
7.14.5.4 Ion abundance ratios - The relative ion abundance criteria listed
in Table 9 for unlabeled analytes and internal standards must be met using peak areas
to calculate ratios.
7.14.5.4.1 If interferences are present, and ion abundance
ratios are not met using peak areas, but all other qualitative identification
criteria are met (RT, S/N, presence of all 3 ions), then use peak heights to
evaluate the ion ratio.
7.14.5.4.2 If, in the judgement of the analyst, the peak is a
PCDD/PCDF, then report the ion abundance ratios determined using peak
heights, quantitate the peaks using peak heights rather than areas for both
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the target anatyte and the internal standard, and flag the result on the report
form.
7.14.S.S Polychlorinated diphenyl ether (PCDPE) interferences.
The identification of a GO peak as a PCDF cannot be made if a signal having
S/N greater than 2.5 is detected at the same retention time (± 2 seconds) in the
corresponding PCDPE channel (Table 8). If a PCDPE is detected, an Estimated
Maximum Possible Concentration (EMPC) should be calculated for this GC peak
according to Sec. 7.15.7, regardless of the ion abundance ratio, and reported.
7.14.6 When peaks are present that do not meet all of the identification criteria in
Sec. 7.14.5 and the reporting of an estimated maximum possible concentration according to
Sec. 7.15.7 will not meet the specific project objectives, then the analyst may need to take
additional steps to resolve the potential interference problems. However, this decision
generally is project-specific and should not be applied without knowledge of the intended
application of the results. These steps may be most appropriate when historical data indicate
that 2,3,7,8-substHuted PCDDs/PCDFs have been detected in samples from the site or facility,
yet the results from a specific analysis are inconclusive. The additional steps may include the
use of additional or repeated sample cleanup procedures or the use of HRGC/MS/MS (e.g.,
tandem mass spectrometry).
7.15 Calculations
7.15.1 For GC peaks that have met all the identification criteria outlined in Sec.
7.14.5, calculate the concentration of the individual PCDD or PCDF isomers using the
formulae:
ALL MATRICES OTHER THAN WATER:
CL * (An1 + Af)
(pg/kg) =
WM^ A*) x RF„
WATER:
Q. * (A.1 + A„2)
CB (ng/L) = ^
V x {A^ + A^ * RF„ - .. .
where:
A„1 and A/ = integrated ion abundances (peak areas) of the quantitation ions of the isomer
of interest (Table 8).
A^1 and A*2 = integrated ion abundances (peak areas) of the quantitation ions of the
appropriate internal standard (Table 8).
Cn = concentration of unlabeled PCDD/PCDF found in the sample.
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W = weight of sample extracted, in grams.
V = volume of sample extracted, in liters.
Q* = nanograms of the appropriate internal standard added to the sample prior to
extraction.
RF„ = calculated relative response factor from calibration verification (see Sec.
7.13.3.6).
NOTE: In instances where peak heights are used to evaluate ion abundance ratios due to
interferences (Sec. 7.14.5.4), substitute peak heights for areas in the formulae
above.
For solid matrices, the units of ng/g that result from the formula above are equivalent
to |jg/kg. Using isotope dilution techniques for quantitation, the concentration data are
recovery corrected, and therefore, the volume of the final extract and the injection volume are
implicit in the value of Q*.
7.15.1.1 For homologues that contain only one 2,3,7,8-substituted isomer
(TCDD, PeCDD, HpCDD, and TCDF), the RF of the 2,3,7,8-substituted isomer from the
calibration verification will be used to quantitate both the 2,3,7,8-substituted isomers
and the non-2,3,7,8-isomers.
7.15.1.2 For homologues that contain more than one 2,3,7,8-substituted
isomer (HxCDD, PeCDF, HxCDF, and HpCDF), the RF used to calculate the
concentration of each 2,3,7,8-substituted isomers will be the RF determined for that
isomer during the calibration verification.
7.15.1.3 For homologues that contain one or more non-2,3,7,8-substituted
isomer, the RF used to calculate the concentration of these isomers will be the lowest
of the RFs determined during the calibration verification for the 2,3,7,8-substituted
isomers in that homologue. This RF will yield the highest possible concentration for the
non-2,3,7,8-substituted isomers.
NOTE: The relative response factors of given isomers within any homologue may be
different. However, for the purposes of these calculations, it will be assumed
that every non-2,3,7,8-substituted isomer for a given homologue has the same
relative response factor. In order to minimize the effect of this assumption on
risk assessment, the 2,3,7,8-substituted isomer with the lowest RF was
chosen as representative of each homologue. All relative response factor
calculations for the non-2,3,7,8-substituted isomers in a given homologue are
based on that isomer.
7.15.2 In addition to the concentrations of specific isomers, the total homologue
concentrations are also reported. Calculate the total concentration of each homologue of
PCDDs/PCDFs as follows:
Total concentration = sum of the concentrations of every positively identified isomer of each
PCDD/PCDF homologue.
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The total concentration must include the non-2,3,7,8-substituted isomers as well as the
2,3,7,&-substituted isomers that are also reported separately. The total number of GC peaks
included in the total homologue concentration should be reported.
7.15.3 If the area of any internal standard in a diluted sample is less than 10% of the
area of that internal standard in the calibration verification standard, then the unlabeled
PCDD/PCDF concentrations in the sample shall be estimated using the recovery standard,
using the formulae that follow. The purpose is to ensure that there is an adequate MS
response for quantitation in a diluted sample. While use of a smaller aliquot of the sample
might require smaller dilutions and therefore yield a larger area for the internal standard in the
diluted extract, this practice leads to other concerns about the homogeneity of the sample and
the representativeness of the aliquot taken for extraction.
ALL MATRICES OTHER THAN WATER:
Q_ * (A + An2) x D
C (pg/kg) - ra
w * (A^ + A^) * RFre
WATER:
Cn»,
V > (Ai ~ A»> « RF„
where:
D = the dilution factor (Sec. 7.14.4.2).
An1, A„2, K\ V. Qmr RF,,, W, and V are defined in Sees. 7.13.3.2 and 7.15.1.
7.15.4 Report results for soil/sediment, fly ash, and chemical waste samples in
micrograms per kilogram (pg/kg) and water samples in nanograms per liter (ng/L).
7.15.5 Calculate the percent recovery, R^, for each internal standard and the cleanup
standard in the sample extract, using the formula:
(A,! + A if) * Q_
R (%) = L*. Hi Z5 X 100 - ~ •
(Ara ~ AS » » Q«
where:
A*1, Ajg2, A,,1, A*2, Q*, Q„, and RF„ are defined in Sees. 7.13.3.2 and 7.15.1.
NOTE: When calculating the recovery of the 3TCI4-2,3,7,8-TCDD cleanup standard, only one
m/z is monitored for this standard; therefore, only one peak area will be used in the
numerator of this formula. Use both peak areas of the 13C,2-1,2,3,4-TCDD recovery
standard in the denominator.
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7.15.5.1 The ^12-1,2,3,4-TCDD Is used to quantitate the TCDD arid TCDF
internal standards and the cleanup standard, and the 1?C12-1,2,3,7,8,9-HxCDD is used
to quantitate the HxCDD, HpCDF and OCDO internal standards (Table 10).
7.15.5.2 If the original sample, prior to any dilutions, has any internal
standard with a percent recovery of less than 25% or greater than 150%, re-extraction
and reanalysis of that sample is necessary.
7.15.6 Sample specific estimated detection limit - The sample specific estimated
detection limit (EDL) is the estimate made by the laboratory of the concentration of a given
analyte required to produce a signal with a peak height of at least 2.5 times the background
signal level. The estimate is specific to a particular analysis of the sample, and will be affected
by sample size, dilution, etc.
7.15.6.1 An EDL is calculated for each 2,3,7,8-substituted isomer that is
not identified, regardless of whether or not non-2,3,7,8-substituted isomers in that
homologue are present. The EDL is also calculated for 2,3,7,8-substituted isomers1
giving responses for both the quantitation ions that are less than 2.5 times the
background level.
7.15.6.2 Use the formulae below to calculate an EDL for each absent
2,3,7,8-substituted PCDD/PCDF. The background level (l-y is determined by
measuring the height of the noise at the expected retention times of both the
quantitation ions of the particular 2,3,7,8-substituted isomer.
ALL MATRICES OTHER THAN WATER:
2.5 x QjX (Hn + H.2) x D
EDL (pg/kg) = B 1 " n/
W * (Hi * H|) x RF„
WATER:
2.5 x Q x + H2) x D
EDL (ng/L) -=5—^ -
V * (Hb + H^) x RFn
where:
H,,1 and H„2 = The peak heights of the noise for both of the quantitation ions of the
2,3,7,8-substituted isomer of interest
H^and Hjg2 — The peak heights of both the quantitation ions of the appropriate
internal standards
D = dilution factor (Sec. 7.14,4.2).
Qb, RFjj, W and V are defined in Sees. 7.13.3.2 and 7.15.1.
7.15.6.3 If none of the isomers within a homologue are detected, then the
EDL for the "total" homologue concentration is the lowest EDL for any of the 2,3,7,8-
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substituted isomers that were not detected. Do not add together the EDLs for the
various isomers. If a 2,3,7,8-substituted isomer is reported in the homologue, then no
EDL for the "total" is calculated.
7.15.7 Estimated maximum possible concentration - An estimated maximum possible
concentration (EMPC) Is calculated for 2,3,7,8-substituted isomers that are characterized by
a response with an S/N of at least 2.5 for both the quantitation ions, and meet all of the
identification criteria in Sec. 7.14.5 except the ion abundance ratio criteria in Sec. 7.14.5.4 or
when a peak representing a PCDPE has been detected (7.14.5.5). An EMPC is a worst-case
estimate of ttie concentration. Calculate the EMPC according to the following formulae:
ALL MATRICES OTHER THAN WATER:
Q„ * (A^ + A„2) x D
EMPCn (Mg/kg)
WATER:
EMPC„ (ng/L)
W * (Al + A*) x RF„
Qb - (A„1 ~ An2) x p
Vx(A^A^)x RF„
where:
A,1 and A^2 = Areas of both the quantitation ions.
Ajg1, A*2, Qb, RF, D, W, and V are defined in Sees. 7.13,3.2 and 7.15.1.
7.15.8 Toxic equivalent concentration (TEQ) calculation - The 2,3,7,8-TCDD toxic
equivalent concentration of PCDDs/PCDFs present in the sample is calculated according to
the method recommended by the Chlorinated Dioxins Workgroup (CDWG) of the EPA and the
Centers for Disease Control (CDC). This method assigns a 2,3,7,8-TCDD toxicity equivalency
factor (TEF) to each of the seventeen 2,3,7,8-substituted PCDDs/PCDFs shown in Table 11
("Update of Toxicity Equivalency Factors [TEFs] for Estimating Risks Associated with
Exposures to Mixtures of Chlorinated Dibenzo-p-Dioxins and -Dibenzofurans [CDDs/CDFs]"
March 1989 [EPA 625/3-89/016]).
7.15.8.1 The 2,3,7,8-TCDD TEQ Of the PCDDs/PCDFs present in the
sample is calculated by summing the product of the concentration for each of the
compounds listed in Table 11 and the TEF for each compound. The principal purpose
of making this calculation is to provide the data user with a single value, normalized to
the toxicity of 2,3,7,8-TCDD, that can more readily be used in decisions related to
mixtures of these highly toxic compounds.
7.15.8.1.1 The exclusion of homologues such as mono-, di-, tri-
and the non-2,3,7,8-substituted isomers in the higher homologues does not
mean that they are not toxic Their toxicity, as estimated at this time, is much
less than the toxicity of the compounds listed in Table 11. Hence, only the
2,3,7,8-substituted isomers are included in the TEF calculations. The
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procedure for calculating the 2,3,7,8-TCDD toxic equivalence cited above is
riot claimed by the CDWG to be based on a thoroughly established scientific
foundation. Rather, the procedure represents a "consensus recommendation
on science policy."
7.15.8.1.2 When calculating the TEQ of a sample, include only
those 2,3,7,8-substituted isomers that were detected in the sample and met
all of the qualitative identification criteria in Sec. 7.14.5. Do not include
EM PC or EDL values in the TEQ calculation.
7.15.8.2 The TEQ of a sample is also used in this analytical procedure to
determine when second column confirmation may be necessary. The need for second
column confirmation is based on the known difficulties in separating 2,3,7,8-TCDF from
other isomers. Historical problems have been associated with the separation of
2,3,7,8-TCDF from 1,2,4,9-, 1,2,7,9-, 2,3,4,6-, 2,3,4,7- and 2,3,4,8-TCDF. Because of
the toxicological concern associated with 2,3,7,8-TCDF, additional analyses may be
required for some samples as described below. If project-specific requirements do not
include second column confirmation or specify a different approach to confirmation,
then this step may be omitted and the project-specific requirements take precedence.
7.15.8.2.1 If the TEQ calculated in Sec. 7.15.8.1 is greaterthan
0.7 ppb for soil/sediment or fly ash, 7 ppb for chemical waste, or 7 ppt for an
aqueous sample, and 2,3,7,8-TCDF is either detected or reported as an
EMPC, then better isomer specificity may be required than can be achieved
on the DB-5 column. The TEQ values listed here for the various matrices are
equivalent to 70% of the historical "Action Level" set by the CDC for soil
concentrations of 2,3,7,8-TCDD at Superfund sites. As such, it provides a
conservative mechanism for determining when the additional specificity
provided by a second column confirmation may be required.
7.15.8.2.2 The sample extract may be reanalyzed on a 60 m
SP-2330 or SP-2331 GC column (or equivalent) in order to achieve better GC
resolution, and therefore, better identification and quantitation of 2,3,7,8-
TCDF. Other columns that provide better specificity for 2,3,7,8-TCDF than
the DB-5 column may also be used.
7.15.8.2.3 Regardless of the GC column used, for a gas
chromatographic peak to be identified as a 2,3,7,8-substituted PCDD/PCDF
isomer during the second column confirmation, it must meet the ion
abundance, signal-to-noise, and retention time criteria listed in Sec. 7.14.5.
7.15.8.2.4 The second column confirmation analysis may be
optimized for the analysis of 2,3,7,8-TCDF, arid need not be used to confirm
the results for any other 2,3,7,8-substituted PCDDs/PCDFs identified during
the original analysis.
8.0 QUALITY CONTROL
8.1 Refer to Chapter One and Method 8000 for specific quality control (QC) procedures.
Each laboratory should maintain a formal quality assurance program. The laboratory should also
maintain records to document the quality of the data generated.
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8.2 Quality control procedures necessary to evaluate the GC/MS system operation include
evaluation of chromatographic resolution, retention time windows, calibration verification and
chromatographic analysis of samples. Performance criteria are given in the following sections of this
method:
8.2.1 GC resolution criteria for the DB-5 or equivalent column are given in Sec.
7.12.1.
8.2.2 GC resolution criteria for SP-2331 or equivalent column are given in Sec.
7.12.2.
8.2.3 Initial calibration criteria are given in Sec. 7.13.3.1.
8.2.4 Response factor criteria for the initial calibration are given in Sec. 7.13.3.4.
8.2.5 Calibration verification criteria are given in Sec. 7.13.3.6.
8.2.6 Ion abundance criteria are given in Sees. 7.13.3.1, 7.13.3.6, and 7.14.5.4.
8.2.7 Instrument sensitivity criteria are given in Sec. 7.13.3.6.
8.2.8 Response factor criteria for the calibration verification are given in Sec.
7.13.3.6.
8.2.9 Identification criteria are given in Sec, 7.14.5.
8.2.10 Criteria for isotope ratio measurements for PCDDs/PCDFs are given in Sees.
7.13.3.1, 7.13.3.6, and Table 9.
8.3 Initial Demonstration of Proficiency - Each laboratory must demonstrate initial
proficiency with each sample preparation and determinative method combination it utilizes, by
generating data of acceptable accuracy and precision for target analytes in a dean matrix. The
laboratory must also repeat the following operations whenever new staff are trained or significant
changes in instrumentation are made. See Method 8000, Sec. 8.0 for information on how to
accomplish this demonstration.
8.4 Sample Quality Control for Preparation and Analysis - The laboratory must also have
procedures for documenting the effect of the matrix on method performance (precision, accuracy,
and detection limit). At a minimum, this includes the analysis of QC samples including a method
blank, a matrix spike, a duplicate, and a laboratory control sample (LCS) in each analytical batch.
8.4.1 Documenting the effect of the matrix should include the analysis of at least
one matrix spike and one duplicate unspiked sample or one matrix spike/matrix spike duplicate
pair. The decision on whether to prepare and analyze duplicate samples or a matrix
spike/matrix spike duplicate must be based on a knowledge of the samples in the sample
batch. If samples are expected to contain target analytes, then laboratories may use one
matrix spike and a duplicate analysis of an unspiked field sample. If samples are not expected
to contain target analytes, laboratories should use a matrix spike and matrix spike duplicate
pair. Consult Sec. 8 of Method 8000 for information on developing acceptance criteria for the
MS/MSD.
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8.4.2 A laboratory control sample (LCS) should be included with each analytical
batch. The LCS consists of an aliquot of a clean (control) matrix similar to the sample matrix
and of the same weight or volume. The LCS is spiked with the same analytes at the same
concentrations as the matrix spike. When the results of the matrix spike analysis indicate a
potential problem due to the sample matrix itself, the LCS results are used to verify that the
laboratory can perform the analysis in a clean matrix. Consult Sec. 8 of Method 8000 for
information on developing acceptance criteria for the LCS.
8.4.3 The analysis of method blanks is critical to the provision of meaningful sample
results.
8.4.3.1 Method blanks should be prepared at a frequency of at least 5%,
that is, one method blank for each group of up to 20 samples prepared at the same
time, by the same procedures.
8.4.3.2 When sample extracts are subjected to cleanup procedures, the
associated method blank must also be subjected to the same cleanup procedures.
8.4.3.4 As described in Chapter One, the results of the method blank
should be:
8.4.3.4.1 Less than the MDL for the analyte.
8.4.3.4.2 Less than 5% of the regulatory limit associated with
an analyte.
8.4.3.4.3 Or less than 5% of the sample result for the same
analyte, whichever is greater.
8.4.3.4.4 If the method blank results do not meet the
acceptance criteria above, then the laboratory should take corrective action
to locate and reduce the source of the contamination and to re-extract and
reanalyze any samples associated with the contaminated method blank.
8.4.4 The laboratory should not subtract the results of the method blank from those
of any associated samples. Such "blank subtraction" is inappropriate and often leads to
negative sample results. If the method blank results do not meet the acceptance criteria in
8.4.3 and reanalysis is not practical, then the data user should be provided with the sample
results, the method blank results, and a discussion of the corrective actions undertaken by the
laboratory.
8.5 It is recommended that the laboratory adopt additional quality assurance practices for
use with this method. The specific practices that are most productive depend upon the needs of the
laboratory and the nature of the samples. Whenever possible, the laboratory should analyze
standard reference materials and participate in relevant performance evaluation studies.
9.0 METHOD PERFORMANCE
No performance data are available at this time.
8280B - 35
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10.0 REFERENCES
1 "update of Toxicity Equivalency Factors (TEFs) for Estimating Risks Associated with
Exposures to Mixtures of Chlorinated Dibenzo-p-Dioxins and Dibenzofurans (CDDs/CDFs),"
March 1989 (EPA 6251/3-89/016).
2. "Method 8290: Polychlorinated Dibenzodioxins (PCDDs) and Polychlorinated Dibenzofurans
(PCDFs) by High Resolution Gas Chromatography/High Resolution Mass Spectrometry
(HRGC/HRMS)," Test Methods for Evaluating Solid Waste, Physical/Chemical Methods (EPA
OSW SW-846).
3. "Statement of Work for Analysis of Polychlorinated Dibenzo-prdioxins (PCDD) and
Polychlorinated Dibenzofurans, Multi-Media, Multi-Concentration, DFLM01.1," September 1991.
4. "Method 613:2,3,7,8-Tetrachlorodibenzo-p-Dioxin," 40 CFR Part 136, Guidelines Establishing
Test Procedures for the Analysis of Pollutants Under the Clean Water Act, October 26,1984.
5. "Extraction of Polychlorinated Dibenzo-p-Dioxins and Polychlorinated Dibenzofurans from
Environmental Samples Using Accelerated Solvent Extraction (ASE)," B. E. Richter, J. L.
Ezzell, D, E. Knowles, and F. Hoefler, Chemosphem, 34 (5-7), 975-987,1997,
11.0 RECOMMENDED SAFETY AND HANDLING PROCEDURES FOR PCDDs/PCDFs
11.1 The following safety practices are excerpts from EPA Method 613, Sec. 4 (July 1982
version) and amended for use in conjunction with this method. The 2,3,7,8-TCDD isomer has been
found to be acnegenic, carcinogenic, and teratogenic in laboratory animal studies. Other PCDDs
and PCDFs containing chlorine atoms in positions 2,3,7,8 are known to have toxicities comparable
to that of 2,3,7,8-TCDD. The analyst should note that finely divided dry soils contaminated with
PCDDs and PCDFs are particularly hazardous because of the potential for inhalation and ingestion.
It is recommended that such samples be processed in a confined environment, such as a hood or
a glove box Laboratory personnel handling these types of samples should wear masks fitted with
charcoal filters to prevent inhalation of dust.
11.2 The toxicity or carcinogenicity of each reagent used in this method is not precisely
defined; however, each chemical compound should be treated as a potential health hazard. From
this viewpoint, exposure to these chemicals must be kept to a minimum. The laboratory is
responsible for maintaining a current awareness file of OSHA regulations regarding the safe handling
of the chemicals listed in this method. A reference file of material safety data sheets should be
made available to all personnel involved in the chemical analysis of samples suspected to contain
PCDDs and/or PCDFs.
11.3 Each laboratory must develop a strict safety program for the handling of PCDDs and
PCDFs. The laboratory practices listed below are recommended.
11.3.1 Contamination of the laboratory will be minimized by conducting most of the
manipulations in a hood.
11.3.2 The effluents of sample splitters for the gas chromatograph and roughing
pumps on the HRGC/HRMS system should pass through either a column of activated charcoal
or be bubbled through a trap containing oil or high boiling alcohols.
8280B - 36
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11.3.3 Liquid waste should be dissolved in methanol or ethanof and irradiated with
ultraviolet light at a wavelength less than 290 nm for several days (use F 40 BL lamps, or
equivalent). Using this analytical method, analyze the irradiated liquid wastes and dispose of
the solutions when 2,3,7,8-TCDD and -TCDF congeners can no longer be detected.
11.4 The following precautions were issued by Dow Chemical U.S.A. for safe handling of
2,3,7,8-TCDD in the laboratory and amended for use in conjunction with this method. The following
statements on safe handling are as complete as possible on the basis of available toxicological
information. The precautions for safe handling and use are necessarily general in nature since
detailed, specific recommendations can be made only for the particular exposure and circumstances
of each individual use. Assistance in evaluating tie health hazards of particular plant conditions may
be obtained from certain consulting laboratories and from State Departments of Health or of Labor,
many of which have an industrial health service. The 2,3,7,8-TCDD isomer is extremely toxic to
certain kinds of laboratory animals. However, it has been handled for years without injury in
analytical and biological laboratories. Many techniques used in handling radioactive and infectious
materials are applicable to 2,3,7,8-TCDD.
11.4.1 Protective Equipment - Disposable plastic gloves, apron or lab coat, safety
glasses and laboratory hood adequate for radioactive work. However, PVC gloves should not
be used.
11.4.2 Training - Workers must be trained in the proper method of removing
contaminated gloves and clothing without contacting the exterior surfaces.
11.4.3 Personal Hygiene - Thorough washing of hands and forearms after each
manipulation and before breaks (coffee, lunch, and shift).
11.4.4 Confinement - Isolated work area, posted with signs, segregated glassware
and tools, plastic backed absorbent paper on bench tops.
11.4.5 Waste - Good technique includes minimizing contaminated waste. Plastic
bag liners should be used in waste cans.
11.4.6 Disposal of Hazardous Wastes - Refer to the November 7,1986 issue of the
Federal Register on Land Ban Rulings for details concerning the handling of dioxin containing
wastes.
11.4.7 Decontamination of Personnel - apply a mild soap with plenty of scrubbing
action. Glassware, tools and surfaces - Chlorothene NU Solvent (Trademark of the Dow
Chemical Company) is the least toxic solvent shown to be effective. Satisfactory cleaning may
be accomplished by rinsing with Chlorothene, then washing with a detergent and water. Dish
water may be disposed to the sewer after percolation through a charcoal bed filter. If is
prudent to minimize solvent wastes because they require special disposal through commercial
services that are expensive.
11.4.8 Laundry - Clothing known to be contaminated should be disposed with the
precautions described under "Disposal of Hazardous Wastes". Laboratory coats or other
clothing worn in 2,3,7,8-TCDD work area may be laundered. Clothing should be collected in
plastic bags. Persons who convey the bags and launder the clothing should be advised of the
hazard and trained in proper handling. The clothing may be put into a washer without contact
if the launderer knows the problem. The washer should be run through one full cycle before
being used again for other clothing.
8280B - 37
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11.4.9 Wipe Tests - A useful method for determining cleanliness of work surfaces
and tools is to wipe the surface with a piece of filter paper, extract the filter paper and analyze
the extract.
11.4.10 Inhalation - Any procedure that may generate airborne contamination must
be carried out with good ventilation. Gross losses to a ventilation system must not be allowed.
Handling of the dilute solutions normally used in analytical arid animal work presents no
significant inhalation hazards except in case of an accident.
11.4.11 Accidents - Remove contaminated clothing immediately, taking precautions
not to contaminate skin or other articles. Wash exposed skin vigorously and repeatedly until
medical attention is obtained.
11.5 It is recommended that personnel working in laboratories where PCDD/PCDF are
handled be given periodic physical examinations (at least annually). Such examinations should
include specialized tests, such as those for urinary porphyrins and for certain Mood parameters
which, based upon published clinical observations, are appropriate for persons who may be exposed
to PCDDs/PCDFs. Periodic facial photographs to document the onset of dermatologic problems are
also advisable.
8280B- 38
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TABLE 1
CALIBRATION SOLUTIONS
Concentration of Standard In ng/^L
Analyte
CC1
CC2
CC3
CC4
CCS
2,3,7,8-TCDD
0.1
0.25
0.5
1.0
2.0
2,3,7,8-TCDF
0.1
0.25
0.5
1.0
2.0
1,2,3,7,8-PeCDF
0.1
0.25
0.5
1.0
2.0
1,2,3,7,8-PeCDD
0.1
0.25
0.5
" 1.0
2.0
2,3,4,7,8-PeCDF

0.5

1,2,3,4,7,8-HxCDF

1.25

1,2,3,6,7,8-HxCDF
0.25
0.625
1.25
2.5
5.0
1,2,3,4,7,8-HxCDD

1.25

1,2,3,6,7,8-HxCDD
0.25
0.625
1.25
2.5
5.0
1,2,3,7,8,9-HxCDD

1.25

2,3,4,6,7,8-HxCDF

1.25

1,2,3,7,8,9-HxCDF

1.25

1,2,3,4,7,8,9-HpCDF

1.25

1,2,3,4,6,7,8-HpCDF
0.25
0.625
1.25
2.5
5.0
1,2,3,4,6,7,8-HpCDD
0.25
0.625
1.25
2.5
5.0
OCDD
0.5
1.25
2.5
5.0
10.0
OCDF
0.5
1.25
2.5
5.0
10.0
13C12-2,3,7,8-TCDD
0.5
0.5
0.5
0.5
0.5
13C12-2,3,7,8-TCDF
0.5
0.5
0.5
0.5
0.5
13C12-1,2,3,6,7,8-HxCDD
0.5
0.5
0.5
0.5
0.5
13Cir1,2,3,4,6,7,8-HpCDF
1.0
1.0
1.0
1.0
1.0
13C12-OCDD
1.0
1.0
1.0
1.0
1.0
13C12-1234-TCDD
0.5
0.5
0.5
0.5
0.5
13C12-123789-HxCDD
0.5
0.5
0.5
0.5
0.5
37CI4-2378-TCDD

0.25

* These compounds are only required in the CC3 solution. Therefore, do not perform % RSD
calculations on these analytes unless they are present in all five solutions.
8280B - 39
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TABLE 2
QUANTITATION LIMITS FOR TARGET COMPOUNDS
Anaiyte
CAS Number
Water
(ng/L)
Fly
Soil
(Mfl/kg)
Chemical
Ash
(M9/kg)
Waste*
(HQ/kg)
2,3,7,8-TCDD
1746-01-6
10
1.0
1.0
10
2,3,7,8-TCDF
51207-31-9
10
1.0
1.0
10
1,2,3,7,8-PeCDF
57117-41-6
25
2.5
2.5
25
1,2,3,7,8-PeCDD
40321-76-4
25
2.5
2.5
25
2,3,4,7,8-PeCDF
57117-31-4
25
2.5
2.5
25
1,2,3,4,7,8-HxCDF
70648-26-9
25
2.5
2.5
25
1,2,3,6,7,8-HxCDF
57117-44-9
25
2.5
2.5
25
1,2,3,4,7,8-HxCDD
39227-28-6
25
2.5
2.5
25
1,2,3,6,7,8-HxCDD
57653-85-7
25
2.5
2.5
25
1,2,3,7,8,9-HxCDD
19408-74-3
25
2.5
2.5
25
2,3,4,6,7,8-HxCDF
60851-34-5
25
2.5
2.5
25
1,2,3,7,8,9-HxCDF
72918-21-9
25
2.5
2.5
25
1,2,3,4,6,7,8-HpCDF
67562-39-4
25
2.5
2.5
25
1^,3,4,6,7,8-HpCDD
35822-46-9
25
2.5
2.5
25
1,2,3,4,7,8,9-HpCDF
55673-89-7
25
2.5
2.5
25
OCDD
3268-87-9
50
5.0
5.0
50
OCDF
39001-02-0
50
5.0
5.0
50
* "Chemical waste" includes the matrices of oils, still bottoms, oily sludge, wet fuel oil, oil-laced soil,
and surface water heavily contaminated with these matrices.
8280B - 40
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TABLE 3
INTERNAL STANDARD, RECOVERY STANDARD, AND CLEANUP STANDARD SOLUTIONS
INTERNAL STANDARD SOLUTION
Internal Standards
Concentration
"0,2-2,3,7,8-TCDD
5 ng/pL
13C|2-2,3,7,8-TCDF
5 ng/pL
13Cc-1,2,3,6,7,8-HxCDD
5 ng/pL
13C|2-1,2,3,4,6,7,8-HpCDF
10 ng/ML
13Cc-OCDD
10 ng/pL
RECOVERY STANDARD SOLUTION
Recovery Standards
Concentration
13CE-1,2,3,4-TCDD
5 ng/ML
13Cc-1i2,3,7,8,9-HxCDD
5 ng/ML
CLEANUP STANDARD SOLUTION
CleanuD Standard
Concentration
37CI4-2,3,7,8-TCDD
5 ng/pL
8280B- 41
Revision 2
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TABLE 4
CALIBRATION VERIFICATION SOLUTION
Volume Solution
500 pL CC4 (Table 1 j
125 pL Supplemental Calibration solution (below)
50 jjL Internal Standard solution (Table 3)
50 }jL Recovery Standard solution (Table 3)
50 \il Cleanup Standard solution (Table 3)
225 Ml- Tridecane (or nonane)
This solution will yield a final volume of 1.0 mL at the concentrations specified for the CC3 solution
in Table 1.
Supplemental Calibration Solution Prepared from Commercially-Available Materials
Analyte Concentration (ng/|iL)
2,3,4,7,8-PeCDF
4
1,2,3,7,8,9-HxCDD
10
1,2,3,4,7,8-HxCDD
10
1,2,3,4,7,8-HxCDF
10
1,2,3,7,8,9-HxCDF
10
2,3,4,6,7,8-HxCDF
10
1,2,3,4,7,8,9-HpCDF
10
8280B - 42
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TABLES
MATRIX SPIKING SOLUTION
Analyte Concentration (ng/|iL)
2,3,7,8-TCDD
2.5
2,3,7,8-TCDF
2.5
1,2,3,7,8-PeCDF
6.25
1,2,3,7,8-PeCDD
6.25
1,2,3,6,7,8-HxCDF
6.25
1,2,3,6,7,8-HxCDD
6.25
1,2,3,4,6,7,8-HpCDF
6.25
1,2,3,4,6,7,8-HpCDD
6.25
OCDD
12.5
OCDF
12.5
This solution is prepared in tridecane (or nonane) and diluted with acetone prior to use (see Sec.
5.16).
8280B - 43
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TABLE 6
PCDD/PCDF ISOMERS IN THE WINDOW DEFINING MIX FOR A 60 m DB-5 COLUMN

First
Last
Approximate
Homologue
Eluled
Eluted
Concentration (Mg/mL)
TCDD
1,3,6,8-
1,2,8,9-
1.0
TCDF
1,3,6,8-
1,2,8,9-
1.0
PeCDD
1,2,4,7,9-
1,2,3,8,9-
1.0
PeCDF
1,3,4,6,8-
1,2,3,8,9-
1.0
HxCDD
1,2,4,6,7,9-
1,2,3,4,6,7-
. 1.0
HxCDF
1,2,3,4,6,8-
1,2,3,4,8,9-
1.0
HpCDD
1,2,3,4,6,7,9-
1,2,3,4,6,7,8-
1.0
HpCDF
1,2,3,4,6,7,8-
1,2,3,4,7,8,9-
1.0
8280B - 44
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TABLE 7
RECOMMENDED SELECTED ION MONITORING DESCRIPTORS
Descriptor 1 Descriptor 2 Descriptors Descriptor 4
243
277
311
345
259
293
327
361
277
311
345
379
293
327
361
395
304
338
374
408
306
340
376
410
316
342
390
420
318
354
392
422
320
356
402
424
322
358
404
426
328
374
408
442
332
376
410
AAA
"¦ 1 "1
334
390
420
458
340
392
422
460
342
402
424
470
356
404
426
472
358
410
446
480
376
446
480
514
The ions at m/z 376 (HxCDPE), 410 (HpCDPE), 446 (OCDPE), 480 (NCDPE) and 514 (DCDPE)
represent the polychlorinated diphenyl ethers.
The ions in each of the four recommended descriptors are arranged so that there is overlap between
the descriptors. The ions for the TCDD, TCDF, PeCDD, and PeCDF isomers are in the first
descriptor, the tens for the PeCDD, PeCDF, HxCDD and HxCDF isomers are in the second
descriptor, the ions for the HxCDD, HxCDF, HpCDD and HpCDF isomers are in the third, and the
ions for the HpCDD, HpCDF, OCDD and OCDF isomers are in the fourth descriptor.
B280B - 45
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TABLE 8
IONS SPECIFIED FOR SELECTED ION MONITORING FOR PCDDs/PCDFs
Analyte Quantitation Ions M-[COCI]*
TCDD
320
322
259
PeCDD
356
358
293
HxCDD
396
392
327
HpCDD
424
426
361
OCDD
458
460
395
TCDF
304
306
243
PeCDF
340
342
277
HxCDF
374
376
311
HpCDF
408
410
345
OCDF
442
444
379
Internal Standards

"02-2,3,7,8-TCDD
332
334
—
"C|2-1 ,2,3,6,7,8-HxCDD
402
404
—
"Cc-OCDD
470
472
—
13Cn-2,3,7,8-TCDF
316
318
—
%r1,2,3,4,6,7,8-HPCDF
420
422
—
Recovery Standards

13Ce-1,2,3,4-TCDD
332
334
_
"Cc-I ,2,3,7,8,9-HxCDD
402
404
—
Cleanup Standard

,3,7,8-TCDD
328
(1)
265
Polychlorinated diphenyf ethers

-
HxCDPE
376
—
_
HpCDPE
410

—
OCDPE
446

—
NCDPE
480

—
DCDPE
514

(1) There is only one quantitation ion monitored for the cleanup standard.
8280B - 46
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TABLE 9
CRITERIA FOR ISOTOPIC RATIO MEASUREMENTS FOR PCDDs/PCDFs
Analyte
Selected
Ions
Theoretical
Ion
Abundance
Control
limits
TCDD
PeCDD
HxCDD
HpCDD
OCDD
320/322
356/358
390/392
424/426
458/460
0.77
1.55
1.24
1.04
0.89
0.65 - 0.89
1.32-1.78
1.05-1.43
0.88-1.20
0.76-1.02
TCDF
PeCDF
HxCDF
HpCDF
OCDF
304/306
340/342
374/376
408/410
442/444
0.77
1.55
1.24
1.04
0.89
0.65 - 0.89
1.32-1.78
1.05-1.43
0.88-1.20
0.76-1.02
Internal Standards
13Ce-1,2,3,4-TCDD
13C,2-1,2,3,6,7,8-HxCDD
13Cq-OCDD
"0,2-2,3,7,8-TCDF
"Cc-1,2,3,4,6,7,8-HPCDF
Recovery Standards
13C|2-,1,2,3,4-TCDD
13C|2-1,2,3,7,8,9-HxCDD
332/334
402/404
470/472
316/318
420/422
332/334
402/404
0.77
1.24
0.89
0.77
1.04
0.77
1.24
0.65 - 0.89
1.05-1.43
0.76-1.01
0.65 - 0.89
0.88-1.20
0.65 - 0.89
1.05-1.43
8280B - 47
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TABLE 10
RELATIONSHIP OF INTERNAL STANDARDS TO ANALYTES, AND RECOVERY
STANDARDS TO INTERNAL STANDARDS, CLEANUP STANDARD, AND ANALYTES
INTERNAL STANDARDS VS. ANALYTES
Internal Standard
Analyte
13C12-TCDD
13C12-HxCDD
13c12-ocdd
13C12-TCDF
13C12-HpCDF
2,3,7,8-TCDD
1,2,3,7,8-PeCDD
1.2.3.6.7.8-HxCDD
1.2.3.7.8.9-HxCDD
1,2,3,4,7,8-HxCDD
1.2.3.4.6.7.8-HpCDD
1,2,3,4,6,7,8,9-OCDD
1,2,3,4,6,7,8,9-OCDF
2,3,7,8-TCDF
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCDF
1.2.3.6.7.8-HxCDF
1.2.3.7.8.9-HxCDF
1,2,3,4,7,8-HxCDF
2,3,4,6,7,8-HxCDF
1.2.3.4.5.8.9-HpCDF
1,2,3,4,7,8,9-HpCDF
8280B - 48
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TABLE 10 (cont.)
RECOVERY STANDARDS VS. ANALYTES, INTERNAL STANDARDS, AND CLEANUP STANDARD
Recovery Standard
Analyte, Internal Standard
13Ci2"1,2,3,4-TCDD
2,3,7,8-TCDD

1,2,3,7,8-PeCDD

2,3,7,8-TCDF

1,2,3,7,8-PeCDF

2,3,4,7,8-PeCDF

13C12-2,3,7,8-TCDD

,3C12-2378-TCDF

37CI4-2378-TCDD
13C12-1,2,3,7,8,9-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,8,9-HxCDD
1,2,3,4,7,8-HxCDD
1.2.3.6.7.8-HxCDF
1.2.3.7.8.9-HxCDF
"1,2,3,4,7,8-HxCDF
2,3,4,6,7,8-HxCDF
1,2,3,4,5,8,9-HpCDF
1,2,3,4,7,8,9-HpCDF
1,2,3,4,6,7,8,9-OCDD
1,2,3,4,6,7,8,9-OCDF
13C12-1,2,3,6,7,8-HxCDD
13C12-1,2,3,4,6,7,8-HpCDF
13C12-OCDD
82808 - 49
Revision 2
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TABLE 11
2,3,7,8-TCDD TOXICITY EQUIVALENCY FACTORS (TEFs) FOR THE PCDDs/PCDFs
Compound Toxicity Equivalency Factor (TEF)
Mono-, di-, and trichloro dibenzo-p-dioxins 0.0
2,3,7,8-tetrachloro-dibenzo-p-dioxin 1.0
All other tetrachloro-cfibenzo-/>.dioxins 0.0
1,2,3,7,8-pentachloro-dibenzo-/>dioxin 0.5
All other pentachloro-dibenzo-p-dioxins 0.0
1,2,3,4,7,8-hexachloro-dibenzo-p-dioxin 0.1
1.2.3.6.7.8-hexachloro-dibenzo-p-dioxin 0.1
1.2.3.7.8.9-hexachIoro-dibenzo-p-dioxin 0.1
All other hexachloro-dibenzo-p-dioxins 0.0
1,2,3,4,6,7,8-heptachloro-diberizo-p-dioxin 0.01
All other heptachloro-dibenzo-p-dioxins 0.0
Octachloro-dibenzo-p-dioxin 0.001
All mono-, di-, and trichloro dibenzofurans 0.0
2,3,7,8-tetrachlorodibenzofuran 0.1
All other tetrachforodibenzofurans 0.0
1,2,3,7,8-pentachlorodibenzofu ran 0.0S
2,3,4,7,8-pentachlorodibenzofuran 0.5
All other pentachlorodibenzofurans 0.0
1,2,3,4,7,8-hexachlorodibenzofviran 0.1
1.2.3.6.7.8-hexachlorodibenzofuran 0.1
1.2.3.7.8.9-hexachlorodibenzofuran 0.1
2,3,4,6,7,8-hexachlorodibenzofuran 0.1
All other hexachlorodibenzofurans 0.0
1.2.3.4.6.7.8-heptachlorodibenzofuran 0.01
1.2.3.4.7.8.9-heptachlorodibenzofuran - 0.01
All other heptachlorodibenzofurans 0.0
Octachlorodibenzofuran 0.001
8280B - 50
Revision 2
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FIGURE 1
GENERAL STRUCTURES OF PCDDs (top) AND PCDFs (bottom)
8280B - 51
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FIGURE 2
VALLEY BETWEEN 2,3,7,8-TCDD AND OTHER CLOSELY ELUTING
ISOMERS ON A DB-5 GC COLUMN
<•> pj
100
CM
CD
22:30
24:00
2S:30
27:00
Time
Selected "m current profile for m/z 322 (TCDD) produced by MS analysis of GC performance check
solution on a 60 m x 0.32 mm DB-5 fused silica capillary column with 0.25 ym film thickness.
Injector temp: 270°C
Starting temp: 200°C for 2 min
200 to 220 °C @ 57min and held for 16 min
220 to 235°C @ 5°/min and held for 7 min
235 to 330°C @ 5°/min and held for 5 min
Splitless valve time: 45 sec
Total time: 60 min
8280B - 52
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FIGURE 3
EXAMPLE OF THE ANALYTICAL SEQUENCE FOR CALIBRATING AN SP-2331 COLUMN
Time Analysis
Window Defining Mix
Column Performance Solution (SP-2331)
Hour 0 CC3
CC1 (initial Calibration)
CC2
CC4
CCS
Blanks and Samples
CC1 (must be injected within the 12-hour period.)
Hour 12 Column Performance Solution (SP-2331)
CC3
Blanks and Samples
CC1 (must be injected within the 12-hour period.)
Hour 24 Column Performance Solution (SP-2331)
CCS
Blanks and Samples
CC1 (must be injected within the 12-hour period.)
NOTE: When a column other than SP-2331 is employed, the column performance solution
need not be analyzed.
8280B - S3
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FIGURE 4
MANUAL DETERMINATION OF S/N
E,
100
90
80
70-
80-
50
117
= 19.5
40
30
20
10
20:00
22:00
24:00
26:00
30:00
28:00
nniiiwrwwi»r"»n"

Mi'vrmnei i fir rnenTi:
1 ^
28:00
The peak height (S) is measured between the mean noise (lines C and D). These mean
signal values are obtained by tracing the One between the baseline average noise extremes,
E1 and E2, and between the apex average noise extremes, E3 and E4, at the apex of the
signal.
NOTE: It is imperative that the instrument interface amplifier electronic zero offset be set
high enough so that negative going baseline noise is recorded.
8280B- 54
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METHOD 82806
POLYCHLORINATED DIBENZO-p-DIOXINS AND POLYCHLORINATED DIBENZOFURANS BY
HIGH RESOLUTION GAS CHROMATOGRAPHY/LOW RESOLUTION MASS SPECTROMETRY
(HRGC/LRMS)
Start
Stop
7.7 When necessary, continue
concentration to final volume of
1.0 mL.
7.14 Analyze samples.
7.6 Concentrate sample to
approximately 10 mL.
7.15 Calculate Results
7.8 Acid - base cleanup
and sample concentration.
7.13 Calibrate GC/MS system.
7.12 Set GC conditions.
7.10 Carbon Column
Chromatography Cleanup
and sample concentration.
7.11 Perform final
concentration down to
100 uL.
7.9 Silica gel and Alumina
Column chromatography
cleanup and sample concentration.
7.0 - 7.5 Extract sample
using technique appropriate
for matrix. Prepare additional
samples for spiking.
8280B - 55
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METHOD 8290A
POLYCHLORINATED DIBENZODIOXINS fPCDDsl AND POLYCHLORINATED
DIBENZOFURANS fPCDFs) BY HIGH-RESOLUTION GAS CHROMATOGRAPHY/HIGH-
RESOLUTION MASS SPECTROMETRY fHRGC/HRMS)
1.0 SCOPE AND APPLICATION
1.1 This method provides procedures for the detection and quantitative measurement of
poiychlorinated dibenzo-p-dioxins (tetra- through octachlorinated homologues; PCDDs), and
polychlorinated dibenzofurans (tetra- through octachlorinated homologues; PCDFs) in a variety of
environmental matrices and at part-per-trillion (ppt) to part-per-quadrillion (ppq) concentrations. The
following compounds can be determined by this method:
Analyte
CAS Registry No.
2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)
1746-01-6
1,2,3,7,8-Pentachlorodibenzo-p-dioxin (PeCDD)
40321-76-4
1,2,3,4,7,8-Hexachlorodibenzo-p-dioxin (HxCDD)
39227-28-6
1,2,3,6,7,8-Hexachiorodibenzo-p-dioxin (HxCDD)
57653-85-7
1,2,3,7,8,9-Hexachlorodibenzo-p-dioxin (HxCDD)
19408-74-3
1,2,3,4,6,7,8-Heptachlorodibenzo-p-dioxin (HpCDD)
35822-46-9
1,2,3,4,5,6,7,8-Octachlorodibenzo-p-dioxin (OCDD)
3268-87-9
2,3,7,8-Tetrachlorodibenzofuran (TCDF)
51207-31-9
1,2,3,7,8-Pentachlorodibenzofuran (PeCDF)
57117-41-6
2,3,4,7,8-Pentachlorodibenzofuran (PeCDF)
57117-31-4
1,2,3,4,7,8-Hexachlorodibenzofuran (HxCDF)
70648-26-9
1,2,3,6,7,8-Hexachlorodibenzofuran (HxCDF)
57117-44-9
1,2,3,7,8,9-Hexachlorodibenzofuran (HxCDF)
72918-21-9
2,3,4,6,7,8-Hexachlorodibenzofuran (HxCDF)
60851-34-5
1,2,3,4,6,7,8-Heptachlorodibenzofuran (HpCDF)
67562-39-4
1,2,3,4,7,8,9-Heptachlorodibenzofuran (HpCDF)
55673-89-7
1,2,3,4,5,6,7,8-Octachlorodibenzofuran (OCDF)
39001-02-0
41903-57-5
36088-22-9
34465-46-8
37871-00-4
55722-27-5
30402-15-4
55684-94-1
38998-75-3
Total T etrachlorodibenzo-p-dioxin (TCDD)
Total Pentachlorodibenzo-p-dioxin (PeCDD)
Total Hexachlorodibenzo-p-dioxin (HxCDD)
Total Heptachlorodibenzo-p-dioxin (HpCDD)
Total Tetrachiorodibenzofuran (TCDF)
Total Perrtachlorodibenzofuran (PeCDF)
Total Hexachlorodibenzofuran (HxCDF)
Total Heptachlorodibenzofuran (HpCDF)
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1.2 The analytical method calls for the use of high-resolution gas chromatography and high-
resolution mass spectrometry (HRGC/HRMS) on purified sample extracts. Table 1 lists the various
sample types covered by this analytical protocol, the 2,3,7,8-TCDD-based method calibration Rmits
(MCLs), and other pertinent information. Samples containing concentrations of specific congeneric
analytes (PCDDs and PCDFs) considered within the scope of this method that are greater than
ten times the upper MCLs must be analyzed by a protocol designed for such concentration levels,
e.g., Method 8280. An optional method for reporting the analytical results using a 2,3,7,8-TCDD
toxicity equivalency factor (TEF) is described.
1.3 The sensitivity of this method is dependent upon the level of interferences within a
given matrix. The calibration range of the method for a 1-L water sample is 10 to 2000 ppq for
TCDD/TCDF and PeCDD/PeCDF, and 1.0 to 200 ppt for a 10-g soil, sediment, fly ash, or tissue
sample for the same analytes (Table 1). Analysis of a one-tenth aliquot of the sample permits
measurement of concentrations up to 10 times the upper MCL. The actual limits of detection and
quantitation will differ from the lower MCL, depending on the complexity of the matrix.
) 1.4 This method is designed for use by analysts who are experienced with residue analysis
and ikiDed in HRGC/HRMS.
1.5 Because of the extreme toxicity of many of these compounds, the analyst must take
the necessary precautions to prevent exposure to materials known or believed to contain PCDDs
or PCDFs. It is the responsibility of the laboratory personnel to ensure that safe handling procedures
are employed. Sec. 11 of this method discusses safety procedures.
2.0 SUMMARY OF METHOD
2.1 This procedure uses matrix-specific extraction, arialyte-spedfic cleanup, and
HRGC/HRMS analysis techniques.
2.2 If interferences are encountered, the method provides selected cleanup procedures to
aid the analyst in their elimination. A simplified analysis flow chart is presented at the end of this
method.
2.3 A specified amount (see Table 1) of soil, sediment, fly ash, water, sludge (including
paper pulp}, still bottom, fuel oil, chemical reactor residue, fish tissue, or human adipose tissue is
spiked with a solution containing specified amounts of each of the nine isotopically f3C12) labeled
FCDDs/PCDFs listed in Column 1 of Table 2. The sample is then extracted according to a matrix*
specific extraction procedure. Aqueous samples that are judged to contain 1 percent or more solids,
and solid samples that show an aqueous phase, are fitters*!, the solid phase (including the filter) and
the aqueous phase extracted separately, and the extracts combined before, extract cleanup. The
extraction procedures are:
a) Toluene: Soxhlet extraction for soil, sediment, fly ash, and paper pulp samples;
b) Methylene chloride: liquid-liquid extraction for water samples;
c) Toluene: Dean-Stark extraction for fuel oil, and aqueous sludge samples;
d) Toluene extraction for still bottom samples;
e) Hexane/methytene chloride: Soxhlet extraction or methylene chloride: Soxhlet extraction
for fish tissue samples; and
f) Methylene chloride extraction for human adipose tissue samples.
8290A - 2
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9) As an option, all solid samples (wet or dry) may be extracted with toluene using a
Soxhlet/Dean Stark extraction system or using pressurized fluid extraction (PFE) (Method
3545).
The decision for the selection of an extraction procedure for chemical reactor residue samples
is based on the appearance (consistency, viscosity) of the samples. Generally, they can be handled
according to the procedure used for still bottom (or chemical sludge) samples.
2.4 The extracts are submitted to an acid-base washing treatment and dried. Following
a solvent exchange step, the extracts are cleaned up by column chromatography on alumina, silica
gel, and activated carbon.
2.4.1 The extracts from adipose tissue samples are treated with silica gel
impregnated with sulfuric acid before chromatography on acidic silica gel, neutral alumina, and
activated carbon.
2.4.2 Fish tissue and paper pulp extracts are subjected to an acid wash treatment
only, prior to chromatography on alumina and activated carbon.
2.5 The preparation of the final extract for HRGC/HRMS analysis is accomplished by adding
10 to 50 mL (depending on the matrix) of a nonane solution containing 50 pg/pL of the recovery
standards 13C,2-1,2,3,4-TCDD and t3C ^,2,3,7,8,9-HxCDD (Table 2). The former is used to
determine the percent recoveries of tetra- and pentachlorinated PCDD/PCDF congeners, white the
latter is used to determine the percent recoveries of the hexa-, hepta- and octachlorinated
PCDD/PCDF congeners.
2.6 A 2-\iL aliquot of the concentrated extract are injected into an HRGC/HRMS system
capable of performing selected ion monitoring at resolving power of at least 10,000 (10 percent
valley definition).
2.7 The identification of OCDD and nine of the fifteen 2,3,7,8-substituted congeners (Table
3), for which 13C-labeled standards are available in the sample fortification and recovery standard
solutions (Table 2), is based on their elution at their exact retention time (within 0.005 retention time
units measured in the routine calibration) and the simultaneous detection of the two most abundant
ions in the molecular ion region. The remaining six 2,3,7,8-substituted congeners (i.e., 2,3,4,7,8-
PeCDF; 1,2,3,4,7,8-HxCDD; 1,2,3,6,7,8-HxCDF; 1,2,3,7,8,9-HxCDF; 2,3,4,6,7,8-HxCDF, and
1,2,3,4,7,8,9-HpCDF), for which no carbon-labeled internal standards are available in the sample
fortification solution, and all other PCDD/PCDF congeners are identified when their relative retention
times fell within tfieir respective PCDD/PCDF retention time windows, as established from the routine
calforation data, and the simultaneous detection of the two most abundant ions in the molecular ion
region. The identification of OCDF is based on its retention time relative to 13C12-OCDD and the
simultaneous detection of the two most abundant ions in the molecular ion region. Identification also
is based on a comparison of the ratios of the integrated ion abundance of the molecular ion species
to their theoretical abundance ratios.
2.8 Quantitation of the individual congeners, total PCDDs, and total PCDFs is achieved in
conjunction with the establishment of a multipoint (five points) calibration curve for each homologue,
during which each calibration solution is analyzed once.
8290A - 3
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3.0 INTERFERENCES
3.1 Solvents, reagents, glassware and other sample processing hardware may yield
discrete artifacts or elevated baselines that may cause misinterpretation of the chromatographic data
(see references 1 and 2.) All of these materials must be demonstrated to be free from interferants
under the conditions of analysis by performing laboratory method blanks. Analysts should avoid
using PVC gloves.
3.2 The use of high purity reagents and solvents helps minimize interference problems.
Purification of solvents by distillation in all-glass systems may be necessary.
3.3 Interferants coextracted from the sample will vary considerably from matrix to matrix.
PCDDs and PCDFs are often associated with other interfering chlorinated substances such as
polychlorinated biphenyfs (PCBs), polychlorinated diphenyi ethers (PCDPEs), polychlorinated
naphthalenes, and polychlorinated alkyldibenzofurans, that may be found at concentrations several
orders of magnitude higher than the analytes of interest. Retention times of target anatytes must be
verified using reference standards. These values must correspond to the retention time windows
established in Sec. 8.1.1.3. While cleanup techniques are provided as part of this method, unique
samples may require additional cleanup steps to achieve lower detection limits.
3.4 A high-resolution capillary column (60-m DB-5, J&W Scientific, or equivalent) is used
in this method. However, no single column is known to resolve all isomers. The 60-m DB-5 GC
column is capable of 2,3,7,8-TCDD isomer specificity (Sec. 8.1.1). In order to determine the
concentration of the 2,3,7,8-TCDF (if detected on the DB-5 column), the sample extract must be
reanalyzed on a column capable of 2,3,7,8-TCDF isomer specificity (e.g., DB-225, SP-2330, SP-
2331, or equivalent).
4.0 APPARATUS AND MATERIALS
4.1 High-resolution gas chromatograph/high-resolution mass spectrometer/data system
(HRGC/HRMS/DS) - The GC must be equipped for temperature programming, and all required
accessories must be available, such as syringes, gases, and capillary columns.
4.1.1 GC injection port - The GC injection port must be designed for capillary
columns. The use of splitless injection techniques is recommended. On-column 1-|iL
injections can be used on the 60-m DB-5 column. The use of a moving needle injection port
is also acceptable. When using the method described in this protocol, a 2-pL injection volume
is used consistently (i.e., the injection volumes for all extracts, blanks, calibration solutions and
the performance check samples are 2 |jL). The use of 1-|jL injections is allowed; however,
laboratories must remain consistent throughout the analyses by .using the same injection
volume at all times.
4.1.2 GC/MS interface - The GC/MS interface components should withstand 350°C.
The interface must be designed so that the separation of 2,3,7,8-TCDD from the other TCDD
isomers achieved in the gas chromatographic column is not appreciably degraded. Cold spots
or active surfaces (adsorption sites) in the GC/MS interface can cause peak tailing and peak
broadening. It is recommended that the GC column be fitted directly into the mass
spectrometer ion source without being exposed to the ionizing electron beam. Graphite
ferrules should be avoided in the injection port because they may adsorb the PCDDs and
PCDFs. Vespel™, or equivalent, ferrules are recommended.
8290A-4
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4.1.3 Mass spectrometer - The static resolving power of the instrument must be
maintained at a minimum of 10,000 (10 percent valley).
4.1.4 Data system - A dedicated data system is employed to control the rapid
multiple-ion monitoring process and to acquire the data. Quantitation data (peak areas or peak
heights) and SIM traces (displays of intensities of each ion signal being monitored including
the lock-mass ion as a function of time) must be acquired during the analyses and stored.
Quantitations may be reported based upon computer-generated peak areas or upon measured
peak heights (chart recording). The data system must be capable of acquiring data at a
minimum of 10 ions in a single scan. It is also recommended to have a data system capable
of switching to different sets of ions (descriptors) at specified times during an HRGC/HRMS
acquisition. The data system should be able to provide hard copies of individual ion
chromatograms for selected gas chromatographic time intervals. It should also be able to
acquire mass spectral peak profiles (Sec. 8.1.2.3) and provide hard copies of peak profiles to
demonstrate the required resolving power. The data system should permit the measurement
of noise on the base line.
NOTE: The detector ADC zero setting must allow peak-to-peak measurement of the noise
on the base line of every monitored channel and allow for good estimation of the
instrument resolving power. The effect of different zero settings on the measured
resolving power is shown in Figure 2.
4.2 GC columns
Fused-silica capillary columns are needed. The columns shall demonstrate the required
separation of all 2,3,7,8-specific isomers whether a dual-column or a single-column analysis is
chosen. Chromatographic performance must be demonstrated and documented (Sec. 8.2.2) at the
beginning of each 12-hour period (after mass resolution and GC resolution are demonstrated) during
which sample extracts or concentration calibration solutions will be analyzed. Recommended
operating conditions for the recommended columns are shown in Sec. 7.6.
4.2.1 60-m DB-5 (J&W Scientific) or equivalent fused-silica capillary column
In order to have an isomer-specific determination of 2,3,7,8-TCDD and to allow the
detection of OCDD/OCDF within a reasonable time interval in one HRGC/HRMS analysis, use
of this column is recommended. Isomer specificity for all 2,3,7,8-substituted PCDDs/PCDFs
cannot be achieved on the 60-m DB-5 column. Problems have been associated with the
separation of 2,3,7,8-TCDD from 1,2,3,7-TCDD and 1,2,6,8-TCDD, and separation of 2,3,7,8-
TCDF from 1,2,4,9-, 1,2,7,9-, 2,3,4,6-, 2,3,4,7-, and 2,3,4,8-TCDF. Because of the toxicologic
concern associated with 2,3,7,8-TCDD and 2,3,7,8-TCDF, additional analyses may be
necessary for some samples.
4.2.2 30-m DB-225 (J&W Scientific) or equivalent fused-silica capillary column
For the DB-225 column, problems are associated with the separation of 2,3,7,8-TCDF
from 2,3,4,7-TCDF and a combination of 1,2,3,9- and 2,3,4,8-TCDF.
8290A-5
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4.3 Miscellaneous equipment and materials - The following list of Hems does not
necessarily constitute an exhaustive compendium of the equipment needed for this analytical
method.
NOTE: Reuse of glassware should be minimized to avoid the risk of contamination. All glassware
that is reused should be scrupulously cleaned as soon as possible after use, according to
the following procedure: Rinse glassware with the last solvent used in it. Wash with hot
detergent water, then rinse with copious amounts of tap water and several portions of
organic-free reagent water. Rinse with high purity acetone and hexane and store it
inverted or capped with solvent rinsed aluminum foil in a clean environment.
4.3.1 Nitrogen evaporation apparatus with variable flow rate.
4.3.2 Balances capable of accurately weighing to 0.01 g and 0.0001 g.
4.3.3 Centrifuge.
4.3.4 Water bath - equipped with concentric ring covers and capable of being
temperature controlled within ± 2°C.
4.3.5 Stainless steel or glass container large enough to hold contents of one-pint
sample containers.
4.3.6 Glove box.
4.3.7 Drying oven.
4.3.8 Stainless steel spoons and spatulas.
4.3.9 Laboratory hoods.
4.3.10 Pipets - disposable, Pasteur, 150 mm long x 5 mm ID.
4.3.11 Pipets - disposable, serological, 10-mL, for the preparation of the carbon
columns specified in Sec. 7.5.3.
4.3.12 Reaction vial - 2-mL, silanized amber glass (Reacti-vial, or equivalent).
4.3.13 Stainless steel meat grinder with a 3 to 5 mm hole size inner plate.
4.3.14 Separatory funnels - 125-ml and 2000-mL.
4.3.15 Kudema-Danish concentrator - 500-mL, fitted with 10-mL concentrator tube
and three-ball Snyder column.
4.3.16 PTFE or Carborundum (silicon carbide) boiling chips (or equivalent), washed
with hexane before use.
NOTE: PFTE boiling chips may float in methylene chloride, may not woric in the presence of
any water phase, and may be penetrated by nonpolar organic compounds.
8290A-6
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4.3.17 Chromatographic columns - glass, 300 mm x 10.S mm, fitted with PTFE
stopcock.
4.3.18 Adapters for concentrator tubes.
4.3.19 Glass fiber filters - 0.70-Mm, Whatman GFF, or equivalent.
4.3.20 Dean-Stark trap - 5- or 10-mL, with T-joints, condenser and 125-mL flask.
4.3.21 Continuous liquid-liquid extractor - 1-L sample capacity, suitable for use with
heavier than water solvents.
4.3.22 All glass Soxhlet apparatus with 500-mL flask.
4.3.23 Soxhlet/Dean-Stark extractor (optional) - all glass, 500-mL flask.
4.3.24 Glass funnels - sized to hold 170 mL of liquid.
4.3.25 Desiccator.
4.3.26 Solvent reservoir (125-mL) - compatible with gravity carbon column.
4.3.27 Rotary evaporator witfi a temperature-controlled water bath.
4.3.28 High speed tissue homogenizer - equipped with an EN-8 probe, or equivalent.
4.3.29 Glass wool - extract with methylene chloride, dry, and store in a glass jar.
4.3.30 Extraction jars - glass, 250-mL, with PTFE-lined screw cap.
4.3.31 Volumetric flasks - Class A, 10-mL to 1000-mL.
4.3.32 Glass vials - 1-dram (or metric equivalent).
5.0 REAGENTS AND STANDARD SOLUTIONS
5.1 Organic-free reagent water - All references to water in this method refer to organic-free
reagent water, as defined in Chapter One.
5.2 Column chromatography reagents
5.2.1 Alumina • neutral, 80/200 mesh (Super 1, Woelm®, or equivalent). Store in
a sealed container at room temperature, in a desiccator, over self-indicating silica gel.
5.2.2 Alumina - acidic AG4, (Bio Rad Laboratories catalog #132-1240, or
equivalent). Soxhlet extract with methylene chloride for 24 hours if blanks show contamination,
and activate by heating in a foil covered glass container for 24 hours at 190°C. Store in a
glass bottle sealed with a PTFE-lined screw cap.
5.2.3 Silica gel - high purity grade, type 60, 70-230 mesh. Soxhlet extract with
methylene chloride for 24 hours if blanks show contamination, and activate by heating in a foil
8290A - 7
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covered glass container for 24 hours at 190°C. Store in a glass bottle sealed with a PTFE-
lined screw cap.
5.2.4 Silica gel impregnated with sodium hydroxide. Add one part (by weight) or 1
M NaOH solution to two parts (by weight) silica gel (extracted and activated) in a screw cap
bottle and mix with a glass rod until free of lumps. Store in a glass bottle sealed with a PTFE-
iined screw cap.
5.2.5 Silica gel impregnated with 40 percent (by weight) sulfuric acid. Add two parts
(by weight) concentrated sulfuric add to three parts (by weight) silica gel (extracted and
activated), mix with a glass rod until free of lumps, and store in a screw capped glass bottle.
Store in a glass bottle sealed with a PTFE-lined screw cap.
5.2.6 Celite 545® (Supelco), or equivalent.
5.2.7 Charcoal carbon - Activated carbon, Carbopak C (Supelco) or equivalent,
prewashed with methanol and dried in vacuo at 110°C. Store in a glass bottle sealed with a
PTFE-lined screw cap. (Note: AX-21 [Anderson Development Company] carbon is no longer
available, but existing stocks may be utilized).
5.3 Reagents
5.3.1 Sulfuric acid, H2SO4, concentrated, ACS grade, specific gravity 1.84.
5.3.2 Potassium hydroxide, KOH, ACS grade, 20 percent (w/v) in organic-free
reagent water.
5.3.3 Sodium chloride, NaCI, analytical reagent, 5 percent (w/v) in organic-free
reagent water.
5.3.4 Potassium carbonate, K2C03, anhydrous, analytical reagent.
5.4 Sodium sulfate (powder, anhydrous), Na2S04 - Purify by heating at 400°C for 4 hours
in a shallow tray. If, after heating, the sodium sulfate develops a noticeable grayish cast (due to the
presence of carbon in the crystal matrix) that batch of sodium sulfate is not suitable for use and
should be discarded. Extraction with methylene chloride may produce sodium sulfate that is suitable
for use in such instances, but following extraction, a reagent blank must be analyzed that
demonstrates that there is no interference from the sodium sulfate.
5.5 Solvents - all solvents must be (at a minimum) pesticide grade or equivalent, distilled-
in-glass.
5.5.1
Methylene chloride, CHjClj.
5.5.2
Hexane, CgH14.
5.5.3
Methanol, CH3OH.
5.5.4
Nonane, CgH^.
5.5.5
Toluene, C^CHa.
8290A - 8
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5.5.6 Cyclohexane, C6H12.
5.5.7 Acetone, CH3COCH3.
5.6 High-Resolution Concentration Calibration Solutions (Table 5) - Five nonane solutions
containing 17 unlabeled and 11 carbon-labeled PCDDs and PCDFs at known concentrations are
used to calibrate the instrument. The concentration ranges are homologue-dependent, with the
lowest values for the tetrachlorinated dioxin and furan (1.0 pg/pL) and the highest values for the
octachlorinated congeners (1000 pg/pL). Standards containing more carbon-labeled PCDDs and
PCDFs may also be employed.
5.7 GC Column Performance Check Solution - This solution contains the first and last
eluting isomers for each homologous series from tetra- through heptachlorinated congeners. The
solution also contains a series of other TCDD isomers for the purpose of documenting the
chromatographic resolution. The 13C1z-2,3,7,8-TCDD is also present. The laboratory is required to
use nonane as the solvent and adjust the volume so that the final concentration does not exceed
100 pg/pL per congener. Table 7 summarizes the qualitative composition (minimum requirement)
of this performance evaluation solution.
5.8 Sample Fortification Solution - This nonane solution contains the nine internal
standards at the nominal concentrations that are listed in Table 2. The solution contains at least one
carbon-labeled standard for each homologous series, and it is used to measure the concentrations
of the native substances. (Note that "C12-OCDF is not present in the solution.) Standards
containing more carbon-labeled PCDDs and PCDFs may also be employed, provided that the same
labeled compounds are contained in the calibration standards in Sec. 5.6.
5.9 Recovery Standard Solution - This nonane solution contains two recovery standards,
13C12-1,2,3,4-TCDD and 1t12-1,2,3,7,8,9-HxCDD, at a nominal concentration of 50 pg/pL per
compound. 10 to 50 pL of this solution will be spiked into each sample extract before the final
concentration step and HRGC/HRMS analysis.
5.10 Matrix Spike Fortification Solution - Solution used to prepare the MS and MSD samples.
It contains all unlabeled analytes listed in Table 5 at concentrations corresponding to the HRCC 3.
6.0 SAMPLE COLLECTION, PRESERVATION, AND HANDLING
6.1 See the introductory material to this chapter, Organic Analytes, Sec. 4.1.
6.2 Sample collection
6.2.1 Sample collection personnel should, to the extent possible, homogenize
samples in the field before filling the sample containers. This should minimize or eliminate the
necessity for sample homogenization in the laboratory. The analyst should make a judgment,
based on the appearance of the sample, regarding the necessity for additional mixing. If the
sample is clearly not homogeneous, the entire contents should be transferred to a glass or
stainless steel pan for mixing with a stainless steel spoon or spatula before removal of a
sample portion for analysis.
6.2.2 Grab and composite samples must be collected in glass containers.
Conventional sampling practices must be followed. The bottle must not be prewashed with
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sample before collection. Sampling equipment must be free of potential sources of
contamination.
6.3 Grinding or blending of fish samples - If not otherwise specified in a project plan, the
whole fish (frozen) should be blended or ground to provide a homogeneous sample. The use of a
stainless steel meat grinder with a 3 to 5 mm hole size inner plate is recommended. In some
circumstances, analysis of fillet or specific organs of fish may be requested. If so requested, the
above whole fish requirement is superseded.
6.4 Storage and holding times - All samples, except fish and adipose tissue samples, must
be stored at 4°C in the dark, and should be extracted within 30 days and completely analyzed within
45 days of extraction. Fish and adipose tissue samples must be stored at -20°C in the dark, and
should be extracted within 30 days and completely analyzed within 45 days of collection.
NOTE: The holding times listed in Sec. 6.4 are recommendations. PCDDs and PCDFs are very
stable in a variety of matrices, and holding times under the conditions listed in Sec. 6.4
may be as high as a year for certain matrices.
6.5 Phase separation
This is a guideline for phase separation for very wet (>25 percent water) soil, sediment and
paper pulp samples. Place a 50-g portion in a suitable centrifuge bottle and centrifuge for
30 minutes at 2,000 rpm. Remove the bottle and mark the interface level on the bottle. Estimate
the relative volume of each phase. With a disposable pipet, transfer the liquid layer into a clean
bottle. Mix the solid with a stainless steel spatula and remove a portion to be weighed and analyzed
(percent dry weight determination, extraction). Return the remaining solid portion to the original
sample bottle (empty) or to a clean sample bottle that is properly labeled, and store it as appropriate.
Analyze the solid phase by using only the soil, sediment and paper pulp method. Take note of, and
report, the estimated volume of liquid before disposing of the liquid as a liquid waste.
6.6 Soil, sediment, or paper sludge (pulp) percent dry weight determination
When results are to be reported on a dry-weight basis, the percent dry weight of soil, sediment
or paper pulp samples may be determined according to the following procedure. Weigh a 10-g
portion of the soil or sediment sample (± 0.5 g) to three significant figures. Dry it to constant weight
at 110 °C in an adequately ventilated oven. Allow the sample to cool in a desiccator. Weigh the
dried solid to three significant figures. Calculate and report the percent dry weight. Do not use this
solid portion of the sample for extraction, but instead dispose of it as hazardous waste.
% dry weight = 9 °* d>y sample * 100
g of sample
CAUTION: Finely divided soils and sediments contaminated with PCDDs/PCDFs are hazardous
because of the potential for inhalation or ingestion of particles containing
PCDDs/PCDFs (including 2,3,7,8-TCDD). Such samples should be handled in a
confined environment (i.e., a closed hood or a glove box).
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6.7 Lipid content determination
6.7,1 Fish tissue - To determine the lipid content of fish tissue, concentrate 125 mL
of the fish tissue extract (Sec. 7.2.2), in a tared 200-mL round-bottom flask, on a rotary
evaporator until a constant weight (W) is achieved.
lipid - we'9ht residue * 2 x ^
weight of sample
The factor of 2 accounts for the use of half of the extract (e.g., 125 mL of 250 mL total volume)
for the lipid determination.
Dispose of the lipid residue as a hazardous waste if the results of the analysis indicate
the presence of PCDDs or PCDFs.
Other procedures and other extract volumes may be employed for the lipid
determination, provided that they are clearly described and documented. Adjustments to the
amount of internal standards spiked in Sec. 7.1 will be required if different volumes are
employed.
6.7.2 Adipose tissue - Details for the determination of the adipose tissue lipid
content are provided in Sec. 7.3.3.
PROCEDURE
7.1 Internal standard addition
The sample fortification solution (Sec. 5.8) containing the carbon-labeled internal
standards is added to each sample prior to extraction.
7.1.1 Select an appropriate size sample aliquot Typical sample size requirements
for different matrices are given in Sec. 7.4 and in Table 1. Transfer the sample portion to a
tared flask and determine its weight.
7.1.2 Except for adipose tissue, add an appropriate quantity of the sample
fortification mixture (Sec. 5.8) to the sample. Ail samples should be spiked with 100 jjL of the
sample fortification mixture to give internal standard concentrations, as indicated in Table 1.
As an example, for 13C12-2,3,7,8-TCDD, a 10-g soil sample requires the addition of 1000 pg of
1Jfc12-2,3,7,8-TCDD to give the required 100 ppt fortification level. The fish tissue sample (20
g) must be spiked with 200 pL of the internal standard solution, because half of the extract will
be used to determine the lipid content (Sec. 6.7.1).
7.1.2.1 For the fortification of soil, sediment, fly ash, water, fish tissue,
paper pulp and wet sludge samples, mix the sample fortification solution with 1.0 mL
acetone.
7.1.2.2 Do not dilute the nonane solution for the other matrices.
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7.1.2.3 The fortification of adipose tissue is carried out at the time of
homogenization (Sec. 7.3.2.3).
7.2 Extraction and purification of fish and paper pulp samples
7.2.1 Add 60 g of anhydrous sodium sulfate to a 20-g portion of a homogeneous
fish sample (Sec. 6.3) and mix thoroughly with a stainless steel spatula. After breaking up any
lumps, place the fish/sodium sulfate mixture in the Soxhlet apparatus on top of a glass wool
plug. Add 250 mL methylene chloride or hexane/methylene chloride (1:1) to the Soxhlet
apparatus and reflux for 16 hours. "Hie solvent must cycle completely through the system five
times per hour. Follow the same procedure for the partially dewatered paper pulp sample
(using a 10-g sample, 30 g of anhydrous sodium sulfate and 200 ml of toluene).
NOTE: As an option, a Soxhlet/Dean-Stark extractor system may be used, with toluene as
the solvent. No sodium sulfate is added when using this option.
7.2.2 Transfer the fish extract from Sec. 7.2.1 to a 250-mL volumetric flask and fill
to the mark with methylene chloride. Mix well, then remove 125 mL for the determination of
the lipid content (Sec. 6.7.1). Transfer the remaining 125 mL of the extract, plus two 15-mL
hexane/methylene chloride rinses of the volumetric flask, to a K-D apparatus equipped with a
Snyder column. Quantitatively transfer all of the paper pulp extract to a K-D apparatus
equipped with a Snyder column.
mm. as an option, a rotary evaporator may be used in place of the K-D apparatus for the
concentration of the extracts.
7.2.3 Add a PTFE (or equivalent) boiling chip. Concentrate the extract in a water
bath to an apparent volume of 10 mL. Remove the apparatus from the water bath and allow
to cool for 5 minutes.
7.2.4 Add 50 mL hexane and a new boiling chip to the K-D flask. Concentrate in
a water bath to an apparent volume of 5 mL. Remove the apparatus from the water bath and
allow to cool for 5 minutes.
NOTE: The methylene chloride must have been completely removed before proceeding with
the next step.
7.2.5 Remove and invert the Snyder column and rinse it into the K-D apparatus with
two 1-mL portions of hexane. Decant the contents of the K-D apparatus and concentrator tube
into a 125-mL separatory funnel. Rinse the K-D apparatus with two additional 5-mL portions
of hexane and add the rinses to the funnel. Proceed with the cleanup according to the
instructions in Sec. 7.5.1.1, but omit the procedures described in Sees. 7.5.1.2 and 7.5.1.3.
7.3 Extraction and purification of human adipose tissue
7.3.1 Human adipose tissue samples must be stored at a temperature of -20°C or
lower from the time of collection until the time of analysis. The use of chlorinated materials
during the collection of the samples must be avoided. Samples are handled with stainless
steel forceps, spatulas, or scissors. All sample bottles (glass) are cleaned as specified in the
note at the end of Sec. 4.3. PTFE-lined caps should be used.
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NOTE: The specified storage temperature of -20°C is the maximum storage temperature
permissible for adipose tissue samples. Lower storage temperatures are
recommended.
7.3.2 Adipose tissue extraction
7.3.2.1 Weigh a 10-g portion of a frozen adipose tissue sample to the
nearest 0.01 g, into a culture tube (2.2 x 15 cm).
NOTE: The sample size may be smaller, depending on availably, in such situations,
the analyst is required to adjust the volume of the internal standard solution
added to the sample to meet the fortification level stipulated in Table 1.
7.3.2.2 Allow the adipose tissue specimen to reach room temperature (up
to 2 hours).
7.3.2.3 Add 10 mL of methylene chloride and "100 pL of the sample
fortification solution. Homogenize the mixture for approximately 1 minute with a tissue
homogenizes
7.3.2.4 Allow the mixture to separate, then remove the methylene chloride
extract from the residual solid material with a disposable pipet. Percolate the methylene
chloride through a filter funnel containing a clean glass wool plug and 10 g of
anhydrous sodium sulfate. Collect the dried extract in a graduated 100-mL volumetric
flask.
7.3.2.5 Add a second 10 mL portion of methylene chloride to the sample
and homogenize for 1 minute. Decant the solvent, dry it, and transfer it to the 100-mL
volumetric flask (Sec. 7.3.2.4).
7.3.2.6 Rinse the culture tube with at least two additional portions of
methylene chloride (10-mL each), and transfer the entire contents to the filter funnel
containing the anhydrous sodium sulfate. Rinse the filter funnel and the anhydrous
sodium sulfate contents with additional methylene chloride (20 to 40 mL) into the 100-
mL flask. Discard the sodium sulfate.
7.3.2.7 Adjust the volume to the 100-mL marie with methylene chloride.
7.3.3 Adipose tissue lipid content determination
7.3.3.1 Preweigh a clean 1-dram (or metric equivalent) glass vial to the
nearest 0.0001 g on an analytical balance tared to zero.
7.3.3.2 Accurately transfer 1.0 mL of the final extract (100 mL) from Sec.
7.3.2.7 to the vial. Reduce the volume of the extract on a water bath (50-60°C) by a
gentle stream of purified nitrogen until an oily residue remains. Nitrogen evaporation
is continued until a constant weight is achieved.
NOTE: When the sample size of the adipose tissue is smaller than 10 g, then the
analyst may use a larger portion (up to 10 percent) of the extract defined in
Sec. 7.3.2.7 for the lipid determination.
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7.3.3.3 Accurately weigh the vial with the residue to the nearest 0.0001
g and calculate the weight of the lipid present in the vial based on the difference of the
weights.
7.3.3.4 Calculate the percent lipid content of the original sample to the
nearest 0.1 percent as shown below:
W„ *
% ypjd = —1 St x 100
W* * V„
where:
W,r = weight of the lipid residue to the nearest 0.0001 g calculated from Sec.
7.3.3.3,
= total volume (100 mL) of the extract in mL from Sec. 7.3.2.7,
= weight of the original adipose tissue sample to the nearest 0.01 g from Sec.
7.3.2.1, and
V* = volume of the aliquot of the final extract in mL used for the quantitative
measure of the lipid residue (1.0 mL) from Sec. 7.3.3.2.
7.3.3.5 Record the weight of the lipid residue measured in Sec. 7.3.3.3
and the percent lipid content from Sec. 7.3.3.4.
7.3.4 Adipose tissue extract concentration
7.3.4.1 Quantitatively transfer the remaining extract from Sec. 7,3.3.2
(99,0 mL) to a 500-mL Erlenmeyer flask. Rinse the volumetric flask with 20 to 30 ml
of additional methylene chloride to ensure quantitative transfer.
7.3.4.2 Concentrate the extract on a rotary evaporator and a water bath
at 40 °C until an oily residue remains.
7.3.5 Adipose tissue extract cleanup
7.3.5.1 Add 200 mL of hexane to the lipid residue in the 500-mL
Erlenmeyer flask and swirl the flask to dissolve the residue.
7.3.5.2 Slowly add, with stirring, 100 g of 40 percent (w/w) sulfuric acid-
impregnated silica gel. Stir with a magnetic stirrer for two hours at room temperature.
7.3.5.3 Allow the solid phase to settle, and decant the liquid through a
filter funnel containing 10 g of anhydrous sodium sulfate on a glass woo! plug, into
another 500-mL Erlenmeyer flask.
7.3.5.4 Rinse the solid phase with two 50-mL portions of hexane. Stir
each rinse for 15 minutes, decant, and dry as described under Sec. 7.3.5.3, Combine
the hexane extracts from Sec. 7.3.5.3 with the rinses.
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7.3.5.5 Rinse the sodium sulfate in the filter funnel with an additional
25 mL of hexane and combine this rinse with the hexane extracts from Sec. 7.3.5.4.
7.3.5.6 Prepare an acidic silica column as follows: Pack a 2 cm x 10 cm
chromatographic column with a glass wool plug, add approximately 20 mL of hexane,
add 1 g of silica gel and allow to settle, then add 4 g of 40 percent (w/w) sulfuric acid-
impregnated silica gel and allow to settle. Elute the excess hexane from the column
until the solvent level reaches the top of the chromatographic packing. Verify that the
column does not have any air bubbles and channels.
7.3.5.7 Quantitatively transfer the hexane extract from the Erlenmeyer
flask (Sees. 7.3.5.3 through 7.3.5.5) to the silica gel column reservoir. Allow the
hexane extract to percolate through the column and collect the eluate in a 500-mL K-D
apparatus.
7.3.5.8 Complete the elution by percolating 50 mL of hexane through the
column into the K-D apparatus. Concentrate the eluate on a steam bath to about 5 mL.
Use nitrogen evaporation to bring the final volume to about 100 jiL.
NOTE: If the silica gel impregnated with 40 percent sulfuric add is highly discolored
throughout the length of the adsorbent bed, the cleaning procedure must be
repeated beginning with Sec. 7.3.5.1.
7.3.5.9 The extract is ready for the column cleanups described in
Sees. 7.5.2 through 7.5.3,6.
Extraction and purification of environmental and waste samples
7.4.1 Sludge/wet fuel oil
7.4.1.1 Extract aqueous sludge or wet fuel oil samples by refluxing a
sample (e.g., 2 g) with 50 mL of toluene in a 125-mL flask fitted with a Dean-Stark
water separator. Continue refluxing the sample until all the water is removed.
NOTE: If the sludge or fuel oil sample dissolves in toluene, treat it according to the
instructions in Sec. 7.4.2 below. If the sludge sample originates from pulp
(paper mills), beat it according to the instructions starting in Sec. 7.2, but
without the addition of sodium sulfate.
7.4.1.2 Cool the sample, filter the toluene extract through a glass fiber
filter, or equivalent, into a 100-mL round-bottom flask.
7.4.1.3 Rinse the filter with 10 mL of toluene and combine the extract with
the rinse.
7.4.1.4 Concentrate the combined solutions to near dryness on a rotary
evaporator at 50°C or using nitrogen evaporation. Proceed with Sec. 7.4.4.
7.4.2 Still bottom/oil
7.4.2.1 Extract still bottom or oil samples by mixing a sample portion (e.g.,
1.0 g) with 10 mL of toluene in a small beaker and filtering the solution through a glass
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fiber filter (or equivalent) into a 50-mL round-bottom flask. Rinse the beaker and filter
with 10 mL of toluene.
7.4.2.2 Concentrate the combined toluene solutions to near dryness on
a rotary evaporator at 50° C or using nitrogen evaporation. Proceed with Sec. 7.4.4.
7.4.3 Fly ash
NOTE: Because of the tendency of fly ash to "fly", all handling steps should be
performed in a hood in order to minimize contamination.
7.4.3.1 Weigh about 10 g of % ash to two decimal places and transfer to
an extraction jar. Add 100 (jL of the sample fortification solution (Sec. 5.8), diluted to
1 mL with acetone, to the sample. Add 150mLof 1 M HCI to the fly ash sample. Seal
the jar with the PTFE-lined screw cap and shake for 3 hours at room temperature.
7.4.3.2 Rinse a glass fiber filter with toluene, and flter the sample through
the filter paper, placed in a Buchner funnel, into a 1-L flask. Wash the fly ash cake with
approximately 500 mL of organic-free reagent water and dry the filter cake overnight
at room temperature in a desiccator.
7.4.3.3 Add 10 g of anhydrous powdered sodium sulfate, mix thoroughly,
let sit in a closed container for one hour, mix again, let sit for another hour, and mix
again.
7.4.3.4 Place the sample and the filter paper into an extraction thimble,
and extract in a Soxhlet extraction apparatus charged with 200 mL of toluene for
16 hours using a five cycle/hour schedule.
NOTE: As an option, a Soxhletf Dean-Stark extractor system may be used, with
toluene as the solvent. No sodium sulfate is added when using this option.
7.4.3.5 Cool and filter the toluene extract through a glass fiber filter into
a 500-mL round-bottom flask. Rinse the filter with 10 mL of toluene. Add the rinse to
the extract and concentrate the combined toluene solutions to near dryness on a rotary
evaporator at 50°C or using nitrogen evaporation. Proceed with Sec. 7.4.4.
7.4.3.6 Alternatively, fly ash samples may be extracted with a
toluene/acetic acid mixture using pressurized fluid extraction (PFE), as described in
Method 3545. When using PFE, the HCI pretreatment in Sec. 7.4.3.1 may be omitted.
7.4.4 Transfer the concentrate to a 125-mL separately funnel using 15 mL of
hexane. Rinse the flask with two 5-mL portions of hexane and add the rinses to the funnel.
Shake the combined solutions in the separatory funnel for two minutes with 50 mL of 5 percent
sodium chloride solution, discard the aqueous layer, and proceed with Sec. 7.5.
7.4.5 Aqueous samples
7.4.5.1 Allow the sample to come to ambient temperature, then mark the
water meniscus on the side of the 1-L sample bottle for later determination of the exact
sample volume. Add the required acetone diluted sample fortification solution
(Sec. 5.8).
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7.4.5.2 When the sample is judged to contain 1 percent or more solids,
the sample must be filtered through a glass fiber filter that has been rinsed with
toluene. If the suspended solids content is too great to filter through the 0.45-jjm filter,
centrifuge the sample, decant, and then filter the aqueous phase.
NOTE: Paper mill effluent samples normally contain 0.02%-0.2% solids, and would
not require filtration. However, for optimum analytical results, all paper mill
effluent samples should be filtered, the isolated solids and filtrate extracted
separately, and the extracts recombined.
7.4.5.3 Combine the solids from the centrifuge bottle(s) with the
particulates on the filter and with the filter itself and proceed with the Soxhlet extraction
as specified in Sees. 7.4.6.1 through 7.4.6.4.
NOTE: Pressurized fluid extraction has not been evaluated for the extraction of the
particulate fraction.
Remove and invert the Snyder column and rinse it down into the K-D apparatus with
two 1-mL portions of hexane.
7.4.5.4 Pour the aqueous filtrate into a 2-1 separately funnel. Add 60 mL
of methylene chloride to the sample bottle, seal and shake for 30 seconds to rinse the
inner surface. Transfer the solvent to the separatory funnel and extract the sample by
shaking the funnel for two minutes with periodic venting.
7.4.5.5 Allow the organic layer to separate from the water phase for a
minimum of 10 minutes. If the emulsion interface between layers is more than one
third the volume of the solvent' layer, the analyst must employ mechanical techniques
to complete the phase separation (e.g., glass stimng rod).
7.4.5.6 Collect the methylene chloride in a K-D apparatus (mounted with
a 10-mL concentrator tube) by passing the sample extracts through a filter funnel
packed with a glass wool plug and 5 g of anhydrous sodium sulfate.
NOTE: As an option, a rotary evaporator may be used in place of the K-D apparatus
for the concentration of the extracts.
7.4.5.7 Repeat the extraction twice with fresh 60-mL portions of
methylene chloride. After the third extraction, rinse the sodium sulfate with an
additional 30 mL of methylene chloride to ensure quantitative transfer. Combine all
extracts and the rinse in the K-D apparatus.
NOTE: A continuous liquid-liquid extractor may be used in place of a separatory
funnel when experience with a sample from a given source indicates that a
serious emulsion problem will result or an emulsion is encountered when
using a separatory funnel. Add 60 mL of methylene chloride to the sample
bottle, seal, and shake for 30 seconds to rinse the inner surface. Transfer
the solvent to the extractor. Repeat the rinse of the sample bottle with an
additional 50- to 100-mL portion of methylene chloride and add the rinse to
the extractor. Add 200 to 500 mL of methylene chloride to the distilling flask,
add sufficient organic-free reagent water (Sec. 5.1) to ensure proper
operation, and extract for 24 hours. Allow to cool, then detach the distilling
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flask. Dry and concentrate the extract as described in Sees. 7.4.5.6 and
7.4.5.8 through 7.4.5.10. Proceed with Sec. 7.4.5.11.
7.4.5.8 Attach a Snyder column and concentrate the extract on a water
bath until the apparent volume of the liquid is 5 mL. Remove the K-D apparatus and
allow it to drain and cool for at least 10 minutes.
7.4.5.9 Remove the Snyder column, add 50 mL of hexane, add the
concentrate obtained from the Soxhlet extraction of the suspended solids
(Sec. 7.4.5.3}, if applicable, re-attach the Snyder column, and concentrate to
approximately 5 mL. Add a new boiling chip to the K-D apparatus before proceeding
with the second concentration step.
7.4.5.10 Rinse the flask and the lower joint with two 5-mL portions of
hexane and combine the rinses with the extract to give a final volume of about 15 mL
7.4.5.11 Determine the original sample volume by filling the sample bottle
to the mark with water and transferring the water to a 1000-mL graduated cylinder.
Record the sample volume to the nearest 5 mL Proceed with Sec. 7.5.
7.4.6 Soil/sediment
7.4.6.1 Add 10 g of anhydrous powdered sodium sulfate to the sample
aliquot (10 g or less) and mix thoroughly with a stainless steel spatula. After breaking
up any lumps, place the soil/sodium sulfate mixture in the Soxhlet apparatus on top of
a glass wool plug (the use of an extraction thimble is optional).
NOTE: As an option, a Soxhlet/Dean-Stark extractor system may be used, with
toluene as the solvent. No sodium sulfate is added when using this option.
7.4.6.2 Add 200 to 250 mL of toluene to the Soxhlet apparatus and reflux
for 16 hours. The solvent must cycle completely through the system Ave times per
hour.
NOTE: If the dried sample is not of free flowing consistency, more sodium sulfate
must be added.
7.4.6.3 Cool and filter the extract through a glass fiber filter into a 500-mL
round-bottom flask for evaporation of the toluene. Rinse the Alter with 10 mL of
toluene, and concentrate the combined fractions to near dryness on a rotary evaporator
at 50 °C. Remove the flask from the water bath and allow to cool for 5 minutes.
7.4.6.4 Transfer the residue to a 125-mL separatory funnel, using 15 mL
of hexane. Rinse the flask with two additional portions of hexane, and add the rinses
to the funnel. Proceed with Sec. 7.5.
7.4.6.5 Alternatively, soil/sediment samples may be extracted with
toluene using pressurized fluid extraction (PFE), as described in Method 3545.
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7.5 Cleanup
7.5.1 Acid-base washing
7.5.1.1 Partition the hexane extract against 40 mL of concentrated sulfuric
acid. Shake for two minutes. Remove and discard the sulfuric acid layer (bottom).
Repeat the acid washing until no color is visible in the add layer (perform a maximum
of four add washings).
7.5.1.2 Omit this step for the fish sample extract. Partition the extract
against 40 mL of 5 percent (w/v) aqueous sodium chloride. Shake for two minutes.
Remove and discard the aqueous layer (bottom).
7.5.1.3 Omit this step for the fish sample extract. Partition the extract
against 40 mL of 20 percent (w/v) aqueous potassium hydroxide (KOH), Shake for
two minutes. Remove and discard the aqueous layer (bottom). Repeat the base
washing until no color is visible in the bottom layer (perform a maximum of four base
washings). Strong base (KOH) is known to degrade certain PCDDs/PCDFs, so contact
time must be minimized.
7.5.1.4 Partition the extract against 40 mL of 5 percent (w/v) aqueous
sodium chloride. Shake for two minutes. Remove and discard the aqueous layer
(bottom). Dry the extract by pouring it through a Alter funnel containing anhydrous
sodium sulfate on a glass wool plug, and collect it in a 50-mL round-bottom flask.
Rinse the funnel with the sodium sulfate with two 15-mL portions of hexane, add the
rinses to the 50-mL flask, and concentrate the hexane solution to near dryness on a
rotary evaporator (35°C water bath) or nitrogen evaporation, making sure ail traces of
toluene (when applicable) are removed.
7.5.2 Silica/alumina column deanup
7.5.2.1 Pack a gravity column (glass, 30 cm x 10.5 mm), fitted with a
PTFE stopcock, with of silica gel as follows: Insert a glass wool plug into the bottom
of the column. Place 1 g of silica gef in the column and tap the column gently to settle
the silica gel. Add 2 g of sodium hydroxide-impregnated silica gel, 4 g of sulfuric add-
impregnated silica gel, and 2 g of silica gel. Tap the column gently after each addition.
A small positive pressure (5 psi) of dean nitrogen may be used if needed. Bute with
10 mL of hexane and close the stopcock just before exposure of the top layer of silica
gel to air. Discard the eluate. Check the column for channeling. If channeling is
observed, discard the column. Do not tap the wetted column.
7.5.2.2 Pack a gravity column (glass, 300 mm x 10.5 mm), fitted with a
PTFE stopcock, with alumina as follows: Insert a glass wool plug into the bottom of
the column. Add a 4 g layer of sodium sulfate. Add a 4 g layer of Woelm® Super 1
neutral alumina. Tap the top of the column gently. Woelm® Super 1 neutral alumina
need not be activated or deaned before use, but it should be stored in a sealed
desiccator. Add a 4 g layer of anhydrous sodium sulfate to cover the alumina. Elute
with 10 mL hexane and dose the stopcock just before exposure of the sodium sulfate
layer to air. Discard the eluate. Check the column for channeling. If channeling is
observed, discard the column. Do not tap a wetted column.
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NOTE: Alternatively, acidic alumina (Sec. 5.2.2} may be used in place of neutral
alumina.
7.5.2.3 Dissolve the residue from Sec. 7.5.1.4 in 2 mL of hexane and
apply the hexane solution to the top of the silica gel column. Rinse the flask with
enough hexane (3-4 mL) to quantitatively transfer of the sample to the surface of the
silica gel.
7.5.2.4 Elute the sflica gel column with 90 mL of hexane, concentrate the
eluate on a rotary evaporator (35°C water bath} to approximately 1 mL, and apply the
concentrate to the top of the alumina column (Sec. 7.5.2.2). Rinse the rotary
evaporator flask twice with 2 mL of hexane, and add the rinses to the top of the
alumina column.
7.5.2.5 Add 20 mL of hexane to the alumina column and elute until the
hexane level is just below the top of the sodium sulfate. Do not discard the eluted
hexane, but collect it in a separate flask and store N for later use, as it may be useful
in determining where the labeled analytes are being lost if recoveries are not
satisfactory.
7.5.2.6 Add 15 mL of 60 percent methylene chloride in hexane (v/v) to the
alumina column and collect the eluate in a conical-shaped (15-mL) concentration tube.
With a carefully regulated stream of nitrogen, concentrate the 60 percent methylene
chloride/hexane fraction to about 2 mL.
7.5.3 Carbon column cleanup
7.5.3.1 Thoroughly mix 9.0 g of activated carbon (Sec. 5.2.7} and 41.0 g
of Celite 545® to produce an 18% w/w mixture. Activate the mixture at 130°C for 6
hours, and store in a desiccator.
NOTE: Check each new batch of the carbon/Celite mixture by adding 50 mL of the
calibration verification solution to 950 mL of hexane. Take this solution
through the carbon column cleanup step, concentrate to 50 |jL and analyze.
If the recovery of any of the analytes is less than 80%, this batch of
carbon/Celite mixture may not be used.
7.5.3.2 Prepare a 4-inch long glass column by cutting off each end of a
10-mL disposable serological pipet. Fire polish both ends and flare if desired, insert
a glass wool plug at one end of the column, and pack it with 1 g of the carbon/Celite
mixture. Insert an additional glass wool plug in the other end.
CAUTION: It is very important that the column be packed properly to ensure that
carbon fines are not earned into the eluate. PCDDs/PCDFs will adhere
to the carbon fines and greatly reduce recovery. If carbon fines are
carried into the eluate, filter the eluate, using a 0.7-pm Alter (pre-rinsed
with toluene), then proceed to Sec. 7.5.3.6,
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7.5.3.3
Rinse the column with:
4 mL of toluene
2 mL of methylene chloride/methanol/toJuene (75:20:5 v/v)
4 mL of eydohexane/methylene chloride {50:50 v/v)
The flow rate should be less than 0.5 mUmin. Discard all the column rinsates.
7.5.3.4 While the column is still wet, transfer the concentrated eluate from
Sec. 7.5.26 to the prepared carbon column. Rinse the eluate container with two 0.5-
mL portions of hexane and transfer the rinses to the carbon column. Elute the column
with the following sequence of solvents.
10 mL of cyclohexane/methylene chloride (50:50 v/v).
5 mL of methylene chloride/methanol/toluene (75:20:5 v/v).
NOTE: The above two eluates may be collected arid combined, and used as a check
on column efficiency.
7.5.3.5 Once the solvents have eluted through the column, turn the
column over, and elute the PCDD/PCDF fraction with 20 mL of toluene, and collect the
eluate.
7.5.3.6 Concentrate the toluene fraction to about 1 mL on a rotary
evaporator by using a water bath at 50°C or with nitrogen evaporation. Carefully
transfer the concentrate into a 1-mL minivial and, again at elevated temperature
(50°C), reduce the volume to about 100 pL using a stream of nitrogen and a sand bath.
Rinse the rotary evaporator flask three times with 300 pL of a solution of 1 percent
toluene in methylene chloride, and add the rinses to the concentrate. Add 10 |jL of the
nonane recovery standard solution (Sec. 5.9) for soil, sediment, water, fish, paper pulp
and adipose tissue samples, or 50 mL of the recovery standard solution for sludge, still
bottom and fly ash samples. Store the sample at room temperature in the dark.
Chromatographic/mass spectrometric conditions and data acquisition parameters
7.6.1 Gas chromatograph operating conditions
Column coating: DB-5
Film thickness: 0.25 pm
Column dimension: 60-m x 0.32 mm
injector temperature: 270°C
Splitless valve time: 45 s
Interface temperature: Function of the final temperature
Temperature program
Initial temperature: 200°C
Initial hold time: 2 min
1 st temp, ramp: 5 °C/min to 220°C, hold for 16 minutes
2nd temp, ramp: 5 °C/min to 235°C, hold for 7 minutes
3rd temp, ramp: 5 °C/min to 330 °C, hold for 5 minutes
Total time: 60 min
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7.6.2 Mass spectrometer
7.6.2.1 The mass spectrometer must be operated in a selected ion
monitoring (SIM) mode with a total cycle time (including the voltage reset time) of one
second or less (Sec. 7.6.3.1). At a minimum, the ions listed in Table 6 for each of the
five SIM descriptors must be monitored. Note that with the exception of the last
descriptor (OCDD/OCDF), all descriptors contain 10 ions. The selection (Table 6) of
the molecular ions M and M+2 for "C-HxCDF and 1^C-HpCDF rather than M+2 and
M+4 (for consistency) was made to eliminate, even under high-resolution mass
spectrometric conditions, interferences occurring in these two ion channels for samples
containing high levels of native HxCDDs and HpCDDs. It is important to maintain the
same set of ions for both calibration and sample extract analyses. The selection of the
lock-mass ion is left to the performing laboratory.
NOTE: At the option of the analyst, the tetra- and pentachlorinated dioxins and
furans may be combined into a single descriptor.
7.6.2.2 The recommended mass spectrometer tuning conditions are
based on the groups of monitored ions shown in Table 6. By using a PFK molecular
leak, tune the installment to meet the minimum required resolving power of 10,000 (10
percent valley) at m/z 304.9824 (PFK) or any other reference signal close to m/z
303.9016 (from TCDF). By using peak matching conditions and the aforementioned
PFK reference peak, verify that the exact mass of m/z 380.9760 (PFK) is within 5 ppm
of the required value. Note that the selection of the low- and high-mass ions must be
such that they provide the largest voltage jump performed in any of the five mass
descriptors (Table 6).
7.6.3 Data acquisition
7.6.3.1 The total cycle time for data acquisition must be < 1 second. The
total cycle time includes the sum of all the dwell times and voltage reset times.
7.6.3.2 Acquire SIM data for all of the ions in the descriptors in Table 6.
7.7 Calibration
7.7.1 Initial calibration
Initial calibration is required before any samples are analyzed for PCDDs and PCDFs
and must meet the acceptance criteria in Sec. 7.7.2. Initial calibration is also required if any
routine calibration (Sec. 7.7.3) does not meet the required criteria listed in Sec. 7.7.2.
7.7.1.1 All five high-resolution concentration calibration solutions listed in
Table S must be used for the initial calibration.
7.7.1.2 Tune the instrument with PFK, as described in Sec. 7.6.2.2.
7.?.1.3 Inject 2 jjL of the GC column performance check solution
(Sec. 5.7) and acquire SIM mass spectral data as described earlier in Sec. 7.6.2. The
total cycle time must be < 1 second. The laboratory must not perform any further
analysis until it is demonstrated and documented that the criteria listed in Sec. 8.2.1
were met.
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7.7.1.4 By using the same GC (Sec. 7.6.1) and MS (Sec. 7.6.2) conditions
that produced acceptable results with the column performance check solution, analyze
a 2-jjL portion of each of the five concentration calibration solutions once with the
following mass spectrometer operating parameters.
7.7.1.4.1 The ratio of integrated ion current for the ions
appearing in Table 8 (homologous series quantitation ions) must be within the
indicated control limits (set for each homologous series) for all unlabeled
calibration standards in Table 5.
7.7.1.4.2 The ratio of integrated ion current for the ions
belonging to the carbon-labeled internal and recovery standards (Table 5)
must be within the control limits stipulated in Table 8.
NOTE: Sees. 7.7.1.4.1 and 7.7.1.4.2 require that 17 ion ratios from Sec.
7.7.1.4.1 and 11 ion ratios from Sec. 7.7.1.4.2 be within the
specified control limits simultaneously in one run. It is the
laboratory's responsibility to take corrective action if tine ion
abundance ratios are outside the limits.
7.7.1.4.3 For each selected ion current profile (SICP) and for
each GC signal corresponding to the elution of a target analyte and of its
labeled standards, the signal-to-noise ratio (S/N) must be better than or equal
to 10. Measurement of S/N is required for any GC peak that has an apparent
S/N of less than 5:1. The result of the calculation must appear on the SICP
above the GC peak in question.
7.7.1.4.4 Referring to Table 9, calculate the 17 relative
response factors (RF) for unlabeled target analytes fRF(n); n = 1 to 17]
relative to their appropriate internal standards (Table 5) and the nine RFs for
the ,3C12-labeled internal standards [RF(is); is = 18 to 26)] relative to the two
recovery standards (Table 5) according to the following formulae:
RF - * An2) * Qb
(A*1 + A5 x Qn
. <*sLlOjl5-
-------
Aj and = sum of the integrated ion abundances of the quantitation
ions (Tables 6 and 9} for the labeled recovery standards,
Qto = quantity of the internal standard injected (pg),
Qn = quantity of the recovery standard injected (pg), and
Q„ a quantity of the unlabeled PCDD/PCDF analyte injected (pg).
The RF„ and RF* are dimensionless quantities; the units used to express Qm,
Qm and Qn must be the same.
7.7.1.4.5 Calculate the values and Mr respective percent
relative standard deviations (%RSD) for the five calibration solutions:
RF. = £!
where n represents a particular PCDD/PCDF (2,3,7,8-substituted) congener
(n = 1 to 17; Table 9), and j is the injection number (or calibration solution
number; j = 1 to 5).
7.7.1.4.6 The relative response factors to be used for the
determination of the concentration of total isomers in a homologous series
(Table 9) are calculated as follows:
7.7.1.4.6.1 For congeners that belong to a
homologous series containing only one isomer (e.g., OCDD and
OCDF) or only one 2,3,7,8-substituted isomer (Table 4; TCDD,
PeCDD, HpCDD, and TCDF), the RF used will be the same as
the RF determined in Sec. 7.7.1.4.5.
NOTE: The calibration solutions do not contain 13C12-OCDF as
an internal standard. This is because a minimum
resolving power of 12,000 is required to resolve the
(M+6T ion of 13C12-OCDF from the [M+2]* ion of OCDD
(and [M+4F from 13Ci2-OCDF with [M] * of OCDD).
Therefore, the RF or OCDF is calculated relative to
13C12-OCDD.
7.7.1.4.6.2 For congeners that belong to a
homologous series containing more than one 2,3,7,8-substituted
isomer (Table 4), the RF used for those homologous series will
be the mean of the RFs calculated for all individual 2,3,7,8-
substituted congeners using the equation below:
RF„ . *2
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where:
k = 27 to 30 (Table 9), with 27 = PeCDF; 28 = HxCDF; 29 =
HxCDD; and 30 = HpCDF,
t = total number of 2,3,7,8-substituted isomers present in the
calibration solutions (Table 5) for each homologous series
(e.g., two for PeCDF, four for HxCDF, three for HxCDD,
two for HpCDF),
NOTE: Presumably, the HRGC/HRMS response factors of
different isomers within a homologous series are
dffferent However, this analytical protocol will make the
assumption that the HRGC/HRMS responses of all
isomers in a homologous series that do not have the
2,3,7,8-substitution pattern are the same as the
responses of one or more of the 2,3,7,8-substituted
isomer(s) in that homologous series.
7.7.1.4.7 Relative response factors (RF„) to be used for the
determination of the percent recoveries for the nine internal standards are
calculated as follows:
Au"*CL
RF_ = — -5
ITi
ERF*
RF" = J~5 '
where:
m = 18 to 26 (congener type) and j = 1 to 5 (injection number),
A*m = sum of the integrated ion abundances of the quantitation ions
(Tables 6 and 9) for a given internal standard (m = 18 to 26),
- sum of the integrated ion abundances of the quantitation ions
(Tables 6 arid 9) for the appropriate recovery standard (see
Table 5, footnotes),
Qm, Qtem = quantities of, respectively, the recovery standard (rs) and a
particular internal standard (is = m) injected (pg),
RFm = relative response factor of a particular internal standard (m)
relative to an appropriate recovery standard, as determined from
one injection, and
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RFm = calculated mean relative response factor of a particular internal
standard (m) relative to an appropriate recovery standard, as
determined from the five initial calibration injections (j).
7.7.2 Criteria for acceptable calibration - The criteria listed below for acceptable
calibration must be met before sample analyses are performed.
7.7.2.1 The percent relative standard deviations for the mean response
factors (RFn andKFj from the 17 unlabeled standards must not exceed ± 20 percent,
and those for the nine labeled reference compounds must not exceed *: 30 percent.
7.7.2.2 The S/N for the GC signals present in every SICP (including the
ones for the labeled standards) must be > 10.
7.7.2.3 The ion abundance ratios (Table 8} must be within the specified
control limits.
NOTE: if the criterion for acceptable calibration listed in Sec. 7.7.2.1 is met, the
analyte-specific RF can then be considered independent of the analyte
quantity for the calibration concentration range. The mean RFs will be used
for all calculations until the routine calibration criteria (Sec. 7.7.4) are no
longer met. At such time, new RF values will be calculated from a new set
of injections of the calibration solutions.
7.7.3 Routine calibration (continuing calibration check) - Routine calibrations must
be performed at the beginning of a 12-hour period, after successful mass resolution and GC
resolution performance checks. A routine calibration is also required at the end of a 12-hour
shift. Inject 2 mL of the concentration calibration solution HRCC-3 standard (Table 5). By using
the same HRGC/HRMS conditions as used in Sees. 7.6.1 and 7.6.2, determine and document
an acceptable calibration as provided in Sec. 7.7.4.
7.7.4 Criteria for acceptable routine calibration - The following criteria must be met
before further analysis is performed.
7.7.4.1 The measured RFs [RFn for the unlabeled standards] obtained
during the routine calibration runs must be within ± 20 percent of the mean values
established during the initial calibration (Sec. 7.7.1.4.5).
7.7.4.2 The measured RFs [RFm for the labeled standards] obtained
during the routine calibration runs must be within ± 30 percent of the mean values
established during the initial calibration (Sec. 7.7.1.4.7).
7.7.4.3 The ion abundance ratios (Table 8) must be within the allowed
control limits.
7.7.4.4 If either one of the criteria in Sees. 7.7.4.1 and 7.7.4.2 is not
satisfied, repeat one more time. If these criteria are still not satisfied, the entire routine
calibration process (Sec. 7.7.1) must be reviewed. If the ion abundance ratio criterion
(Sec. 7.7.4.3) is not satisfied, refer to the note in Sec. 7.7.1.4.2 for resolution.
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NOTE; An initial calibration must be earned out whenever the HRCC-3, the sample
fortification, or the recovery standard solution is replaced by a new solution
from a different lot.
7.8 Analysis
7.8.1 Remove the sample or blank extract (from Sec. 7.5.3.6) from storage. With
a stream of dry, purified nitrogen, reduce the extract volume to 10 pL to 50 pL.
NOTE: A final volume of 20 pL or more should be used whenever possible. A 10-pL final
volume is difficult to handle, and injection of 2 pL out of 10 pL leaves little sample for
confirmations and repeat injections, and for archiving.
7.8.2 Inject a 2-pL aliquot of the extract into the GC, operated under the conditions
that have been established to produce acceptable results with the performance check solution
(Sees. 7.6.1 and 7.6.2).
7.8.3 Acquire SIM data according to Sees. 7.6.2 and 7.6.3. Use the same acquisi-
tion and mass spectrometer operating conditions previously used to determine the relative
response factors (Sees. 7.7.1.4.4 through 7.7.1.4.7). Ions characteristic of polychlorinated
diphenyl ethers are included in the descriptors listed in Table 6.
NOTE: The acquisition period must at least encompass the PCDD/PCDF overall retention
time window previously determined (Sec. 8.2.1.3). Selected ion current profiles
(SIGP) for the lock-mass ions (one per mass descriptor) must also be recorded and
included in the data package. These SICPs must be true representations of the
evolution of the lock-mass ions amplitudes during the HRGC/HRMS run (see Sec.
8.2.2 for the proper level of reference compound to be metered Wo the ion
chamber.) The analyst may be required to monitor a PFK ion, not as a lock-mass,
but as a regular ion, in order to meet this requirement. It is recommended to
examine the lock-mass ion SICP for obvious basic sensitivity and stability changes
of the instrument during the GC/MS run that could affect the measurements.
7.8.4 Identification criteria - For a gas chromatographic peak to be identified as a
PCDD or PCDF, it must meet all of the following criteria;
7.8.4.1 Retention times
7.8.4.1.1 For 2,3,7,8-substituted congeners, which have an
isotopically-labeled internal or recovery standard present in the sample
extract (this represents a total of 10 congeners including OCDD; Tables 2
and 3), the retention time (RRT; at maximum peak height) of the sample
components (i.e., the two ions used for quantitation purposes listed in Table
6) must be within -1 to +3 seconds of the isotopically-labeled standard.
7.8.4.1.2 For 2,3,7,8-substituted compounds that do not have
an isotopically-labeled internal standard present in the sample extract (this
represents a total of six congeners; Table 3), the retention time must fall
within 0.005 retention time units of the relative retention times measured in
the routine calibration. Identification of OCDF is based on its retention time
relative to 13C12-OCDD as determined from the daily routine calibration
results.
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7.8.4.1.3 For non-2,3,7,8-substituted compounds (tetra
through octa; totaling 119 congeners), the retention time must be within the
corresponding homologous retention time windows established by analyzing
the column performance check solution (Sec. 8.1.3).
7.8.4.1.4 The ion current responses for both ions used for
quantitative purposes (e.g., for TCDDs: m/z 319.8965 and 321.8936) must
reach maximum simultaneously (± 2 seconds).
7.8.4.1.5 The ion current responses for both ions used for the
labeled standards (e.g., for 13C12-TCDD: m/z 331.9368 and m/z 333.9339)
must reach maximum simultaneously (± 2 seconds).
NOTE: The analyst is required to verity the presence of 1,2,8,9-TCDD and
1,3,4,6,8-PeCDF (Sec. 8.1.3) in the SICPs of the daily performance
checks. Should either one compound be missing, the analyst is
required to take corrective action as it may indicate a potential
problem with the ability to detect all the PCDDs/PCDFs.
7.8.4.2 Ion abundance ratios
The integrated ion currents for the two ions used for quantitation
purposes must have a ratio between the lower and upper limits established
for the homologous series to which the peak is assigned. See Sees. 7.7.1.4.1
and 7.7.1.4.2 and Table 8 for details.
7.8.4.3 Signal-to-noise ratio
All ion current intensities must be > 2.5 times noise level for posi-
tive identification of an unlabeled PCDD/PCDF compound or a group of
coeluting isomers. Figure 6 describes the procedure to be followed for the
determination of the S/N. Labeled analytes must have a S/N > 10.
7.8.4.4 Polychlorinated diphenyl ether interferences
In addition to the above criteria, the identification of a GC peak as
a PCDF can only be made if no signal having a S/N > 2.5 is detected at the
same retention time (± 2 seconds) in the corresponding polychlorinated
diphenyl ether (PCDPE, Table 6) channel.
7.9 Calculations
7.9.1 For gas chromatographic peaks that have met the criteria outlined in Sec.
7.8.4, calculate the concentration of the PCDD or PCDF compounds using the formula:
c K * Q«
X Ah * W * RFn
where:
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C„ = concentration of unlabeled PCDD/PCDF congeners (or group of coeluting isomer
within an homologous series) in pg/g,
A* = sum of the integrated ion abundances of the quantitation ions (Table 6) for unlabeled
PCDDs/PCDFs,
A* = sum of the integrated ton abundances of the quantitation ions (Table 6) for the labeled
internal standards,
Qto = quantity, in pg, of the internal standard added to the sample before extraction,
W = weight, in g, of the sample (solid or organic liquid), or volume in mL of an aqueous
sample, and
RF„ = calculated mean relative response factor for the analyte (RFn with n = 1 to 17; Sec.
7.7.1.4.5).
If the analyte is identified as one of the 2,3,7,8-subst'rtuted PGDDs or PCDFs, KF„ Is
the value calculated using the equation in Sec. 7.7.1.4.5. However, if it is a non-2,3,7,8-
substituted congener, the RF(k) value is the one calculated using the equation in
Sec. 7.7.1.4.6.2. (RFk, for k = 27 to 30).
7.9.2 Calculate the percent recovery of the nine internal standards measured in the
sample extract, using the formula:
A., * Q
percent recovery = — * 100
Ob * x RFb
where;
A* = sum of the integrated ion abundances of the quantitation ions (Table 6) for the
labeled internal standard,
A,s = sum of the integrated ion abundances of the quantitation ions (Table 6) for the
labeled recovery standard; the selection of the recovery standard depends on the
type of congeners (see Table 5, footnotes),
Qs = quantity, in pg, of the internal standard added to the sample before extraction,
Qre = quantity, in pg, of the recovery standard added to the cleaned-up sample residue
before HRGC/HRMS analysis, and
RF m = calculated mean relative response factor for the labeled internal standard relative to
the appropriate (see Table 5, footnotes) recovery standard. This represents the
mean obtained in Sec. 7.7.1.4.7 (RF m with m - 18 to 26).
NOTE: For human adipose tissue, adjust the percent recoveries by adding 1 percent to the
calculated value to compensate for the 1 percent of the extract diverted for the lipid
determination.
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7.9.3 If the concentration in the final extract of any of the fifteen 2,3,7,8-substituted
PCDD/PCDF compounds (Table 3) exceeds the upper method calibration limits (MCL) listed
in Table 1 (e.g., 200 pg/pL for TCDD in soil), the linear range of response versus concentration
may have been exceeded, and a second analysis of the sample (using a one-tenth aliquot)
should be undertaken. The volumes of the internal and recovery standard solutions should
remain the same as described for the sample preparation (Sees. 7.1 to 7.9.3).
if a smaller sample size would not be representative of the entire sample, one of the
following options is recommended:
(1) Re-extract an additional aliquot of sufficient size to insure that it is representative
of the entire sample. Spike it with a higher concentration of internal standard.
Prior to GC/MS analysis, dilute the sample so that it has a concentration of
internal standard equivalent to that present in the calibration standard. Then,
analyze the diluted extract.
(2) Re-extract an additional aliquot of sufficient size to insure that it is representative
of the entire sample. Spike it with a higher concentration of internal standard.
Immediately following extraction, transfer the sample to a volumetric flask and
dilute to known volume. Remove an appropriate aliquot and proceed with cleanup
and analysis.
(3) Use the original analysis data to quantitate the internal standard recoveries.
Respike the original extract (note that no additional cleanup is necessary) with 100
times the usual quantity of internal standards. Mute the re-spiked extract by a
factor of 100. Reanalyze the diluted sample using the internal standard recoveries
calculated from the initial analysis to correct the results for losses during isolation
and cleanup.
7.9.4 The total concentration for each homologous series of PCDD and PCDF is
calculated by summing up the concentrations of all positively identified isomers of each
homologous series. Therefore, the total should also include the 2,3,7,8-substituted congeners.
The total number of GC signals included in the homologous total concentration value must be
specified in the report. If an isomer is not detected, use zero (0) in this calculation.
7.9.5 Sample specific estimated detection limit - The sample specific estimated
detection limit (EDI) is the concentration of a given analyte required to produce a signal with
a peak height of at least 2.5 times the background signal level. An EDL is calculated for each
2,3,7,8-substituted congener that is not identified, regardless of whether or not other non-
2,3,7,8-substHuted isomers are present. Two methods of calculation can be used, as follows,
depending on the type of response produced during the analysis of a particular sample.
7.9.5.1 Samples giving a response for both quantitation ions (Tables 6
and 9) that is less than 2.5 times the background level.
Use the expression below to calculate an EDL for each 2,3,7,8-substituted
PCDD/PCDF that does not have a response with S/N ;> 2.5). The background level is
determined by measuring the range of the noise (peak to peak) for the two quantitation
ions (Table 6) of a particular 2,3,7,8-substituted isomer within an homologous series,
in the region of the SICP trace corresponding to the elution of the internal standard (if
the congener possesses an internal standard) or in the region of the SICP where the
congener is expected to elute by comparison with the routine calibration data (for those
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congeners that do not have a 13C-labeled standard), multiplying that noise height by
2.5, and relating the product to an estimated concentration that would produce that
peak height. Use the formula:
EDL = S—
H,, * W x RFn
where:
EDL « estimated detection limit for homologous 2,3,7,8-substituted PCDDs/PCDFs.
= sum ofthe height of the noise level for each quantitation ion (Table 6) for the
unlabeled PCDDs/PCDFs, measured as shown in Figure 6.
H„ = sum of the height of the noise level for each quantitation ion (Table 6) for the
labeled internal standard, measured as shown in Figure 6.
W, RFm and Q* retain the same meanings as defined in Sec. 7.9.1.
7.9.5.2 Estimated maximum possible concentration - An estimated
maximum possible concentration (EMPC) is calculated for 2,3,7,8-substituted isomers
that are characterized by a response with an S/N of at least 2.5 for both the
quantitation ions, and meet all ofthe identification criteria in Sec. 7.8.4 except the ion
abundance ratio criteria or when a peak representing a PCDPE has been detected.
An EMPC is a worst-case estimate of the concentration. Calculate the EMPC
according to the expression shown in Sec. 7.9.1.
7.9.6 The relative percent difference (RPD) of any duplicate sample results are
calculated as follows:
RPD = * " 82 * x 100
si + S2
2
where Si and Sj represent sample and duplicate sample results.
7.9.7 The 2,3,7,8-TCDD toxicity equivalents (TE) of PCDDs and PCDFs present in
the sample are calculated, if requested by the data user, according to the method
recommended by the Chlorinated Dioxins Workgroup (CDWG) of the EPA and the Center for
Disease Control (CDC). This method assigns a 2,3,7,8-TCDD toxicity equivalency factor (TEF)
to each of the fifteen 2,3,7,8-substituted PCDDs and PCDFs (Table 3) and to OCDD and
OCDF, as shown in Table 10. The 2,3,7,8-TCDD equivalent of the PCDDs and PCDFs present
in the sample is calculated by summing the TEF times their concentration for each of the
compounds or groups of compounds fisted in Table 10. The exclusion of other homologous
series such as mono-, di-, and tri- chlorinated dibenzodioxins and dibenzofurans does not
mean that they are non-toxic. However, their toxicity, as known at this time, is much lower
than the toxicity ofthe compounds listed in Table 10. The above procedure for calculating the
2,3,7,8-TCDD toxicity equivalents is not claimed by the CDWG to be based on a thoroughly
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established scientific foundation. The procedure, rather, represents a "consensus recom-
mendation on science policy." Since the procedure may be changed in the future, reporting
requirements for PCDD and PCDF data would still include the reporting of the analyte
concentrations of the PCDD/PCDF congener as calculated in Sees. 7.9.1 and 7.9.4.
7.9.8 Two GC column TEF determination
7.9.8.1 The concentration of 2,3,7,8-TCDD (see note below), is calculated
from the analysis of the sample extract on the 60-m DB-5 (or equivalent) fused-siiica
capillary column. The experimental conditions remain the same as the conditions
described previously in Sec. 7.8, and the calculations are performed as outlined in Sec.
7.9. The chromatographic separation between the 2,3,7,8-TCDD and its close eiuters
(1,2,3,7/1,2,3,8-TCDD and 1,2,3,9-TCDD) must be equal or less than 25 percent valley.
7.9.8.2 The concentration of the 2,3,7,8-TCDF is obtained from the
analysis of the sample extract on the 30-m DB-225 (or equivalent) fused-silica capillary
column. However, the GC/MS conditions must be altered so that: (1) only the first
three descriptors (i.e., tetra-, penta-, and hexachlorinated congeners) of Table 6 are
used; and (2) the switching time between descriptor 2 (pentachlorinated congeners)
and descriptor 3 (hexachlorinated congeners) takes place following the elution of 13C12-
1,2,3,7,8-PeCDD. Hie concentration calculations are performed as outlined in Sec. 7.9.
The chromatographic separation between the 2,3,7,8-TCDF and its close eiuters
(2,3,4,7-TCDF and 1,2,3,9-TCDF) must be equal or less than 25 percent valley.
NOTE: The confirmation and quantitation of 2,3,7,8-TCDD (Sec. 7.9.7.1.1) may be
accomplished on the SP-2330 GC column instead of the DB-5 column,
provided the criteria listed in Sec. 8.2.1 are met and the requirements
described in Sec. 8.3.2 am followed.
7.9.8.3 For a gas chromatographic peak to be identified as a 2,3,7,8-
substituted PCDD/PCDF congener, it must meet the ion abundance and signal-to-noise
ratio criteria listed in Sees. 7.8.4.2 and 7.8.4.3, respectively. In addition, the retention
time identification criterion described in Sec. 7.8.4.1.1 applies here for congeners for
which a cartoon-labeled analogue is available in the sample extract. However, the
relative retention time (RRT) of the 2,3,7,8-substituted congeners for which no cartoon-
labeled analogues are available must fall within 0.006 units of the carbon-labeled
standard RRT. Experimentally, this is accomplished by using the attributions described
in Table 11 and the results from the routine calibration run on the SP-2330 column.
8,0 QUALITY CONTROL
8.1 Refer to Chapter One for specific quality control (QC) procedures. Quality control to
validate sample extraction is covered in Method 3500. If extract cleanup was performed, follow the
QC in Method 3600 and in the specific cleanup method.
8.2 System performance criteria - System performance criteria are presented below. The
laboratory may use the recommended GC column described in Sec. 4.2. ft must be documented that
all applicable system performance criteria (specified in Sees. 8.2.1 and 8.2.2) were met before
analysis of any sample is performed. Sec. 7.6.1 provides recommended GC conditions that can be
used to satisfy the required criteria. Figure 3 provides a typical 12-hour analysis sequence, whereby
the response factors and mass spectrometer resolving power checks must be performed at the
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beginning and the end of each 12-hour period of operation. A GC column performance check is only
required at the beginning of each 12-hour period during which samples are analyzed. An
HRGC/HRMS method blank run is required between a calibration run and the first sample run. The
same method blank extract may thus be analyzed more than once if the number of samples within
a batch requires more than 12 hours of analyses.
8.2.1 GC column performance
8.2.1.1 Inject 2 |jL (Sec. 4.1.1) of the column performance check solution
(Sec. 5.7) and acquire selected ion monitoring (SIM) data as described in Sec. 7.6.2
within a total cycle time of« 1 second (Sec. 7.6.3.1).
8.2.1.2 The chromatographic separation between 2,3,7,8-TCDD and the
peaks representing any either unlabeled TCDD isomers must be resolved with a valley
of <25 percent (Figure 4), where:
Valley percent = (x/y) x (100)
x = measured as in Figure 4 from the 2,3,7,8-closest TCDD eluting isomer
y s the peak height of 2,3,7,8-TCDD
It is the responsibility of the laboratory to verily the conditions suitable for the
appropriate resolution of 2,3,7,8-TCDD from all other TCDD isomers. The GC column
performance check solution also contains the known first and last PCDD/PCDF eluters
under the conditions specified in this protocol. Their retention times are used to
determine the eight homologue retention time windows that are used for qualitative
(Sec. 7.8.4.1) and quantitative purposes. All peaks (that includes 13C12-2,3,7,8-TCDD)
should be labeled and identified on the chromatograms. Furthermore, all first eluters
of a homologous series should be labeled with the tetter F, and all last eluters of a
homologous series should be labeled with the letter L (Figure 4 shows an example of
peak labeling for TCDD isomers). Any individual selected ion current profile (SICP) (for
the tetras, this would be the SICP for m/z 322 and m/z 304) or the reconstructed
homologue ion current (for the tetras, this would correspond to m/z 320 + m/z 322 +
m/z 304 + m/z 306) constitutes an acceptable form of data presentation. An SICP for
the labeled compounds (e.g., m/z 334 for labeled TCDD) is also required.
8.2.1.3 The retention times for the switching of SIM ions characteristic of
one homologous series to the next higher homologous series must be indicated in the
SICP. Accurate switching at the appropriate times is absolutely necessary for accurate
monitoring of these compounds. Allowable tolerance on the daily verification with the
GC performance check solution should be better than 10 seconds for the absolute
retention times of all the components of the mixture. Particular caution should be
exercised for the switching time between the last tetrachlorinated congener (i.e.,
1,2,8,9-TCDD) and the first pentachlorinated congener (i.e., 1,3,4,6,8-PeCDF), as
these two compounds elute within 15 seconds of each other on the 60-m DB-5 column.
A laboratory with a GC/MS system that is not capable of detecting both congeners
(1,2,8,9-TCDD and 1,3,4,6,8-PeCDF) within one analysis must take corrective action.
If the recommended column is not used, then the first-and last-eluting isomer of each
homologue must be determined experimentally on the column which is used, and the
appropriate isomers must then be used for window definition and switching times.
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8.2.2 Mass spectrometer performance
8.2.2.1 The mass spectrometer must be operated in the electron
ionization mode. A static resolving power of at least 10,000 (10 percent valley
definition) must be demonstrated at appropriate masses before any analysis is
performed (Sec. 7.8). Static resolving power checks must be performed at the
beginning and at the end of each 12-hour period of operation. However, it is
recommended that a check of the static resolution be made and documented before
and after each analysis. Corrective action must be implemented whenever the
resolving power does not meet the requirement.
8.2.2.2 Chromatography time for PCDDs and PCDFs exceeds the long
term mass stability of the mass spectrometer. Because the instrument is operated in
the high-resolution mode, mass drifts of a few ppm (e.g., 5 ppm in mass) can have
serious adverse effects on instrument performance. Therefore, a mass drift correction
is mandatory. To that effect, it is recommended to select a lock-mass ton from the
reference compound (PFK is recommended) used for tuning the mass spectrometer.
The selection of the lock-mass ion is dependent on the masses of the ions monitored
within each descriptor. Table 6 offers some suggestions for the lock-mass ions.
However, an acceptable lock-mass ion at any mass between the lightest and heaviest
ion in each descriptor can be used to monitor and correct mass drifts. The level of the
reference compound (PFK) metered into the ion chamber during HRGC/HRMS
analyses should be adjusted so that the amplitude of the most intense selected lock-
mass ion signal (regardless of the descriptor number) does not exceed 10 percent of
the full scale deflection for a given set of detector parameters. Under those conditions,
sensitivity changes that might occur during the analysis can be more effectively
monitored.
NOTE: Excessive PFK (or any other reference substance) may cause noise
problems and contamination of the ion source resulting in an increase in
downtime for source cleaning.
8.2.2.3 Documentation of the instrument resolving power must then be
accomplished by recording the peak profile of the high-mass reference signal (m/z
380.9760) obtained during the above peak matching experiment by using the low-mass
PFK ion at m/z 304.9824 as a reference. The minimum resolving power of 10,000
must be demonstrated on the high-mass ion while it is transmitted at a lower
accelerating voltage than the low-mass reference ion, which is transmitted at full
sensitivity. The format of the peak profile representation (Figure 5) must allow manual
determination of the resolution, i.e., the horizontal axis must be a calibrated mass scale
(amu or ppm per division). The result of the peak width measurement (performed at
5 percent of the maximum, which corresponds to the 10 percent valley definition) must
appear on the hard copy and cannot exceed 100 ppm at m/z 380.9760 (or 0.038 amu
at that particular mass).
8.3 Quality control samples
8.3.1 Performance evaluation samples - When available, performance evaluation
(PE) samples containing known amounts of unlabeled 2,3,7,8-substituted PCDDs/PCDFs or
other PCDD/PCDF congeners should be analyzed alongside routine field samples.
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8.3.2
Performance check solutions
8.3.2.1 At the beginning of each 12-hour period during which samples are
to be analyzed, an aliquot of the 1) GC column performance check solution and 2)
high-resolution concentration calibration solution No. 3 (HRCC-3; see Table 5) shall be
analyzed to demonstrate adequate GC resolution and sensitivity, response factor repro-
ducibility, and mass range calibration, and to establish the PCDD/PCDF retention time
windows. A mass resolution check shall also be performed to demonstrate adequate
mass resolution using an appropriate reference compound (PFK is recommended).
If the required criteria are not met, remedial action must be taken before any samples
are analyzed.
8.3.2.2 To validate positive sample data, the routine or continuing
calibration (HRCC-3; Table 5) and the mass resolution check must be performed also
at the end of each 12-hour period during which samples are analyzed. Furthermore,
an HRGC/HRMS method blank run must be recorded following a calibration run and
the first sample run.
8.3.2.2.1 If the laboratory operates only during one period
(shift) each day of 12 hours or less, the GC performance check solution must
be analyzed only once (at the beginning of the period) to validate the data
acquired during the period. However, the mass resolution and continuing
calibration checks must be performed at the beginning as well as at the end
of the period.
8.3.2.2.2 If the laboratory operates during consecutive 12-hour
periods (shifts), analysis of the GC performance check solution must be
performed at the beginning of each 12-hour period. The mass resolution and
continuing calibration checks from the previous period can be used for the
beginning of the next period.
8.3.2.3 Results of at least one analysis of the GC column performance
check solution and of two mass resolution and continuing calibration checks must be
reported with the sample data collected during a 12-hour period.
8.3.2.4 Deviations from criteria specified for the GC performance check
or for the mass resolution check invalidate all positive sample data collected between
analyses of the performance check solution, and the extracts from those positive
samples shall be reanalyzed.
If the routine calibration run fails at the beginning of a 12-hour shift, the
instructions in Sec. 7.7.4.4 must be followed. If the continuing calibration check
performed at the end of a 12 hour period fails by no more than 25 percent RPD for the
17 unlabeled compounds and 35 percent RPD for the 9 labeled reference compounds,
use the mean to the two "daily" RF values from the two daily routine calibration runs
to compute the analyte concentrations, instead of the RF values obtained from the
initial calibration. A new initial calibration (new RFs) is required immediately (within two
hours) following the analysis of the samples, whenever the RPD from the end-of-shift
routine calibration exceeds 25 percent or 35 percent, respectively. Failure to perform
a new initial calibration immediately following the analysis of the samples will
automatically require reanalysis of all positive sample extracts analyzed before the
failed end-of-shift continuing calibration check.
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8.3.3 The GC column performance check mixture, high-resolution concentra-
tion calibration solutions, and the sample fortification solutions may be obtained from
commercial sources.
8.3.4 Field blanks - Batches of field samples may contain a field blank sample of
uncontaminated soil, sediment or water that is to be fortified before analysis according to
Sec. 8.3.4.1. in addition to this field blank, a batch of samples may include a rinsate, which
is a portion of the solvent that was used to rinse sampling equipment. The rinsate is analyzed
to assure that the samples were not contaminated by the sampling equipment.
8.3.4.1 Fortified field blank
8.3.4.1.1 Weigh a 10-g portion or use 1-L (for aqueous
samples) of the specified field blank sample and add 100 mL of the solution
containing the nine internal standards (Table 2} diluted with 1.0 mL acetone
(Sec. 7.1).
8.3.4.1.2 Extract by using the procedures beginning in Sees.
7.4.5 or 7.4.6, as applicable, add 10 jiL of the recovery standard solution
(Sec. 7.5.3.6) and analyze a 2-|jL aliquot of the concentrated extract.
8.3.4.1.3 Calculate the concentration (Sec. 7.9.1) of 2,3,7,8-
substituted PCDDs/PCDFs and the percent recovery of the internal standards
(Sec. 7.9.2).
8.3.4.1.4 Extract and analyze a new simulated fortified field
blank whenever new lots of solvents or reagents are used for sample
extraction or for column chromatographic procedures.
8.3.4.2 Rinsate sample
8.3.4.2.1 Take a 100-mL (± 0.5 mL) portion of the sampling
equipment rinse solvent (rinsate sample), filter, if necessary, and add 100 mL
of the solution containing the nine internal standards (Table 2).
8.3.4.2.2 Using a K-D apparatus, concentrate to about 5 mL.
NOTE: As an option, a rotary evaporator may be used in place of the K-D
apparatus for the concentration of the rinsate.
8.3.4.2.3 Transfer the 5 mL concentrate from the K-D
concentrator tube in 1-mL portions to a 1-mL minivial, reducing the volume
in the minivial as necessary with a gentle stream of dry nitrogen.
8.3.4.2.4 Rinse the K-D concentrator tube with two 0.5 mL
portions of hexane and transfer the rinses to the 1 mL minivial. Concentrate
with dry nitrogen, as necessary.
8.3.4.2.5 Just before analysis, add 10 jjL recovery standard
solution (Table 2) and reduce the volume to its final volume, as necessary
(Sec. 7.8.1). No column chromatography is required.
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8.3.4.2.6 Analyze an aliquot of the solution following the same
procedures used to analyze samples.
8.3.4.2.7 Report percent recovery of the internal standard and
the presence of any PCDD/PCDF compounds in (jg/L of rinsate solvent.
8.3.5 Duplicate analyses
In each batch of samples, locate the sample specified for duplicate analysis,
and analyze a second 10-g soil or sediment sample portion or 1-L water sample, or an
appropriate amount of the type of matrix under consideration.
8.3.5.1 The results of the laboratory duplicates (percent recovery and
concentrations of 2,3,7,8-substituted PCDD/PCDF compounds) should agree within
25 percent relative difference (difference expressed as percentage of the mean).
Report all results.
8.3.5.2 Recommended actions to help locate problems
Verify satisfactory instrument performance (Sees. 8.2 and 8.3).
If possible, verily that no error was made while weighing the sample portions.
Review the analytical procedures with the performing laboratory personnel.
8.3.6 Matrix spike and matrix spike duplicate
8.3.6. 1 Locate the sample for the MS and MSD analyses (the sample may
be labeled "double volume").
8.3.6.2 Add an appropriate volume of the matrix spike fortification solution
(Sec. 5.10) and of the sample fortification solution (Sec. 5.8), adjusting the fortification
level as specified in Table 1 under IS Spiking Levels.
8.3.6.3 Analyze the MS and MSD samples as described in Sec. 7.
8.3.6.4 The results obtained from the MS and MSD samples
(concentrations of 2,3,7,8-substituted PCDDs/PCDFs) should agree within 20 percent
relative difference.
8.4 Percent recovery of the internal standards - For each sample, method blank and
rinsate, calculate the percent recovery (Sec. 7.9.2). The percent recovery should be between
40 percent and 135 percent for all 2,3,7,8-substituted internal standards.
NOTE: A low or high percent recovery for a blank does not require discarding the analytical data
but it may indicate a potential problem with future analytical data.
8.5 Identification criteria
8.5.1 If either one of the identification criteria appearing in Sees. 7.8.4.1.1 through
7.8.4.1.4 is not met for an homologous series, it is reported that the sample does not contain
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unlabeled 2,3,7,8-substituted PCDD/PCDF isomers for that homologous series at the
calculated detection limit (Sec. 7.9.5)
8.5.2 If the first initial identification criteria (Sees. 7.8.4.1.1 through 7.8.4.1.4) are
met, but the criteria appearing in Sees. 7.8.4.1.5 and 7.8,4.2 are not met, that sample is
presumed to contain interfering contaminants. This must be noted on the analytical report
form, and the sample should be rerun or the extract reanalyzed.
8.6 Unused portions of samples and sample extracts should be preserved for six months
after sample receipt to allow further analyses.
8.7 Reuse of glassware is to be minimized to avoid the risk of contamination.
9.0 METHOD PERFORMANCE
9.1 Table 12 provides data from a comparison of Soxhlet extraction and pressurized fluid
extraction of samples of ground chimney brick. The data are taken from Reference 8.
9.2 Table 13 provides data from a comparison of Soxhlet extraction and pressurized fluid
extraction of samples of urban dust. The data are taken from Reference 8.
9.3 Table 14 provides data from a comparison of Soxhlet extraction and pressurized fluid
extraction of samples of fly ash. PFE data are provided for samples that were pretreated with an
HC1 wash and for samples that were not pretreated, but were extracted with a mixture of toluene and
acetic acid. The data are taken from Reference 8.
9.4 Table 15 provides data from a comparison of Soxhlet extraction and pressurized fluid
extraction of a soil sample (EC-2) from the National Water Research Institute (Burlington, Ontario,
Canada) that contains high levels of PCDDs and PCDFs. The data are taken from Reference 8.
9.5 Table 16 provides data from a comparison of Soxhlet extraction and pressurized fluid
extraction of a sediment sample (HS-2) from the National Research Council institute for Marine
Biosciences (Halifax, Nova Scotia, Canada) that contains iow levels of PCDDs and PCDFs. The
data are taken from Reference 8.
9.6 Table 17 provides data from a comparison of Soxhlet extraction and pressurized fluid
extraction for two field-contaminated sediment samples. The data are taken from Reference 8.
10.0 REFERENCES
1. "Control of Interferences in the Analysis of Human Adipose Tissue for 2,3,7,8-
Tetrachlorodibenzo-p-dioxin". D. G. Patterson, J.S. Holler, D.F. Grote, L.R. Alexander, C.R.
Lapeza, R.C. O'Connor and J.A. Liddle. Environ. Toxicol. Chem. 5, 355-360 (1986).
2. "Method 8290; Analytical Procedures and Quality Assurance for Multimedia Analysis of
Polychlorinated Dibenzop-Dioxins and Dibenzofurans by High-Resolution Gas
Chromatography/High-Resolution Mass Spectrometry". Y. Tondeur and W.F. Beckert. U.S.
Environmental Protection Agency, Environmental Monitoring Systems Laboratory, Las Vegas,
NV.
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3. "Carcinogens - Working with Carcinogens", Department of Health, Education, and Welfare,
Public Health Service, Center for Disease Control. National Institute for Occupational Safety
and Health. Publication No. 77-206, August 1977.
4. "OSHA Safety and Health Standards, General Industry", (29 CFR 1910), Occupational Safety
and Health Administration, OSHA 2206 (revised January 1976),
5. "Safety in Academic Chemistry Laboratories", American Chemical Society Publication,
Committee on Chemical Safety (3rd Edition, 1979.)
6. "Hybrid HRGC/MS/MS Method for the Characterization of Tetrachlorinated Dibenzo-p-diroans
in Environmental Samples." Y. Tondeur, W.J. Niedertiut, S.R. Missler, and J.E. Campana,
Mass Spectrom. 14, 449-456 (1987).
7. USEPA National Dioxin Study - Phase II, "Analytical Procedures and Quality Assurance Plan
for the Determination of PCDD/PCDF in Fish", EPA-Duluth, October 26,1987.
8. "Extraction of Polychlorinated Dibenzo-p-Dioxi ns and Polychlorinated Dibenzofurans from
Environmental Samples Using Accelerated Solvent Extraction (ASE)," B. E. Richter, J. L.
Ezzell, D. E. Knowles, and F. Hoefler, Chemosphem, 34 (5-7), 975-987,1997.
11.0 SAFETY
11.1 The following safety practices are excerpts from EPA Method 613, Sec. 4 (July 1982
version) and amended for use in conjunction with this method. The 2,3,7,8-TCDD isomer has been
found to be acnegenic, carcinogenic, and teratogenic in laboratory animal studies. Other PCDDs
and PCDFs containing chlorine atoms in positions 2,3,7,8 are known to have toxicities comparable
to that of 2,3,7,8-TCDD. The analyst should note that finely divided dry soils contaminated with
PCDDs and PCDFs are particularly hazardous because of the potential for inhalation and ingestion.
It is recommended that such samples be processed in a confined environment, such as a hood or
a glove box. Laboratory personnel handling these types of samples should wear masks fitted with
charcoal filters to prevent inhalation of dust.
11.2 The toxicity or carcinogenicity of each reagent used in this method is not precisely
defined; however, each chemical compound should be treated as a potential health hazard. From
this viewpoint, exposure to these chemicals must be kept to a minimum. The laboratory is
responsible for maintaining a current awareness file of OSHA regulations regarding the safe handling
of the chemicals specified in this method. A reference file of material safety data sheets should be
made available to all personnel involved in the chemical analysis of samples suspected to contain
PCDDs/PCDFs. Additional references to laboratory safety are given in references 3, 4 and 5.
11.3 Each laboratory must develop a strict safety program for the handling of PCDDs and
PCDFs. The laboratory practices listed below are recommended.
11.3.1 Contamination of the laboratory will be minimized by conducting most of the
manipulations in a hood.
11.3.2 The effluents of sample splitters for the gas chromatograph and roughing
pumps on the HRGC/HRMS system should pass through either a column of activated charcoal
or be bubbled through a trap containing oil or high boiling alcohols.
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11.3.3 Liquid waste should be dissolved in methanol or ethanol and irradiated with
ultraviolet light at a wavelength less than 290 nm for several days (use F 40 BL lamps, or
equivalent). Using this analytical method, analyze the irradiated liquid wastes and dispose of
the solutions when 2,3,7,8-TCDD and 2,3,7,8-TCDF congeners can no longer be detected.
11.4 The following precautions for safe handling of 2,3,7,8-TCDD in the laboratory were
issued by Dow Chemical U.S.A, (revised 11/78) and amended for use in conjunction with this
method. They are as complete as possible on the basis of available toxicologica! information. The
precautions for safe handling and use are necessarily general in nature since detailed, specific
recommendations can be made only for the particular exposure and circumstances of each individual
use. Assistance in evaluating the health hazards of particular plant conditions may be obtained from
certain consulting laboratories and from State Departments of Health or of Labor, many of which
have an industrial health service. The 2,3,7,8-TCDD isomer is extremely toxic to certain kinds of
laboratory animals. However, it has been handled for years without injury in analytical and biological
laboratories. Many techniques used in handling radioactive and infectious materials are applicable
to 2,3,7,8-TCDD.
11.4.1 Protective equipment Throw away plastic gloves, apron or lab coat, safety
glasses and laboratory hood adequate for radioactive work. However, PVC gloves should not
be used.
11.4.2 Training: Workers must be trained in the proper method of removing
contaminated gloves and clothing without contacting the exterior surfaces.
11.4.3 Personal hygiene: Thorough washing of hands and forearms after each
manipulation and before breaks (coffee, lunch, and shift).
11.4.4 Confinement: Isolated work area, posted with signs, segregated glassware
and tools, plastic backed absorbent paper on benchtops.
11.4.5 Waste: Good technique includes minimzing contaminated waste. Plastic bag
liners should be used in waste cans.
11.4.6 Disposal of hazardous wastes: Refer to the November 7,1986 issue of the
Federal Register on Land Ban Rulings for details concerning the handling of dioxin-containing
wastes.
11.4.7 Personnel decontamination: Apply a mild soap with plenty of scrubbing action.
Glassware, tools and surfaces - Chlorothene NU Solvent™ (Dow Chemical Company) is the
least toxic solvent shown to be effective. Satisfactory cleaning -may be accomplished by
rinsing with Chlorothene, then washing with a detergent and water. Dishwater may be
disposed to the sewer after percolation through a charcoal bed filter. It is prudent to minimize
solvent wastes because they require costly special disposal through commercial services.
11.4.8 Laundry: Clothing known to be contaminated should be disposed with the
precautions described under "Disposal of Hazardous Wastes". Laboratory coats or other
clothing worn in 2,3,7,8-TCDD work area may be laundered. Clothing should be collected in
plastic bags. Persons who convey the bags and launder the clothing should be advised of the
hazard and trained in proper handling. The clothing may be put into a washer without contact
if the launderer knows the problem. The washer should be run through one full cycle before
being used again for other clothing.
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11.4.9 Wipe tests: A useful method for determining cleanliness of work surfaces and
tools is to wipe the surface with a piece of filter paper, extract the filter paper and analyze the
extract.
NOTE: A procedure for the collection, handling, analysis, and reporting requirements of wipe
tests performed within the laboratory is described in Appendix A. The results and
decision making processes are based on the presence of 2,3,7,8-substituted
PCDDs/PCDFs.
11.4.10 Inhalation: Any procedure that may generate airborne contamination must
be carried out with good ventilation. Gross losses to a ventilation system must not be allowed.
Handling of the dilute solutions normally used in analytical and animal work presents no
significant inhalation hazards except in case of an accident.
11.4.11 Accidents: Remove contaminated clothing immediately, talcing precautions
not to contaminate skin or other articles. Wash exposed skin vigorously and repeatedly until
medical attention is obtained.
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APPENDIX A
PROCEDURES FOR THE COLLECTION, HANDLING, ANALYSIS, AND
REPORTING OF WIPE TESTS PERFORMED WITHIN THE LABORATORY
This procedure is designed for the periodic evaluation of potential contamination by 2,3,7,8-
substituted PCDD/PCDF congeners of the working areas inside the laboratory.
A, 1 Perform the wipe tests on surface areas of two inches by one foot with glass fiber paper
saturated with distilled in glass acetone using a pair of clean stainless steel forceps. Use one wiper
for each of the designated areas. Combine the wipers to one composite sample in an extraction jar
containing 200 mL of distilled-in-glass acetone. Place an equal number of unused wipers in 200 mL
acetone and use this as a control. Add 100 pL of the sample fortification solution (Sec. 5.8) to each
jar containing used or unused wipers.
A. 1.1 Close the jar containing the wipers and the acetone and extract for 20 minutes
using a wrist action shaker. Transfer the extract into a K-D apparatus fitted with a
concentration tube and a three-ball Snyder column. Add two PTFE or Carborundum ™ boiling
chips and concentrate the extract to an apparent volume of 1.0 mL on a steam bath. Rinse
the Snyder column and the K-D assembly with two 1-mL portions of hexane into the
concentrator tube, and concentrate its contents to near dryness with a gentle stream of
nitrogen. Add 1.0 mL of hexane to the concentrator tube and swirl the solvent on the walls.
A. 1.2 Prepare a neutral alumina column as described in Sec. 7.5,2.2 and follow the
steps outlined in Sees. 7.5.2.3 through 7.5.2.5.
A. 1.3 Add 10 mL of the recovery standard solution as described in Sec. 7.5.3.6.
A.2 Concentrate the contents of the vial to a final volume of 10 pL (either in a miniviai or
in a capillary tube). Inject 2 pL of each extract (wipe and control) onto a capillary column and
analyze for 2,3,7,8-substituted PCDDs/PCDFs as specified in the analytical method in Sec. 7.8.
Perform calculations according to Sec. 7.9.
A.3 Report the presence of 2,3,7,8-substituted PCDDs and PCDFs as a quantity (pg or ng)
per wipe test experiment (WTE). Under the conditions outlined in this analytical protocol, a lower
limit of calibration of 10 pg/WTE is expected for 2,3,7,8-TCDD. A positive response for the blank
(control) is defined as a signal in the TCDD retention time window at any of the masses monitored
which is equivalent to or above 3 pg of 2,3,7,8-TCDD per WTE. For other congeners, use the
multiplication factors listed in Table 1, footnote (a) (e.g., for OCDD, the lower MCL is 10 x 5 = 50
pg/WTE and the positive response for the blank would be 3 x 5 = 15 pg). Also, report the recoveries
of the internal standards during the simplified cleanup procedure.
A.4 At a minimum, wipe tests should be performed when there is evidence of contamination
in the method blanks.
A.5 An upper limit of 25 pg per TCDD isomer and per wipe test experiment is allowed (use
multiplication factors listed in footnote (a) from Table 1 for other congeners). This value corresponds
to 214 times the lower calibration limit of the analytical method. Steps to correct the contamination
must be taken whenever these levels are exceeded. To that effect, first vacuum the working places
(hoods, benches, sink) using a vacuum cleaner equipped with a high efficiency particulate absorbent
(HEPA) filter and then wash with a detergent. A new set of wipes should be analyzed before anyone
is allowed to work in the dioxin area of the laboratory after corrective action has been taken.
8290A - 42
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January 1998
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TABLE 1
TYPES OF MATRICES, SAMPLE SIZES, AND 2,3,7,8-TCDD-BASED
METHOD CALIBRATION LIMITS (PARTS PER TRILLION)

Water
Soil
Sediment
Paper Pulp6
Fly Ash
Fish
Tissue"
Human
Adipose
Tissue
Sludge
Fuel Oil
Still
Bottom
Lower MCL"
0.01
1.0
1.0
1.0
1.0
5.0
10
Upper MCL"
2
200
200
200
200
1000
2000
Sample
Weight (g)
1000
10
10
20
10
2
1
IS Spiking
Level (ppt)
1
100
100
100
100
500
1000
Final Ext.
Vol. (mL)«
10-50
10-50
50
10-50
10-50
50
50
8 For other congeners multiply the values by 1 for TCDF/PeCDD/PeCDF, by 2.5 for
HxCDD/HxCDF/HpCDD/HpCDF, and by 5 for OCDD/OCDF.
b Sample dewatered according to Sec. 6.5.
c One half of the extract from the 20 g sample is used for determination of lipid content (Sec. 7.2.2).
" See Sec. 7.8.1.
NOTE: Chemical reactor residues are treated as still bottoms, if their appearances so suggest.
8290A - 43
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TABLE 2
COMPOSITION OF THE SAMPLE FORTIFICATION
AND RECOVERY STANDARD SOLUTIONS3
Analyte
Sample Fortification
Solution Concentration
(pgW
Recovery Standard Solution
Concentration (pg/^L)
13C12-2,3,7,8-TCDD
10
—
13C12-2,3,7,8-TCDF
10
—
13C12-1,2,3,4-TCDD
-
50
13C12-1,2,3,7,8-PeCDD
10
—
13C12-1,2,3,7,8-PeCDF
10
—
13C12-1,2,3,6,7,0-HxCDD
25
—
13C12-1,2,3,4,7,8-HxCDF
25
—
13C12-1,2,3,7,8,9-HxCDD
-
50
13C12-1,2,3,4,6,7,8-HpCDD
25
—
13C12-1,2,3,4,6,7,8-HpCDF
25
—
13C12-OCDD
50

'These solutions should be made freshly every day in nonane or other appropriate solvent because
of the possibility of adsorptive losses to glassware. If these solutions are to be kept for more than
one day, then the sample fortification solution concentrations should be increased ten fold, and the
recovery standard solution concentrations should be doubled. Corresponding adjustments of the
spiking volumes must then be made.
8290A-44
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January 1998
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TABLE 3
THE FIFTEEN 2,3,7,8-SUBSTITUTED PCDD AND PCDF CONGENERS
PCDD
PCDF
2,3,7,8-TCDDf)
2,3,7,8-TCDF(*)
1,2,3,7,8-PeCDD(*)
1,2,3,7,8-PeCDFf)
1,2,3,6,7,8-HxCDD(#)
2,3,4,7,8-PeCDF
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDF
1,2,3,7,8,9-HxCDD(+)
1,2,3,7,8,9-HxCDF
1,2,3,4,6,7,8-HpCDD(*)
1,2,3,4,7,8-HxCDFC)

2,3,4,6,7,8-HxCDF

1,2,3,4,6,7,8-HpCDFC*)

1,2,3,4,7,8,9-HpCDF
* The 13C-labeled analogue is used as an internal standard.
+ The 13C-labeled analogue is used as a recovery standard.
8290A - 45
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TABLE 4
ISOMERS OF CHLORINATED DIOXINS AND FURANS
AS A FUNCTION OF THE NUMBER OF CHLORINE ATOMS
Number of
Number of
Number of
Number of
Number of
Chlorine Atoms
Dioxin Isomers
2,3,7,8-Dioxins
Furan Isomers
2,3,7,8-Furans
1
2
—
4
_
2
10
—
16

3
14
—
28
—
4
22
1
38
1
5
14
1
28
2
6
10
3
16
4
7
2
1
4
2
8
1
1
1
1
Total
75
7
135
10
8290A - 46
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TABLES
HIGH-RESOLUTION CONCENTRATION CALIBRATION SOLUTIONS
Analyte
Unlabeled Analytes

Concentration (pg/^L)

5
4
3
"" 2
1
2,3,7,8-TCDD
200
50
10
2.5
1
2,3,7,8-TCDF
200
50
10
2.5
1
12,3,7,8-PeCDD
500
125
25
6.25
2.5
12,3,7,8-PeCDF
500
125
25
6.25
2.5
2,3,4,7,8-PeCDF
500
125
25
6.25
2.5
1,2,3,4,7,8-HxCDD
500
125
25
6.25
2.5
1^,3,6,7,8-HxCDD
500
125
25
6.25
2.5
1,2,3,7,8,9-HxCDD
500
125
25
6.25
2.5
1,2,3,4,7,8-HxCDF
500
125
25
6.25
2.5
1^,3,6,7,8-HxCDF
500
125
25
6.25
2.5
12,3,7,8,S-HxCDF
500
125
25
625
2.5
2,3,4,6,7,8-HxCDF
500
125
25
6.25
2.5
1,2,3,4,6,7,8-HpCDD
500
125
25
6.25
2.5
1,2,3,4,6,7,8-HpCDF
500
125
25
6.25
2.5
1,2,3,4,7,8,9-HpCDF
500
125
25
625
2.5
OCDD
1,000
250
50
12.5
5
OCDF
1,000
250
50
12.5
5
Internal Standards

13Cl2-2,3,7,8-TCDD
50
50
50
50
50
13C«-2,3,7,8-TCDF
50
50
50
50
50
13Ch-1 ,2,3,7,8-PeCDD
50
50
50
50
50
13Cl2-1,2,3,7,8-PeCDF
50
50
50
50
50
1SC12-1,2,3,6,7,8-HXCDD
125
125
125
125
125
13C«-1,2,3,4,7,8-HxCDF
125
125
125
125
125
1sCn2-1,2,3,4,6,7,8-HpCDD
125
125
125
125
125
,3C12-1,2,3,4,6,7,8-HpCDF
125
125
125
125
125
13C12-OCDD
250
250
250
250
250
Recoverv Standards

13C12-1,2,3,4-TCDD
50
50
50
50
50
13C12-1,2,3,7,8,9-HxCDD
125
125
125
125
125
8290A - 47
Revision 1
January 1998
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TABLE 6
IONS MONITORED FOR HRGC/HRMS ANALYSIS OF PCDDS/PCDFS
Descriptor Accurate Mass* ion ID Elemental Composition Analyte
303.9016
M
C^H^Cip
TCDF
305.8987
M+2
C^H^CI/CIO
TCDF
315.9419
M
<%2h*c\4o
TCDF (S)
317.9389
M+2
13C12H43SCI337CIO
TCDF (S)
319.8965
M
C^H^CIA
TCDD
321.8936
M+2
c^h^ci/'ciOj
TCDD
331.9368
M
13c12h435ci4o2
TCDD (S)
333.9338
M+2
1®C12H43WCK>i
TCDD (S)
375.8364
M+2
C12H435CI537CIO
HxCDPE
[354.9792]
LOCK
c9f13
PFK
339.8597
M+2
C12H3^CI437CIO
PeCDF
341.8567
M+4

PeCDF
351.9000
M+2
"CnH*CUvClO
PeCDF (S)
353.8970
M+4

PeCDF (S)
355.8546
M+2
c.jH^a^cio,
PeCDD
357.8516
M+4
c12h335ci337ci2o2
PeCDD
367.8949
M+2
^i^Ci^CIOj
PeCDD (S)
369.8919
M+4
13C12H3®CI337CI202
PeCDD (S)
409.7974
M+2
Ci2H335C^37CIO
HpCDPE
P54.9792J
LOCK
V.
PFK
373.8208
M+2
C^H^C^CIO
HxCDF
375.8178
M+4
CuH^Cl/CljO
HxCDF
383.8639
M
13c12H23Sa6o
HxCDF (S)
385.8610
M+2
"c^H^i^ao
HxCDF (S)
389.8156
M+2
C12H235Cl537CI02
HxCDD
391.8127
M+4
Ci2H235CI437CI202
HxCDD
8290A-48
Revision 1
January 1998
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TABLE 6
(continued)
Descriptor
Accurate Mass*
Ion ID
Elemental Composition
Analyte

401.8559
M+2
13c12h235ci537cio2
HxCDD(S)

403.8529
M+4
13c12h235ci437ci2o2
HxCDD (S)

445.7555
M+4
CizH^Cl/Cip
OCDPE

[430.9728]
LOCK
CgFtf
PFK
4
407.7818
M+2
c^h^ci^cio
HpCDF

409.7788
M+4
C^H^CIg^CljO
HpCDF

417.8250
M
"C^H^O
HpCDF (S)

419.8220
M+2
"C^H^Ci/CIO
HpCDF

423.7767
M+2
C12H35C!B37CI02
HpCDD

425.7737
M+4
p- u36/-*i 37/"*! n
V^i2ii Wig
HpCDD

435.8169
M+2
13/** L|35f-M
Wig wlv/2
HpCDD (S)

437.8140
M+4
13f* lj35/^i 37^m f\
v^n wig V/12U2
HpCDD (S)

479.7165
M+4
p lj35/-M 37i-sj
W^" W17 vljU
NCDPE

[430.97281
LOCK
C9F17
PFK
5
441.7428
M+2
c^fao
OCDF

443.7399
M+4
Cu^a^CiaO
OCDF

457.7377
M+2
a 35/-M 37mA
W17 viVJj
OCDD

459.7348
M+4
r* 35fM 37/-st
w^2 vig W12W2
OCDD

469.7780
M+2
13/"* 35/"M
w^2 W17 W1U2
OCDD (S)

471.7750
M+4
13/"* 35/-»| 37*"m f\
W-J2 Wig WI2W2
OCDD (S)

513.6775
M+4
c^a/c^o
DCDPE

[442.97281
LOCK

PFK
S = internal/recovery standard
a The following nuclidic masses were used:
H = 1.007825 O = 15.994915
C = 12.000000 ^Cl = 34.968853
13C - 13.003355 37Ci = 36.965903
F = 18.9984
8290A - 49
Revision 1
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taoi e f
I
-------
TABLE 8
THEORETICAL ION ABUNDANCE RATIOS AND THEIR CONTROL LIMITS
FOR PCDDS AND PCDFS
# Chlorine
Atoms
Ion Type
Theoretical Abundance Ratio
Control Limits
Lower
Upper
4
M/M+2
0.77
0.65
0.89
5
M+2/M+4
1.55
1.32
1.78
6
M+2/M+4
1.24
1.05
1.43
g(a)
M/M+2
0.51
0.43
0.59
7
-------
TABLE 9
RELATIVE RESPONSE FACTOR [RF (NUMBER)] ATTRIBUTIONS
Number
Specific Congener Name
1
2,3,7,8-TCDD (and total TCDDs)
2
2,3,7,8-TCDF (and total TCDFs)
3
1,2,3,7,8-PeCDD(and total PeCDDs)
4
1,2,3,7,8-PeCDF
5
2,3,4,7,8-PeCDF
6
1,2,3,4,7,8-HxCDD
7
1,2,3,6,7,8-HxCDD
8
1,2,3,7,8,9-HxCDD
9
1,2,3,4,7,8-HxCDF
10
1,2,3,6,7,8-HxCDF
11
1,2,3,7,8,9-HxCDF
12
2,3,4,6,7,8-HxCDF
13
1,2,3,4,6,7,8-HpCDD (and total HpCDDs)
14
1,2,3,4,6,7,8-HpCDF
15
1,2,3,4,7,8,9-HpCDF
16
OCDD
17
OCDF
18
13C12-2,3,7,8-TCDD
19
13C12-2,3,7,8-TCDF
20
13C12-1,2,3,7,8-PeCDD
21
13C12-1,2,3,7,8-PeCDF
22
13C12-1,2,3,6,7,8-HxCDD
23
13C12-1,2,3,4,7,8-HxCDF
24
13C12-1,2,3,4,6,7,8-HpCDD
25
13C12-1,2,3,4,6,7,8-HpCDF
26
13C12-OCDD
27
Total PeCDFs
28
Total HxCDFs
29
Total HxCDDs
30
Total HpCDFs
8290A-52
Revision 1
January 1998
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TABLE 10
2,3,7,8-TCDD TOXICITY" EQUIVALENCY FACTORS (TEFS)
FOR THE POLYCHLORINATED DIBENZODIOXINS AND DlBENZOFURANS
Analyte
TEP
2,3,7,8-TCDD
1.00
1,2,3,7,8-PeCDD
0.50
1,2,3,6,7,8-HxCDD
0,10
1,2,3,7,8,9-HxCDD
0.10
1,2,3,4,7,8-HxCDD
0.10
1,2,3,4,6,7,8-HpCDD
0.01
1,2,3,4,6,7,8,9-OCDD
0.001
2,3,7,8-TCDF
0.1
1,2,3,7,8-PeCDF
0.05
2,3,4,7,8-PeCDF
0.5
1,2,3,6,7,8-HxCDF
0.1
1,2,3,7,8,9-HxCDF
0.1
1,2,3,4,7,8-HxCDF
0.1
2,3,4,6,7,8-HxCDF
0.1
1,2,3,4,6,7,8-HpCDF
0.01
1,2,3,4,7,8,9-HpCDF
0.01
1,2,3,4,6,7,8,9-OCDF
0.001
'Taken from "Interim Procedures for Estimating Risks Associated with Exposures to Mixtures of
Chlorinated Dibertzo-p-Dioxin and -Dibenzofurans (CDDs and CDFs) and 1989 Update", (EPA/625/3-
89/016, March 1989).
8290A - 53
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TABLE 11
ANALYTE RELATIVE RETENTION TIME REFERENCE ATTRIBUTIONS
Analyte
Analyte RRT Reference8
1,2,3,4,7,8-HxCDD
13012-1,2,3,6,7,8-HxCDD
1,2,3,6,7,8-HxCDF
13C12-1 ,2,3,4,7,8-HxCDF
1,2,3,7,8,9-HxCDF
"C12-1,2,3,4,7,8-HxCDF
2,3,4,6,7,8-HxCDF
13Cl2-1,2,3,4,7,8-HxCDF
The retention time of 2,3,4,7,8-PeCDF on the DB-5 column, is measured
relative to 13Cir1,2,3,7t8-PeCDF and the retention time of 1,2,3,4,7,8,9-HpCDF
relative to 13C12-1,2,3,4,6,7,8-HpCDF.
8290A - 54
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TABLE 12
COMPARISON OF SOXHLET AND PRESSURIZED FLUID EXTRACTION (PFE)
FOR EXTRACTION OF GROUND CHIMNEY BRICK
Soxhiet (n=1) PFE (n=2)*
Analyle (ng/kg) (rig/kg)
2,3,7,8-TCDD
6
6
1,2,3,7,8-PeCDD
52
57
1,2,3,4,7,8-HxCDD
46
52
1,2,3,6,7,8-HxCDD
120
130
1,2,3,7,9,9-HxCDD
97
1000
1,2,3,4,6,7,8-HpCDD
1000
820
OCDD
2900
2600
2,3,7,8-TCDF
160
180
1,2,3,7,8 (+ 1,2,3,4,8)-PeCDF
430
470
2,3,4,7,9-PeCDF
390
390
1,2,3,4,7,8 (+ 1,2,3,4,7,9)-HxCDF
1100
1100
1,2,3,6,7,8-HxCDF
540
570
2,3,4,6,7,8-HxCDF
400
360
1,2,3,7,8,9-HxCDF
42
42
1,2,3,4,6,7,8-HpCDF
2100
2000
1,2,3,4,7,8,9-HpCDF
140
120
OCDF
2000
2000
Total TCDD
440
530
Total PeCDD
900
940
Total HxCDD
1800
2000
Total HpCDD
2000
2100
Total TCDF
2300
2600
Total PeCDF
4100
4300
Total HxCDF
4700
4700
Total HpCDF
2800
2600
* Sum of two extractions of each sample
Data from Reference 8
8290A - 55
Revision 1
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TABLE 13
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF URBAN DUST
Soxhlet (n=1) PFE (n=2)*
Analyte (ng/kg) (ng/kg)
_____ , — ~ 32
1.2.3.7.8-PeCDD 11.8 13.1
1,2,3,4,7,8-HxCDD 9.8 8.0
1.2.3.6.7.8-HxCDD 11.5 9.5
1.2.3.7.9.9-HxCDD ND(8) ND (8)
1,2,3,4,6,7,8-HpCDD 113 107
OCDD 445 314
2,3,7,8-TCDF 12.5 18.6
1,2,3,7,8 (1,2,3,4,8)«PeCDF 9.9 12.0
2.3.4.7.9-PeCDF 13.9 18.1
1,2,3,4,7,8 ( + 1^,3,4,7.9D-HXCOF 18.7 23.7
1,2,3,6,7,8-HxCDF 10.7 15.8
2.3.4.6.7.8-HxCDF 3.3 8.7
1.2.3.7.8.9-HxCDF ND (2) ND(2)
1.2.3.4.6.7.8-HpCDF 13.2 29.4
1.2.3.4.7.8.9-HpCDF ND(3) ND(3)
OCDF ND (10) ND (10)
Total TCDD 182 325
Total PeCDD 175 281
Total HxCDD 86.7 81.7
Total HpCDD 221 217
Total TCDF 333 419
Total PeCDF 146 179
Total HxCDF 65.9 122
Total HpCDF 13.2 29,4
ND = Not detected, with detection limit given in parentheses
* Sum of two extractions of each sample
Data from Reference 8
8290A - 56
Revision 1
January 1998
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TABLE 14
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF FLY ASH
(with and without HCl pretreatrnent for PFE)
Soxhlet (n=1) PFE (n=2)* PFE (n=2)*
with HCl (fjg/lcg)t with HCl w/o HCl
Analyte

*
2,3,7,8-TCDD
0.32
0.36
0.28
1,2,3,7,8-PeCDD
1.6
2.1
"1.7
1,2,3,4,7,8-HxCDD
1.2
14
1.2
1,2,3,6,7,8-HxCDD
2.4
2.7
2.4
1,2,3,7,9,9-HxCDD
2.4
2.3
2.2
1,2,3,4,6,7,8-HpCDD
8.2
9.6 *
8.1
OCDD
11.4
12.8
10.6
2,3,7,8-TCDF
3.7
4.3
3.4
1,2,3,7,8 (+ 1,2,3,4,8)-PeCDF
4.2
4.6
3.9
2,3,4,7,9-PeCDF
5.6
6.6
5.8
1,2,3,4,7,8 (+ 1,2,3,4,7,9)-HxCDF
7.8
8.7
5.4
1,2,3,6,7,8-HxCDF
7.2
8.5
5.3
2,3,4,6,7,8-HxCDF
6.6
7.2
4.5
1,2,3,7,8,9-HxCDF
0.43
0.56
0.30
1,2,3,4,6,7,8-HpCDF
18.0
17.6
16.8
1,2,3,4,7,8,9-HpCDF
2.3
2.4
2.0
OCDF
13.5
15.8
13.9
Total TCDD
12.0
12.4
10.5
Total PeCDD
16.6
20.5
16.2
Total HxCDD
38.2
42.4
36.7
Total HpCDD
15.0
19.8
16.0
Total TCDF
60.5
67.5^
56.1
Total PeCDF
83.5
87.3
77.4
Total HxCDF
65.2
73.5
46.1
Total HpCDF
28.1
32.2
26.5
f Fly ash was pretreated with HCl, followed by a water rinse, and extracted with toluene,
j These samples received no HCl pretreatrnent, and were extracted with a mixture of toluene
and acetic acid.
* Sum of two extractions of each sample
Data from Reference 8
8290A - 57
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TABLE 15
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF SOIL (EC-2)
Analyte
Soxhlet Results (n=10)
PFE Results (n=2)
ng/kg
% RSD
ng/kg
% RSD
2,3,7,8-TCDD
270
9.1
270
0.0
1,2,3,7,8-PeCDD
24
12
22
3.3
1,2,3,4,7,8-HxCDD
23
8.3
24
3.0
1,2,3,6,7,8-HxCDD
83
3.6
87
0.8
1,2,3,7,9,9-HxCDD
60
6.2
57
7.4
1,2,3,4,6,7,8-HpCDD
720
6.7
720
1.0
OCDD
4000
6.2
4200
0.0
2,3,7,8-TGDF *
100
7.3
82
2.6
1,2,3,7,8 (+ 1,2,3,4,8)-PeCDF
39
14
36
3.9
2,3,4,7,9-PeCDF
62
5.5
60
0.0
1,2,3,4,7,8 (+ 1,2,3,4,7,9)-HxCDF
740
5.3
690
0.0
1,2,3,6,7,8-HxCDF
120
6.2
120
0.0
2,3,4,6,7,8-HxCDF
45
9.0
60
1.2
1,2,3,7,8,9-HxCDF
4.9
31
5.3
15
1,2,3,4,6,7,8-HpCDF
2600
6.7
2500
0.0
1,2,3,4,7,8,9-HpCDF
160
5.5
160
0.0
OCDF
7800
8.3
7000
3.1
Total TCDD
430
9.7
370
1.9
Total PeCDD
300
3.7
280
7.7
Total HxCDD
720
5.8
690
2.0
Total HpCDD
1300
7.0
1300
0.0
Total TCDF
620
12
380
18
Total PeCDF
820
9.4
710
7.0
Total HxCDF
1900
5.7
1900
0.0
Total HpCDF
3800
8.2
3900
3.6
* Single-column analysis only, may include contributions from other isomers that may co-elute.
Data from Reference 8
8290A - 58
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TABLE 16
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF SEDIMENT (HS-2)
Soxhlet Results (n=10) PFE Results (r>=2)
Analyte ng/kg % RSD rig/kg % RSD
2,3,7,8-TCDD
ND (1)
-
ND (1)
-
1,2,3,7,8-PeCDD
1.6
4.6
ND (1)
_
1,2,3,4,7,8-HxCDD
4.5
4.8
5.2
11
1,2,3,6,7,8-HxCDD
19
4.3
21
0.0
1,2,3,7,9,9-HxCDD
24
4.3
28
2.6
1,2,3,4,6,7,8-HpCDD
1200
8.1
1300
0.0
Apnn
6500
4.2
7100
0.0
2,3,7,8-TCDF *
8.5
11
6.6
5.4
1,2,3,7,8 ( + 1,2,3,4,8)-PeCDF
1.9
17
2.0
0.0
2,3,4,7,9-PeCDF
3.7
7.9
3.7
3.8
1,2,3,4,7,8 (+ 1,2,3,4,7,9}-HxCDF
17
7.3
17
4.3
1,2,3,6,7,8-HxCDF
3.7
5.6
4.0
5.4
2,3,4,6,7,8-HxCDF
3.7
18
4.4
3.2
1,2,3,7,8,9-HxCDF
ND (1)
-
ND (1)

1,2,3,4,6,7,8-HpCDF
91
1.6
96
3.7
1,2,3,4,7,8,9-HpCDF
5.2
6.7
5.3
6.7
OCDF
300
3.8
280
2.6
Total TCDD
3.9
14
2.5
34
Total PeCDD
17
7.8
10
10
Total HxCDD
510
5.6
570
1.3
Total HpCDD
4700
8.3
5100
11
Total TCDF
39
11
24
3.0
Total PeCDF
33
13
" 28
0.0
Total HxCDF
89
3.2
87
12
Total HpCDF
293
3.3
310
0.0
* Single-column analysis only, may include contributions from other isomers that may co-elute.
, ND * Not detected, with detection limit given in parentheses
. Data from Reference 8
8290A-59
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TABLE 17
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF CONTAMINATED SEDIMENTS

Hamilton Harbor
Parrots Bay
Analyte
Soxhlet
PFE
Soxhlet
PFE
2,3,7,8-TCDD
3.7
3,1
19
19
1,2,3,7,8-PeCDD
5.1
5.4
8.3
6.0
1,2,3,4,7,8-HxCDD
6.4
7.2
8.6
6.7
1,2,3,6,7,8-HxCDD
27
26
26
17
1,2,3,7,9,9-HxCDD
20
28
24
18
1,2,3,4,6,7,8-HpCDD
460
430
280
250
OCDD
3100
3100
1900
1600
2,3,7,8-TCDF *
61
44
80
48
1,2,3,7,8 (+1,2,3,4,8)-PeCDF
14
14
ND (20)
9.8
2,3,4,7,9-PeCDF
26
25
22
14
1,2,3,4,7,8 (+ 1,2,3,4,7,9)-HxCDF
27
37
79
59
1,2,3,6,7,8-HxCDF
17
16
ND (20)
15
2,3,4,6,7,8-HxCDF
14
14
21
11
1,2,3,7,8,9-HxCDF
ND(2)
1.6
4.9
ND (1)
1,2,3,4,6,7,8-HpCDF
130
130
270
220
1,2,3,4,7,8,9-H pCDF
14
13
17
12
OCDF
270
210
510
370
Total TCDD
50
14
39
48
Total PeCDD
63
15
87
66
Total HxCDD
220
180
230
200
Total HpCDD
850
810
580
530
Total TCDF
370
130
400
270
Total PeCDF
290
110
180
170
Total HxCDF
240
160
230
230
Total HpCDF
350
290
400
360
* Single-column analysis only, may include contributions from other isomers that may co-etute.
. ND = Not detected, with detection limit given in parentheses
. Data from Reference 8
8290A - 60
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FIGURE 1
GENERAL STRUCTURES OF DIBENZO-p-DIOXIN (TOP) AND DIBENZOFURAN (BOTTOM)
8
o
o
8
o
o
8290A - 61
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FIGURE 2
M/AM
I
W 1
5.600
5,600
8 650
, W W \J
Peak profile displays demonstrating the effect of (he detector zero on the measured resolving power.
In this example, the true resolving power is 5,600.
A) The zero was set too high; no effect is observed upon the measurement of the resolving
power.
B) The zero was adjusted property.
C) The zero was set too low; this results in overestimating the actual resolving power because
the peak-to-peak noise cannot be measured accurately.
8290A - 62 Revision 1
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FIGURE 3
TYPICAL 12-HOUR ANALYSIS SEQUENCE OF EVENTS.
Analytical Procedure
8:00 AM
8:00 PM
9:00 AM
11:00 AM
Samples
Method
Blank
Concentrate to 10 mL
Initial or
Routine
Calibration
Mass
Resolution
Thaw Sample Extract
Routine
Calibration
GC Column
Performance
Mass Resolution
Mass Accuracy
8290A- 63
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FIGURE 4
oo ....
100

22:30
25:30
24:00
27:00
Time
Selected ion current profile for m/z 322 (TCDDs) produced by MS analysis of the GC performance
check solution on a 60 m DB-5 fused-siiica capillary column under the conditions listed in Sec. 7.6.
8290A - 64
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FIGURE 5
95 ppm
Ref. mass 304.9824 Peak top
Span. 200 ppm
System file name YVES1S0
Data file name A:85Z567
Resolution 10000
Group number 1
ionization mode EI +
Switching VOLTAGE
Ref. masses 304.9824
380.S260
M/ AM—10.500
Channel B 380.9260 Lock mass
Span 200 ppm
Peak profiles representing two PFK reference ions at m/z 305 and 381. The
resolution of the high-mass signal is 95 ppm at 5 percent of the peak height;
this corresponds to a resolving power M/CsM of 10,500 {10 percent valley definition).
8290A-65
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FIGURE 6
MANUAL DETERMINATION OF S/N.
Ei
T
_s
N
117
19.5
, i
E,
»
Jtu.Mi lilMl Li... J... ¦( t1 fli. il vm LVI.i li'.i I
*-c
0+—
20:00
1 1 ' I
22:00 24:00
26:00
I
28:00
. -Z_*.Q *
C]
30.00
The peak height (S) is measured between the mean noise (lines C and D). These mean signal
values are obtained by tracing the line between the baseline average noise extremes, E1 and
E2, and between the apex average noise extremes, E3 and E4, at the apex of the signal.
NOTE: It is imperative that the instalment interface amplifier electronic zero offset be set high
enough so that negative going baseline noise is recorded.
8290A - 66
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METHOD 829QA
POLYCHLORINATED DIBENZODIOXINS (PCDDs) AND POLYCHLORINATED
DIBENZOFURANS (PCDFs) BY HIGH-RESOLUTION GAS
CHROMATOGRAPHY/HIGH-RESOLUTION MASS SPECTROMETRY (HRGC/HRMS)
Start
_5_
7.1 Add internal standard
to sample.
7.2 - 7.4 Prepare sample and extract
using appropriate procedure
- Fish and Pulp/Paper
- Human Adipose Tissue
- Sludge/Wet Fuel Oil
- Still Bottom/Oil
- Fly Ash
- Aqueous Samples
- Soil/Sediment
7.S Cleanup Samples
- Partition
- Silica/Aluminum Column
• Carbon Column
>

7.6 Establish GC/MS and data
acquisition settings.

r
7,7 Calibrate GC/MS system.

r
7.8 Analyze samples and identify
compounds.

f
7.9 Perform calculations.
Stop
8290A - 67
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4.3 DETERMINATION OF ORGANIC ANALYTES
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the
allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to
be used by the analyst and the regulated community in making judgements necessary to meet
the data quality objectives or needs for the intended use of the data.
4.3.3 HIGH PERFORMANCE LIQUID CHROMATOGRAPHIC METHODS
The following methods are included in this section:
Method 8310: Polynuclear Aromatic Hydrocarbons
Method 8315A: Determination of Carbonyl Compounds by High Performance
Liquid Chromatography (HPLC)
Appendix A: Recrystallization of 2,4-Dinitrophenylhydrazine
(DNPH)
Acrylamide, Acrylonitrile and Acrolein by High Performance
Liquid Chromatography (HPLC)
N-Methyicarbamates by High Performance Liquid
Chromatography (HPLC)
Solvent-Extractable Nonvolatile Compounds by High
Performance Liquid Chromatography/Thermospray/Mass
Spectrometry (HPLC/TS/MS) or Ultraviolet (UV) Detection
Solvent Extractable Nonvolatile Compounds by High
Performance Liquid Chromatography/Particle Beam/Mass
Spectrometry (HPLC/PB/MS)
Nitroaromatics and Nitramines by High Performance Liquid
Chromatography (HPLC)
Tetrazene by Reverse Phase High Performance Liquid
Chromatography (HPLC)
Nitroglycerine by High Performance Liquid Chromatography
Method 8316:
Method 8318:
Method 8321B:
Method 8325:
Method 8330A:
Method 8331:
Method 8332:
FOUR-12 Revision 4
January 1998
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METHOD 8321B
SOLVENT-EXTRACTABLE NONVOLATILE COMPOUNDS BY
HIGH PERFORMANCE LIQUID CHROMATOGRAPHY/THERMOSPRAY/MASS
SPECTROMETRY /HPLC/TS/MS) OR ULTRAVIOLET fUV> DETECTION
1.0 SCOPE AND APPLICATION
1.1 This method covers the use of high performance liquid chromatography (HPLC),
coupled with both thermospray-mass spectrometry (TS-MS) and an ultraviolet (UV) detector, for the
determination of disperse azo dyes, organophosphorus compounds, tris(2,3-dibromopropyt)
phosphate, chlorinated phenoxyacid compounds and their esters, and carbamates in wastewater,
ground water, and soil/sediment matrices. Data are also provided for the determination of
chlorophenoxy acid herbicides in fly ash (Table 12), however, recoveries for most compounds are
very low, indicating poor extraction efficiency for these analytes using the extraction procedure
included in this method. The following compounds may be determined by this method, although not
all of the compounds are amenable to UV detection:
Analyte
CAS No.
Azo Dves
Disperse Red 1
Disperse Red 5
Disperse Red 13
Disperse Yellow 5
Disperse Orange 3
Disperse Orange 30
Disperse Brown 1
Solvent Red 3
Solvent Red 23
2872-52-8
3769-57-1
126038-78-6
6439-53-8
730-40-5
5261-31-4
17464-91-4
6535-42-8
85-86-9
Anthraouinone Dves
Disperse Blue 3
Disperse Blue 14
Disperse Red 60
Coumarin Dyes
2475-46-9
2475-44-7
17413-58-5
Fluorescent Briahteners
Fluorescent Brightener61
Fluorescent Brightener 236
8066-05-5
3333-62-8
Alkaloids
Caffeine
Strychnine
58-08-2
57-24-9
8321B -1
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Analyte
CAS No.
Oraanophosphorus Compounds
Methomyl
Thiofanox
Famphur
Asulam
Dichlorvos
Dimethoate
Disulfoton
Fensulfothion
Merphos
Parathion methyl
Monocrotophos
Naled
Phorate
Trichlorfon
Tris(2,3-dibromopropyl) phosphate (Tris-BP)
Chlorinated Phenoxvacid Compounds
Dalapon
Dicamba
2,4-D
MCPA
MCPP
Dichlorprop
2,4,5-T
Si I vex (2,4,5-TP)
Dinoseb
2,4-DB
2,4-D, butoxyethanol ester
2,4-D, ethylhexyl ester
2,4,5-T, butyl ester
2,4,5-T, butoxyethanol ester
Carbamates
Aldicarb*
Aldicarb sulfone
Aldicarb sulfoxide
Aminocarb
83210 -2
16752-77-5
39196-18-4
52-85-7
3337-71-1
62-73-7
60-51-5
298-04-4
115-90-2
150-50-5
298-00-0
6923-22-4
300-76-5
298-02-2
52-68-6
126-72-7
75-gg-O
1918-00-9
94-75-7
94-74-6
7085-19-0
120-36-5
93-76-5
93-72-1
88-85-7
94-82-6
1929-73-3
1928-43-4
93-79-8
2545-59-7
116-06-3
1646-88-4
1646-87-3
2032-59-9
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Analyte
CAS No.
Barban
101-27-9
Benomyl
17804-35-2
Bromadl
314-40-9
Bendiocarb*
22781-23-3
Cartoaryl*
63-25-2
Carbendazim*
10605-21-7
3-Hydroxycarbofuran
16655-82-6
Carbofuran*
1563-66-2
Chloroxuron
1982-47-4
Chloropropham
101-21-3
Diuron*
330r54-1
Fenuron
101-42-8
Fluometuron
2164-17-2
Linuron*
330-55-2
Methiocarb
2032-65-7
Methomyl*
16752-77-5
Mexacarbate
315-18-4
Monuron
150-68-5
Neburon
555-37-3
Oxamyl*
23135-22-0
Propachlor
1918-16-7
Propham
122-42-9
Propoxur
114-26-1
Siduron
1082-49-6
Tebuthiuron
34014-18-1
8 Chemical Abstract Service Registry Number.
* These carbamates were tested in a multi-laboratory evaluation.
All others were tested in a single-laboratory evaluation. - -
1.2 This method may be applicable to the analysis of other non-volatile or semivolatile
compounds that are solvent-extractable, are amenable to HPLC, and can be ionized under
thermospray introduction for mass spectrometric detection or can be determined by a UV detector.
1.3 Method 8321 is designed to detect the chlorinated phenoxyacid compounds (free add
form) and their esters without the use of hydrolysis and esterification in the extraction procedure,
although hydrolysis to the add form will simplify quantitation.
1.4 The compounds listed in this method were chosen for analysis by HPLC/MS because
they have been designated as problem compounds that are hard to analyze by gas chromatographic
8321B - 3
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methods. The sensitivity of this method is dependent upon the level of interferants within a given
matrix, and varies with compound class and even by compound within a class. Additionally, the
sensitivity is dependent upon the mode of operation of the mass spectrometer, with the selected
reaction monitoring (SRM) mode providing greater sensitivity than single quadrupole scanning.
1.5 For further compound identification, MS/MS (CAD - Collision Activated Dissociation)
can be used as ari optional extension of this method.
1.6 Tris-BP has been classified as a carcinogen. Purified standard material and stock
standard solutions should be handled in a hood.
1.7 This method is restricted to use by, or under the supervision of, analysts experienced
in the use of high performance liquid chromatography using mass spectrometers or ultraviolet
detectors. Analysts should also be skilled in the interpretation of liquid chromatograms and mass
spectra. Each analyst must demonstrate the ability to generate acceptable results with this method.
2.0 SUMMARY OF METHOD
2.1 This method provides reversed-phase high performance liquid chromatographic
(RP/HPLC) and thermospray (TS> mass spectrometry (MS) conditions and ultraviolet (UV)
conditions for the detection of the target analytes.
2.1.1 Sample extracts can be analyzed by direct injection into the thermospray or
onto a liquid chromatographic-thermospray interface
2.1.2 A gradient elution program is used on the chromatograph to separate the
compounds.
2.1.3 Quantitative analysis may be performed by either TS/MS or UV detection,
using either an external or internal standard approach. TS/MS detection may be performed
in either a negative ionization (discharge electrode) mode or a positive ionization mode, with
a single quadrupole mass spectrometer.
2.1.4 In some cases, the thermospray interface may introduce variability that leads
to less precise quantitation. In such instances, the MS response may be used to identify the
analytes of Interest while the quantitative results are derived from the response of the UV
detector.
2.2 Prior to analysis, appropriate sample preparation techniques must be used.
2.2.1 Samples for analysis of chlorinated phenoxyacid compounds may be prepared
by a modification of Method 8151 (see Sec. 7.3) or other appropriate extraction technique. In
general, the pHofa 1-L aqueous sample or 50-g solid sample is adjusted and the sample is
extracted with diethyl ether, concentrated, and the solvent exchanged to acetonitrile. Samples
for these analytes may also be extracted using solid-phase extraction after a pH adjustment,
as described in Method 3535.
2.2.2 For carbamates, 1-L aqueous samples or 40-g solid samples are extracted
with methylene chloride (refer to appropriate 3500 series method), concentrated (preferably
using a rotary evaporator with adapter) and the solvent exchanged to methanol.
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2.2.3 Samples for analysis of the other target analytes are prepared by established
extraction techniques. In general, water samples are extracted at a neutral pH with methylene
chloride, using an appropriate 3500 series method. Solid samples are extracted with a mixture
of methylene chloride/acetone (1:1), using an appropriate 3500 series method. Extract may
require concentration and solvent exchange prior to analysis.
2.2.4 A micro-extraction technique for the extraction of Tris-B P from aqueous and
non-aqueous matrices is included in this method (see Sec. 7.2).
2.3 An optional thermospray-mass spectrometry/mass spectrometry (TS-MS/MS)
confirmatory procedure is provided in this method (see Sec. 7.11). That procedure employs MS/MS
Collision Activated Dissociation (CAD) or wire-repeiier CAD.
3.0 INTERFERENCES
3.1 Refer to Methods 3500, 3600, 8000 and 8151.
3.2 The use of Florisil Column Cleanup (Method 3620) has been demonstrated to yield
recoveries less than 85% for some of the compounds in this method, and is therefore not
recommended for all compounds. Refer to Table 2 of Method 3620 for recoveries of
organophosphorus compounds as a function of Florisil fractions.
3.3 Compounds with high proton affinity may mask the MS response of some of the target
analytes. Therefore, except when the thermospray MS/MS system is used for rapid screening of
samples (see Sec. 7.11.1), an HPLC must be used to perform the chromatographic separations
necessary for quantitative analyses.
3.4 Analytical difficulties encountered with specific organophosphorus compounds, as
applied in this method, may include, but are not limited to, the following:
3.4.1 Methyl parathion shows some minor degradation during analysis.
3.4.2 Naled can undergo debromination to form dichlorvos. This reaction may
occur during sample preparation and extraction, and the extent may depend of the nature of
the sample matrix. The analyst should consider the potential for debromination of Naled when
this compound is to be determined.
3.4.3 Merphos often contains contamination from merphos oxide. Oxidation of
merphos can occur during storage, and possibly upon introduction into the mass spectrometer.
3.4.4 The water solubility of dichlorvos (DDVP) is 10 g/L at 20°C, and as a result,
recovery of the this compound by solvent extraction from aqueous solutions is poor.
3.4.5 Trichloron rearranges and undergoes dehydrochlorination (loss of HO) in
acidic, neutral, or basic media, forming dichlorvos (DDVP). When either of these compounds
are to be determined, the analyst should be aware of the possibility of this rearrangement in
order to prevent misidentifications.
3.5 The chlorinated phenoxy acid compounds, being strong organic adds, react readily with
alkaline substances and may be lost during analysis. Therefore, glassware and glass wool must be
8321B - 5
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acid-rinsed, and sodium sulfate must be acidified with sulfuric add prior to use, to avoid this
possibility.
3.6 Due to the reactivity of the chlorinated herbicides, the standards must be prepared in
acetonitiile. Methyiation mil occur slowly, if prepared in methanol.
v. 3.7 Benomyl quickly degrades to carbendazim in the environment (Reference 21).
3.8 Solvents, reagents, glassware, and other sample processing hardware may yield
discrete artifacts or elevated baselines, or both, causing misinterpretation of chromatograms or
spectra. All of these materials must be demonstrated to be free from interferences under the
conditions of the analysis by running reagent blanks. Specific selection of reagents and purification
of solvents by distillation in all-glass systems may be required.
3.9 Interferants co-extracted from the sample wili vary considerably from source to source.
Retention times of target analytes must be verified by using reference standards.
3.10 The optional use of HPLC/MS/MS methods aids in the confirmation of specific analytes.
These methods are less subject to chemical noise than other mass spectrometric methods.
4.0 APPARATUS AND MATERIALS
4.1 HPLC/MS
The following apparatus and materials are necessary for the use of the HPLC/MS portions of
this method.
4.1.1 High performance liquid chromatograph (HPLC) - An analytical system with
programmable solvent delivery system and all required accessories, including injection loop
(with a minimum 10-jjL loop volume), analytical columns, purging gases, etc. At a minimum,
the solvent delivery system must be capable of delivering a binary solvent system. The
chromatographic system must be capable of being interfaced with a mass spectrometer (MS).
4.1.2 HPLC post-column addition pump - If post-column addition of reagents is
employed, a pump is required. Ideally, this pump should be a syringe pump, and does not
have to be capable of solvent programming. It is also possible to add the ionization reagents
to the solvents and not perform post-column addition (see Sec. 7.6).
4.1.3 HPLC/MS interface
4.1.3.1 Interface - Thermospray ionization interface and source that will
give acceptable calibration response for each analyte of interest at the concentration
required. The source must be capable of generating both positive and negative ions,
and have a discharge electrode or filament.
4.1.3.2 Micromixer - 10-mL, connects HPLC column system with HPLC
post-column addition solvent system, if post-column addition is used.
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4.1.4 Mass spectrometer system
4.1.4.1 A angle quadrupole mass spectrometer capable of scanning from
1 to 1000 amu. The spectrometer must also be capable of scanning from 150 to 450
amu in 1.5 sec. or less, using 70 volts (nominal) electron energy in the positive or
negative electron impact modes. In addition, the mass spectrometer must be capable
of producing a calibrated mass spectrum for PEG 400, 600, or 800 (see Sec. 5.14) or
other compounds used for mass calibration.
4.1.4.2 Optional triple quadrupole mass spectrometer - capable of
generating daughter ion spectra with a collision gas in the second quadrupole and
operation in the single quadrupole mode.
4.1.5 Data system - A computer system that allows the continuous acquisition and
storage on machine-readable media of all mass spectra obtained throughout the duration of
the chromatographic program must be connected to the mass spectrometer. The computer
must have software that allows any MS data file to be searched for ions of a specified mass,
and such ion abundances to be plotted versus time or scan number. This type of plot is
defined as an Extracted Ion Current Profile (EICP). Software must also be available that
allows integration of the abundances in any EICP between specified time or scan-number
limits. There must be computer software available to operate the specific modes of the mass
spectrometer.
4.2 HPLC with UV detector
An analytical system with solvent programmable pumping system for at least a binary solvent
system, and all required accessories including syringes, 10-pL injection loop, analytical columns,
purging gases, etc. An automatic injebtor is optional, but is useful for multiple samples. The
columns specified in Sec. 4.3 are also used with this system.
If the UV detector is to be used in tandem with the thenmospray interface, then the detector cell
must be capable of withstanding high pressures (up to 6000 psi). However, the UV detector may
be attached to an HPLC independent of the HPLC/TS/MS and, in that case, standard HPLC
pressures are acceptable.
4.3 HPLC columns - A guard column and an analytical column are necessary.
The columns listed in this section were those used to develop the method. The mention of
these columns is not intended to exclude the use of other columns that are available or that may be
developed. Laboratories may use columns of other dimensions and/or packed with different
stationary phases, provided that they document method performance data (e.g., chromatographic
resolution, anaiyte breakdown, and quantitation limits) that provide analytical performance that is
appropriate for the intended application.
4.3.1 Guard Column - C18 reversed-phase guard column, 10 mm x 2.6 mm ID, 0.5-
|im frit, or equivalent. The guard column should be packed with the same or similar stationary
phase as the analytical column.
4.3.2 Analytical Column - C1fl reversed-phase column, 100 mm x 2 mm ID, 5-jjm
particle size of ODS-Hypersil; or Cb reversed phase column, 100 mm x 2 mm ID, 3-|jm particle
size of MOS2-Hypersil, or equivalent.
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4.4 Purification equipment for azo dye standards
4.4.1 Soxhlet extraction apparatus
4.4.2 Extraction thimbles - single thickness, 43 x 123 mm
4.4.3 Filter paper, 9.0 cm (Whatman qualitative No. 1 or equivalent).
4.4.4 Silica-gel column - 3 in. x 8 in., packed with silica gel (Type 60, EM reagent
70/230 mesh).
4.5 Extraction equipment for chlorinated phenoxyacid compounds
4.5.1 Erienmeyer flasks - 500-mL wide-mouth glass, 500-mL glass, with 24/40
ground-glass joint, 1000-mL glass.
4.5.2 Separatory funnel - 2000-mL.
4.5.3 Graduated cylinder -1000-mL.
4.5.4 Funnel - 75-mm diameter.
4.5.5 Wrist shaker - Bunnell Model 75 or equivalent.
4.5.6 pH meter.
4.6 Kudema-Danish (K-D) apparatus (optional).
4.6.1 Concentrator tube - 10-mL graduated. A ground-glass stopper is used to
prevent evaporation of extracts.
4.6.2 Evaporation flask - 500-mL. Attach to concentrator tube with springs,
clamps, or equivalent.
4.6.3 Two-ball micro-Snyder column
4.6.4 Springs - % in.
4.6.5 Solvent vapor recovery system (Kontes K-545000-1006 or K-547300-0000,
Ace Glass 6614-30, or equivalent).
NOTE: This glassware is recommended for the purpose of solvent recovery during the
concentration procedures requiring the use of Kudema-Danish evaporative
concentrators. Incorporation of this apparatus may be required by State or local
municipality regulations that govern air emissions of volatile organics. EPA
recommends the incorporation of this type of reclamation system as a method to
implement an emissions reduction program. Solvent recovery is a means to
conform with waste minimization and pollution prevention initiatives.
4.7 Disposable serological pipets
4.8 Collection tube - 15-mL conical, graduated.
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4.9 Vials - 5-mL conical, glass, with PTFE-Iined screw-caps or crimp tops.
4.10 Glass wool
4.11 Microsyringes - 100-pL, 50-mL, 10-pL (Hamilton 701 N or equivalent), and 50 jjL
(Blunted, Hamilton 705SNR or equivalent).
4.12 Rotary evaporator - Equipped with 1000-mL receiving flask.
4.13 Balances - Analytical, 0.0001 g, top-loading, 0.01 g.
4.14 Volumetric flasks, Class A - 10-m L to 1000-mL.
4.15 Graduated cylinder -100-mL.
4.16 Separatory funnel - 250-mL.
4.17 Separatory funnel - 2-L, with PTFE stopcock.
4.18 Concentrator adaptor (optional) - for carbamate extraction.
4.19 Nitrogen evaporation apparatus - N-Evap Analytical Evaporator Model 111,
Organomation Association Inc., Northborough, MA, or equivalent.
5.0 REAGENTS
5.1 Reagent grade chemicals shall be used in all tests. Unless otherwise indicated, it is
intended that all reagents shall conform to the specifications of the Committee on Analytical
Reagents of the American Chemical Society, where such specifications are available. Other grades
may be used, provided it is first ascertained that the reagent is of sufficiently high purity to permit its
use without lessening the accuracy of the determination.
5.2 Organic-free reagent water. All references to water in this method refer to organic-free
reagent water, as defined in Chapter One.
5.3 Sodium sulfate (granular, anhydrous), Na2S04. Purify by heating at 400°C for 4 hours
in a shallow tray, or by precleaning the sodium sulfate with methylene chloride.
5.4 Ammonium acetate, N^OOCCHa, solution (0.1 M). Filter through a 0.45-Mm
membrane filter (Millipore HA or equivalent).
5.5 Acetic acid, CH3C02H
5.6 Sulfuric acid solution
5.6.1 (1:1, v/v) - Slowly add 50 mL H2S04 (sp. gr. 1.84) to 50 mL of water.
5.6.2 (1:3, v/v) - Slowly add 25 mL H2SO< (sp. gr. 1.84) to 75 mL of water.
5.7 Argon gas, 99+% pure.
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5.8 Solvents - Unless otherwise noted, all solvents must be pesticide quality or equivalent.
5.8.1 Methylene chloride, CHjCIj
5.8.2 Toluene, CeH5CH3
5.8.3 Acetone, CHgCOCHa
5.8.4 Diethyl Ether, CjHgOCjHs - Must be free of peroxides as indicated by test
strips (EM Quant, or equivalent). Procedures for removal of peroxides are provided with the
test strips. After cleanup, 20 mL of ethyl alcohol preservative must be added to each liter of
ether,
5.8.5 Methanol, CH3OH - HPLC quality or equivalent.
5.8.6 Acetonitrile, CH3CN - HPLC quality or equivalent.
5.8.7 Ethyl acetate, CHaCOjCjHg
5.9 Standard materials - pure standard materials or certified solutions of each analyte
targeted for analysis. Disperse azo dyes must be purified before use according to Sec. 5.10.
WARNING: Tris-BP has been classified as a carcinogen. Purified standard material and stock
standard solutions should be handled in a hood.
5.10 Disperse azo dye purification
Two procedures are involved. The first step is the Soxhlet extraction of the dye for 24 hours
with toluene and evaporation of the liquid extract to dryness, using a rotary evaporator. The solid
is then recrystallized from toluene, and dried in an oven at approximately 100°C. If this step does
not give the required purity, column chromatography should be employed. Load the solid onto a 3
x 8 inch silica gel column (Sec. 4.4.4), and elute with diethyl ether. Separate impurities
chromatographically, and collect the major dye fraction.
5.11 Stock standard solutions - Standards may be prepared from pure standard materials
or may be purchased as certified solutions. Commercially-prepared stock standards may be used
if they are certified by the manufacturer and verified against a standard made from pure material.
5.11.1 Prepare stock standard solutions by accurately weighing 0.0100 g of pure
material. Dissolve the material in methanol or other suitable solvent (e.g., prepare Tris-BP in
ethyl acetate), and dilute to known volume in a volumetric flask. ~ .
NOTE: Due to the reactivity of the chlorinated herbicides, the standards must be prepared
in acetonitrile. Methylation will occur if standards are prepared in methanol.
If compound purity is certified at 96% or greater, the weight can be used without
correction to calculate the concentration of the stock standard. Commercially-prepared stock
standards can be used at any concentration if they are certified by the manufacturer or by an
independent source.
5.11.2 Transfer the stock standard solutions into glass vials with PTFE-lined
screw-caps or crimp-tops. Store at 4°C and protect from light. Stock standard solutions
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should be checked frequently for signs of degradation or evaporation, especially just prior to
preparing calibration standards.
5.12 Calibration standards - A minimum of five different concentrations for each parameter
of interest should be prepared through dilution of the stock standards with methanol (or other
suitable solvent). At least one of the calibration standards should correspond to a sample
concentration at or below that necessary to meet the data quality objectives of the project. The
remaining concentrations should correspond to the expected range of concentrations found in real
samples, or should define the working range of the HPLC-UV or HPLC-TS/MS system (see Method
8000). Calibration standards must be replaced after one or two months, or sooner if comparison
with check standards indicates a problem.
5.13 Surrogate standards - The analyst should monitor the performance of the extraction,
cleanup (when used), and analytical system, along with the effectiveness of the method in dealing
with each sample matrix, by spiking each sample, standard, and blank with one or two surrogates
(e.g., organophosphorus or chlorinated phenoxyadd compounds not expected to be present in the
sample).
5.14 HPLC/MS tuning standard - Polyethylene glycol 400 (PEG-400), PEG-600, or PEG-800
are recommended as tuning standards. However, analysts may use other tuning standards as
recommended by the instrument manufacturer or other documented source. If one of the PEG
solutions is used, dilute to 10 percent (v/v) in methanol. Which PEG is used will depend upon
analyte molecular weight range: m.w. <500, use PEG-400; m.w. >500, use PEG-600 or PEG-800.
5.15 Internal standards - When the internal standard calibration option is used for HPLC/MS
analyses, it is recommended that analysts use stable isotopically-labeled compounds of the same
chemical class when they are available (e.g., 13C6-carbofuran may be used as an internal standard
in the analysis of carbamates).
5.16 Matrix spiking standards - Consult Method 3500 for information on matrix spiking
solutions. Prepare a solution containing the analytes of interest in a suitable solvent.
NOTE: The form of the compounds used for spiking should be identical to the form of the target
analytes. For the phenoxyacid herbicides in particular, use the acid form of the acid
analytes, rot the ester form or an ether, as use of these other forms mil not represent the
performance of the overall extraction, cleanup, and determinative methods relative to the
target analytes. Conversely, when the ester forms are of the analytes of interest, e.g., 2,4-
D, butoxyethanol ester, use the ester form of the analyte for preparing matrix spiking
solutions.
6.0 SAMPLE COLLECTION, PRESERVATION, AND HANDLING
See the introductory material to Chapter Four, Organic Analytes, Sec. 4.1.
7.0 PROCEDURE
7.1 Sample preparation
Prior to analysis, samples must be extracted using either an appropriate 3500 series method
or using specific procedures described in this method.
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7.1.1 Samples for analysis of disperse azo dyes and organophosphorus compounds
must be prepared by an appropriate 3500 series method prior to HPLC/MS analysis.
7.1.2 Samples for the analysis of Tris(2,3-dibromopropyl)phosphate (Tris-BP) must
be prepared according to Sec. 7.2, prior to HPLC/MS analysis.
7.1.3 Samples for the analysis of chlorinated phenoxyacid compounds and their
esters should be prepared according to Sec. 7.3, or other appropriate technique, prior to
HPLC/MS analysis. TCLP leachates to be analyzed for the phenoxyacid herbicides may also
be prepared using solid-phase extraction (SPE), as described in Method 3535.
7.2 Microextraction of T ris-BP
7.2.1 Solid samples
7.2.1.1 Weigh a 1-g portion of the sample into a tared beaker, if the
sample appears moist, add an equivalent amount of anhydrous sodium sulfate and mix
well. Add 100 mL of Tris-BP (approximate concentration 1000 mg/L) to the sample
selected for spiking; the amount added should result in a final concentration of 100
ng/pL in the 1-mL extract.
7.2.1.2 Remove the glass wool plug from a disposable serological pipet.
insert a 1 cm plug of clean silane treated glass wool to the bottom (narrow end) of the
pipet. Pack 2 cm of anhydrous sodium sulfate onto the top of the glass wool. Wash
pipet and contents with 3 - 5 mL of methanol.
7.2.1.3 Pack the sample into the pipet prepared according to Sec. 7.2.1.2.
If packing material has dried, wet with a few mL of methanol first, then pack sample
into the pipet.
7.2.1.4 Extract the sample with 3 mL of methanol followed by 4 mL of 50%
(v/v) methanol/methylene chloride (rinse the sample beaker with each volume of
extraction solvent prior to adding it to the pipet containing the sample). Collect the
extract in a 15-mL graduated glass tube.
7.2.1.5 Evaporate the extract to 1 mL using the nitrogen evaporation
technique (Sec. 7.5). Record the volume. It may not be possible to evaporate some
sludge samples to a reasonable concentration.
7.2.1.6 Determination of percent dry weight - When sample results are to
be calculated on a dry weight basis, a second portion of sample should be weighed at
the same time as the portion used for analytical determination.
WARNING: The drying oven should be contained in a hood or be vented. Significant
laboratory contamination may result from drying a heavily contaminated
sample.
Immediately after weighing the sample for extraction, weigh 5 -10 g of the
sample into a tared crucible. Dry this aliquot overnight at 105°C. Allow to cool in a
desiccator before weighing. Calculate the % dry weight as follows:
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% dry weight = 9 <* dry sample x10Q
g of sample
This oven-dried aliquot is not used for the extraction and should be disposed of
appropriately once the dry weight has been determined.
7.2.2 Aqueous samples
7.2.2.1 Using a 100-mL graduated cylinder, measure 100 mL of sample
and transfer it to a 250-mL separatory funnel. Add 200 pL of Tris-BP (approximate
concentration 1000 mg/L) to the sample selected for spiking; the amount added should
result in a final concentration of 200 ng/pL in the 1-mL extract.
7.2.2.2 Add 10 mL of methylene chloride to the separatory funnel. Seal
and shake the separatory funnel three times, approximately 30 seconds each time, with
periodic venting to release excess pressure.
NOTE: Methylene chloride creates excessive pressure rapidly; therefore, initial
venting should be done immediately after the separatory funnel has been
sealed and shaken once. Methylene chloride is a suspected carcinogen, use
necessary safety precautions.
7.2.2.3 Allow the organic layer to separate from the water phase for a
minimum of 10 minutes. If the emulsion interface between layers is more than
one-third the size of the solvent layer, the analyst must employ mechanical techniques
to complete phase separation. See Section 7 of Method 3510.
7.2.2.4 Collect the extract in a 15-mL graduated glass tube. Concentrate
the extract to 1 mL, using nitrogen evaporation (Sec. 7.5).
7.3 Extraction for chlorinated phenoxyadd compounds
Preparation of soil, sediment, and other solid samples should follow the procedures outlined
in Method 8151, or other appropriate technique, with the exception of no hydrolysis or esterification
is generally performed. However, if the analyst desires to determine all of the phenoxyadd moieties
as the add, hydrolysis may be performed. Sec. 7.3.1 presents an outline of the procedure with the
appropriate changes necessary for determination by Method 8321. Sec. 7.3.2 describes the
extraction procedure for aqueous samples. TCLP leachates may be extracted using solid-phase
extraction, as described in Method 3535.
7.3.1 Extraction of solid samples
7.3.1.1 Add 50 g of soil/sediment sample to a 500-mL, wide-mouth
Ertenmeyer flask. Add spiking solutions, if required, mix well and allow to stand for 15
minutes. Add 50 mL of organic-free reagent water and stir for 30 minutes. Determine
the pH of the sample with a glass electrode and pH meter, while stirring. Adjust the
pH to 2 with cold H2S04 (1:1) and monitor the pH for 15 minutes, with stirring. If
necessary, add additional H2S04 until the pH remains at 2.
7.3.1.2 Add 20 mL of acetone to the flask, and mix the contents with the
wrist shaker for 20 minutes. Add 80 mL of diethyl ether to the same flask, and shake
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again for 20 minutes. Decant the extract and measure the volume of solvent
recovered.
7.3.1.3 Extract the sample twice more using 20 mL of acetone followed
by 80 mL of diethyl ether. After addition of each solvent, the mixture should be shaken
with the wrist shaker for 10 minutes and the acetone-ether extract decanted.
7.3.1.4 After the third extraction, the volume of extract recovered should
be at least 75% of the volume of added solvent. If this is not the case, additional
extractions may be necessary. Combine the extracts in a 2000-mL separator/ funnel
containing 250 mL of 5% acidified sodium sulfate. If an emulsion forms, slowly add 5
g of acidified sodium sulfate (anhydrous) until the solvent-water mixture separates. A
quantity of acidified sodium sulfate equal to the weight of the sample may be added,
if necessary.
7.3.1.5 Check the pH of the extract If it is not at or below pH 2, add more
concentrated H2S04 until the extract is stabilized at the desired pH. Gently mix the
contents of the separatory funnel for 1 minute and allow the layers to separate. Collect
the aqueous phase in a clean beaker, and the extract phase (top layer) in a 500-mL
Erienmeyer flask with a ground-glass stopper. Race the aqueous phase back into the
separatory funnel and re-extract using 25 mL of diethyl ether. Allow the layers to
separate and discard the aqueous layer. Combine the ether extracts in the 500-mL
Erlenmeyer flask.
7.3.1.6 Add 45 - 50 g acidified anhydrous sodium sulfate to the combined
ether extracts. Allow the extract to remain in contact with the sodium sulfate for
approximately 2 hours.
NOTE: The drying step is very critical. Any moisture remaining in the ether will result
in low recoveries. The amount of sodium sulfate used is adequate if seme
free flowing crystals are visible when swirling the flask. If all of the sodium
sulfate solidifies in a cake, add a few additional grams of acidified sodium
sulfate and again test by swirling. The 2-hour drying time is a minimum;
however, the extracts may be held overnight in contact with the sodium
sulfate,
7.3.1.7 Transfer the ether extract, through a funnel plugged with
acid-washed glass wool, into a 500-mL K-D flask equipped with a 10-mL concentrator
tube. Use a glass rod to crush caked sodium sulfate during the transfer. Rinse the
Erlenmeyer flask and column with 20-30 mL of diethyl ether to complete the
quantitative transfer. Reduce the volume of the extract using the macro K-D technique
(Sec. 7.5).
7.3.2 Extraction of aqueous samples
7.3.2.1 Using a 1000-mL graduated cylinder, measure 1 liter (nominal) of
sample, record the sample volume to the nearest 5 mL, and transfer it to a separatory
funnel. If high concentrations are anticipated, a smaller volume may be used and then
diluted with organic-free reagent water to 1 liter. Adjust the pH to less than 2 with
sulfuric acid (1:1).
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7.3.2.2 Add 150 mL of diethyl ether to the sample bottle, seal, and shake
for 30 seconds to rinse the walls. Transfer the solvent wash to the separatory funnel
and extract the sample by shaking the funnel for 2 minutes with periodic venting to
release excess pressure. Allow the organic layer to separate from the water layer for
a minimum of 10 minutes. If the emulsion interface between layers is more than
one-third the size of the solvent layer, the analyst must employ mechanical techniques
to complete the phase separation. The optimum technique depends upon the sample,
and may include stirring, filtration of the emulsion through glass wool, centrifugation,
or other physical methods. Drain the aqueous phase into a 1000-mL Erienmeyer flask.
7.3.2.3 Repeat the extraction two more times using 100 mL of diethyl
ether each time. Combine the extracts in a 500-mL Erienmeyer flask, (Rinse the 1 (Wo-
rn L flask with each additional aliquot of extracting solvent to make a quantitative
transfer.)
7.3.2.4 Proceed to Sec. 7.5 for drying, K-D concentration, solvent
exchange, and final volume adjustment.
7.4 Extraction of carbamates
Preparation of aqueous, soil, sediment, and other solid samples must follow an appropriate
3500 series method. The following sections provide general considerations.
7.4.1 One-liter aqueous samples are extracted with methylene chloride using an
appropriate 3500 series method.
7.4.2 Forty-gram quantities of solid samples are extracted with methylene chloride
using an appropriate 3500 series method.
7.4.3 Concentration steps can be performed using a rotary evaporator or K-D,
reducing the final extract to 5-10 mL.
7.4.4 Final concentration of the extract and exchanging the solvent to a 1-mL final
volume of methanol may be accomplished using an adaptor on the rotary evaporator. If an
adaptor is unavailable, the final concentration may be performed using nitrogen evaporation,
in a fume hood.
7.5 Extract concentration techniques
Two procedures are provided for the concentration of extracts: macro-concentration by
Kudema-Danish (K-D) and micro-concentration by nitrogen evaporation.
7.5.1 Macro-concentration by K-D
Add one or two clean boiling chips to the flask and attach a three-ball macro-Snyder
column. Attach the solvent vapor recovery glassware (condenser and collection device, Sec.
4.6.5) to the Snyder column of the K-D apparatus following manufacturer's instructions.
Prewet the Snyder column by adding about 1 mL of diethyl ether to the top. Place the
apparatus on a hot water bath (60°-65°C) so that the concentrator tube is partially immersed
in the hot water and the entire lower rounded surface of the flask is bathed in vapor. Adjust
the vertical position of the apparatus and the water temperature, as required, to complete the
concentration in 15-20 minutes. At the proper rate of distillation the balls of the column will
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actively chatter, but the chambers will not flood. When the apparent volume of liquid reaches
5 mL, remove the K-D apparatus from the water bath and allow it to drain and cool for at least
10 minutes.
7.5.2 Solvent exchange
Prior to analysis, the final extract solvent must be exchanged to methanol or
acetonitrile.
7.5.2.1 Transfer the concentrator tube to a nitrogen evaporation device.
Add a total of 5 mL of the final solvent of choice (methanol or acetonitrile).
7.5.2.2 Reduce the extract volume according to Sec. 7.5.3 and adjust the
final volume to 1 mL (or other volume necessary to achieve the required sensitivity).
7.5.3 Micro-concentration by nitrogen evaporation
7.5.3.1 Place the concentrator tube in a warm water bath (approximately
35°C) and evaporate the solvent volume to the required level using a gentle stream of
clean, dry nitrogen (filtered through a column of activated carbon).
CAUTION: Do not use piasticized tubing between the carbon trap and the sample.
7.5.3.2 The internal wall of the tube must be rinsed down several times
with the final solvent during the operation. During evaporation, the solvent level in the
tube must be positioned to prevent water from condensing into the sample (i.e., the
solvent level should be below the level of the water bath). Under normal operating
conditions, the extract should not be allowed to become dry.
7.5.4 Transfer the extract to a glass vial with a PTFE-lined screw-cap or crimp-top
and store refrigerated at 4°C. Proceed with HPLC analysis.
7.6 HPLC chromatographic conditions
7.6.1 Recommended mobile phases and elution gradients for some groups of
analytes are shown in Tables 1 and 2. Analysts should also consult the instrument
manufacturer's instructions. In the absence of specific recommendations, the following
conditions may be a useful starting point.
Flow rate 0.8 mL/min
Post-column mobile phase 0.1 M ammonium acetafe (1% methanol)/(0.1 M
ammonium acetate for phenoxyacid compounds)
Post-column flow rate 0.4 mL/min
Optimize the instrumental conditions for resolution of the target analytes and sensitivity. Post-
column addition of the MS ionization reagents may not be necessary in all instances, and these
reagents may be added to the elution solvents, provided that adequate performance can be
demonstrated.
NOTE: Once established, the same operating conditions must be used for both calibrations
and sample analyses.
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7.6.2 If there is a chromatographic problem from compound retention when
analyzing disperse azo dyes, organophosphorus compounds, or tris(2,3-dibromo-
propyl)phosphate, a 2% constant flow of methylene chloride may be applied as needed.
Methylene chloride/aqueous methanol solutions must be used with caution as HPLC eluants.
Acetic acid (1%), another mobile phase modifier, can be used with compounds with acid
functional groups.
7.6.3 A total flow rate of 1.0 to 1.5 mL/min may be necessary to maintain
thermospray ionization, however, consult the instrument manufacturer's instructions and adjust
the flow rate as needed.
7.7 Recommended thermospray/MS operating conditions
Prior to analysis of samples, the analyst should evaluate the relative sensitivity of the target
compounds to each ionization mode to determine which may provide better sensitivity during
analyses. This evaluation may be based on the structures of the anaiytes or by conducting analyses
in each of the two ionization modes. Some groups of target compounds will have much better
sensitivity using either positive or negative ionization (e.g., carbamates are generally more sensitive
to the positive ionization mode and phenoxyacids are generally more sensitive to the negative
ionization mode). When alt the anaiytes of interest for a given application respond adequately in a
given ionization mode, a single analysis using that mode may be employed.
7,7.1 Positive ionization mode conditions
Scan time
Optional repeller wire or plate
Discharge electrode
Filament
Mass range
Off
On or off (optional, analyte dependent)
150 to 450 amu (analyte dependent, expect 1 to
18 amu higher than molecular weight of the
compound).
1.50 sec/scan
170 to 250 v (sensitivity optimized). See Figure 2
for schematic of source with wire repeller.
7.7.2 Negative ionization mode conditions
Discharge electrode
Filament
Mass Range
Scan time
On
Off
135 to 450 amu
1.50 sec/scan
7.7.3 Thermospray temperatures
Vaporizer control
Vaporizer tip
Jet
Source block
110 to 130°C
200 to 215°C
210 to 220°C
230 to 265°C. (Some compounds may degrade in
the source block at higher temperatures, operator
should use knowledge of chemical properties to
estimate proper source temperature).
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7.7.4 Sample injection volume
An injection volume of 20 to 100 mL is normally used. The injection loop must be
overfilled by, minimally, a factor of two (e.g., 20-pL sample used to overfill a 10-pL injection
loop) when manual injections are performed. If solids are present in the extract, allow them
to settle or centrifuge the extract and withdraw the injection volume from the clear layer,
7.8 Calibration
7.8.1 Thermospray/MS system
When an MS detector is employed, the system must be tuned on quadrupole 1 (and
quadrupole 3 for triple quadaipoles) for accurate mass assignment, sensitivity, and resolution.
It is recommended that this be accomplished using polyethylene glycol (PEG) 400,600, or 800
(see Sec. 5.14) which have average molecular weights of 400, 600, and 800, respectively.
Analysts may use other tuning standards as recommended by the instrument manufacturer or
other documented source. If PEGS are used, a mixture of these PEGs can be made such that
it will approximate the expected working mass range for the analyses. Use PEG 400 for
analysis of chlorinated phenoxyacid compounds. The PEG is introduced via the thenmospray
interface, circumventing the HPLC.
7.8.1.1 The mass calibration parameters are as follows:
PEG 400 and 600 PEG 800
Mass range 15 to 765 amu Mass range 15 to 900 amu
Scan time 0.5 to 5.0 sec/scan Scan time 0.5 to 5.0 sec/scan
Approximately 100 scans should be acquired, with 2 to 3 injections made.
The scan with the best fit to the accurate mass table (see Tables 7 and 8) should be
used as the calibration table. If calibrants other than PEG are used, the mass range
should be from 15 to approximately 20 amu higher than the highest mass used for
calibration. A scan time should be chosen which will give at least 6 scans across the
calibrant peak.
7.8.1.2 The low mass range from 15 to 100 amu is covered by the ions
from the ammonium acetate buffer used in the thermospray process.
NH/
18 amu
NH/-H20
36 amu
ch3oh-nh/
50 amu (methanol)
CHaCN-NH/
59 amu (acetonitrile)
CH3OOH -NH/
78 amu (acetic acid)
The appearance of m/z 50 or 59 depends upon the use of methanol or acetonitrile as
the organic modifier. The higher mass range is covered by the ammonium ion adducts
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of the various ethylene glycols (e.g., H(OCH2CH2)„OH where n=4, gives the
H(OCH2CH2)4OH*NH/ ion at m/z 212).
7.8.2 Liquid chromatographic system
7.8.2.1 Choose the proper ionization conditions for the MS detector, as
outlined in Sec. 7.7. When UV detection is employed in conjunction with the MS
detector, establish appropriate operating conditions for the UV detector.
7.8.2.2 Prepare five calibration standards (see Sec. 5.12 and Method
8000). Inject each calibration standard onto the HPLC, using the chromatographic
conditions outlined in Table 1. Refer to Sec. 7.0 of Method 8000 for guidance on
external and internal calibration options and calibration acceptance criteria. In most
cases the (M*H)+ and (M*NH^)+ adduct ions are the only ions of significant abundance.
For example, Table 9 lists the retention times and the major ions (>5%) present in the
positive ionization thenmospray single quadrupole spectra of the organophosphorus
compounds.
7.8.2.3 The use of selective ion monitoring (SIM) is acceptable in
situations requiring detection limits below the normal range of full spectra analysis.
However, SIM may provide a lesser degree of confidence in the compound
identification unless multiple ions are monitored for each compound.
7.8.2.4 The use of selective reaction monitoring (SRM) is also acceptable
when using triple-quad MS/MS and enhanced sensitivity is needed.
7.8.2.5 If UV detection is being used, integrate the area under the full
chromatographic peak for each concentration. Quantitation by HPLC-UV may be
preferred if it is know that sample interference and/or analyte coetution are not a
problem, or when response of the MS detector is not sufficiently stable for quantitative
analyses. In these instances, the MS response may be used for positive qualitative
identification of the analytes while the UV response is used for quantitation.
7.8.2.6 The retention time of the chromatographic peak is an important
variable in analyte identification. Therefore, the relative retention time of the analyte
(versus the internal standard) should be in the range of 0.9 to 1.1.
7.8.3 Calibration verification
At the beginning of each analytical shift, the response of the instrument system must
be verified by the analysis of a single standard at the approximate mid-point of the initial
calibration range. Consult Method 8000 for information on performing this demonstration and
the acceptance criteria that should be employed.
7.9 Sample Analysis
Once the LC system has been calibrated as outlined in Sec 7.8, it is ready for sample analysis,
employing both MS and UV detectors. Depending on the sensitivity necessary for a given project,
analyses may be conducted using the MS detector in either the positive or negative ionization
modes. The positive ionization mode generally provides greater sensitivity, and may be more
appropriate for samples containing very low concentrations of the analytes of interest. However,
analyst are advised that some compounds may be detectable in only the negative ionization mode.
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7.9.1 An instrument blank (methanol) should be analyzed after the standards, in
order to demonstrate that the system is free from contamination.
7.9.2 if performing manual injections, take an appropriate aliquot of the sample as
per Sec. 7.7.4. Start the HPLC gradient elution, load and inject the sample aliquot, and start
the mass spectrometer data system analysis.
7.9.3 If using an autoinjector, ensure that it is set up property according to the
manufacturer's instructions and that all samples arid standards are loaded in the proper order.
Start the autoinjector, the HPLC gradient elution, and the mass spectrometer data system.
7.9.4 The concentration of the analyte is determined by using the initial calibration
data (see Method 8000) from either the MS or UV detector response. Samples whose
concentrations exceed the calibration range must be diluted to fall within the range.
7.9.5 When using MS or MS/MS, and when it is appropriate for the compounds of
interest and the project objectives, determinations in both positive and negative ionization
analyses may be done on each sample extract.
7.10 Calculations
7.10.1 Using the external or internal standard calibration procedure (Method 8000),
determine the identity and quantity of each component peak in the sample reconstructed ion
chromatogram which corresponds to the compounds used for calibration processes. See
Method 8000 for calculations.
7.10.2 The retention time of the chromatographic peak is an important parameter for
the identity of the analyte. However, because matrix interferences can change
chromatographic column conditions, the absolute retention times are not as significant as
relative retention times (when using internal standards), and the mass spectral patterns are
important criteria for analyte identification.
7.10.3 In instances when the TS/MS response exhibits higher variability, the MS
response may be used to identify the analytes of interest while the quantitative results are
derived from the response of the UV detector.
7.11 Optional MS/MS confirmation
With respect to this method, MS/MS shall be defined as the daughter ion collision activated
dissociation acquisition with quadrupole one set on one mass (parent ion), quadruple two
pressurized with argon and with a higher offset voltage than normal, and quadrupole three set to
scan desired mass range.
7.11.1 Since the thermospray process often generates only one or two ions per
compound, the use of MS/MS is a more specific mode of operation yielding molecular
structural information. In this mode, samples can be rapidly screened through direct injection
of the sample into the thermospray (e.g., without using the HPLC to separate the sample
components).
7.11.2 When using MS/MS, the first quadrupole should be set to the protonated
molecule or ammoniated adduct of the analyte of interest. The third quadrupole should be set
to scan from 30 amu to just above the mass region of the protonated molecule.
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7.11.3 The collision gas pressure (Ar) should be set at about 1.0 mTorr and the
collision energy at 20 eV. If these parameters fail to give considerable fragmentation, the
settings may be increased to create more and stronger collisions.
7.11.4 For analytical determinations, the base peak of the collision spectrum shall
be taken as the quantitation ion. For extra specificity, a second ion should be chosen as a
backup quantitation ion.
7.11.5 Perform an initial calibration, as outlined in Sec. 7.8.
7.11.6 MS/MS contamination and interferences
7.11.6.1 If the MS/MS mode is to be used without chromatographic
separation (rapid screening), then the method blank analysis must show that the
sample preparation and analysis procedures are free of contamination by the analyte
of interest or by interfering compounds. Refer to Sec. 8.0 of Method 8000 for guidance
on acceptable method blank performance. If contamination is detected in the method
blank above acceptable limits, re-extraction and reanalysis of the affected samples is
necessary.
7.11.6.2 The MS/MS spectra of a calibration standard and the sample
should be compared and the ratios of the three major (most intense) ions examined.
These ratios should be approximately the same unless there is an interference. If an
interference appears, chromatographic separation must be utilized.
7.11.6.3 The signal of the target analyte in a sample may be suppressed
by co-extracted interferences which do not give a signal in the monitored ions. In order
to monitor such signal suppression, an internal standard may be spiked into ail
standards, blanks, and sample extracts at a consistent concentration prior to analysis.
The internal standard may be any compound which responds well in the appropriate
ionization mode and which is not likely to be found in nature. (Note: Atrazine-ds has
been used successfully for positive ion analysis, while 2,6-dinitrotoluene-d3 has been
used successfully for negative ion analysis.) The amount spiked should be chosen
such that the signal produced is at least 100 times the noise level for the appropriate
ion. The signal of the internal standard should be monitored. Reanalysis is required
for any sample in which the internal standard peak height varies by more than 30%
from the average internal standard height obtained during the five-point calibration. If
reanalysis confirms this variance in signal, the sample should be reanalyzed using a
chromatographic separation. Quantitation of analyte concentration may be performed
using this internal standard. External standard quantitation is also allowed.
7.11.7 The total area of the quantitation ion(s) is calculated and the initial calibration
is used to calculate sample results.
7.11.8 MS/MS techniques can also be used to perform structural analysis on ions
represented by unassigned m/z ratios. The procedure for compounds of unknown structures
is to set up a CAD experiment on the ion of interest. The spectrum generated from this
experiment will reflect the structure of the compound by its fragmentation pattern. A trained
mass spectroscopist and some history of the sample are usually needed to interpret the
spectrum. (CAD experiments on actual standards of the expected compound are necessary
for confirmation or denial of that substance.)
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7.12 Optional wire-repeller CAD confirmation
7.12.1 See Figure 3 for the correct position of the wire-repeller in the thermospray
source Mock.
7.12.2 Once the wire-repeller is inserted into the thermospray flow, the voltage can
be increased to approximately 500 - 700 v. Enough voltage is necessary to create fragment
ions, but not so much that shorting occurs.
7.12.3 Continue as outlined in Sec. 7.9.
8.0 QUALITY CONTROL
8.1 Refer to Chapter One and Method 8000 for specific quality control (QC) procedures.
Each laboratory should maintain a formal quality assurance program. The laboratory should also
maintain records to document the quality of the data generated.
8.2 Quality control procedures necessary to evaluate the HPLC system operation are found
in Method 8000 and include evaluation of retention time windows, calibration verification and
chromatographic analysis of samples. Check the performance of the entire analytical system daily
using data gathered from analyses of blanks, standards, and replicate samples. If any of the
chromatographic QC limits are not met, the analyst should examine the LC system for
• Leaks,
• Proper pressure delivery,
• A dirty guard column; may need replacing or repacking, and
Possible partial thermospray plugging.
Checking any of tfie above items will necessitate shutting down the HFLC/TS system, making repairs
and/or replacements, and then restarting tfie analyses. A calibration verification standard should be
reanalyzed before any sample analyses, as described in Sec. 7.8.3.
8.3 Initial demonstration of proficiency - Each laboratory must demonstrate initial proficiency
with each sample preparation and determinative method combination it utilizes, by generating data
of acceptable accuracy and precision for target analytes in a clean matrix. The laboratory must also
repeat the following operations whenever new staff are trained or significant changes in
instrumentation are made. See Method 8000, Sec. 8.0 for information on how to accomplish this
demonstration.
8.4 Sample quality control for preparation and analysis - The laboratory must also have
procedures for documenting the effect of the matrix on method performance (precision, accuracy,
and detection limit). At a minimum, this includes the analysis of QC samples including a method
blank, matrix spite, a duplicate, and a laboratory control sample (LCS) in each analytical batch and
the addition of surrogates to each field sample and QC sample.
8.4.1 Documenting the effect of the matrix should include the analysis of at least
one matrix spike and one duplicate unspiked sample or one matrix spike/matrix spike duplicate
pair. The decision on whether to prepare and anafyze duplicate samples or a matrix
spike/matrix spike duplicate must be based on a knowledge of the samples in the sample
batch. If samples are expected to contain target analytes, then laboratories may use one
matrix spike and a duplicate analysis of an unspiked field sample. If samples are not expected
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to contain target analytes, laboratories should use a matrix spike and matrix spike duplicate
pair.
8.4.2 A laboratory control sample (LCS) should be included with each analytical
batch. The LCS consists of an aliquot of a clean (control) matrix similar to the sample matrix
and of the same weight or volume. The LCS is spiked with the same analytes at the same
concentrations as the matrix spike. When the results of the matrix spike analysis indicate a
potential problem due to the sample matrix itself, the LCS results are used to verify that the
laboratory can perform the analysis in a clean matrix.
8.4.3 See Method 8000, Sec. 8.0 for the details on carrying out sample quality
control procedures for preparation and analysis.
8.5 Surrogate recoveries - The laboratory must evaluate surrogate recovery data from
individual samples versus the surrogate control limits developed by the laboratory. See Method
8000, Sec. 8.0 for information on evaluating surrogate data and developing and updating surrogate
limits.
8.6 It is recommended that the laboratory adopt additional qualify assurance practices for
use with this method. The specific practices that are most productive depend upon the needs of the
laboratory and the nature of the samples. Whenever possible, the laboratory should analyze
standard reference materials and participate in relevant performance evaluation studies.
9.0 METHOD PERFORMANCE
9.1 Single operator accuracy and precision studies have been conducted using spiked
sediment, wastewater, sludge, and water samples. Tables 4,5, and 6 provide single-laboratory data
for Disperse Red 1. Table 1 provides the data for organophosphorus pesticides, Table 11 for
Tris-BP, Table 12 for chlorophenoxyacid herbicides, and Tables 14 ami 15 for carbamates.
9.2 Table 13 presents multi-laboratory accuracy and precision data for the chlorinated
phenoxyacid herbicides. The data summary is based on data from three laboratories that analyzed
duplicate solvent solutions at each concentration specified in the table.
9.3 Tables 16 and 17 present the multi-laboratory accuracy and precision data for the
carbamates. The data summary is based on data from nine laboratories that analyzed triplicate
solvent solutions at each concentration level specified in the tables.
9.4 Table 18 provides data for solid-phase extraction of 2,4-D and 2,4,5-TP spiked into
TCLP buffers at two different spiking levels.
10.0 REFERENCES
1. Voyksner, R.D., Haney, CA, "Optimization and Application of Thermospray High-Performance
Liquid Chromatography/Mass Spectrometry", Anal. Chem.. 1985, 57. 991-996.
2. Blakley, C.R., Vestal, M.L., Thermospray Interface for Liquid Chromatography/Mass
Spectrometry", Anal. Chem.. 1983, 55, 750-754.
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3. Taylor, V.» Hickey, D.M., Marsden, P.J., "Single Laboratory Validation of EPA Method 8140",
EPA-600/4-87/009, U.S. Environmental Protection Agency, Las Vegas, NV, 1987, 144 pp.
4. "Guidelines for Data Acquisition and Data Quality Evaluation in Environmental Chemistry",
Anal. Chem- 1980, 52, 2242-2249.
5. Betowski, L.D., Jones, T.L., "The Analysis of Organophosphorus Pesticide Samples by
HPLC/MS and HPLC/MS/MS", Environmental Science and Technology. 1988.
8. U.S. ERA: 2nd Annual Report on Carcinogens. NTP 81-43, Dec. 1981, pp. 236-237.
9. Blum, A., Ames, B.N., Science 195.1977,17.
10. Zweidinger, RA, Cooper, S.D., Pellazari, E.D., Measurements of Organic Pollutants In Water
and Wastewater. ASTM 686.
11. Cremlyn, R., Pesticides: Preparation and mode of Action. John Wiley and Sons, Chichester,
1978, p. 142.
12. Cotterill, E.G., Byast, T.H., "HPLC of Pesticide Residues in Environmental Samples", In Liouid
Chromatography in Environmental Analysis. Laurence, J.F., Ed., Humana Press, Clifton, NJ,
1984.
13. Voyksner, R.D., "Thermospray HPLC/MS for Monitoring the Environment", In Applications of
New Mass Spectrometry Technioues in Pesticide Chemistry: Rosen, J.D., Ed., John Wiley and
Sons: NewYork, 1987.
14. Yinon, J., Jones, T.L., Betowski, L.D., Rap. Comm. Mass Spectrom., 1989, 3, 38.
15. Shore, F.L., Amick, E.N., Pan, S.T., Gurka, D.F., "Single Laboratory Validation of EPA Method
8150 for the Analysis of Chlorinated Herbicides in Hazardous Waste", EPA/600/4-85/060, U.S.
Environmental Protection Agency, Las Vegas, NV, 1985.
16. "Development and Evaluations of an LC/MS/MS Protocol", EPA/600/X-86/328, Dec. 1983.
17. "An LC/MS Performance Evaluation Study of Organophosphorus Pesticides",
EPA/600/X-89/006, Jan. 1989.
18. "A Performance Evaluation Study of a Liquid Chromatography/Mass Spectrometry Method for
Tris-(2,3-Dibromopropyl) Phosphate", EPA/600/X-89/135, June 1989.
19. "Liquid Chromatography/Mass Spectrometry Performance Evaluation of Chlorinated
Phenoxyacid Herbicides and Their Esters", EPA/600/X-89/176, July 1989.
20. "An Interiaboratory Comparison of an SW-846 Method for the Analysis of the Chlorinated
Phenoxyacid Herbicides by LC/MS", EPA/600/X-90/133, June 1990.
21. Somasundaram, L., and J.R. Coates, Ed,, "Pesticide Transformation Products Fate and
Significance in the Environment", ACS Symposium Series 459, Ch. 13,1991.
22. Single-Laboratory Evaluation of Carbamates, APPL, Inc., Fresno, CA.
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23. "Interlaboratory Calibration Study of a Thermospray-Liquid Chromatography/ Mass
Spectrometry (TS-LC/MS) Method for Selected Carbamate Pestiddes", EPA/600/X-92/102,
August 1992.
24. Marked, C., "3M Data Submission to EPA," letter to B. Lesnik, June 27,1995.
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TABLE 1
RECOMMENDED HPLC CHROMATOGRAPHIC CONDITIONS
Analytes
Initial Mobile
Phase
(%)
Initial
Time
(min)
Final
Gradient
(linear)
Final Mobile
Phase(%)
Time
(min)
Organophosphorus
Compounds
50/50
(water/methanol)
0
10
100
(methanol)
5
Azo Dyes
50/50
(Watei/CHgCN)
0
5
100
(CH3CN)
5
Tris(2,3-
dibromopropyl)
phosphate
50/50
(water/methanol)
0
10
100
(methanol)
5
Chlorinated
phenoxyatid
compounds
75/25
(0.1 M NH4
acetate in 1%
acetic acid/
methanol)
2
15
40/60
(0.1 M NH4
acetate in 1%
acetic acid/
methanol)

40/60
(0.1 M
Ammonium
acetate in 1 %
acetic acid/
methanol)
3
5
75/25
(0.1 M Ammonium
acetate in 1%
acetic acid/
methanol)
10
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TABLE 2
RECOMMENDED HPLC CHROMATOGRAPHIC CONDITIONS FOR CARBAMATES
Time (min) Mobile phase A (percent) Mobile phase B (percent)
Option AO 95 5
30 20 80
35 0 100
40 95 5
45 95 5
A = 5 mM ammonium acetate with 0.1 M acetic acid, and
B = methanol, with optional post-column addition of 0.5 M ammonium acetate
Time (min) Mobile phase A (percent) Mobile phase B (percent)
Option B 0 95 5
30 0 100
35 0 100
40 95 5
45 95 5
A = water with 0.1 M ammonium acetate with 1% acetic acid
B = methanol with 0.1 M ammonium acetate with 1% acetic add, with
optional post-column addition of 0.1 M ammonium acetate.
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TABLE 3
COMPOUNDS AMENABLE TO THERMOSPRAY MASS SPECTROMETRY
Disperse Azo Dyes Alkaloids
Methine Dyes Aromatic ureas
Arylmethane Dyes Amides
Coumarin Dyes Amines
Anthraquinorie Dyes Amino acids
Xanthene Dyes Organophosphorus Compounds
Flame retardants Chlorinated Phenoxyaeid Compounds
Carbamates
TABLE 4
PRECISION AND ACCURACY COMPARISONS OF MS AND MS/MS WITH
HPLC/UV FOR ORGANIC-FREE REAGENT WATER SPIKED WITH DISPERSE RED 1
Percent Recovery
HPLC/UV MS CAD SRM
Spike 1 82.2 ± 0.2 92.5 ±3.7 87.6 ± 4.6 95.5 ±17.1
Spike 2 87.4 ± 0.6 90.2 ±4.7 90.4 ±9.9 90.0 ±5.9
RPD 6.1% 2.5% 3.2% 5.9%
Data from Reference 16.
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TABLE 5
PRECISION AND ACCURACY COMPARISONS OF MS AND MS/MS WITH
HPLC/UV FOR MUNICIPAL WASTEWATER SPIKED WITH DISPERSE RED 1
Percent Recovery
HPLC/UV MS CAD
Spike 1 93.4 ±0.3 102.0 ±31 82.7 ±13
Spike 2 96.2 ±0.1 79.7 ±15 83.7 ±5.2
RPD 3.0% 25% 1.2%
Data from Reference 16.
TABLE 6
RESULTS FROM ANALYSES OF ACTIVATED SLUDGE PROCESS WASTEWATER
Recovery of Disperse Red 1 (mg/L)
HPLC/UV MS CAD
5 mg/L Spiking Concentration
1 0.721 ± 0.003 0.664 ± 0.030 0.796 ± 0.008
1-D 0.731 ± 0.021 0.600 ± 0.068 0.768 ± 0.093
2 0.279 ± 0.000 0.253 ± 0.052 0.301 ± 0.042
3 0.482 ±0.001 0.449 ±0.016 0.510 ±0.091
RPD 1.3% 10.1% 3.6%
0 mg/L Spiking Concentration
1 0.000 0.005 ±0.0007 <0.001
1-D 0.000 0.006 ±0.001 <0.001
2 0.000 0.002 ±0.0003 <0.001
3 0.000 0.003 ± 0.0004 <0.001
RPD - 18.2%
Data from Reference 16.
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TABLE 7
CALIBRATION MASSES AND % RELATIVE ABUNDANCES OF PEG 400
Mass % Relative Abundance8
18.0 32.3
35.06 13.5
36.04 40.5
50.06 94.6
77.04 27.0
168.12 5.4
212.14 10.3
256.17 17.6
300.20 27.0
344.22 45.9
388.25 64.9
432.28 100
476.30 94.6
520-33 81.1
564.35 67.6
608.38 32.4
652.41 16.2
653.41 4.1
696.43 8.1
697.44 2.7 _
a Intensities are normalized to mass 432.
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TABLE 8
CALIBRATION MASSES AND % RELATIVE ABUNDANCES OF PEG 600
Mass % Relative Abundance8
18.0
4.7
36.04
11.4
50.06
64.9
77.04
17.5
168.12
9.3
212.14
43.9
256.17
56.1
300.20
22.8
344.22
28.1
388.25
38.6
432.28
54.4
476.30
64.9
520.33
86.0
564.35
100
608.38
63.2
652.41
17.5
653,41
5.6
696.43
1.8
* Intensities are normalized to mass 564.
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TABLE 9
RETENTION TIMES AND THERMOSPRAY MASS SPECTRA
OF ORGANOPHOSPHORUS COMPOUNDS
Compound
Retention Time (min)
Mass (% Relative Abundance)8
Monocrotophos
Trichlorfon
Dimethoate
Dichlorvos
Naled
Fensulfothion
Parathion methyl
Phorate
Disulfoton
Merphos
1:09
1:22
1:28
4:40
9:16
9:52
10:52
13:30
13:55
18:51
241 (100), 224 (14)
274 (100), 257 (19). 238 (19)
230 (100), 247 (20)
238 (100), 221 (40)
398 (100), 381 (23), 238 (5), 221, (2)
" 326 (10), 309 (100)
281 (100), 264 (8), 251 (21), 234 (48)
278 (4), 261 (100)
292(10), 275(100)
315 (100), 299 (15)
a For molecules containing a, Br and S, only the base peak of the isotopic cluster is listed.
Data from Reference 17.
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TABLE 10
SINGLE OPERATOR ACCURACY AND PRECISION FOR LOW CONCENTRATION DRINKING
WATER, LOW CONCENTRATION SOIL, MEDIUM CONCENTRATION DRINKING
WATER, MEDIUM CONCENTRATION SEDIMENT
Mean Std. Spike Recovery #
Matrix Compound Rec. (%) Dev. Cone. Range (%) Analyses
Low conc.
drinking water
~imethoate
70
7.7
5
85-54
15
ftjg/L)
Dichlorvos
40
12
5
64-14
15

Naled
0.5
1.0
5
2-0
15

Fensulfothion
112
3.3
5
119-106
15

Parathion
methyl
50
28
10
105-0
15

Phonate
16
35
5
86-0
15

Disulfoton
3.5
8
5
19-0
15

Merphos
237
25
5
287 -187
15
Low com. soil
Dimethoate
16
4
50
24-7
15
ftjg/kg)
Dichlorvos
ND

Naled
ND

Fensulfothion
45
5
50
56-34
15

Parathion
methyl
ND

100

Phorate
78
15
50
109-48
15

Disulfoton
36
7
50
49-22
15

Merphos
118
19
50
155-81
15
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TABLE 10
(continued)
Matrix
Compound
Mean
Rec. (%)
Std.
Dev.
Spike
Conc.
Recovery
Range {%)
#
Analyses
Medium conc.
drinking water
Dimethoate
52
4
50
61-43
12
(pg/L)
Dichlorvos
146
29
50
204-89
12

Naled
4
3
50
9-0
12

Fensulfothion
65
7
50
79-51
12

Parathion
methyl
85
24
100
133-37
12

Phorate
10
15
50
41-0
12

Disulfoton
2
1
50
4-0
12

Merphos
101
13
50
126- 75
12
Medium conc.
sediment
Dimethoate
74
8.5
2
91-57
15
(mg/kg)
Dichlorvos
166
25
2
216-115
15

Naled
ND

Fensulfothion
72
8.6
2
90-55
15

Parathion
methyl
84
9
3
102-66
15

Phorate
58
6
2
70-46
15

Disulfoton
56
5
2
66-47
15

Merphos
78
4
2
66-70
12
Data from Reference 17.
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TABLE 11
SINGLE OPERATOR ACCURACY AND PRECISION FOR TRIS-BP IN
MUNICIPAL WASTE WATER, DRINKING WATER, CHEMICAL SLUDGE
Compound
Matrix
Mean
Rec. (%) Std. Dev.
Spike
Cone.
Recovery
Range (%)
#
Analyses
Tris-BP
Municipal
wastewater
25 8.0
2
41 - 9.0
15

Drinking
water
40 5.0
2
50-30
12

Chemical
sludge
63 11
100
84-42
8
Data from Reference 18.
8321B - 35
Revision 2
January 1998
-------
TABLE 12
SINGLE LABORATORY OPERATOR ACCURACY AND PRECISION
FOR THE CHLORINATED PHENOXYACiD HERBICIDES
Compound
Mean
Recovery %
Std. Dev.
Spike Cone.
Recovery
Range (%)
#
Analyse
LOW LEVEL DRINKING WATER

pg/L

Dicamba
63
22
5
86-33
9
2,4-D
26
13
5
37- 0
9
MCPA
60
23
5
92-37
9
MCPP
78
21
5
116-54
9
Dichlorprop
43
18
5
61-0
9
2,4,5-T
72
31
5
138-43
9
Silvex
62
14
5
88-46
9
2,4-DB
29
24
5
62- 0
9
Dinoseb
73
11
5
85-49
9
Dalapon
ND
ND
5
ND
9
2,4-D,ester
73
17
5
104-48
9
HIGH LEVEL DRINKING WATER

Dicamba
54
30
50
103-26
9
2,4-D
60
35
50
119-35
9
MCPA
67
41
50
128-32
9
MCPP
66
33
50
122-35
9
Dichlorprop
66
33
50
116-27
9
2,4,5-T
61
23
50
99-44
9
Silvex
74
35
50
132 -45
9
2,4-DB
83
25
50
120 - 52
9
Dinoseb
91
10
50
102-76
9
Dalapon
43
9.6
50
56-31
9
2,4-D,ester
97
19
50
130-76
9
8321B - 36
Revision 2
January 1998
-------
TABLE 12
(continued)
Mean Recovery #
Compound
Recovery %
Std. Dev.
Spike Cone.
Range (%)
Analyses
LOW LEVEL SAND

Mg/g

Dicamba
117
26
0.1
147 - 82
10
2,4-D
147
23
0.1
180-118
*10
MCPA
167
79
0.1
280 - 78
10
MCPP
142
39
0.1
192 - 81
10
Dichiorprop
ND
ND
0.1
ND
10
2,4,5-T
134
27
0.1
171.-99
10
Silvex
121
23
0.1
154-85
10
2,4-DB
199
86
0.1
245- 0
10
Dinoseb
76
74
0.1
210-6
10
Dalapon
ND
ND
0.1
ND
10
2,4-D,ester
180
58
0.1
239 - 59
7
HIGH LEVEL SAND

Mg/g

Dicamba
153
33
1
209-119
9
2,4-D
218
27
1
276-187
9
MCPA
143
30
1
205-111
9
MCPP
158
34
1
226-115
9
Dichiorprop
92
37
1
161 - 51
9
2,4,5-T
160
29
1
204 -131
9
Siivex
176
34
1
225 -141
9
2,4-DB
145
22
1
192--110
9
Dinoseb
114
28
1
140-65
9
Dalapon
287
86
1
418-166
9
2,4-D,ester
20
3.6
1
25-17
7
83210-37
Revision 2
January 1998
-------
TABLE 12
(continued)
Compound
Mean
Recovery %
Std. Dev.
Spike Cone.
Recovery
Range(%)
#
Analyses
LOW LEVEL MUNICIPAL ASH

Mg/g

Dicamba
83
22
0.1
104-48
9
2,4-D
ND
ND
0.1
ND
9
MCPA
ND
ND
0.1
ND
9
MCPP
ND
ND
0.1
ND
9
Dichlorprop
ND
ND
0.1
ND
9
2,4,5-T
27
25
0.1
60- 0
9
Siivex
68
38
0.1
128 - 22
9
2,4-DB
ND
ND
0.1
ND
9
Dinoseb
44
13
0.1
65-26
9
Dalapon
ND
ND
0.1
ND
9
2,4-D,ester
29
23
0.1
53-0
6
HIGH LEVEL MUNICIPAL ASH

Mg/g

Dicamba
66
21
1
96-41
9
2,4-D
8.7
4.8
1
21 - 5
9
MCPA
3,2
4.8
1
10- 0
9
MCPP
10
4.3

16-4.7
9
Dichlorprop
ND
ND
1
ND
9
2,4,5-T
2.9
1.2
1
3.6- 0
9
Siivex
6.0
3.1
1
12-2.8
9
2,4-DB
ND
ND
1
-ND
9
Dinoseb
16
6.8
1
23-0
9
Dalapon
ND
ND
1
ND
9
2,4-D,ester
1.9
1.7
1
6.7-0
6
Source: Reference 19,
All recoveries are in negative ionization mode, except for 2,4-D, ester.
ND = Not Detected,
8321B-38
Revision 2
January 1998
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TABLE 13
MULTI-LABORATORY ACCURACY AND PRECISION DATA
FOR THE CHLORINATED PHENOXYACID HERBICIDES
Compound Spiking Concentration Mean (% Recovery)* RSP"
2,4,5-T 500 mg/L 90 23
2,4,5-T.butoxy ester 90 29
2,4-D 86 17
2,4-DB 95 22
Dalapon 83 13
Dicamba 77 - 25
Dichlorprop 84 20
Dinoseb 78 15
MCPA 89 11
MCPP 86 12
Silvex 96 27
2,4,5-T 50 mg/L 62 68
2,4,5-T,butoxy ester 85 9
2,4-D 64 80
2,4-DB 104 28
Dalapon 121 99
Dicamba 90 23
Dichlorprop 96 15
Dinoseb 86 57
MCPA 96 20
MCPP 76 74
Silvex 65 71
8321B - 39
Revision 2
January 1998
-------

TABLE 13
(continued)

Compound
Spiking Concentration
Mean (% Recovery)*
RSD"
2,4,5-T
5 mg/L
90

28
2,4,5-T,butoxy ester

17
2,4-D

103

31
2,4-DB

OA
9V

" 21
Datapon

150

4
Dicamba

105

12
Dichlorprop

102

22
Dinoseb

108

30
MCPA

18
MCPP

15
Silvex

15
a Mean of duplicate data from 3 laboratories,
b Relative standard deviation of duplicate data from 3 laboratories.
Data from Reference 20.
8321B - 40
Revision 2
January 1998
-------
TABLE 14
SINGLE-LABORATORY EVALUATION OF AVERAGE RECOVERY
AND PRECISION DATA FOR WATER0
Analyte
Average % Recovery"
Standard Deviation
%RSD
Aldicarb sulfoxide
7.6
2.8
37.0
Aldicarb sulfone
56.0
27.1
48.5
OxamyP
38.9
17.9
45.9
Methomyi
52.0
19.6
37.7
3-Hydroxycarbofuran8
22.2
9.3
41.7
Fenuron
72.5
22.0
30.3
Benomyl/Carbendazim
47.3
14.7
31.0
Aldicarb
81.0
13.7 "
16.9
Aminocarb
109
38.3
35.1
Carbofuran
85.5
10.0
11.7
Propoxur
79.1
13.7
17.3
Monuron
91.8
11.3
12.3
Bromadl
87.6
12.1
13.8
Tebuthiuron
87.1
9.0
10.3
Carbaryl
82.1
13.5
16.5
Fluometuron
84.4
8.3
9.8
Propham
80.7
13.8
17.1
Propachlor
84.3
10.0
11.9
Diuron
90.8
14.1
15.6
Siduron
88.0
9.5
10.8
Methiocarb
93.3
12.8
13.8
Barban
88.1
11.2
12.7
Linuron
87.1
16.8
19.3
Chloropropham
94.9
15.3
16.1
Mexacarbate
79.8
12.9
16.2
Chloroxuron
106
24.9
23.5
Neburon
85.3
12.6
14.8
a Values generated from internal response factor calculations.
b Nine spikes were performed at three concentrations. The concentrations for Aldicarb sulfoxide,
Barban, Chloropropham, and Mexacarbate spike levels were at 25 pg/L, 50 pg/L, and 100 pg/L
All other analyte concentrations were 5 pg/L, 10 pg/L, and 20 pg/L. One injection was
disregarded as an outlier. The total number of spikes analyzed was 26.
c Data from Reference 22.
8321B-41
Revision 2
January 1998
-------
TABLE 15
SINGLE-LABORATORY EVALUATION OF AVERAGE RECOVERY
AND PRECISION DATA FOR SOIL"
Analyte
Average % Recovery
Standard Deviation
%RSD
Aldicarb sulfoxide
66.9
31.3
46.7
Aldicarb sulfone
162
51.4
31.7
Oxamyl
78.9
46.1
58.5
Methomyl
84.9
25.8
30.4
3-Hydroxycarbofuran
105
36.3
34.5
Fenuron
91.9
16.7
18.1
Benomyl/Carbendazim
95.6
18.2
19.0
Aldicarb
97.9
17.0
17.4
Aminocarb
133
44.7
33.6
Carbofuran
109
14.4
13.2
Propoxur
104
16.5
15.9
Monuron
101
12.4
12.3
Bromacil
100
9.0
9.0
Tebuthiuron
104
11.9
11.5
Carbaryl
102
15.5
15.2
Fluometuron
94.5
15.7
16.7
Propham
92.8
12.0
12.9
Propachlor
94.6
10.3
10.9
Diuron
107
17.4
16.2
Siduron
100
12.0
12.0
Methiocarb
107
14.2
13.2
Barban
92.3
15.6
16.9
Linuron
104
13.6
13.1
Chloropropham
105
9.3
8.9
Mexacarbate
77.2
9.8
12.7
Chloroxuron
121
27.3
22.5
Neburon
92.1
16.5
17.9
a Nine spikes were performed at three concentrations. The concentrations for Aldicarb sulfoxide,
Barban, Chloropropham, and Mexacarbate spike levels were at 0.625 pg/g, 1.25 jjg/g, and 2.5
pg/g. All other analyte concentrations were 0.125 pg/g, 0.25 pg/g, and 0.50 pg/g. One injection
was disregarded as an outlier. Hie total number of spikes analyzed was 26.
b Data from Reference 22.
8321B - 42
Revision 2
January 1998
-------
TABLE 16
MULTI-LABORATORY EVALUATION OF METHOD ACCURACY
(AFTER OUTUER REMOVAL)*
Analyte

Percent Recovery

High-Concentration
Samples"
Medium-Concentration
Samples"
Low-Concentration
Samples"
Aldicarb
98.7
110
52.0
Bendiocarb
81.4
95.0
52.0
Carbaryt
92.0
108
62.0
Carbendazim
125
138
128
Carbofuran
87.8
92.3
72.0
Diuron
79.9
98.8
66.0
Linuron
84.8
93.0
82.0
Methomyl
93.3
90.8
90.0
Oxamyl
83.8
88.0
98.0
a Three replicates per laboratory; eight to nine laboratories (per Table 26 of Reference 23). The true
concentration is 90 mg/L per compound, except Carbendazim at 22.5 mg/L.
b Two replicates per laboratory; eight to nine laboratories (per Table 26 of Reference 23). The true
concentration is 40 mg/L per compound except Carbendazim at 10 mg/L.
c Three replicates per laboratory; eight to nine laboratories (per Table 26 of Reference 23). The true
concentration is 5 mg/L per compound, except Carbendazim at 1,25 mg/L
6 Data from Reference 23.
8321B - 43
Revision 2
January 1998
-------
TABLE 17
MULTI-LABORATORY EVALUATION OF METHOD PRECISION (AFTER OUTLIER REMOVAL)'
High Concentration Medium Concentration Low Concentration
Analyte
Avg.
Sr
SR
%RSDr
%RSDr
Avg.
Sr
sr
%RSDr
%RSDr
Avg
Sr
Sr
%RSDr
%RSDr
Aldicarb
88.8
11.4
34.4
12.9
38.8
44.1
7.7
17.0
17.5
38.5
2.6
0.9
2.6
33.1
98.2
Bendiocarb
73.3
16.1
39.3
21.9
53.6
38.0
6.6
16.6
17.3
43.7
2.6
0.6
1.6
21.3
61.9
Carbaryl
82.8
11.7
34.0
14.2
41.1
43.1
3.0
15.7
7.0
36.4
3.1
0.7
2.3
23.3
75.8
Carbendazim
28.1
5.6
15.3
19.9
54.4
13.8
1.4
8.9
10.4
64.2
1.6
0.4
1.1
26.1
68.2
Carbofuran
79.0
16.7
35.2
21.2
44.5
36.9
5.0
16.3
13.6
44.3
3.6
0.9
3.3
25.2
91.6
Diuron
71.9
13.1
26.1
18.2
36.3
39.5
2.6
11.8
6.5
29.8
3.3
0.5
2.6
16.2
77.9
Linuron
76.3
8.3
32.5
10.9
42.6
37.2
3.9
13.4
10.5
35.9
4.1
0.6
2.1
15.7
51.4
Methomyl
84.0
10.8
29.4
12.9
35.0
36.3
2.8
15.0
7.8
41.2
4.5
0.7
4.1
15.3
92.9
Oxamyl
75.5
12.4
37.0
16.4
49.1
35.2
3.7
20.8
10.4
59.1
4.9
0.5
4.6
9.7
93.6
Average

16.5
43.9

11.2
43.7

20.7
79.1
Std. Dev.

4.0
I
7.1

4.1
11.2

7.1
16.3
sr and Sp are the standard deviations for repeatability and reproducibility, respectively. RSDr and RSDr are the corresponding relative standard deviations
for repeatability and reproducibility, respectively. The units for average, sr and sR are mg/L.
8 Data from Reference 23.
8321B - 44
Revision 2
January 1998
-------
TABLE 18
SINGLE LABORATORY RECOVERY DATA FOR SOLID-PHASE EXTRACTION OF
CHLORINATED HERBICIDES FROM SPIKED TCLP BUFFERS
Compound
Buffer 1
Buffer 2
Spike Level (pg/L) Recovery (%)
RSD
Recovery (%)
RSD
2,4-D
2,4,5-TP
5,000
500
91
93
2
9
79
92*
6
2*
2,4-D
2,4,5-TP
20,000
2000
100
103*
3
2*
99*
78
1*
7
Except where noted with an asterisk, all results are from seven replicates. Those marked with an asterisk are from
three replicates.
Data are from Reference 24,
8321B - 45
Revision 2
January 1998
-------
FIGURE 1
SCHEMATIC OF THE THERMOSPRAY PROBE AND ION SOURCE
Rang*
Ion Sampling
Con*
Sourea
Mounting
Wat*
Electron Vaporizer
•mik Probo
Mechanical
Sourea
Block
Vaponitr
CofltroUr
Vapor || Haatar Vaporizer
Temperature | Coupling
Block
Tamparature
T-
8321B -46
Revision 2
January 1998
-------
FIGURE 2
THERMOSPRAY SOURCE WITH WIRE-REPiLLER
(High sensitivity configuration)

r
¦ V
1

\
CERAMIC INSULATOR
WIRE REPELLER

83218 - 47
Revision 2
January 1998
-------
FIGURE 3
THERMOSPRAY SOURCE WITH WiRE-REPELLER
(CAD configuration)

\
CERAMIC INSULATOR
* WIRE REPELLER

83215 - 48
Revision 2
January 1998
-------
METHOD 8321B
SOLVENT-EXTRACTABLE NONVOLATILE COMPOUNDS BY
HIGH PERFORMANCE LIQUID CHROMATOGRAPHY/THERMOSPRAY/MASS
SPECTROMETRY (HPLCfTSIMS) OR ULTRAVIOLET fUV) DETECTION
Is sampis
analyzed for
Tns-8P?
7.1 Prepare sample
for •xtrietinn.
Yes
7.2 Prepiir* sample
for Tfla-BP
microextracti on,
is
•ample high"
concentration
waati?
7.2.1.1 weigh out a
19 •ample end
•pike the s«l«6t«d
•ample withTris-BP
Um dilution
Method 3580.
7.2.2.1 Spike
The selected sample
with Tris-BP
Is sample
aqueous?
7.2.2.2 Add
CH jCI 2» sea
and shake
three times
7.2.1.1 Add
equivalent amount
of anhydrous
I*
•ample
moist?
Chlorinated Phinoxyacid
Compound*
Typa
of
Analyte
7.2,2.3
Allow organic
and water
layers to separate
7.3 Use modified
Method 8151
Carbamate
Pesticides
7.2.1.3 Pack
•ample in pipet
is
•ample solid
or aqueous?
7.4 Use appropriate
3500 series method
on 40 f sampi
Solid
7.2.1.4
Extract sampl
first with CH3OH
followed by
CH3OH/CH 2CU
7,2.1.S
Reduce volume
by K*D or
evaporation
ISec, 7,51.
7.2.2.4
Collect the
extract
Aqueous
7.4 Use appropriate
3500 sariee method!
on 1 L sample
7.5 Concentrate
•ample and
exchange
extraction solvent
to methanol or
ecetonitrile during
K-D procedures.
7.8 Select and
implement instrument
calibration
procedure
7.9 Sample
analysis
using selected
LC detector
7.6 Set HPie
Chromatographic
condition*.
7.7 Set HPLC/T5/M5
operating
conditions
7.10 Use
Method 3000 »o
calculate analyte(s)
concentration
8321B-49
Revision 2
January 1998
-------
METHOD 8330A
NITROAROMATICS AND NITRAMINES BY HIGH
PERFORMANCE LIQUID CHROMATOGRAPHY MPLO
1.0 SCOPE AND APPLICATION
1.1 Method 8330 is intended for the trace analysis of explosives residues by high
performance liquid chromatography using a UV detector. This method is used to determine the
concentration of the following compounds in a water, soil, or sediment matrix:
Anatyte
Abbreviation
CAS Number
Octahydro-1,3,5,7-tetranitro-l ,3,5,7-tetrazocine
HMX
2691-41-0
Hexahydro-1,3,5-trinitro-1,3,5-triazine
RDX
121-82-4
1,3,5-Trinitrobenzene
1,3,5-TNB
99-35-4
1,3-Dinitrobenzene
1,3-DNB
99-65-0
Methyl-2,4,6-trinitrophenylnitramine
Tetryl
479-45-8
Nitrobenzene
NB
98-95-3
2,4,6-T rinitrotoluene
2,4,6-TNT
118-96-7
4-Amino-2,6-dinitrotoluene
4-Am-DNT
1946-51-0
2-Amino-4, 6-dinitrotoluene
2-Am-DNT
35572-78-2
2,4-Dinitrotoluene
2,4-DNT
121-14-2
2,6-Dinitrotoluene
2,6-DNT
606-20-2
2-Nitrotoluene
2-NT
88-72-2
3-Nitrotoluene
3-NT
99-08-1
4-Nitrotoluene
4-NT
99-99-0
1.2 Method 8330 provides a salting-out extraction procedure for low concentrations (parts
per trillion, or ng/L) of explosives residues in surface or ground water. Direct injection of diluted and
filtered water samples can be used for water samples of higher concentration (See Table 1). Solid-
phase extraction, using Method 3535, may also be applied to aqueous samples.
1.3 All of these compounds are either used in the manufacture of explosives or are the
degradation products of compounds used for that purpose. When making stock solutions for
calibration, treat each explosive compound with caution. See NOTE in Sec. 5.3.1 and Sec. 11.
1A The estimated quantitation limits (EQLs) of target analytes determined by Method 8330
in water and soil are presented in Table 1.
8330A-1
Revision 1
January 1998
-------
1.5 This method is restricted to use by or under the supervision of analysts experienced
in the use of HPLC, skilled in the interpretation of chromatograms, and experienced in handling
explosive materials. (See Sec. 11.0 on SAFETY.) Each analyst must demonstrate the ability to
generate acceptable results with this method.
2.0 SUMMARY OF METHOD
2.1 Method 8330 provides high performance liquid chromatographic (HPLC) conditions for
the detection of ppb levels of certain explosive residues in water, soil, and sediment. Prior to use
of this method, appropriate sample preparation techniques must be used.
2.2 Low-level salting-out method with no evaporation - Aqueous samples of low
concentration are extracted by a salting-out extraction procedure with acetonitrile and sodium
chloride. The small volume of acetonitrile that remains undissolved above the salt water Is drawn
off and transferred to a smaller volumetric flask. It is back-extracted by vigorous stirring with a
specific volume of salt water. After equilibration, the phases are allowed to separate and the small
volume of acetonitrile residing in the narrow neck of the volumetric flask is removed using a Pasteur
pipet. The concentrated extract is diluted 1:1 with reagent grade water. An aliquot is separated on
a C-18 reversed-phase column, determined at 254 nm, and confirmed on a CN reversed-phase
column.
2.3 Solid-phase extraction method - Aqueous samples may also be prepared using solid-
phase extraction, as described in Method 3535.
2.4 High-level direct injection method - Aqueous samples of higher concentration can be
diluted 1/1 (v/v) with methanol or acetonitrile, filtered, separated on a C-18 reversed-phase column,
determined at 254 nm, and confirmed on a CN reversed-phase column. If HMX is an important
target analyte, methanol is preferred.
2.5 Soil and sediment samples are extracted using acetonitrile in an ultrasonic bath, filtered
and analyzed as described in Sec. 2.3.
3.0 INTERFERENCES
3.1 Solvents, reagents, glassware and other sample processing hardware may yield
discrete artifacts and/or elevated baselines, causing misinterpretation of the chromatograms. All of
these materials must be demonstrated to be free from interferences.
3.2 2,4-DNT and 2,6-DNT elute at similar retention times (retention time difference of 0.2
minutes). A large concentration of one isomer may mask the response of the other isomer. If it is
not apparent that both isomers are present (or are not detected), an isomeric mixture should be
reported.
3.3 Tetryl decomposes rapidly in methanol/water solutions, as well as with heat. All
aqueous samples expected to contain tetryl should be diluted with acetonitrile prior to filtration and
acidified to pH <3. All samples expected to contain tetryl should not be exposed to temperatures
above room temperature.
8330A-2
Revision 1
January 1998
-------
3.4 Degradation products of tetryl appear as a shoulder on the 2,4,6-TNT peak. Peak
heights rather than peak areas should be used when tetryl is present in concentrations that are
significant relative to the concentration of 2,4,6-TNT.
4.0 APPARATUS AND MATERIALS
4.1 HPLC system
4.1.1 HPLC - equipped with a pump capable of achieving 4000 psi, a 100-pL
loop injector and a 254-nm UV detector (Perkirt-Elmer Series 3, or equivalent). . For the low
concentration option, the detector must be capable of maintaining a stable baseline at 0.001
absorbance units full scale.
4.1.2 Recommended columns
4.1.2.1 Primary column -C-18 Reversed-phase HPLC column, 25-cm x
4.6-mm (5 |jm) (Supelco LC-18, or equivalent).
4.1.2.2 Secondary column - CN Reversed-phase HPLC column, 2S-cm
x 4.6-mm (5 ^m) (Supelco LC-CN, or equivalent).
4.1.3 Strip chart recorder
4.1.4 Digital integrator (optional)
4.1.5 Autosampler (optional)
4.2 Other equipment
4.2.1 Temperature-controlled ultrasonic bath
4.2.2 Vortex mixer
4.2.3 Balance - capable of weighing ± 0.0001 g
4.2.4 Magnetic stirrer with PTFE stirring bars
4.2.5 Water bath - Heated, with concentric ring cover, capable of temperature
control (± 5°C). The bath should be used in a hood.
4.2.6 Oven - Forced air, without heating.
4.3 Materials
4.3.1 High-pressure injection syringe - 500-pL (Hamilton liquid syringe, or
equivalent).
4.3.2 Disposable cartridge Alters - 0.45-ym PTFE Alter.
4.3.3 Pipets - Class A, glass, appropriate sizes.
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4.3.4
Pasteur pipets
4.3.5 Scintillation vials - 20-mL, glass.
4.3.6 Vials - 15-mL, glass, PTFE-lined cap.
4.3.7 Vials - 40-mL, glass, PTFE-lined cap.
4.3.8 Disposable syringes - Plastipak, 3-mL and 10-mL or equivalent.
4.3.9 Volumetric flasks - 10-mL, 25-mL, 10Q-mL, and 1-L, with ground-glass
stoppers, Class A.
NQS: The 100-mL and 1-L volumetric flasks used for magnetic stirrer extraction must be
round.
4.3.10 Vacuum desiccator-Glass.
4.3.11 Mortar and pestle - Steel.
4.3.12 Sieve - 30-mesh.
4.3.13 Graduated cylinders - 10-mL, 25-mL, and 1-L.
5.0 REAGENTS
5.1 Reagent grade inorganic chemicals shall be used in all tests. Unless otherwise
indicated, it is intended that all reagents shall conform to the specifications of the Committee on
Analytical Reagents of the American Chemical Society, where such specifications are available.
Other grades may be used, provided it is first ascertained that the reagent is of sufficiently high purity
to permit its use without towering the accuracy of the determination.
5.1.1 Acetonitrile, CH3CN - HPLC grade.
5.1.2 Methanol, CH3OH - HPLC grade.
5.1.3 Calcium chloride, CaCI2 - Reagent grade. Prepare an aqueous solution
containing 5 g/L of calcium chloride.
5.1.4 Sodium chloride, NaCI, shipped in glass bottles - reagent grade.
5.2 Organic-free reagent water - All references to water in this method refer to organic-free
reagent water, as defined in Chapter One.
5.3 Stock standard solutions
Dry each solid analyte standard to constant weight in a vacuum desiccator in the dark. Place
about 0.100 g (weighed to 0.0001 g) of a single analyte into a 100-mL volumetric flask and dilute to
volume with acetonitrile. Invert flask several times until dissolved. Store in refrigerator at 4°C in the
dark. Calculate the concentration of the stock solution from the actual weight used (nominal
concentration - 1,000 mg/L). Stock solutions may be used for up to one year.
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NOTE: The HMX, RDX, Tetryl, and 2,4,6-TNT are explosives and the neat material should be
handled carefully. See SAFETY in Sec. 11 for guidance, HMX, RDX, and Tetryl reference
materials are shipped under water. Drying at ambient temperature requires several days.
DO NOT DRY AT ELEVATED TEMPERATURES!
5.4 Intermediate standards solutions
5.4.1 if both 2,4-DNT and 2,6-DNT are to be determined, prepare two separate
intermediate stock solutions containing (1) HMX, RDX, 1,3,5-TNB, 1,3-DNB, NB, 2,4,6-TNT,
and 2,4-DNT and (2) Tetryl, 2,6-DNT, 2-NT, 3-NT, and 4-NT. Intermediate stock standard
solutions should be prepared at 1,000 pg/L, in acetonitrite when analyzing soil samples, and
in methanol when analyzing aqueous samples.
5.4.2 Dilute the two concentrated intermediate stock solutions, with the appropriate
solvent, to prepare intermediate standard solutions that cover the range of 2.5 -1,000 pg/L
These solutions should be refrigerated on preparation, and may be used for 30 days.
5.4.3 For the low-level method, the analyst must conduct a detection limit study and
devise dilution series appropriate to the desired range. Standards for the low level method
must be prepared immediately prior to use.
5.5 Working standards - Calibration standards at a minimum of five concentration levels
should be prepared by the dilution of the intermediate standards solutions by 50% (v/v) with 5 g/L
calcium chloride solution (Sec. 5,1.3). These solutions must be refrigerated and stored in the dark,
and prepared fresh on the day of calibration.
5.6 Surrogate spiking solution - The analyst should monitor the performance of the
extraction and analytical system as well as the effectiveness of the method in dealing with each
sample matrix by spiking each sample, standard and reagent water blank with one or two surrogates
(e.g., analytes not expected to be present in the sample).
5.7 Matrix spiking solutions - Prepare matrix spiking solutions in methanol such that the
concentration in the sample is five times the Estimated Quantitation Limit (Table 1). Ml target
analytes should be included.
5.8 HPLC mobile phase - To prepare 1 Lof mobile phase, add 500 mL of methanol to 500
mL of organic-free reagent water.
6.0 SAMPLE COLLECTION, PRESERVATION, AND HANDLING
6.1 Follow conventional sampling and sample handling procedures as specified for
semivolatile organics in Chapter Four.
6.2 Samples and sample extracts must be stored in the dark at 4°C. Holding times are the
same as for semivolatile organics.
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7.0
PROCEDURE
7.1 Sample preparation
This method addresses both aqueous and solid samples. There are three extraction
procedures that may be applied to aqueous samples, depending on the expected level of
explosive residue in the sample and the available equipment: a low-level salting-out extraction,
a high-level extraction, and solid-phase extraction. It is highly recommended that aqueous
process waste samples be screened with the high-level method to determine If the low-level
method (1-50 nfl/L) is required. Most groundwater samples will fall into the low-level method.
7.1.1 Aqueous low-level method (salting-out extraction)
7.1.1.1 Add 251.3 g of sodium chloride to a 1-L volumetric flask (round).
Measure 770 mL of a water sample (using a 1-L graduated cylinder) and transfer it to
the volumetric flask containing the salt. Add a stir bar and mix the contents at
maximum speed on a magnetic stirrer until the salt is completely dissolved.
7.1.1.2 Add 164 mL of acetonltrile (measured with a 250-mL graduated
cylinder) while the solution is being stirred and stir for an additional 15 minutes. Turn
off the stirrer and allow the phases to separate for 10 minutes.
7.1.1.3 Remove the acetonitrile (upper) layer (about 8 mL) with a Pasteur
pipet and transfer it to a 100-mL volumetric flask (with a round bottom). Add 10 mL of
fresh acetonitrile to the water sample in the 1-L flask. Again stir the contents of the
flask for 15 minutes followed by 10 minutes for phase separation. Combine the second
acetonitrile portion with the initial extract. The inclusion of a few drops of salt water at
this point is unimportant.
7.1.1.4 Add 84 mL of salt water (325 g NaCI per 1000 mL of reagent
water) to the acetonitrile extract in the 100-mL volumetric flask. Add a stir bar and stir
the contents on a magnetic stirrer for 15 minutes, followed by 10 minutes for phase
separation. Carefully transfer the acetonitrile phase to a 10-mL graduated cylinder
using a Pasteur pipet. At this stage, the amount of water transferred with the
acetonitrile must be minimized. The water contains a high concentration of NaCI that
produces a large peak at the beginning of the chromatogram, where it could interfere
with the HMX determination.
7.1.1.5 Add an additional 1.0 mL of acetonitrile to the 100-mL volumetric
flask. Again stir the contents of the flask for 15 minutes, followed by 10 minutes for
phase separation. Combine the second acetonitrile portion with the initial extract in the
10-mL graduated cylinder (transfer to a 25-mL graduated cylinder if the volume
exceeds 5 mL). Record the total volume of acetonitrile extract to the nearest 0.1 mL.
(Use this as the volume of total extract [VJ in the calculation of concentration after
converting to nL). The resulting extract, about 5-6 mL, is then diluted 1:1 with
organiofree reagent water (with pH <3 if tetryl is a suspected analyte) prior to analysis.
7.1.1.6 If the diluted extract is turbid, filter it through a 0.45-^m PTFE filter
using a disposable syringe. Discard the first 0.5 mL of filtrate, and retain the remainder
in a PTFE-capped vial for RP-HPLC analysis in Sec. 7.4.
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7.1.2 Aqueous high-level method
7.1.2.1 Sample filtration
Place a 5-mL aliquot of each water sample in a scintillation vial, add 5 mL of
acetonitrile, shake thoroughly, and filter through a 0.45-pm PTFE filter using a
disposable syringe.
7.1.2.2 Discard the first 3 mL of filtrate, and retain the remainder in a
PTFE-capped vial for RP-HPLC analysis in Sec. 7.4. HMX quantitation can be
improved with the use of methanol rather than acetonitrile for dilution before filtration.
7.1.3 Solid-phase extraction
Aqueous samples containing nitroaromatics and nitramines may also be extracted
using solid-phase extraction (SPE) in both disk and cartridge formats. Consult Method 3535
for the procedures to be employed and the apparatus and materials that are required.
7.1.4 Soil and sediment samples
7.1.4.1 Sample homogenization
Pry soil samples in air at room temperature (or less) to a constant weight,
being careful not to expose the samples to direct sunlight. Grind and homogenize the
dried sample thoroughly in an aceton'itrile-rinsed mortar to pass a 30-mesh sieve.
NOTE: Soil samples should be screened by Method 8515 prior to grinding in a mortar
and pestle (See Safety Sec. 11.2).
7.1.4.2 Sample extraction
7.1.4.2.1 Place a 2.0-g subsample of each soil sample in a 15-
mL glass vial. Add 10.0 mL of acetonitrile, cap with PTFE-lined cap, vortex
swirl for one minute, and place in a cooled ultrasonic bath for 18 hours.
7.1.4.2.2 After sonication, allow sample to settle for 30
minutes. Remove 5.0 mL of supernatant, and combine with 5.0 mL of
calcium chloride solution (Sec. 5.1.3) in a 20-mL vial. Shake, and let stand
for 15 minutes.
7.1.4.2.3 Place supernatant in a disposable syringe and filter
through a 0.45-jjm PTFE filter. Discard first 3 mL and retain remainder in a
PTFE-capped vial for RP-HPLC analysis in Sec. 7.4.
7.2 Chromatographic conditions (recommended)
Primary Column: C-18 reversed-phase HPLC column, 25-cm x
4,6-mm, 5 Mm (Supelco LC-18 or equivalent).
Secondary Column: CN reversed-phase HPLC column, 25-cm x 4.6-mm,
5 Mm (Supelco LC-CN or equivalent).
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Mobile Phase:
50/50 (v/v) methanol/organic-fnee reagent water.
1.5 mL/min
Flow Rate:
Injection volume: 100-pL
UV Detector 254 nm
254 nm
7.3 Calibration of HPLC
7.3.1 All electronic equipment is allowed to warm up for 30 minutes. During this
period, at least 15 void volumes of mobile phase are passed through the column
(approximately 20 min at 1.5 mL/min) and continued until the baseline is level at the UV
detector's greatest sensitivity.
7.3.2 Initial calibration - Injections of each calibration standard over the
concentration range of interest are made sequentially into the HPLG in random order. Peak
heights or peak areas are obtained for each analyte. Employ one of the calibration options
described in Method 8000.
7.3.3 Calibration verification - Analyze one mid-point calibration standard, at a
minimum, at the beginning of the day, and after every 20 sample extracts (recommended after
every 10, in order to minimize the number of samples that may be affected by a failing
standard), and after the last sample of the day. Calculate the calibration factor for each analyte
from the peak height or peak area and compare it with the mean calibration factor obtained for
the initial calibration, as described in Method 8000. The calibration factor for the calibration
verification must agree within ±15% of the mean calibration factor of the initial calibration. If
this criterion is not met, a new initial calibration must be performed, or another of the
calibration options described in Method 8000 must be employed.
7.4 HPLC analysis
7.4.1 Analyze the samples using the chromatographic conditions given in Sec. 7.2.
All positive measurements observed on the C-18 column must be confirmed by injection onto
the CN column.
7.4.2 Method 8000 provides instructions on the analysis sequence, appropriate
dilutions, establishing daily retention time windows, and identification criteria. Include a
mid-level standard after each group of 20 samples in the analysis sequence, if column
temperature control is not employed, special care must be taken to ensure that temperature
shifts do not cause peak misidentification.
7.4.3 Table 2 summarizes the estimated retention times on both C-18 and CN
columns for a number of analytes anatyzable using this method. An example of the separation
achieved by Column 1 is shown in Figure 1.
7.4.4 Record the resulting peak sizes in peak heights or area units. The use of
peak heights is recommended to improve reproducibility of low level samples.
7.4.5 The calculation of sample concentrations is described in Method 8000.
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8,0 QUALITY CONTROL
8.1 Refer to Chapter One arid Method 8000 for specific quality control (QC) procedures.
Quality control procedures to ensure the proper operation of the various sample preparation and/or
sample introduction techniques can be found in Method 3500. Each laboratory should maintain a
formal quality assurance program. The laboratory should also maintain records to document the
quality of the data generated.
8.2 Quality control procedures that are necessary to validate the HPLC system operation
are found in Method 8000, Sec. 8.0.
8.3 Initial demonstration of proficiency
Each laboratory must demonstrate initial proficiency with each sample preparation and
determinative method combination it utilizes, by generating data of acceptable accuracy and
precision for target analytes in a clean matrix. The laboratory must also repeat the following
operations whenever new staff are trained or significant changes in instrumentation are made. See
Method 8000, Sec. 8.0 for information on how to accomplish this demonstration.
8.4 Sample quality control for preparation and analysis
The laboratory must also haw procedures for documenting the effect of the matrix on method
performance (precision, accuracy, and quantitation limit). At a minimum, this includes the analysts
of QC samples including a method blank, matrix spike, a duplicate, and a laboratory control sample
(LCS) in each analytical batch and the addition of surrogates to each field sample and QC sample.
8.4.1 Before processing any samples, the analyst should demonstrate, through the
analysis of a method blank, that interferences from the analytical system, glassware, and
reagents are under control. Each time a set of samples is analyzed or there is a change in
reagents, a method blank should be analyzed as a safeguard against chronic laboratory
contamination. The blanks should be earned through all stages of sample preparation and
measurement.
8.4.2 Documenting the effect of the matrix should include the analysis of at least
one matrix spike and one duplicate unspiked sample or one matrix spike/matrix spike duplicate
pair. The decision on whether to prepare and analyze duplicate samples or a matrix
spike/matrix spike duplicate must be based on a knowledge of the samples in the sample
batch. If samples are expected to contain target analytes, then laboratories may use one
matrix spike and a duplicate analysis of an unspiked field sample. If samples are not expected
to contain target analytes, laboratories should use a matrix spike and matrix spike duplicate
pair.
8.4.3 A Laboratory Control Sample (LCS) should be included with each analytical
batch. The LCS consists of an aliquot of a clean (control) matrix similar to the sample matrix
and of the same weight or volume. The LCS is spiked with the same analytes at the same
concentrations as the matrix spike. When the results of the matrix spike analysis indicate a
potential problem due to the sample matrix itself, the LCS results are used to verify that the
laboratory can perform the analysis in a clean matrix.
8.4.4 See Method 8000, Sec. 8.0, for the details on carrying out sample quality
control procedures for preparation and analysis.
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B.5 Surrogate recoveries
The laboratory must evaluate surrogate recovery data from individual samples versus the
surrogate control limits developed by the laboratory. See Method 8000, Sec. 8.0, for information on
evaluating surrogate data and developing and updating surrogate limits.
8.6 It is recommended that the laboratory adopt additional quality assurance practices for
use with this method. The specific practices that are most productive depend upon the needs of the
laboratory and the nature of the samples. Whenever possible, the laboratory should analyze
standard reference materials and participate in relevant performance evaluation studies.
9.0 METHOD PERFORMANCE
9.1 Table 3 provides the single-laboratory precision based on data from the analysis of
blind duplicates of four spiked soil samples and four field-contaminated samples analyzed by seven
laboratories.
9.2 Table 4 provides the multi-laboratory error based on data from the analysis of blind
duplicates of four spiked soil samples and four field-contaminated samples analyzed by seven
laboratories.
9.3 Table 5 provides the multi-laboratory variance of the high-level method for water based
on data from nine laboratories.
9.4 Table 6 provides multi-laboratory recovery data from the analysis of spiked soil samples
by seven laboratories.
9.5 Table 7 provides a comparison of method accuracy for soil and aqueous samples (high-
level method).
9.6 Table 8 provides precision and accuracy data for the salting-out extraction method.
9.7 Table 9 provides data from a comparison of direct injection of groundwater samples
with both the salting-out extraction and the solid-phase extraction techniques.
9.8 Table 10 provides data comparing the precision of duplicate samples analyzed by direct
injection of groundwater samples with both the salting-out extraction and the solid-phase extraction
techniques.
9.9 Table 11 provides a comparison of recovery data for spiked samples analyzed by direct
injection of groundwater samples with both the salting-out extraction and the solid-phase extraction
techniques.
10.0 REFERENCES
1. Bauer, C.F., T.F. Jenkins, S.M. Koza, P.W. Schumacher, P.H. Miyares and M.E. Walsh (1989).
Development of an analytical method for the determination of explosive residues in soil. Part
3. Collaborative test results and final performance evaluation. USACE Cold Regions
Research and Engineering Laboratory, CRREL Report 89-9.
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2. Grant, C.L., A.D. Hewitt and T.F. Jenkins (1989), Comparison of low concentration
measurement capability estimates in trace analysis: Method Detection Limits and Certified
Reporting Limits. USACE Cold Regions Research and Engineering Laboratory, Special Report
89-20.
3. Jenkins, T.F., C.F. Bauer, D.C. Leggett and C.L. Grant (1984). Reverse-phased HPLC method
for analysis of TNT, RDX, HMX and 2,4-DNT in munitions wastewater. USACE Cold Regions
Research and Engineering Laboratory, CRREL Report 84-29.
4. Jenkins, T.F. and M.E. Walsh (1987). Development of an analytical method for explosive
residues in soil. USACE Cold Regions Research and Engineering Laboratory, CRREL Report
87-7.
5. Jenkins, T.F., P.M. Miyares and ME. Walsh (1988a). An improved RP-HPLC method for
determining nitroaromatics and nitramines in water. USACE Cold Regions Research and
Engineering Laboratory, Special Report 88-23.
6. Jenkins, T.F. and P.H. Miyares (1992). Comparison of Cartridge and Membrane Solid-Phase
Extraction with Salting-out Solvent Extraction for Preconcentration of Nitroaromatic and
Nitramine Explosives from Water. USACE Cold Regions Research and Engineering
Laboratory, Draft CRREL Special Report.
7. Jenkins, T.F., P.W. Schumacher, M.E. Walsh and C.F. Bauer (1988b). Development of an
analytical method for the determination of explosive residues in soil. Part II: Further
development and ruggedness testing. USACE Cold Regions Research and Engineering
Laboratory, CRREL Report 88-8.
8. Leggett, D.C., T.F. Jenkins and P.H. Miyares (1990). Salting-out solvent extraction for
preconcentration of neutral polar organic solutes from water. Analytical Chemistry, 62:
1355-1356.
9. Miyares, P.H. and T.F. Jenkins (1990). Satting-out solvent extraction for determining low levels
of nitroaromatics and nitramines in water. USACE Cold Regions Research and Engineering
Laboratory, Special Report 90-30.
10. Jenkins, T. F., Thome, P. G., Myers, K. F., McCormick, E. F., Parker, D. E., and B. L Escalon
(1995). Evaluation of Clean Solid Phases for Extraction of Nitroaromatics and Nitramines from
Water. USACE Cold Regions Research and Engineering Laboratory, Special Report 95-22.
11.0 SAFETY
11.1 Standard precautionary measures used for handling other organic compounds should
be sufficient for the safe handling of the analytes targeted by Method 8330. The only extra caution
that should be taken is when handling the analytical standard neat material for the explosives
themselves and in rare cases where soil or waste samples are highly contaminated with the
explosives. Follow the note for drying the neat materials at ambient temperatures.
11.2 It is advisable to screen soil or waste samples using Method 8515 to determine whether
high concentrations of explosives are present. Soil samples containing as much as 2% of 2,4,6-TNT
have been safely ground. Samples containing higher concentrations should not be ground in the
mortar and pestle. Method 8515 is for 2,4,6-TNT, however, the other nitroaromatics will also cause
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a color to be developed and provide a rough estimation of their concentrations. 2,4,6-TNT is the
analyte most often detected in high concentrations in soil samples. Visual observation of a soil
sample is also important when the sample is taken from a site expected to contain explosives.
Lumps of material that have a chemical appearance should be suspect and not ground. Explosives
are generally a very finely ground grayish-white material.
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TABLE 1
ESTIMATED QUANTITATION LIMITS
Water (m§/L)
Analytes
Low-Level
High-Level
Soil (mg/kg)
HMX
-
13.0
2.2
RDX
0.84
14.0
1.0
1,3,5-TNB
0.26
7.3
0.25
1,3-DNB
0.11
4.0
0,25
Tetryl
-
4.0
0.65
NB
-
6.4
0.26
2,4,6-TNT
0.11
6.9
0.25
4-Am-DNT
0.060
-
-
2-Am-DNT
0.035
-
-
2,6-DNT
0.31
9.4
0.26
2,4-DNT
0.020
5.7
0.25
2-NT
-
12.0
0.25
4-NT
-
8.5
0.25
3-NT
-
7.9
0.25
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TABLE 2
RETENTION TIMES AND CAPACITY FACTORS ON LC-18 AND LC-CN COLUMNS
Retention time (min) Capacity factor (k)*
Analyte
LC-18
LC-CN
LC-18
LC-CN
HMX
2.44
8.35
0.49
2.52
RDX
3.73
6.15
1.27
1.59
1,3,5-TNB
5.11
4.05
2.12
0.71
1,3-DNB
6.16
4.18
2.76
0,76
Tetryl
6.93
7.36
3.23
2.11
NB
7.23
3.81
3.41
0.61
2,4,6-TNT
8.42
5.00
4.13
1.11
4-Am-DNT
8.88
5.10
4.41
1.15
2-Am-DNT
9.12
5.65
4.56
1.38
2,6-DNT
9.82
4.61
4.99
0.95
2,4-DNT
10.05
4.87
5.13
1.05
2-NT
12.26
4.37
6.48
0.84
4-NT
13.26
4.41
7.09
0.86
3-NT
14.23
4,45
7.68
0.88
•Capacity factors are based on an unretained peak for nitrate at 1.71 min on LC-18 and at 2.00 min
on LC-CN.
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TABLE 3
SINGLE LABORATORY PRECISION OF METHOD FOR SOIL SAMPLES
Spiked Soils Field-Contaminated Soils
Mean Cone. Mean Cone.
Analyte
(mg/kg)
SD
%RSD
(mg/kg)
SD
%RSD
HMX
46
1.7
3.7
14
1.8
12.8

153
21.6
14.1
RDX
60
1.4
2.3
104
12
11.5

877
29.6
3.4
1,3,5-TNB
8.6
0.4
4.6
2.8
0.2
7.1

46
1.9
4.1
72
6.0
8.3
2,4,6-TNT
40
1.4
3.5
7.0
0.61
9.0

669
55
8.2
1,3-DNB
3.5
0.14
4.0
1.1
0.11
9.8
2,4-DNT
5.0
0.17
3.4
1.0
0.44
42.3
Tetryl
17
3.1
17.9
2.3
0.41
18.0
Source: Reference 1.
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TABLE 4
MULTILABORATORY ERROR OF METHOD FOR SOIL SAMPLES
Spiked Soils Field-Contaminated Soils
Analyte
Mean Cone,
(mg/kg)
SD
%RSD
Mean Cone,
(mg/kg)
SD
%RSD
HMX
46
2.6
5.7
14
3.7
26.0

153
37.3
24.0
RDX
60
2.6
4.4
104
17.4
17.0

877
67.3
7.7
1,3,5-TNB
8.6
0.61
7.1
2.8
0.23
8.2

46
2.97
6.5
72
8.8
12.2
2,4,6-TNT
40
1.88
4.7
7.0
1.27
18.0

669
63.4
9.5
1,3-DNB
3.5
0.24
6.9
1.1
0.16
14.5
2,4-DNT
5.0
0.22
4.4
1.0
0.74
74.0
Tetryl
17
5.22
30.7
2.3
0.49
21.3
Source: Reference 1.
TABLES
MULTILABORATORY VARIANCE OF METHOD FOR WATER SAMPLES3
Analyte
Mean Cone. (ng/L)
SD
%RSD
HMX
203
14.8
7.3
RDX
274
20.8
7.6
2,4-DNT
107
7 7
7.2
2,4,6-TNT
107
in
10.4
8 Nine Laboratories
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TABLE 6
MULTILABORATORY RECOVERY DATA FOR SPIKED SOIL SAMPLES

Concentration (yg/g)

Laboratory
HMX
RDX
1,3,5-TNB
1,3-DNB
Tetryi
2,4,6-TNT
2,4-DNT
1
44.97
48.78
48.99
49.94
32.48
49.73
51.05
3
50.25
48.50
45.85
45.96
47.91
46.25
48.37
4
42.40
44.00
43.40
49.50
31.60
53.50
50.90
5
46.50
48.40
46.90
48.80
32.10
55.80
49.60
6
56.20
55.00
41.60
46.30
13.20
56.80
45.70
7
41.50
41.50
38.00
44.50
2.60
3S.00
43.50
8
52.70
52.20
48.00
48.30
44.80
51.30
49.10
True Cone
50.35
50.20
50.15
50.05
50.35
50.65
50.05
Mean Cone
47.79
48.34
44.68
47.67
29.24
49.91
48.32
Std. Dev.
5.46
4.57
3.91
2.09
16.24
7.11
2.78
% RSD
11.42
9.45
8.75
4.39
55.53
14.26
5.76
% Diff.*
5.08
3.71
10.91
4.76
41.93
1.46
3.46
Mean %
Recovery
95
96
89
95
58
98
96
* Between true value and mean determined value.
Source: Reference 1.
8330A- 17
Revision 1
January 1998
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TABLE 7
COMPARISON OF METHOD ACCURACY FOR SOIL AND AQUEOUS SAMPLES
(HIGH CONCENTRATION METHOD)
Analyte
Recovery (%)
Soil Method*
Aqueous Method*
2,4-DNT
2,4,6-TNT
RDX
HMX
96.0
96.8
96.8
95.4
98.6
94^4
99.6
95.5
Data from Reference 1.
' Data from Reference 3.
8330A -18
Revision 1
January 1995
-------
TABLE 8
PRECISION AND ACCURACY DATA FOR THE SALTING-OUT EXTRACTION METHOD
Analyte
# Samples
%RSD
Mean Recovery (%)
Highest
Concentration Tested
HMX
20
10.5
106
1.14
RDX
20
8.7
106
1.04
1,3,5-TNB
20
7.6
119
0.82
1,3-DNB
20
6.6
102
1.04
Tetryl
20
16.4
93
0.93
2,4,6-TNT
20
7.6
105
0.98
2-Am-DNT
20
9.1
102
1.04
2,4-DNT
20
5.8
101
1.01
1,2-NT
20
9.1
102
1.07
1,4-NT
20
18.1
96
1.06
1,3-NT
20
12.4
97
1.23
All tests were performed in reagent water.
Source: Reference 6.
8330A -19
Revision 1
January 1998
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TABLE 9
COMPARISON OF DIRECT ANALYSIS OF GROUNDWATER SAMPLES CONTAINING
NITROAROMATICS WITH SALTING-OUT AND SOLID-PHASE EXTRACTION TECHNIQUES
Analvte Concentration fyg/L)
imole
Techniaue
HMX
RDX
TNB DNB
DNA
TNT 24D
4A
2A
1
Direct

Salting-out
1.04
2.45

0.47
0.36
0.32

SPE-Cart.
1.00
1.33

0.44
0.29
0.30

SPE-Disk
0.93
1.35

0.57 -
0.28
0.56
2
Direct
94
79

Salting-out
54.2
63.8

0.3
0.33
3.08
1.36

SPE-Cart.
64.0
83.1

0.3
0.34
3.34
2.27

SPE-Disk
57.1
71.8

0.3
0.29
2.89
2.05
3
Direct
93
91

Salting-out
85.7
75.3

0.2
0.19
2.43
1.31

SPE-Cart.
93.1
88.8

0.2
0.17
2.49
1.65

SPE-Disk
78.9
74.7

0.2
0.13
1.99
1.89
4
Direct
45
14

Salting-out
45.7
16.4
0.17
0.3
0.13
2.18
1.21

SPE-Cart.
48.0
21.6

0,2
0.19
2.31
1.42

SPE-Disk
40.8
18.9

0.2
0.13
2.07
1.64
5
Direct

Salting-out
0.76
5.77

-
0.13
0.05

SPE-Cart.
1.16
6.48

0.16
0.05

SPE-Disk
1.19
6.11

0.16
0.14
6
Direct

Salting-out
10.5
6.17

0.10
0.71
0.33

SPE-Cart.
11.5
7.03

0.10
0.79
0,40

SPE-Disk
10.3
6.34

0.07
0.82
0.70
8330A-20
Revision 1
January 1998
-------
TABLE 9
(continued)
AnaMe Concentration fuo/L)
Sample
Techniaue
HMX
RDX
TNB DNB DMA
TNT 24D 4A
2A
7
Direct
134
365

Salting-out
75.4
202

0.98 8.12
1.80

SPE-Cart.
115
308

1.51 11.3
3.44

SPE-Disk
109
291

1.41 9.81
3.30
8
Direct

Salting-out
0.61
10.9

SPE-Cart.
0.64
11.9

SPE-Disk
0.64
11.0

9
Direct
25
13

Salting-out
30.2
12.1

1.14
0.56

SPE-Cart.
31.2
12.7

1.50
0.79

QDC riielf
Or t-LUSK
27.5
11.0 "

1.34
0.79
10
Direct

Salting-out
0.33
7.12

SPE-Cart.
0.62
8*23

SPE-Disk
0.26
7.60

14
Direct

Salting-out

5.98

SPE-Cart.

12.0

SPE-Disk

11.6

16
Direct

Salting-out
0.58
28.7
0.04
0.39
0.13

SPE-Cart.
0.77
33.8
0.03
0.43
0.17

SPE-Disk
0.66
32.7
0.03
0.44
0.22
18
Direct
165
58

9
7

Salting-out
141
39.1
0.80
0.96 8.5
5.62
8330A - 21
Revision 1
January 1998
-------
TABLE 9
(continued)
Analvte Concentration fua/LI
Sample
Techniaue
HMX
RDX
TNB
DNB
DNA
TNT
24D
4A
2A

SPE-Cart.
152
44.4

0.93
0.88

9.5
7.01

SPE-Disk
138
40.9

0.90
0.99

9.3
6.03
19
Direct
173
76

59
54

Salting-out
172
69.5

2.6
23.1
1.20
65.2
56.4

SPE-Cart.
142
75.6

0.11
2.5
20.9
1.08
57.7
50.5

SPE-Disk
136
72.7

0.11
2.4
20.3
1.23
55.0
48.0
21
Direct
252
157
5

110

47
65

Salting-out
227
132
6.62
0.30

102

42.6
56.5

SPE-Cart.
238
146
6.90
0.33

104

48.0
63.5

SPE-Disk
226
141
6.45
0.31

102

47.0
61.8
22
Direct
218
40

Salting-out
201
35.9

2,20
1.90

SPE-Cart.
203
36.5

2.74
2.24

SPE-Disk
199
35.8

2.78
2.08
24
Direct

Salting-out
2.15
7.54

SPE-Cart.
2.47
8.91

SPE-Disk
2.34
8.84

25
Direct

Salting-out

SPE-Cart.

0.59

SPE-Disk

0.63

27
Direct
112
608
8

180

10
8

Salting-out
82.8
429
4.45
0.79

137

7.71
6.20

SPE-Cart.
91.0
510
9.53
0.90

149

8.25
7.67

SPE-Disk
77.3
445
7.37
0.79

128

8.16
6.33
8330A - 22
Revision 1
January 1998
-------
TABLE 9
(continued)
Analvte Concentration (uq/LI
Sample Technique HMX RDX TNB DNB DNA TNT 24D 4A
0.37 0.10
0.87 0.17
0.65 0.13
28
Direct
325
102

Salting-out
290
87.5

SPE-Cart.
319
109

SPE-Disk
249
85.7
29
Direct

Salting-out

SPE-Cart

0.43

SPE-Disk

0.28
31
Direct

Salting-out

SPE-Cart.

0.21

SPE-Disk

0.23
32
Direct

Salting-out

SPE-Cart.

SPE-Disk
0.38

TNT
24D 4A
2A
14
51
40
13.9
42.3
33.5
22.0
56.2
45.0
17.2
43.0
34.5
An additional 11 samples (11,12, 13, 15,17,20, 23,26, 30, 31, and 33) were analyzed in which
none of the analytes were detected by any of the techniques. Therefore, the non-detect results are
not shown hem. Similarly, for those samples that are shown here, the fields are left blank for the
analytes that were not detected.
All data are taken from Reference 10.
8330A - 23
Revision 1
January 1998
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TABLE 10
RELATIVE PERCENT DIFFERENCE BETWEEN DUPLICATE SAMPLE ANALYSES
Relative Percent Difference (%1
Sample
Techniaue
HMX
RDX
TNB
DNB
DNA
TNT
24D
4A
2A
4
Direct
0
24

Salting-out
0
15

6
100
8

18
11

SPE-Cait.
1
12

0
45

8
5

SPE-Dlsk
3
8

0
17

2
1
29
Direct
Salting-out
SPE-Cart.
SPE-Disk

26
7

LCS
Direct
1
0
0

1
1

Salting-out
4
4
4

3
3

SPE-Cart.
6
1 -
7

6
6

SPE-Disk
5
7
7

13
6

Alt data are taken from Reference 10.
8330A-24
Revision 1
January 1998
-------
TABLE 11
RECOVERY OF ANALYTES FROM SPIKED SAMPLES

Percent Recovery (%)

Sample
Techniaue
HMX
RDX
TNB
TNT
24D
LCS1
Direct
99.5
98.5
95.6
96.5
98.1

Salting-out
94.2
91.2
92.9
83.2
92.1

SPE-Cart.
99.0
101.0
96.6
94.1
95.1

SPE-Disk
92.5
95.6
89.3
88.6
86.9
LCS2
Direct
98.8
98.2
95.9
97.2
99.2

Salting-out
91.0
95.0
89.0
81.0
89.0

SPE-Cart.
93.5
100.0
83.0
89.1
89.3

SPE-Disk
88.0
102.0
83.0
78.0
82.0
29
Direct
95.0
95.5
95.2
92.8
93.0

Salting-out
107.0
89.0
85.0
89.0
65.0

SPE-Cart.
103.0
107.0
104.0
05.0
102.0

SPE-Disk
80.0
78.0
76.0
78.0
77.0
4
Direct
105,5
105.0
103.0
104.0
105.0

Salting-out
23*
191*
76.0
83.0
76.0

SPE-Cart.
351*
95*
92.2
91.1
93.7

SPE-Disk
308*
49.5*
87.4
85.6
90.8
All data are taken from Reference 10.
* Results for these analytes in Sample 4 are believed to result from spiking levels that are very
similar to the background concentrations of these analytes in this sample (see Reference 10).
8330A - 25
Revision 1
January 1998
-------
FIGURE 1
EXAMPLE CHROMATOGRAMS
EXPLOSIVES ON A
CIS COLUMN
i.
I
X
J,
JL
J.
EXPLOSIVES OH *
ON COLUMN
*•
a
.A
4
i.
±
!
0
2
6
8
8330A - 26
Revision 1
January 1998
-------
METHOD 8330A
NITROAROMATICS AND NITRAMINES BY HIGH
PERFORMANCE LIQUID CHROMATOGRAPHY (HPLC)
/ Uh \
Liquid-Liquid X Liquid-Liquid
V Extraction or
\ SP6 V
/ 7.1 \
I* it high or
High
Low
7.1.3 Solid Phasa
Extraction (SPE).
Go to Mathod 3S35.
7.1.1.8 Filtar if turbid.
Transfer to a vial for
RP-HPLC analysis.
7.1.1.2 Add 164 ibL of
acatonitrlia (ACN) and
stir. Allow ptiasas to
so parati.
7.1.14 Add 84 nriL of
•alt wafer ts tho ACN
attract and stir. Traiwfar
ACN attract ts 10 mL
grad. cyltndar.
SalHno Out
7.1.1.1 Add 251.3 o of salt
and 770 mL of watw
•am pit to a 1 I vol. flask.
Mis tha contents.
7.1.1.5 *
100 mL
1 mL of ACN to
I. flask. Stir and
tnnsfar to tha 10 mL grad.
eylindat. Racord voluma.
Diluta 1:1
7.1.2 Sam pit Filtration:
Placa S mL ai
scmtilstjcn vial,
tnathansl, sliaka,
&
filter.
Oiscard first 3 mL. Ratain
twmaindar for usa.
7.1.1.3 Transfer ACN layar
to 100 mL vol. flask. Add
10 mL of frash ACN to 1 L
Rask and stir. Transfer 2nd
wrtiorr and comtrina with 1st
100 mL flask.
8330A - 27
Revision 1
January 1998
-------
METHOD 833QA
(continued)
o
7.1,4.1 Sample Homogenization
Air dry sample at room Temp,
or below. Avoid exposure to
direct sunlight. Grind sample
in an acetonftrile rinsed mortar.

r
7.1.4.2 Sample Extraction.
>
r
7.1.4.2.1 Place 2 g soil
subsample, 10 mis
acetonitrile in 15 mL
glass vial. Cap. vortex
swirl, place in room Temp,
or below ultrasonic bath
for 18 hrs.
7.1.4.2.2 Let soln. settle.
Add 5 mL supernatant to
5 mL calcium chloride
soln. in 20 mL vial. Shake,
Let stand 15 mins.
7.1.4.2.3 Filter supernatant
through 0.5 um filter. Discard
initial 3 mL, retain remainder
for analysis.
7.4 Sample Analysis.
7.3 Calibration of
HPLC.
7,2 Set Chromatographic
Conditions.
7.4.3 Refer to Table 2 for
typical anlayte retention
times.
7.4.1 Analyze samples. Confirm
measurement w/injection onto
_ CN column.
7.3.2 Run working stds. in
triplicate. Calculate calibration
factors as described in
Method SOOO.
7.3.3 Analyze midrange
calibration std. at beginning,
after every 20 sample extracts
and end of sample analyses.
Redo Section 7.3.1 if mean
calibration factors do not
agree to w/in +/-15% of
initial calibration.
J
C Stop )
8330A - 28
Revision 1
January 1998
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4.3 DETERMINATION OF ORGANIC ANALYTES
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the
allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to
be used by the analyst and the regulated community in making judgements necessary to meet
the data quality objectives or needs for the intended use of the data.
4.3.4 INFRARED METHODS
The following methods are included in this section:
Method 8410:
Method 8430:
Method 8440:
Gas Chromatography/Fourier Transform Infrared (GC/FT-IR)
Spectrometry for Semivolatile Organics: Capillary Column
Analysis of Bis(2-chloroethyi) Ether and Hydrolysis Products
by Direct Aqueous Injection GC/FT-IR
Total Recoverable Petroleum Hydrocarbons by Infrared
Spectrophotometry
FOUR-13 Revision 4
January 1998
-------
4.3 DETERMINATION OF ORGANIC ANALYTES
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the
allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to
be used by the analyst and the regulated community in making judgements necessary to meet
the data qualify objectives or needs for the intended use of the data.
4.3.5 MISCELLANEOUS SPECTROMETRY METHODS
The following method is included in this section:
Method 8520: Continuous Measurement of Formaldehyde in Ambient Air
FOUR-14
Revision 4
January 1998
-------
4.4 IMMUNOASSAY METHODS
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the
allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to
be used by the analyst and the regulated community in making judgements necessary to meet
the data quality objectives or needs for the intended use of the data.
The following methods are included in this section:
Method
4000:
Method
4010A:
Method
4015:
Method
4020:
Method
4030:
Method
4035:
Method
4040:
Method
4041:
Method
4042:
Method
4050:
Method
4051:
Method
4670:
Immunoassay
Screening for Pentachlorophenoi by Immunoassay
Screening for 2,4-Dichlorophenoxyacetic Acid by
Immunoassay
Screening for Polychlorinated Biphenyls by Immunoassay
Soil Screening for Petroleum Hydrocarbons by Immunoassay
Soil Screening for Polynuclear Aromatic Hydrocarbons by
Immunoassay
Soil Screening for Toxaphene by Immunoassay
Soil Screening for Chlordane by Immunoassay
Soil Screening for DDT by Immunoassay
TNT Explosives in Soil by Immunoassay
Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) in Soil by
Immunoassay
Triazine Herbacides as Atrazine in Water by Quantitative
Immunoassay
FOUR-15
Revision 4
January 1998
-------
METHOD 4670
TRIAZINE HERBICIDES AS ATRAZ1NE IN WATER
BY QUANTITATIVE IMMUNOASSAY
1.0 SCOPE AND APPLICATION
1.1 This method describes a procedure for the quantitative determination of atrazine
(CAS# 1912-24-9) and other triazine herbicides in water using a competitive immunoassay. The
method provides a single quantitative result, reported as atrazine, for all compounds detected.
However, the extent to which other triazine herbicides and other compounds are detected may vary
between commercial testing products (see Sees. 1.4 and 4.0).
1.2 Testing products are commercially available from several manufacturers. The testing
product evaluated by EPA for this method employs a competitive immunoassay. Other products
differ in a number of respects, including the format of the test (tubes versus microtiter plates), the
reagents used, and the specific steps in the test procedure.
1.3 The method detection limit (MDL) submitted by the manufacturer of the testing product
described in Sec. 6.2 was 0.03 |jg/L for drinking water samples. The actual detection limit may be
highly dependent on the sample matrix and analyst's performance.
1.4 Since immunoassay methods use antibody molecules that can bind to more than the
target analyte, an immunoassay has a tendency to overestimate the concentration of the target
analyte when other analytes are present that may bind with the antibody. The commercially-available
testing product evaluated for this method is based on an immunochemical reaction that will also
respond to other triazine compounds. These other triazine compounds are often included in
pesticide formulations containing atrazine. Thus, the specificity of this procedure for atrazine is
partly a function of the cross-reactivity of those other compounds (see Table 1). Therefore, as with
other analytical techniques such as single-column gas chromatography, it is advisable to confirm
positive test results near or above a regulatory action limit when the presence of other triazines is
suspected.
1.5 This method is restricted to use by or under the supervision of analysts trained in the
performance and interpretation of immunoassay methods. Each analyst must demonstrate the ability
to generate acceptable results with this method (see Sec. 9.5).
2.0 SUMMARY OF METHOD
2.1 An accurately measured volume of sample (as little as 200 mL for some testing
products) is mixed with a volume of enzyme-atrazine conjugate reagent in a test tube or a microtiter
plate that has an anti-atrazine antibody immobilized on the surface, or in a vessel to which particles
(magnetic particles for one testing product) with an immobilized antibody on the surface are added.
The conjugate "competes" with the atrazine present in the sample for binding to the immobilized
anti-atrazine antibody. The mixture is incubated at the temperature, and for the time, described in
the manufacturer's instructions. (Testing products may employ other solid-phase support
configurations, or even eliminate the solid-phase support. The summary here is intended to be
generic and not to limit the development of other testing products).
4670-1
Revision 0
January 1998
-------
2.2 Unbound conjugate and sample analyte that may be present in the tubes or wells are
removed by washing with organic-free reagent water or wash solution specified by the manufacturer.
A signal-generating substrate/chromogen reagent is added and the tube or plate is incubated as
described in the manufacturer's instructions, in the case of the testing product described in Sec. 6.2,
a magnetic field is applied to the tubes to retain the magnetic particle coated with antibody and any
bound enzyme conjugate present during the wash step. (Other testing products may use different
configurations).
2.3 In an enzyme immunoassay, a stop solution is added to the tubes or wells of the plate
to terminate the signal generating activity of the enzyme conjugate reagent. The absorbance is
measured at a wavelength specified by the manufacturer. The test is interpreted by measuring the
signal produced by a sample and determining the concentration from a dose-response curve
constructed from standards tested at the same time. For a competitive immunoassay, the color
(signal) developed during the test is inversely proportional to the concentration of atrazine in the
sample.
3.0 DEFINITIONS
The definitions associated with immunoassay procedures are given in Method 4000 and in
the glossary at the end of this method.
4.0 INTERFERENCES
4.1 Compounds that are chemically similar may cause a positive test result (false positive)
for atrazine. This phenomenon is known as cross-reactivity. The testing product used in preparation
of this method has been evaluated for cross-reactivity by the manufacturer. Table 1 provides the
concentration at which known cross-reactants will give a comparable response to that of atrazine
when present in the sample.
4.1.1 The presence of cross-reacting compounds will result in an increase in the
calculated concentration of the sample being analyzed and therefore influence the incidence
of false positive results. Thus, from the standpoint of monitoring compliance with a regulatory
action limit, cross-reactivity is not a significant concern for test results below the action limit.
4.1.2 As with techniques such as single-column gas chromatography, in instances
where the presence of other triazine compounds is known or suspected, it may be advisable
to confirm positive results near or above the regulatory action limit using another analytical
technique. However, false negative results are generally not a concern with immunoassay
techniques.
4.2 Non-specific interferences such as sample pH, temperature, osmolality, solvents,
surfactants, and the presence of metal ions can effect immunoassay performance. Samples should
be tested at the pH and temperature range specified by the testing product manufacturer. Review
the product literature with regard to other potential interferences.
4.3 Storage temperatures may alter the useful life of the testing product reagents and
supplies. Follow the manufacturer's directions for storage and use of all reagents and supplies.
4670-2
Revision 0
January 1998
-------
5.0 SAFETY
No extraordinary safety measures are required. However, safety procedures consistent with
good laboratory practices should be employed. Some reagents may contain dilute add solutions.
Avoid contact with eyes, skin, and mucous membranes.
6.0 EQUIPMENT AND SUPPLIES
6.1 Each commercially-available testing product will supply or specify the apparatus and
materials necessary for successful completion of the test. Most testing products supply the
equipment and supplies specific to the immunoassay, including the tubes or plates containing the
immobHized antibody, and the immunochemical reagents. Do not mix the equipment, supplies, and
reagents from the testing products for different analytes, or from the testing products from different
manufacturers. Testing products contain immunochemical reagents that are evaluated by the
manufacturer on a lot-specific basis. Do not mix the reagents from one lot with those from another
lot unless expressly allowed by the manufacturer. Other equipment that may be required, but is not
supplied with the testing product, includes common laboratory items such as precision pipetting
devices, vortex mixers, etc.
6.2 The immunoassay testing product listed below has been submitted to EPA, evaluated
by the Agency, and found to meet the performance specifications necessary for inclusion in SW-846.
Additional testing products may be available from other manufacturers or in different formats. As
additional testing products are evaluated by EPA and found to provide equivalent performance,
information will be made available by the Office of Solid Waste regarding all those testing products
that are capable of meeting the performance specifications in this method. However, this procedure
mil not be revised solely to include information on additional testing products.
Atrazine RaPID Assay® (Ohmicron Environmental Diagnostics, Inc.).
7.0 REAGENTS AND STANDARDS
As with the equipment and supplies, each commercially-available testing product will supply
or specify the reagents necessary for successful completion of the test. This includes the calibrators
(standards) employed in the immunoassay. As noted in Sec. 6.1, do not mix the equipment,
supplies, and reagents from the testing products for different analytes, or from the testing products
from different manufacturers. Store all reagents and standards according to the manufacturer's
instructions, and, where applicable, discarding any which have exceeded the expiration date
assigned by the manufacturer.
In addition, in order to demonstrate the method performance described in Sec. 9, the following
reagents and standards mil be required.
7.1 Organic-free reagent water - All references to water in this method refer to organic-free
reagent water, as defined in Chapter One. Organic-free reagent water is used for the preparation
of the initial demonstration of capability test, the laboratory control sample, and other quality control
tests. These tests are in addition to any control material(s) supplied by the manufacturer.
7.2 Atrazine spiking solution - a solution of atrazine in a water-miscible solvent is required
for spiking into organic-free reagent water to prepare the initial demonstration of proficiency test, the
laboratory control sample, and other quality control tests. This solution may be provided by the
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manufacturer. If not provided, the laboratory should prepare a spiking solution or purchase one from
a commercial source. Consult the manufacturer's instructions regarding solvents that may interfere
with the testing product and do not use them. The concentration of this solution should be
approximately 0.3 pg/mL, such that a 100 mL volume spiked into a 10 mL volume of reagent water
will yield a concentration of 3 ms'L. Other volumes and concentrations may be employed, provided
that the laboratory can demonstrate that the volume of solvent used does not affect the test
performance.
7.3 Solutions for adjusting the pH of samples before extraction, where such pH adjustment
is specified by the manufacturer.
7.3.1 Sulfuric acid solution (1:1 v/v), H2S04 - Slowly add 50 mL of HjS04 (sp, gr.
1.84) to 50 mL of organic-free reagent water.
7.3.2 Sodium hydroxide solution (2N), NaOH - Dissolve 8 g NaOH in organic-free
reagent water and dilute to 100 mL.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
Sample collection, preservation, and storage requirements may vary by EPA program and
may be specified in the regulation that requires compliance monitoring for a given contaminant
Where such requirements are specified in the regulation, follow those requirements. In the absence
of specific regulatory requirements, use the following Information as guidance in determining the
sample collection, preservation, and storage requirements.
8.1 Sample Collection
The immunoassay testing products employ very small (< 1 mL) sample volumes. Therefore,
sample collection procedures should focus on the volume necessary to ensure that the sample
represents the source. «
8.1.1 Samples should be collected in pre-cleaned glass containers.
8.1.2 When sampling from a water tap, open the tap and allow the system to flush
until the water temperature has stabilized (usually 2 to 5 minutes). Adjust the flow to about
500 mL/min, and collect samples from the flowing stream. When sampling from an open
body of water, fill the sample container with water from a representative area.
8.2 Sample preservation
8.2.1 if residual chlorine is present, add 80 mg of sodium thiosulfate per liter of
sample to the bottle, prior to collecting the sample.
8.2.2 Retard microbiological degradation by adjusting the pH of the samples to <2
with hydrochloric acid at the time of sample collection. Before analysis, readjust the pH of the
samples to the pH specified by the manufacturer with 2N NaOH. The pH of the entire
collected sample should be adjusted, not just the small volume utilized for the analysis.
8.3 Sample storage - Samples should be stored at 4 ± 2°C until analysis, but must be
warmed to the temperature specified by the manufacturer for analysis.
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8.4 EPA has not conducted holding time studies relative to immunoassay.
9.0 QUALITY CONTROL
As noted in Sec. 1.2, the specific formats of the commercially-available testing products vary
by manufacturer. As a result, those testing products evaluated and accepted by EPA represent
performance-based analytical methods. Therefore, it is imperative that the manufacturer's
instructions and specifications be followed closely. Follow the manufacturer's instructions for
the testing product being used for quality control procedures specific to the testing product used.
The following discussion of quality control requirements relies heavily of the analyst's knowledge
and understanding of the manufacturer's instructions.
9.1 Initial calibration
An initial calibration must be performed concurrent with the analysis of any samples, as
described in Sec. 10.
9.2 Calibration verification
Calibration verification is not performed in the traditional sense because the initial calibration
standards are analyzed with each batch of samples each time the analyses are performed.
9.3 Routine Quality Control
Routine quality control procedures associated with this method include the analyses of
standards, matrix spike samples, laboratory control samples, method blanks, and duplicate or
replicate analyses (as specified by the manufacturer). All of the analyses described below must be
conducted simultaneously, e.g., as part of the same batch of samples. A batch of samples consists
of up to 20 field samples prepared and analyzed at the same time, or the maximum number of
samples that can be analyzed along with the standards, controls, and other analyses specified by
the manufacturer using a single testing product, whichever is fewer. The batch must include any
duplicate or replicate analyses specified by the manufacturer as well as all additional quality control
tests specified by EPA in this procedure.
9.3.1 Calibration standards must be analyzed concurrently with each batch of
samples processed.
9.3.2 Matrix spike (MS) samples must be analyzed with each batch of samples
processed. The matrix spike samples should contain atrazine at the regulatory limit of
interest (e.g., the MCL for the Drinking Water Program). The sample chosen for spiking
should be representative of the field samples being analyzed.
9.3.3 The analyst must evaluate the accuracy of the assay by analyzing a
laboratory control sample (LCS) consisting of organic-free reagent water sample spiked at the
regulatory limit of concern for atrazine. For the Drinking Water Program, the LCS must be
spiked at 3 pg/L (the MCL for atrazine) with the spiking solution in Sec. 7.2. The mean
recovery (bias) of the assay must be between 80-120%. If the manufacturer does not supply
the spiking solution described in Sec. 7.2, or if another regulatory limit is relevant, then the
laboratory is responsible for purchasing or preparing an appropriate spiking solution and
performing this test. An LCS must be prepared and analyzed with each batch of samples
analyzed.
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NOTE: Spiking at 3 M9^- may require that the sample be diluted to be within the calibration range
for some testing products, however, it provides data regarding the Was (if any) at the
regulatory threshold, as well as indications of the analyst's proficiency at making dilutions.
9.3.4 A method blank, consisting of a volume of organic-free reagent water (see
7.1) equal to that of a field sample, must be analyzed with each batch of samples processed.
The method blank should not contain any detectable atrazine.
9.3.5 Samples should be analyzed in duplicate or triplicate, as instructed by the
manufacturer. The number of replicate analyses is specified by the manufacturer, and is a
function of the overall precision of the particular testing product. If the manufacturer
determines that, in order to achieve the precision claimed by the manufacturer, a given
number of replicate analyses must be performed, then the laboratory must employ the
specified number of replicate analyses.
9.4 Sample Dilutions
If the sample concentration is outside of the calibrated range demonstrated by the initial
calibration and as specified by the manufacturer, then the sample must be diluted to within the
calibration range and re-tested. As employed in these testing products, the calibration range
specified by the manufacturer is based on a B/B0 in the 0.2-0.8 (20-80%) range. Given the nature
of the competitive immunoassay, the sample cannot be diluted after color development. Thus, a
diluted aliquot of the original sample must be prepared and analyzed.
NOTE: The B/Bq range of 0.2-0.8 is narrower than the simple concentration range of the calibration
standards. Therefore, the decision to dilute a sample for reanalysis must be based on an
evaluation of the B/B0 value of the sample, and not on a simple comparison of the
concentration in the sample and the highest standard in the calibration.
9.5 Initial demonstration of proficiency
Each laboratory must demonstrate initial proficiency with the testing product that it utilizes,
by generating data of acceptable accuracy and precision for a reference sample containing atrazine
in a dean matrix. The laboratory must also repeat this demonstration whenever new staff are trained
or significant changes in instrumentation are made.
9.5.1 The reference sample is prepared from a spiking solution containing the
anaiyte of interest (see Sec. 7.2). Given the very small sample volume required for the
immunoassay, a single 10-mL aliquot will provide sufficient volume for multiple tests and
minimizes the difficulties involved in spiking small volumes of organic-free reagent water.
Prepare a new aliquot each time the initial demonstration is to be performed.
9.5.2 Prepare an aliquot of organic-free reagent water, spiking it with the solution
in Sec. 7.2 to yield a concentration of 3 M9/L. Mix the aliquot well and allow the spiked
sample to stand for at least one hour.
9.5.3 Analyze at least four replicate subsamples of the spiked organic-free reagent
water aliquot using the same procedures used to analyze actual samples (Sec. 11). Analyze
the number of replicates of each subsample specified by the manufacturer, e.g., if the
manufacturer specifies triplicate analyses of samples, then analyze 12 replicates (4 x 3) of
the spiked sample.
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9.5.4 Calculate the mean recovery (X), and the standard deviation of the
recoveries using the total number of replicate results, as described in Sec. 12.6.
9.5.5 Given the total number of replicate analyses performed, the mean recovery
(X) should be in the range of 90-110% and the relative standard deviation should be no more
than 10% of the mean recovery. If the results fall outside of these acceptance limits, recheck
all calculations. If no errors are found, repeat the demonstration until the specifications are
met
9.6 Other Quality Control Considerations
9.6.1 Do not use testing products past their expiration date.
9.6.2 Do not mix the equipment, supplies, and reagents from the testing products
for different analytes, or from the testing products from different manufacturers.
9.6.3 Use the testing products within the storage temperature and operating
temperature limits specified by the manufacturer.
10.0 CALIBRATION AND STANDARDIZATION
10.1 The analyst must perform an initial calibration. This calibration is performed
concurrently with the analysis of samples.
10.1.1 The initial calibration must consist of standards (calibrators) at a minimum
of three concentrations that describe the quantitation range of the assay and should
preferably span the regulatory limit of interest (e.g., for drinking water, the maximum
contaminant level [MCL] is 3.0 vgll). The standards must fall within the B/Bq range of 0.2 to
0.8. The calibrators are generally provided by the product manufacturer. Calibration curves
where all the calibrators are below the regulatory limit are allowed, but will require dilution and
reanalysis of samples when the sample concentration is near the regulatory limit. Calibration
curves where all the calibrators are above the applicable regulatory limit may not be employed
for compliance monitoring.
10.1.2 The testing product must also contain a "zero standard" or diluent solution
that contains none of the target analytes. This solution is used to generate the B0 value, but
must not be used as one of the three standards specified in Sec. 10.1.1.
10.1.3 When the entire dose response of a competitive immunoassay testing
product (the absorbance of the solution or other signal specified-by the manufacturer) is
plotted on the y-axis against the concentration of the calibration standard on the x-axis, the
resulting calibration curve will be hyperbolic when plotted on rectilinear paper, sigmoidal when
plotted on semi-log paper, and linear when a Log'rt-log transformation of the data is employed
and plotted on rectilinear paper. In addition, since the immunoassay is competitive, the blank
(zero standard) will yield the highest response, with the color development inversely
proportional to the standard concentration.
A plot of either the Logit B/Bq or the Log it of the signal (absorbance units) versus the
natural log of concentration is a widely used representation of the calibration data that
generally yields a linear response curve. It is the basis of most computerized data analysis
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algorithms for competitive binding assays. The Logit B/B0 is calculated according to the
following formula:
loge = Natural log or logarithm base e
B = Response of the standard or sample
Bq - Response of the zero standard
When Logit B/Bc is plotted against the natural log (loge) of concentration, the results
approximate a straight line with a negative slope (see Figure 1c). The transformed calibration
data can then be characterized by the slope, intercept, correlation coefficient, and standard
error of the line. The following sections describe the use of the Logit-log transformation of
the data to prepare a calibration curve. Manufacturer's may provide software that performs
these calculations and, if provided, such software should be employed according to the
manufacturer's instructions.
10.1.3.1 The commercially-available testing products may specify the
analysis of standards in duplicate, or even in triplicate in some testing products. Thus,
a three-point initial calibration may generate six to nine calibration points. Calculate the
mean response (absorfeance) at each concentration, and use this in all subsequent
calculations.
10.1.3.2 Following the Logit B/B0 and log transformations described in
Sec. 10.1.3, construct a first order regression line (e.g., y = mx + b) using Logit B/B0
as the dependent variable (y-axis) and the loge concentration as the independent
variable (x-axis). Since the slope of the line is negative, the regression cannot be
forced through the origin, as the zero standard will yield the highest response and a
value of 1.0 for B/B0. The standards used to construct the regression line all must
have B/B0 values (prior to the Logit transformation) that fall within the 0.2-0.8 range.
The correlation coefficient of the regression (r) must be at least 0.98 in order
to employ the calibration curve (manufacturers may provide more stringent linearity
requirements for their testing products). If r is less than 0.98, check the expiration
dates of all reagents, review the procedures to ensure-that all standards were
incubated for the same time specified by the manufacturer, and perform a new
calibration.
10.2 By convention, the working range of an immunoassay calibration curve is defined as
the range of B/Bq from 0.2 to 0.8 (or %B/B0 from 20% to 80%). Samples may be quantitated only
within the working range of the curve.
10.3 As noted in Sec. 9, a new initial calibration curve must be constructed with each batch
of samples assayed.
B
Logit (B/Bq) = log.
where:
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11.0 PROCEDURE
Follow the manufacturer's instructions for the test being used. These instructions are
summarized in Sees. 11.1 through 11.3, however, given the difference in test formats and reagents,
the discussion is generic in nature. Where the manufacturer's instructions contradict these
instructions or where these instructions do not apply to a specific testing product, follow the
manufacturer's instructions.
11.1 Prepare the samples and standards
11.1.1 Bring samples, controls, and reagents to ambient temperature. Verify that
the ambient temperature is consistent with the manufacturer's recommendations and
imitations for the method. Do not attempt to perform tests outside of the temperature range
specified by the manufacturer.
11.1.2 Check the pH of the samples. If necessary, adjust the pH to the range
specified by the manufacturer, using 2N NaOH.
11.2 Prepare the spectrophotometer, photometer, or signal measurement equipment
specified by the manufacturer.
11.3 Assay samples
11.3.1 Dispense the standards, controls, and samples into the container specified
by the manufacturer. Be certain to include the replicate analyses specified by the
manufacturer and the routine quality control samples specified in Sec. 9.3 (also in replicate
if samples are analyzed in replicate). Determine the maximum number of standards, controls,
and samples that can be analyzed simultaneously and limit the number of field samples
accordingly.
11.3.2 Dispense the enzyme conjugate reagent into each container as specified by
the manufacturer.
11.3.3 Dispense the antibody capture reagent (where appropriate) as specified by
the manufacturer.
11.3.4 Immunoassay methods employ kinetic and chromogenic reactions that are
temperature sensitive. As a result, take care to perform the assay in the temperature range
recommended by the manufacturer. Failure to follow temperature recommendations can lead
to anomalous test results.
NOTE: Do not attempt to process more samples simultaneously than specified by the manufacturer,
as the additional processing time will lead to different incubation times for the samples and
standards being tested and will produce erroneous results.
11.3.5 Wash each tube or well with washing reagents, as directed by the
manufacturer.
11.3.6 Dispense the signal generating and signal terminating reagents (e.g.,
substrate/chromogen reagent and stop solutions) to each container in accordance with the
manufacturer's instructions. Pay careful attention to the incubation times specified by the
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manufacturer. Failure to follow incubation time recommendations can lead to erroneous
results.
11.3.7 Interpret the test results within the time specified by the manufacturer.
Follow the manufacturer's instructions for determining the sample concentration. For
instance, read absorbance values (or optical density) at wavelength(s) specified by the
manufacturer. Follow the manufacturer's quality control and data acceptance instructions.
12.0 DATA ANALYSIS AND CALCULATIONS
As with the specific formats of the testing products and the reagents and supplies, the
specifics of the required calculations may vary by manufacturer. Some testing products may provide
measuring devices such as optical density readers or spectrophotometers and may include software
for performing all the necessary calculations. Other testing products may require the analyst to plot
results manually, using graph paper that may or may not be provided with the testing product, and
determine sample results by interpolation from a standard curve. Whichever approach is used, the
laboratory records (bench notes, etc.) should clearly indicate how the results were obtained and
records specific to each determination, whether in hard copy or in electronic form, should be retained
by the laboratory to substantiate the results.
12.1 Follow the manufacturer's instructions regarding calculation of all testing product
results. Use the calibration curve generated concurrently with the sample analyses.
12.2 Where replicate test results are generated for samples or standards, calculate the
mean concentration (C) as:
n
mean concentration = C = ——
where C, is the concentration in each replicate and n is the number of replicate analyses.
12.3 For duplicate test results, calculate the relative percent difference (RPD) according to
the following equation:
RPD = J—! Ll x 100
Pi + C,)
2
where C, and G, are the concentrations of the two replicate determinations.
12.4 When the manufacturer's instructions specify the analyses of three or more replicates,
calculate the standard deviation (SD) and the relative standard deviation (RSD) of the replicate
results for each sample, according to the following equations:
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£(Ci-c)2
M
SD = — * 100
where C, is the concentration in each replicate, € is the mean concentration, and n is the number
of replicate analyses.
12.5 Accuracy is estimated from the recovery of spiked analytes from the matrix Of interest.
Laboratory performance in a dean matrix is estimated from the recovery of analytes in the LCS.
Calculate the recovery of each spiked analyte in the matrix spike, matrix spike duplicate (if
performed) and LCS according to the following formula.
C, = Measured concentration of the spiked sample aliquot
Cu = Measured concentration of the unspiked sample aliquot (use 0 for the LCS)
Cn = Nominal (theoretical) concentration of the spiked sample aliquot
12.6 For the initial demonstration of proficiency (Sec. 9.6) calculate the mean recovery (X),
and the standard deviation of the recoveries, using the results from ail replicate analyses of the four
subsamples. Use the equation in Sec. 12.4 for standard deviation, substituting recovery for
concentration.
13.0 METHOD PERFORMANCE
13.1 Table 1 summarizes the cross-reactivity of other triazines relative to atrazine for the
testing product listed in Sec. 6.2. Other testing products may have different cross-reactivity
characteristics.
13.2 Table 2 summarizes the single laboratory MDL data submitted by the manufacturer for
the testing product in Sec. 6.2.
13.3 Table 3 summarizes the results of a collaborative study of the immunoassay testing
product described in Sec. 6.2 conducted under the auspices of the AOAC and described in
Reference 3.
13.4 Figure 1 (a-c) provides throe graphical representations of the calibration of atrazine
using a competitive binding immunoassay such as those described here.
C - C
Recovery = %R = — a x 100
C„
where:
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14.0 POLLUTION PREVENTION
Analysis for atrazine using immunoassay conforms with EPA's pollution prevention goals.
Little, if any, solvent is used and minimal waste is generated.
15.0 WASTE MANAGEMENT
Laboratory waste management procedures must be consistent with federal, state, and local
regulations.
16.0 REFERENCES
1. "Principles of Competitive Protein-Binding Assays," Dell, W.O,, Franchimont, P„ John Wiley
and Sons, New York, 1983.
2. "Immunoassay Analysis and GC/MS Confirmation for Residues of Atrazine in Water Samples
from a Field Study Conducted by the State of Wisconsin," Project No. 101174, Report No.
ABR-91069, CIBA-GEIGY Corporation, April 6,1992.
3. "Determination of Atrazine in Water by a Magnetic Particle Immunoassay; Collaborative
Study," Hayes, Mary C., Jourdan, Scott W., and Herzog, David P., JAOAC, 79(2): 530-538,
1996.
4. "Performance Characteristics of a Novel Magnetic-particle-based Enzyme-linked
Immunosorbent Assay for the Quantitative Analysis of Atrazine and Related Triazines in
Water Samples," Rubio, Fernando M., Itak, Jeanne M,, Scutellaro, Adele M., Selisker, Michele
Y.f and Herzog, David P., Food & Agricultural Immunology, 3:113-125,1991.
5. "Comparison of an Enzyme immunoassay and Gas Chromatography/Mass Spectrometry for
the Detection of Atrazine in Surface Waters," Gruessner, Barry, Shambaugh, Nathaniel C.,
and Watzin, Mary, C., Environmental Science and Technology, 29:251-254,1995.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 through 3 and Figure 1.
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TABLE 1
CROSS-REACTIVITY OF RaPID ASSAY TO RELATED COMPOUNDS
Percent Reactivity
Compound CAS # Relative to Atrazine
Atrazine
1912-24-9
100
Ametryn
834-12-8
185
Prometryn
7287-19-6
113
Propazine
139-40-2
?7
Prometon
1610-18-0
32
De-ethylated atrazine
6190-65-4
22
Simazine
122-34-9
15
Terbutryn
886-50-0
13
Terbuthylazlne
5915-41-3
5
Hydroxy atrazine
2163-68-0
0.5
De-isopropylated atrazine
1007-28-9
0.3
Cyanazine
21725-46-2
<0.1
TABLE 2
METHOD DETECTION LIMIT (jjg/L)
Product
n
Spike Level
Std. Dev.
MDL1
RaPID Assay
10
0.1
0.0105
0.03
1 The manufacturer reported MDL results for 10 replicates but used the Student's t value of 3.143,
for seven replicates, in performing the calculations. The value shown above was corrected to the
appropriate t value of 2.821.
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TABLE 3
SUMMARY STATISTICS OF COLLABORATIVE STUDY OF RaPID ASSAY (Source: Ref. 3)
Sample Type
Spike
Cone.
(lig/L)1
n
Mean Cone.
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FIGURE 1
CALIBRATION DATA FROM A COMPETITIVE IMMUNOASSAY
Figure 1a - Generalized plot of immunoassay signal (test response) versus concentration of
calibration standard (pg/L).
J
2
1
0
•24 SJB .1J .U 41 M U iJB « ZO
ift| ft—li atfam
Figure 1b - Generalized plot of B/(B0-B) versus log concentration of calibration standard.
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FIGURE 1
(continued)
CALIBRATION DATA FROM A COMPETITIVE IMMUNOASSAY
110
1J
IS
Loffc(W»o-*) U
M
m
A»
A*
•fct 44 -M -4* 4S «0 AS 10 M 19
lofCooontndca
Figure 1c - Generalized plot of Logit [B/(B0-B)] versus log concentration of calibration standard.
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GLOSSARY OF TERMS
Antibody - A binding protein which is produced in response to an antigen, and which has the ability
to bond with the antigen that stimulated its production.
%B/B0 - an indication of the displacement characteristics of the conjugate from the antibody at
specified concentrations of the target compound.
%_B_ _ Response of the standard or sample x ^
B0 Response of the zero standard
Competitive Immunoassay - An immunoassay method involving an in-vitro competitive binding
reaction.
Cross-Reactivity - The relative concentration of an untargetecf substance that would produce a
response equivalent to a specified concentration of the targeted compound. In a quantitative
immunoassay, it provides an indication of the concentration of cross-reactant that would produce
a positive response. Cross-reactivity for individual compounds is often calculated as the ratio of
target substance concentration to the cross-reacting substance concentration at 50% inhibition of
the immunoassay's maximum signal times 100%.
Dose-Response Curve - Representation of the signal generated by an immunoassay (y axis) plotted
against the concentration of the target compound (x axis) in a series of standards of known
concentration. When plotting a competitive immunoassay in a rectilinear format, the dose-response
will have a hyperbolic character. When the log of concentration is used, the plot assumes a
sigmoidal shape, and when the log of signal is plotted against the Logit transformation of
concentration, a straight line plot is produced.
ELISA - Enzyme Linked Immunosorbent Assay is an enzyme immunoassay method that uses an
immobilized reagent (e.g. .antibody adsorbed to a plastic tube), to facilitate the separation of targeted
anaiytes (antibody-bound components) from non-target substances (free reaction components) using
a washing step, and an enzyme conjugate to generate the signal used for the interpretation of
results.
Enzyme Conjugate - A molecule produced by the coupling of an enzyme molecule to an
immunoassay component that is responsible for acting upon a substrate to produce a detectable
signal.
Enzyme Immunoassay - An immunoassay method that uses an enzyme conjugate reagent to
generate the signal used for interpretation of results. The enzyme mediated response may take the
form of a chromogenic, fluorogenic, chemiluminescent or potentiometric reaction, (see Immunoassay
and ELISA)
False Negatives - A negative interpretation of the method containing the target anaiytes at or above
the detection level. Ideally, an immunoassay test product should produce no false negatives. The
false negative rate can be estimated by analyzing split samples using both the test product and a
reference method.
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False Positives - A positive interpretation for a sample is defined as a positive response for a sample
that contains analytes below the action level.
Immunoassay - An analytical technique that uses an antibody molecule as a binding agent in the
detection and quantitation of substances in a sample, (see Enzyme Immunoassay and ELISA)
Immunogen - A substance having a minimum size and complexity, and that is sufficiently foreign to
a genetically competent host to stimulate an immune response.
Logit - A logarithmic transformation of data normalized to the highest observed response. For the
competitive immunoassay described in this procedure, the Logit transformation is calculated as:
Natural Log - The logarithm, base e, of a number. The natural logarithm may also be represented
as "In" or 'log,."
B
Bq/
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4.5 MISCELLANEOUS SCREENING METHODS
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the
allowed flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified
in a regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to
be used by the analyst and the regulated community in making judgements necessary to meet
the data quality objectives or needs for the intended use of the data.
The following methods are included in this section:
Method 3820;
Method 8515:
Method 9074:
Method 9078:
Method 9079:
Hexadecane Extraction and Screening of Purgeable Organics
Colorimetric Screening Method for Trinitrotoluene (TNT) in
Soil
Turbidmetric Screening Method for Total Recoverable
Pertoleum Hydrocarbons in Soil
Screening Test Method for Polychlorinated Biphenyls in Soil
Screening Test Method for Polychlorinated Biphenyls in
Transformer Oil
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METHOD 9074
TURBIDIMETRIC SCREENING METHOD FOR TOTAL RECOVERABLE
PETROLEUM HYDROCARBONS IN SOIL
1.0 SCOPE AND APPLICATION
1.1 This method may be used to screen soil samples to determine the total amount of
recoverable petroleum hydrocarbon contamination in soil including a wide range of fuels, oils, and
greases. The turbidimetric approach in this method is designed to quickly screen soil samples using
a system calibrated with a blank and a single calibration standard.
1.2 The definition of total recoverable petroleum hydrocarbons for this method can be found
in the section on definitions (Sec. 3.0).
1.3 This screening technique is specifically designed to be used in the field but may also
have some screening applications in the laboratory. The system analysis range is 10-2000 ppm for
most hydrocarbons.
1.4 This method is considered a screening technique because of the broad spectrum of
hydrocarbons it detects. The method may be especially useful in quickly determining that a site does
not contain hydrocarbon contamination. However, it cannot be used to determine specific
hydrocarbon compounds or groups of compounds that may be part of a larger hydrocarbon mixture.
As with other screening techniques, it is advisable to confirm a certain percentage of both positive
and negative test results, especially when near or above a regulatory action limit or when the
presence of background or interfering hydrocarbons is suspected. The limitations of this procedure
are described in more detail in the section on interferences (Sec. 4.0).
1.5 This method does not address the evaporation of volatile petroleum hydrocarbon
mixtures {i.e. gasoline) during sample collection, preparation, and analysis. Although the screening
kit can be used to qualitatively detect volatile hydrocarbons, it is NOT recommended that the system
be used to quantitatively determine volatile petroleum hydrocarbons unless evaporation during
sample handling is addressed, appropriate response factor corrections are made, and method
performance is demonstrated on real world samples.
1.6 This method is restricted to use by or under the supervision of trained analysts. Each
analyst must demonstrate the ability to generate acceptable results with this method.
2.0 SUMMARY OF METHOD ...
2.1 A 10 ± 0.1 g sample of soil is extracted with a solvent mixture composed primarily of
methanol. The resulting mixture is allowed to settle and the free liquid is decanted into the barrel
of a filter-syringe assembly. The liquid is filtered through a 0.2-pm filter into a vial containing an
aqueous emulsifier development solution. The filtered sample is allowed to develop for 10 minutes.
During the development, any hydrocarbons present precipitate out arid become suspended in
solution.
2.2 The developed sample is placed in a turbidimeter that has been calibrated using a
blank and a single calibration standard. A beam of yellow light at 585 nm is passed through the
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sample arid the scattering of light through the suspension at 90° is measured. Hie concentration
of total recoverable petroleum hydrocarbons present is calculated relative to the standard curve.
3.0 DEFINITIONS
3.1 See Sec. 5.0 of Chapter 1 and the manufacturer's instructions for definitions associated
with this analytical procedure.
3.2 For the purpose of this method, "total recoverable petroleum hydrocarbons" is defined
as those hydrocarbons that are recovered using the solvent-specific extraction procedure provided
with this kit. Since there is no cleanup step to separate any co-extracted naturally occurring
hydrocarbons from the petroleum hydrocarbons, elevated turbidimetric readings are likely without
performing background correction. See the interferences section (Sec. 4.0) for additional details.
4.0 INTERFERENCES
4.1 This method is considered a screening technique because of the broad spectrum of
hydrocarbons it detects. It cannot distinguish between co-extracted naturally occurring hydrocarbons
and petroleum hydrocarbons. Using background correction and/or a selected response factor
discussed in the manufacturer's instructions, an analyst may be able to eliminate some of the
interferences caused by co-extracted naturally occurring hydrocarbons. However, it is very difficult
to find a truly clean, representative sample for use as a background.
4.2 This method has been shown to be susceptible to interference from vegetable oils
(positive interference). It is anticipated that co-extracted naturally occurring oils from vegetative
materials would be one of the most probable positive interferants found in the field. To demonstrate
this interference, standard soil samples were spiked with com oil at levels of 50 to 1000 ppm and
tested with PetroFLAG™ system. Soil samples spiked with mineral oil were also analyzed for
comparison. These data indicate that, over the range tested, the slope of the PetroFLAG™
vegetable oil response is approximately 18% of the response of the mineral oil standard. Supporting
data are presented in Table 2.
4.3 This method has been shown to be susceptible to interference from water (negative
interference). To demonstrate this interference, soils were spiked with diesel fuel at 100 ppm. The
samples were then spiked with varying amounts of water, up to saturation. The samples were
analyzed using the PetroFLAG™ system and the results were below that expected for the spike
added. The low bias may be due to a decrease in extraction efficiency in samples containing large
amounts of water, as a result of dilution of the extraction solvent. Supporting data are presented in
Table 3.
4.4 This method has been shown to NOT be significantly affected by up to 5% sodium
chloride contamination. Supporting data are presented in Table 6.
4.5 This method has been shown to NOT be significantly affected by up to 1000 ppm of
common surfactants such as trisodium phosphate (TSP), soap, and sodium dodecyf sulfate (SDS).
Supporting data are presented in Tables 7, 8, and 9.
4.6 Polycyclic aromatic hydrocarbons (PAHs) are a class of compounds present in many
hydrocarbon mixtures that are detected by the PetroFLAG system. These compounds are often
targeted because of their toxic characteristics and may be present individually as soil contaminants.
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However, the response of the individual PAHs varies greatly from compound to compound.
Therefore, use erf the PetroFLAG system to quantitate individual PAHs is not recommended without
good knowledge of the ate and after adjusting the analytical approach. Quantitation of PAHs as part
of a larger hydrocarbon fraction, such as diesel fuel, is recommended. Supporting data are
presented in Table 12.
4.7 The PetroFLAG™ analyzer can be used at temperatures from 4°C to 45°C. The
analyzer is equipped with an on-board temperature sensor to measure the ambient temperature at
which measurements are being made. The software uses this temperature reading to correct the
optical drift caused by temperature fluctuations.
4.8 Temperature at which the calibration is run should be recorded because of the effect
temperature has on the suspension. This can be done by taking a reading without inserting a vial.
If, during sample analysis, the temperature fluctuates more than ±10°C from the temperature at the
calibration, the calibration should be rerun at the new temperature.
5.0 SAFETY
Safety practices appropriate for handling potentially contaminated hazardous or toxic samples
and extraction solvents should be employed.
6.0 EQUIPMENT AND SUPPLIES
PetroFLAG™ Hydrocarbon Analysis System, (Dexsil Corporation, One Hamden Parte Drive,
Hamden, CT), or equivalent. Each commercially-available test kit will supply or specify the apparatus
and materials necessary for successful completion of the test.
7.0 REAGENTS AND STANDARDS
Each commercially-available test kit will supply or specify the reagents necessary for
successful completion of the test. Reagents should be labeled with appropriate expiration dates,
and reagents should not be employed beyond such dates.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 See the introductory material to this chapter, Organic Analytes, Sec. 4.1.
8.2 Soil samples may be contaminated, and should therefore be considered hazardous and
handled accordingly. All samples should be collected using a sampling plan that addresses the
considerations discussed in Chapter Nine.
8.3 To achieve accurate analyses, soil samples should be well homogenized prior to
testing. The hydrocarbons may not be evenly distributed in a soil sample and extensive mixing is
necessary to assure homogeneity.
NOTE: It is strongly recommended that any free aqueous liquid be decanted from samples prior
to analysis with the PetroFLAG system. Free aqueous liquid will dilute the extraction
solvent and produce a negative interference.
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NOTE: When users of the PetroFLAG system wish to report their results on a dry weight basis,
additional representative samples should be collected for percent moisture determination.
See the extraction Methods 3540 or 3550 for the procedure for determining percent
moisture.
9.0 QUALITY CONTROL
9.1 Follow the manufacturer's instructions for quality control procedures specific to the test
kit used. Additional guidance on qualify control is provided in Chapter One.
9.2 Use of replicate analyses, particularly when results indicate concentrations near the
action level, is recommended to refine information gathered with the kit.
9.3 Method 9074 is intended for use as a screening procedure in either the field or a fixed
laboratory. Wherever it is employed, a quality assurance program appropriate for a screening
procedure should be employed as a means of documenting the quality of the resulting data.
10.0 CALIBRATION AND STANDARDIZATION
See the PetroFLAG™ Hydrocarbon Analyzer User's Manual for instruction on generating an
initial calibration curve using the PetroFLAG™ analyzer. Contact the manufacturer for specific
details on the calibration calculations programmed into the PetroFLAG™ analyzer.
11.0 PROCEDURE
Follow the manufacturer's instructions in the PetroFLAG™ Hydrocarbon Analyzer User's
Manual to extract, develop, and analyze soil samples. Those test kits used must meet or exceed
the performance specifications indicated in Tables 1 through 3.
12.0 DATA ANALYSIS AND CALCULATIONS
Consult the PetroFLAG™ Hydrocarbon Analyzer User's Manual for the procedure used to
generate concentration readings from samples using the PetroFLAG™ analyzer. Contact the
manufacturer for specific details on the concentration calculations programmed into the
PetroFLAG™ analyzer.
13.0 METHOD PERFORMANCE
13.1 Method Detection Limits were determined using a modification of the procedures in
Chapter One and in 40 CFR, Part 136. The procedure was modified slightly because the instrument
automatically subtracts an average blank value for each analysis (blank analysis is part of the
calibration procedure of the PetroFLAG™ test system). Two sets of seven samples each were
prepared, one set spiked with 30 ppm of diesel fuel, and one set spiked with 30 ppm of used motor
oil. The standard deviation (SD) of the results for each oil type were calculated. The method
detection limit (MDL) was determined by multiplying the SD by the Student's t value (3.143). These
data are presented in Table 1. The MDL for diesel fuel was 13 ppm and for used motor oil was 18.6
ppm (Ref 1).
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13.2 Samples of a standard soil were prepared by spiking with either diesel fuel or used
motor oil at 100 ppm intervals from 100 ppm to 1000 ppm. Each sample was analyzed in duplicate
by the PetroFLAG™ system and by Methods 3550 and 8015B. The results are shown in Table 4.
These data were analyzed using regression analysis. The results of the regression analysis are also
provided in Table 4. In addition, an analysis of variance (ANOVA) analysis was performed. The F-
statistic from the ANOVA revealed a significant bias between the two methods, with the
PetroFLAG™ providing consistently higher values for both types of contamination. The results
confirm that the kit design is intentionally conservative, in that it favors a high bias in order to avoid
reporting false negative results (Ref. 1).
13.3 Precision and bias were determined by analysis of variance (ANOVA) of the results
obtained from spiked soil samples. Four sets of spiked samples were prepared, containing either
diesel fuel or used motor oil at two different concentrations (200 and 1000 ppm). Each analyte at
each concentration was analyzed in duplicate 10 times (e.g., 20 replicates of each). The results
were transformed into recovery data. The ANOVA used these transformed data. The results are
presented in Table 5. The F-statistic for the diesel fuel analysis indicate a slight day effect for these
samples. The F-statistic seems to be driven more by the very low value of'the mean square error
within days rather than by any large value for the mean square error between days (Ref. 1).
13.4 The response of the PetroFLAG System to a soil spiked with 500 ppm ofdiesei fuel and
0 to 5% of dry sodium chloride is provided in Table 6 (Ref. 2).
13.5 The responses of the PetroFLAG System to a soil spiked with 500 ppm of diesel fuel
and up to 1000 ppm of common surfactants such as trisodium phosphate (TSP), soap, and sodium
dodecyi sulfate (SDS), are presented in Tables 7, 8, and 9 (Ref. 2).
13.6 Performance of the PetroFLAG™ system on anthracene from 100 to 2000 ppm and on
creosote from 100 to 1000 ppm are presented in Tables 10 and 11, respectively. An explanation of
the erratic performance of anthracene is provided in the Table 10 narrative (Ref. 2).
13.7 The performance of the PetroFLAG system for several PAHs relative to the mineral oil
calibrator on soil is presented in Table 12 (Ref. 4).
13.8 Performance of the PetroFLAG™ system on Jet-A from 40 to 2808 ppm (Ref. 4) and
on gasoline from 1000 to 4070 ppm (Ref. 2) are provided in Tables 13 and 14, respectively. An
explanation of the performance of Jet-A and gasoline are provided in the narrative in Tables 13 and
14.
14.0 POLLUTION PREVENTION
This method does not use any halogenated solvents and may be used to help reduce the
number of samples sent to the laboratory under certain project scenarios. Traditional laboratory
extraction methods (i.e. Soxhlet or sonication) would generally require much larger volumes of
solvent to extract the sample.
15.0 WASTE MANAGEMENT
Waste management procedures must be consistent with federal, state, and local regulations.
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16.0 REFERENCES
1. Data Validation Package, Testing for Petroleum Hydrocarbons in Soil by Turbimetric Analysis,
PetroFLAG™ Test System, DEXSILGorp., Hamden, CT.
2. Supplementary Validation Data, Additional Analyte and Contaminant Testing Data tor the
PetroFLAG™ Hydrocarbon Analysis System, DEXSIL Corp., Hamden, CT, August 24,1995.
3. PetroFLAG1* Hydrocarbon Analyzer User's Manual, DEXSIL Corp., Hamden, CT.
4. Supplementary Data Validation Package III, Additional Analyte Testing Data for Petroleum
Hydrocarbons in Soil by Turbimetric Analysis - PetroFLAG™ Test System, DEXSIL Corp;,
Hamden, CT, June 20,1997.
5. Supplementary Data VaHdation Package IV, Polycydic Aromatic Hydrocarbon Response data
for Method 9074 Petroleum Hydrocarbons in Soil by Turbimetric Analysis - PetroFLAGm Test
System, DEXSIL Corp., Hamden, CT, August 22,1997.
17. TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Tables 1 through 14.
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TABLE 1
METHOD DETECTION LIMIT FOR PetroFLAG TEST SYSTEM
Trial#
30 ppm diesel fuel
30 ppm motor oil
1
34
35
2
24
41
3
28
40
4
34
53
5
36
46
6
32
48
7
30
42
Average (ppm)
31.03
43.6
SD (ppm)
4.12
5.91
MDL (ppm)
13.0
18.6
Data from Reference 1.
TABLE 2

RELATIVE RESPONSE OF VEGETABLE OILS AS AN INTERFERANT
Analyte Spike
Concentration (ppm)
Mineral Oil
Response (ppm)
Vegetable Oil
Response® (ppm)
50
55
30
100
100
45
200
189
94
500
504
111
1000
947
208
a The vegetable oil samples were analyzed using the PetroFLAG system set to response
factor 10. The slope of the PetroFLAG vegetable oil response is approximately 18% of the
response of the mineral oil standard. This means that a sample containing 5,560 ppm
vegetable oil would provoke a response equivalent to that given by 1,000 ppm mineral oil.
Data from Reference 1.
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TABLE 3
EFFECT OF WATER ON PetroFLAG RESULTS
% Water Saturation (% Water)
% Recovery of Mineral Oil*
0(0)
100
5(1)
94
25(5)
98
50(10)
95
100 (20)
85
1 Soil sample spiked with 100 ppm of mineral oil. (Ref. 1)
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TABLE 4
COMPARISON OF PetroFLAG AND GC TEST RESULTS
Spike Cone.

PetroFLAG
3550/8015B
(MB/9)

(M0/g)

(M9/0)

Diesel Fuel
Trial 1

Trial 2 •
Trial 1

Trial 2
100
112

116
73

82
200
230

248
158

156
300
312

370
242

218
400
420

455
299

275
500
538

564
342

344
600
626

654
460

439
700
774

790
509
'
494
800
910

900
612

607
900
1091

977
678

614
1000
1182

1062
646

649
Corr Coef

0.999

0.992

Slope

1.126

0.679

Intercept

-2.8

30.5

Motor Oil
Trial 1

Trial 2
Trial 1

Trial 2
100
121

128
123

82
200
243

292
200

200
300
381

408
301

275
400
428

497
341

343
500
531

554
441

452
600
654

668
534

528
700
717

771
609

652
800
880

883
711

746
900
931

1052
835

881
1000
1014

1098
887

846
Corr Coef

0.998

0.997

Slope

1.02

0.887

Intercept

50.9

20.5

Data from Reference 1.
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TABLE 5
ANOVA RESULTS FOR SPIKED PETROLEUM HYDROCARBON SAMPLES

Mean
Variance
Standard
Standard
Analyte/Concentration
n
<*>
(%i2)
Deviation (%,)
Error (
-------
TABLE 8
RESPONSE OF PetroFLAG SYSTEM WITH VARIOUS SOAP CONCENTRATIONS®
Soap Concentration (ppm)

0
100
200
500
1000
PetroFLAG Response (ppm)
500
494
488
502
528
a Response of the PetroFLAG system for soil containing 500 ppm of dieset fuel and various levels
of soap (non-ionic and anionic surfactants). The samples were analyzed using the PetroFLAG
system set to response factor 5 (Ref. 2).
TABLE 9
RESPONSE OF PetroFLAG SYSTEM WITH VARIOUS SDS CONCENTRATIONS3
SDS Concentration (ppm)

0
100
200
500
1000
PetroFLAG Response (ppm)
472
474
488
486
496
a Response of the PetroFLAG system for soil containing 500 ppm of diesel fuel and various
levels of sodium dodecyl sulfate, a surfactant. The samples were analyzed using the
PetroFLAG system set to response factor 5 (Ref. 2).
TABLE 10
RESPONSE OF PetroFLAG SYSTEM WITH VARIOUS AMOUNTS OF ANTHRACENE8
Anthracene Cone, (ppm)

100
200
500
1000
2000
PetroFLAG Response (ppm)
798
1376
1641
1380
1735
3 Response of the PetroFLAG system for soil containing various levels of anthracene. The results
show that the PetroFLAG system returns a strong response to anthracene. The response to
anthracene is higher than response to the calibrator, therefore, the meter displays a reading over-
estimating the concentration. For concentrations greater than 200 ppm, the turbidity developed
exceeds the recommended level (i.e. a reading greater than 1000 on response factor 10). To
obtain accurate results the user should rerun the sample using a smaller sample size. This will
bring the results into linear range. The samples were analyzed using the PetroFLAG system set
to response factor 10 (Ref. 2).
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TABLE 11
RESPONSE OF PetroFLAG SYSTEM WITH VARIOUS AMOUNTS OF CREOSOTE*
Creosote Cone, (ppm)

100 200
500
1000
PetroFLAG Response (ppm)
103 210
538
1043
8 Response of the PetroFLAG system for soil containing various levels of creosote. The samples
were analyzed using the PetroFLAG system set to response factor 8 (Ref. 2).
TABLE 12
RELATIVE RESPONSE OF PetroFLAG SYSTEM TO VARIOUS
POLYCYCLIC AROMATIC HYDROCARBONS3
Compound
Spike Level in ppm
(Matrix Used)
PetroFLAG Reading
in ppm (Rf 10)
Response Relative to
Mineral Oil Calibrator
Anthracene
100 (Soil)
798
8
Benzo[a]pyrene
50 (Soil)
180
3.6
Chrysene
16 (Solvent)
172
11
Fluoranthene
200 (Solvent)
101
0.5
Pyrene
200 (Solvent)
216
1.1
* The data for anthracene and benzo(a)pyrene were generated by spiking each compound onto a
composite sandy clay loam soil and homogenizing the sample for later analysis. The soil sample
size was 10 g. The soil spiking procedure used for anthracene and benzo(a)pyrene produced
inconsistent results for the other PAH compounds. These compounds (chrysene, flouranthene,
and pyrene), which are very soluble in the extraction solvent, were spiked directly into the
extraction solvent and analyzed. All of the PAHs samples were analyzed on response factor 10
(the correct response factor for mineral oil). The data indicate that, for example, using a standard
sample size analyzed on response factor 10 (the correct response factor for mineral oil), a 100
ppm anthracene sample read 798 ppm. The PetroFLAG response to the above analytes is equal
to or greater than the calibrator in all cases except for fluoranthene which has a response
equivalent to diesel fuel.
NOTE: When analyzing soils containing anthracene, benzo(a)pyrene, or chrysene the PetroFLAG
meter will read over range for concentrations of 250, 550, and 180 ppm respectively. These soils
can be analyzed using a 1 gram sample size to increase the ma>dmum quantifiable concentration.
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TABLE 13
RESPONSE OF PetroFLAG SYSTEM WITH VARIOUS AMOUNTS OF JET-A"
Jet-AConc. (ppm)
0 40 79 198 397 793 1586 2776
PetroFLAG Response (ppm) 54 110 162 208 368 700 1592 2808
a Response of the PetroFLAG system for soil containing various levels of Jet-A. The composite
soils were prepared from two types of clay-loam soil and sand. The component soils were air dried
and sieved to remove particles larger than 850 jjm and then mixed in the ratio 2:1:1, followed by
tumbling for one hour. The soil was weighed out into 10 g aiiquots. Each of the soil aliquot* was
spiked by direct injection of Jet-A fuel onto the soil using a microliter syringe, mixed, and analyzed
by the PetroFLAG system with the instrument set to response factor-4. The coefficient of
determination (r2) for the Jet-A data was 0.997, indicating that the PetroFLAG response was linear
over the range 40 ppm to 2808 ppm (Ref. 4).
TABLE 14
RESPONSE OF PetroFLAG SYSTEM WITH VARIOUS AMOUNTS
OF WEATHERED GASOLINE3
Weathered Gasoline Cone, (ppm)
1000 2040 3050 4070
PetroFLAG Response (ppm) 285 1780 4335 6870
a Response of the PetroFLAG system for soil containing various levels of weathered gasoline (50%
evaporated). The manufacturer recommends that PetroFLAG be used to qualitatively detect
gasoline at these levels. It is not recommended that PetroFLAG be used quantitatively for gasoline
unless significant response factor corrections are made and evaporation of the target
hydrocarbons is addressed. The samples were analysed using the PetroFLAG system set to
response factor 2 (Ref. 2).
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CHAPTER FIVE
MISCELLANEOUS TEST METHODS
Prior to employing the methods in this chapter, analysts are advised to consult the disclaimer
statement at the front of this manual and the information in Chapter Two for guidance on the allowed
flexibility in the choice of apparatus, reagents, and supplies. In addition, unless specified in a
regulation, the use of SW-846 methods is not mandatory in response to Federal testing
requirements. The information contained in each procedure is provided by EPA as guidance to be
used by the analyst and the regulated community in making judgements necessary to meet the data
quality objectives or needs for the intended use of the data.
The following methods are found in Chapter Five:
Method
5Q5Q:
Method
9010B:
Method
9012A:
Method
9013:
Method
9014:
Method
9020B:
Method
9021:
Method
9022:
Method
9023:
Method
9030B:
Method
9031:
Method
9034:
Method
903S:
Method
9036:
Method
9038:
Method
90S6:
Method
9057:
Method
9060:
Method
9065:
Method
9066:
Method
9067:
Method
9070:
Method
9071A:
Method
9075:
Method
9076:
Bomb Preparation Method for Solid Waste
Total and Amenable Cyanide: Distillation
Total and Amenable Cyanide (Automated Colorimetric, with
Off-Line Distillation)
Cyanide Extraction Procedure for Solids and Oils
Titrimetric and Manual Spectrophotometry Determinative
Methods for Cyanide
Total Organic Halides (TOX)
Purgeable Organic Halides (POX)
Total Organic Halides (TOX) by Neutron Activation Analysis
Extractable Organic Halides (EOX) in Solids
Acid-Soluble and Acid-Insoluble Sulfides: Distillation
Extractable Sulfides
Titrimetric Procedure for Acid-Soluble and Acid-Insoluble
Sulfides
Sulfate (Colorimetric, Automated, Chloranilate)
Sulfate (Colorimetric, Automated, Methylthymol Blue, AAII)
Sulfate (Turbidimetric)
Determination of Inorganic Anions by Ion Chromatography
Determination of Chloride from HC1/CI2 Emission Sampling
Train (Methods 0050 and 0051) by Anion Chromatography
Total Organic Carbon
Phenolics (Spectrophotometry, Manual 4-AAP with
Distillation)
Phenolics (Colorimetric, Automated 4-AAP with Distillation)
Phenolics (Spectrophotometric, MBTH with Distillation)
Total Recoverable Oil & Grease (Gravimetric, Separatory
Funnel Extraction)
Oil and Grease Extraction Method for Sludge and Sediment
Samples
Test Method for Total Chlorine in New and Used Petroleum
Products by X-Ray Fluorescence Spectrometry (XRF)
Test Method for Total Chlorine in New and Used Petroleum
Products by Oxidative Combustion and Microcoulometry
FIVE -1
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Method 9077:
Method A:
Method B:
Method C:
Method
9131
Method
9132
Method
9210
Method
9211
Method
9212
Method
9213
Method
9214
Method
9215
Method
9216
Method
9250
Method
9251
Method
9253
Method
9320
Test Methods for Total Chlorine in New and Used Petroleum
Products (Field Test Kit Methods)
Fixed End Point Test Kit Method
Reverse Titration Quantitative End Point Test Kit
Method
Direct Titration Quantitative End Point Test Kit Method
Total Coliform: Multiple Tube Fermentation Technique
Total Coliform: Membrane-Filter Technique
Potentiometric Determination of Nitrate in Aqueous Samples
with Ion-Selective Electrode
Potentiometric Determination of Bromide in Aqueous Samples
with Ion-Selective Electrode
Potentiometric Determination of Chloride in Aqueous Samples
with Ion-Selective Electrode
Potentiometric Determination of Cyanide in Aqueous Samples
and Distillates with Ion-Selective Electrode
Potentiometric Determination of Fluoride in Aqueous Samples
with ion-Selective Electrode
Potentiometric Determination of Sulfide in Aqueous Samples
and Distillates with Ion-Selective Electrode
Potentiometric Determination of Nitrate in Aqueous Samples
with Ion-Selective Electrode
Chloride (Colorimetric, Automated Ferricyanide AAI)
Chloride (Colorimetric, Automated Ferri cyanide AAI I)
Chloride (Titrimetric, Silver Nitrate)
Radium-228
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METHOD 9216
FOTENTIOMETRIC DETERMINATION OF NITRITE
IN AQUEOUS SAMPLES WITH ION-SELECTIVE ELECTRODE
1.0 SCOPE AND APPLICATION
1.1 This method can be used for measuring nitrite in drinking water, wastewater, and
reagent waters. If this method is used for other types of water samples (i.e., surface water, ground
water, etc.), method precision and accuracy must be demonstrated for each matrix type.
1.2 The method detection limit is 0.05 mg/L of nitrite as nitrogen. Nitrite concentrations
from 0.05 to 20 mg/L may be measured.
1.3 Ion selective electrodes (ISEs) must be used carefully and results must be interpreted
cautiously. An ISE may be affected by numerous analytical interferences which may either increase
or decrease the apparent analyte concentration, or which may damage the ISE. Effects of most
interferences can be minimized or eliminated by adding appropriate chemical reagents to the
sample. Obtaining the most accurate results, therefore, requires some knowledge of the sample
composition.
NOTE: Manufacturers usually include a list of interferences in the instruction manual
accompanying an ISE, along with recommended methods for minimizing or eliminating effects
of these interferences.
2.0 SUMMARY OF METHOD
2.1 This method uses a nitrite-selective electrode. All standards and samples are mixed
with an equal volume of nitrite interference suppressor solution (NISS). A calibration curve is
constructed by recording the nitrite calibration standard readings using an appropriate meter or by
manual plotting. Samples are then read in the same manner, and the concentrations reported by
the meter or read from the graph.
3.0 DEFINITIONS
Refer to Chapter Three for the applicable definitions.
4.0 INTERFERENCES
4.1 Some anions, if present at high enough levels, are electrode interferences and will
cause measurement errors. Table 1 displays the levels of possible interferences causing a 10%
error. NISS is mixed in an equal volume with standards as well as with samples. For example, 25
mL of sample would be mixed with 25 mL of NISS. This procedure ensures that samples and
standards are properly buffered, have a similar background and that no correction factor is needed
for the dilution. Figure 1 shows how the nitrite electrode response changes with pH. This is
compensated for by the addition of the NISS. Selectivity is mathematically demonstrated by the
following equation:
E - E' + s log[c, + £Kjj c,23]
Where: E = Reference potential
9216-1
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s = Slope
Cj = Primary ion concentration
Kjj = Selectivity Coefficient
^^Interfering ion concentration
Zj = charge ratio of interfering ion
Successful analytical conditions depend upon:
Cj» JKjj c,25
4.2 Temperature changes affect electrode potentials. Therefore, standards and samples
must be equilibrated at the same constant temperature (± 1 °C).
CAUTION: Use hood to avoid exposure to toxic gases released during acidification,
4.3 The user should be aware of the potential of interferences from colloidal substances
and that, if necessary, the samples may be filtered.
4.4 Standard electrode filling solutions containing high levels of KCI should not be used as
the reference electrode filling solution.
4.5 If electrodes are exposed to samples with nitrite concentrations greater than 20 mg/L,
their response may become very sluggish when again measuring at a lower concentration. If this
occurs, soak the electrodes for 8-12 hours in a mixture of the 0.5 mg/L standard and NISS.
5.0 SAFETY
5.1 Refer to Chapter Three for additional guidance on safety protocols.
5.2 it is the responsibility of the user to prepare, handle, and dispose of electrolyte solutions
in accordance with all applicable federal, state, and local regulations.
6.0 EQUIPMENT AND SUPPLIES
6.1 A pH/mV meter capable of reading to 0.1 mV or an ISE meter.
6.2 Nitrite ISE (Orion 93-46 or equivalent) and double-junction reference electrode (Orion
90-01 or equivalent).
6.3 Thermally isolated magnetic stirrer, fluorocarbon (PFA or TFM)-coated magnetic stir
bar, and stopwatch.
6.4 Volumetric flasks, 100 mL and 1 L - Class A.
6.5 Volumetric pipets, 5 mL, 10 mL and 50 mL - Class A
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7.0 REAGENTS AND STANDARDS
7.1 Reagent grade chemicals shall be used in all tests. Unless otherwise indicated, it is
intended that all reagents shall conform to the specifications of the Committee on Analytical
Reagents of the American Chemical Society, where such specifications are available. Other grades
may be used, provided it is first ascertained that the reagent is of sufficiently high purity to permit its
use without lessening the accuracy of the determination.
7.2 Reagent water. All references to water in this method refer to reagent water, as defined
in Chapter One.
7.3 ISA adjuster solution (2M, (NH4)2SO<): Dissolve 26.4 g of ammonium sulfate in reagent
water to make 100 mL of solution.
7.4 Nitrite reference electrode filling solution (Orion 900046 or equivalent).
7.5 Nitrite interference suppressor solution (NISS) (Orion 934610 or equivalent).
7.6 1000 mg/L nitrite as N, stock standard - Weigh out 4.93 g of ACS reagent grade sodium
nitrite that has been dried for 24 hours in a desiccator. Place in a clean one L volumetric flask. Add
approximately 200 mL of reagent water and mix to dissolve. Add two drops of NaOH and make to
volume. Mix by inverting 20 times.
7.7 100 mg/L nitrite as N, stock standard - Pipet 10.0 mL of the 1000 mg/L standard into
a clean 100 mL volumetric flask. Make to volume and mix well. Replace this standard monthly.
7.8 10 mg/L nitrite as N, stock standard - Pipet 10.0 mL of the 100 mg/L standard into a
dean 100 mL volumetric flask- Make to volume and mix well. Replace this standard weekly.
7.9 5 mg/L nitrite as N, stock standard - Pipet 50.0 mL of the 10 mg/L standard solution into
a clean 100 mL volumetric flask. Make to volume with reagent water and mix well. This standard
should be replaced daily.
7.10 1 mg/L nitrite as N, stock standard - Pipet 10.0 mL of the 10.0 mg/L standard into a
clean 100 mL volumetric flask. Make to volume with reagent water and mix well. This standard
should be replaced daily.
7.11 0.5 mg/L nitrite as N, stock standard - Pipet 5.00 mL of the 10.0 mg/L standard into a
clean 100 mL volumetric flask. Make to volume with reagent water and mix well. This standard
should be replaced daily.
8.0 SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 All samples must have been collected using a sampling plan that addresses the
considerations discussed in Chapter Nine of this manual.
8.2 Samples should be stored at 4 °C and must be analyzed within 48 hours of collection.
9.0 QUALITY CONTROL
9.1 Refer to Chapter One for specific quality control procedures.
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9.2 Initial calibration verification standard (ICV): After performing the calibration step
(Section 10.0), verify calibration by analyzing an ICV. The ICV contains a known nitrite concentration
at the mid-range of the calibration standards and is from an independent source. ICV recovery must
be 90-110 percent. If not, the source of error must be found and corrected. An acceptable ICV must
be analyzed prior to sample analysis. The ICV also serves as a laboratory control sample.
9.3 Continuing calibration verification standard (CCV): After every 10 samples, and after
the final sample, a CCV must be analyzed. The CCV contains a known nitrite concentration at mid-
calibration range. CCV recovery must be 90-110 percent. If not, the error source must be found and
corrected. If I8E calibration has changed, all samples analyzed since the last acceptable CCV must
be re-analyzed.
9.4 Reagent blank: After the ICV and after every CCV, a reagent blank must be analyzed.
A reagent blank is 25 mL of reagent water with 25 mL of NISS added. The indicated reagent blank
concentration must be less than 1 mg/L nitrite. If not, the contamination source must be found and
corrected. Ail samples analyzed since the last acceptable reagent blank must be re-analyzed.
9.5 Matrix spike: Follow the matrix spike protocols presented in Chapter One. The spike
concentration must be 10 times the detection limit and the volume added must be negligible (less
than or equal to one-thousandth the sample aliquot volume). Spike recovery must be 75-125
percent. If not, samples must be analyzed by the method of standard additions.
10.0 CALIBRATION AND STANDARDIZATION
10.1 When using a nitrate ISE and a separate double-junction reference electrode, ensure
that reference electrode inner and outer chambers are filled with solutions recommended by the
manufacturer. Equilibrate the electrodes for at least one hour in a 100 mg/L nitrite standard before
use.
10.2 Calibrate the nitrite ISE using standards that narrowly bracket the expected sample
concentration. If the sample concentration is unknown, calibrate with 0.5 mg/L, 1.0 mg/L, and 5.0
mg/L nitrite standards Add 25.0 mL of a standard solution and 25 mL of the NISS into a 100 mL
beaker to make each calibration standard.
10.3 Add a fluorocarbon (PFA or TFM)-coated magnetic stir bar, place the beaker on a
magnetic stir plate, and stir at slow speed (no visible vortex). Immerse the electrode tips to just
above the rotating stir bar. If using an ISE meter, calibrate the meter in terms of nitrite concentration
following the manufacturer's instructions. If using a pH/mV meter, record the meter reading (mV)
as soon as the reading is stable, but in no case should the time exceed five minutes after immersing
the electrode tips.
10.4 Prepare a calibration curve by plotting measured potential (mV) as a function of the
logarithm of nitrite concentration. For corrective action, consult the ISE operating manual.
11.0 PROCEDURE
11.1 Allow samples and standards to equilibrate to room temperature.
11.2 Prior to and between analyses, rinse the electrodes thoroughly with reagent water and
gently shake off excess water. Low-level measurements are faster if the electrode tips are first
immersed five minutes in reagent water.
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11.3 Add 25.0 mL of sample and 25.0 mL of NISS to a 100-mL beaker. Add a fluorocarbon
(PFA orTFM)-coated magnetic stir bar. Place the beaker on a magnetic stir plate and stir at a slow
speed (no visible vortex). Immerse the electrode tips to just above the rotating stir bar. Record the
meter reading (mV or concentration) as soon as the reading is stable, but in no case should the time
exceed five minutes after immersing the electrode tips. If reading mV, determine nitrite-nitrogen
concentration from the calibration curve.
11.4 When analyses have been completed, rinse the electrodes thoroughly and store them
in a 100 mg/L nitrate standard solution. If the electrodes will not be used more than one day, drain
the reference electrode internal Ailing solutions, rinse with reagent water, and store dry.
12.0 DATA ANALYSIS AND CALCULATIONS
Results must be reported in units commensurate with their intended use and all dilutions must
be taken into account when computing final results.
13.0 METHOD PERFORMANCE
13.1 Figure 2 displays a typical calibration curve for nitrite at 0.5 mg/L, 1 mg/L, and 5 mg/L.
Figure 3 displays a low level calibration curve for nitrite at 0.05 mg/L, 0.1 mg/L, 0.2 mg/L, and 0.5
mg/L
13.2 Table 1 displays the levels at which known interferences may impact the analysis.
Refer to Sec. 4.0 for a discussion on interferences.
13.3 The following documents may provide additional guidance and insight on this method
and technique:
13.3.1 "Determination of Nitrite in Foods and Wastewater Using a Nitrite-Selective
Electrode", S.J.West, X.Wen, M.S.Frant, NAChaniotakis, Pittsburgh Conference, March 1994.
13.3.2 "Determination of Nitrate, Nitrite, and Ammonia in Advanced Secondary
Effluent by Means of Ion-Selective Electrodes", S.J.West, X.Wen, Pittsburgh Conference,
March 1994.
13.3.3 Model 93-46 Nitrite Electrode Instruction Manual, ATI Orion, Boston MA,
1994.
14.0 POLLUTION PREVENTION
14.1 Pollution prevention encompasses any technique that reduces.or eliminates the quantity
and/or toxicity of waste at the point of generation. Numerous opportunities for pollution prevention
exist in laboratory operation. The EPA has established a preferred hierarchy of environmental
management techniques that places pollution prevention as the management option of first choice.
Whenever feasible, laboratory personnel should use pollution prevention techniques to address their
waste generation. When wastes cannot be feasibly reduced at the source, the Agency recommends
recycling as the next best option.
14.2 For information about pollution prevention that may be applicable to laboratories and
research institutions consult Less is Better Laboratory Chemical management for Waste Reduction
available from the American Chemical Society's Department of Government Relations and Science
Policy, 1155 16th St., N.W. Washington, D.C. 20036, (202) 872-4477.
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15.0 WASTE MANAGEMENT
The Environmental Protection Agency requires that laboratory waste management practices
be conducted consistent with all applicable rules and regulations. The Agency urges laboratories
to protect the air, water, and land by minimizing and controlling all releases from hoods and bench
operations, complying with the letter and spirit of any sewer discharge permits and regulations, and
by complying with all solid and hazardous waste regulations, particularly the hazardous waste
identification rules and land disposal restrictions. For further information on waste management,
consult The Waste Management Manual for Laboratory Personnel available from the American
Chemical Society at the address fisted in Sec. 14.2.
16.0 REFERENCES
1. Applications Laboratory Report, Tests in Water Samples by Nitrite Electrode and 'Standard
Methods' Colorimetric Analysis", ATI Orion, Boston MA, April 1995.
17.0 TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
The pages to follow contain Table 1, Figures 1 through 3, and a flow diagram of the method
procedures.
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TABLE 1
NITRITE ELECTRODE SELECTIVITY DATA
Interfering Ion
log Kjj
10% Error Ratio (ppm)
Hydroxide
2.8
-
Fluoride
-3.1
170
Chloride
-3.1
320
* Chlorate
-3.4
1600
Perchlorate
-3.1
830
Bromide
o
CO
1
570
Iodide
-1.2
15
Sulfate
-4.1
1100
Nitrate (N)
-3,3
200
Phosphate
-4.0
9500
Polyphosphate
-4.4
3400
Bicarbonate
-3.3
870
Acetate
-3.2
720
Lactate
-4.9
Very high
Phthaiate
-2.5
380
Ascorbate
-4.2
Very high
Salicylate
-0.8
7.0
Source: Reference 1.
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FIGURE 1
NITRITE ELECTRODE pH RESPONSE
|vp||&5 ^>||f§Ss S«
¦•• ><;•>' :\S..
miglpg
tmmmmwm
f FffcpPRfHsKS =-«6':^,:-
mxmmwm
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FIGURE 2
CALIBRATION CURVE FOR STANDARD LEVEL OF NITRITE
150
140-
130
> 120
110-
100-
0.1
Nitrite, mg/L as N
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FIGURE 3
CALIBRATION CURVE FOR LOW LEVEL NITRITE METHOD
200
190
180
> 170
160
150
140
0.1
Nitrite, mg/L as N
0.01
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METHOD 9216
POTENTIOMETRIC DETERMINATION OF NITRITE
m AQUEOUS SAMPLES WITH ION-SELECTIVE ELECTRO PE
Start
11.1 Allow Standards
and samples to
equilibrate to
room temperature.
11.2 Rinse electrode
with reagent water.
11.4 Drain reference
electrode and clean.
10.1 - 10.2
Calibrate
Nitrite ISE,
11.3 Add 25 mL sample
and 25 mL NISS ts beaker,
and measure concentration
using electrode meter
and calculate concentration.
fi U. S. GOVERNiMEW PRINTING OFFICE; 1998-439-410/90222
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