SW 846.3.4A
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, II, HA, HE, and ffl. 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 H).
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 II/IIA, a copy of Final Update IIB, a copy
of Update III, 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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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 QA 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
Method 3500B: Organic Extraction and Sample Preparation
Method 351OC: Separatory Funnel Liquid-Liquid Extraction
Method 3520C: Continuous Liquid-Liquid Extraction
Method 3535A: Solid-Phase Extraction (SPE)
Method 3540C: Soxhlet Extraction
Method 3541: Automated Soxhlet Extraction
Method 3542: Extraction of Semivolatile Analytes Collected Using Method 0010
(Modified Method 5 Sampling Train)
Method 3545A: Pressurized Fluid Extraction (PFE)
Method 3550B: Ultrasonic Extraction
Method 3560: Supercritical Fluid Extraction of Total Recoverable Petroleum
Hydrocarbons
Method 3561: Supercritical Fluid Extraction of Polynuclear Aromatic Hydrocarbons
Method 3562: Supercritical Fluid Extraction of Polychlorinated Biphenyls (PCBs) and
Organochlorine Pesticides
Method 3580A: Waste Dilution
Method 3585: Waste Dilution for Volatile Organics
Method 5000: Sample Preparation for Volatile Organic Compounds
Method 5021: Volatile Organic Compounds in Soils and Other Solid Matrices Using
Equilibrium Headspace Analysis
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4.3.5 Miscellaneous Spectrometric Methods
Method 8520: Continuous Measurement of Formaldehyde in Ambient Air
4.4 Immunoassay Methods
Method 4000: Immunoassay
Method 401OA: Screening for Pentachlorophenol by Immunoassay
Method 4015: Screening for 2,4-Dichlorophenoxyacetic Acid by Immunoassay
Method 4020: Screening for Polychlorinated Biphenyls by Immunoassay
Method 4030: Soil Screening for Petroleum Hydrocarbons by Immunoassay
Method 4035: Soil Screening for Polynuclear Aromatic Hydrocarbons by
Immunoassay
Method 4040: Soil Screening for Toxaphene by Immunoassay
Method 4041: Soil Screening for Chlordane by Immunoassay
Method 4042: Soil Screening for DDT by Immunoassay
Method 4050: TNT Explosives in Soil by Immunoassay
Method 4051: Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) in Soil by Immunoassay
Method 4670: 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 TWO
DISCLAIMER
ABSTRACT
TABLE OF CONTENTS
METHOD INDEX AND CONVERSION TABLE
PREFACE
CHAPTER ONE, REPRINTED - QUALITY CONTROL
1.0 Introduction
2.0 QA Project Plan
3.0 Field Operations
4.0 Laboratory Operations
5.0 Definitions
6.0 References
PART III 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:
Modified Method 5 Sampling Train
Preparation of XAD-2 Sorbent Resin
Total Chromatographable Organic Material Analysis
Sampling for Selected Aldehyde and Ketone Emissions from
Stationary Sources
Source Assessment Sampling System (SASS)
Sampling Method for Polychlorinated Dibenzo-p-Dioxins and
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/CI2 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
CONTENTS -11
Revision 5
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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 Sludge 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. Centrifugation: 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 semivolatile 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 Analytes
All atomic absorption analyses should employ appropriate background correction systems
whenever spectral interferences could be present. Several background correction techniques are
employed in modern 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 verify 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.)
Element(s) Modifier(s)
As and Se Nickel nitrate, palladium
Pb Phosphoric acid, ammonium phosphate, palladium
Cd Ammonium phosphate, palladium
Sb Ammonium nitrate, palladium
Tl Platinum, palladium
ICP. AA, and GFAA calibration standards must match the acid composition and strength of
the acids 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 Water"; Ground Water 1984, 22(1). 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 by Ion Chromatoqraphv:
(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
Acenaphthylene 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
Allyl alcohol 8015, 8260
Allyl chloride 8021, 8260
2-Aminoanthraquinone 8270
Aminoazobenzene 8270
4-Aminobiphenyl 8270
Aminocarb 8321
2-Amino-4,6-dinitrotoluene (2-Am-DNT) 8330
4-Amino-2,6-dinitrotoluene (4-Am-DNT) 8330
3-Amino-9-ethylcarbazole 8270
Anilazine 8270
Aniline 8131, 8270
o-Anisidine 8270
Anthracene 8100, 8270, 8275, 8310, 8410
Aramite 8270
Aroclor-1016 (PCB-1016) 8082, 8270
Aroclor-1221 (PCB-1221) 8082, 8270
Aroclor-1232 (PCB-1232) 8082, 8270
Aroclor-1242 (PCB-1242) 8082, 8270
Aroclor-1248 (PCB-1248) 8082, 8270
Aroclor-1254 (PCB-1254) 8082, 8270
Aroclor-1260 (PCB-1260) 8082, 8270
Aspon 8141
Asulam 8321
Atrazine 8141
Azinphos-ethyl 8141
Azinphos-methyl 8141, 8270
Barban 8270, 8321
Baygon (Propoxur) 8318, 8321
Bendiocarb 8321
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TABLE 2-1. (Continued)
Analyte Applicable Method(s)
Benefin 8091
Benomyl 8321
Bentazon 8151
Benzal chloride 8121
Benzaldehyde 8315
Benz(a)anthracene 8100, 8270, 8275, 8310, 8410
Benzene 8021, 8260
Benzenethiol (Thiophenol) 8270
Benzidine 8270, 8325
Benzo(b)fluoranthene 8100, 8270, 8275, 8310
Benzo(j)fluoranthene 8100
Benzo(k)fluoranthene 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 i 8270
6-BHC (5-Hexachlorocyclohexane) 80811 81211 8270
Y-BHC (Lindane, Y-Hexachlorocyclohexane) 8081 i 8121 i 8270
Bis(2-chloroethoxy)methane 8111, B27Q, 8410
Bis(2-chloroethyl) ether 8111, 8270, 841o| 8430
Bis(2-chloroethyl)sulfide 8260
Bis(2-chloroisopropyl) ether 8021, 8111, 8270, 8410
Bis(2-n-butoxyethyl) phthalate 8061
Bis(2-ethoxyethyl) phthalate 8061
Bis(2-ethylhexyl) phthalate 8061, 8270, 8410
Bis(2-methoxyethyl) 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 i 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)
Butanal 8315
1-Butanol (n-Butyl alcohol) 8015
n-Butanol 8260
2-Butanone (Methyl ethyl ketone, MEK) 8015, 8260
Butralin 8091
n-Butyl alcohol (1-Butanol) 8015
t-Butyl alcohol 8015
n-Butylbenzene 8021, 8260
sec-Butylbenzene 8021, 8260
tert-Butylbenzene 8021, 8260
Butyl benzyl phthalate 8061, 8270, 8410
2-sec-Butyl-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-Chlorobutane 8260
Chlorodibromomethane (Dibromochloromethane) 8021, 8260
2-Chloro-4,6-dinitroaniline 8131
1-Chloro-2,4-dinitrobenzene 8091
1-Chloro-3,4-dinitrobenzene 8091
Chloroethane 8021, 8260
2-Chloroethanol 8021, 8260, 8430
2-(2-Chloroethoxy)ethanol 8430
2-Chloroethyl vinyl ether 8021, 8260
Chloroform 8021, 8260
1-Chlorohexane 8260
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TABLE 2-1. (Continued)
Analyte Applicable Method(s)
Chloromethane 8021 8260
5-Chloro-2-methylaniline .' 8270
Chloromethyl methyl ether 8021
2-Chloro-5-methylphenol 8041
4-Chloro-2-methylphenol ' ' 8041
4-Chloro-3-methylphenol 8041, 8270, 8410
3-(Chloromethyl)pyridine hydrochloride .' 8270
1-Chloronaphthalene 8270, 8275
2-Chloronaphthalene 8121, 827oi 8410
Chloroneb 8081
2-Chloro-4-nitroaniline 8131
4-Chloro-2-nitroaniline 8131
1-Chloro-2-nitrobenzene 8091
1-Chloro-4-nitrobenzene 8091
2-Chloro-6-nitrotoluene 8091
4-Chloro-2-nitrotoluene 8091
4-Chloro-3-nitrotoluene 8091
2-Chlorophenol 8041, 8270, 8410
3-Chlorophenol 8041
4-Chlorophenol ' " '' 8041 8410
4-Chloro-1,2-phenylenediamine ' 8270
4-Chloro-1,3-phenylenediamine 8270
4-Chlorophenyl 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-Chloropropionitrile ' 8260
Chloropropham 8321
Chloropropylate 8081
Chlorothalonil ' ' 8081
2-Chlorotoluene 8021 8260
4-Chlorotoluene 8021' 8260
Chloroxuron ' 3321
Chlorpyrifos 8141
Chlorpyrifos methyl 8141
Chrysene 8100, 8270, 8275, 8310, 8410
Coumaphos 8141, 8270
Coumarin Dyes ' 3321
p-Cresidine '_ 8270
o-Cresol (2-Methylphenol) 8041, 8270, 8410
m-Cresol (3-Methylphenol) 8041', 8270
p-Cresol (4-Methylphenol) 8041, 8270, 8275, 8410
Crotonaldehyde 8015, 8260 8315
Crotoxyphos 8141 8270
TWO-15 Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte
Applicable Method(s)
Cyclohexanone 8315
2-Cyclohexyl-4,6-dinitrophenol 8041, 8270
2,4-D 8151, 8321
Dalapon 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',3,314,4'5,51,6,6'-Decachlorobiphenyl 8275
Decanal 8315
Demeton-O, and Demeton-S 8141, 8270
2,4-D, ethylhexyl ester 8321
Diallate 8081, 8270
Diamyl phthalate 8061
2,4-Diaminotoluene 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)carbazole 8100
Dibenzofuran 8270, 8275, 8410
Dibenzo(a,e)pyrene .8100, 8270
Dibenzo(a,h)pyrene 8100
Dibenzo(a,i)pyrene 8100
Dibenzothiophene 8275
Dibromochloromethane (Chlorodibromomethane) 8021, 8260
1,2-Dibromo-3-chloropropane (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-Dichlorobenzene 8021, 8121, 8260, 8270, 8410
1,3-Dichlorobenzene 8021, 8121, 8260, 8270, 8410
1,4-Dichlorobenzene 8021, 8121, 8260, 8270, 8410
3,3'-Dichlorobenzidine 8270,8325
TWO-16
Revision 4
January 1998
-------
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
Dichloromethane (DCM, Methylene chloride) 8021, 8260
2,6-Dichloro-4-nitroaniline 8131
2,3-Dichloronitrobenzene 8091
2,4-Dichloronitrobenzene 8091
3,5-Dichloronitrobenzene 8091
3,4-Dichloronitrobenzene 8091
2,5-Dichloronitrobenzene 8091
2,3-Dichlorophenol 8041
2,4-Dichlorophenol 8041, 8270, 8410
2,5-Dichlorophenol 8041
2,6-Dichlorophenol 8041, 8270
3,4-Dichlorophenol 8041
3,5-Dichlorophenol 8041
2,4-Dichlorophenol 3-methyl-4-nitrophenyl ether 8111
2,6-Dichlorophenyl 4-nitrophenyl ether 8111
3,5-Dichlorophenyl 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-propanol 8021, 8260
1,1-Dichloropropene 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
TWO-17 Revision 4
January 1998
-------
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, 827o| 8410
Diethylstilbestrol 8270
Diethyl sulfate 8270
Dihexyl phthalate 8061
Diisobutyl 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-Dimethylphenol 8041
2,4-Dimethylphenol 8041, 8270
2,5-Dimethylphenol 8041
2,6-Dimethylphenol 8041
3,4-Dimethylphenol 8041
Dimethyl phthalate 8061, 8270, 8410
Dinitramine 8091
2,4-Dinitroaniline 8131
1,2-Dinitrobenzene 8091, 8270
1,3-Dinitrobenzene (1,3-DNB) 8091, 827o| 8330
1,4-Dinitrobenzene 8091, 8270
4,6-Dinitro-2-methylphenol 8270, 8410
2,4-Dinitrophenol 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-Diphenylhydantoin 8270
1,2-Diphenylhydrazine 8270
Disperse Blue 3 8321
Disperse Blue 14 8321
TWO-18 Revision 4
January 1998
-------
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
Disulfoton 8141, 8270, 8321
Diuron 8321, 8325
1,3-DNB (1,3-Dinitrobenzene) 8091, 8270, 8330
DNBP (2-sec-Butyl-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
ERN 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
Fluoranthene 8100, 8270, 8275, 8310, 8410
Fluorene 8100, 8270, 8275, 8310, 8410
Fluorescent Brightener 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
Heptachlor 8081, 8270
2,2',3,3',4,4',5-Heptachlorobiphenyl 8082, 8275
2,2',3,414I,515'-Heptachlorobiphenyl 8082, 8275
2,2',3,4,4',5',6-Heptachlorobiphenyl 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',3,3,4,4l-Hexachlorobiphenyl 8275
2,21,3,4,4',5'-Hexachlorobiphenyl 8082, 8275
2,2',3,4,5,51-Hexachlorobiphenyl 8082
2,2',3,5,5',6-Hexachlorobiphenyl 8082
2,2',4,4',5,5'-Hexachlorobiphenyl 8082
Hexachlorobutadiene 8021, 8121, 8260, 8270, 8410
ot-Hexachlorocyclohexane (a-BHC) 8081, 8121, 8270
P-Hexachlorocyclohexane (P-BHC) 8081, 8121, 8270
5-Hexachlorocyclohexane (5-BHC) 8081, 8121, 8270
Y-Hexachlorocyclohexane (v-BHC, Lindane) 8081, 8121, 8270
Hexachlorocyclopentadiene 8081, 8121, 8270, 8410
Hexachloroethane 8121, 8260, 8270, 8410
Hexachlorophene 8270
Hexachloropropene 8270
Hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) 8330
Hexamethylphosphoramide (HMPA) 8141, 8270
Hexanal 8315
2-Hexanone 8260
Hexyl 2-ethylhexyl phthalate 8061
HMPA (Hexamethyl phosphoramide) 8141, 8270
HMX (Octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine) 8330
1,2,3,4,6,7,8-HpCDD 8280, 8290
HpCDD, total 8280, 8290
1,2,3,4,6,7,8-HpCDF 8280, 8290
1,2,3,4,7,8,9-HpCDF 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-HxCDF 8280, 8290
2,3,4,6,7,8-HxCDF 8280, 8290
HxCDF 8280, 8290
Hydroquinone 8270
3-Hydroxycarbofuran 8318, 8321
5-Hydroxydicamba 8151
2-Hydroxypropionitrile 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
Isophorone 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-dinitrophenol 8041
4,4'-Methylenebis(2-chloroaniline) 8270
4,4'-Methylenebis(N,N-dimethylaniline) 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 methanesulfonate 8270
2-Methylnaphthalene 8270, 8410
Methyl parathion (Parathion, methyl) 8270, 8141, 8321
4-Methyl-2-pentanone (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-Methyl-2-pentanone) 8015, 8260
Mirex 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-Naphthylamine 8270
2-Naphthylamine 8270
Neburon 8321
Nicotine 8270
5-Nitroacenaphthene 8270
2-Nitroaniline 8131, 8270, 8410
3-Nitroaniline 8131, 8270, 8410
4-Nitroaniline 8131, 8270, 8410
5-Nitro-o-anisidine 8270
Nitrobenzene (NB) 8091, 8260, 8270, 8330, 8410
4-Nitrobiphenyl 8270
Nitrofen 8081, 8270
Nitroglycerin 8332
2-Nitrophenol 8041, 8270, 8410
3-Nitrophenol 8041
4-Nitrophenol 8041, 8151, 8270, 8410
4-Nitrophenyl phenyl ether 8111
TWO - 22 Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte Applicable Method(s)
2-Nitropropane 8260
Nitroquinoline-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-n-propylamine 8070, 8270, 8410
N-Nitrosomethylethylamine 8270
N-Nitrosomorpholine 8270
N-Nitrosopiperidine 8270
N-Nitrosopyrrolidine 8270
2-Nitrotoluene (o-Nitrotoluene, 2-NT) 8091, 8330
3-Nitrotoluene (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
2,2'3,3'4,415,5'6-Nonachlorobiphenyl 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'I4,4'5,51-Octachlorobiphenyl 8275
Octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (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 8280, 8290
2,3,4,7,8-PeCDF 8280, 8290
PeCDF, total 8280, 8290
Pendimethaline (Penoxalin) 8091
Penoxalin (Pendimethaline) 8091
Pentachlorobenzene 8121, 8270
2,2',3,4,5'-Pentachlorobiphenyl 8082
2,2',4,5,51-Pentachlorobiphenyl 8082, 8275
2,3,3',4',6-Pentachlorobiphenyl , 8082
2,3',4,4',5-Pentachlorobiphenyl 8275
Pentachloroethane 8260
Pentachloronitrobenzene 8091, 8270
Pentachlorophenol 8041, 8151, 8270, 8410
Pentafluorobenzene 8260
Pentanal (Valeraldehyde) 8315
2-Pentanone 8015, 8260
Permethrin (c/s and trans) 8081
Perthane 8081
Phenacetin 8270
Phenanthrene 8100, 8270, 8275, 8310, 8410
Phenobarbital 8270
Phenol 8041, 8270, 8410
1,4-Phenylenediamine 8270
Phorate 8141, 8270, 8321
Phosalone 8270
Phosmet 8141, 8270
Phosphamidon 8141, 8270
Phthalic anhydride 8270
Picloram 8151
2-Picoline (2-Methylpyridine) 8015, 8260, 8270
Piperonyl sulfoxide 8270
Profluralin 8091
Promecarb 8318
Pronamide 8270
Propachlor 8081, 8321
Propanal (Propionaldehyde) 8315, 8321
1-Propanol 8015, 8260
2-Propanol (Isopropyl alcohol) 8015, 8260
Propargyl alcohol 8260
Propenal (Acrolein) 8260, 8315
Propham 8321
B-Propiolactone 8260
Propionaldehyde (Propanal) 8315
Propionitrile (Ethyl cyanide) 8015, 8260
Propoxur (Baygon) 8318, 8321
n-Propylamine 8260
TWO - 24
Revision 4
January 1998
-------
TABLE 2-1. (Continued)
Analyte
Applicable Method(s)
n-Propylbenzene 8021, 8260
Propylthiouracil 8270
Prothiophos (Tokuthion) 8141
Pyrene 8100, 8270, 8275, 8310, 8410
Pyridine '.'.'. 8015, 8260
RDX (Hexahydro-1,3,5-trinitro-1,3,5-triazine) 8330
Resorcinol 8270
Ronnel 8141
Rotenone 8325
Safrole 8270
Sevin (Carbaryl) 8270, 8318, 8321, 8325
Siduron 8321, 8325
Simazine 8141
Silvex (2,4,5-TP) 8151, 8321
Solvent Red 3 8321
Solvent Red 23 8321
Stirophos (Tetrachlorvinphos) 8141, 8270
Strobane 8081
Strychnine 8270, 8321
Styrene 8021, 8260
Sulfallate 8270
Sulfotepp 8141
Sulprofos (Bolstar) 8141
2,4,5-T 8151, 8321
2,4,5-T, butoxyethanol ester 8321
2,4,5-T, butyl ester 8321
2,3,7,8-TCDD 8280, 8290
TCDD, total „ 8280, 8290
2,3,7,8-TCDF 8280, 8290
TCDF, total 8280, 8290
Tebuthiuron 8321
Temik (Aldicarb) 8318, 8321
Terbufos 8141, 8270
1,2,3,4-Tetrachlorobenzene 8121
1,2,3,5-Tetrachlorobenzene 8121
1,2,4,5-Tetrachlorobenzene 8121, 8270
2,2',3,5'-Tetrachlorobiphenyl 8082, 8275
2,2',4,5'-Tetrachlorobiphenyl 8275
2,2',5,5'-Tetrachlorobiphenyl 8082, 8275
2,3',4,4'-Tetrachlorobiphenyl 8082, 8275
1,1,1,2-Tetrachloroethane 8021, 8260
1,1,2,2-Tetrachloroethane 8021, 8260
Tetrachloroethene 8021, 8260
2,3,4,5-Tetrachlorophenol 8041
2,3,4,6-Tetrachlorophenol 8041, 8270
2,3,5,6-Tetrachlorophenol 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
Tetraethyl dithiopyrophosphate 8270
Tetraethyl pyrophosphate (TEPP) 8141, 8270
Tetrazene 8331
Tetryl (Methyl-2,4,6-trinitrophenylnitramine) 8330
Thiofanox 8321
Thionazin (Zinophos) 8141, 8270
Thiophenol (Benzenethiol) 8270
1,3,5-TNB (1,3,5-Trinitrobenzene) 8270, 8330
2,4,6-TNT (2,4,6-Trinitrobenzene) 8330
TOCP (Tri-o-cresylphosphate) 8141
Tokuthion (Prothiophos) 8141
m-Tolualdehyde 8315
o-Tolualdehyde 8315
p-Tolualdehyde 8315
Toluene 8021, 8260
Toluene diisocyanate 8270
o-Toluidine 8015, 8260, 8270
Toxaphene 8081, 8270
2,4,5-TP (Silvex) 8151, 8321
2,4,6-Trichloroaniline 8131
2,4,5-Trichloroaniline 8131
1,2,3-Trichlorobenzene 8021, 8121, 8260
1,2,4-Trichlorobenzene 8021, 8121, 8260, 8270, 8275, 8410
2,2',5-Trichlorobiphenyl 8082, 8275
2,3',5-Trichlorobiphenyl 8275
2,4',5-Trichlorobiphenyl 8082, 8275
1,3,5-Trichlorobenzene 8121
1,1,1-Trichloroethane 8021, 8260
1,1,2-Trichloroethane 8021, 8260
Trichloroethene 8021, 8260
Trichlorofluoromethane 8021, 8260
Trichlorfon 8141, 8321
Trichloronate 8141
1,2,3-Trichloro-4-nitrobenzene 8091
1,2,4-Trichloro-5-nitrobenzene 8091
2,4,6-Trichloronitrobenzene 8091
2,3,4-Trichlorophenol 8041
2,3,5-Trichlorophenol 8041
2,3,6-Trichlorophenol 8041
2,4,5-Trichlorophenol 8041, 8270, 8410
2,4,6-Trichlorophenol 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
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TABLE 2-1. (Continued)
Analyte Applicable Method(s)
2,3,5-Trichlorophenyl 4-nitrophenyl ether 8111
2,4,5-Trichlorophenyl 4-nitrophenyl 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-Trimethylaniline 8270
1,2,4-Trimethylbenzene 8021, 8260
1,3,5-Trimethylbenzene 8021, 8260
Trimethyl phosphate 8270
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-tolyl 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
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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
Allyl 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
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TABLE 2-4
METHOD 8021 (GC, PHOTOIONIZATION AND ELECTROLYTIC
CONDUCTIVITY DETECTORS) - AROMATIC AND HALOGENATED VOLATILES
Allyl chloride
Benzene
Benzyl chloride
Bis(2-chloroisopropyl)
ether
Bromoacetone
Bromobenzene
Bromochloromethane
Bromodichloromethane
Bromoform
Bromomethane
n-Butylbenzene
sec-Butylbenzene
tert-Butylbenzene
Carbon tetrachloride
Chlorobenzene
Chlorodibromomethane
Chloroethane
2-Chloroethanol
2-Chloroethyl vinyl ether
Chloroform
Chloromethyl methyl ether
Chloroprene
Chloromethane
2-Chlorotoluene
4-Chlorotoluene
1,2-Dibromo-3-chloropropane
1,2-Dibromoethane
Dibromomethane
1,2-Dichlorobenzene
1,3-Dichlorobenzene
1,4-Dichlorobenzene
Dichlorodifluoromethane
1,1-Dichloroethane
1,2-Dichloroethane
1,1 -Dichloroethene
cis-1,2-Dichloroethene
trans-1,2-Dichloroethene
1,2-Dichloropropane
1,3-Dichloropropane
2,2-Dichloropropane
1,3-Dichloro-2-propanol
1,1-Dichloropropene
cis-1,3-Dichloropropene
trans-1,3-Dichloropropene
Epichlorhydrin
Ethylbenzene
Hexachlorobutadiene
Isopropylbenzene
p-lsopropyltoluene
Methylene chloride
Naphthalene
n-Propylbenzene
Styrene
1,1,1,2-Tetrachloroethane
1,1,2,2-Tetrachloroethane
Tetrachloroethene
Toluene
1,2,3-Trichlorobenzene
1,2,4-Trichlorobenzene
1,1,1 -Trichloroethane
1,1,2-Trichloroethane
Trichloroethene
Trichlorofluoromethane
1,2,3-Trichloropropane
1,2,4-Trimethylbenzene
1,3,5-Trimethylbenzene
Vinyl chloride
o-Xylene
m-Xylene
p-Xylene
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
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TABLE 2-6
METHOD 8041 (GC) - PHENOLS
2-Chloro-5-methylphenol 2,4-Dinitrophenol
4-Chloro-2-methylphenol 2,5-Dinitrophenol
4-Chloro-3-methylphenol Dinoseb
2-Chlorophenol 2-Methyl-4,6-dinitrophenol
3-Chlorophenol 2-Methylphenol (o-Cresol)
4-Chlorophenol 3-Methylphenol (m-Cresol)
2-Cyclohexyl-4,6-dinitro- 4-Methylphenol (p-Cresol)
phenol 2-Nitrophenol
2,3-Dichlorophenol 3-Nitrophenol
2,4-Dichlorophenol 4-Nitrophenol
2,5-Dichlorophenol Pentachlorophenol
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-Dimethylphenol 2,3,4-Trichlorophenol
2,5-Dimethylphenol 2,3,5-Trichlorophenol
2,6-Dimethylphenol 2,3,6-Trichlorophenol
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 Diisobutyl 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
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TABLE 2-9
METHOD 8081 (GC) - ORGANOCHLORINE PESTICIDES AND PCBs
Alachlor
Aldrin
a-BHC
P-BHC
6-BHC
Y-BHC (Lindane)
Captafol
Chlorobenzilate
a-Chlordane
y-Chlordane
Chlordane (NOS)
Chloroneb
Chloropropylate
Chlorothalonil
DBCP
DCPA
4,4'-DDD
4,4'-DDE
4,4'-DDT
Diallate
Dichlone
Dicofol
Dieldrin
Endosulfan I
Endosulfan II
Endosulfan sulfate
Endrin
Endrin aldehyde
Endrin ketone
Etridiazole
Halowax-1000
Halowax-1001
Halowax-1013
Halowax-1014
Halowax-1051
Halowax-1099
Heptachlor
Heptachlor
epoxide
Hexachlorobenzene
Hexachlorocyclo-
pentadiene
Isodrin
Methoxychlor
Mirex
Nitrofen
frans-Nonachlor
PCNB
Permethrin (c/s and
trans)
Perthane
Propachlor
Strobane
Toxaphene
Trifluralin
TWO - 31
Revision 4
January 1998
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TABLE 2-10
METHOD 8082 (GC) - POLYCHLORINATED BIPHENYLS
Aroclor 1016
Aroclor 1221
Aroclor 1232
Aroclor 1242
Aroclor 1248
Aroclor 1254
Aroclor 1260
2-Chlorobiphenyl
2,3-Dichlorobiphenyl
2,2',5-Trichlorobiphenyl
2,4', 5-Trichlorobiphenyl
2,2',3,51-Tetrachlorobiphenyl
2,2',5,5'-Tetrachlorobiphenyl
2,3' ,4,4'-Tetrachlorobiphenyl
2,2',3,4,5l-Pentachlorobiphenyl
2,2',4,5,5l-Pentachlorobiphenyl
2,3,3',4',6-Pentachlorobiphenyl
2,2',3,4,41,51-Hexachlorobiphenyl
2,2l,3,4,5,5'-Hexachlorobiphenyl
2,2',3,5,5',6-Hexachlorobiphenyl
212I1414'15,5'-Hexachlorobiphenyl
2,2',3,3',4,4',5-Heptachlorobiphenyl
2,2',3,4,41I5,5I-Heptachlorobiphenyl
2,2',3,4,41,5I,6-Heptachloro-
biphenyl
2,2',3,4',5,5',6-Heptachlorobiphenyl
2,21,3,31,4,41,5,5'16-Nonachloro-
biphenyl
TABLE 2-11
METHOD 8091 (GC) - NITROAROMATICS AND CYCLIC KETONES
Benefin
Butralin
1-Chloro-2,4-dinitrobenzene
1-Chloro-3,4-dinitrobenzene
1-Chloro-2-nitrobenzene
1-Chloro-4-nitrobenzene
2-Chloro-6-nitrotoluene
4-Chloro-2-nitrotoluene
4-Chloro-3-nitrotoluene
2,3-Dichloronitrobenzene
2,4-Dichloronitrobenzene
3,5-Dichloronitrobenzene
3,4-Dichloronitrobenzene
2,5-Dichloronitrobenzene
Dinitramine
1,2-Dinitrobenzene
1,3-Dinitrobenzene
1,4-Dinitrobenzene
2,4-Dinitrotoluene
2,6-Dinitrotoluene
Isopropalin
1,2-Naphthoquinone
1,4-Naphthoquinone
Nitrobenzene
2-Nitrotoluene
3-Nitrotoluene
4-Nitrotoluene
Penoxalin [Pendimethalin]
Pentachloronitrobenzene
Profluralin
2,3,4,5-Tetrachloronitrobenzene
2,3,5,6-Tetrachloronitrobenzene
1,2,3-Trichloro-4-nitrobenzene
1,2,4-Trichloro-5-nitrobenzene
2,4,6-Trichloronitrobenzene
Trifluralin
TWO-32
Revision 4
January 1998
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TABLE 2-12
METHOD 8100 - POLYNUCLEAR AROMATIC HYDROCARBONS
Acenaphthene
Acenaphthylene
Anthracene
Benz(a)anthracene
Benzo(b)fluoranthene
Benzo(j)fluoranthene
Benzo(k)fluoranthene
Benzo(g,h,i)perylene
Benzo(a)pyrene
Chrysene
Dibenz(a,h)acridine
Dibenz(a,j)acridine
Dibenz(a,h)anthracene
7H-Dibenzo(c,g)carbazole
Dibenzo(a,e)pyrene
Dibenzo(a,h)pyrene
Dibenzo(a,i)pyrene
Fluoranthene
Fluorene
lndeno(1,2,3-cd)pyrene
3-Methylcholanthrene
Naphthalene
Phenanthrene
Pyrene
TABLE 2-13
METHOD 8111 (GC) - HALOETHERS
Bis(2-chloroethoxy)methane
Bis(2-chloroethyl) ether
Bis(2-chloroisopropyl) ether
4-Bromophenyl phenyl ether
4-Chlorophenyl phenyl ether
2-Chlorophenyl 4-nitrophenyl ether
3-Chlorophenyl 4-nitrophenyl ether
4-Chlorophenyl 4-nitrophenyl ether
2,4-Dibromophenyl 4-nitrophenyl
ether
2,4-Dichlorophenyl 3-methyl-4-
nitrophenyl ether
2,6-Dichlorophenyl 4-nitrophenyl
ether
3,5-Dichlorophenyl 4-nitrophenyl
ether
2,5-Dichlorophenyl 4-nitrophenyl
ether
2,4-Dichlorophenyl 4-nitrophenyl
ether
2,3-Dichlorophenyl 4-nitrophenyl
ether
3,4-Dichlorophenyl 4-nitrophenyl
ether
4-Nitrophenyl phenyl ether
2,4,6-Trichlorophenyl 4-nitrophenyl
ether
2,3,6-Trichlorophenyl 4-nitrophenyl
ether
2,3,5-Trichlorophenyl 4-nitrophenyl
ether
2,4,5-Trichlorophenyl 4-nitrophenyl
ether
3,4,5-Trichlorophenyl 4-nitrophenyl
ether
2,3,4-Trichlorophenyl 4-nitrophenyl ether
TWO-33
Revision 4
January 1998
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TABLE 2-14
METHOD 8121 (GC) - CHLORINATED HYDROCARBONS
Benzal chloride 5-Hexachlorocyclohexane
Benzotrichloride [5-BHC]
Benzyl chloride Y-Hexachlorocyclohexane [v-BHC]
2-Chloronaphthalene Hexachlorocyclopentadiene
1,2-Dichlorobenzene Hexachloroethane
1,3-Dichlorobenzene Pentachlorobenzene
1,4-Dichlorobenzene 1,2,3,4-Tetrachlorobenzene
Hexachlorobenzene 1,2,3,5-Tetrachlorobenzene
Hexachlorobutadiene 1,2,4,5-Tetrachlorobenzene
a-Hexachlorocyclohexane 1,2,3-Trichlorobenzene
[a-BHC] 1,2,4-Trichlorobenzene
3-Hexachlorocyclohexane 1,3,5-Trichlorobenzene
[p-BHC]
TABLE 2-15
METHOD 8131 (GC) - ANILINE AND SELECTED DERIVATIVES
Aniline 2,6-Dibromo-4-nitroaniline
4-Bromoaniline 3,4-Dichloroaniline
2-Bromo-6-chloro-4-nitroanilne 2,6-Dichloro-4-nitroaniline
2-Bromo-4,6-dintroaniline 2,4-Dinitroaniline
2-Chloroaniline 2-Nitroaniline
3-Chloroaniline 3-Nitroaniline
4-Chloroaniline 4-Nitroaniline
2-Chloro-4,6-dinitroaniline 2,4,6-Trichloroaniline
2-Chloro-4-nitroaniline 2,4,5-Trichloroaniline
4-Chloro-2-nitroaniline
TWO - 34 Revision 4
January 1998
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TABLE 2-16
METHOD 8141 (GC) - ORGANOPHOSPHORUS COMPOUNDS
Aspon Fenthion
Atrazine Fonophos
Azinphos-ethyl Hexamethyl phosphoramide (HMPA)
Azinphos-methyl 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
Chloramben acid Pentachiorophenol
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
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TABLE 2-18
METHOD 8260 (GC/MS)- VOLATILE ORGANIC COMPOUNDS
Acetone
Acetonitrile
Acrolein (Propenal)
Acrylonitrile
Ally! alcohol
Allyl chloride
Benzene
Benzyl chloride
Bis(2-chloroethyl)-
sulfide
Bromoacetone
Bromobenzene
Bromochloromethane
Bromodichloromethane
Bromoform
Bromomethane
n-Butanol
2-Butanone (MEK)
t-Butyl alcohol
n-Butylbenzene
sec-Butylbenzene
tert-Butylbenzene
Carbon disulfide
Carbon tetrachloride
Chloral hydrate
Chloroacetonitrile
Chlorobenzene
1-Chlorobutane
Chlorodibromomethane
Chloroethane
2-Chloroethanol
2-Chloroethyl vinyl
ether
Chloroform
1-Chlorohexane
Chloromethane
Chloroprene
3-Chloropropionitrile
2-Chlorotoluene
4-Chlorotoluene
Crotonaldehyde
1,2-Dibromo-3-
chloropropane
1,2-Dibromoethane
Dibromofluoromethane
Dibromomethane
1,2-Dichlorobenzene
1,3-Dichlorobenzene
1,4-Dichlorobenzene
cis-1,4-Dichloro-
2-butene
trans-1,4-Dichloro-2-
butene
Dichlorodifluoromethane
1,1-Dichloroethane
1,2-Dichloroethane
1,1-Dichloroethene
cis-1,2-Dichloroethene
trans-1,2-Dichloro-
ethene
1,2-Dichloropropane
1,3-Dichloropropane
2,2-Dichloropropane
1,3-Dichloro-2-propanol
1,1-Dichloropropene
cis-1,3-Dichloropropene
trans-1,3-Dichloro-
propene
1,2,3,4-Diepoxybutane
Diethyl ether
1,4-Dioxane
Epichlorohydrin
Ethanol
Ethyl acetate
Ethylbenzene
Ethylene oxide
Ethyl methacrylate
Hexachlorobutadiene
Hexachloroethane
2-Hexanone
2-Hydroxypropionitrile
lodomethane
Isobutyl alcohol
Isopropylbenzene
p-lsopropyltoluene
Malononitrile
Methacrylonitrile
Methanol
Methyl-t-butyl ether
Methylene chloride
Methyl acrylate
Methyl methacrylate
4-Methyl-2-pentanone
(MIBK)
Naphthalene
Nitrobenzene
2-Nitropropane
N-Nitroso-di-n-
butylamine
Paraldehyde
Pentachloroethane
Pentafluorobenzene
2-Pentanone
2-Picoline
1-Propanol
2-Propanol
Propargyl alcohol
(i-Propiolactone
Propionitrile (Ethyl
cyanide)
n-Propylamine
n-Propylbenzene
Pyridine
Styrene
1,1,1,2-Tetrachloro-
ethane
1,1,2,2-Tetrachloro-
ethane
Tetrachloroethene
Toluene
o-Toluidine
1,2,3-Trichlorobenzene
1,2,4-Trichlorobenzene
1,1,1 -Trichloroethane
1,1,2-Trichloroethane
Trichloroethene
Trichlorofluoromethane
1,2,3-Trichloropropane
1,2,4-Trimethylbenzene
1,3,5-Trimethylbenzene
Vinyl acetate
Vinyl chloride
o-Xylene
m-Xylene
p-Xylene
TWO - 36
Revision 4
January 1998
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TABLE 2-19
METHOD 8270 (GC/MS) - SEMIVOLATILE ORGANIC COMPOUNDS
Acenaphthene
Acenaphthylene
Acetophenone
2-Acetylaminofluorene
1-Acetyl-2-thiourea
Aldrin
2-Aminoanthraquinone
Aminoazobenzene
4-Aminobiphenyl
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
P-BHC
6-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
Chlordane (NOS)
Chlorfenvinphos
4-Chloroaniline
Chlorobenzilate
5-Chloro-2-methyl-
aniline
4-Chloro-3-methylphenol
3-(Chloromethyl)-
pyridine hydro-
chloride
1 -Chloronaphthalene
2-Chloronaphthalene
2-Chlorophenol
4-Chloro-1,2-phenylene-
diamine
4-Chloro-1,3-phenylene-
diamine
4-Chlorophenyl phenyl
ether
Chrysene
Coumaphos
p-Cresidine
Crotoxyphos
2-Cyclohexyl-4,6-
dinitrophenol
4,4'-DDD
4,4'-DDE
4,4'-DDT
Demeton-O
Demeton-S
Diallate (cis or trans)
2,4-Diaminotoluene
Dibenz(a,j)acridine
Dibenz(a,h)anthracene
Dibenzofuran
Dibenzo(a,e)pyrene
1,2-Dibromo-3-
chloropropane
Di-n-butyl phthalate
Dichlone
1,2-Dichlorobenzene
1,3-Dichlorobenzene
1,4-Dichlorobenzene
3,3'-Dichlorobenzidine
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-Dimethylphenethyl-
amine
2,4-Dimethylphenol
Dimethyl phthalate
1,2-Dinitrobenzene
1,3-Dinitrobenzene
1,4-Dinitrobenzene
4,6-Dinitro-2-methyl-
phenol
2,4-Dinitrophenol
2,4-Dinitrotoluene
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
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TABLE 2-19 (CONTINUED)
Fenthion
Fluchloralin
Fluoranthene
Fluorene
Heptachlor
Heptachlor epoxide
Hexachlorobenzene
Hexachlorobutadiene
Hexachlorocyclo-
pentadiene
Hexachloroethane
Hexachlorophene
Hexachloropropene
Hexamethylphosphoramide
Hydroquinone
lndeno(1,2,3-cd)pyrene
Isodrin
Isophorone
Isosafrole
Kepone
Leptophos
Malathion
Maleic anhydride
Mestranol
Methapyrilene
Methoxychlor
3-Methylcholanthrene
4,4'-Methylenebis-
(2-chloroaniline)
4,4'-Methylenebis-
(N,N-dimethylaniline)
Methyl methanesulfonate
2-Methylnaphthalene
Methyl parathion
2-Methylphenol
3-Methylphenol
4-Methylphenol
Mevinphos
Mexacarbate
Mi rex
Monocrotophos
Naled
Naphthalene
1,4-Naphthoquinone
1-Naphthylamine
2-Naphthylamine
Nicotine
5-Nitroacenaphthene
2-Nitroaniline
3-Nitroaniline
4-Nitroaniline
5-Nitro-o-anisidine
Nitrobenzene
4-Nitrobiphenyl
Nitrofen
2-Nitrophenol
4-Nitrophenol
Nitroquinoline-1-oxide
N-Nitrosodi-n-
butylamine
N-Nitrosodiethylamine
N-Nitrosodimethylamine
N-Nitrosodiphenylamine
N-Nitrosodi-n-propyl-
amine
N-Nitrosomethylethyl-
amine
N-Nitrosomorpholine
N-Nitrosopiperidine
N-Nitrosopyrrolidine
5-Nitro-o-toluidine
Octamethyl pyrophos-
phoramide
4,4'-Oxydianiline
Parathion
Pentachlorobenzene
Pentachloronitrobenzene
Pentachlorophenol
Phenacetin
Phenanthrene
Phenobarbital
Phenol
1,4-Phenylenediamine
Phorate
Phosalone
Phosmet
Phosphamidion
Phthalic anhydride
2-Picoline (2-Methylpyridine)
Piperonyl sulfoxide
Pronamide
Propylthiouracil
Pyrene
Resorcinol
Safrole
Strychnine
Sulfallate
Terbufos
1,2,4,5-Tetrachloro
benzene
2,3,4,6-Tetrachloro-
phenol
Tetrachlorvinphos
Tetraethyl dithio-
pyrophosphate
Tetraethyl
pyrophosphate
Thionazine
Thiophenol
(Benzenethiol)
Toluene diisocyanate
o-Toluidine
Toxaphene
1,2,4-Trichlorobenzene
2,4,5-Trichlorophenol
2,4,6-Trichlorophenol
O,O,O-Triethyl
phosphorothioate
Trifluralin
2,4,5-Trimethylaniline
Trimethyl phosphate
1,3,5-Trinitrobenzene
Tris(2,3-dibromopropyl)
phosphate
Tri-p-tolyl phosphate
TWO-38
Revision 4
January 1998
-------
TABLE 2-20
METHOD 8275 (TE/GC/MS) - SEMIVOLATILE ORGANIC COMPOUNDS
Acenaphthene
Acenaphthylene
Anthracene
Benz(a)anthracene
Benzo(a)pyrene
Benzo(b)fluoranthene
Benzo(g,h,i)perylene
Benzo(k)fluoranthene
4-Bromophenyl phenyl ether
1 -Chloronaphthalene
Chrysene
Dibenzofuran
Dibenz(a,h)anthracene
Dibenzotniophene
Fluoranthene
Fluorene
Hexachlorobenzene
lndeno(1,2,3-cd)pyrene
Naphthalene
Phenanthrene
Pyrene
1,2,4-Trichlorobenzene
2-Chlorobiphenyl
3,3'-Dichlorobiphenyl
2,2',5-Trichloro-
biphenyl
2,3',5-Trichloro-
biphenyl
2,4',5-Trichloro-
biphenyl
2,2',515l-Tetrachloro-
biphenyl
2,2'4,51-Tetrachloro-
biphenyl
2,2'3,5I-Tetrachloro-
biphenyl
2,3II4,4'-Tetrachloro-
biphenyl
2,2',415,5'-Penta-
chlorobiphenyl
2,3',4,4',5-Penta-
chlorobiphenyl
2,2',3,4141,5I-
Hexachlorobiphenyl
2,2',3,3',4,41-
Hexachlorobiphenyl
2,2',3,41,5,5'16-
Heptachlorobiphenyl
2,21,3,4,41,515I-
Heptachlorobiphenyl
2,2',3,3t,4I4',5-
Heptachlorobiphenyl
2,2',3,3f)4,4',5,5'-
Octachlorobiphenyl
2,2',3,314141,5,51,6-
Nonachlorobiphenyl
2,2',313'4,4',5,5',6,61-
Decachlorobiphenyl
TABLE 2-21
METHODS 8280 (HRGC/LRMS) AND 8290 (HRGC/HRMS) -
POLYCHLORINATED DIBENZO-p-DIOXINS (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
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
Acenaphthylene
Anthracene
Benzo(a)anthracene
Benzo(a)pyrene
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
Acetaldehyde
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
Acrylonitrile
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
-------
TABLE 2-26. METHOD 8321 (HPLC/TS/MS) - NONVOLATILE ORGANIC COMPOUNDS
Azo Dyes
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, ethylhexyl 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
Organophosphorus Compounds
Asulam
Fensulfothion
Dichlorvos
Dimethoate
Disulfoton
Parathion methyl
Merphos
Methomyl
Monocrotophos
Famphur
Naled
Phorate
Trichlorfon
Thiofanox
Tris(2,3-dibromopropyl) phosphate
(Tris-BP)
Anthraquinone Dyes
Disperse Blue 3
Disperse Blue 14
Disperse Red 60
Coumarin Dyes
Fluorescent Brighteners
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 3,3'-Dimethylbenzidine
Benzoylprop ethyl Diuron
Carbaryl Linuron (Lorox)
o-Chlorophenyl thiourea Monuron
3,3'-Dichlorobenzidine Rotenone
3,3'-Dimethoxybenzidine Siduron
TABLE 2-28
METHOD 8330 (HPLC) - NITROAROMATICS AND NITRAMINES
4-Amino-2,6-dinitrotoluene Nitrobenzene (NB)
(4-Am-DNT) 2-Nitrotoluene (2-NT)
2-Amino-4,6-dinitrotoluene 3-Nitrotoluene (3-NT)
(2-Am-DNT) 4-Nitrotoluene (4-NT)
1,3-Dinitrobenzene (1,3-DNB) Octahydro-1,3,5,7-tetranitro-
2,4-Dinitrotoluene (2,4-DNT) 1,3,5,7-tetrazocine (HMX)
2,6-Dinitrotoluene (2,6-DNT) 1,3,5-Trinitrobenzene (1,3,5-TNB)
Hexahydro-1,3,5-trinitro- 2,4,6-Trinitrotoluene (2,4,6-TNT)
1,3,5-triazine(RDX)
Methyl-2,4,6-trinitrophenyl-
nitramine (Tetryl)
TABLE 2-29
METHOD 8331 (REVERSE PHASE HPLC)
Tetrazene
TABLE 2-30
METHOD 8332 (HPLC)
Nitroglycerine
TWO - 43 Revision 4
January 1998
-------
TABLE 2-31
METHOD 8410 - SEMIVOLATILE ORGANIC COMPOUNDS
Acenaphthene
Acenaphthylene
Anthracene
Benzo(a)anthracene
Benzo(a)pyrene
Benzole acid
Bis(2-chloroethoxy)methane
Bis(2-chloroethyl) ether
Bis(2-chloroisopropyl) ether
Bis(2-ethylhexyl) phthalate
4-Bromophenyl phenyl ether
Butyl 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-Hexachlorobutadiene
Hexachlorocyclopentadiene
Hexachloroethane
Isophorone
2-Methylnaphthalene
2-Methylphenol
4-Methylphenol
Naphthalene
2-Nitroaniline
3-Nitroaniline
4-Nitroaniline
Nitrobenzene
2-Nitrophenol
4-Nitrophenol
N-Nitrosodimethylamine
N-Nitrosodiphenylamine
N-Nitroso-di-n-propylamine
Pentachlorophenol
Phenanthrene
Phenol
Pyrene
1,2,4-Trichlorobenzene
2,4,5-Trichlorophenol
2,4,6-Trichlorophenol
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)ethanol
Diethylene glycol
Ethylene glycol
TWO-44
Revision 4
January 1998
-------
TABLE 2-33. DETERMINATIVE METHODS FOR INORGANIC ANALYTES
Analyte 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
TWO - 45 Revision 4
January 1998
-------
TABLE 2-34
CONTAINERS, PRESERVATION TECHNIQUES, AND HOLDING TIMES
FOR AQUEOUS MATRICESA
Name
Container
Preservation
Maximum holding time
Inorganic Tests:
Chloride
Cyanide, total and
amenable
to chlorination
P,G
P,G
None required
Cool to 4°C; if oxidizing
agents present add 5 mL
0.1N NaAsO2 per L or
0.06 g of ascorbic acid
per L; adjust pH>12 with
50% NaOH.
See Method 9010 for
other interferences.
28 days
14 days
Hydrogen ion (pH)
Nitrate
Sulfate
Sulfide
Metals:
Chromium VI
Mercury
Metals, except chromium VI
and mercury
Organic Tests:
Acrolein and acrylonitrile
Benzidines
Chlorinated hydrocarbons
Dioxins and Furans
Haloethers
Nitroaromatics and
cyclic ketones
Nitrosamines
P, G
P,G
P, G
P,G
P,G
P,G
P, G
G, PTFE-lined
septum
G, PTFE-lined
cap
G, PTFE-lined
cap
G, PTFE-lined
cap
G, PTFE-lined
cap
G, PTFE-lined
cap
G, PTFE-lined
cap
None required
Cool to 4°C
Cool to 4°C
Cool to 4°C, add zinc
acetate
Cool to 4°C
HNO3 to pH<2
HNO3 to pH<2
Cool to 4°C,
0.008% Na2S2O33,
Adjust pH to 4-5
Cool to 4°C,
0.008% Na2S2O33
Cool to 4°C,
0.008% Na2S2O33
Cool to 4°C,
0.008% Na2S2O33
Cool to 4°C,
0.008% Na2S2O33
Cool to 4°C,
0.008% Na2S2O33,
store in dark
Cool to 4°C,
0.008% Na2S2O33,
store in dark
24 hours
48 hours
28 days
7 days
24 hours
28 days
6 months
14 days
7 days until extraction,
40 days after extraction
7 days until extraction,
40 days after extraction
30 days until extraction,
45 days after extraction
7 days until extraction,
40 days after extraction
7 days until extraction,
40 days after extraction
7 days until extraction,
40 days after extraction
(continued on next page)
TWO - 46
Revision 4
January 1998
-------
TABLE 2-34 (continued)
Name
Container1 Preservation
Maximum holding time
Oil and grease G
Organic carbon, total (TOC) P, G
Organochlorine pesticides
Organophosphorus
pesticides
PCBs
Phenols
Phthalate esters
Polynuclear aromatic
hydrocarbons
Purgeable aromatic
hydrocarbons
Cool to 4°C,
add 5 mL diluted HCI
Cool to 4°C,
store in dark2
G, PTFE-lined Cool to 4°C
cap
Cool to 4°C"
G, PTFE-lined
cap
G, PTFE-lined
cap
G, PTFE-lined
cap
G, PTFE-lined
cap
G, PTFE-lined
cap
G, PTFE-lined
septum
Purgeable Halocarbons G, PTFE-lined
septum
Total organic halides (TOX) G, PTFE-lined
cap
Radiological Tests:
Alpha, beta and radium P, G
Cool to 4°C
Cool to 4°C,
0.008% Na2S2O33
Cool to 4°C
Cool to 4°C,
0.008% Na2S2O33,
store in dark
Cool to 4°C,
0.008% Na2S2032'3
Cool to 4°C,
0.008% Na2S2O33
Cool to 4°C, Adjust to
pH<2 with H2SO4
HNO3 to pH<2
28 days
28 days
7 days until extraction,
40 days after extraction
7 days until extraction,
40 days after extraction
7 days until extraction,
40 days after extraction
7 days until extraction,
40 days after extraction
7 days until extraction,
40 days after extraction
7 days until extraction,
40 days after extraction
14 days
14 days
28 days
6 months
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 H2SO4, HCI or solid NaHSO4. Free chlorine must be removed prior to adjustment.
3 Free chlorine must be removed by the appropriate addition of Na2S2O3.
4 Adjust samples to pH 5-8 using NaOH or H2SO4
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.)
Analyte Type
Acid Extractable
Acrolein, Acrylonitrile, and
Acetonitrile
Acrylamide
Aniline and Selected Derivatives
Aromatic Volatiles
Base/Neutral Extractable
Carbamates
Chlorinated Herbicides
Chlorinated Hydrocarbons
Dyes
Explosives
Formaldehyde
Haloethers
Matrix
Aqueous1
3510
3520
(PH * 2)
5031
80324
3510
3520
(pH>11)
5031 11
5021
5030
5032
3510
3520
(pH>11)
831 85
8321
81516
(PH * 2)
8321
3510
3520
(pHas
received)
3510
3520
83307
8331 8
831 59
3510
3520
Solids
3540
3541
3545
3550
5031
3540
3541
3545
3550
5021
5032
5035
3540
3541
3545
3550
831 85
8321
81516
8321
3540
3541
3550
3540
3541
3545
3550
83307
83318
831 59
3540
3541
3545
3550
Sludges and
Emulsions12
3520
(pH < 2)
5031
3520
(pH>11)
5030
5032
3520
(pH>11)
831 85
81516
(pH < 2)
3520
(pHas
recieved)
Organic
Liquids,
Tars, Oils
3650
35803
3585
35803
3585
3650
35803
831 85
35803
35803
TWO - 48
Revision 4
January 1998
-------
TABLE 2-35
PREPARATION METHODS FOR ORGANIC ANALYTES
(continued)
Analyte Type
Halogenated Volatiles
Nitroaromatic and Cyclic
Ketones
Nitrosamines
Non-halogenated Volatiles
Organochlorine Pesticides
Organophosphorus Pesticides
Phenols
Phthalate Esters
Polychlorinated Biphenyls
PCDDs and PCDFs
Matrix
Aqueous1
5021
5030
5032
3510
3520
(pH 5-9)
3510
3520
5021
5031
5032
3510
3520
3535
(pH 5-9)
3510
3520
(pH 5-8)
3510
3520
(PH * 2)
3510
3520
3535
(PH 5-7)
3510
3520
3535
(pH 5-9)
828010
829010
Solids
5021
5032
5035
3540
3541
3545
3550
3540
3541
3545
3550
5021
5031
5032
3540
3541
3545
3550
3540
3541
3545
3540
3541
3545
3550
3562
3540
3541
3545
3550
3540
3541
3545
3562
828010
829010
Sludges and
Emulsions1'2
5030
3520
(pH 5-9)
5021
5031
5032
3520
(pH 5-9)
3520
(pH 5-8)
3520
(PH * 2)
3520
(PH 5- 7)
3520
(pH 5-9)
828010
829010
Organic
Liquids,
Tars, Oils
3585
35803
5032
3585
35803
35803
3650
35803
35803
35803
828010
829010
TWO-49
Revision 4
January 1998
-------
TABLE 2-35
PREPARATION METHODS FOR ORGANIC ANALYTES
(continued)
Analyte Type
Polynuclear Aromatic
Hydrocarbons
Volatile Organics
Matrix
Aqueous1
3510
3520
(pHas
received)
5021
5030
5031
5032
Solids
3540
3541
3545
3550
3561
5021
5031
5032
5035
Sludges and
Emulsions1'2
3520
(pHas
received)
5021
5030
5031
5032
Organic
Liquids,
Tars, Oils
35803
3585
Footnotes for Table 2-35
9
10
11
The pH at which extraction should be performed is shown in parentheses.
If attempts to break an emulsion are unsuccessful, these methods may be used.
Method 3580 is only appropriate if the sample is soluble in the specified solvent.
Method 8032 contains the extraction, cleanup, and determinative procedures for this analyte.
Method 8318 contains the extraction, cleanup, and determinative procedures for these analytes.
Method 8151 contains the extraction, cleanup, and determinative procedures for these analytes.
Method 8330 contains the extraction, cleanup, and determinative procedures for these analytes.
Method 8331 is for Tetrazene only, and contains the extraction, cleanup, and determinative procedures for
this analyte.
Method 8315 contains the extraction, cleanup, and determinative procedures for this analyte.
Methods 8280 and 8290 contain the extraction, cleanup, and determinative procedures for these analytes.
Method 5031 may be used when only aniline is to be determined.
TWO-50
Revision 4
January 1998
-------
TABLE 2-36. CLEANUP METHODS FOR ORGANIC ANALYTE EXTRACTS
Analyte Type
Acid Extractable
Base/Neutral Extractable
Carbamates
Chlorinated Herbicides
Chlorinated Hydrocarbons
Haloethers
Nitroaromatics & Cyclic Ketones
Nitrosamines
Organochlorine Pesticides
Organophosphorus Pesticides
Phenols
Phthalate Esters
Polychlorinated Biphenyls
Polychlorinated Dibenzo-p-Dioxins and
Polychlorinated Dibenzofurans
Polynuclear Aromatic Hydrocarbons
Method
3650, 3640
3650, 3640
831 81
81512
3620
3640
3620
3640
3620
3640
3610, 3620, 3640
3620
3630
3640
3660
3620
3630
3640
3650
80413
3610
3611
3620
3640
3620
3630
3640
3660
3665
82804
8290"
3610
3611
3630
3640
3650
Method 8318 contains the extraction, cleanup, and determinative procedures for these analytes.
Method 8151 contains the extraction, cleanup, and determinative procedures for these analytes.
Method 8041 includes a dervatization technique followed by GC/ECD analysis, if interferences
are encountered using GC/FID.
Methods 8280 and 8290 contain the extraction, cleanup, and determinative procedures for these
analytes.
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TABLE 2-37. DETERMINATIVE METHODS ORGANIC ANALYTES
Analyte Type
Acid Extractable
Acrolein, Acrylonitrile, Acetonitrile
Acrylamide
Aniline and Selected Derivatives
Aromatic Volatiles
Base/Neutral Extractable
Carbamates
Chlorinated Herbicides
Chlorinated Hydrocarbons
Dyes
Explosives
Formaldehyde
Haloethers
Halogenated Volatiles
Nitroaromatics and Cyclic Ketones
Nitrosoamines
Non-halogenated Volatiles
Organochlorine Pesticides
Organophosphorus Pesticides
Phenols
Petroleum Hydrocarbons
Phthalate Esters
Polychlorinated Biphenyls
PCDDs and PCDFs
Polynuclear Aromatic Hydrocarbons
Volatile Organics
GC/MS
Method
8270
8260
8260
8270
8260
8270
82703
8270
8270
8260
8270
8270
8260
82703
82703
8270
8270
82703
8280
8290
8270
8260
Specific GC
Method
8031
80331
8032
8131
8021
8151
8121
8111
8011,8021
8091
8070
8015
8081
8141
8041
8015
8061
8082
8100
8011,8015,
8021,8031,
8032, 8033
HPLC
Method
831 52
8316
8316
83254
8318, 8321
8321
8321
8330,
8331,8332
8315
83305
8315
8321
8310
8315
8316
Of these analytes, Method 8033 is for acetonitrile only.
Of these analytes, Method 8315 is for acrolein only.
This method is an alternative confirmation method, not the method of choice.
Benzidines and related compounds.
Nitroaromatics (see "Explosives").
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TABLE 2-38
PREPARATION METHODS FOR INORGANIC ANALYSES1
MATRIX
Surface Water
Ground Water
Extracts
Aqueous samples containing suspended
solids
Oils
Oil Sludges
Tars
Waxes
Paints
Paint Sludges
Petroleum Products
Sediments
Sludges
Soil Samples
Ashes
Biological Tissues
METHOD
3005, 3010, 3015,
3005, 3010, 3015,
3020
3020
3010, 3015, 3020
3010, 3015, 3020
3031,3040,3051,
30522
3031, 30522
3031, 30522
3031, 3040, 30522
3031 , 30522
3031, 30522
3031,3040, 30522
3050,3051, 30522
3050, 3051, 30522
3050, 3051, 30522
, 30603
, 30603
, 30603
30522
30522
1lt 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.
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TABLE 2-39
USE OF LEACHING, EXTRACTION AND DIGESTION METHODS
FOR INORGANIC ANALYSIS
(Generally ordered by increasing strength)
METHOD
1310
1311
1312
1320
3040
3005
3020
3010
3060A
3015
3050
3051
3031
3052
REAGENTS & CONDITIONS
dilute acetic acid (synthetic municipal
solid waste leachate)
dilute acetic acid (synthetic municipal
solid waste leachate)
dilute H2SO4 and HNO3 (synthetic acid
rain)
dilute H2SO4 and HNO3 (synthetic acid
rain)
solvent
HNO3, heat
HN03, heat
HNO3, HCI, heat
Na2CO3/NaOH, heat
HNO3, HCI (optional), pressure, heat
HNO3, H2O2, HCI (optional), heat
HNO3, HCI (optional), pressure, heat
Potassium permanganate, H2SO4, HNO3,
HCI, heat
HN03, HF, HCI (optional) H2O2 (optional),
heat, pressure
USE
Simulate leaching of a waste in a
municipal solid waste landfill
Simulate leaching of a waste in a
municipal solid waste landfill
Simulate acid rain leaching of a
waste
Simulate long-term acid rain
leaching of a waste
Dissolution of oils, oily wastes,
greasses and waxes
Surface and ground waters
Aqueous samples and extracts for
GFAA work only
Aqueous samples and extracts
Soils, sludges, sediments and some
industrial wastes for the analysis of
hexavalent chromium only.
Aqueous samples and extracts
Sediments, soils, and sludges
Sludges, sediments, soils and oils
Oils, oily sludges, tars, waxes, paint,
paint sludge
Siliceous, organic and other
complex matrices for total sample
decomposition
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FIGURE 2-1
ORGANIC ANALYSIS OPTIONS FOR SOILD AND LIQUID MATRICES
to be analyzed
for extractabl
Organic or
Liquid Oil
GC Analysis Procedures:
EDB and DBCP:
Nonhalogenated Volatile Organics:
Halogenated Volatile Compounds:
Acrylamide:
Acetonitrile:
8011
8015
8021
8032
8033
HPLC Analysis Procedures:
Acrolein, Acrylonitrile, Acrylamide
Carbaryl Compounds:
Extractable
Cleanup Procedure:
Alumina Column: 3610
Alumina Column for Petroleum Wastes: 3611
Florisil Column: 3*20
Silica Gel Column: 3630
Gel Permeation: 3640
Acid Base Partioning: 3650
Sulfur: 3660
Sulfuric Acid Cleanup: 3665
HPLC Analysis Procedures:
8310, 8318, 8321, 8325, 8330,
8331, 8332.
GC Analysis Procedures:
Phenols: 8041
Phthalate Esters: 8061
Nitrosamines: 8070
Organochlorine Pesticides: 8081
PCBs: 8082
Nitroaromatics and Cyclic Ketones: 8091
Polynuclear Aromatic Hydrocarbons: 8100
Haloethers: 8111
Chlorinated Hydrocarbons: 8121
Organophosphorus Pesticides: 8141
Chlorinated Herbicides: 8151
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FIGURE 2-2
SCHEMATIC OF SEQUENCE TO DETERMINE
IF A WASTE IS HAZARDOUS BY CHARACTERISTIC
~-\
Nonhazardous by >.
reason of )
ignitability J
characteristic ./
^-^^
DOT (49 CFR 173.300)
Is waste
ignitable?
Is
waste
reactive to
air and/or
water'
Is waste
explosive?
Generator Knowledge
DOT (49 CFR 173.151)
What is
physical state
of waste?
Is waste
ignitable?
Perform Paint
Filter Test
(Method 9095)
Methods 1110 and 9040
Yes
Is waste
corrosive?
f Nonhazardous "\
( for corrosivity )
X^characteristjc,/
Methods 1010 or 1020
Yes
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FIGURE 2-2
(Continued)
Q Hazardous ")
Nonhazardous
for ignitability
characteristic
Reactive CN
and Sulfide Tests
Does waste
generate toxic
gas?
Nonhazardous
for toxic gas generation
(reactivity) characteristic
Is total
concen. of TC
constituents •+• 20 <
TC regulatory
limit?
Nonhazardous
for toxicity
characteristic
Is waste
leachable and
toxic?
(Method 1311)
Nonhazardous
for toxicity
characteristic
(~~ Hazardous J)
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FIGURE 2-3A
RECOMMENDED SW-846 METHODS FOR ANALYSIS OF EP LEACHATES
Sample
Leach Method
1310
Prep. Methods
3010, 3015
(7760 Ag)
Determinative
Methods
6010
6020
7000
7010
3510/3520/3535
Neutral
7470
Hg
8081
Pesticides
8151/8321
Herbicides
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FIGURE 2-3B
RECOMMENDED SW-846 METHODS FOR ANALYSIS OF TCLP LEACHATES
Sample
Leach
Method
Prep.
Methods
TCLP
Determinative
Methods
3010
3015
6010
6020
7000
7010
7470
Hg
3510/35207
3535
Neutral
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
VOA
Semivolatiles
8260
3510 or
3520
8270
Organic
Sample
Pesticides
3510, 3520,
or 3535
Neutral
3620, 3640,
and/or 3660
8081
Herbicides
Dioxins
8151 or
8321
8280 or
8290
1 - Optional: Cleanup required only if interferences prevent analysis.
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FIGURE 2-4B.
GROUND WATER ANALYSIS: INDICATOR ANALYTES
1 - Barcelona, 1984, (See Reference 1)
2 - Riggin, 1984, (See Reference 2)
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FIGURE 2-4C.
GROUND WATER ANALYSIS: INORGANIC ANALYTES
GROUND WATER1
SAMPLE
SAMPLE PREPARATION
3005 OR 3015
SAMPLE PREPARATION
3015 OR 3020
Ag, Al, As, Ba,
Be, Cd, Co, Cr,
Cu, Fe, Mg, Mn,
Mo, Ni, Pb, Sb,
Se, Tl, V, Zn
Ag, Al, As, Ba,
Be, Cd, Co, Cr,
Cu, Mn, Ni, Pb,
Sb, Tl, 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
Ag, As, Ba, Be,
Cd, Co, Cr, Cu,
Fe, Pb, Mn, Mo,
Ni, Sb, Se, Tl,
V, Zn
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 Introduction
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-um filter (Method 3005).
Suspended metals: The concentration of metals determined in the portion of a sample that is
retained by a 0.45-um 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 (IDL): 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): 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 (\C\Ti standard: A certified or independently prepared solution used
to verify the accuracy of the initial calibration. For ICP analysis, it must be run at each wavelength
used in the analysis.
Continuing calibration verification (CCW 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 (MSA): 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 all 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 borosilicate 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 acid if adequate cleaning is documented by an analytical quality
control program. (Chromic acid should also not be used with plastic bottles.)
3.1.4 Safety
The toxicity or carcinogenicity 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, 1986.
4. "Safety in Academic Chemistry Laboratories," American Chemical Society Publication,
Committee on Chemical Safety.
3.1.5 Sample Preparation
For all non-speciated 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, reducing its particle 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 oven 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 instruments 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 instrument 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 analytical
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(±sa) - b(±sb)
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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).
Sy = /S2 + Sb2 + Sc2
In this case, "a" represents the standard deviation of the measurement, "b" represents the
standard deviation of the blank, that must be subtracted, and "c" represents the uncertainty
associated with the sampling error. This example will only consider the uncertainty of the
measurement "a" and the blank "b", as the sampling uncertainty "c" is beyond the scope of this
discussion (presented previously and may be found in other literature, References 7, 8).
The following example illustrates a common relationship that causes the imprecision of the
blank to be the limiting factor in the overall uncertainty of the analysis. Analysis of a set of samples
determines the mean value to be 55.5 ± 0.3, with an analytical blank of 11 ± 5 (which is too large to
be ignored). The uncertainties of these results are the standard deviations of the replicate
measurements. The analytical blank becomes the dominant uncertainty in calculating the uncertainty
of the final result. Here the blank subtracted mean y = 55.5 -11= 44.5, and the standard deviation
is 5. The net result of the analysis is 44.5 ± 5. Essentially, the entire uncertainty is due to the
uncertainty of the analytical blank.
It has been suggested by some environmental laboratories that having a blank concentration
below the instrumental detection limit while the measurement is detectable provides a more
convenient measurement. This is, however, not an appropriate approach for minimizing contribution
of the blank. A blank value below the limit of detection does not remove its influence. Just because
the blank is not detectable does not mean it is not influencing the measurement. An accurate
measurement of the blank value with high precision provides the most accurate overall analytical
estimate of the concentration.
The four primary areas that effect the analytical blank can be demonstrated using standard
reference materials in analysis. Table 3-2 illustrates and isolates the main blank influencing
parameters: environment, reagents, materials, and analyst skills. The skill of the analyst was kept
constant as the same analyst changed the environment, reagents, and combinations of these
parameters in the analysis (3). Trace elements in glass (TEG) National Institute of Standards and
Technology (MIST) standard reference material was used to keep sample homogeneity constant and
to permit removal of the sampling error by using sample sizes where appropriate homogeneity had
previously been demonstrated.
It is important to note that the relationship of the precision and measurement remained relatively
constant. This relationship yields no information about the accuracy of the data. The significance
of the first two major sources of contamination, environment and reagents, can be evaluated. In the
example above, the contamination in the laboratory air and in the acid used for the reagent blank
altered the accuracy of the example above by over two orders of magnitude for both lead and silver.
The larger influence of the two sources in this example is the laboratory environment in which the
samples were prepared.
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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 clean
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 Mm, for the high efficiency p.articulate 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 acid 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 "Brownian movement" 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 all 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 acids 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 the specific types of materials of non-contaminating nature and chemical inertness to most
acid 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 closed vessel liners that come in
contact with the sample. These materials are the most stable to acid reactions (with the exception
of quartz and glass if hydrofluoric acid is used). Fluoropolymers are the most common and were
adapted from other chemical uses for application in pressure systems. The fluoropolymers, 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 acid has a boiling point of approximately 330 °C and can damage all fluoropolymers
by melting them. Quartz and glass can safely contain sulfuric acid 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
THREE - 7 Revision 4
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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 Trace Elements; Alfaassi, 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) Rabinowitz, M. B.; Wetherill, G. W. "Identifying Sources of Lead Contamination by Stable
Isotope Techniques", Environ. Sci. Technol. 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 Analysis; 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. R. Soc. London 1982, 305, 669-680.
THREE - 9 Revision 4
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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) Skelly, E. M.; DiStefano, F. T. "Clean Room and Microwave Digestion Techniques: Improvement
in Detection Limits for Aluminum Determination by GF-AAS", Appl. Spectrosc. 1988, 42,1302-1306.
(19) Prevatt, F. J. "Clean Chemistry for Trace Analysis", Environmental Testing and Analysis 1995,
4, 24-27.
THREE-10 Revision 4
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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
Treatment/
Preservative
Holding Time"
Inorganic Analvtes (except hexavalent chromium and mercury):
Aqueous
Total 100 600
Solid
Dissolved
Suspended
Total
Hexavalent Chromium:
Aqueous
Solid
100
100
2g
100
2.5 g
600
600
200 g
400
100 g
HNO3 to pH <2
6 months
Filter on site;
HN03topH<2
6 months
Filter on site
6 months
6 months
24 hours
Store at 4°±2°C
until analyzed
One month
to extraction, 4 days
after extraction
Store at 4°±2°C
until analyzed
Mercury:
Aqueous
Total
Dissolved
Solid
Total
100 400 HNO3topH<2
28 days
100 400 Filter;
HNO3 to pH <2
28 days
0.2 g 200 g 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 the 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
Initial analysis of TEG* standard
Analysis using sub-boiled distilled acids
Analysis in class 100 hood
Analysis using sub-boiled acids in class 100 hood
Pb (ng)
3301250
260±200
20±8
2±1
Ag (ng)
970±500
2071200
312
* TEG - Trace Element in glass, SUMS 610 - 619,1 s.
TABLE 3-3
EXAMPLES OF LEAD CONCENTRATIONS IN AIR
SITE
LEAD CONCENTRATION (ug m'3)
Downtown Air, St. Louis, MO
Rural Park Air, Southeastern MO
Laboratory Air, NIST, MD
18.84 (Ref. 9)
0.77 (Ref. 10)
0.4 (Ref.3)
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CLASS*
TABLE 3-4
CLEANLINESS LEVELS IN FEDERAL STANDARD 209a (Ref. 11)
MAXIMUM CONTAMINATION IN WORK AREA (particles ff3)
100
10,000
100,000
100 particles > 0.5 um
0 particles > 5.0
10,000 particles > 0.5 um
65 particles > 5.0 um
100,000 particles > 0.5 um
700 particles > 5.0 um
AThe standard required laminar-flow equipment to attain this level of cleanliness.
Since measurement of dust particles smaller than 0.5 um introduces substantial errors, 0.5 um has
been adopted as the criterion of measurement.
TABLE 3-5
PARTICULATE CONCENTRATIONS IN LABORATORY AIR (Ref. 10)
SITE
Ordinary Laboratory
Clean Room
Clean Hood
Iron
0.2
0.001
0.0009
CONCENTRATION (ua m*)
Copper Lead
0.02 0.4
0.002 0.0002
0.007 0.0003
Cadmium
0.002
ND
0.0002
ND - Not Detected
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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*. Tefzel*
Quartz - Synthetic
Polyethylene (suitable for storgage only, not for acid digestion)
Quartz - Natural
Borosilicate Glass
* Various forms of Teflon®
THREE -14 Revision 4
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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)
•g
o
U
'•5
OH
O)
• vH
*^^
•^
7
E
3
U
610°
5106
f
4106
3106
2106
HO6
n
—
r
i
i
i
I
I
-i
+
*
•
•
_!
^
•
•
»
•
— •
•
•
•
«
Laboratory >.
Particle count 5.8 x 106
Volume of air 100 L
Clean Room
Particle count 504
Volume of air 378,000 L
0
10 20 30
Particle Diameter (jam)
40
THREE-15
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FIGURE 2
PARTICLE SIZE COMPARISON CHART FOR COMMON PARTICULATES (Refs. 1, 12)
Scanning Electron Microscope Optical Microscope ^,,^°
Naked Eye
Molecular Range Macro Molecular Range
small
organic
molecules
(sugars)
seasc
tobaccc
carbon black
virus _
• ^>
metallurj
^ photochi
It nuclei '
r -^ —
I
smoke
I ^
I
paint pig
I
I ^
fumes
•^ ^
I
I
oil smoke _
I
;ical dust) and fun
mical smog
I
I i
I
Macro
Micro Particle Range Particle
Range
fly as
bacteria
nent
^
cement c
h
^
human
hair
t pollens
list
"
milled flour
^ 1 Mi
coal
red
blood
cells
es ^ ^
nsecticide dusts
dust
^
0.001
0.01
0.10 0.5 1.0
Average particle diameter (|im)
10.0
100.0
THREE - 16
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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. The unfiltered or filtered sample is
heated with dilute HCI and HNO3 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 acid 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 acid 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
acid. 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 acid, hydrochloric acid, 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 acid and hydrogen peroxide followed by dilution with either nitric or hydrochloric
acid. 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 acid 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 including ash, biological tissue,
oil, oil contaminated soil, sediment, sludge, and soil for total analysis by FLAA, CVAA, GFAA, ICP-
AES, and ICP-MS. Nitric acid and hydrofluoric acid 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(lll).
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.
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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 acid
extraction/dissolution of available metals in aqueous samples, drinking water, mobility-procedure
extracts, and wastes that contain suspended solids for the following elements:
Element
CASRN*
Aluminum
Antimony
Arsenic
Barium
Beryllium
Boron
Cadmium
Calcium
Chromium
Cobalt
Copper
Iron
Lead
Magnesium
Manganese
Mercury
Molybdenum
Nickel
Potassium
Selenium
Silver
Sodium
Strontium
Thallium
Vanadium
Zinc
(Al)
(Sb)
(As)
(Ba)
(Be)
(B)
(Cd)
(Ca)
(Cr)
(Co)
(Cu)
(Fe)
(Pb)
(Mg)
(Mn)
(Hg)
(Mo)
(Ni)
(K)
(Se)
(Ag)
(Na)
(Sr)
(Tl)
(V)
(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
aChemical Abstract Service Registry Number
*Elements which typically require the addition of HCI for optimum recoveries. 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 (see Sec. 9.0).
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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 acid, 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 HCI may limit the methods of detection, or increase the difficulties
of detection with some techniques.
Due to the rapid advances in microwave technology, consult 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
will 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 fluorocarbons 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 acids 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 acid 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 acid 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 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 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 loss 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 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 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.
3015A-4 Revision 1
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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 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 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 (HNO3). 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 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 HNO3.
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.
3015A-5 Revision 1
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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 acid 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.
3015A-6 Revision 1
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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 watts, specified in the procedure to reproduce the heating profile
specified in Section 1 1 .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 ± 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
t
Where:
P = the apparent power absorbed by the sample in watts (W) (joule sec*1)
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'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 °C~1), 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
3015A-7 Revision 1
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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 acid (greater than 80 °C, but less than boiling) for a minimum of two hours
followed by hot (1:1) nitric acid (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 acids
(approximately 10% V/V) appropriate for the specific material used and then rinsed with reagent
water and dried in a clean 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 acid or, alternatively, 4 ± 0.1 mL
concentrated nitric acid and 1 ± 0.1 mL concentrated hydrochloric acid to the vessel in a fume
hood (or fume exhausted enclosure). The addition of concentrated hydrochloric acid 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
and silver upon addition of HCI have been described in the literature (Reference 7). The
addition of hydrochloric acid may, however, limit the detection techniques or increase the
difficulties of analysis for some detection systems.
CAUTION: The addition of hydrochloric acid must be in the form of concentrated
hydrochloric acid and not from a premixed combination of acids. A build-up 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.
3015A-8 Revision 1
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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. Properly 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 acids.
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 remaining 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
3015A- 9 Revision 1
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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 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 clog
nebulizers or interfere with injection of the sample into the instrument, 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 SiO2, TiO2, 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 for
concentration of analytes prior to analysis. The chemistry and volatility of the analytes 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.
3015A-10 Revision 1
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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) = (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 (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 acid 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.
3015A-11 Revision 1
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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, LB. 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 acid 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.
SamplePrep 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.
3015A-12 Revision 1
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TABLE 1
RESULTS OF VALIDATION STUDY FOR METHOD 3015 (NITRIC ONLY)
(REFERENCE 6)
Element
Al
As
Ba
Be
Ca
Cd
Co
Cr
Cu
Fe
K
Mg
Mn
Na
Ni
Pb
Se
V
Zn
TM-11
Mean
480
140
45
240
64
78
290
61
280
280
530
56
Std
Dev
26
23
2
14
4
4
16
3
16
32
26
3
TM-12
Mean
2800
2400
240
1150
350
320
1180
300
1290
1360
2400
520
Std
Dev
88.
70
8
36
10
9
43
9
39
35
61
9
T-95
Mean
5000
35000
20000
65.97
Std
Dev
784
1922
10690
2.65
T-107
Mean
210
13
200
11.3
12000
12
23
42
60
2200
53
2300
30.1
13
31
Std
Dev
19
1
16
0.5
783
1
1
4
9
110
3
1056
0.2
1
3
T-109
Mean
120
90
26
59000
10
30
34
130
2600
10200
47
13800
61
39
70
Std
Dev
31
11
1
999
2
6
4
7
383
218
3
516
2
1
4
Element
Sb
Tl
Aq
WP980 #1
Mean
18.0
55
Std
Dev
0.5
2
WP980 #2
Mean
110
7.0
Std
Dev
34
0.5
WS378 #4
Mean
ND
Std
Dev
WS378#12
Mean
19
Std
Dev
5
3015A-13
Revision 1
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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
Ag
B
Be
Co
Hg
Mo
Ni
Sr
V
Zn
5 mL HNO3
digest
0.31 ± 0.05
23.8 ±3.1
0.81 ±0.13
12.0 ±0.30
—
2.97 ± 0.72
39.6 ±2.5
41.9 ±1.3
6.18 ±2.5
418 ±12
4 mL HNO, +
1 mL HCI digest
0.41 ± 0.09
30.6 ± 8.3
0.91 ±0.19
11. 5 ±0.98
1.49 ±0.03
3.15 ±0.28
41.3 ±1.7
49.0 ±1.6
14.6 ±2.4
412 ±31
Total Analyte
Concentration
<4
_»*
__ _*
14.0 ± 0.6
1.44 ±0.07
___*
44.1 ±3.0
(130)
95 ±4
438 ± 12
Results reported in ug/g analyte (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 analyte 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
Ag
B
Co
Mo
Ni
Pb
Sb
Sr
5 mL HNO,
digest
1.3110.12
32.9 ±2.1
10.5 ±0.34
0.99 ± 0.06
12.2 ±1.2
135 ±4
3.7 ± 0.30
140 ±6
4 mL HNO, +
1 mL HCI digest
1.62 ±0.11
31.8 ±2.7
10.4 ±0.41
1.1 ±0.11
13.1 ±1.9
136 ±4
5.2 ± 0.53
143 ±7
Total Analyte
Concentration
(1.9)*
(63)*
14.8 ± 0.76
(1.7)*
(13)*
129 ±26
14.3 ±2.2
(330)
Results reported in ug/g analyte (mean ± 95% confidence limit).
Total concentrations are taken from NIST SRM Certificate of Analysis.
* Values in parenthesis are reference concentrations.
3015A-15
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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 HNO3 AND 4 ML HNO3 + 1 ML HCL)
OF METHOD 3015.
u
o
0)
5
13
O)
1
H
180
160
140
120
100
80
60
40
20
r-
-
-
-
-
-
-
-
—
7j
J
temperature
5:0 and 4:1
digest pressure
0:0 (water only)
digest pressure
n 20
- 15
- 10
- 5
*3
i^
en
fD
»T
rf
3
0
0
10 15 20
Time (min)
25
30
3015A-16
Revision 1
January 1998
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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 (HNO3), 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:
Element
CASRN8
Aluminum
Antimony
Arsenic
Barium
Beryllium
Boron
Cadmium
Calcium
Chromium
Cobalt
Copper
Iron
Lead
Magnesium
Manganese
Mercury
Molybdenum
Nickel
Potassium
Selenium
Silver
Sodium
Strontium
Thallium
Vanadium
Zinc
(Al)
(Sb)
(As)
(Ba)
(Be)
(B)
(Cd)
(Ca)
(Cr)
(Co)
(Cu)
(Fe)
(Pb)
(Mg)
(Mn)
(Hg)
(Mo)
(Ni)
(K)
(Se)
(Ag)
(Na)
(Sr)
(TO
(V)
(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
"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 oUs, 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
3051A-1
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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
80
Acid Ratio (NitricrHydrochloric)
3051A - 22
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FIGURE 7
PERCENT RECOVERY OF ANTIMONY AND IRON, RESPECTIVELY, FROM SRM 4355
(PERUVIAN SOIL) USING BOTH DIGEST OPTIONS
(10 ML HN03 AND 9 ML HNO3 + 3 ML HCL ) OF METHOD 3051
(N=6) (Refs. 6, 7)
100
80
> 60
o
u
40
20
9:3 Sb
9:3 Fe
10:0 Fe
10:OVSb
Acid Ratio (NitricHydrochloric)
3051A - 23
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METHOD 3051A
MICROWAVE ASSISTED ACID DIGESTION OF SEDIMENTS. SLUDGES SOILS AND Oil
10 1 Calibrate
microwave equipment.
11 2 Acid wash and water nrtM
all digestion vessels and
glassware
11 31 Weigh aliquot into the
digestion vessel.
11.3.2 Add 10* 0.1 mL of cone .
alternatively, 9*0.1 mL of cone. HNCg
and 3*0.1 ml of cone. HCI.
1135 Heat samples according to time
or temperatute and oretsure prefile.
1136 Allow vessels to cool
to room temperature
11 37 Vent each vessel. Transfer
sample to an actd cleaned bottle
11.3.8 May remove or reduce HNO3
and HCI by evaporation near
dryness with controlled purge gas.
11 3.9 Use appropriate elemental
analysis technique and/or SW-846
methods.
12.0 Calculate concentrations based
on ongmal sample weight.
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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
atomic 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.
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
nitrous-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.
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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
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 7741 A:
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 Voltammetrv
(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 Voltammetrv
(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 SOIL BY 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.
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.
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, H 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 Revision 0
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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 Iog10 of concentration is used, the plot
assumes a sigmoidal shape, when the log 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 analytes (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 chromogenic, fluorogenic, chemiluminescent or potentiometric reaction, (see
Immunoassay and ELISA)
3.7 False Negatives - A negative interpretation of the sample containing the target analytes
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 analytes 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.
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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 uL.
6.8 Squirt bottle - 500 mL or equivalent.
6.9 Graduated cylinder - 500 mL or equivalent.
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.
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 HNO3
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.
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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 uL) into the corresponding dilution bottles.
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11.4 Sample Analysis
11.4.1 Add diluted samples to the black line on each mercury assay tube (500 uL)
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 uL), 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 uL), 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 uL), and incubate
for 5 minutes at ambient temperature. Add three drops (105 uL) 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 "<[control 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 ">[control 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 Revision 0
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be determined to be a false negative. None of the experiments yielded false positives or false
negative results.
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., Carlson, L.D., 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, L.D., 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 letter and spirit of any sewer discharge permits and regulations, and
4500 - 6 Revision 0
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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 (ELISA) 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.
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TABLE 1
CORRELATION OF IMMUNOASSAY AND CVAA ANALYSES
FOR MERCURY IN SOIL AT A SUPERFUND SITE
Hg cone, by CVAA (ppm)
1.0
0.2
0.02
0.03
<0.02
<0.02
0.02
36.2
7.4
0.03
0.3
0.03
0.03
0.1
<0.03
0.9
0.03
0.04
<0.02
39.4
46.5
18.2
139
106
4.7
0.4
Hg cone, by Immunoassay
(ppm)
<5
<5
<5
<5
<5
<5
<5
>15
5-15
<5
<5
<5
<5
<5
<5
<5
<5
<5
<5
>15
>15
>15
>15
>15
>15
<5
Agreement a
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
N-FP
Y
(uontmueoj
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TABLE 1 (Continued)
Hg cone, by CVAA (ppm)
1.0
0.2
0.3
4.1
<0.02
0.05
56.6
0.5
0.2
0.1
0.3
0.02
0.04
0.08
0.03
0.02
<0.01
0.2
0.06
<0.01
<0.01
28.1
51.8
21.8
7.7
0.4
18.0
Hg cone, by Immunoassay
(ppm)
<5
<5
<5
5-15
<5
<5
>15
<5
<5
<5
<5
<5
<5
<5
<5
<5
<5
<5
<5
<5
<5
5-15
>15
5-15
5-15
<5
>15
Agreement*
Y
Y
Y
N-FP
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
Y
N-FN
Y
N-FN
Y
Y
Y
(Continued)
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TABLE 1 (Continued)
Hg cone, by CVAA (ppm)
0.8
2.2
4.4
1.1
0.05
1.3
0.06
<0.02
<0.01
3.1
3.4
0.3
3.4
2.0
0.13
0.06
Hg cone, by Immunoassay
(ppm)
<5
<5
5-15
5-15
<5
5-15
<5
<5
<5
5-15
5-15
<5
5-15
5-15
<5
<5
Agreement a
Y
Y
N-FP
N-FP
Y
N-FP
Y
Y
Y
N-FP
N-FP
Y
N-FP
N-FP
Y
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)
1.0 0.50 0.25 0.00
SAMPLE CONC. (ppm)
1.0 0.50 0.25 0.00
#1)
#2)
#3)
#4)
#5)
A)
B)
C)
D)
A)
B)
C)
D)
A)
B)
C)
D)
A)
B)
C)
D)
A)
B)
C)
D)
0.97
0.93
0.84
0.88
1.05
0.99
1.00
1.08
1.17
1.11
0.95
0.99
0.91
0.87
0.78
0.90
1.15
1.11
1.07
1.09
0.75
0.74
0.73
0.70
0.81
0.83
0.65
0.88
0.80
0.82
0.62
0.80
0.76
0.66
0.67
0.69
0.61
0.67
0.66
0.54
0.53
0.50
0.51
0.51
0.58
0.43
0.57
0.48
0.58
0.52
0.43
0.51
0.36
0.49
0.42
0.39
0.46
0.35
0.48
0.50
0.08
0.06
0.08
0.08
0.10
0.10
0.10
0.12
0.08
0.11
0.10
0.11
0.09
0.07
0.06
0.06
0.07
0.07
0.07
0.09
0.99
1.00
0.98
0.98
1.02
0.90
0.97
0.94
1.06
1.03
1.00
1.03
1.07
1.15
1.26
1.16
0.88
0.94
0.90
0.88
0.48
0.50
0.53
0.49
0.52
0.50
0.49
0.53
0.55
0.54
0.52
0.54
0.57
0.58
0.57
0.57
0.47
0.48
0.47
0.47
0.24
0.25
0.25
0.24
0.24
0.24
0.25
0.25
0.26
0.28
0.28
0.27
0.28
0.31
0.30
0.28
0.24
0.23
0.26
0.24
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
Source: Reference 5
4500-11
Revision 0
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TABLE 3
CROSS-REACTIVITY OF ELEMENTS WITH THE MERCURY IMMUNOASSAY
ELEMENT
Mercury
Arsenic
Barium
Cadmium
Chromium
Copper
Gold
Iron
Lead
Nickel
Silver
Sodium
Strontium
Thallium
Zinc
SOIL EQUIVALENT CONCENTRATION
REQUIRED TO YIELD A POSITIVE RESULT
(ppm)
0.36
>55,000
>1 00,000
>82,000
38,000
47,000
144,000
>41,000
>1 50,000
>43,000
79,000
>17.000
>64,000
>1 50,000
>48,000
Source: Reference 3
4500 - 12
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TABLE 4
ANALYSIS OF CERTIFIED REFERENCE SOILS USING IMMUNOASSAY
REFERENCE SAMPLE
ERA Inorganic Blank Soil
NIST 2709
NIST 2704
ERA CLP Lot #216
ERA Custom Mercury Std.1
dil. 1
dil. 2
dil. 3
NIST 8408
[Hg]
(ppm)
<0.10
1.40
1.47
2.36
4
15
50
122
ABSORBANCE @ 405-nm
exp.1
0.12
1.01
0.78
1.54
1.76
1.99
2.04
2.55
exp.2
0.05
0.64
0.41
0.84
1.01
1.45
1.73
2.55
exp.3
0.08
0.47
0.47
0.932
0.83
1.59
2.02
2.55
INTERPRETATION
3
<4
<4
<4
3
3
>15
>15
Source: Reference 2
1 dilutions from 107 ppm.
2 only value that gives incorrect conclusion.
3 standard reference point, no interpretation.
TABLE 5
ANALYSIS OF MULTIPLE MATRICES AT AN ABANDONED BATTERY RECLAMATION SITE
USING IMMUNOASSAY
SAMPLE DESCRIPTION
IMMMUNOASSAY RESULTS
CVAA RESULTS
Process Room
Dust from process room
Groundwater - unfiltered
Soil, alkaline
Sludge from tank
Sump sludge
Cinder block
Cinderblock duplicate
Soil
Paint
Background cinderblock
Background paint
Debris from CO2 blast
<5ppm
<5ppm
< 0.5 ppb
<5ppm
> 15 ppm
5 > 15 ppm
< 5 ppm
< 5 ppm
5 > 1 5 ppm
> 15 ppm
< 5 ppm
> 15 ppm
> 15 ppm
0.83 ppm
> 4.5 ppm
< 0.4 ppb
0.93 ppm
4,400 ppm
14 ppm
3 ppm
14 ppm
34 ppm
1 .4 ppm
14 ppm
19 ppm
Source: Reference 2
4500-13
Revision 0
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FIGURE 1
TEN-MONTH IMMUNOASSAY KIT STABILITY
2.50
Log[Hg](ppb)
4500 - 14
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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-ug/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-leachable) elements are required.
1.2 ICP-MS has been applied to the determination of over 60 elements in various matrices.
Analytes for which EPA has demonstrated the acceptability of Method 6020 in a multi-laboratory
study on solid and aqueous wastes are listed below.
Element
CASRN"
Aluminum
Antimony
Arsenic
Barium
Beryllium
Cadmium
Calcium
Chromium
Cobalt
Copper
Iron
Lead
Magnesium
Manganese
Mercury
Nickel
Potassium
Selenium
Silver
Sodium
Thallium
Vanadium
Zinc
(Al)
(Sb)
(As)
(Ba)
(Be)
(Cd)
(Ca)
(Cr)
(Co)
(Cu)
(Fe)
(Pb)
(Mg)
(Mn)
(Hg)
(Ni)
(K)
(Se)
(Ag)
(Na)
(Tl)
(V)
(Zn)
7429-90-5
7440-36-0
7440-38-2
7440-39-3
7440-41-7
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
7440-02-0
7440-09-7
7782-49-2
7440-22-4
7440-23-5
7440-28-0
7440-62-2
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
6020A -1
Revision 1
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TABLE 7
COMPARISON OF TOTAL MERCURY RESULTS IN HEAVILY CONTAMINATED SOILS
Soil Sample
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
Mercury in ug/g
ICP-MS
27.8
442
64.7
339
281
23.8
217
157
1670
73.5
2090
96.4
1080
294
3300
301
2130
247
2630
CVAA
29.2
376
58.2
589
454
21.4
183
129
1360
64.8
1830
85.8
1190
258
2850
281
2020
226
2080
Source: Reference 16.
6020A - 21
Revision 1
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METHOD 6020A
INDUCTIVELY COUPLED PLASMA - MASS SPECTROMETRY
7.1 Use
appropriate
digestion
procedure
(Chapter Three.)
for the analytes
Readjust
instrument per
manufacturers'
recommendations
all masses that
could affect data
configuration of
Are
results
outside +/-
10% range of
actual
value?
7.3 Set up
and stabilize
instrument.
check standard
blank after each
Does
response
exceed
calibration
curve
range?
7.11 Calculate
concentration.
^
f
6020A-22
Revision 1
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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 This 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 clean 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.
6200 -1 Revision 0
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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, for 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 ((3), 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 Kp line is produced by
a vacancy in the K shell filled by an M shell electron. The KQ transition is on average 6 to 7 times
more probable than the K^ transition; therefore, the K,,, line is approximately 7 times more intense
than the K,, 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 (La 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 elements 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 kiloelectron 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
6200 - 2 Revision 0
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proportional to the energy of the x-rays. An electronic multichannel analyzer (MCA) measures 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 FP: 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 gV_: 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
6200 - 3 Revision 0
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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 Kp line of element Z-1 with the K,, line of
element Z. This is called the ^/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 K,, energy is 5.41 keV. The Fe K., and Kp energies are 6.40
and 7.06 keV, respectively, and the Co K,, energy is 6.92 keV. The difference between the V Kp and
Cr ^energies is 20 eV, and the difference between the Fe Kp and the Co K,, 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 Cr or 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)-
6200 - 4 Revision 0
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4.7 Other interferences can arise from K/L, K/M, and L/M line overlaps, although these
overlaps are less common. Examples of such overlap involve arsenic (As) Ka/lead (Pb) L^ and sulfur
(S) Ke/Pb Ma. In the As/Pb case, Pb can be measured from the Pb Lp line, and As can be measured
from either the As «„ or the As KB 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 and 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 Revision 0
January 1998
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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, this 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 instrument 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 all 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.
The 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 Tnode, 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
cooling via the Peltier effect. Instruments 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)-170 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 amplitudes (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 (urn) thick.
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6.5 Mortar and pestle: glass, agate, or aluminum oxide; for grinding soil and sediment
samples.
6.6 Containers: glass or plastic to store samples.
6.7 Sieves: 60-mesh (0.25 mm), 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 (MIST), 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 would 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 instalments, 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 verify 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 assess 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
SD = Standard deviation of the concentration for the analyte
Mean Concentration = Mean concentration for the analyte
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)*, 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 SD 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. The
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 with 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 instruments: 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 Trail! 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 = ((C8-Ck)/Ck)x100
where:
%D = Percent difference
Ck = Certified concentration of standard sample
Cs = 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
(Rayleigh) backscatter peaks of primary radiation. These peak intensities are known to be a
function of sample composition, and the ratio of the Compton to Rayleigh 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 ICP 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 ICP or AA. A total acid 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. The 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
analysis.
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
only 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
this method, this modest amount of sample preparation was found to take less than 5 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 and 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 for 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 pm 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.
6200-17 Revision 0
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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 instruments 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 All 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 analyte.
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 analyte
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 Revision 0
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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 analytes such as mercury, selenium, silver, and thorium were
not detected in any of the precision samples so these analytes are not listed in Table 5. Some
analytes 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
these 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 of the 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 analytes. 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 of the low or nondetectable concentrations of these three analytes in the soil and sediment
SRMs.
Only 12 analytes are presented in Table 7. These are the analytes 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 Revision 0
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value, and 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-silty 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-sample 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 and 1.00 indicating
the data would need to be corrected very little or not at all to match the confirmatory laboratory data.
The r2 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.
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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 and 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.
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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 PRO 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.
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TABLE 1
INTERFERENCE FREE DETECTION LIMITS
Analyte
Antimony (Sb)
Arsenic (As)
Barium (Ba)
Cadmium (Cd)
Calcium (Ca)
Chromium (Cr)
Cobalt (Co)
Copper (Cu)
Iron (Fe)
Lead (Pb)
Manganese (Mn)
Mercury (Hg)
Molybdenum (Mo)
Nickel (Ni)
Potassium (K)
Rubidium (Rb)
Selenium (Se)
Silver (Ag)
Strontium (Sr)
Thallium (Tl)
Thorium (Th)
Tin (Sn)
Titanium (Ti)
Vanadium (V)
Zinc (Zn)
Zirconium (Zr)
Chemical
Abstract
Series Number
7440-36-0
7440-38-0
7440-39-3
7440-43-9
7440-70-2
7440-47-3
7440-48-4
7440-50-8
7439-89-6
7439-92-1
7439-96-5
7439-97-6
7439-93-7
7440-02-0
7440-09-7
7440-17-7
7782-49-2
7440-22-4
7440-24-6
7440-28-0
7440-29-1
7440-31-5
7440-32-6
7440-62-2
7440-66-6
7440-67-7
Detection Limit in
Quartz Sand
(milligrams per kilogram)
40
40
20
100
70
150
60
50
60
20
70
30
10
50
200
10
40
70
10
20
10
60
50
50
50
10
Source: References 1, 2, and 3
6200 - 23
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TABLE 2
SUMMARY OF RADIOISOTOPE SOURCE CHARACTERISTICS
Source
Fe-55
Cd-109
Am-241
Cm-244
Activity
(mCi)
20-50
5-30
5-30
60-100
Half-Life
(Years)
2.7
1.3
458
17.8
Excitation Energy
(keV)
5.9
22.1 and 87.9
26.4 and 59.6
14.2
Elemental Analysis Range
Sulfur to Chromium K Lines
Molybdenum to Barium L Lines
Calcium to Rhodium K Lines
Tantalum to Lead K Lines
Barium to Uranium L Lines
Copper to Thulium K Lines
Tungsten to Uranium L Lines
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
Cu
Mo
Ag
Recommended
Voltage Range
(kV)
18-22
40-50
50-65
K-alpha
Emission
(keV)
8.04
17.4
22.1
Elemental Analysis Range
Potassium to Cobalt K Lines
Silver to Gadolinium L Lines
Cobalt to Yttrium K Lines
Europium to Radon L Lines
Zinc to Technicium K Lines
Ytterbium to Neptunium 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.
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TABLE 4
FIELD-BASED METHOD DETECTION LIMITS (mg/kg)1
Analyte
Antimony
Arsenic
Barium
Chromium
Cobalt
Copper
Lead
Manganese
Molybdenum
Nickel
Rubidium
Strontium
Tin
Zinc
Instrument
TN
9000
55
60
60
200
330
85
45
240
25
100
30
35
85
80
40
TN Lead
Analyzer
NR
50
NR
460
NR
115
40
340
NR
NR
NR
NR
NR
95
NR
X-MET 920
(SiLi
Detector)
NR
55
30
210
NR
75
45
NR
NR
NA
NR
NR
NR
70
NR
X-MET 920
(Gas-Filled
Detector)
NR
50
400
110
NR
100
100
NR
NR
NA
NR
NR
NR
NA
NR
XL
Spectrum
Analyzer
NR
110
NR
900
NR
125
75
NR
30
NA
45
40
NR
110
25
MAP
Spectrum
Analyzer
NR
225
NR
NR
NR
525
165
NR
NR
NR
NR
NR
NR
NA
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.
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TABLE 5
PRECISION
Analyte
Antimony
Arsenic
Barium
Cadmium
Calcium
Chromium
Cobalt
Copper
Iron
Lead
Manganese
Molybdenum
Nickel
Potassium
Rubidium
Strontium
Tin
Titanium
Zinc
Zirconium
Average Relative Standard Deviation for Each Instrument
at 5 to 10 Times the MDL
TN
9000
6.54
5.33
4.02
29.84"
2.16
22.25
33.90
7.03
1.78
6.45
27.04
6.95
30.85"
3.90
13.06
4.28
24.32"
4.87
7.27
3.58
TN Lead
Analyzer
NR
4.11
NR
NR
NR
25.78
NR
9.11
1.67
5.93
24.75
NR
NR
NR
NR
NR
NR
NR
7.48
NR
X-MET 920
(SiLi
Detector)
NR
3.23
3.31
24.80"
NR
22.72
NR
8.49
1.55
5.05
NR
NR
24.92"
NR
NR
NR
NR
NR
4.26
NR
X-MET 920
(Gas-Filled
Detector)
NR
1.91
5.91
NR
NR
3.91
NR
9.12
NR
7.56
NR
NR
20.92"
NR
NR
NR
NR
NR
2.28
NR
XL
Spectrum
Analyzer
NR
12.47
NR
NR
NR
30.25
NR
12.77
2.30
6.97
NR
12.60
NA
NR
32.69"
8.86
NR
NR
10.95
6.49
MAP
Spectrum
Analyzer
NR
6.68
NR
NR
NR
NR
NR
14.86
NR
12.16
NR
NR
NR
NR
NR
NR
NR
NR
0.83
NR
Source: Reference 4
" 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.
NR Not reported.
NA Not applicable; analyte was reported but was below the method detection limit.
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TABLE 6
PRECISION AS AFFECTED BY SAMPLE PREPARATION
Analyte
Antimony
Arsenic
Barium
Cadmium8
Calcium
Chromium
Cobalt
Copper
Iron
Lead
Manganese
Mercury
Molybdenum
Nickel8
Potassium
Rubidium
Selenium
Silver3
Strontium
Thallium
Thorium
Tin
Titanium
Vanadium
Zinc
Average Relative Standard Deviation for Each Preparation Method
In Situ-Field
30.1
22.5
17.3
41.2
17.5
17.6
28.4
26.4
10.3
25.1
40.5
ND
21.6
29.8
18.6
29.8
ND
31.9
15.2
39.0
NR
ND
13.3
NR
26.6
20.2
Intrusive-
Undried and Unground
15.0
5.36
3.38
30.8
1.68
28.5
31.1
10.2
1.67
8.55
12.3
ND
20.1
20.4
3.04
16.2
20.2
31.0
3.38
16.0
NR
14.1
4.15
NR
13.3
5.63
Intrusive-
Dried and Ground
14.4
3.76
2.90
28.3
1.24
21.9
28.4
7.90
1.57
6.03
13.0
ND
19.2
18.2
2.57
18.9
19.5
29.2
3.98
19.5
NR
15.3
3.74
NR
11.1
5.18
Source: Reference 4
8 These values may be biased high because the concentration of these analytes in the soil
samples was near the detection limit.
ND Not detected.
NR Not reported.
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TABLE 7
ACCURACY
Analyte
Sb
As
Ba
Cd
Cr
Cu
Fe
Pb
Mn
Ni
Sr
Zn
Instrument
TN 9000
n
2
5
9
2
2
8
6
11
4
3
8
11
Range
of
% Rec.
100-149
68-115
98-198
99-129
99-178
61-140
78-155
66-138
81-104
99-122
110-178
41-130
Mean
%Rec.
124.3
92.8
135.3
114.3
138.4
95.0
103.7
98.9
93.1
109.8
132.6
94.3
SD
NA
17.3
36.9
NA
NA
28.8
26.1
19.2
9.70
12.0
23.8
24.0
TN Lead Analyzer
n
5
__
__
6
6
11
3
__
—
10
Range
of
% Rec.
__
44-105
,,
__
38-107
89-159
68-131
92-152
—
__
81-133
Mean
%
Rec.
__
83.4
„
_
79.1
102.3
97.4
113.1
__
—
100.0
SD
_
23.2
__
_
27.0
28.6
18.4
33.8
_
_
19.7
X-MET 920 (SiLi Detector)
n
4
9
6
7
11
6
12
__
__
_„
12
Range
of
% r\ec.
_„
9.7-91
18-848
81-202
22-273
10-210
48-94
23-94
,,
— _
__
46-181
Mean
%
Rec
_
47.7
168.2
110.5
143.1
111.8
80.4
72.7
__
_
106.6
SD
__
39.7
262
45.7
93.8
72.1
16.2
20.9
_
„
«
34.7
XL Spectrum Analyzer
n
— .
5
_
—
3
8
6
13
__
3
7
11
Range
of
%Rec.
__
38-535
„
__
98-625
95-480
26-187
80-234
„
57-123
86-209
31-199
Mean
%
Rec.
189.8
_
^^
279.2
203.0
108.6
107.3
,__
87.5
125.1
94.6
SD
_
206
„
_
300
147
52.9
39.9
__
33.5
39.5
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
Revision 0
January 1°°B
-------
TABLc 8
ACCURACY FOR TN 9000'
Standard
Reference
Material
RTC CRM-021
RTC CRM-020
BCRCRM143R
BCRCRM141
USGS GXR-2
USGS GXR-6
NIST2711
NIST2710
NIST 2709
NIST 2704
CNRC PACS-1
SARM-51
SARM-52
Arsenic
Cert.
Cone.
24.8
397
_
_
25.0
330
105
626
17.7
23.4
211
—
—
Meas.
Cone.
NO
429
_
_
ND
294
104
722
ND
ND
143
_
—
%Rec.
NA
92.5
_
—
NA
88.9
99.3
115.4
NA
NA
67.7
—
—
Barium
Cert.
Cone.
586
22.3
—
—
2240
1300
726
707
968
414
—
335
410
Meas.
Cone.
1135
ND
—
—
2946
2581
801
782
950
443
772
466
527
%Rec.
193.5
NA
—
—
131.5
198.5
110.3
110.6
98.1
107.0
NA
139.1
128.5
Copper
Cert.
Cone.
4792
753
131
32.6
76.0
66.0
114
2950
34.6
98.6
452
268
219
Meas.
Cone.
2908
583
105
ND
106
ND
ND
2834
ND
105
302
373
193
AtlxOC.
60.7
77.4
80.5
NA
140.2
NA
NA
96.1
NA
106.2
66.9
139.2
88.1
Lead
Cert.
Cone.
144742
5195
180
29.4
690
101
1162
5532
18.9
161
404
5200
1200
Meas.
Cone.
149947
3444
206
ND
742
80.9
1172
5420
ND
167
332
7199
1107
%Rec.
103.6
66.3
114.8
NA
107.6
80.1
100.9
98.0
NA
103.5
82.3
138.4
92.2
Cert.
Cone.
546
3022
1055
81.3
530
118
350
6952
106
438
824
2200
264
Zinc
Meas.
Cone.
224
3916
1043
ND
596
ND
333
6476
98.5
427
611
2676
215
%Rec.
40.9
129.6
99.0
NA
112.4
NA
94.9
93.2
93.0
97.4
74.2
121.6
81.4
Source: Reference 4
' All concentrations in milligrams per kilogram.
%Rec. Percent recovery.
ND Not detected.
NA Not applicable.
— No data.
6200 - 29
Revision 0
January 1998
-------
TABLE 9
REGRESSION PARAMETERS FOR COMPARABILITY1
All Data
SoiM
Soil 2
Soil3
Prepl
Prep 2
Prep 3
Prep 4
All Data
SoiM
Soil 2
Soil 3
Prepl
Prep 2
Prep 3
Prep 4
Arsenic
n
824
368
453
—
207
208
204
205
r2
0.94
0.96
0.94
—
0.87
0.97
0.96
0.96
Int.
1.62
1.41
1.51
—
2.69
1.38
1.20
1.45
Slope
0.94
0.95
0.96
—
0.85
0.95
0.99
0.98
Lead
n
1205
357
451
397
305
298
302
300
r2
0.92
0.94
0.93
0.90
0.80
0.97
0.98
0.96
Int.
1.66
1.41
1.62
2.40
2.88
1.41
1.26
1.38
Slope
0.95
0.96
0.97
0.90
0.86
0.96
0.99
1.00
Barium
n
1255
393
462
400
312
315
315
313
r2
0.71
0.05
0.56
0.85
0.64
0.67
0.78
0.81
Int.
60.3
42.6
30.2
44.7
53.7
64.6
64.6
58.9
Slope
0.54
0.11
0.66
0.59
0.55
0.52
0.53
0.55
Zinc
n
1103
329
423
351
286
272
274
271
r2
0.89
0.93
0.85
0.90
0.79
0.95
0.93
0.94
Int.
1.86
1.78
2.57
1.70
3.16
1.86
1.32
1.41
Slope
0.95
0.93
0.90
0.98
0.87
0.93
1.00
1.01
Copper
n
984
385
463
136
256
246
236
246
r2
0.93
0.94
0.92
0.46
0.87
0.96
0.97
0.96
Int.
2.19
1.26
2.09
16.60
3.89
2.04
1.45
1.99
Slope
0.93
0.99
0.95
0.57
0.87
0.93
0.99
0.96
Chromium
n
280
—
—
186
105
77
49
49
r2
0.70
—
—
0.66
0.80
0.51
0.73
075
Int.
64.6
—
—
38.9
66.1
81.3
53.7
31 6
Slope
0.42
—
—
0.50
0.43
0.36
0.45
056
Source: Reference 4
1 Log-transformed data
n Number of data points
r2 Coefficient of determination
Int. Y-intercept
— No applicable data
6200 - 30
Revision 0
January 1998
-------
-------
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 uL 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 back 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.
6500 -1 Revision 0
January 1998
-------
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
Effluent
Drinking Water
Landfill
Leachate
Sample #
1
2
3
4
5
6
7
1
2
3
4
5
6
7
1
2
Cd Red'n1
0.3
-.
—
—
—
2.4
0.7
0.6
0.6
0.4
0.6
0.6
0.3
0.5
__
—
1C2
ND
ND
ND
ND
2.1
1.9
0.3
0.3
0.3
0.3
0.3
0.3
0.1
0.3
ND
ND
CIE
ND
ND
ND
0.5
2.4
2.2
0.4
0.4
4.4
4.4
0.3
0.4
0.1
0.4
ND
ND
Source: Reference 2.
1 Total nitrite + nitrate determined using 4500-NO3 F, Cadmium Reduction Method
2 Nitrite + nitrate determined using 4110 C, Single Column Ion chromatorgraphy 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
Revision 0
January 1998
-------
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
Effluent
Drinking Water
Landfill
Leachate
Sample #
1
2
3
4
5
6
7
1
2
3
4
5
6
7
1
2
Ascorbic Acid1
3.4
4.9
4.7
5.3
3.0
2.9
<0.1
<0.1
<0.1
—
<0.1
<0.1
—
—
<0.1
2.2
1C2
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
1.6
CIE
2.8
4.4
4.5
4.2
3.0
2.3
<0.1
ND
ND
ND
ND
ND
ND
ND
<0.1
1.4
Source: Reference 2.
1 Phosphate determined using 4500 PO4 E, Ascorbic Acid Method
2 Phosphate determined using 4110 C, Single Column Ion Chromatography Using Direct
Conductivity Detection
6500 - 21
Revision 0
January 1998
-------
TABLE 11
CAPILLARY ION ELECTROPHORESIS ANION ANALYSIS ROUND ROBIN1
USING CHROMATE ELECTROLYTE (mg/L)
Sample
1. Bleachwaste
2. Creekwater
3. Wastewater
4. Wastewater
5. Wastewater
6. Wastewater
7. Wastewater
8. Wastewater
9. Wastewater
10. Wastewater
1 1 . Surfacewater
12. Wastewater
13. Drinking Water
14. Drinking Water
Chloride
<0.046
3.06±0.27
24.6±0.62
59.7±2.9
63.8±2.0
72.0±5.4
139110.0
51.4±7.7
29.9±4.3
766±44
3.71 ±0.39
22.1 ±0.62
5.15±0.35
4.95±0.24
Bromide
<0.046
<0.046
<0.046
0.85±0.52
0.68±0.52
0.05±0.01
<0.046
<0.046
<0.046
<0.046
<0.046
8.47±0.30
<0.046
<0.046
Nitrite
<0.072
<0.072
<0.072
<0.072
<0.072
<0.072
4.0±1.3
<0.072
2.14±1.35
<0.072
<0.072
<0.072
<0.072
<0.072
Sulfate
0.30±0.37
3.00±0.30
2.02±0.56
109±4.4
115±3.9
144±11.8
584±35
40.2±6.1
217±19
489±46
2.70±0.39
133±4.4
2.64±0.26
2.62±0.21
Nitrate
<0.84
0.37±0.19
<0.084
44.9±1.6
44.3±1.06
5.38±2.57
353±25.5
39.9±7.9
13.9±4.9
12.9±6.9
0.23±0.20
<0.084
0.50±0.27
0.54±0.25
Fluoride
<0.020
0.11 ±0.09
0.08±0.08
0.988±0.21
1.04±0.17
0.57±0.21
3.01±0.80
1.17±0.24
1.33±0.28
<0.020
0.11 ±0.097
0.76±0.11
0.59±0.097
0.56±0.09
Phosphate
<0.041
<0.061
3.74±0.75
4.94±1.32
4.78±1.55
1.18±1.01
9.34±5.17
6.99±1.31
9.95±5.04
41.3±8.5
<0.041
<0.041
<0.041
<0.041
Source: Reference 2.
1 Five laboratory Intel-laboratory precision.
6500 - 22
Revision 0
January 1998
-------
FIGURE 1
HARDWARE SCHEMATIC OF A CAPILLARY
ION ELECTROPHORESIS SYSTEM
75 (im x 60 cm
Silica Capillary
Constant
Temperature
Compartment,
FIGURE 2
PICTORIAL DIAGRAM OF ANION MOBILITY AND
ELECTROOSMATIC FLOW MODIFIER
Cathode
Injection Side
Detection Side
— Si Si Si Si Si Si Si -Si -._ Si
<}. <4. d> ^o--" a 6. (4. ciH
N N N
N N N
N N N
6500 - 23
Revision 0
January 1998
-------
FIGURE 3
SELECTIVITY DIAGRAM OF ANION MOBILITY
USING CAPILLARY ION ELECTROPHORESIS
All
Cations
.^ ,
Inorganic
Anions
Cl Br
NO2,SO4,
NO3
SO3, SzOs
DiValent
Org Acids,
Oxymetals
F, PO4,
CIO2. CIO3.
Formate
0)
rbonal
MonoValent
Organic Acids
C2 thru Ce
Water
and All
Neutral
Organics
Migration Time
MT=0
High Mobility
Anions
Low Mobility
Anions
MT >7 min
FIGURE 4
PICTORIAL DIAGRAM OF INDIRECT UV DETECTION
e e
eeeeeAAAAeeeeeee
eeeeeeAAAAeeeeeee
High UV Absorbing
Electrolyte
Analyte ion (A) displaces electrolye ion (e)
charge for charge or transfer ratio causing
a net decrease in background absorbance.
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
11
3.000
3.500
Minutes
4.000
4.500
Electrolyte: 4.7 mM NajCrCy4.0 mM TTAOH /10 mM CHES / 0.1 mM Calcium Gluconate
Capillary: 75 \im (id) x 375 pm x 60 cm (length), Uncoated Silica
Voltage: 15 kV using a Negative Power Supply
Current 14 ± uA, Constant Current
Sampling: Hydrostatic at 10 cm for 30 seconds
Detection: Indirect UV using a Hg Lamp and 254 nm Filter
Anion
1 . Chloride
2. Bromide
3. Nitrite
4. Sulfate
5. Nitrate
6. Oxalate
7. Fluoride
8. Formate
9. O-Phosphate
10. Carbonate
11. Acetate
Cone.
Mg/L
2.0
4.0
4.0
4.0
4.0
5.0
1.0
5.0
4.0
5.0
Migration
Time in
Mintues
3.200
3.296
3.343
3.465
3.583
3.684
3.823
3.873
4.004
4.281
4.560
Migration
Time Ratio to
Cl
1.000
1.030
1.045
1.083
1.120
1.151
1.195
1.210
1.251
1.338
1.425
Peak
Area
1204
1147
2012
1948
1805
3102
1708
1420
2924
3958
Time
Corrected
Peak Area
376.04
348.05
601.72
562.05
503.69
842.14
446.65
366.61
730.25
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
Nitrate = 0.084
Bromide = 0.090
Fluoride = 0.020
Nitrite = 0.072
phosphate = 0.041
Sulfate = 0.032
3.200
3.400
3.600
Minutes
3.600
4.000
FIGURE 7
ELECTROPHEROGRAM OF TYPICAL DRINKING WATER
USING CHROMATE ELECTROLYTE
a
a = 24.72 mg/L
SO. = 7.99
NO, = 0 36
F < 0.10
HCO, = Natural
3000
4000
Minutes
6500 • 26
Revision 0
January 1998
-------
FIGURE 8
ELECTROPHEROGRAM OF TYPICAL MUNICIPAL WASTEWATER DISCHARGE
USING CHROMATE ELECTROLYTE
Anions in ma/L. No Dilution
1 Chloride = 93.3
2 Nitrite = 0.46
3 Sulfate =60.3
4 Nitrate =408
5 Carbonate = Natural
3.000
3.500
Minutes
4.000
4.500
FIGURE 9
ELECTROPHEROGRAM OF TYPICAL INDUSTRIAL WASTEWATER DISCHARGE
USING CHROMATE ELECTROLYTE
Aninng in mq/L. No Dilution
1 Chloride = 2.0
2 Nitrite = 16
= 34.7
= 16 5
< 0.05
6 Phosphate = 12.3
7 Carbonate = Natural
3 Sulfate
4 Nitrate
5 Formate
3.000
3500 Minutes
4500
6500 - 27
Revision 0
January 1998
-------
FIGURE 10
LINEARTY CALIBRATION CURVE FOR CHLORIDE, BROMIDE, AND SULFATE
USING CHROMATE ELECTROLYTE
10
7.5
(0
!
o
E
2.5
0
TCPA = 167 62 |CI| + 32.93; R2 = 0.9996
TCPA = 126.76(SO<1 -16.12; R2 = 0.9998
TCPA = 7B.23[Br| + 11.76; R2= 0 9995
0
3 Data Points per Concentration
Based upon Youden Pair Design
10
20 30
mg/L Anion
—r ~
40
50
FIGURE 11
LINEARTY CALIBRATION CURVE FOR FLUORIDE AND O-PHOSPHATE
USING CHROMATE ELECTROLYTE
3 Data Points per Concentration
Based upon Youden Pair Design
TCPA = 37667|FJ*1005; R2= 09985
TCPA = 158.25[PO«| - 19 68; R2= 0 9996
20 30
mg/L Anion
40
50
6500 - 28
Revision 0
January 1998
-------
FIGURE 12
LINEARITY CALIBRATION CURVE FOR NITRITE AND NITRATE
USING CHROMATE ELECTROLYTE
to
I
JC
CD
o> in
Q.-0
10
7.5
0)
.i 2.5
0
3 Data Points per Concentration
Based upon Youden Pair Design
TCPA = 130 95(NOj] * 28 93, R3 = 0 9996
TCPA = 106.82|NO3l » 126.61; RJ= 0 9992
NO2.
20 30
mg/L Anion
6500 - 29
Revision 0
January 1998
-------
METHOD 6500
DISSOLVED INORGANIC ANIONS IN AQUEOUS MATRICES
BY CAPILLARY ION ELECTRQPHQRFSIS
— '"•
Start
^
r
11.1 Set-up Capillary
Electrophoresis system
according to manufacturers
instructions.
^
r
11.2.1 Condition capillary
with NaOH for 5 min.
followed by chromate
electrolyte soln. for 5 min.
^
r
11.2.2 Program system for
hydrostatic sampling for
30 seconds.
>
r
11.2.3 Program system for
constant current 14 uj^ and
a run time of 5 min.
>
r
11.2.4 Program system for
data acquisition rate of
20 points per second.
>
r
11.2.5 Monitor UV response
at 254 nm.
^
r
11.3 Analyze all standards
and samples.
>
r
(^ Stop J)
6500 - 30
Revision 0
January 1998
-------
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 ug/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
Antimony
Boron
Barium
Cadmium
Calcium
Chromium
Copper
Iron
Lead
Magnesium
Mercury
Molybdenum
Nickel
Potassium
Selenium
Silver
Strontium
Thallium
Vanadium
Zinc
(Sb)
(B)
(Ba)
(Cd)
(Ca)
(Cr)
(Cu)
(Fe)
(Pb)
(Mg)
(Hg)
(Mo)
(Ni)
(K)
(Se)
(Ag)
(Sr)
(Tl)
(V)
(Zn)
CASRN3
7440-36-0
7440-42-8
7440-39-3
7440-43-9
7440-70-2
7440-47-3
7440-50-8
7439-89-6
7439-92-1
7439-95-4
7439-97-6
7439-98-7
7440-02-0
7440-09-7
7782-49-2
7440-22-4
7440-24-6
7440-28-0
7440-62-2
7440-66-6
a 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 solubilize 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(lll) and Cr(VI) will be used for demonstration purposes. Two isotopic
spikes are prepared and characterized: ^C^lll) spike enriched in 50Cr and 53Cr(VI) enriched in
53Cr. The dominant natural isotope for Cr is 52Cr, at 83.79% (^Cr, 4.35%; 53Cr, 9.50%; ^Cr,
2.36%). A measured amount of a representative aqueous sample is mixed well with an
appropriate amount of ^CrOII) and 53Cr(VI) spike solutions. The spiked sample is then
separated into Cr(lll) and Cr(VI) using chromatography or other separation method. Four
isotope ratios are measured: "Crpy^Crtlll), 53Cr(lll)/52Cr(lll), "CitVIJ/^CrfVI), and
^CrO/IJ/^CrO/l). 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(lll) or from Cr(lll) 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 deconvoluted.
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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 if 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 * «(!-«T)
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 o Isotopel _
^sample/standard ~ ^background
Isotope2_
^sample/standard ~ background
Rm is the dead-time-corrected isotope ratio;
and lsotope2S sampfe/standard and are the integrated dead-time-corrected-
counts for the sample or standard of Isotopel and Isotope2, respectively;
0 I80tope1 Background and ISOt0pe2Sbaekground are the integrated dead-time-corrected-counts for the
background of Isotopel and Isotope2, respectively.
As shown in Figure 1, which displays the ^Cr/^Cr ratios for SRM 979 (Cr(NO3)3«9H2O) 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 5.8x105. 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 material 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 Rn, 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:
RC = mass bias factor x Rm
where:
• Rc and Rm are the corrected isotope ratio and the measured dead-time-corrected-
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 argon 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(VI) and Cr(lll), including natCr(VI) and natCr(lll) with natural abundance, ^CrO/l) enriched in
enriched in ^Cr, and isotopic-abundance-certified Cr standard solution.
7.3.2 1 mg/mL Cr(VI) and Cr(lll) standards are commercially available. natCr(VI) and
natCr(lll) can also be prepared from K2Cr2O7 and Cr metal, respectively.
7.3.2.1 natCr(VI) standard solution, stock, 1 g = 1 mg Cr: Dissolve 0.2829
grams of K2Cr2O7 in about 80 mL of reagent water and dilute to 100 g with reagent
water.
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7.3.2.2 natCr(lll) standard solution, stock, 1 g = 1 mg Cr: Dissolve 0.1 g
Cr metal in a minimum amount of 6M HCI and dilute the solution with 1% HNO3 to 100
grams.
7.3.3 ^rfVI) standard solution, 1 g « 10 ug Cr: The following procedure describes
chromium oxide as the source material. A 150 mL glass or quartz beaker is used for the
dissolution. Weigh 5.8 mg (the exact amount should be calculated based on the content of Cr
in the material) ^Cr-enriched oxide into the beaker and add 8 ml concentrated HCIO4. Slowly
heat the beaker on a hot plate until bubbles form on the bottom; the solution should not boil.
Keep heating the solution for up to 6 hours until all solids are dissolved and only 1 to 2 mL of
the solution remains. Turn off the hot plate and wait until the beaker cools down. Rinse the
beaker and watch glass with 10 mL reagent water; the solution should turn intense yellow.
Add 50 uL of 30% H2O2 and 4.5 mL of concentrated NH4OH. Slowly heat the vessel until the
solution gently boils to oxidize all Crto Cr(VI). Allow the solution to boil for at least 15 minutes
to remove the excessive H2O2. Transfer the solution to a 500 mL polymeric (Teflon,
polyethylene, polypropylene, etc.) bottle and dilute the solution to 400 g. The exact
concentration of the 53Cr(VI) spike must be calibrated with natCr(VI) standard as described in
Section 7.4.
NOTE: The procedure may be simpler when the isotope-enriched materials are
available in other forms. For example, when K2Cr2O7 enriched in 53Cr is available, the
solid can be dissolved in reagent water without further conversion; when Cr metal is
available, the metal can be dissolved in 6M HCI as described in Section 7.3.2.2,
followed by the addition of H2O2 and NH4OH to oxidize Cr(lll) to Cr(VI) as described
above.
CAUTION: Concentrated HCIO4 is a very strong oxidizer. Safety protocols require this
reagent only be used in a perchloric acid hood or equivalent solution and vapor
handling system.
7.3.4 ^CrOII) standard solution, 1 g » 10 ug Cr: The following procedure describes
chromium metal as the source material. Weigh 4 mg of the metal into a 30 mL Teflon vessel.
Add 4 mL of 6M HCI and gently heat the solution but do not boil it until the solid is dissolved.
Continue to heat the solution until only 1 to 2 mL of the solution remains. The solution is then
cooled and transferred to a 500 mL polymeric bottle. Dilute the solution with 1% HNO3 to 400
grams. The exact concentration of the ^C^lll) spike must be calibrated with natCr standard as
described in Section 7.4.
NOTE: The procedure depends on the form of the material. For example, when
K2Cr2O7 enriched in ^Cr is available, the solid can be dissolved in 1% HNO3, followed
by the addition of H2O2 to reduce Cr(VI) to Cr(lll). The excessive H2O2 can be removed
by boiling the solution.
7.3.5 Isotopic-abundance-certified standard solution, stock, 1 g « 10 ug Cr: Weigh
31 mg Cr(NO3)3«9H2O (SRM 979) into a 500 mL polymeric container. Dissolve the solid and
dilute it with 1% HNO3 to 400 g.
7.4 The isotope-enriched spikes require characterization since a limited amount of material
is usually weighed, complex treatment is involved, or the purity of the source material is limited
(frequently <99% ). For the SIDMS method, in addition to the total concentration of the standard,
the distribution of the species must be determined before it can be used. Inverse IDMS and inverse
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SIDMS measurement is used to calibrate the isotope-enriched spike and to determine the species
distribution. The characterization of "CrO/l) spike solution will be illustrated as an example.
7.4.1 Calibration of total concentration of spike solution with natural material: Weigh
the proper amount (Wx) of 10 ug/g (Cg,,^) ""Cr standard and the proper amount (W?) of the
53Cr(VI) spike (nominal concentration is 10 ug/g ) into a polymeric container, and dilute the
mixture with 1% HNO3 to a concentration suitable for isotope ratio measurement. Use direct
aspiration mode to determine the isotope ratio of ^Cr/^Cr (R 53/52)- The concentration of the
spike, CSp|ke, can be calculated using the following equations.
CSpike = CSMS
c c*w*
s~~wT
CX = Cstandard
where, Cs and Cx 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. "Ag and ^Ax are the atomic
fractions of ^r for the spike and standard, respectively. HAS and 52AX are the atomic fractions
of 52Cr 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 ^Cr to ^Cr.
NOTE: Average atomic weight = Z(atomic weight of the isotope x atomic fraction)
7.4.2 Calibration of the concentration of the Cr(VI) in the ^(Vl) spike with natCr(VI):
Weigh the proper amount (Wx) of 10 ug/g (C*{andard)natCr(VI) standard and the proper amount
(Ws) of the ^CrfVI) spike (nominal concentration is 10 ug/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 HNO3. 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, 0^, can be calculated using the following equations.
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VI VI
..
M
W
w
53.
VI 52
p
^53/52
^53/52
'X = ^Standard /Mx
where, Cg and GX are the concentrations of Cr(VI) in the isotope-enriched spike and standard
with natural isotopic abundance in umole/g, respectively. Msand Mx are the average atomic
weights of the spike and the standard in g/mol, respectively. "^ and ^x are the atomic
fractions of ^r for the spike and standard, respectively. KAS and 52>AX are the atomic fractions
of 52Cr for the spike and standard, respectively.
NOTE: This set of equations is similar to those used in the determination of total Cr in
^(Vl) standard (Section 7.4.1). The general equations for inverse SIDMS are not so
simple. However, for speciation 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(lll) 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 ^CrfVI) spike can be calculated as:
percentage of Cr(VI) =
Spike
100%
percentage of Cr(lll) =
N VI
'Spike
'Spike
X 100%
NOTE: No determination of the species distribution in ^Crflll) 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% HNO3 (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 % HNO3 (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 speciation 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 quality 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(lll) 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 (Cr(NO3)3»9H2O) 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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1 1 .2.2 Prepare the isotope-enriched spikes in species forms and 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.
1 1 .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 ug/g 53Cr(VI) spike to a concentration so
that the isotope ratio of ^CrFCr in Cr(VI) will be approximately 1:1. Thoroughly mix the spike
and the sample. The isotope ratios ^Cr/^Cr for samples must be within the range of 0.1:1 to
10:1, except for blanks or samples with extremely low concentrations.
1 1 .2.5 Dilute the ^(-(V^-spiked sample with reagent water. If the solution is strongly
basic, neutralize the sample with concentrated HNOs to avoid the hydrolysis of Cr(lll). Spike
the diluted sample with 10 ug/g ^C^lll) 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 ^Cr/^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
instead of double-spiking with both ^C^lll) and 53Cr(VI). However, this is based on the
assumption that only unidirectional conversion, the reduction Cr(VI) to Cr(lll), 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
Cr(VI)). 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 KMnO4 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(lll) and 53Cr/52Cr 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-Calculations: The quantitative values shall be reported in appropriate units, such
as micrograms per liter (ug/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, CSamp|e(ug/g), in the final sample solutions.
CSample = CXMX
CS = CSpike /MS
csws
wx
AS °53/S2 ^5
R 52A -53A
^53/52 rtX ^X
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 in g/mole, respectively. ^As and 53AX are the atomic fractions of 53Cr for
the isotope-enriched spike and sample, respectively. 52AS and 52AX are the atomic fractions of
ffiCr for the isotope-enriched spike and sample, respectively. Cspike is the concentration of the
isotope-enriched spike in [ig/g.
NOTE: When isotope ^Cr 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.
Q
Concentration (dry weight)(mg/kg) = SamP|e
where, CSam^i = Concentration based on the wet sample (ug/g)
% Solids
S =
100
12.2 SIDMS-Calculations: The quantitative values shall be reported in appropriate units,
such as micrograms per liter (ug/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.
cX+X'cX') (i-g)
,„
53/52 ~
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where,
R'^/52 is the measured isotope ratio of ^Cr to 52Cr of Cr(lll) in the spiked sample
MAX is the atomic fraction of ^Cr in the sample (usually a constant in nature)
CJJ is the concentration of Cr(lll) in the sample (umole/g, unknown)
Wx is the weight of the sample (g)
50 A.™ is the atomic fraction of ^Cr in the ^Crflll) spike
Cs" is the concentration of Cr(lll) in the ^Crflll) spike (umole/g)
Ws" is the weight of the ^CrflM) spike (g)
C^1 is the concentration of Cr(VI) in the sample (umole/g, unknown)
a is the percentage of Cr(lll) oxidized to Cr(VI) after spiking (unknown)
(3 is the percentage of Cr(VI) reduced to Cr(lll) after spiking (unknown)
NOTE: The unit of the concentrations shown above is umole/g. The conversion factor
from umole/g to ug/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 (umole/g)x M = Concentration (ug/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 ^Crflll) and 53Cr(VI)
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 = XX
-A/1 /MVI
'Spike'MS
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53. vi DVI 52.VI
AS~ K53/52 AS
pVI 52. 53A
^53/52 MX MX
where, C™ and C^ are the concentrations of the isotope-enriched spike and the sample
in umole/g, respectively. Ms and Mx are the average atomic weight of the isotope-
enriched spike and the sample in g/mole, respectively. ^Ag and KAX are the atomic
fraction 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.
Cspike is the concentration of the isotope-enriched spike in ug/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, dx, C^', 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
W -
W ~
HIW - NIH C^ W = N
SVVS ~ INS' °S VVS IN
At the beginning of the iteration, arbitrary values can be assigned to Nx and a. For example,
both of them are assigned as Os. Now we need to know the expression of Nx' and p. After careful
derivation, we can get the following equations:
P = -
P = -
These equations can be rewritten as:
The solutions are
B2
and
P =
A2 C
B,
B
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Use these two values in the following equations to solve NX and a
52A -53 " VI 52 '" 52
A
2A N'" + 52AIIINIII\
AXNX + ASNs) ~
a - I-RVI 5V
u ' ^53/52 MS
Rewrite the equation as:
B3a = C3
B4« = C4
again
'3
'4 B4
A3 B3
B
and
a =
V3 °3
^4 C4
A.3 B
Repeating the calculation, the variables N^', N^' , a and (3 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 IDMS 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
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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 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. Fasset, J. D.; Paulsen, P. J. Isotope Dilution Mass Spectrometry Anal. Chem. 1989, 61, 643A-
649A.
2. Kingston, H. M. Method ofSpeciated 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. James, B. R.; Petura, J. C.; Wale, R. J.; Mussoline, G. R. Environ. Sci. Tech. 1995, 29, 2377.
6800-21 Revision 0
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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.sampleprep.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
(ng/g)
Days after
Aliquot spiking
1
1 4
13
1
2 4
13
1
3 4
13
True
Cr(lll)
69.8 ± 0.3
69.2 ± 0.6
70.5 ± 0.9
69.6 ± 0.2
69.3 ± 0.7
70.7 ± 0.4
69.8 ± 0.6
69.0 ± 0.8
70.4 ± 0.5
69.67
Cr(VI)
68.8 ± 0.3
69.4 ± 0.3
68.5 ± 0.4
68.8 ± 0.4
69.6 ± 0.6
68.8 ± 0.3
69.0 ± 0.2
69.6 ± 0.3
68.9 ± 0.8
68.63
Conversion
(%)
Cr(lll) to
Cr(VI)
4.87 ± 0.22
3.47 ±0.11
2.80 ±0.1 3
17.6 ±0.1
14.6 ±1.3
12.8 ±0.1
23.8 ± 0.3
21 .6 ±0.2
17.6 ±0.3
Cr(VI) to
Cr(lll)
3.57 ± 0.03
11.9±0.5
22.4 ± 0.2
2.95 ± 0.02
11.4 ±0.7
22.1 ± 0.3
2.76 ± 0.08
10.2 ±0.1
22.1 ± 0.1
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(VI). Measurements were
done on different days to check the stability of the species during storage. Despite the different degrees of
interconversion, the deconvoluted concentrations for both Cr(lll) 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)
sample
COPR1
COPR3
COPR4
Method
Cone, of Cr(VI)
(ug/g)
1330
1410
1500
91.2
81.5
83.1
408.9
414.4
400.2
7196
Average
(mean ±std)
1410 ±85
85.3 ± 5.2
407.8 ± 7.2
SIDMS
Cone, of Cr(VI)
(ug/g)
1373
1449
1512
93.9
82.1
90.4
419.8
426.1
408.0
Average
(mean ±std)
1445 ±70
88.8 ±6.1
41 8.0 ±9.2
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
1
2
3
4
Mass of Soil (g)
0
1.53
3.06
3.12
Spiked n*Cr(VI)
(ng/g)
2.997
3.033
2.993
1.587
Recovery (%)
Method 71 96
101 ±0.4
91 .8 ±1.7
81 .9 ±1.1
71 .6 ±2.5
SIDMS
100 ±1.3
100 ±0.3
101 ±0.3
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 5.8X105.
0.054 -i dead time (ns)
0.053 -
O
o °-052 '
o 0.051 -
S
S. 0.050 -
o
o
V)
~ 0.049-
0.048
Gain loss
OE+0 2E+5 4E+5 6E+5 SE-t-5
Counts per second for S2Cr
1E+6
FIGURE 2
IDMS DETERMINATION OF VANADIUM IN CRUDE OIL. NUMBERS SHOWN ABOVE THE BARS ARE
THE ATOMIC FRACTION
(Revised from Reference 1)
(D
CL
3
o
a>
c
o
E
CD
_eo
0)
CC
50 51
Isotope
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FIGURE 3
SEPARATION AND DETECTION OF CR(III) AND CR(VI) WITH ION-EXCHANGE CHROMATOGRAPHY
COUPLED WITH AN ICP-MS
(Reference 5)
100-
25.00
Cr(lll)
Cr(III): lOOppb
Cr(VI): lOOppb
Flow rate: 1.0 mL/min
Eluent: 0.06M NO3', pH = 3
Column: CETAC ANX 4605 Cr
50.00
time (s)
75.00 100.00
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 isotopic abundance.
(b): Chromatograms of the same solution spiked with isotope-enriched spikes ^Crflll) and KCr(VI)
(a)
100%
20 30 40 50 60 70 80 90 100 110 120
tifn»
-------
FIGURE 5
GRAPHIC CALCULATED ILLUSTRATION OF THE APPLICATION OF SIDMS TO THE SIMULTANEOUS
DETERMINATION OF CR(III) AND CR(VI)
Cr(III)
(a Isotop* distribution of th* unsplked sample
140 -
120 •
S 100 •
sŁ so-
jjeo.
I 40 •
' 20 .
i
ES560559
50 52 53
Isotope
• Cr(VI)
spite
HCr(VI)
natural
D Cr(lll)
spite
B Cr(lll)
natural
(c) Isotope distribution of the spiked sample
8 8 8 § fi
• Cr(VI)
spike
HCr(VI)
natural
D Cr(IH)
spite
BCr(IIO
natural
50
52
Isotop*
53
(•) 1
140 .
120-
1 100-
1| 80-
I 40 •
• 20 -
totop* distribution of th* spiked sample vrfth
Interconversion
1§H
1
50 52 53
Isotop*
20*
• Cr(VI)
spite
BCr(VI)
natural
DCr(l«)
spite
B Cr(lll)
natural
Cr(VI)
(b
140-
120-
1 100-
1| 80
I 40-
• 20-
) Isotop* distribution of th* unsplkcd sample
i
50 52 53
Isotope
DCr(IB)
spite
acr(ii)
natural
• Cr(VO
spite
BCr(VI)
natural
(d) Isotop* distribution of th* splk*d sample
140.
120
100.
60
40-
20-
0
0 Cr(ll)
spite
BCr(ll)
natural
• Cr(VI)
spite
BCr(V1)
natural
50
52
Itotop*
53
(f) Isotop* distribution of th* splk*d sunpl* «4th JOS
lnt*reonv*rslon
140.
120-
r 100..
!Ą B0'
jleo.
S 40-
' 20.
0- •
DCr(H)
spite
BCr(ll)
natural
• Cr(Vl)
spite
SCrfVI)
natural
so
52
Isotop*
53
(a) and (b) show the initial natural isotopic abundance of species Cr(lll) and Cr(VI) in a 50^l 200 ppb Cr solution
in which the concentrations of both Cr(lll) and Cr(VI) are 100 ppb. In (c) and (d), the sample is spiked with 100
ppb ^CrOII) On which ^r is enriched) and 100 ppb ^(Vl) On which 53Cr is enriched), there is no interconversion
between Cr(lll) and Cr(VI). In (e) and (f), 20% of Cr(lll) is converted to Cr(VI), and 20% of Cr(VI) is converted
to Cr(lll). Different degrees of interconversion results in different isotopic abundances, so the change of the
relative isotopic abundance can be applied to the determination of the species and the degree of the
interconversion.
6800-27
Revision 0
January 1998
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METHOD 6800
ELEMENTAL AND SPECIATED ISOTOPE DILUTION MASS SPECTROMETRY-
ISOTOPE DILUTION MASS SPECTROMETRY
11.2 Speciated Istotope
Dilution Mass
Spectrometry (SIDMS).
7.4 Prepare isotope
enriched standards.
11.1.2 Spike the isotope
enriched standards with
standard solution that
has natural isotopic
abundance.
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
s the instrument
ready for
measuring isotopic
ratios?
10.1.1 Set-up and tune
instrument according to
Method 6020.
10.1.2 Determine the
dead time of the detector.
11.1.5 Measure the isotope
ratios of the samples. Measure
mass bias factors with an
interval of several hours.
11.1.5
Are the isotopic
ratios outside 0.1 to 10
for samples containing
significant amounts
of analyte?
11.1.5.2
Is the
response so high
that the gain
depression might
occur?
12.1.1 Integrate the
counts, perform dead
time correction and mass
bias correction.
12.1.1 Calculate the
isotope ratios.
12.1.2 Calculate the
concentrations using
the measured isotope
ratios.
12.1.3 If required,
calculate results for
solids on a dry-
weight basis.
6800-28
Revision 0
January 1998
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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 CASRNa
Aluminum
Antimony
Barium
Beryllium
Cadmium
Calcium
Chromium
Cobalt
Copper
Iron
Lead
Lithium
Magnesium
Manganese
Molybdenum
Nickel
Osmium
Potassium
Silver
Sodium
Strontium
Thallium
Tin
Vanadium
Zinc
(Al)
(Sb)
(Ba)
(Be)
(Cd)
(Ca)
(Cr)
(Co)
(Cu)
(Fe)
(Pb)
(Li)
(Mg)
(Mn)
(Mo)
(Ni)
(Os)
(K)
(Ag)
(Na)
(Sr)
(Tl)
(Sn)
(V)
(Zn)
7429-90-5
7440-36-0
7440-39-3
7440-41-7
7440-43-9
7440-70-2
7440-47-3
7440-48-4
7440-50-8
7439-89-6
7439-92-1
7439-93-2
7439-95-4
7439-96-5
7439-98-7
7440-02-0
7440-04-2
7440-09-7
7440-22-4
7440-23-5
7440-24-6
7440-28-0
7440-31-5
7440-62-2
7440-66-6
a 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 direct aspiration technique. For clean
aqueous samples, the detection limits shown in the table by direct aspiration may be extended
7000B -1
Revision 2
January 1998
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-------
METHOD 7010
GRAPHITE FURNACE ATOMIC ABSORPTION SPECTROPHOTOMETRY
1.0 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
Arsenic
Barium
Beryllium
Cadmium
Chromium
Cobalt
Copper
Iron
Lead
Manganese
Molybdenum
Nickel
Selenium
Silver
Thallium
Vanadium
Zinc
(Sb)
(As)
(Ba)
(Be)
(Cd)
(Cr)
(Co)
(Cu)
(Fe)
(Pb)
(Mn)
(Mo)
(Ni)
(Se)
(Ag)
(Tl)
(V)
(Zn)
7440-36-0
7440-38-2
7440-39-3
7440-41-7
7440-43-9
7440-47-3
7440-48-4
7440-50-8
7439-89-6
7439-92-1
7439-96-5
7439-98-7
7440-02-0
7782-49-2
7440-22-4
7440-28-0
7440-62-2
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
7010-1 Revision 0
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(196.0 nm). 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:HNO3 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. Electrodeless 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 uL as required.
Pipet tips should be checked as a possible source of contamination when contamination is
7010-6 Revision 0
January 1998
-------
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 - All 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 acid 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 , HNO3: 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 acid (1:1), HCI: 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.
7.5 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 acids. (See individual methods for specific instructions.) Sulfuric or phosphoric
acids should be avoided as they produce an adverse effect on many elements. The stock solutions
are prepared at concentrations of 1.000 mg of the metal per 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. Examples of appropriate standard preparations can be found in Sections 7.6.1
through 7.6.18.
7010-7 Re vision 0
January 1998
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7.6.1 Antimony: Carefully weigh 2.743 g of antimony potassium tartrate,
K(SbO)C4H4O6»1/2H2O, and dissolve in reagent water. Dilute to 1 L with reagent water;
7.6.2 Arsenic: Dissolve 1.320 g of arsenic trioxide, As2O3, or equivalent in 100 mL
of reagent water containing 4 g NaOH. Acidify the solution with 20 mL cone. HNO3 and dilute
to 1 L with reagent water.
7.6.3 Barium: Dissolve 1.779 g barium chloride, BaCI2»2H2O, in reagent water and
dilute to 1 L with reagent water.
7.6.4 Beryllium: Dissolve 11.659 g beryllium sulfate, BeSO4, in reagent water
containing 2 mL nitric acid (cone.) and dilute to 1 L with reagent water.
7.6.5 Cadmium: Dissolve 1.000 g cadmium metal in 20 mL of 1:1 HNO3 and dilute
to 1 L with reagent water.
7.6.6 Chromium: Dissolve 1.923 g of chromium trioxide, CrO3l in reagent water,
acidify with redistilled HNO3, 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 HNO3 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.
7.6.8 Copper: Dissolve 1.000 g of electrolytic copper in 5 mL of redistilled HNO3
and dilute to 1 L with reagent water.
7.6.9 iron: Dissolve 1.000 g iron wire in 10 mL redistilled HNO3 and reagent water
and dilute to 1 L with reagent water. Note that iron passivates in cone. HNO3, and therefore
some water should be present.
7.6.10 Lead: Dissolve 1.599 g of lead nitrate, PbfNO^, in reagent water, acidify with
10 mL redistilled HNO3, and dilute to 1 L with reagent water.
7.6.11 Manganese: Dissolve 1.000 g manganese metal in 10 mL redistilled HNO3
and dilute to 1 L with reagent water.
7.6.12 Molybdenum: Dissolve 1.840 g of ammonium molybdate, (NH4)6Mo7O24.4H2O,
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(NO3)2»6H2O
in 10 mL HNO3 and dilute to 1 L with reagent water.
7.6.14 Selenium: Dissolve 0.345 g of selenious acid (actual assay 94.6% H2SeO3)
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, AgNO3, in reagent water.
Add 10 mL of HNO3 (cone.) and dilute to 1 L with reagent water. Because this standard is light
sensitive, store in a amber glass bottle in a refrigerator.
7010-8 Revision 0
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7.6.16 Thallium: Dissolve 1.303 g thallium nitrate, TINO3, in reagent water, acidify
with 10 mL cone. HNO3, and dilute to 1 L with reagent water.
7.6.17 Vanadium: Dissolve 1.785 g of vanadium pentoxide, V2O5, in 10 mL of cone.
HNO3 and dilute to 1 L with reagent water.
7.6.18 Zjnc: Dissolve 1.000 g zinc metal in 10 ml of cone. HNO3 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
analyte. 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 HNO3 (1 mL of HNO3, adding 0.1 mL of cone. HCI, if necessary). Dissolve 200
mg of Mg(NO3)2 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 Ni(NO3)2»6H2O 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)2HPO4, in reagent water and dilute to 100 mL.
7.7.5 Palladium chloride: Weigh 0.25 g of PdCI2 to the nearest 0.0001 g and
dissolve in 10 mL of 1:1 HNO3. 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 Wank 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 contaminated samples after the last acceptable method blank must be
7010-9 Revision 0
January 1998
-------
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 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-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, 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 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.
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
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 <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 for the undiluted sample and five
7010-10 Revision 0
January 1998
-------
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 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-analyte signals. The unknown sample
concentration Cx is calculated:
- SBVSCS
cx=T^"
where SA and SB are the analytical signals (corrected for the blank) of solutions A and B,
respectively. V, and C8 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.
7010-11 Revision 0
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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 quality 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 calibration 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
7010-12 Revision 0
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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 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 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 acid
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:
M9/L metal in sample = A
where:
A = ug/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 x V
mg metal/kg sample =
W
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 Revision 0
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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.
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TABLE 1
FURNACE ATOMIC ABSORPTION DETECTION LIMITS FOR ANALYTES
IN REAGENT WATER
Furnace Procedure"'"
Detection Limit
Metal (ug/L)
Antimony 3
Arsenic 1
Barium(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-uL 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
ELEMENT
Sb
As
Ba
Be
Cd
Cr
Co
Cu
Fe
Pb
Mn
Mo
Ni
Se
Ag
Tl
V
Zn
WAVELENGTH (nm)
ZILfi,
231.1
193.7
553.6
234.9
228.8
357.9
240.7
324.7
248,3,
248.8,271.8,
302.1,252.7
283,3,
217.0
Z®Ł,
403.1
313.3
232.0.
352.4
196.0
328.1
276.8
318.4
213.9
PURGE GAS1
argon or nitrogen
argon
argon
argon
argon
argon
argon
argon or nitrogen
argon or nitrogen
argon
argon or nitrogen
argon
argon or nitrogen
argon
argon
argon or nitrogen
argon
argon or nitrogen
COMMENTS
nitrogen should not
be used
nitrogen should not
be used
nitrogen should not
be used
nitrogen should not
be used
Note: If more than one wavelength is listed, the primary line is underlined.
1The argon/H2 purge gas is also applicable.
Source: Reference 1
7010-17
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FIGURE 1
STANDARD ADDITION PLOT
Zero
Absorbance
Cone, of
Sample
AddnO
No Addn
AddrM
Addn of 50%
of Expected
Amount
Addn 2 Addn 3
Addn of 100% Addn of 150%
of Expected of Expected
Amount Amount
7010-18
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METHOD 7471B
MERCURY IN SOLID OR SEMISOLID WASTE (MANUAL COLD-VAPOR TECHNIQUE)
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 Revision 2
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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 cell) occurs at -20 seconds.
7473-10
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Deleclor
nlerference
] filler
vessel
jt
S1
Cuvette h
Spectromete
Long
measuring
cuvette
Short
measuring
cuvette
Amalgamator
Oxygen
flow
regulato
Sample
boat
release
J^x
"^^
1 "
y
"^ ^
—
^v.
^
'
^
Citalyit
furnace
mpot.it ion Dosing
"^ >e equipment
Catalyst furnace decomposition furna
Mercury release furnace
FIGURE 2
DIAGRAM OF THE MERCURY ANALYSIS SYSTEM
7473-11
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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 cell.
Primary Calibration Curve
>w Range
01
u
c
ra
Ł>
O
ft
0.8
0.6
0.4
0.2 -
Low Range
1 ' ' i ' • • ' i • ' • • r*
i • • ' • i • • •
y = 0.023725 + 0.023015x R= 0.99782
0 5 10 15 20 25 30 35 40
ngHg
Primary Calibration Curve
High Range
1 ' ' i ' • • • i • ' • • i • • • • 1 • • ' • i • ' '
0.8
I °'6
m
X)
1 0.4
0.2
0
• y = 0.046033 + 0.0014433x R= 0.9Łc53j
100 200 300 400 500 600
ngHg
7473 - 12
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FIGURE 4
PRIMARY CALIBRATION CURVE USING THE DMA 80 IN FIELD ANALYSIS CONDITIONS
•fi
o
CO
0.8
0.6
0.4
0.2
0
y = 0.023495 + 0.020552x R= 0.99766 J
0 5 10 15 20 25 30 35 40
ngHg
7473-13
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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)
u
C
«
-a
o
CO
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
—I I I—I I I I I I—I—I—I—I—I—I—I—I—p
y = 0.05268 + 0.0014197x R= 0.99765
y = 0.072404 + 0.001448x R= 0.9^698
Aqueous Standards
Solid SRM Standards t
100 200
300 400
ngHg
500
600
7473-14
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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 acids 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 dryer 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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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 MuKiphasic 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, the 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 all 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 Revision 0
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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(43.3811.29%).
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 Revision 0
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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.
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TABLE 1
DETERMINATION OF WATER IN USED OIL
(w/w %)
Expected
0
0.1
0.2
0.5
1.0
2.0
5.0
10.0
20.0
25.0
50.0
Method 9001
0.161
0.149
0.226
0.459
0.948
2.36
5.03
9.82
20.2
26.37
50.05
Method 9000
0.061
0.145
0.255
0.561
1.07
2.46
5.05
9.97
20.0
26.05
50.60
Source: Referenced
TABLE 2
ANALYSIS OF USED OIL CERTIFIED REFERENCE MATERIALS3
CRM
ERM-34
ERM-35
ERM-36
ERM-41
Certified Value, wt %
1.95
5.86
10.3
87.4
Method 9001, wt%
1.92±0.02
5.91±0.61
10.30±0.85
88.4±6.7
Method 9000, wt %
1.86±0.09
6.13±0.55
10.310.81
86.4±6.6
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
0
10
20
30
40
Method 9001
1.14
10.06
18.99
28.52
38.47
Method 9000
0.579
9.74
19.67
29.95
40.34
Source: Reference 2.
TABLE 4
DETERMINATION OF ALCOHOL IN WATER/ALCOHOL MIXTURES
Expected % Alcohol, v/v
0
10
25
40
50
80
100
Vodka, 40
Whiskey, 40
Gin, 47
Method 9001 (% v/v)
0
10.0
25.6
40.9
48.5
80.6
99.9
41.9
40.0
47.2
Method 9000 (% v/v)
0
10.3
25.0
38.7
49.1
79.8
100.0
42.0
41.9
48.7
Source: Reference 2
9001-7
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METHOD 9001
DETERMINATION OF WATER IN WASTE MATERIALS BY QUANTITATIVE
CALCIUM HYDRIDE REACTION
11.1 Follow directions
provided by kit
manufacturer.
11.3 Extract 1g sample
with dilution solvent. React
O.SmL aliquot of extract
with the calcium hydride
reagent.
11.2 React measured
0.4 - 0.8 mL or 1g sample
with the calcium hydride
reagent.
11.3 Record results
in w/w percent water.
11.3 Record results in
v/v or w/w percent water.
9001-8
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CHAPTER FOUR
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 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 all 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.
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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 that 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 Oraanics (including Pesticides, PCBs and Herbicides.)
Containers used to collect samples for the determination of semivolatile organic compounds
should be soap and water washed followed 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 may NOT
be used for the storage of samples due to the possibility of sample contamination from 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.
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 acid 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 acid solution. They are alkaline, equivalent to
ca. 0.1 N NaOH upon dilution, and are claimed to remove dried blood, silicone greases,
distillation residues, insoluble organic residues, etc. They are further claimed to remove
radioactive traces and will not attack glass or exert a corrosive effect on skin or clothing.
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, and 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 5021: 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
H2SO4, HCI, or solid NaHSO4.
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 VGA vial.
Cool to 4°C and adjust pH to less than 2 with
H2SO4, HCI, or solid NaHSO4.
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. Coolto4°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 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.
4.2.1 EXTRACTIONS AND PREPARATIONS
The following methods are included in this section:
Method
Method
Method
Method
Method
Method
Method
3500B:
351OC:
3520C:
3535A:
3540C:
3541:
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 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
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 analyte 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 not 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 divinylbenzene (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-placel
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, clamp, 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
(Supelco) 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 C18 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 divinylbenzene (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-um 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
4.8 Solvent Vapor Recovery System - Kontes 545000-1 006 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 Kuderna-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. 1 1 Nitrogen evaporation apparatus (optional) - N-Evap, 1 2- 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 cans
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 inches)
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 it 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 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), Na2SO4 - 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), H2SO4 - Slowly add 50 mL of concentrated
H2SO4 (sp. gr. 1.84) to 50 ml of organic-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, (CH3)2CO.
5.5.7 Methyl-terf-butyl ether (MTBE), C5H12O.
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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 very 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 all target analytes. The procedures
for disk washing, disk conditioning, sample extraction, and sample elution 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 the 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 not 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 blank 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 the pH of 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 cis
Organochlorine pesticides cie
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 prefilter 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 not 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 1st solvent wash volume
Phthalate esters 20 ml methylene chloride
Organochlorine pesticides 20 ml_ methylene chloride
Polychlorinated biphenyls (PCBs) 20 mL methylene chloride
Organophosphorus pesticides 5 mL acetone
Nitroaromatics and nitramines 5 mL acetonitrile
TCLP leachates containing organochlorine pesticides 5 mL acetone
TCLP leachates containing semivolatiles 5 mL acetone
TCLP leachates containing phenoxyacid herbicides 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 all 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 2nd solvent wash volume
Phthalate esters 10mL acetone
Organochlorine pesticides 10 mL acetone
Polychlorinated biphenyls (PCBs) not required
Organophosphorus pesticides 5 mL methanol
Nitroaromatics and nitramines 15 mL acetonitrile
TCLP leachates containing organochlorine pesticides 5 mL ethyl acetate
TCLP leachates containing semivolatiles 5 mL ethyl acetate
TCLP leachates containing phenoxyacid herbicides not required
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7.5 Disk conditioning
The extraction disks are composed of hydrophobia 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.
Analvte Group Conditioning steps
Phthalate esters 20 ml_ methanol, soak 1 min,
20 mL reagent water
Organochlorine pesticides 20 mL methanol, soak 1 min,
20 mL reagent water
Polychlorinated biphenyls (PCBs) 20 mL methanol, soak 1 min,
20 mL reagent water
Organophosphorus pesticides 5 mL methanol, soak 1 min,
20 mL reagent water
Nitroaromatics and nitramines 15 mL acetonitrile, soak 3 min
30 mL reagent water
TCLP leachates containing organochlorine pesticides 5 mL methanol soak 1 min,
15 mL reagent water
TCLP leachates containing semivolatiles 5 mL methanol soak 1 min,
15 mL reagent water
TCLP leachates containing phenoxyacid herbicides 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 just 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 just 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 and 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 not allow the
cartridge to go dry.
7.6.2 When only a thin layer of solvent remains above the sorbent 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 flexible 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 mL/min, 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 blanks 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, swirl 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.
recommended elution solvent for each group of analytes is listed below.
The
Analvte Group
Phthalate esters
Organochlorine pesticides
Polychlorinated 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 twice 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 water 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 reaches
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 clean 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. Place 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 semivolatile
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 Any 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. al., "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., 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. USAGE 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
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)
Extraction pH
5-7
5-9
5-9
as received
as received
as produced by TCLP
as produced by TCLP
1.0
Disk Medium3
C18
C18
C«
SDB-RPS
SDB-RPS
SDB-XC
SDB-XC
SDB-XC
Elution Solvent
acetonitrile
methylene' chloride
methylene chloride
MTBE
acetonitrile
ethyl acetate
ethyl acetate
acetonitrile
Final Extract
Volume for
Exchange Solvent Analysis (mL)b
hexane
hexane
hexane
hexane
acetonitrile
hexane
methylene chloride
hexane
10.0
10.0
10.0
10.0
10.0
10.0
1.0
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
-Clamp
.TM
Empore
Extraction Disk
Base
(Fritted or with Screen)
Drip Tube
Filter Flask or Manifold
FIGURE 1
DISK EXTRACTION APPARATUS
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METHOD 3535A
SOLID-PHASE EXTRACTION (SPE)
7.6 Prepare cartridge and
load sample on cartridge.
7.9 Elute target compounds
from cartridge.
7.1 Prepare the sample.
7.2 Adjust the pH.
7.3 Select appropriate disk
(or cartridge) and set-up
apparatus.
7.8 Elute analytes from the disk
7.10 Perform KD concentration
7.12 Go to appropriate cleanup
method or sample analysis
technique.
Stop
—- .. -—
3535A-18
7.1 1 Perform Micro-Snyder or
N2 evaporation.
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METHOD 3545A
PRESSURIZED FLUID EXTRACTION (PFE)
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 ug/kg of
semivolatile organic compounds, 250 to 2500 ug/kg of organophosphorus pesticides, 5 to 250 ug/kg
of organochlorine pesticides, 50 to 5000 ug/kg of chlorinated herbicides, 1 to 1400 ug/kg of PCBs,
and 1 to 2500 ng/kg of 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 pelletized 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. In such
cases, proceed with Method 3620 and/or Method 3660.
3.3 Samples for PCDD/PCDF analysis should be subjected to the various cleanup
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).
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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 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 Drying agents
5.3.1 Sodium sulfate (granular anhydrous), Na2SO4.
5.3.2 Pallatized 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, clean 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 (H3PO4) in organic-free reagent water.
5.4.2 Trifluoroacetic acid solution (see Sec. 5.5.5). Prepare a 1% (v/v) solution of
trifluoroacetic acid 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), CH3COCH3/CH2CI2.
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), CH3COCH3/C6H14.
5.5.3 PCBs may be extracted with acetone/hexane (1:1, v/v), CH3COCH3/C6H14 or
acetone/methylene chloride (1:1, v/v), CH3COCH3/CH2CI2 or hexane, C6H14.
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5.5.4 Organophosphorus pesticides may be extracted with methylene chloride,
CH2CI2 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), CH3COCH3/CH2CI2/H3PO4l or an
acetone/methylene chloride/trifluoroacetic acid solution (250:125:1, v/v/v),
CHaCOCHa/C^CIj/CFgCOOH. (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,
nonpolar organics, e.g., 4,4'-DDT, PCBs, etc. Air-drying may not be appropriate for
the analysis of the more volatile organochlorine 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 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 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:
o/o dry weight - 9 <* dry sample x 1(JO
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: 1500 - 2000 psi
Static time: 5 min (after 5 min pre-heat equilibration)
Flush volume: 60% of the cell volume
Nitrogen purge: 60 sec at 150 psi (purge time may be extended for larger cells)
Static Cycles: 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, cleanup, 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 ug/kg for the chlorinated pesticides and from 250 to 12500 ug/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 low 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 ug/kg for the OPPs and from 50 to 5000 ug/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 Dionex 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 ug/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
spike duplicates were included. The data are reported in Reference 2, and represent seven replicate
extractions and 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 ug/kg levels.
Samples were extracted by the Dionex Accelerated Solvent Extractor and by traditional Soxhlet
techniques. Extracts were analyzed by a high resolution mass spectrometric 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 TCUPPL (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)."
Chemosphers, 34(5-7), pp. 975-987, 1997.
3545A - 8 Revision 1
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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
instrument. 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.
3545A - 9 Revision 1
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METHOD 3545A
PRESSURIZED FLUID EXTRACTION (PFE)
^
r
7.1 Prepare
sample.
>
r
7.2 Determine
sample % dry
weight.
>
r
7.3 Grind sufficient
weight of the dried
sample.
>
r
7.4 Transfer ground
sample to an
extraction cell.
^
r
7.5 Add surrogates
and matrix spiking
standards.
w
^
7.6 Place extraction
cells into auto
sampling train.
>
f
7.7 Load
collection tray.
^
t
7.8 Optimize
conditions of
extractor.
>
f
7.9 Begin
extraction.
>
r
7.10 Collect
extracts and
allow to cool.
>
r
7.11 Perform
cleanup or
determinative
method.
3545A-10
Revision 1
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METHOD 3562
SUPERCRITICAL FLUID EXTRACTION OF POLYCHLORINATED BIPHENYLS (PCBs)
AND ORGANOCHLORINE PESTICIDES
1.0 SCOPE AND APPLICATION
1.1 Method 3562 describes the extraction with supercritical fluids of polychlorinated
biphenyls (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
2,2',5,5'-Tetrachlorobiphenyl
2,2',4,5,5l-Pentachlorobiphenyl
2,3,3',4,4I-Pentachlorobiphenyl
2,3',4,4',5-Pentachlorobiphenyl
2,2I,3,3',4,4l-Hexachlorobiphenyl
2,2',3,4,41,51-Hexachlorobiphenyl
2,2',3,4',5',6-Hexachlorobiphenyl
2,2',4,4l,5,5'-Hexachlorobiphenyl
2,3I3',4,4'I5'-Hexachlorobiphenyl
2,2',3,3',4,4',5-Heptachlorobiphenyl
2)2'13,4,4',5,51-Heptachlorobiphenyl
Aldrin
p-Hexachlorocyclohexane (P-BHC)
6-Hexachlorocydohexane (6-BHC)
Y-Hexachlorocyclohexane (v-BHC, or Lindane)
a-Chlordane
4,4'-DDD
4,4'-DDE
4,4'-DDT
Dieldrin
Endosulfan II
Endrin
Endrin aldehyde
Heptachlor
Heptachlor epoxide
3562 - 1
7012-37-5
35693-99-3
37680-73-2
32598-14-4
31508-00-6
38380-07-3
35065-28-2
38380-04-0
35065-27-1
38380-08-4
35065-30-6
35065-29-3
309-00-2
319-85-7
319-86-8
58-89-9
5103-71-9
72-54-8
72-55-9
50-29-3
60-57-1
33213-65-9
72-20-8
7421-93-4
76-44-8
1024-57-3
28
52
101
105
118
128
138
149
153
156
170
180
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1.2 Method 3562 is not 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 solid-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 CO2 "snow". A 1 - 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 nonpolar.
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 CO2 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 dean 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 CO2 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 CO2 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
all 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 2000 psi or more.
5.4 When liquid CO2 comes in contact with skin, it can cause "bums" because of its low
temperature (-70 °C). Bums 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 CO2 is used for cryogenic cooling, typical coolant consumption is 5 L/min, which
results in a CO2 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 urn 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 urn particle diameter (commonly used in SPE
cartridges), as a solid trap for the PCBs.
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6.1.3.2 For organochlorine pesticides, octadecyl 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,
or equivalent, may be used to monitor the fluid usage. Such a device is useful because CO2 tanks
used for SFE 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 urn 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 CO2 is absolutely necessary for use in SFE. Aluminum cylinders are
preferred to steel cylinders. The cylinders must be fitted with eductor tubes.
7.2 CO2 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 CO2 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 - The 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-ambient 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
the choice of internal standards, where applicable. However, note that for PCBs, 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 hexyl ether, 2,4-
dichlorobenzyl heptyl ether, 1,2,3,4-tetrachloronaphthalene, hexabromobenzene, and
octachloronaphthalene.
7.4.2 Internal standard for organochlorine pesticides - Pentachloronitrobenzene
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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octachloronaphthalene. Prepare a stock solution of 10 mg/mL. Apply 150-|jL aliquots to the soil
samples within the extraction vessels at the exit end of the flow-through vessels. It has been
observed that a very small volume (10 uL) 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 Celite 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 and
rinsing the trapping material prior to reuse. All solvents should be pesticide-grade or equivalent.
7.9.1 n-Heptane,
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. 1 1) 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 105°C. Allow to cool in a desiccator before weighing.
11.2.2 Calculate the % dry weight as follows:
% dry weight = 9 °* dry sample x
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 CO2 solid "snow"
prepared from the extraction grade CO2. 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 CO2 to
sublime away. As soon as the sample appears free-flowing and solid CO2 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-deaned 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 the sample and CO2 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 CO2. 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
follows:
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 material. 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 uL 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-cleaned Pyrex glass wool, Celite, etc.
11.6 Sample extraction conditions
11.6.1 Recommended conditions for PCBs
11.6.1.1 Extraction conditions
Pressure: 4417 psi (305 bar)
Extraction chamber temperature: 80°C
Density: 0.75 g/mL
Extraction fluid composition: CO2
Static equilibration time: 10 minutes
Dynamic extraction time: 40 minutes
Extraction fluid flow rate: 2.5 mUminute
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 CO2 at a reference temperature
of 4.0°C and a density 0.92 g/mL, or 92 g of CO2).
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11.6.1.2 Collection conditions (during extraction)
Trap packing: Florisil
Trap temperature: 15-20°C
Nozzle temperature: 45-55 °C (variable restrictor)
11.6.1.3 Reconstitution conditions for collected 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: 4330 psi (299 bar)
Extraction chamber temperature: 50°C
Density: 0.87 g/mL
Extraction fluid composition: CO2
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Static equilibration time: 20 minutes
Dynamic extraction time: 30 minutes
Extraction fluid flow rate: 1.0 mL/minute
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 CO2 at a reference temperature of
4.0°C and a density 0.92 g/mL, or 28 g of CO2).
11.6.2.2 Collection conditions (during extraction)
Trap packing: ODS
Trap temperature: 20°C
Nozzle temperature: 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: n-Hexane
Collected fraction volume: 1.3 ml_
Trap temperature: 50 °C
Nozzle temperature: 30 °C (variable restrictor)
Rinse solvent flow rate: 2 mUminute
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 copper 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 Celite 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
organochlorine 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% clay, 21%
sand, 5.4% organic matter, 46% silt, and 2.2% moisture. Seven replicate 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 (Ftorisil). 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 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.
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," 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.
11.1 Use appropriate
sample handling
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
copper powder
to sample
11.5 Transfer weighed sample to
extraction vessel and add surrogates
i
0
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METHOD 3562
(Continued)
11.6.1 Collection
of PCBs
O
>
1
11.6 Sample
Extraction
11.6.2 Collection
of organochloride
pesticides
11.7 Label extract
11.8 Additional sulfur cleanup
of extract if necessary
11.9 Perform SFE
system maintenance
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METHOD 8081B
ORGANOCHLORINE PESTICIDES BY GAS CHROMATOGRAPHY
1.0 SCOPE AND APPLICATION
1.1 Method 8081 may be used to determine the concentrations of various organochlorine
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
a-BHC
P-BHC
Y-BHC (Lindane)
6-BHC
Chlorobenzilate
a-Chlordane
y-Chlordane
Chlordane - not otherwise specified
DBCP
4,4'-DDD
4,4'-DDE
4,4'-DDT
Diallate
Dieldrin
Endosulfan I
Endosulfan II
Endosulfan sulfate
Endrin
Endrin aldehyde
Endrin ketone
Heptachlor
Heptachlor epoxide
Hexachlorobenzene
Hexachlorocyclopentadiene
Isodrin
Methoxychlor
Toxaphene
309-00-2
319-84-6
319-85-7
58-89-9
319-86-8
510-15-6
5103-71-9
5103-74-2
57-74-9
96-12-8
72-54-8
72-55-9
50-29-3
2303-16-4
60-57-1
959-98-8
33213-65-9
1031-07-8
72-20-8
7421-93-4
53494-70-5
76-44-8
1024-57-3
118-74-1
77-47-4
465-73-6
72-43-5
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 analyte, 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:
Alachlor
Captafol
Chloroneb
Chloropropylate
Chlorothalonil
DCPA
Dichlone
Dicofol
Etridiazole
Halowax-1000
Halowax-1001
Halowax-1013
Halowax-1014
Halowax-1051
Halowax-1099
Mirex
Nitrofen
PCNB
Permethrin (c/s + trans)
Perthane
Propachlor
Strobane
foans-Nonachlor
Trifluralin
15972-60-8
2425-06-1
2675-77-6
5836-10-2
1897-45-6
1861-32-1
117-80-6
115-32-2
2593-15-9
58718-66-4
58718-67-5
12616-35-2
12616-36-3
2234-13-1
39450-05-0
2385-85-5
1836-75-5
82-68-8
52645-53-1
72-56-0
1918-16-7
8001-50-1
39765-80-5
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 appropnate
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 Soxhlet), 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-uL sample into a gas
chromatograph with a narrow- or wide-bore fused-silica 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 interfere with the
detection of early-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/lsodrin
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 Il/Chloropropylate/Chlorobenzilate
4,4'-DDT/Endosulfan sulfate
Methoxychlor/Dicofol
DB-1701 Chlorothalonil/p-BHC
6-BHC/DCPA/Permethrin
a-Chlordane/frans-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 splrt-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 in 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 urn 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 urn coating thickness, 1 um 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 um or 0.83 um 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 um
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 polysiloxane (DB-5, SPB-5, RTx-
5, or equivalent), 1.5 um 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 urn 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 urn 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 um 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 um 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-trimethylpentane) 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 weight 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 (3-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/uL) of pentachloronitrobenzene or 1-
bromo-2-nitrobenzene. Spike 10 uL 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/uL) of 1-bromo-2-nitrobenzene.
Spike 10 uL 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 surrogates 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-elution 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/uL. Use a spiking
volume of 100 uL 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 phthalate 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 (<;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.
7.3.1.2 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 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 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 chromatogram 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 the 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 analytes employs a single-point calibration. A single calibration
standard near the mid-point of the expected calibration range of each multi-component
analyte is included with the initial calibration of the single component analytes 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 analytes
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 analytes 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-uL 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 analytes, 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:
CF = Peak 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:
where n is the number of standards analyzed.
7.4.5.3 Calculate the standard deviation (SD) and the RSD of the
calibration factors for each analyte as:
SD
E(CFrCF)2
n-1
RSD = -^ 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 Verify 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 analytes
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.
CF - CF
% 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
all analytes. If the average of the responses for aN 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-uL aliquot of the concentrated sample extract. Record the volume
injected to the nearest 0.05 uL and the resulting peak size in area units.
755 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 (ug/L) =
where:
A,, = Area (or height) of the peak for the analyte in the sample.
V, = Total volume of the concentrated extract (uL).
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.
CF = Mean calibration factor from the initial calibration (area/ng).
Vi = Volume of the extract injected (uL). 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 .
V8 = 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 yg/L.
7.5.6.2 For non-aqueous samples
Concentration (ug/kg) =
where A*, Vt, D, CF, and V, are the same as for aqueous samples, and
W8 = 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 ug/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 coeluting 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 analyte) 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
analyte that is above the acceptance limit, i.e., >15%, and the analyte 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 analyte
would have been detected were it present. In contrast, if an analyte above the QC limits was
detected in a sample extract, then re-injection is necessary to ensure accurate quantitation.
If an analyte 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 analyte 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.
759 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 8000.
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 the retention time windows. If any of these subsequent
standards fall outside their absolute retention time windows, the GC 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
m 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
pmenes. 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 that the major Toxaphene 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 study 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 disproportionally larger or smaller in the sample compared to the
standard.
7.6.1.3.5 The heights or areas of the 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 Chlordane 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 Registry number for Technical Chlordane is properly 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 matenal 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
otherwise 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 subtract their areas from the 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.
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.
Construct the baseline of Technical Chlordane in the standard chromatogram between
the 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 57-74-9.
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 hexachloride. 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, P, y, and 5) 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 ODD
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/uL in the final extract for each single-component compound. Ion trap or selected ion
monitoring will normally require a concentration of approximately 1 ng/uL.
7.7.2 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.
7.7.3 GC/MS is not recommended for confirmation when concentrations are below
1 ng/uL 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 analytes. Endrin and DDT breakdown to endrin aldehyde,
endrin ketone, ODD, 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 It 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 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
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
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. 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'-DDE, 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 = sum of degradation peak areas (ODD * PPE),1(X)
sum of all peak areas (DDT + DDE + ODD)
% breakdown of endrin - sum of degradation peak areas (aldehyde + ketone) K1(?0
sum of all peak areas (endrin + aldehyde + ketone)
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8.4.6.2 The breakdown of DDT 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 Florisil (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 Florisil 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 method 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 ug/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 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% clay, 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., Benedict©, 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 - Pesticides 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 Pesticide Analysis", Bull. Environ. Contam. Toxicol., 1971, 6, 9.
4. Jensen, S., Renberg, L, Reutergardth, 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,
Cincinnati, OH, 45268.
6. Pionke, H.B., Chesters, G., Armstrong, D.E., "Extraction of Chlorinated Hydrocarbon
Insecticides from Soil", Agron. J., 1968, 60, 289.
7. Burke, J.A., Mills, P.A., Bostwick, D.C., "Experiments with Evaporation of Solutions of
Chlorinated Pesticides", J. Assoc. Off. Anal. Chem., 1966, 49, 999.
8. Glazer, J.A., et al., "Trace Analyses for Wastewaters", Environ. Sci. and Techno!., 1981, 15,
1426.
9. 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.
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. Markell, 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 Organochlorine Pesticide
Residues from Soils," Proceedings of the Tenth Annual Waste Testing and Quality Assurance
Symposium, Arlington, VA, July, 1994.
13. Markell, 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
Compound
Aldrin
a-BHC
P-BHC
6-BHC
Y-BHC (Lindane)
a-Chlordane
y-Chlordane
4,4'-DDD
4,4'-DDE
4,4'-DDT
Dieldrin
Endosulfan I
Endosulfan II
Endosulfan sulfate
Endrin
Endrin aldehyde
Heptachlor
Heptachlor epoxide
Methoxychlor
Toxaphene
Retention
DB-6088
11.84
8.14
9.86
11.20
9.52
15.24
14.63
18.43
16.34
19.48
16.41
15.25
18.45
20.21
17.80
19.72
10.66
13.97
22.80
MR
Time (min)
DB-17018
12.50
9.46
13.58
14.39
10.84
16.48
16.20
19.56
16.76
20.10
17.32
15.96
19.72
22.36
18.06
21.18
11.56
15.03
22.34
MR
MR = Multiple response compound.
a See Table 4 for GC
operating conditions.
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TABLE 2
GAS CHROMATOGRAPHIC RETENTION TIMES FOR THE ORGANOCHLORINE PESTICIDES
USING NARROW-BORE CAPILLARY COLUMNS, SINGLE-COLUMN METHOD OF ANALYSIS
Compound
Aldrin
a-BHC
P-BHC
5-BHC
Y-BHC (Lindane)
a-Chlordane
y-Chlordane
4,4'-DDD
4,4'-DDE
4,4'-DDT
Dieldrin
Endosulfan 1
Endosulfan II
Endosulfan sulfate
Endrin
Endrin aldehyde
Heptachlor
Heptachlor epoxide
Methoxychlor
Toxaphene
NA = Data not available
MR = Multiple response
Retention
DB-6088
14.51
11.43
12.59
13.69
12.46
NA
17.34
21.67
19.09
23.13
19.67
18.27
22.17
24.45
21.37
23.78
13.41
16.62
28.65
MR
compound.
Time (min)
DB-5a
14.70
10.94
11.51
12.20
11.71
NA
17.02
20.11
18.30
21.84
18.74
17.62
20.11
21.84
19.73
20.85
13.59
16.05
24.43
MR
a See Table 4 for GC operating conditions.
8081 B- 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. 5.0 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
-------
TABLE 4
GC OPERATING CONDITIONS FOR ORGANOCHLORINE 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 urn film thickness.
Carrier gas
Carrier gas pressure
Injector temperature
Detector temperature
Initial temperature
Temperature program
Final temperature
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
270°C
Column 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), 25 urn coating
thickness, 1 urn film thickness.
Carrier gas
Carrier gas pressure
Injector temperature
Detector temperature
Initial temperature
Temperature program
Final temperature
Nitrogen
20 psi
225°C
300°C
160°C, hold 2 minutes
160°C to 290°C at 5°C/min
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-silica capillary column chemically bonded with 35
percent phenyl methylpolysiloxane (DB-608, SPB-608, RTx-35, or equivalent), 0.5 urn or
0.83 urn film 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 urn 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
1500Cto270°Cat5°C/min
270°C, hold 10min
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 urn film thickness.
Carrier gas
Carrier gas flow rate
Makeup gas
Makeup gas flow rate
Injector temperature
Detector temperature
Initial temperature
Temperature program
Final temperature
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
265°C, hold 18 min
8081B-30
Revision 2
January 1998
-------
TABLE 6
RETENTION TIMES OF THE ORGANOCHLORINE PESTICIDES8
DUAL-COLUMN METHOD OF ANALYSIS
Compound
DBCP
Hexachlorocyclopentadiene
Etridiazole
Chloroneb
Hexachlorobenzene
Diallate
Propachlor
Trifluralin
a-BHC
PCNB
Y-BHC
Heptachlor
Aldrin
Alachlor
Chlorothalonil
Alachlor
p-BHC
Isodrin
DCPA
5-BHC
Heptachlor epoxide
Endosulfan-l
Y-Chlordane
a-Chlordane
frans-Nonachlor
4,4'-DDE
Dieldrin
Perthane
Endrin
Chloropropylate
Chlorobenzilate
Nitrofen
4,4'-DDD
Endosulfan II
4,4'-DDT
Endrin aldehyde
DB-5 RT (min)
2.14
4.49
6.38
7.46
12.79
12.35
9.96
11.87
12.35
14.47
14.14
18.34
20.37
18.58
15.81
18.58
13.80
22.08
21.38
15.49
22.83
25.00
24.29
25.25
25.58
26.80
26.60
28.45
27.86
28.92
28.92
27.86
29.32
28.45
31.62
29.63
DB-1701 RT(min)
2.84
4.88
8.42
10.60
14.58
15.07
15.43
16.26
17.42
18.20
20.00
21.16
22.78
24.18
24.42
24.18
25.04
25.29
26.11
26.37
27.31
28.88
29.32
29.82
30.01
30.40
31.20
32.18
32.44
34.14
34.42
34.42
35.32
35.51
36.30
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
Captafol 32.65 41.42
Endrin ketone 33.79 42.26
Permethrin 41.50 45.81
Kepone 31.10 b
Dicofol 35.33 b
Dichlone 15.17 b
a,a'-Dibromo-m-xylene 9.17 11.51
2-Bromobiphenyl 8.54 12.49
3 See Table 7 for GC operating conditions.
b Not detected at 2 ng per injection.
8081 B-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:
Column 2:
DB-1701 or equivalent
30-m x 0.53 mm ID
1.0 um film thickness
DB-5 or equivalent
30-m x 0.53 mm ID
0.83 um film thickness
Carrier gas
Carrier gas flow rate
Makeup gas
Makeup gas flow rate
Injector temperature
Detector temperature
Initial temperature
Temperature program
Final temperature
Helium
6 mL/minute
Nitrogen
20 mL/min
250°C
320°C
140°C, hold 2 minutes
140°C to 270°C at 2.8°C/min
270°C, hold 1 minute
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:
Column 2:
DB-1701 (J&W) or equivalent
30 m x 0.53 mm ID
1.0 um film thickness
DB-5 (J&W) or equivalent
30 m x 0.53 mm ID
1.5 um film thickness
Carrier gas:
Carrier gas flow rate:
Makeup gas:
Makeup gas flow rate:
Injector temperature:
Detector temperature:
Initial temperature:
Temperature program:
Final temperature
Helium
6 mL/minute
Nitrogen
20 (mL/min)
250°C
320°C
150°C, hold 0.5 min
150°C to 190°C at 12°C/min, hold 2 min190°C to 275°C
at 4°C/min
275°C, hold 10 min
8081B-34
Revision 2
January 1998
-------
TABLE 9
ANALYTE RECOVERY FROM SEWAGE SLUDGE
Compound
Hexachloroethane
2-Chloronapthalene
4-Bromodiphenyl ether
a-BHC
Y-BHC
Heptachlor
Aldrin
P-BHC
6-BHC
Heptachlor epoxide
Endosulfan 1
Y-Chlordane
a-Chlordane
DDE
Dieldrin
Endrin
Endosulfan II
DDT
Endrin aldehyde
ODD
Tetrachloro-m-xylene
Decachlorobiphenyl
Ultrasonic
% Recovery
80
50
118
88
55
60
92
351
51
54
52
50
49
52
89
56
52
57
45
57
71
26
Extraction
%RSD
7
56
4
25
9
13
33
71
11
11
11
9
8
11
19
10
10
10
6
11
19
23
Soxhlet
% Recovery
79
67
nd
265
155
469
875
150
57
70
70
65
66
74
327
92
88
95
42
99
82
28
%RSD
1
8
nd
18
29
294
734
260
2
3
4
1
0
1
7
15
11
17
10
8
1
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 m x 0.53 mm ID.
8081B- 35
Revision 2
January 1998
-------
TABLE 10
ANALYTE RECOVERY FROM DICHLOROETHANE STILLBOTTOMS
Compound
Hexachloroethane
2-Chloronapthalene
4-Bromodiphenyl ether
a-BHC
P-BHC
Heptachlor
Aldrin
(3-BHC
6-BHC
Heptachlor epoxide
Endosulfan I
Y-Chlordane
a-Chlordane
DDE
Dieldrin
Endrin
Endosulfan II
DDT
Endrin aldehyde
ODD
Tetrachloro-m-xylene
Decachlorobiphenyl
Ultrasonic
% Recovery
70
59
159
55
43
48
48
51
43
47
47
48
45
45
45
50
49
49
40
48
49
17
Extraction
%RSD
2
3
14
7
6
6
5
7
4
6
4
5
5
4
5
6
5
4
4
5
2
29
Soxhlet
%Recovery
50
35
128
47
30
55
200
75
119
66
41
47
37
70
58
41
46
40
29
35
176
104
%RSD
30
35
137
25
30
18
258
42
129
34
18
13
21
40
24
23
17
29
20
21
211
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.
8081B-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)3
% Recovery
Compound Name DB-5 DB-1701
a-BHC
P-BHC
Heptachlor
Aldrin
Heptachlor epoxide
trans-Chlordane
Endosulfan I
Dieldrin
Endrin
Endosulfan II
4,4'-DDT
Mirex
89
86
94
ND
97
94
92
ND
111
104
ND
108
94
ND
95
92
97
95
92
113
104
104
ND
102
a 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.
All compounds were spiked at 500 ug/kg.
Data taken from Reference 10.
8081B-37 Revision 2
January 1998
-------
TABLE 12
SINGLE LABORATORY RECOVERY DATA FOR SOLID-PHASE EXTRACTION OF
ORGANOCHLORINE PESTICIDES FROM TCLP BUFFERS SPIKED AT TWO LEVELS
Compound
Spike Level
(M9/L)
Buffer 1 (pH =
Recovery (%)
2.886)
RSD
Buffer 2 (pH =
Recovery (%)
: 4.937)
RSD
Low Level Spike
Toxaphene
Chlordane
Y-BHC (Lindane)
Heptachlor
Heptachlor
epoxide
Endrin
Methoxychlor
250
15
200
4
4
10
5000
86
88
115
95
107
89
97
13
7
7
11
9
5
8
77
95
98
77
104
100
95
17
6
5
23
12
6
6
High Level Spike
Toxaphene
Chlordane
Y-BHC (Lindane)
Heptachlor
Heptachlor
epoxide
Endrin
Methoxychlor
1000
60
800
16
16
40
20,000
106
116
109
113*
82
84
100
7
12
19
18*
17
19
4
Results were from seven replicate spiked buffer samples, except where
that only three replicates were analyzed.
8081 B- 38
85
107
112
93
91
82
87
15
12
5
3
7
4
8
noted with *, which indciates
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
Compound
Spike Level
(ug/L)*
%R
Labi
RSD
n
%R
Lab 2
RSD
n
%R
Lab 3
RSD
n
Buffer 1 pH = 2.886
Toxaphene
Chlordane
Y-BHC (Lindane)
Heptachlor
Heptachlor epoxide
Endrin
Methoxychlor
500
30
400
8
8
20
10,000
75
80
104
88
92
106
107
25
15
11
13
13
12
12
7
7
7
7
7
7
7
95.4
57.8
99.3
70.8
108.7
110
86.7
2.4
12.0
0.6
20.4
6.9
0
2.2
3
3
3
3
3
3
3
86.0
73.8
86.6
88.0
75.0
78.3
84.8
4.3
0.9
6.4
9.1
2.8
4.6
8.5
3
3
3
3
3
3
3
Buffer 2 pH = 4.937
Toxaphene
Chlordane
Y-BHC (Lindane)
Heptachlor
Heptachlor epoxide
Endrin
Methoxychlor
* 250-mL aliquots of leachate
500
30
400
8
8
20
10,000
were spiked
87
91
74
71
118
124
73
by Labs
9
8
20
21
1
7
22
7
7
7
7
3
3
7
98
66.7
102.7
62.5
113
111.7
88.8
2 and 3 at the levels show. Lab
8081 B- 39
4.1
5.0
2.2
20
0
2.6
2.7
1 spiked
3
3
3
3
3
3
3
88.8
73.7
89.3
85.0
81.3
83.0
89.6
4.1
11.5
3.1
1.5
2.7
3.4
2.7
3
3
3
3
3
3
3
at one-half these levels.
Revision 2
January 1998
-------
TABLE 14
SINGLE LABORATORY ACCURACY AND PRECISION DATA FOR SOLID-PHASE EXTRACTION BY METHOD 35351
Bias (%)
Compound
Aldrin
P-BHC
6-BHC
a-Chlordane
Y-Chlordane
Dieldrin
Endosulfan I
Endosulfan II
Endrin
Endrin aldehyde
Heptachlor
Heptachlor
epoxide
LJndane
p.p'-DDE
p.p'-DDT
p,p'-TDE (ODD)
Ground
water
(low)
37.3
89.2
106.2
75.4
70.7
83.4
79.6
94.5
88.3
87.5
43.1
76.4
81.3
80.3
86.6
90.5
Ground
water
(high)
93.5
107.8
86.0
112.3
98.9
96.1
99.1
101.6
98.4
99.9
95.4
97.6
115.2
96.0
105.4
101.1
Waste
water
(low)
79.3
79.7
88.9
78.9
79.9
81.2
79.6
82.7
85.1
69.0
71.8
75.3
82.1
85.1
105
74.9
Waste
water
(high)
94.0
82.3
83.4
89.5
93.9
93.3
87.9
93.5
89.6
80.2
78.6
83.4
85.3
97.9
111
79.6
Precision (%)
Ground Ground Waste
water water water
(low) (high) (low)
23.7
6.5
5.6
12.8
15.8
7.1
10.6
5.8
6.2
6.0
19.2
12.1
11.1
8.3
4.4
4.8
5.5
2.5
2.4
2.7
2.7
2.3
2.3
2.8
2.3
4.0
3.9
2.4
3.2
2.5
2.7
2.4
6.7
1.6
2.5
4.7
4.6
3.8
4.1
4.2
3.1
3.3
5.0
2.9
2.4
4.4
4.3
4.6
Waste
water
(high)
3.4
4.2
4.2
2.4
2.9
3.6
3.8
4.1
2.9
5.9
2.8
3.3
3.1
2.4
4.7
2.9
MDL
Ground Waste
water water
(pg/L) (ug/L)
1.4
0.91
0.93
1.5
1.8
0.9
1.3
0.9
1.7
0.8
1.3
1.5
1.4
1.0
0.6
1.4
0.83
0.20
0.35
0.58
0.58
0.49
0.51
0.54
0.82
0.36
0.56
0.34
0.32
0.59
0.71
0.85
1AII results determined from seven replicates of each sample type. Two spiking levels were used. "Low" samples were spiked at 5-10 pg/L for
each analyte, while "high" samples were spiked at 250 - 500 ug/L. MDL values were determined from the "low" samples without further
consideration of the spiking level.
8081 B-
40
Revision 2
January 1998
-------
TABLE 15
RECOVERY (BIAS) OF ORGANOCHLORINE PESTICIDES USING SFE METHOD 3562
(Seven replicates)
Compounds
Y-BHC
(3-BHC
Heptachlor
6-BHC
Aldrin
Heptachlor epoxide
a-Chlordane
4,4'DDE
Dieldrin
Endrin
4,4'-DDD
Endosulfan II
4,4'-DDT
Endrin aldehyde
Matrix Mean Recovery
Delphi3
250 ug/kg
102.6
101.9
101.3
120.9
56.7
102.3
106.4
110.9
106.9
211.0
93.0
105.6
126.7
64.3
107.9
Delphi-5 a
5 ug/kg
66.4
73.0
61.6
82.3
28.7
71.9
87.1
75.7
80.4
87.0
80.4
89.9
81.3
74.0
74.3
McCarthy"
250 ug/kg
80.7
86.1
78.0
90.4
52.1
87.1
88.1
88.4
88.1
111.7
85.0
92.1
110.9
63.0
85.9
McCarthy"
5 ug/kg
82.7
85.1
79.1
89.6
77.1
87.4
105.9
118.7
140.8
98.7
88.1
88.6
199.7
86.7
102.0
Auburn0
250 ug/kg
86.0
87.4
83.3
92.9
42.1
89.6
91.7
83.6
90.6
90.5
83.7
87.7
83.6
21.0
79.8
Auburn0
5 ug/kg
86.1
86.3
80.4
89.4
74.6
91.1
97.1
110.9
80.1
87.6
90.4
92.9
124.3
38.3
87.8
Mean
Recovery
84.1
86.6
80.6
94.2
55.2
88.2
96.1
98.0
97.8
114.4
86.8
92.5
121.1
37.9
89.5
8 Delphi: Loamy sand soil
b McCarthy: Sandy loamy-organic rich soil
c Auburn: Clay-loamy soil
8081 B- 41
Revision 2
January 1998
-------
TABLE 16
RELATIVE STANDARD DEVIATION (PRECISION) OF ORGANOCHLORINE PESTICIDES USING SFE METHOD 3562
(Seven replicates)
Compounds
Y-BHC
P-BHC
Heptachlor
6-BHC
Aldrin
Heptachlor epoxide
a-Chlordane
4,4'DDE
Dieldrin
Endrin
4,4'-DDD
Endosulfan II
4,4'-DDT
Endrin aldehyde
Matrix Mean Recovery
Delphi3
250 ug/kg
3.9
6.5
4.4
5.3
2.9
3.0
3.6
5.2
4.3
7.2
6.9
5.1
12.5
3.9
5.3
Delphi8
5 ug/kg
3.3
3.0
2.1
3.1
5.5
2.7
5.7
15.3
4.5
6.0
3.1
4.7
6.2
7.5
5.2
McCarthy"
250 ug/kg
3.3
3.0
4.3
3.3
2.8
3.6
4.8
4.8
2.9
4.5
3.7
3.2
6.6
4.7
4.0
McCarthy"
5 ug/kg
6.5
4.3
5.0
7.1
4.6
4.3
13.8
4.2
23.9
6.0
3.5
3.3
5.9
11.6
7.4
Auburn0
250 ug/kg
4.0
4.6
4.4
4.1
1.6
4.7
4.2
7.7
5.0
4.3
4.3
5.5
4.9
1.9
4.4
Auburn6
5 ug/kg Mean
1.6
2.0
2.6
3.5
1.9
4.2
2.5
3.4
3.1
10.5
7.4
4.6
3.4
26.0
5.5
3.7
3.9
3.8
4.4
3.2
3.8
5.8
6.8
7.3
6.4
4.8
4.4
6.6
9.3
5.3
a Delphi: Loamy sand soil
b McCarthy: Sandy loamy-organic rich soil
c Auburn: Clay-loamy soil
8081 B- 42
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FIGURE 1
GAS CHROMATOGRAM OF THE MIXED ORGANOCHLORINE PESTICIDE STANDARD
Start Time : 0.00 min
Scale Factor: 0
O-
:o
m
it"
Ss-
H
§'
L/1"
Ul
O"
End Time : 33.00 min Low Point : 20.00 MV High Point : 420.00 i
Plot Offset: 20 mV Plot Scale: 400 mV
Response [mV]
KJ rO OJ (jt
O
-------
FIGURE 2
GAS CHROMATOGRAM OF INDIVIDUAL ORGANOCHLORINE PESTICIDE STANDARD MIX A
Start Time : 0.00 min
Scale Factor: 0
Ul
End Time : 33.00 min
Plot Offset: 20 mV
Lou Point : 20.00 mV
Plot Scale: 250 mV
High Point : 270.00 i
Response [mV]
o.
0
L_J
1
§
^
:-5.13
c
tŁL
12.33
14.27
-10.98
CD
K3
17-08
20.22
20.77
-19.78
.13
-23.08
Ul
L-J
28.52
-30.05
Column:
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- 44
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FIGURE 3
GAS CHROMATOGRAM OF INDIVIDUAL ORGANOCHLORINE PESTICIDE STANDARD MIX B
Start Time : 0.00 min End Time : 33.00 min
Scale Factor: 0 Plot Offset: 20 mV
L.1—
ID
fD
iiT
H
3'
ID
78-
Ul
O"
Low Point : 20.00 mV High Point : 270.00 mV
Plot Scale: 250 mV
Response
en
o
o
o
o
ro
o
O
I I I I I I I I I I I I I I l l l l I l I l l I l l
ro
(_n
O
--2.74
-6.97
^9.
60
— 10.71
-11.73
•14.27
^-15.24
fi. ns
-14.84
-16.23
—17.08
-17.63
1.11
-18.31
19.54
^20.69
-20.19
21.03
22.00
--22.68
-S.95
-30.04
Column:
Temperature program:
30 m x 0.25 mm ID, DB-5
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.
8081 B-45
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FIGURE 4
GAS CHROMATOGRAM OF TOXAPHENE
8.00-
0.00
2.00
4.00 6.00 8.00 10.00 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 min hold) to 290°C at 6°C/min.
8081B-46
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FIGURE 5
GAS CHROMATOGRAM OF STROBANE
at
DB-1701
o ®
10 t\l
• r-
JuC
DB-5
cu
Strobane 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 (IS-pm film thickness) and 30 m x 0.53 mm ID DB-1701 (1 0-um
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
8081B-47
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FIGURES
GAS CHROMATOGRAM OF ORGANOCHLORINE PESTICIDES
DB-5
! I
,
J J
JM
A
2J
24
|2S
20
1 2
3 4 SU IS S
DB-1701
10 11 12 IS
14
1 35 3» 4
3t
4}
20.
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-um film thickness) and 30 m x 0.53
mm ID DB-1701 (1.0-um film thickness) connected to an 8 in. injection tee (Supeico Inc.). Temperature
program: 140°C (2 min hold) to 270°C (1 min hold) at 2.8°C/min.
8081B-48
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METHOD 8082A
POLYCHLORINATED BIPHENYLS (PCBs) 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
Aroclor 1016
Aroclor 1221
Aroclor 1232
Aroclor 1242
Aroclor 1248
Aroclor 1254
Aroclor 1260
2-Chlorobiphenyl
2,3-Dichlorobiphenyl
2,2',5-Trichlorobiphenyl
2,4',5-Trichlorobiphenyl
2,2',3,51-Tetrachlorobiphenyl
2,2',5,5'-Tetrachlorobiphenyl
2,3',4,4'-Tetrachlorobiphenyl
2,2',3,4,5I-Pentachlorobiphenyl
2,2',4,5,5'-Pentachlorobiphenyl
2,3,3',4',6-Pentachlorobiphenyl
2,2'13,4,4',51-Hexachlorobiphenyl
2,21,3I4,5,51-Hexachlorobiphenyl
2,2',3,5,5',6-Hexachlorobiphenyl
2,2l,4,41,5,5'-Hexachlorobiphenyl
2,2',3,3',4,4',5-Heptachlorobiphenyl
2,2',3,4,4I,5,51-Heptachlorobipheny!
2,2',3,4,4',5',6-Heptachlorobiphenyl
2,2113,4I,5,5',6-Heptachlorobiphenyl
2>2',3,3l,4,4I,5,5t,6-Nonachlorobiphenyl
CAS Registry No.
12674-11-2
11104-28-2
11141-16-5
53469-21-9
12672-29-6
11097-69-1
11096-82-5
2051-60-7
16605-91-7
37680-65-2
16606-02-3
41464-39-5
35693-99-3
32598-10-0
38380-02-8
37680-73-2
38380-03-9
35065-28-2
52712-04-6
52663-63-5
35065-27-1
35065-30-6
35065-29-3
52663-69-1
52663-68-0
40186-72-9
IUPAC*
_
-
_
.
_
.
_
1
5
18
31
44
52
66
87
101
110
138
141
151
153
170
180
183
187
206
8082A- 1
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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 Verify 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 analyte 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 = ———- x 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.
RF - RF
% Difference = ———- x 100
RF
8082A-12 Revision 1
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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. 8.2) based on the last initial calibration, then a new
initial calibration must be performed.
7.6.3 Inject a 2-uL aliquot of the concentrated sample extract. Record the volume
injected to the nearest 0.05 uL 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.
8082A-13 Revision 1
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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 analyzed 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.
8082A-14 Revision 1
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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 Aroclor 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 alter the PCBs to the point that the pattern of a specific Aroclor 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
8082A-15 Revision 1
January 1998
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be as high as 10 ng/|jL in the final extract, while ion trap or SIM may only require a
concentration of 1 ng/uL.
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, clean 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 dimethyldichlorosilane.
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
8082A-16 Revision 1
January 1998
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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 (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.
8.2.1 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 RGBs
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 ug/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,
8082A-17 Revision 1
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and detection limit). At a minimum, this includes the analysis of QC samples including a method
blank, a matn'x 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 ug/L in water and 57 to 70 ug/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 PCBs 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 Revision 1
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9.4 During method performance studies, the concentrations determined as Aroclors 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 Aroclor 1254 and
Aroclor 1260 were between 80% and 90%. Recoveries of congeners from environmental reference
materials ranged from 51 - 66% of the certified Aroclor 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 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:
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 Aroclor 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 Specific Organic Toxic Substances in
Wastewaters. Category 10 - Pesticides 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
-------
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* PCS 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 600/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. Bawadt, B. Johansson, S. Wunderii, 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," Anal. 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 Revision 1
January 1998
-------
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 Revision 1
January 1998
-------
TABLE 2
GC OPERATING CONDITIONS FOR PCBs AS AROCLORS
SINGLE COLUMN ANALYSIS
Narrow-bore columns
Narrow-bore 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 urn film thickness.
Carrier gas (He) 16 psi
Injector temperature 225°C
Detector temperature 300°C
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 5°C/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 methylpolysiloxane (DB-608, SPB-608, or equivalent) 25 urn coating thickness, 1 urn
film thickness
Carrier gas (N2) 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 urn or 0.83 urn
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 urn film thickness.
Carrier gas (He) 5-7 mL/minute
Makeup gas (argon/methane
[P-5 or P-10] or N2) 30 mL/min
Injector temperature 250°C
Detector temperature 290°C
Initial temperature 150°C, hold 0.5 minute
Temperature program 150°C to 270°C at 5°C/min
Final temperature 270°C, hold 10 min
8082A - 22 Revision 1
January 1998
-------
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 Mm film thickness.
Carrier gas (He) 6 mL/minute
Makeup gas (argon/methane
[P-5 or P-1 0] or N2) 30 mL/min
Injector temperature 205°C
Detector temperature 290°C
Initial temperature 140°C, hold 2 min
Temperature program 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
8082A - 23 Revision 1
January 1998
-------
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 m x 0.53 mm ID, 1.0 um film thickness.
Column 2 - DB-5 or equivalent, 30 m x 0.53 mm ID, 1.5 um 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 190°C, at 12°C/min, 2 min hold
190°C to 275°C, at 4°C/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
-------
TABLE 4
SUMMARY OF RETENTION TIMES OF AROCLORS
ON THE DB-5 COLUMN8, DUAL-COLUMN ANALYSIS
Peak
No.b
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
Aroclor
1016
8.41
8.77
8.98
9.71
10.49
10.58
10.90
11.23
11.88
11.99
12.27
12.66
12.98
13.18
13.61
13.80
13.96
14.48
14.63
14.99
15.35
16.01
16.27
Aroclor
1221
5.85
7.63
8.43
8.77
8.99
10.50
10.59
11.24
12.29
12.68
12.99
Aroclor
1232
5.85
7.64
8.43
8.78
9.00
10.50
10.59
10.91
11.24
11.90
12.00
12.29
12.69
13.00
13.19
13.63
13.82
13.97
14.50
14.64
15.02
15.36
16.14
16.29
17.04
17.22
17.46
18.41
18.58
Aroclor
1242
7.57
8.37
8.73
8.94
9.66
10.44
10.53
10.86
11.18
11.84
11.95
12.24
12.64
12.95
13.14
13.58
13.77
13.93
14.46
14.60
14.98
15.32
15.96
16.08
16.26
17.19
17.43
17.92
18.16
18.37
18.56
Aroclor
1248
8.95
10.45
10.85
11.18
11.85
12.24
12.64
12.95
13.15
13.58
13.77
13.93
14.45
14.60
14.97
15.31
16.08
16.24
16.99
17.19
17.43
17.69
17.91
18.14
18.36
18.55
Aroclor
1254
13.59
13.78
13.90
14.46
14.98
15.32
16.10
16.25
16.53
16.96
17.19
17.44
17.69
17.91
18.14
18.36
18.55
Aroclor
1260
13.59
16.26
16.97
17.21
18.37
18.68
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 - 25 Revision 1
January 1998
-------
TABLE 4
(continued)
Peak Aroclor Aroclor
No." 1016 1221
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
Aroclor Aroclor Aroclor Aroclor
1232 1242 1248 1254
18.83 18.80 18.78 18.78
19.33 19.30 19.29 19.29
19.48
19.81
20.03 19.97 19.92 19.92
20.28
20.46 20.45
20.57
20.85 20.83 20.83
21.18 21.14 21.12 20.98
21.36 21.38
21.78
22.08 22.05 22.04
22.38
22.74
22.96
23.23
23.75
23.99
24.27
24.61
24.93
26.22
Aroclor
1260
18.79
19.29
19.48
19.80
20.28
20.57
20.83
21.38
21.78
22.03
22.37
22.73
22.95
23.23
23.42
23.73
23.97
24.16
24.45
24.62
24.91
25.44
26.19
26.52
26.75
27.41
28.07
28.35
29.00
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 - 26 Revision 1
January 1998
-------
TABLE 5
SUMMARY OF RETENTION TIMES OF AROCLORS
ON THE DB-1701 COLUMN", DUAL COLUMN ANALYSIS
Peak
No.b
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
Aroclor
1016
6.33
6.78
6.96
7.64
8.23
8.62
8.88
9.05
9.46
9.77
10.27
10.64
11.01
11.09
11.98
12.39
12.92
12.99
13.14
13.49
13.58
Aroclor
1221
4.45
5.38
5.78
5.86
6.34
6.78
6.96
8.23
8.63
9.06
9.79
10.29
10.65
Aroclor
1232
4.45
5.86
6.34
6.79
6.96
8.23
8.63
8.89
9.06
9.47
9.78
10.29
10.66
11.02
11.10
11.99
12.39
12.77
13.00
13.16
13.49
13.61
14.08
14.30
14.49
15.38
Aroclor
1242
6.28
6.72
6.90
7.59
8.15
8.57
8.83
8.99
9.40
9.71
10.21
10.59
10.96
11.02
11.94
12.33
12.71
12.94
13.09
13.44
13.54
13.67
14.03
14.26
14.46
15.33
Aroclor
1248
6.91
8.16
8.83
8.99
9.41
9.71
10.21
10.59
10.95
11.03
11.93
12.33
12.69
12.93
13.09
13.44
13.54
14.03
14.24
14.39
14.46
15.10
15.32
Aroclor
1254
10.95
11.93
12.33
13.10
13.24
13.51
13.68
14.03
14.24
14.36
14.56
15.10
15.32
Aroclor
1260
13.52
14.02
14.25
14.56
aGC 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
and are not isomer numbers.
8082A - 27 Revision 1
January 1998
-------
TABLE 5
(continued)
Peak Aroclor Aroclor
No.b 1016 1221
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
Aroclor Aroclor
1232 1242
15.65 15.62
15.78 15.74
16.13 16.10
16.77 16.73
17.13 17.09
17.46
17.69
18.48
19.13
Aroclor
1248
15.62
15.74
16.10
16.74
17.07
17.44
17.69
18.19
18.49
19.13
20.57
Aroclor
1254
15.61
15.74
16.08
16.34
16.44
16.55
16.77
17.07
17.29
17.43
17.68
18.17
18.42
18.59
18.86
19.10
19.42
19.55
20.20
20.34
20.55
20.62
20.88
21.53
21.83
23.31
Aroclor
1260
16.61
15.79
16.19
16.34
16.45
16.77
17.08
17.31
17.43
17.68
18.18
18.40
18.86
19.09
19.43
19.59
20.21
20.43
20.66
20.87
21.03
21.53
21.81
23.27
23.85
24.11
24.46
24.59
24.87
25.85
27.05
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 Revision 1
January 1998
-------
TABLE 6
PEAKS DIAGNOSTIC OF PCBs OBSERVED ON 0.53 mm ID COLUMN
DURING SINGLE COLUMN ANALYSIS
Peak
No.8
1
II
III
IV
V
VI
VII
VIII
IX
X
XI
RTon
DB-608"
4.90
7.15
7.89
9.38
10.69
14.24
14.81
16.71
19.27
21.22
22.89
RTon
DB-1701b
4.66
6.96
7.65
9.00
10.54
14.12
14.77
16.38
18.95
21.23
22.46
Aroclor0
1221
1221, 1232, 1248
1061. 1221. 1232, 1242
1016, 1232, 1242, 1248
1016. 1232. 1242
1248. 1254
1254
1254
1254, 1260
1260
1260
a Peaks are sequentially numbered in elution order and are not isomer numbers
Temperature program: T, = 150°C, hold 30 seconds; 5°C/minute to 275°C.
Underline indicates largest peak in the pattern for that Aroclor
b
c
8082A - 29 Revision 1
January 1998
-------
TABLE 7
SPECIFIC PCB CONGENERS IN AROCLORS
Aroclor
Congener IUPAC number 1016 1221 1232 1242 1248 1254 1260
Biphenyl
2-CB
23-DCB
34-DCB
244--TCB
22'35'-TCB
23'44'-TCB
233'4'6-PCB
23'44'5-PCB
22'44'55'-HCB
22'344'5'-HCB
22'344'55'-HpCB
22'33'44'5-HpCB
1
5
12
28*
44
66*
110
118*
153
138
180
170
X
X X X X
X X X X X
X XXX
X XXX
XXX
X
X
X
X
X
X
X
X
X
X
X
X
X
•Apparent co-elution of: 28 with 31 (2,4',5-trichlorobiphenyl)
66 with 95 (2,2',3,5',6-pentachlorobiphenyl)
118 with 149 (2,2',3,4',5',6-hexachlorobiphenyl)
8082A - 30 Revision 1
January 1998
-------
TABLE 8
RETENTION TIMES OF PCB CONGENERS
ON THE DB-5 WIDE-BORE COLUMN
IUPAC # Retention Time (min)
1
5
18
31
52
44
66
101
87
110
151
153
138
141
187
183
180
170
206
209
6.52
10.07
11.62
13.43
14.75
15.51
17.20
18.08
19.11
19.45
19.87
21.30
21.79
22.34
22.89
23.09
24.87
25.93
30.70
32.63 (internal standard)
8082A - 31 Revision 1
January 1998
-------
TABLE 9
SINGLE LABORATORY RECOVERY DATA FOR EXTRACTION OF
RGBs FROM CLAY AND SOIL BY METHOD 3541a (AUTOMATED SOXHLET)
Matrix
Clay
Clay
Clay
Clay
Spike Level
Aroclor (ppm) Trial
1254 5 1
2
3
4
5
6
1254 50 1
2
3
4
5
6
1260 5 1
2
3
4
5
6
1260 50 1
2
3
4
5
6
Percent
Recovery"
87.0
92.7
93.8
98.6!
79.4
28.3
65.3
72.6
97.2
79.6
49.8
59.1
87.3
74.6
60.8
93.8
96.9
113.1
73.5
70.1
92.4
88.9
90.2
67.3
8082A - 32 Revision 1
January 1998
-------
Matrix
Soil
Soil
Soil
Soil
TABLE 9
(continued)
Spike Level
Aroclor (ppm) Trial
1254 5 1
2
3
4
5
1254 50 1
2
3
4
5
6
1260 5 1
2
3
4
5
6
7
1260 50 1
2
3
4
5
6
Percent
Recovery1*
69.7
89.1
91.8
83.2
62.5
84.0
77.5
91.8
66.5
82.3
61.6
83.9
82.8
81.6
96.2
93.7
93.8
97.5
76.9
69.4
92.6
81.6
83.1
76.0
a The operating conditions for the automated Soxhiet
Immersion time: 60 min
Reflux time: 60 min
b Multiple results from two different extractors
Data from Reference 9
8082A - 33
Revision 1
January 1998
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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 (M9/kg) Concentration (M9/kg) Recovery
Labi
Lab 2
Lab 3
Lab 4
Lab 5
Lab 6
Lab 7
Lab 8
All
Labs
n
Mean
S. D.
n
Mean
S. D.
n
Mean
S. D.
n
Mean
S. D.
n
Mean
S. D.
n
Mean
S. D.
n
Mean
S. D.
n
Mean
S. D.
n
Mean
S. D.
5
3
101.2
34.9
3
72.8
10.8
6
112.6
18.2
2
140.9
4.3
3
100.1
17.9
3
65.0
16.0
20
98.8
28.7
50
3
74.0
41.8
6
56.5
7.0
3
63.3
8.3
6
144.3
30.4
3
97.1
8.7
3
127.7
15.5
3
123.4
14.6
3
38.3
21.9
30
92.5
42.9
500 5
3
83.9
7.4
6
66.9
15.4
3
70.6
2.5
6
100.3
13.3
3
80.1
5.1
3
138.7
15.5
3
82.1
7.9
3
92.8
36.5
9 21
71.3 95.5
14.1 25.3
50
3
78.5
7.4
6
70.1
14.5
3
57.2
5.6
6
84.8
3.8
3
79.5
3.1
4
105.9
7.9
3
94.1
5.2
3
51.9
12.8
31
78.6
18.0
500 All Levels
12
84.4
26.0
6 24
74.5 67.0
10.3 13.3
12
66.0
9.1
24
110.5
28.5
3 12
77.0 83.5
9.4 10.3
12
125.4
18.4
12
99.9
19.0
12
62.0
29.1
9 120
75.3 87.6
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. ''
28
52
101
149
118
153
105°
138
128
156°
180
170
Matrix
Mean
SRM1941 CRM481"
EC-1C Dump Marine EC-5° Lake European
Site Soil Sediment Sediment Soil
1 Low#1 Low #2 Low #3 High#1
148.4
88.5
93.3
92.6
89.9
90.8
89.1
90.1
90.8
90.6
92.4
91.3
95.7
63.3
106.6
91.2
105.1
66.1
65.1
72.6
57.4
69.9
88.9
142.4
101.1
85.8
a Congeners which are either certified or have
b Parts per million (ug/g)
0 Congener 105 was not resolved from congener
EC-1 and EC-5
147.7
115.8
100.2
101.5
108.9
95.1
96.6
97.9
101.2
94.3
93.3
95.2
104.0
67.3
84.5
84.5
73.2
82.1
82.8
83.4
76.9
65.9
85.2
82.2
80.5
79.0
had Soxhlet confirmation
132 and congener 156 was
8082A - 35
Mich Bay CRM 392
Saginaw Sewage
Sediment Sludge
High #2 High #3
114.7 89.2
111.1 96.2
111.5 93.9
111.2
110.8 73.5
118.6 97.3
111.8
126.9
87.6
101.1
109.2 100.5
108.7 91.8
not resolved from congener 171
SRM 2974
Fish Tissue
Mussel
Low #4 Congener Mean
101.7
131.4
133.2
69.4
82.7
107.5
79.4
73.1
62.5
59.3
65.7
33.0
83.2
104.6
104.9
101.1
92.2
87.7
94.0
88.8
87.1
79.7
86.6
98.0
81.8
92.2
by the GC method used for samples
Revision 1
January 1998
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TABLE 12
% RELATIVE STANDARD DEVIATION (PRECISION) OF PCBs USING SFE METHOD 3562
PCB No. '
28
52
101
149
118
153
105°
138
128
156 c
180
170
Matrix
Mean
EC-1C Dump
Site Soil
Low#1
11.5
9.1
9.1
7.1
9.8
8.4
6.6
9.2
6.0
8.3
8.0
5.7
8.2
a Congeners which are either
b Parts per
c Congener
EC-1 and
SRM 1941
Marine EC-5° Lake
Sediment
Low #2
1.5
3.3
2.9
0.7
1.9
1.5
3.7
1.8
5.3
0.0
1.3
2.3
2.2
certified or have
Sediment
Low #3
3.8
3.9
2.8
3.8
4.5
3.0
2.7
3.1
3.3
5.1
3.6
3.6
3.6
had Soxhlet
CRM 481 b
European
Soil
High #1
5.6
5.4
4.9
3.9
5.4
4.3
4.3
4.7
4.9
4.5
4.3
3.9
4.7
confirmation.
million (mg/kg)
105 was not resolved from congener 132 and congener 156 was
EC-5.
8082A - 36
Mich Bay
Saginaw
Sediment
High #2
2.4
2.2
1.4
3.4
2.0
4.3
2.7
2.3
2.8
1.9
3.1
2.3
2.6
not resolved from
CRM 392
Sewage
Sludge
High #3
1.9
2.9
5.2
3.3
9.5
9.6
2.7
congener 171
SRM 2974
Fish Tissue
Mussel
Low #4
2.7
3.1
2.9
2.2
2.4
3.0
2.5
2.9
3.3
3.8
2.7
4.0
3.0
Congener Mean
4.2
4.3
4.2
3.0
4.2
4.9
3.2
3.4
3.7
3.4
4.7
3.1
3.8
by the GC method used for samples
Revision 1
January 1998
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TABLE 13
METHOD DETECTION LIMITS (MDLs)a OF PCBs USING SFE METHOD 3562
PCB No.
28
52
101
149
118
153
105 d
138
128
156 d
180
170
Matrix
Mean
SRM1941 CRM481b
EC-1cDump Marine EC-5° Lake European
Site Soil Sediment Sediment Soil
a Low#1 Low #2 Low #3 High #1
13.2
22.3
23.9
7.1
9.8
8.4
6.6
9.2
6.0
8.3
8.0
5.7
10.7
0.5
0.6
0.9
0.7
1.9
1.5
3.7
1.8
5.3
0.0
1.3
2.3
1.7
0.6
1.9
1.9
3.8
4.5
3.0
2.7
3.1
3.3
5.1
3.6
3.6
3.1
n/a
n/a
n/a
n/a
n/a
n/a
n/a
n/a
n/a
n/a
n/a
n/a
n/a
Mich Bay CRM 392
Saginaw Sewage
Sediment Sludge
High #2 High #3
15.4 5.3
29.9 6.9
3.3 20.6
3.7
3.3 7.5
3.5 97.3
2.6
1.7
0.5
0.2
1.9 94.5
0.6
5.6 19.3
a MDLs are highly matrix-dependant. MDLs provided in SW-846 are for guidance purposes and may not
establish their own in-house MDLs to document method performance.
b Congeners which are either certified or have had Soxhlet confirmation.
c Parts per million (mg/kg), THEREFORE LOW MDL IS NOT APPROPRIATE - Use mean ug/kg value.
d Congener 105 was not resolved from congener 132 and congener 156 was not resolved from congener 171
EC-1 and EC-5.
8082A - 37
SRM 2974
Fish Tissue
Mussel
Low #4 Congener Mean
5.0
9.1
9.7
4.1
6.9
9.
3.
7.
0.
0.
0.
3.
5.
always be
by the GC
4
1
2
6
6
9
1
0
achievable.
method used
6.6
11.8
10.1
3.2
5.7
20.5
3.1
3.8
2.6
2.4
18.4
2.6
7.6
Labs should
for samples
Revision 1
January
1998
-------
TABLE 14
SINGLE LABORATORY RECOVERY DATA FOR SPE (METHOD 3535)
OF AROCLOR 1254 FROM WASTEWATER MATRICES SPIKED AT 2 ug/L
Wastewater Type
Chemical Industry
Chemical Industry
Paper Industry
Paper Industry
Pharmaceutical Industry
Pharmaceutical Industry
Refuse
Refuse
POTW
POTW
Mean Cone.
(ug/L)
2.39
0.56
3.00
2.30
1.52
1.02
0.54
0.63
1.92
2.10
Percent
Recovery
120
28
150
115
76
51
27
31
96
105
Std. Dev.
(ug/L)
0.41
0.03
0.56
0.08
0.03
0.03
0.04
0.10
0.15
0.04
RSD
(%)
17.2
5.4
18.5
3.7
1.7
2.9
6.7
16.0
7.8
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
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TABLE 15
SINGLE LABORATORY RECOVERY DATA FOR SPE (METHOD 3535)
OF AROCLOR 1254 FROM WASTEWATER MATRICES SPIKED AT 10 ug/L
Wastewater Type
Chemical Industry
Chemical Industry
Paper Industry
Paper Industry
Pharmaceutical Industry
Pharmaceutical Industry
Refuse
Refuse
POTW
POTW
Mean Cone.
(M9/L)
8.75
8.08
8.88
10.14
9.19
8.42
8.80
8.00
9.52
8.18
Percent
Recovery
88
81
889
101
92
84
88
80
82
82
Std. Dev.
(ug/L)
1.07
0.06
0.71
0.15
0.24
0.17
0.49
1.44
0.17
0.17
RSD
(%)
12.2
0.7
7.9
1.4
2.6
2.0
5.6
18.0
2.1
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.
8082A - 39 Revision 1
January 1998
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TABLE 16
SINGLE LABORATORY RECOVERY DATA SPE (METHOD 3535)
OF AROCLOR 1254 FROM WASTEWATER MATRICES SPIKED AT 100 ug/L
Wastewater Type
Chemical Industry
Chemical Industry
Paper Industry
Paper Industry
Pharmaceutical Industry
Pharmaceutical Industry
Refuse
Refuse
POTW
POTW
Mean Cone.
(ug/L)
81.72
89.71
73.73
95.26
86.41
79.16
85.70
71.50
87.76
80.59
Percent
Recovery
82
90
74
95
86
79
86
72
88
81
Std. Dev.
(M9/L)
1.46
0.66
3.94
1.89
1.95
3.92
1.59
1.61
1.76
0.40
RSD
(%)
1.8
0.7
5.3
2.0
2.3
4.9
1.9
2.2
2.0
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
C18 extraction disks.
8082A - 40 Revision 1
January 1998
-------
o
CD
(M
DB-170
-O
O-
r*
DB-5
0
° »
Ct
JL ^ 1 1
i-
0
III
Ul
1
<
1
t
1.
I
(
t
1
o
Kl'
kr
1
1
III
1
M
1
o
(0
10
o>
u
r- u
FIGURE 1. GC/ECD chromatogram of Aroclor 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-um film thickness) and 30 m x 0.53 mm ID DB-1701 (1.0-um
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
-------
J'JL
r-o-r.
JU
08-1701
•CD
&IV
.V
-a «,
DB-5
1*1
r. i'i
• I-
o •
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-um film thickness) and 30 m x 0.53 mm ID DB-1701 (1.0-um
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 - 42
Revision 1
January 1998
-------
-K>
KDnn
DB-1701
r
•v
o
DB-5
FIGURE 3. GC/ECD chromatogram of Aroclor 1232 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-um film thickness) and 30 m x 0.53 mm ID DB-1701 (1.0-um
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
-------
I-
o-
a>
10
o>
ro <•*> (M H
T-O
n
T-
OHM K> T . 'I
UL__jw_JJU
K»
* I I
m
»
IO
V)
OB-1701
DB-5
li IOO
f4 ("^iiKI k»
1- ^ ( -^M>^ • »
rr-i'i --v o k>
• - «mo - r- k
-o«o— . . o o r
-•—»«•? • i-
i-| • r« .
'AA" ""
FIGURE 4. GC/ECD chromatogram of Aroclor 1242 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-|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 min hold) at 12°C/min then
to 275°C (10 min hold) at 4°C/min.
8082A - 44
Revision 1
January 1998
-------
Ill
(r
Cu
O)
r* m
o- —
T
ki
kt
I*
FIGURE 5. GC/ECD chromatogram of Aroclor 1248 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-um film thickness) and 30 m x 0.53 mm ID DB-1701 (1.0-um
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 - 45
Revision 1
January 1998
-------
DB-1701
i
IO
»
•0
li
DB-5
0 lUU lUO —
I) O'l uyi Hi
^ **
k>
f»
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-um film thickness) and 30 m x 0.53 mm ID DB-1701 (1.0-um
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
f» 19 M n»m
DB-5
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 (1.5-um film thickness) and 30 m x 0.53 mm ID DB-1701 (1.0-um
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 - 47
Revision 1
January 1998
-------
METHOD 8082A
POLYCHLORINATED BIPHENYLS (PCBs) BY GAS CHROMATOGRAPHY
('start
7.1 Choose
appropriate extraction
technique.
7.1 Add specified
matrix spike to sample.
7.2 Perform
extract cleanup.
7.3 Set
chromatographic
conditions.
7.4 Perform
initial calibration.
7.5 Establish retention
time windows.
7.6 Perform GC
analysis of sample
extracts.
7.6.3 Inject sample
extract.
7.6.5
Does
response fall
within
calibration
range?
7.6.10
Any sample
peak inter-
ferences?
7.6.5 Dilute
extract.
Take corrective
action. (Cleanup or
system adjustment
may be necessary.)
7.7 - 7.9 Qualitative
and Quantitative peak
identification.
8082A - 48
Revision 1
January 1998
-------
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-elution. 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.
Organophosphorus Pesticides
Asponb
Azinphos-methyl
Azinphos-ethyl8
Bolstar (Sulprofos)
Carbophenothion8
Chlorfenvinphos8
Chlorpyrifos
Chlorpyrifos methyl3
Coumaphos
Crotoxyphos8
Demeton-Oc
Demeton-Sc
Diazinon
Dichlorofenthion3
Dichlorvos (DDVP)
Dicrotophos3
Dimethoate
Dioxathion80
Disulfoton
EPN
Ethion3
Ethoprop
Famphur3
3244-90-4
86-50-0
2642-71-9
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
2104-64-5
563-12-2
13194-48-4
52-85-7
8141B-1
Revision 2
January 1998
-------
Analyte
CAS Registry No.
Fenitrothion3
Fensulfothion
Fenthion
Fonophos3
Leptophosa'd
Malathion
Merphos0
Mevinphos6
Monocrotophos
Naled
Parathion, ethyl
Parathion, methyl
Phorate
Phosmet3
Phosphamidon3
Ronnel
Stirophos (Tetrachlorvinphos)
Sulfotepp
Tetraethyl pyrophosphate (TEPP)d
Terbufos3
Thionazin3 b (Zinophos)
Tokuthion" (Prothiofos)
Trichlorfon3
Trichloronate"
Industrial Chemicals
Hexamethyl phosphoramide3 (HMPA)
Tri-o-cresyl phosphatea'd (TOCP)
Triazine Herbicides (NPD only)
Atrazine3
Simazine3
122-14-5
115-90-2
55-38-9
944-22-9
21609-90-5
121-75-5
150-50-5
7786-34-7
6923-22-4
300-76-5
56-38-2
298-00-0
298-02-2
732-11-6
13171-21-6
299-84-3
22248-79-9
3689-24-5
107-49-3
13071-79-9
297-97-2
34643-46-4
52-68-6
327-98-0
680-31-9
78-30-8
1912-24-9
122-34-9
3 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.
d Compound is extremely toxic or neurotoxic.
e 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 clean
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:
Azinphos-ethyl, Phosphamidon Dioxathion
Ethion Chlorfenvinphos Leptophos
Carbophenothion HMPA TOCP
Famphur Terbufos Phosmet
1.6 When Method 8141 is used to analyze unfamiliar samples, compound identifications
should be supported by confirmatory analysis. Sec. 8.0 provides gas chromatograph/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
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 acid 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 analytes 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)-sulfite option should
be employed, since copper 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 coelute, 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 coelute 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
Terbufos/tri-o-cresyl phosphate
Naled/Simazine/Atrazine
Dichlorofenthion/Demeton-O
Trichloronate/Aspon
Bolstar/Stirophos/Carbophenothion
Phosphamidon/Crotoxyphos
Fensulfothion/EPN
DB-210
Terbufos/tri-o-cresyl phosphate
Dichlorofenthion/Phosphamidon
Chlorpyrifos, methyl/Parathion, methyl
Chlorpyrifos/Parathion, ethyl
Aspon/Fenthion
Demeton-O/Dimethoate
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 lots 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
poor from aqueous solution.
recovery is
3.8.3 Naled is converted to Dichlorvos (DDVP) on column by debromination. 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 of Trichlorfon, 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
O-[2-(ethylthio)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 elutes 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 times 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 Method 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
split/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-column, 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-um 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-um 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 urn 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-um or 1.5-um 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.
2-3665M).
4.3.3 Splitter 3 - Restek Y-shaped fused-silica 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.
5.0 REAGENTS
5.1 Solvents - All solvents must be pesticide quality or equivalent.
5.1.1 Isooctane, (CH3)3CCH2CH(CH3)2
5.1.2 Hexane, C6H14
5.1.3 Acetone, CH3COCH3
5.1.4 Tetrahydrofuran (THF), C4H8O - for triazine standards only.
5.1.5 Methyl tert-butyl-ether (MTBE), CH3Of-C4H9 -for triazine standards only.
5.2 Stock standard solutions (1000 mg/L) - May be prepared from pure standard materials
or 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=S2]).
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 uL/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 analytes 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 the 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-ug/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 Florisil 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 analytes 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 analytes 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
organophosphorus 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-ul_ injections are recommended. Manual injections of no more
than 2 uL may be used if the analyst demonstrates quantitation precision of ^ 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 uL 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 ML 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
Corrective 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 will 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 "Y"s. 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
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. 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 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 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
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to 2500 ug/kg 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.; Mickey, 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
-------
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
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.
8141 B-17 Revision 2
January 1998
-------
TABLE 1
METHOD DETECTION LIMITS IN A WATER AND A SOIL MATRIX
USING 15-m COLUMNS AND A FLAME PHOTOMETRIC DETECTOR
Compound
Azinphos-methyl
Bolster (Sulprofos)
Chlorpyrifos
Coumaphos
Demeton, -O, -S
Diazinon
Dichlorvos (DDVP)
Dimethoate
Disulfoton
EPN
Ethoprop
Fensulfothion
Fenthion
Malathion
Merphos
Mevinphos
Naled
Parathion, ethyl
Parathion, methyl
Phorate
Ronnel
Sulfotepp
TEPP°
Tetrachlorovinphos
Tokuthion (Protothiofos)0
Trichloronate0
Reagent Water8 (ug/L)
0.10
0.07
0.07
0.20
0.12
0.20
0.80
0.26
0.07
0.04
0.20
0.08
0.08
0.11
0.20
0.50
0.50
0.06
0.12
0.04
0.07
0.07
0.80
0.80
0.07
0.80
Soil" (ug/kg)
5.0
3.5
5.0
10.0
6.0
10.0
40.0
13.0
3.5
2.0
10.0
4.0
5.0
5.5
10.0
25.0
25.0
3.0
6.0
2.0
3.5
3.5
40.0
40.0
5.5
40.0
aSample extracted using Method 3510, Separator/ 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.
81 41 B- 18
Revision 2
January 1998
-------
TABLE 2
DETERMINATION OF ESTIMATED QUANTITATION LIMITS (EQLs)
FOR VARIOUS MATRICES"
Matrix Factor
Ground water (Methods 3510 or 3520) 10b
Low-concentration soil by Soxhlet and no cleanup 10°
Non-water miscible waste (Method 3580) 1000°
a EQL = [Method detection limit (see Table 1)] X [Factor found in this table]. For non-aqueous
samples, the factor is on a wet-weight basis. Sample EQLs are highly 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.
81418-19 Revision 2
January 1998
-------
TABLE 3
RETENTION TIMES ON 15-m COLUMNS
Retention Time (min)
Analyte
TEPP
Dichlorvos (DDVP)
Mevinphos
Demeton, -O and -S
Ethoprop
Naled
Phorate
Monochrotophos
Sulfotepp
Dimethoate
Disulfoton
Diazinon
Merphos
Ronnel
Chlorpyrifos
Malathion
Parathion, methyl
Parathion, ethyl
Trichloronate
Tetrachlorovinphos
Tokuthion (Protothiofos)
Fensulfothion
Bolster (Sulprofos)
Famphur*
EPN
Azinphos-methyl
Fenthion
Coumaphos
DB-5
9.63
14.18
18.31
18.62
19.94
20.04
20.11
20.64
23.71
24.27
26.82
29.23
31.17
31.72
31.84
31.85
32.19
34.65
34.67
35.85
36.34
36.40
38.34
38.83
39.83
aGC operating conditions are shown in Table
"Method 8141 has not been
fully validated for
8141B-
SPB-608
6.44
7.91
12.88
15.90
16.48
19.01
17.52
20.11
18.02
20.18
19.96
20.02
21.73
22.98
26.88
28.78
23.71
27.62
28.41
32.99
24.58
35.20
35.08
36.93
37.80
38.04
29.45
38.87
8.
Famphur.
20
DB-210 DB-1
5.12 10.66
12.79
18.44
17.24
18.67
17.40 19.35
18.19
31.42
19.58
27.96
20.66
19.68
32.44
23.19
25.18
32.58
32.17
33.39
29.95
33.68
39.91
36.80
37.55
37.86
36.71 36.74
37.24
28.86
39.47
Revision 2
January 1998
-------
TABLE 4
RETENTION TIMES ON 30-m COLUMNS8
Retention Time (min)
Analyte
Trimethylphosphate
Dichlorvos (DDVP)
Hexamethylphosphoramide
Trichlorfon
TEPP
Thionazin
Mevinphos
Ethoprop
Diazinon
Sulfotepp
Terbufos
Tri-o-cresyl phosphate
Naled
Phorate
Fonophos
Disulfoton
Merphos
Oxidized Merphos
Dichlorofenthion
Chlorpyrifos, methyl
Ronnel
Chlorpyrifos
Trichloronate
Aspon
Fenthion
Demeton-S
Demeton-O
Monocrotophos0
Dimethoate
Tokuthion
Malathion
Parathion, methyl
DB-5
b
7.45
b
11.22
b
12.32
12.20
12.57
13.23
13.39
13.69
13.69
14.18
12.27
14.44
14.74
14.89
20.25
15.55
15.94
16.30
17.06
17.29
17.29
17.87
11.10
15.57
19.08
18.11
19.29
19.83
20.15
8141B-21
DB-210
2.36
6.99
7.97
11.63
13.82
24.71
10.82
15.29
18.60
16.32
18.23
18.23
15.85
16.57
18.38
18.84
23.22
24.87
20.09
20.45
21.01
22.22
22.73
21.98
22.11
14.86
17.21
15.98
17.21
24.77
21.75
20.45
DB-608
6.56
12.69
11.85
18.69
24.03
20.04
22.97
18.92
20.12
23.89
35.16
26.11
26.29
27.33
29.48
30.44
29.14
21.40
17.70
19.62
20.59
33.30
28.87
25.98
DB-1
10.43
14.45
18.52
21.87
19.60
18.78
19.65
21.73
26.23
23.67
24.85
24.63
20.18
19.3
19.87
27.63
24.57
22.97
Revision 2
January 1998
-------
TABLE 4
(continued)
Retention Time (min)
Analyte
Fenithrothion
Chlorfenvinphos
Parathion, ethyl
Bolster
Stirophos
Ethion
Phosphamidon
Crotoxyphos
Leptophos
Fensulfothion
EPN
Phosmet
Azinphos-methyl
Azinphos-ethyl
Famphur
Coumaphos
Atrazine
Simazine
Carbophenothion
Dioxathion
Trithion methyl
Dicrotophos
Internal Standard
1 -Bromo-2-nitrobenzene
Surrogates
Tributyl phosphate
Triphenyl phosphate
4-Chloro-3-nitrobenzotrifluoride
DB-5
20.63
21.07
21.38
22.09
22.06
22.55
22.77
22.77
24.62
27.54
27.58
27.89
28.70
29.27
29.41
33.22
13.98
13.85
22.14
d
8.11
5.73
DB-210
21.42
23.66
22.22
27.57
24.63
27.12
20.09
23.85
31.32
26.76
29.99
29.89
31.25
32.36
27.79
33.64
17.63
17.41
27.92
d
9.07
5.40
DB-608 DB-1
32.05
29.29 24.82
38.10 29.53
33.40 26.90
37.61
25.88
32.65
44.32
36.58 28.58
41.94 31.60
41.24
43.33 32.33
45.55
38.24
48.02 34.82
22.24
36.62
19.33
11.1
33.4
aGC 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 injected (shifts of over 30
seconds have been observed, see Reference 6).
d Shows multiple peaks; therefore, not included in the composite.
8141 B- 22
Revision 2
January 1998
-------
TABLE 5
RECOVERY OF 27 ORGANOPHOSPHATES BY SEPARATORY FUNNEL EXTRACTION
Percent Recovery at Three Spiking Levels
Analyte
Azinphos methyl
Bolstar
Chlorpyrifos
Coumaphos
Demeton
Diazinon
Dichlorvos
Dimethoate
Disulfoton
EPN
Ethoprop
Fensulfonthion
Fenthion
Malathion
Merphos
Mevinphos
Monocrotophos
Naled
Parathion, ethyl
Parathion, methyl
Phorate
Ronnel
Sulfotep
TEPP
Tetrachlorvinphos
Tokuthion
Trichloroate
NR = Not recovered
Low
126
134
7
103
33
136
80
NR
48
113
82
84
NR
127
NR
NR
NR
NR
101
NR
94
67
87
96
79
NR
NR
8141 B- 23
Medium
143 ±8
141 ±8
89 ±6
90 ±6
67 ±11
121 ±9.5
79 ±11
47 ±3
92 ±7
125 ±9
90 ±6
82 ±12
48 ±10
92 ±6
79
NR
18±4
NR
94 ±5
46 ±4
77 ±6
97 ±5
85 ±4
55 ±72
90 ±7
45 ±3
35
High
101
101
86
96
74
82
72
101
84
97
80
96
89
86
81
55
NR
NR
86
44
73
87
83
63
80
90
4
Revision 2
January 1998
-------
TABLE 6
RECOVERY OF 27 ORGANOPHOSPHATES BY CONTINUOUS LIQUID-LIQUID EXTRACTION
Analyte
Azinphos methyl
Bolstar
Chlorpyrifos
Coumaphos
Demeton
Diazinon
Dichlorvos
Dimethoate
Disulfoton
EPN
Ethoprop
Famphur
Fensulfonthion
Fenthion
Malathion
Merphos
Mevinphos
Monocrotophos
Naled
Parathion, ethyl
Parathion, methyl
Phorate
Ronnel
Sulfotep
TEPP
Tetrachlorvinphos
Tokuthion
Trichloroate
NR = Not recovered
Percent Recovery
Low
NR
NR
13
94
38
NR
81
NR
94
NR
39
—
90
8
105
NR
NR
NR
NR
106
NR
84
82
40
39
56
132
NR
8141 B- 24
at Three Spiking
Medium
129
126
82 ±4
79 ±1
23 ±3
128 ± 37
32 ±1
10 ±8
69 ±5
104 ±18
76 ±2
63 ±15
67 ±26
32 ±2
87 ±4
80
87
30
NR
81 ±1
50 ±30
63 ±3
83 ±7
77 ±1
18±7
70 ±14
32 ±14
NR
Levels
High
122
128
88
89
41
118
74
102
81
119
83
—
90
86
86
79
49
1
74
87
43
74
89
85
70
83
90
21
Revision 2
January 1998
-------
TABLE 7
RECOVERY OF 27 ORGANOPHOSPHATES BY SOXHLET EXTRACTION
Analyte
Azinphos methyl
Bolstar
Chlorpyrifos
Coumaphos
Demeton
Diazinon
Dichlorvos
Dimethoate
Disulfoton
EPN
Ethoprop
Fensulfonthion
Fenthion
Malathion
Merphos
Mevinphos
Monocrotophos
Naled
Parathion, ethyl
Parathion, methyl
Phorate
Ronnel
Sulfotep
TEPP
Tetrachlorvinphos
Tokuthion
Trichloroate
NR = Not recovered
Percent Recovery
Low
156
102
NR
93
169
87
84
NR
78
114
65
72
NR
100
62
NR
NR
NR
75
NR
75
NR
67
36
50
NR
56
8141 B- 25
at Three Spiking
Medium
110±6
103 ±15
66±17
89 ±11
64 ±6
96 ±3
39 ±21
48 ±7
78 ±6
93 ±8
70 ±7
81 ±18
43 ±7
81 ±8
53
71
NR
48
80 ±8
41 ±3
77 ±6
83 ±12
72 ±8
34 ±33
81 ±7
40 ±6
53
Levels
High
87
79
79
90
75
75
71
98
76
82
75
111
89
81
60
63
NR
NR
80
28
78
79
78
63
83
89
53
Revision 2
January 1998
-------
TABLE 8
SUGGESTED OPERATING CONDITIONS FOR 15-m COLUMNS
Columns 1 and 2 (DB-210 and SPB-608 or their equivalents^
Carrier gas (He) flow rate 5mL/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 (DB-5 or equivalent)
Carrier gas (He) flow rate
Initial temperature
Temperature program
5mL/min
130°C, hold for 3 minutes
130°C, to 180°C at 5°C/min, hold for 10
minutes, followed by 180°C 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
-------
Column 1
Column 2
TABLE 9
SUGGESTED OPERATING CONDITIONS FOR 30-m COLUMNS
DB-210
Dimensions: 30-m x 0.53-mm ID
Film Thickness (urn): 1.0
DB-5
Dimensions: 30-m x 0.53-mm ID
Film Thickness (urn): 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 pL
Hexane
Flash vaporization
Dual NPD
1
64
Y-shaped or Tee
Integrator
20psi
400°C
4
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
8141B-27
Revision 2
January 1998
-------
TABLE 10
QUANTITATION AND CHARACTERISTIC IONS FOR GC/MS ANALYSIS
OF ORGANOPHOSPHORUS PESTICIDES
Analyte
Azinphos-methyl
Bolstar (Sulprofos)
Chlorpyrifos
Coumaphos
Demeton-S
Diazinon
Dichlorvos (DDVP)
Dimethoate
Disulfoton
EPN
Ethoprop
Fensulfothion
Fenthion
Malathion
Merphos
Mevinphos
Monocrotophos
Naled
Parathion, ethyl
Parathion, methyl
Phorate
Ronnel
Stirophos
Sulfotepp
TEPP
Tokuthion
Quantitation ion
160
156
197
109
88
137
109
87
88
157
158
293
278
173
209
127
127
109
291
109
75
285
109
322
99
113
8141B-28
Characteristic ions
77,132
140,143,113,33
97,199,125.314
97,226,362,21
60,114,170
179,152,93,199,304
79,185,145
93,125,58,143
89,60,61,97,142
169,141,63,185
43,97,41,126
97,125,141,109,308
125,109,93,169
125,127,93,158
57,153,41,298
109,67,192
67,97,192,109
145,147,79
97,109,139,155
125,263,79
121,97,47,260
125,287,79,109
329,331,79
97,65,93,121,202
155,127,81,109
43,162,267,309
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
Aspon
Azinphos-methyl
Azinphos-ethyl
Bolster
Carbophenothion
Chlorfenvinphos
Chlorpyrifos
Chlorpyrifos methyl
Coumaphos
Crotoxyphos
Demeton
Diazinon
Dichlorofenthion
Dichlorvos (DDVP)
Dicrotophos
Dimethoate
Dioxathion
Disulfoton
EPN
Ethion
Ethoprop
Famphur
Fenitrothion
Fensulfothion
% Recovery
85.6
83.0
88.3
96.1
85.6
87.8
98.8
82.5
84.3
86.3
93.6
91.7
85.2
88.1
88.6
99.3
81.6
93.2
73.8
85.5
95.6
85.2
91.2
86.2
RSD
11.5
13.4
10.8
4.2
11.0
10.2
5.7
12.0
8.7
10.5
4.5
4.7
10.9
6.7
10.8
1.8
14.1
7.6
10.6
10.6
4.1
10.2
8.8
6.4
8141 B- 29
% Recovery
77.7
109.7
92.8
78.2
81.7
90.1
77.5
59.4
100.8
89.4
73.8
70.0
75.6
90.1
75.7
76.7
92.7
79.5
67.9
79.2
81.4
75.6
85.0
97.2
RSD
6.8
7.0
8.1
4.3
7.2
6.0
4.2
7.5
13.5
5.9
5.1
5.0
6.0
7.9
5.7
9.5
11.0
6.1
7.9
6.5
3.7
8.3
5.0
6.0
MDL"
1.7
2.4
2.4
1.1
1.9
1.7
1.0
1.4
4.3
1.7
1.2
1.1
1.4
2.2
1.3
2.3
3.2
1.5
1.7
1.6
0.9
2.0
1.3
1.8
Revision 2
January 1998
-------
TABLE II (cont.)
Ground Water spiked at
250 ppb
Analyte
Fenthion
Fonophos
Leptophos
Malathion
Merphos
Mevinphos
Naled
Parathion, ethyl
Parathion, methyl
Phorate
Phosmet
Phosphamidon
Ronnel
Stirophos
Sulfotepp
Terbufos
Thionazin
Tokuthion
Trichlorfon
Trichloronate
% Recovery
91.2
91.0
81.3
79.5
113.1
57.9
90.1
76.7
93.9
92.3
66.1
86.2
94.7
78.6
75.3
87.1
95.1
94.4
72.7
95.3
RSD
5.4
8.0
12.2
6.9
9.3
6.9
6.7
9.6
5.8
7.1
17.7
11.2
5.2
13.1
9.3
10.5
8.0
4.1
13.5
4.5
Ground Water spiked at
10 ppb
% Recovery
79.5
81.6
73.6
78.0
84.6
96.8
88.1
69.6
83.6
70.8
90.3
80.6
77.8
106.3
68.9
78.0
88.6
77.8
45.6
75.7
RSD
4.3
3.6
8.8
8.7
4.5
6.7
7.9
8.1
4.7
6.7
10.7
5.7
4.7
5.9
8.6
3.7
3.4
5.6
6.9
3.9
MDLa
1.7
0.9
2.0
2.1
1.2
2.0
2.2
1.8
1.2
1.5
3.0
1.4
1.2
2.0
1.9
0.9
1.0
1.4
1.0
0.9
a All MDL values are in ug/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
250 ppb
Analyte
Aspon
Azinphos-methyl
Azinphos-ethyl
Bolster
Carbophenothion
Chlorfenvinphos
Chlorpyrifos
Chlorpyrifos methyl
Coumaphos
Crotoxyphos
Demeton
Diazinon
Dichlorfenthion
Dichlorvos (DDVP)
Dicrotophos
Dimethoate
Dioxathion
Disulfoton
EPN
Ethion
Ethoprop
Famphur
Fenitrothion
Fensulfothion
% Recovery
83.7
102.6
79.8
94.4
82.4
81.7
91.0
77.6
100.2
81.3
95.8
91.8
82.5
60.6
82.0
93.5
84.6
92.5
78.1
83.5
96.3
85.9
83.5
101.7
RSD
1.8
18.0
6.8
8.3
2.9
6.5
8.3
2.2
17.2
5.7
5.3
6.5
1.4
11.1
1.6
4.1
5.6
5.3
9.6
2.0
4.7
2.5
4.8
11.4
8141B-31
Wastewater spiked at
10 ppb
% Recovery
76.3
129.9
96.0
84.9
82.1
88.0
86.5
56.7
111.0
87.5
88.5
82.4
76.2
99.7
73.4
115.7
100.4
90.4
80.1
78.4
92.9
78.6
82.3
110.5
RSD
6.7
12.4
6.7
1.4
6.7
7.2
1.7
7.1
8.5
7.0
5.0
3.2
5.5
6.1
6.1
6.7
9.4
2.6
8.6
6.4
3.1
7.9
5.9
6.5
MDL'
1.6
5.1
2.0
0.4
1.7
2.0
0.5
1.3
3.0
1.9
1.4
0.8
1.3
1.9
1.4
2.4
3.0
0.7
2.2
1.6
0.9
1.9
1.5
2.3
Revision 2
January 1998
-------
TABLE 12
PERFORMANCE DATA FOR ORGANOPHOSPHORUS PESTICIDES
IN WASTEWATER USING METHOD 3535
(continued)
Wastewater spiked at
250 ppb
Analyte
Fenthion
Fonophos
Leptophos
Malathion
Merphos
Mevinphos
Naled
Parathion ethyl
Parathion methyl
Phorate
Phosmet
Phosphamidon
Ronnel
Stirophos
Sulfotepp
Terbufos
Thionazin
Tokuthion
Trichlorfon
Trichloronate
% Recovery
91.7
83.4
81.9
94.8
94.5
62.6
60.6
80.2
92.9
92.4
63.5
81.1
91.4
101.4
78.7
83.0
85.1
91.8
66.8
91.3
a All MDL values are in ug/L, and are highly
guidance purposes and may not always be
MDLs to document method performance.
RSD
7.3
2.6
3.3
6.7
12.7
11.2
11.1
8.1
6.5
6.4
8.2
3.1
8.4
14.3
10.7
1.5
5.8
8.4
4.6
8.1
Wastewater spiked at
10 ppb
% Recovery
88.2
81.3
73.2
94.7
90.7
109.0
99.7
83.6
93.8
85.6
101.3
78.0
88.3
126.5
87.9
80.1
84.8
83.6
52.2
84.3
RSD
2.7
5.0
7.5
5.5
1.4
4.8
6.1
8.6
4.4
2.4
9.1
5.7
2.2
6.5
8.8
6.4
4.9
1.8
8.7
1.6
MDL"
0.7
1.3
1.7
1.6
0.4
1.6
1.9
2.3
1.3
0.6
2.9
1.4
0.6
2.6
2.4
1.6
1.3
0.5
1.4
0.4
matrix dependant. MDLs provided in SW-846 are for
achievable. Labs should establish their own in-house
8141B-32
Revision 2
January 1998
-------
TABLE 13
RECOVERIES OF ANALYTES FROM SPIKED SOIL SAMPLES
USING PRESSURIZED FLUID EXTRACTION (METHOD 3545)
Clay
Analyte
Dichlorvos
Mevinphos
Demeton O&S
Ethoprop
TEPP
Phorate
Sulfotep
Naled
Diazinon
Disulfoton
Monocrotophos
Dimethoate
Ronnel
Chlorpyrifos
Parathion methyl
Parathion ethyl
Fenthion
Tokuthion
Tetrachlorvinphos
Bolstar
Fensulfothion
EPN
Azinphos-methyl
Coumaphos
Results are expressed
Reference 15.
Low
100.0
100.6
103.7
97.4
100.0
98.8
102.6
100.0
97.6
121.8
100.0
92.5
96.4
98.3
94.9
95.4
95.9
97.1
93.8
99.1
100.0
85.8
100.0
100.0
High
280.0
98.0
106.2
95.8
100.0
96.6
99.3
100.0
96.2
86.8
100.0
90.7
95.3
96.3
97.7
97.4
96.5
95.8
93.8
98.4
89.9
97.3
92.4
94.0
as the percentage of
Loam
Low
135.1
104.0
124.3
101.2
100.0
104.6
113.2
100.0
104.2
112.0
100.0
94.3
102.4
98.0
98.9
99.1
104.0
102.4
144.0
105.1
81.2
88.8
85.0
85.1
High
158.5
99.8
108.4
97.2
100.0
98.5
119.1
100.0
101.7
92.5
100.0
89.0
85.0
97.0
98.5
99.5
100.4
96.5
92.3
97.5
76.8
97.1
81.4
90.6
Sand
Low
103.0
91.5
103.4
90.0
100.0
92.6
129.4
100.0
89.3
76.9
100.0
88.5
94.1
90.2
91.3
87.9
83.1
94.2
95.2
96.6
90.1
92.8
96.3
102.6
High
230.2
107.8
106.0
98.4
100.0
100.0
104.2
100.0
100.4
90.7
100.0
101.8
98.7
100.2
98.3
98.2
99.2
98.2
101.9
102.5
102.1
104.8
103.4
109.8
amount determined by a Soxhiet extraction. Data from
8141B-33
Revision 2
January 1998
-------
TABLE 14
BIAS AND PRECISION OF PRESSURIZED FLUID EXTRACTION
(METHOD 3545) OF THREE SPIKED SOIL SAMPLES
Clay
Low
Analyte
Dichlorvos
Mevinphos
Oemeton O&S
Ethoprop
TEPP
Phorate
Sulfotep
Naled
Diazinon
Disulfoton
Monocrotophos
Dimethoate
Ronnel
Chlorpyrifos
Parathion methyl
Parathion ethyl
Fenthion
Tokuthion
Tetrachlorvinphos
Bolstar
Fensulfothion
EPN
Azinphos-methyl
Coumaphos
Bias
0.0
66.1
79.0
83.0
0.0
67.5
66.6
0.0
80.2
55.9
0.0
87.0
81.3
99.5
82.5
85.0
56.4
96.1
72.1
89.0
0.0
72.6
0.0
0.0
Pre
NA
3.8
3.4
4.7
NA
3.2
3.7
NA
4.7
3.6
NA
5.0
3.7
3.1
3.9
3.8
3.8
4.7
3.3
3.4
NA
44.3
NA
NA
High
Bias
5.6
67.2
80.2
84.8
0.0
79.4
69.4
0.0
80.3
93.9
0.0
86.7
81.1
99.0
84.5
83.5
71.4
97.0
69.7
109.5
69.7
76.9
90.6
79.6
Pre
19.0
4.8
4.2
4.8
NA
5.1
4.7
NA
4.8
4.7
NA
5.3
5.0
5.1
5.2
5.2
5.0
5.7
Loam
Low
Bias Pre
10.4 11.4
57.3 11.2
73.7 10.0
76.1 10.7
0.0 NA
63.4 11.8
62.6 11.0
0.0 NA
74.4 12.0
58.9 11.8
0.0 NA
70.7 12.1
73.1 11.1
81.7 14.1
74.4 11.5
77.3 11.9
44.1 10.8
93.2 12.2
5.6 101.4 12.6
6.8
4.3
8.0
5.3
4.8
NA = not applicable
Bias was calculated as the percent recovery of the
relative standard deviation (RSD). Total number
Data from Reference 15.
82.2 9.9
70.4 9.3
92.9 10.1
69.7 13.9
62.8 13.4
High
Bias
6.5
63.1
77.6
77.0
0.0
73.5
66.8
0.0
75.9
89.4
0.0
71.7
64.7
87.7
79.6
79.6
50.9
93.8
64.7
89.2
52.2
70.4
70.5
6.5
Pre
22.2
6.5
6.4
4.9
NA
5.4
7.3
NA
6.0
6.2
NA
18.8
6.5
16.8
5.8
6.1
6.6
6.1
6.5
6.2
7.1
7.1
8.7
10.2
Sand
Low
Bias
13.9
61.6
60.0
75.5
0.0
62.9
62.1
0.0
73.9
52.2
0.0
75.0
69.0
84.1
74.9
78.0
44.3
81.2
69.3
77.3
63.0
68.6
94.5
74.8
certified spiking value. Precision
of analyses of each sample was
Pre
13.4
14.3
12.5
12.8
NA
13.6
13.8
NA
14.0
15.3
NA
13.1
13.6
13.1
13.2
12.7
12.5
12.5
11.9
11.7
9.2
11.2
12.5
16.1
High
Bias
9.9
64.7
77.6
79.0
0.0
76.2
67.7
0.0
77.4
88.5
0.0
80.6
73.8
91.6
80.3
80.3
51.9
85.4
69.6
94.2
62.0
71.9
82.5
72.9
was calculated
7.
8141 B- 34
Pre
22.2
12.1
12.7
10.6
NA
10.8
13.2
NA
11.2
12.3
NA
12.5
11.6
12.7
11.3
11.5
12.6
11.9
13.0
12.8
13.1
11.6
11.4
9.2
as the
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
I „.
n
.2
i
a
>.
a
r
c
(A
! 8
G
S
V... J
L
1 3 5 7 9 11 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 organophosphorus 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
'K
I
s
a.
UJ
«/>
2
O.
' "' i' • •' t.. -i. •. i n,, ,... ,t..,..,,...,..., .,... i ,...,...,...,
1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45
8141B-36
Revision 2
January 1998
-------
FIGURE 3
Chromatogram of target organophosphorus compounds from a 15-m DB-210 column with NPD
detector. More compounds are shown in Figure 4. See Table 3 for retention times.
300.00
250.00
200.00-
150.00-
100.00-
50.00-
0.00 —
1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45
8141B-37
Revision 2
January 1998
-------
FIGURE 4
Chromatogram of target 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.
300.00-,
250.00-
200.00-
150.00-
100.00-
50.00-
0.00
1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 '
45
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 GO
operating conditions.
-r
DB-210
DB-5
8141B-39
Revision 2
January 1998
-------
FIGURE 6
Chromatogram of target organophosphorus compounds on a 30-m DB-5/DB-210 column pair with NPD
detector, with Simazine, Atrazine and Carbophenothion. See Table 4 for retention times and for GC
operating conditions.
-r
\ .
n
U\-r—•'
DB-210
it
it
n
si
vt&Vj
n
a
41
DB-5
8141B-40
Revision 2
January 1998
-------
-------
-------
METHOD 8270D
SEMIVOLATILE ORGANIC COMPOUNDS
BY GAS CHROMATOGRAPHY/MASS SPECTROMETRY (GC/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"
Compounds
Acenaphthene
Acenaphthylene
Acetophenone
2-Acetylaminofluorene
1-Acetyl-2-thiourea
Aldrin
2-Aminoanthraquinone
Aminoazobenzene
4-Aminobiphenyl
3-Amino-9-ethylcarbazole
Anilazine
Aniline
o-Anisidine
Anthracene
Aramite
Aroclor 1016
Aroclor 1221
Aroclor 1232
Aroclor 1242
Aroclor 1248
Aroclor 1254
Aroclor 1260
Azinphos-methyl
Barban
Benzidine
Benzoic acid
Benz(a)anthracene
Benzo(b)fluoranthene
CAS No'
83-32-9
208-96-8
98-86-2
53-96-3
591-08-2
309-00-2
117-79-3
60-09-3
92-67-1
132-32-1
101-05-3
62-53-3
90-04-0
120-12-7
140-57-8
12674-11-2
11104-28-2
11141-16-5
53469-21-9
12672-29-6
11097-69-1
11096-82-5
86-50-0
101-27-9
92-87-5
65-85-0
56-55-3
205-99-2
8270D - 1
3510
X
X
X
X
LR
X
X
X
X
X
X
X
X
X
HS(43)
X
X
X
X
X
X
X
HS(62)
LR
CP
X
X
X
3520
X
X
ND
ND
ND
X
ND
ND
ND
X
ND
X
ND
X
ND
X
X
X
X
X
X
X
ND
ND
CP
X
X
X
3540/
3541
X
X
ND
ND
ND
X
ND
ND
ND
ND
ND
ND
ND
X
ND
X
X
X
X
X
X
X
ND
ND
CP
ND
X
X
3550
X
X
ND
ND
ND
X
ND
ND
ND
ND
ND
X
ND
X
ND
X
X
X
X
X
X
X
ND
ND
CP
X
X
X
3580
X
X
X
X
LR
X
X
X
X
ND
X
X
X
X
X
X
X
X
X
X
X
X
X
LR
CP
X
X
X
Revision 4
January 1998
-------
Appropriate Preparation Techniques"
Compounds
Benzo(k)fluoranthene
Benzo(g,h,i)perylene
Benzo(a)pyrene
p-Benzoquinone
Benzyl alcohol
a-BHC
P-BHC
6-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
Chlordane (NOS)
Chlorfenvinphos
4-Chloroaniline
Chlorobenzilate
5-Chloro-2-methylaniline
4-Chloro-3-methylphenol
3-(Chloromethyl)pyridine
hydrochloride
1 -Chloronaphthalene
2-Chloronaphthalene
2-Chlorophenol
4-Chloro-1 ,2-phenylenediamine
4-Chloro-1 ,3-phenylenediamine
4-Chlorophenyl phenyl ether
Chrysene
Coumaphos
p-Cresidine
Crotoxyphos
CAS Noa
207-08-9
191-24-2
50-32-8
106-51-4
100-51-6
319-84-6
319-85-7
319-86-8
58-89-9
111-91-1
111-44-4
108-60-1
117-81-7
101-55-3
1689-84-5
85-68-7
2425-06-1
133-06-2
63-25-2
1563-66-2
786-19-6
57-74-9
470-90-6
106-47-8
510-15-6
95-79-4
59-50-7
6959-48-4
90-13-1
91-58-7
95-57-8
95-83-0
5131-60-2
7005-72-3
218-01-9
56-72-4
120-71-8
7700-17-6
8270D - 2
3510
X
X
X
OE
X
X
X
X
X
X
X
X
X
X
X
X
HS(55)
HS(40)
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
3520
X
X
X
ND
X
X
X
X
X
X
X
X
X
X
ND
X
ND
ND
ND
ND
ND
X
ND
ND
ND
ND
X
ND
X
X
X
X
X
X
X
ND
ND
ND
3540/
3541
X
X
X
ND
ND
X
X
X
X
X
X
X
X
X
ND
X
ND
ND
ND
ND
ND
X
ND
ND
ND
ND
X
ND
X
X
X
ND
ND
X
X
ND
ND
ND
3550
X
X
X
ND
X
X
X
X
X
X
X
X
X
X
ND
X
ND
ND
ND
ND
ND
X
ND
ND
ND
ND
X
ND
X
X
X
ND
ND
X
X
ND
ND
ND
3580
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
ND
ND
X
X
X
X
X
Revision 4
January 1998
-------
Appropriate Preparation Techniques"
Compounds
2-Cyclohexyl-4,6-dinitro-phenol
4,4'-DDD
4,4'-DDE
4,4'-DDT
Demeton-O
Demeton-S
Diallate (cis or trans)
2,4-Diaminotoluene
Dibenz(a,j)acridine
Dibenz(a,h)anthracene
Dibenzofuran
Dibenzo(a,e)pyrene
1 ,2-Dibromo-3-chloropropane
Di-n-butyl phthalate
Dichlone
1 ,2-Dichlorobenzene
1 ,3-Dichlorobenzene
1 ,4-Dichlorobenzene
3,3'-Dichlorobenzidine
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-Dimethylphenethylamine
2,4-Dimethylphenol
Dimethyl phthalate
1 ,2-Dinitrobenzene
1 ,3-Dinitrobenzene
1 ,4-Dinitrobenzene
4,6-Dinitro-2-methylphenol
CAS No'
131-89-5
72-54-8
72-55-9
50-29-3
298-03-3
126-75-0
2303-16-4
95-80-7
224-42-0
53-70-3
132-64-9
192-65-4
96-12-8
84-74-2
117-80-6
95-50-1
541-73-1
106-46-7
91-94-1
120-83-2
87-65-0
62-73-7
141-66-2
60-57-1
84-66-2
56-53-1
64-67-5
60-51-5
119-90-4
60-11-7
57-97-6
119-93-7
122-09-8
105-67-9
131-11-3
528-29-0
99-65-0
100-25-4
534-52-1
8270D -
3510
X
X
X
X
HS(68)
X
X
DC,OE(42)
X
X
X
ND
X
X
OE
X
X
X
X
X
X
X
X
X
X
AW,OS(67)
LR
HE,HS(31)
X
X
CP(45)
X
ND
X
X
X
X
HE(14)
X
3
3520
ND
X
X
X
ND
ND
ND
ND
ND
X
X
ND
X
X
ND
X
X
X
X
X
ND
ND
ND
X
X
ND
ND
ND
ND
ND
ND
ND
ND
X
X
ND
ND
ND
X
3540/
3541
ND
X
X
X
ND
ND
ND
ND
ND
X
ND
ND
ND
X
ND
X
X
X
X
X
ND
ND
ND
X
X
ND
ND
ND
ND
ND
ND
ND
ND
X
X
ND
ND
ND
X
3550
ND
X
X
X
ND
ND
ND
ND
ND
X
X
ND
ND
X
ND
X
X
X
X
X
ND
ND
ND
X
X
ND
ND
ND
ND
ND
ND
ND
ND
X
X
ND
ND
ND
X
3580
LR
X
X
X
X
X
X
X
X
X
X
X
ND
X
X
X
X
X
X
X
X
X
X
X
X
X
LR
X
LR
X
CP
X
X
X
X
X
X
X
X
Revision 4
January
1998
-------
Appropriate Preparation Techniques"
Compounds
2,4-Dinitrophenol
2,4-Dinitrotoluene
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
Fenthion
Fluchloralin
Fluoranthene
Fluorene
2-Fluorobiphenyl (surr)
2-Fluorophenol (surr)
Heptachlor
Heptachlor epoxide
Hexachlorobenzene
Hexachlorobutadiene
Hexachlorocyclopentadiene
Hexachloroethane
Hexachlorophene
Hexachloropropene
Hexamethylphosphoramide
Hydroquinone
lndeno(1 ,2,3-cd)pyrene
CAS Noa
51-28-5
121-14-2
606-20-2
39300-45-3
88-85-7
122-39-4
57-41-0
122-66-7
117-84-0
298-04-4
959-98-8
33213-65-9
1031-07-8
72-20-8
7421-93-4
53494-70-5
2104-64-5
563-12-2
51-79-6
62-50-0
52-85-7
115-90-2
55-38-9
33245-39-5
206-44-0
86-73-7
321-60-8
367-12-4
76-44-8
1024-57-3
118-74-1
87-68-3
77-47-4
67-72-1
70-30-4
1888-71-7
680-31-9
123-31-9
193-39-5
8270D
3510
X
X
X
CP,HS(28)
X
X
X
X
X
X
X
X
X
X
X
X
X
X
DC(28)
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
AW,CP(62)
X
X
ND
X
-4
3520
X
X
X
ND
ND
X
ND
X
X
ND
X
X
X
X
X
X
ND
ND
ND
ND
ND
ND
ND
ND
X
X
X
X
X
X
X
X
X
X
ND
ND
ND
ND
X
3540/
3541
X
X
X
ND
ND
X
ND
X
X
ND
X
X
X
X
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
X
X
X
X
X
X
X
X
X
ND
ND
ND
ND
X
3550
X
X
X
ND
ND
X
ND
X
X
ND
X
X
X
X
X
X
ND
ND
ND
ND
ND
ND
ND
ND
X
X
X
X
X
X
X
X
X
X
ND
ND
ND
ND
X
3580
X
X
X
CP
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
CP
X
X
X
X
Revision 4
January 1998
-------
Appropriate Preparation Techniques"
Compounds
Isodrin
Isophorone
Isosafrole
Kepone
Leptophos
Malathion
Maleic anhydride
Mestranol
Methapyrilene
Methoxychlor
3-Methylcholanthrene
4,4'-Methylenebis (2-chloroaniline)
4,4'-Methylenebis(N, N-dimethyl-
aniline)
Methyl methanesulfonate
2-Methylnaphthalene
Methyl parathion
2-Methylphenol
3-Methylphenol
4-Methylphenol
Mevinphos
Mexacarbate
Mirex
Monocrotophos
Naled
Naphthalene
1 ,4-Naphthoquinone
1-Naphthylamine
2-Naphthylamine
Nicotine
5-Nitroacenaphthene
2-Nitroaniline
3-Nitroaniline
4-Nitroaniline
5-Nitro-o-anisidine
Nitrobenzene
4-Nitrobiphenyl
Nitrofen
2-Nitrophenol
CAS No"
465-73-6
78-59-1
120-58-1
143-50-0
21609-90-5
121-75-5
108-31-6
72-33-3
91-80-5
72-43-5
56-49-5
101-14-4
101-61-1
66-27-3
91-57-6
298-00-0
95-48-7
108-39-4
106-44-5
7786-34-7
315-18-4
2385-85-5
6923-22-4
300-76-5
91-20-3
130-15-4
134-32-7
91-59-8
54-11-5
602-87-9
88-74-4
99-09-2
100-01-6
99-59-2
98-95-3
92-93-3
1836-75-5
88-75-5
8270D -
3510
X
X
DC(46)
X
X
HS(5)
HE
X
X
X
X
OE,OS(0)
X
X
X
X
X
X
X
X
HE,HS(68)
X
HE
X
X
X
OS(44)
X
DE(67)
X
X
X
X
X
X
X
X
X
5
3520
ND
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
ND
ND
ND
ND
X
X
X
ND
X
ND
ND
X
35407
3541
ND
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
ND
X
3550 3580
ND
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
ND
ND
ND
ND
X
X
X
ND
X
ND
ND
X
X
X
X
X
X
X
X
X
X
X
X
LR
ND
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
Revision 4
January
1998
-------
Appropriate Preparation Techniques"
Compounds
4-Nitrophenol
5-Nitro-o-toluidine
Nitroquinoline-1 -oxide
N-Nitrosodi-n-butylamine
N-Nitrosodiethylamine
N-Nitrosodimethylamine
N-Nitrosomethylethylamine
N-Nitrosodiphenylamine
N-Nitrosodi-n-propylamine
N-Nitrosomorpholine
N-Nitrosopiperidine
N-Nitrosopyrrolidine
Octamethyl pyrophosphoramide
4,4'-Oxydianiline
Parathion
Pentachlorobenzene
Pentachloronitrobenzene
Pentachlorophenol
Phenacetin
Phenanthrene
Phenobarbital
Phenol
1 ,4-Phenylenediamine
Phorate
Phosalone
Phosmet
Phosphamidon
Phthalic anhydride
2-Picoline (2-Methylpyridine)
Piperonyl sulfoxide
Pronamide
Propylthiouracil
Pyrene
Resorcinol
Safrole
Strychnine
Sulfallate
Terbufos
1 ,2,4,5-Tetrachlorobenzene
CAS No8
100-02-7
99-55-8
56-57-5
924-16-3
55-18-5
62-75-9
10595-95-6
86-30-6
621-64-7
59-89-2
100-75-4
930-55-2
152-16-9
101-80-4
56-38-2
608-93-5
82-68-8
87-86-5
62-44-2
85-01-8
50-06-6
108-95-2
106-50-3
298-02-2
2310-17-0
732-11-6
13171-21-6
85-44-9
109-06-8
120-62-7
23950-58-5
51-52-5
129-00-0
108-46-3
94-59-7
57-24-9
95-06-7
13071-79-9
95-94-3
8270D -
3510
X
X
X
X
X
X
X
X
X
ND
X
X
LR
X
X
X
X
X
X
X
X
DC(28)
X
X
HS(65)
HS(15)
HE(63)
CP,HE(1)
X
X
X
LR
X
DC,OE(10)
X
AW,OS(55)
X
X
X
6
3520
X
X
ND
ND
ND
X
ND
X
X
ND
ND
ND
ND
ND
X
ND
ND
X
ND
X
ND
X
ND
ND
ND
ND
ND
ND
X
ND
ND
ND
X
ND
ND
ND
ND
ND
ND
35407
3541
X
ND
ND
ND
ND
X
ND
X
X
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
X
ND
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
ND
ND
ND
ND
ND
3550
X
ND
ND
ND
ND
X
ND
X
X
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
X
ND
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
ND
ND
ND
ND
ND
ND
3580
X
X
X
X
X
X
X
X
X
X
X
X
LR
X
X
X
X
X
X
X
X
X
X
X
X
X
X
CP
ND
X
X
LR
X
X
X
X
X
X
X
Revision 4
January 1998
-------
Compounds
CAS Noa
Appropriate Preparation Techniques"
3540/
3510 3520 3541 3550 3580
2,3,4,6-Tetrachlorophenol
Tetrachlorvinphos
Tetraethyl dithiopyrophosphate
Tetraethyl pyrophosphate
Thionazine
Thiophenol (Benzenethiol)
Toluene diisocyanate
o-Toluidine
Toxaphene
1 ,2,4-Trichlorobenzene
2,4,5-Trichlorophenol
2,4,6-Trichlorophenol
Trifluralin
2,4,5-Trimethylaniline
Trimethyl phosphate
1 ,3,5-Trinitrobenzene
Tris(2,3-dibromopropyl) phosphate
Tri-p-tolyl phosphate
O,O,O-Triethyl phosphorothioate
58-90-2
961-11-5
3689-24-5
107-49-3
297-97-2
108-98-5
584-84-9
95-53-4
8001-35-2
120-82-1
95-95-4
88-06-2
1582-09-8
137-17-7
512-56-1
99-35-4
126-72-7
78-32-0
126-68-1
X
X
X
X
X
X
HE(6)
X
X
X
X
X
X
X
HE(60)
X
X
X
X
ND
ND
X
ND
ND
ND
ND
ND
X
X
X
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
X
ND
X
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
X
X
X
X
ND
ND
ND
ND
ND
ND
ND
x
X
ND
X
X
X
X
X
X
X
X
X
X
X
X
X
LR
X
X
a Chemical Abstract Service Registry Number
b See Sec. 1.2 for other acceptable preparation methods.
in
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
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 70 percent recovery by this technique.
1.2 In addition to the sample preparation methods listed in the above analyte 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 sharp peaks from a gas chromatographic fused-silica capillary column coated with
a slightly polar silicons. Such compounds include polynuclear aromatic hydrocarbons, chlorinated
hydrocarbons and pesticides, phthalate esters, organophosphate esters, nitrosamines, haloethers,
aldehydes, ethers, ketones, anilines, pyridines, quinolines, 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., Arodors, 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 Arodors, Toxaphene, and Chlordane.
1.4 The following compounds may require spedal 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 II, 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-methylphenol, 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 1998
-------
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 ug/kg (wet weight) for soil/sediment samples, 1-200 mg/kg for
wastes (dependent on matrix and method of preparation), and 10 ug/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 spectra with the electron impact (or electron impact-like) spectra of authentic
standards. Quantitation is accomplished by comparing the response of a major (quantitation) ion
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 corrective 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-um 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 uL
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 GC-to-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
data 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 Spectral Library
should also be available.
8270D-10 Revision 4
January 1998
-------
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-ul_.
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 acid, 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-d4, naphthalene-ds, acenaphthene-d 10, phenanthrene-d 10, chrysene-d 12, and perylene-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/uL. Each 1-mL sample
extract undergoing analysis should be spiked with 10 uL of the internal standard solution,
resulting in a concentration of 40 ng/uL 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/uL of
decafluorotriphenylphosphine (DFTPP) should be prepared. The standard should also contain 50
ng/uL each of 4,4-DDT, 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 Calibration standards - A minimum of five calibration standards should be prepared at
five different concentrations. 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 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, the 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 aliquot of calibration standard should be spiked with 10 uL 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 phenol-d6, 2-fluorophenol,
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, carbon 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 and sorbent resin)
Water (including TCLP leachates)
Soil/sediment
Waste
3542
3510, 3520, 3535
3540, 3541, 3545, 3550, 3560, 3561
3540, 3541, 3545, 3550, 3560, 3561, 3580
7.1.2 In very limited applications, direct injection of the sample into the GC/MS
system with a 10-uL syringe may be appropriate. The detection limit is very high
(approximately 10,000 ug/L). Therefore, it is only permitted where concentrations in excess
of 10,000 [jg/L 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 acid
priority pollutants
Methods
3620
3630, 3640, 8041a
3610, 3620, 3640
3610, 3620, 3640
3610, 3620, 3630, 3660, 3665
3620, 3640
3611,3630,3640
3620, 3640
3620, 3640
3620
3611,3650
3640
a Method 8041 includes a derivatization technique and a GC/ECD analysis, if interferences
are encountered on GC/FID.
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° Cat 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 ML
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 ODD 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 uL 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:
AS = Peak area (or height) of the analyte or surrogate.
Ai8 = Peak area (or height) of the internal standard.
Cs = Concentration of the analyte or surrogate, in ug/L.
Cis = Concentration of the internal standard, in ug/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-propylamine;
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 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.
7.3.5 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%.
n
ERF
i
~ ~ \|
mean RF = RF = — SD
Ł(RFrRF)2
n-1
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RSD = 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
analyte in each calibration standard should agree within 0.06 RRT units. Late-eluting target
analytes usually have much better agreement.
___ _ 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 EICP 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 uL of the internal standard solution to the 1-mL concentrated sample extract obtained
from sample preparation.
7.5.3 Inject a 1-2 uL 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/uL, 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
Isosafrol) 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 chromatographically 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 shoulder(s) 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 (RT) 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 muKicomponent compounds (e.g., Toxaphene, Aroclors, 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/uL in the concentrated
sample extract.
7.7.6 Structural isomers that produce very 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 Isosafrol) 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 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 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 GC/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 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 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
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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 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.
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 property. 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 ug/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 Revision 4
January 1998
-------
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 Soxhlet extraction) are
presented in Table 9. Single laboratory accuracy and precision data were obtained for semivolatile
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/mass 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 and subsequently analyzed by Method 8270.
9.5 Single laboratory precision and bias data using Method 3545 (pressurized fluid
extraction) for semivolatile organic compounds are presented in Table 12. The samples were
conditioned spiked samples prepared and certified by a commercial supplier that contained 57
semivolatile organics at three concentrations (250, 2500, and 12,500 ug/kg) 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 Soxtec™ (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 - CO2; Modifier -10% 1:1 (v/v)
8270D - 24 Revision 4
January 1998
-------
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. "Interlaboratory 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., Kornfeld, R.A., "GC-MS Suitability Testing of RCRA Appendix VIII and Michigan
List Analytes", U.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 Extractability 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 Revision 4
January 1998
-------
10. Lee, H.B., Peart, I.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 III, 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 Revision 4
January 1998
-------
TABLE 1
CHARACTERISTIC IONS FOR SEMIVOLATILE COMPOUNDS
Compound
2-Picoline
Aniline
Phenol
Bis(2-chloroethyl) ether
2-Chlorophenol
1 ,3-Dichlorobenzene
1 ,4-Dichlorobenzene-d4 (IS)
1 ,4-Dichlorobenzene
Benzyl alcohol
1 ,2-Dichlorobenzene
N-Nitrosomethylethylamine
Bis(2-chloroisopropyl) ether
Ethyl carbamate
Thiophenol (Benzenethiol)
Methyl methanesulfonate
N-Nitrosodi-n-propylamine
Hexachloroethane
Maleic anhydride
Nitrobenzene
Isophorone
N-Nitrosodiethylamine
2-Nitrophenol
2,4-Dimethylphenol
p-Benzoquinone
Bis(2-chloroethoxy)methane
Benzoic acid
2,4-Dichlorophenol
Trimethyl phosphate
Ethyl methanesulfonate
1 ,2,4-Trichlorobenzene
Naphthalene-d8 (IS)
Naphthalene
Hexachlorobutadiene
Tetraethyl pyrophosphate
Diethyl sulfate
4-Chloro-3-methylphenol
2-Methylnaphthalene
2-Methylphenol
Hexachloropropene
Hexachlorocyclopentadiene
N-Nitrosopyrrolidine
Acetophenone
Retention
Time (min)
3.75"
5.68
5.77
5.82
5.97
6.27
6.35
6.40
6.78
6.85
6.97
7.22
7.27
7.42
7.48
7.55
7.65
7.65
7.87
8.53
8.70
8.75
9.03
9.13
9.23
9.38
9.48
9.53
9.62
9.67
9.75
9.82
10.43
11.07
11.37
11.68
11.87
12.40
12.45
12.60
12.65
12.67
8270D - 27
Primary
Ion
93
93
94
93
128
146
152
146
108
146
88
45
62
110
80
70
117
54
77
82
102
139
122
108
93
122
162
110
79
180
136
128
225
99
139
107
142
107
213
237
100
105
Secondary lon(s)
66,92
66,65
65,66
63,95
64,130
148,111
150,115
148,111
79,77
148,111
42,43,56
77,121
44,45,74
66,109,84
79,65,95
42,101,130
201,199
98,53,44
123,65
95,138
42,57,44,56
109,65
107,121
54,82,80
95,123
105,77
164,98
79,95,109,140
109,97,45.65
182,145
68
129,127
223,227
155,127,81,109
45,59,99,111,125
144,142
141
108,77,79,90
211,215,117,106,141
235,272
41,42,68,69
71,51,120
Revision 4
January 1998
-------
TABLE 1
(continued)
Compound
4-Methylphenol
2,4,6-Trichlorophenol
o-Toluidine
3-Methylphenol
2-Chloronaphthalene
N-Nitrosopiperidine
1 ,4-Phenylenediamine
1 -Chloronaphthalene
2-Nitroaniline
5-Chloro-2-methylaniline
Dimethyl phthalate
Acenaphthylene
2,6-Dinitrotoluene
Phthalic anhydride
o-Anisidine
3-Nitroaniline
Acenaphthene-d10 (IS)
Acenaphthene
2,4-Dinitrophenol
2,6-Dinitrophenol
4-Chloroaniline
Isosafrole
Dibenzofuran
2,4-Diaminotoluene
2,4-Dinitrotoluene
4-Nitrophenol
2-Naphthylamine
1 ,4-Naphthoquinone
p-Cresidine
Dichlorovos
Diethyl phthalate
Fluorene
2,4,5-Trimethylaniline
N-Nitrosodi-n-butylamine
4-Chlorophenyl phenyl ether
Hydroquinone
4,6-Dinitro-2-methylphenol
Resorcinol
N-Nitrosodiphenylamine
Safrole
Hexamethyl phosphoramide
3-(Chloromethyl)pyridine hydrochloride
Diphenylamine
Retention
Time (min)
12.82
12.85
12.87
12.93
13.30
13.55
13.62
13.65a
13.75
14.28
14.48
14.57
14.62
14.62
15.00
15.02
15.05
15.13
15.35
15.47
15.50
15.60
15.63
15.78
15.80
15.80
16.003
16.23
16.45
16.48
16.70
16.70
16.70
16.73
16.78
16.93
17.05
17.13
17.17
17.23
17.33
17.50
17.548
8270D - 28
Primary
Ion
107
196
106
107
162
114
108
162
65
106
163
152
165
104
108
138
164
154
184
162
127
162
168
121
165
139
143
158
122
109
149
166
120
84
204
110
198
110
169
162
135
92
169
Secondary lon(s)
108,77,79,90
198,200
107,77,51,79
108,77,79,90
127,164
42,55,56,41
80,53,54,52
127,164
92,138
141,140,77,89
194,164
151,153
63,89
76,50,148
80,123,52
108,92
162,160
153,152
63,154
164,126,98,63
129,65,92
131,104,77,51
139
122,94,77,104
63,89
109,65
115,116
104,102,76,50,130
94,137,77,93
185,79,145
177,150
165,167
135,134,91,77
57,41,116,158
206,141
81,53,55
51,105
81,82,53,69
168,167
104,77,103,135
44,179,92,42
127,129,65,39
168,167
Revision 4
January 1998
-------
TABLE 1
(continued)
Compound
1 ,2,4,5-Tetrachlorobenzene
1-Naphthylamine
1 -Acetyl-2-thiourea
4-Bromophenyl phenyl ether
Toluene diisocyanate
2,4,5-Trichlorophenol
Hexachlorobenzene
Nicotine
Pentachlorophenol
5-Nitro-o-toluidine
Thionazine
4-Nitroaniline
Phenanthrene-d10 (IS)
Phenanthrene
Anthracene
1 ,4-Dinitrobenzene
Mevinphos
Naled
1 ,3-Dinitrobenzene
Diallate (cis or trans)
1 ,2-Dinitrobenzene
Diallate (trans or cis)
Pentachlorobenzene
5-Nitro-o-anisidine
Pentachloronitrobenzene
4-Nitroquinoline-1 -oxide
Di-n-butyl phthalate
2,3,4,6-Tetrachlorophenol
Dihydrosaffrole
Demeton-O
Fluoranthene
1 ,3,5-Trinitrobenzene
Dicrotophos
Benzidine
Trifluralin
Bromoxynil
Pyrene
Monocrotophos
Phorate
Sulfallate
Demeton-S
Phenacetin
Dimethoate
Retention
Time (min)
17.97
18.20
18.22
18.27
18.42
18.47
18.65
18.70
19.25
19.27
19.35
19.37
19.55
19.62
19.77
19.83
19.90
20.03
20.18
20.57
20.58
20.78
21.35
21.50
21.72
21.73
21.78
21.88
22.42
22.72
23.33
23.68
23.82
23.87
23.88
23.90
24.02
24.08
24.10
24.23
24.30
24.33
24.70
Primary
Ion
216
143
118
248
174
196
284
84
266
152
107
138
188
178
178
168
127
109
168
86
168
86
250
168
237
174
149
232
135
88
202
75
127
184
306
277
202
127
75
188
88
108
87
Secondary lon(s)
214,179,108,143,218
115,89,63
43,42,76
250,141
145,173,146,132,91
198,97,132,99
142,249
133,161,162
264,268
77,79,106,94
96,97,143,79,68
65,108,92,80,39
94,80
179,176
176,179
75,50,76,92,122
192,109,67,164
145,147,301,79,189
76,50,75,92,122
234,43,70
50,63,74
234,43,70
252,108,248,215,254
79,52,138,153,77
142,214,249,295,265
101,128,75,116
150,104
131,230,166,234,168
64,77
89,60,61,115,171
101,203
74,213,120,91,63
67,72,109,193,237
92,185
43,264,41,290
279,88,275,168
200,203
192,67,97,109
121,97,93,260
88,72,60,44
60,81,89,114,115
180,179,109,137,80
93,125,143,229
8270D - 29
Revision 4
January 1998
-------
TABLE 1
(continued)
Compound
Phenobarbital
Carbofuran
Octamethyl pyrophosphoramide
4-Aminobiphenyl
Dioxathion
Terbufos
a.a-Dimethylphenylamine
Pronamide
Aminoazobenzene
Dichlone
Dinoseb
Disulfoton
Fluchloralin
Mexacarbate
4,4'-Oxydianiline
Butyl benzyl phthalate
4-Nitrobiphenyl
Phosphamidon
2-Cyclohexyl-4,6-Dinitrophenol
Methyl parathion
Carbaryl
Dimethylaminoazobenzene
Propylthiouracil
Benz(a)anthracene
Chrysene-d12 (IS)
3,3'-Dichlorobenzidine
Chrysene
Malathion
Kepone
Fenthion
Parathion
Anilazine
Bis(2-ethylhexyl) phthalate
3,3'-Dimethylbenzidine
Carbophenothion
5-Nitroacenaphthene
Methapyrilene
Isodrin
Captan
Chlorfenvinphos
Crotoxyphos
Phosmet
EPN
Retention
Time (min)
24.70
24.90
24.95
25.08
25.25
25.35
25.43
25.48
25.72
25.77
25.83
25.83
25.88
26.02
26.08
26.43
26.55
26.85
26.87
27.03
27.17
27.50
27.68
27.83
27.88
27.88
27.97
28.08
28.18
28.37
28.40
28.47
28.47
28.55
28.58
28.73
28.77
28.95
29.47
29.53
29.73
30.03
30.11
Primary
Ion
204
164
135
169
97
231
58
173
197
191
211
88
306
165
200
149
199
127
231
109
144
225
170
228
240
252
228
173
272
278
109
239
149
212
157
199
97
193
79
267
127
160
157
Secondary lon(s)
117,232,146,161
149,131,122
44,199,286,153,243
168,170,115
125,270,153
57,97,153,103
91,65,134,42
175,145,109,147
92,120,65,77
163,226,228,135,193
163,147,117,240
97,89,142,186
63,326,328,264,65
150,134,164,222
108,171,80,65
91,206
152,141,169,151
264,72,109,138
185,41,193,266
125,263,79,93
115,116,201
120,77,105,148,42
142,114,83
229,226
120,236
254,126
226,229
125,127,93,158
274,237,178,143,270
125,109,169,153
97,291,139,155
241,143,178,89
167,279
106,196,180
97,121,342,159,199
152,169,141,115
50,191,71
66,195,263,265,147
149,77,119,117
269,323,325,295
105,193,166
77,93,317,76
169,185,141,323
8270D - 30
Revision 4
January 1998
-------
TABLE 1
(continued)
Compound
Tetrachlorvinphos
Di-n-octyl phthalate
2-Aminoanthraquinone
Barban
Aramite
Benzo(b)fluoranthene
Nitrofen
Benzo(k)fluoranthene
Chlorobenzilate
Fensulfothion
Ethion
Diethylstilbestrol
Famphur
Tri-p-tolyl phosphate"
Benzo(a)pyrene
Perylene-d12 (IS)
7, 1 2-Dimethylbenz(a)anthracene
5,5-Diphenylhydantoin
Captafol
Dinocap
Methoxychlor
2-Acetylaminofluorene
4,4'-Methylenebis(2-chloroaniline)
3,3'-Dimethoxybenzidine
3-Methylcholanthrene
Phosalone
Azinphos-methyl
Leptophos
Mirex
Tris(2,3-dibromopropyl) phosphate
Dibenz(a,j)acridine
Mestranol
Coumaphos
lndeno(1 ,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
1 ,2:4,5-Dibenzopyrene
Strychnine
Piperonyl sulfoxide
Hexachlorophene
Aldrin
Aroclor 1016
Aroclor 1221
Retention
Time (min)
30.27
30.48
30.63
30.83
30.92
31.45
31.48
31.55
31.77
31.87
32.08
32.15
32.67
32.75
32.80
33.05
33.25
33.40
33.47
33.47
33.55
33.58
34.38
34.47
35.07
35.23
35.25
35.28
35.43
35.68
36.40
36.48
37.08
39.52
39.82
41.43
41.60
45.15
46.43
47.98
—
—
—
Primary
Ion
329
149
223
222
185
252
283
252
251
293
231
268
218
368
252
264
256
180
79
69
227
181
231
244
268
182
160
171
272
201
279
277
362
276
278
276
302
334
162
196
66
222
190
Secondary lon(s)
109,331,79,333
167,43
167,195
51,87,224,257,153
191,319,334,197,321
253,125
285,202,139,253
253,125
139,253,111,141
97,308,125,292
97,153,125,121
145,107,239,121,159
125,93,109,217
367,107,165,198
253,125
260,265
241,239,120
104,252,223,209
77,80,107
41,39
228,152,114,274,212
180,223,152
266,268,140,195
201,229
252,253,126,134,113
184,367,121,379
132,93,104,105
377,375,77,155,379
237,274,270,239,235
137,119,217,219,199
280,277,250
310,174,147,242
226,210,364,97,109
138,227
139,279
138,277
151,150,300
334,335,333
135,105,77
198,209,211,406,408
263,220
260,292
224,260
8270D - 31
Revision 4
January 1998
-------
TABLE 1
(continued)
Compound
Aroclor 1232
Aroclor 1242
Aroclor 1248
Aroclor 1254
Aroclor 1260
a-BHC
P-BHC
6-BHC
Y-BHC (Lindane)
4,4'-DDD
4,4'-DDE
4,4'-DDT
Dieldrin
1 ,2-Diphenylhydrazine
Endosulfan I
Endosulfan II
Endosulfan sulfate
Endrin
Endrin aldehyde
Endrin ketone
2-Fluorobiphenyl (surr)
2-Fluorophenol (surr)
Heptachlor
Heptachlor epoxide
Nitrobenzene-d5 (surr)
N-Nitrosodimethylamine
Phenol-d6 (surr)
Terphenyl-d14 (surr)
2,4,6-Tribromophenol (surr)
Toxaphene
Retention Primary
Time (min) Ion
190
222
292
292
360
183
181
183
183
235
246
235
79
77
195
337
272
263
67
317
172
112
100
353
82
42
99
244
330
159
Secondary lon(s)
224,260
256,292
362,326
362,326
362,394
181,109
183,109
181,109
181,109
237,165
248,176
237,165
263,279
105,182
339,341
339,341
387,422
82,81
345,250
67,319
171
64
272,274
355,351
128,54
74,44
42.71
122,212
332,141
231,233
IS = internal standard
surr = surrogate
Estimated retention times
"Substitute for the non-specific mixture, tricresyl phosphate
8270D - 32
Revision 4
January 1998
-------
TABLE 2
ESTIMATED QUANTITATION LIMITS (EQLs) FOR SEMIVOLATILE ORGANICS
Estimated Quantitation Limits8
Compound
Acenaphthene
Acenaphthylene
Acetophenone
2-Acetylaminofluorene
1 -Acetyl-2-thiourea
2-Aminoanthraquinone
Aminoazobenzene
4-Aminobiphenyl
Anilazine
o-Anisidine
Anthracene
Aramite
Azinphos-methyl
Barban
Benz(a)anthracene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzoic acid
Benzo(g,h,i)perylene
Benzo(a)pyrene
p-Benzoquinone
Benzyl alcohol
Bis(2-chloroethoxy)methane
Bis(2-chloroethyl) ether
Bis(2-chloroisopropyl) ether
4-Bromophenyl phenyl ether
Bromoxynil
Butyl benzyl phthalate
Captafol
Captan
Carbaryl
Carbofuran
Carbophenothion
Chlorfenvinphos
4-Chloroaniline
Chlorobenzilate
5-Chloro-2-methylaniline
Ground water
(M9/L)
10
10
10
20
1000
20
10
20
100
10
10
20
100
200
10
10
10
50
10
10
10
20
10
10
10
10
10
10
20
50
10
10
10
20
20
10
10
8270D - 33
Low Soil/Sediment"
(ug/kg)
660
660
ND
ND
ND
ND
ND
ND
ND
ND
660
ND
ND
ND
660
660
660
3300
660
660
ND
1300
660
660
660
660
ND
660
ND
ND
ND
ND
ND
ND
1300
ND
ND
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits8
Compound
4-Chloro-3-methylphenol
3-(Chloromethyl)pyridinehydrochloride
2-Chloronaphthalene
2-Chlorophenol
4-Chlorophenyl phenyl ether
Chrysene
Coumaphos
p-Cresidine
Crotoxyphos
2-Cyclohexyl-4,6-dinitrophenol
Demeton-O
Demeton-S
Diallate (cis or trans)
Diallate (trans or cis)
2,4-Diaminotoluene
Dibenz(a,j)acridine
Dibenz(a,h)anthracene
Dibenzofuran
Dibenzo(a,e)pyrene
Di-n-butyl phthalate
Dichlone
1 ,2-Dichlorobenzene
1 ,3-Dichlorobenzene
1 ,4-Dichlorobenzene
3,3'-Dichlorobenzidine
2,4-Dichlorophenol
2,6-Dichlorophenol
Dichlorovos
Dicrotophos
Diethyl phthalate
Diethylstilbestrol
Diethyl sulfate
Dimethoate
3,3'-Dimethoxybenzidine
Dimethylaminoazobenzene
7, 1 2-Dimethylbenz(a)anthracene
3,3'-Dimethylbenzidine
2,4-Dimethylphenol
Ground water
(ug/L)
20
100
10
10
10
10
40
10
20
100
10
10
10
10
20
10
10
10
10
10
NA
10
10
10
20
10
10
10
10
10
20
100
20
100
10
10
10
10
8270D - 34
Low Soil/Sediment"
(ug/kg)
1300
ND
660
660
660
660
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
660
660
ND
ND
ND
660
660
660
1300
660
ND
ND
ND
660
ND
ND
ND
ND
ND
ND
ND
660
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits8
Compound
Dimethyl phthalate
1 ,2-Dinitrobenzene
1 ,3-Dinitrobenzene
1 ,4-Dinitrobenzene
4,6-Dinitro-2-methylphenol
2,4-Dinitrophenol
2,4-Dinitrotoluene
2,6-Dinitrotoluene
Dinocap
Dinoseb
5,5-Diphenylhydantoin
Di-n-octyl phthalate
Oisulfoton
EPN
Ethion
Ethyl carbamate
Bis(2-ethylhexyl) phthalate
Ethyl methanesulfonate
Famphur
Fensulfothion
Fenthion
Fluchloralin
Fluoranthene
Fluorene
Hexachlorobenzene
Hexachlorobutadiene
Hexachlorocyclopentadiene
Hexachloroethane
Hexachlorophene
Hexachloropropene
Hexamethylphosphoramide
lndeno(1 ,2,3-cd)pyrene
Isodrin
Isophorone
Isosafrole
Kepone
Leptophos
Malathion
Ground water
(ug/L)
10
40
20
40
50
50
10
10
100
20
20
10
10
10
10
50
10
20
20
40
10
20
10
10
10
10
10
10
50
10
20
10
20
10
10
20
10
50
8270D - 35
Low Soil/Sediment"
(ug/kg)
660
ND
ND
ND
3300
3300
660
660
ND
ND
ND
660
ND
ND
ND
ND
660
ND
ND
ND
ND
ND
660
660
660
660
660
660
ND
ND
ND
660
ND
660
ND
ND
ND
ND
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits8
Compound
Mestranol
Methapyrilene
Methoxychlor
3-Methylcholanthrene
Methyl methanesulfonate
2-Methylnaphthalene
Methyl parathion
2-Methylphenol
3-Methylphenol
4-Methylphenol
Mevinphos
Mexacarbate
Mirex
Monocrotophos
Naled
Naphthalene
1 ,4-Naphthoquinone
1-Naphthylamine
2-Naphthylamine
Nicotine
5-Nitroacenaphthene
2-Nitroaniline
3-Nitroaniline
4-Nitroaniline
5-Nitro-o-anisidine
Nitrobenzene
4-Nitrobiphenyl
Nitrofen
2-Nitrophenol
4-Nitrophenol
5-Nitro-o-toluidine
4-Nitroquinoline-1 -oxide
N-Nitrosodi-n-butylamine
N-Nitrosodiethylamine
N-Nitrosodiphenylamine
N-Nitroso-di-n-propylamine
N-Nitrosopiperidine
N-Nitrosopyrrolidine
Ground water
(M9/L)
20
100
10
10
10
10
10
10
10
10
10
20
10
40
20
10
10
10
10
20
10
50
50
20
10
10
10
20
10
50
10
40
10
20
10
10
20
40
8270D - 36
Low Soil/Sediment11
(ug/kg)
ND
ND
ND
ND
ND
660
ND
660
ND
660
ND
ND
ND
ND
ND
660
ND
ND
ND
ND
ND
3300
3300
ND
ND
660
ND
ND
660
3300
ND
ND
ND
ND
660
660
ND
ND
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits8
Compound
Octamethyl pyrophosphoramide
4,4'-Oxydianiline
Parathion
Pentachlorobenzene
Pentachloronitrobenzene
Pentachlorophenol
Phenacetin
Phenanthrene
Phenobarbital
Phenol
1 ,4-Phenylenediamine
Phorate
Phosalone
Phosmet
Phosphamidon
Phthalic anhydride
2-Picoline
Piperonyl sulfoxide
Pronamide
Propylthiouracil
Pyrene
Resorcinol
Safrole
Strychnine
Sulfallate
Terbufos
1 ,2,4,5-Tetrachlorobenzene
2,3,4,6-Tetrachlorophenol
Tetrachlorvinphos
Tetraethyl pyrophosphate
Thionazine
Thiophenol (Benzenethiol)
o-Toluidine
1 ,2,4-Trichlorobenzene
2,4,5-Trichlorophenol
2,4,6-Trichlorophenol
Trifluralin
2,4,5-Trimethylaniline
Ground water
(ug/L)
200
20
10
10
20
50
20
10
10
10
10
10
100
40
100
100
ND
100
10
100
10
100
10
40
10
20
10
10
20
40
20
20
10
10
10
10
10
10
8270D - 37
Low Soil/Sediment"
(ug/kg)
ND
ND
ND
ND
ND
3300
ND
660
ND
660
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
660
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
660
660
660
ND
ND
Revision 4
January 1998
-------
TABLE 2
(continued)
Estimated Quantitation Limits3
Compound
Trimethyl phosphate
1 ,3,5-Trinitrobenzene
Tris(2,3-dibromopropyl) phosphate
Tri-p-tolyl phosphate(h)
Ground water
(ug/D
10
10
200
10
Low Soil/Sediment"
(MQ/kg)
ND
ND
ND
ND
a 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 Factor0
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 CRITERIA8 b
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
8 Data taken from Reference 3.
b 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 4-Chloro-3-methylphenol
1,4-Dichlorobenzene 2,4-Dichlorophenol
Hexachlorobutadiene 2-Nitrophenol
Diphenylamine Phenol
Di-n-octyl phthalate Pentachlorophenol
Fluoranthene 2,4,6-Trichlorophenol
Benzo(a)pyrene
8270D - 39 Revision 4
January 1998
-------
TABLE 5
SEMIVOLATILE INTERNAL STANDARDS WITH CORRESPONDING ANALYTES
ASSIGNED FOR QUANTITATION
,4-Dichlorobenzene-d4
Naphthalene-d8
Acenaphthene-d
10
Aniline
Benzyl alcohol
Bis(2-chloroethyl) ether
Bis(2-chloroisopropyl) ether
2-Chlorophenol
1,3-Dichlorobenzene
1,4-Dichlorobenzene
1,2-Dichlorobenzene
Ethyl methanesulfonate
2-Fluorophenol (surr)
Hexachloroethane
Methyl methanesulfonate
2-Methylphenol
4-Methylphenol
N-Nitrosodimethylamine
N-Nitroso-di-n-propylamine
Phenol
Phenol-d6 (surr)
2-Picoline
Acetophenone
Benzoic acid
Bis(2-chloroethoxy)methane
4-Chloroaniline
4-Chloro-3-methylphenol
2,4-Dichlorophenol
2,6-Dichlorophenol
a,a-Dimethyl-
phenethylamine
2,4-Dimethylphenol
Hexachlorobutadiene
Isophorone
2-Methylnaphthalene
Naphthalene
Nitrobenzene
Nitrobenzene-d8 (surr)
2-Nitrophenol
N-Nitrosodi-n-butylamine
N-Nitrosopiperidine
1,2,4-Trichlorobenzene
Acenaphthene
Acenaphthylene
1-Chloronaphthalene
2-Chloronaphthalene
4-Chlorophenyl phenyl ether
Dibenzofuran
Diethyl phthalate
Dimethyl phthalate
2,4-Dinitrophenol
2,4-Dinitrotoluene
2,6-Dinitrotoluene
Fluorene
2-Fluorobiphenyl (surr)
Hexachlorocyclopentadiene
1-Naphthylamine
2-Naphthylamine
2-Nitroaniline
3-Nitroaniline
4-Nitroaniline
4-Nitrophenol
Pentachlorobenzene
1,2,4,5-Tetrachlorobenzene
2,3,4,6-Tetrachlorophenol
2,4,6-Tribromophenol (surr)
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'-Dichlorobenzidine
p-Dimethyl aminoazobenzene
Pyrene
Terphenyl-d14 (surr)
7,12-Dimethylbenz(a)
anthracene
Di-n-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(aj)acridine
Dibenz(a,h)anthracene
(surr) = surrogate
8270D - 41
Revision 4
January 1998
-------
TABLE 6
MULTILABORATORY PERFORMANCE DATA8
Compound
Acenaphthene
Acenaphthylene
Aldrin
Anthracene
Benz(a)anthracene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(a)pyrene
Benzo(g , h, i)perylene
Benzyl butyl phthalate
p-BHC
6-BHC
Bis(2-chloroethyl) ether
Bis(2-chloroethoxy)methane
Bis(2-chloroisopropyl) ether
Bis(2-ethylhexyl) phthalate
4-Bromophenyl phenyl ether
2-Chloronaphthalene
4-Chlorophenyl phenyl ether
Chrysene
4,4'-DDD
4,4'-DDE
4,4'-DDT
Dibenzo(a,h)anthracene
Di-n-butyl phthalate
1 ,2-Dichlorobenzene
1 ,3-Dichlorobenzene
1 ,4-Dichlorobenzene
3,3'-Dichlorobenzidine
Dieldrin
Diethyl phthalate
Dimethyl phthalate
2,4-Dinitrotoluene
2,6-Dinitrotoluene
Di-n-octyl phthalate
Endosulfan sulfate
Endrin aldehyde
Fluoranthene
Test cone. Limit for
(ug/L) s (Mg/L)
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
27.6
40.2
39.0
32.0
27.6
38.8
32.3
39.0
58.9
23.4
31.5
21.6
55.0
34.5
46.3
41.1
23.0
13.0
33.4
48.3
31.0
32.0
61.6
70.0
16.7
30.9
41.7
32.1
71.4
30.7
26.5
23.2
21.8
29.6
31.4
16.7
32.5
32.8
8270D - 42
Range for x
(ug/L)
60.1-132.3
53.5-126.0
7.2-152.2
43.4-118.0
41.8-133.0
42.0-140.4
25.2-145.7
31.7-148.0
D-195.0
D-139.9
41.5-130.6
D-100.0
42.9-126.0
49.2-164.7
62.8-138.6
28.9-136.8
64.9-114.4
64.5-113.5
38.4-144.7
44.1-139.9
D-134.5
19.2-119.7
D-170.6
D-199.7
8.4-111.0
48.6-112.0
16.7-153.9
37.3-105.7
8.2-212.5
44.3-119.3
D-100.0
D-100.0
47.5-126.9
68.1-136.7
18.6-131.8
D-103.5
D-188.8
42.9-121.3
Range
P. Ps(%)
47-145
33-145
D-166
27-133
33-143
24-159
11-162
17-163
D-219
D-152
24-149
D-110
12-158
33-184
36-166
8-158
53-127
60-118
25-158
17-168
D-145
4-136
D-203
D-227
1-118
32-129
D-172
20-124
D-262
29-136
D-114
D-112
39-139
50-158
4-146
D-107
D-209
26-137
Revision 4
January 1998
-------
TABLE 6
(continued)
Test cone. Limit for
Compound
Fluorene
Heptachlor
Heptachlor epoxide
Hexachlorobenzene
Hexachlorobutadiene
Hexachloroethane
lndeno(1 ,2,3-cd)pyrene
Isophorone
Naphthalene
Nitrobenzene
N-Nitrosodi-n-propylamine
Aroclor 1260
Phenanthrene
Pyrene
1 ,2,4-Trichlorobenzene
4-Chloro-3-methylphenol
2-Chlorophenol
2,4-Chlorophenol
2,4-Dimethylphenol
2,4-Dinitrophenol
2-Methyl-4,6-dinitrophenol
2-Nitrophenol
4-Nitrophenol
Pentachlorophenol
Phenol
2,4,6-Trichlorophenol
s = Standard deviation of four
~x. = Average recovery for four i
(M9/L)
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
recovery
recovery
s (ug/L)
20.7
37.2
54.7
24.9
26.3
24.5
44.6
63.3
30.1
39.3
55.4
54.2
20.6
25.2
28.1
37.2
28.7
26.4
26.1
49.8
93.2
35.2
47.2
48.9
22.6
31.7
measurements, in
measurements, in
Range for x
(ug/L)
71.6-108.4
D-172.2
70.9-109.4
7.8-141.5
37.8-102.2
55.2-100.0
D-150.9
46.6-180.2
35.6-119.6
54.3-157.6
13.6-197.9
19.3-121.0
65.2-108.7
69.6-100.0
57.3-129.2
40.8-127.9
36.2-120.4
52.5-121.7
41.8-109.0
D-172.9
53.0-100.0
45.0-166.7
13.0-106.5
38.1-151.8
16.6-100.0
52.4-129.2
M9/L
ug/L
Range
P. P«(%)
59-121
D-192
26.155
D-152
24-116
40-113
D-171
21-196
21-133
35-180
D-230
D-164
54-120
52-115
44-142
22-147
23-134
39-135
32-119
D-191
D-181
29-182
D-132
14-176
5-112
37-144
p, ps = Measured percent recovery
D = Detected; result must be greater than zero
a 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 capillary 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
January 1998
-------
TABLE 7
METHOD ACCURACY AND PRECISION AS FUNCTIONS OF CONCENTRATION8
Compound
Acenaphthene
Acenaphthylene
Aldrin
Anthracene
Benz(a)anthracene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(a)pyrene
Benzo(g , h , i) perylene
Benzyl butyl phthalate
p-BHC
6-BHC
Bis(2-chloroethyl) ether
Bis(2-chloroethoxy)methane
Bis(2-chloroisopropyl) ether
Bis(2-ethylhexyl) phthalate
4-Bromophenyl phenyl ether
2-Chloronaphthalene
4-Chlorophenyl phenyl ether
Chrysene
4,4'-DDD
4,4'-DDE
4,4'-DDT
Dibenzo(a,h)anthracene
Di-n-butyl phthalate
1 ,2-Dichlorobenzene
1 ,3-Dichlorobenzene
1 ,4-Dichlorobenzene
3,3'-Dichlorobenzidine
Dieldrin
Diethyl phthalate
Dimethyl phthalate
2,4-Dinitrotoluene
2,6-Dinitrotoluene
Di-n-octyl phthalate
Endosulfan sulfate
Endrin aldehyde
Fluoranthene
Fluorene
Accuracy, as
recovery, x' (ug/L)
0.96C+0.19
0.89C+0.74
0.78C+1.66
0.80C+0.68
0.88C-0.60
0.93C-1.80
0.87C-1.56
0.90C-0.13
0.98C-0.86
0.66C-1.68
0.87C-0.94
0.29C-1.09
0.86C-1.54
1.12C-5.04
1.03C-2.31
0.84C-1.18
0.91C-1.34
0.89C+0.01
0.91C+0.53
0.93C-1.00
0.56C-0.40
0.70C-0.54
0.79C-3.28
0.88C+4.72
0.59C+0.71
0.80C+0.28
0.86C-0.70
0.73C-1.47
1.23C-12.65
0.82C-0.16
0.43C+1.00
0.20C+1.03
0.92C-4.81
1.06C-3.60
0.76C-0.79
0.39C+0.41
0.76C-3.86
0.81C+1.10
0.90C-0.00
8270D -
Single analyst
precision, sr' (ug/L)
0.157-0.12
0.247-1.06
0.277-1.28
0.217-0.32
0.157+0.93
0.227+0.43
0.197+1.03
0.227+0.48
0.29l(+2.40
0.187+0.94
0.20X-0.58
0.34X+0.86
0.353(-0.99
0.16X+1.34
0.247+0.28
0.267+0.73
0.137+0.66
0.077+0.52
0.207-0.94
0.287+0.13
0.297-0.32
0.267-1.17
0.427+0.19
0.307+8.51
0.137+1.16
0.207+0.47
0.257+0.68
0.247+0.23
0.287+7.33
0.207-0.16
0.287+1.44
0.547+0.19
0.127+1.06
0.147+1.26
0.217+1.19
0.127+2.47
0.187+3.91
0.227-0.73
0.127+0.26
44
Overall precision,
S' (ug/L)
0.217-0.67
0.267-0.54
0.437+1.13
0.277-0.64
0.267-0.21
0.297+0.96
0.357+0.40
0.327+1.35
0.517-0.44
0.537+0.92
0.307+1.94
0.937-0.17
0.357+0.10
0.267+2.01
0.257+1.04
0.367+0.67
0.167+0.66
0.137+0.34
0.307-0.46
0.337-0.09
0.667-0.96
0.397-1.04
0.657-0.58
0.597+0.25
0.397+0.60
0.247+0.39
0.417+0.11
0.297+0.36
0.477+3.45
0.267-0.07
0.527+0.22
1.057-0.92
0.217+1.50
0.197+0.35
0.377+1.19
0.637-1.03
0.737-0.62
0.287-0.60
0.137+0.61
Revision 4
January 1998
-------
TABLE 7
(continued)
Compound
Accuracy, as
recovery, x1 (ug/L)
Single analyst
precision, sr' (ug/L)
Overall precision,
S'(ug/L)
Heptachlor
Heptachlor epoxide
Hexachlorobenzene
Hexachlorobutadiene
Hexachloroethane
lndeno(1 ,2,3-cd)pyrene
Isophorone
Naphthalene
Nitrobenzene
N-Nitrosodi-n-propylamine
Aroclor 1260
Phenanthrene
Pyrene
1 ,2,4-Trichlorobenzene
4-Chloro-3-methylphenol
2-Chlorophenol
2,4-Dichlorophenol
2,4-Dimethylphenol
2,4-Dinitrophenol
2-Methyl-4,6-dinitrophenol
2-Nitrophenol
4-Nitrophenol
Pentachlorophenol
Phenol
2,4,6-Trichlorophenol
0.87C-2.97
0.92C-1.87
0.74C+0.66
0.71C-1.01
0.73C-0.83
0.78C-3.10
1.12C+1.41
0.76C+1.58
1.09C-3.05
1.12C-6.22
0.81C-10.86
0.87C+0.06
0.84C-0.16
0.94C-0.79
0.84C+0.35
0.78C+0.29
0.87C-0.13
0.71C+4.41
0.81C-18.04
1.04C-28.04
0.07C-1.15
0.61C-1.22
0.93C+1.99
0.43C+1.26
0.91C-0.18
0.247-0.56
0.337-0.46
0.187-0.10
0.197+0.92
0.177+0.67
0.297+1.46
0.277+0.77
0.217-0.41
0.197+0.92
0.277+0.68
0.357+3.61
0.127+0.57
0.167+0.06
0.157+0.85
0.237+0.75
0.187+1.46
0.157+1.25
0.167+1.21
0.387+2.36
0.107+42.29
0.167+1.94
0.387+2.57
0.247+3.03
0.267+0.73
0.167+2.22
0.507-0.23
0.287+0.64
0.437-0.52
0.267+0.49
0.177+0.80
0.507-0.44
0.337+0.26
0.307-0.68
0.277+0.21
0.447+0.47
0.437+1.82
0.157+0.25
0.157+0.31
0.217+0.39
0.297+1.31
0.287+0.97
0.217+1.28
0.227+1.31
0.427+26.29
0.267+23.10
0.277+2.60
0.447+3.24
0.307+4.33
0.357+0.58
0.227+1.81
x' = Expected recovery for one or more measurements of a sample containing a concentration of
C, in ug/L.
sr' = Expected single analyst standard deviation of measurements at an average concentration of
7, in ug/L.
S1 = Expected interlaboratory standard deviation of measurements at an average concentration
found of 7, in ug/L.
C = True value for the concentration, in ug/L
7 = Average recovery found for measurements of samples containing a concentration of C, in
Mfl/L.
a 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
Revision 4
January 1998
-------
TABLE 8
EXTRACTION EFFICIENCY AND AQUEOUS STABILITY RESULTS
Compound
Percent Recovery, Day 0 Percent Recovery, Day 7
Mean RSD Mean RSD
3-Amino-9-ethylcarbazole 80
4-Chloro-1,2-phenylenediamine 91
4-Chloro-1,3-phenylenediamine 84
1,2-Dibromo-3-chloropropane 97
Dinoseb 99
Parathion 100
4,4'-Methylenebis(N,N-dimethylaniline) 108
5-Nitro-o-toluidine 99
2-Picoline 80
Tetraethyl dithiopyrophosphate 92
8
1
3
2
3
2
4
10
4
7
73
108
70
98
97
103
90
93
83
70
3
4
3
5
6
4
4
4
4
1
Data taken from Reference 6.
8270D - 46
Revision 4
January 1998
-------
TABLE 9
MEAN PERCENT RECOVERIES AND PERCENT RSD VALUES FOR SEMIVOLATILE ORGANICS
FROM SPIKED CLAY SOIL AND TOPSOIL BY AUTOMATED SOXHLET EXTRACTION
(METHOD 3541) WITH HEXANE-ACETONE (1:1)a
Clay Soil
Compound
1 ,3-Dichlorobenzene
1 ,2-Dichlorobenzene
Nitrobenzene
Benzal chloride
Benzotrichloride
4-Chloro-2-nitrotoluene
Hexachlorocyclopentadiene
2,4-Dichloronitrobenzene
3,4-Dichloronitrobenzene
Pentachlorobenzene
2,3,4,5-Tetrachloronitrobenzene
Benefin
alpha-BHC
Hexachlorobenzene
delta-BHC
Heptachlor
Aldrin
Isopropalin
Heptachlor epoxide
trans-Chlordane
Endosulfan I
Dieldrin
2,5-Dichlorophenyl-4-nitrophenyl ether
Endrin
Endosulfan II
p.p'-DDT
213,6-Trichlorophenyl-4'-nitrophenyl ether
2,3,4-Trichlorophenyl-4'-nitrophenyl ether
Mirex
Mean
Recovery
0
0
0
0
0
0
4.1
35.2
34.9
13.7
55.9
62.6
58.2
26.9
95.8
46.9
97.7
102
90.4
90.1
96.3
129
110
102
104
134
110
112
104
RSD
—
--
—
—
—
—
15
7.6
15
7.3
6.7
4.8
7.3
13
4.6
9.2
12
4.3
4.4
4.5
4.4
4.7
4.1
4.5
4.1
2.1
4.8
4.4
5.3
Topsoil
Mean
Recovery
0
0
0
0
0
0
7.8
21.2
20.4
14.8
50.4
62.7
54.8
25.1
99.2
49.1
102
105
93.6
95.0
101
104
112
106
105
111
110
112
108
RSD
—
—
—
—
—
—
23
15
11
13
6.0
2.9
4.8
5.7
1.3
6.3
7.4
2.3
2.4
2.3
2.2
1.9
2.1
3.7
0.4
2.0
2.8
3.3
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-
Trichlorophenyl-4-nitrophenyl ether, and 2,3,4-Trichlorophenyl-4-nitrophenyl ether at 1500 ng/g,
Nitrobenzene at 2000 ng/g, and 1,3-Dichlorobenzene and 1,2-Dichlorobenzene at 5000 ng/g.
8270D - 47
Revision 4
January 1998
-------
TABLE 10
SINGLE LABORATORY ACCURACY AND PRECISION DATA FOR THE EXTRACTION
OF SEMIVOLATILE ORGANICS FROM SPIKED CLAY BY
AUTOMATED SOXHLET (METHOD 3541)a
Compound
Phenol
Bis(2-chloroethyl)ether
2-Chlorophenol
Benzyl alcohol
2-Methylphenol
Bis(2-chloroisopropyl)ether
4-Methylphenol
N-Nitroso-di-n-propylamine
Nitrobenzene
Isophorone
2-Nitrophenol
2,4-Dimethylphenol
Benzole acid
Bis(2-chloroethoxy)methane
2,4-Dichlorophenol
1 ,2,4-Trichlorobenzene
Naphthalene
4-Chloroaniline
4-Chloro-3-methylphenol
2-Methylnaphthalene
Hexachlorocyclopentadiene
2,4,6-Trichlorophenol
2,4,5-Trichlorophenol
2-Chloronaphthalene
2-Nitroaniline
Dimethyl phthalate
Acenaphthylene
3-Nitroaniline
Acenaphthene
2,4-Dinitrophenol
4-Nitrophenol
Dibenzofuran
2,4-Dinitrotoluene
2,6-Dinitrotoluene
Diethyl phthalate
4-Chlorophenyl-phenyl ether
Fluorene
Mean Recovery
47.8
25.4
42.7
55.9
17.6
15.0
23.4
41.4
28.2
56.1
36.0
50.1
40.6
44.1
55.6
18.1
26.2
55.7
65.1
47.0
19.3
70.2
26.8
61.2
73.8
74.6
71.6
77.6
79.2
91.9
62.9
82.1
84.2
68.3
74.9
67.2
82.1
8270D - 48
RSD
5.6
13
4.3
7.2
6.6
15
6.7
6.2
7.7
4.2
6.5
5.7
7.7
3.0
4.6
31
15
12
5.1
8.6
19
6.3
2.9
6.0
6.0
5.2
5.7
5.3
4.0
8.9
16
5.9
5.4
5.8
5.4
3.2
3.4
Revision 4
January 1998
-------
TABLE 10
(continued)
Compound
4-Nitroaniline
4,6-Dinitro-2-methylphenol
N-Nitrosodiphenylamine
4-Bromophenyl-phenyl ether
Hexachlorobenzene
Pentachlorophenol
Phenanthrene
Anthracene
Di-n-butyl phthalate
Fluoranthene
Pyrene
Butyl benzyl phthalate
3,3'-Dichlorobenzidine
Benzo(a)anthracene
Bis(2-ethylhexyl) phthalate
Chrysene
Di-n-octyl phthalate
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(a)pyrene
lndeno(1 ,2,3-cd)pyrene
Dibenzo(a,h)anthracene
Benzo(g,h,i)perylene
1 ,2-Dichlorobenzene
1 ,3-Dichlorobenzene
1 ,4-Dichlorobenzene
Hexachloroethane
Hexachlorobutadiene
Mean Recovery
79.0
63.4
77.0
62.4
72.6
62.7
83.9
96.3
78.3
87.7
102
66.3
25.2
73.4
77.2
76.2
83.1
82.7
71.7
71.7
72.2
66.7
63.9
0
0
0
0
0
RSD
7.9
6.8
3.4
3.0
3.7
6.1
5.4
3.9
40
6.9
0.8
5.2
11
3.8
4.8
4.4
4.8
5.0
4.1
4.1
4.3
6.3
8.0
—
—
—
—
~~
a 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 Revision 4
January 1998
-------
TABLE 11
PRECISION AND BIAS VALUES FOR METHOD 35421
Compound
2-Fluorophenol
Phenol-d5
Nitrobenzene-d5
2-Fluorobiphenyl
2,4,6-Tribromophenol
Terphenyl-d14
Mean Recovery
74.6
77.8
65.6
75.9
67.0
78.6
Standard Deviation
28.6
27.7
32.5
30.3
34.0
32.4
% RSD
38.3
35.6
49.6
39.9
50.7
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
January 1998
-------
TABLE 12
ACCELERATED SOLVENT EXTRACTION (METHOD 3545) RECOVERY VALUES
AS PERCENT OF SOXTEC™
Compound
Phenol
Bis(2-chloroethyl) ether
2-Chlorophenol
1,3-Dichlorobenzene
1 ,4-Dichlorobenzene
1,2-Dichlorobenzene
2-Methylphenol
Bis(2-chloroisopropyl)ether
o-Toluidine
N-Nitroso-di-n-propylamine
Hexachloroethane
Nitrobenzene
Isophorone
2,4-Dimethylphenol
2-Nitrophenol
Bis(chloroethoxy)methane
2,4-Dichlorophenol
1 ,2,4-Trichlorobenzene
Naphthalene
4-Chloroaniline
Hexachlorobutadiene
4-Chloro-3-methylphenol
2-Methylnaphthalene
Hexachlorocyclopentadiene
2,4,6-Trichlorophenol
2,4,5-Trichlorophenol
2-Chloronaphthalene
2-Nitroaniline
2,6-Dinitrotoluene
Acenaphthytene
3-Nitroaniline
Acenaphthene
4-Nitrophenol
2,4-Dinitrotoluene
Low
93.3
102.1
100.8
127.7
127.9
116.8
98.9
109.4
100.0
103.0
97.1
104.8
100.0
100.0
80.7
94.4
88.9
98.0
101.7
100.0
101.1
90.4
93.2
100.0
94.6
84.4
100.0
90.0
83.1
104.9
*224.0
102.1
0.0
73.9
Clav
Mid
78.7
85.1
82.6
129.7
127.0
115.8
82.1
71.5
89.7
79.1
125.1
82.4
86.4
104.5
80.5
80.6
87.8
97.8
97.2
*150.2
98.7
80.2
89.9
100.0
90.0
91.9
91.3
83.4
90.6
95.9
115.6
92.6
93.2
91.9
High
135.9
109.1
115.0
110.0
110.5
101.3
119.7
108.0
117.2
107.7
111.0
106.6
98.2
140.0
107.9
94.7
111.4
98.8
123.6
•162.4
102.2
114.7
94.6
0.0
112.0
109.6
93.6
97.4
91.6
100.5
97.6
97.6
121.5
100.2
Low
73.9
96.0
93.8
•364.2
*365.9
•159.2
87.6
81.8
100.0
83.9
•245.4
86.8
87.1
100.0
91.4
86.5
85.9
123.0
113.2
100.0
124.1
79.0
104.1
100.0
84.2
96.1
97.6
71.3
86.4
99.0
100.0
97.2
18.1
84.7
Loam
Mid
82.8
88.0
88.9
129.9
127.8
113.4
89.4
81.0
•152.5
88.1
117.1
84.6
87.5
114.4
86.7
84.4
87.6
93.7
102.9
125.5
90.3
85.2
92.2
100.0
91.2
80.7
93.4
88.4
90.6
97.9
111.8
96.9
87.1
93.8
High
124.6
103.6
111.1
119.0
116.4
105.5
111.0
88.6
120.3
96.2
128.1
101.7
109.7
123.1
103.2
99.6
103.5
94.5
129.5
•263.6
98.0
109.8
105.9
6.8
103.6
103.6
98.3
89.9
90.3
108.8
107.8
104.4
116.6
98.9
Low
108.8
122.3
115.0
•241.3
•309.6
•189.3
133.2
118.1
100.0
109.9
•566.7
119.7
135.5
100.0
122.1
130.6
123.3
137.0
•174.5
100.0
134.9
131.6
146.2
100.0
101.6
108.9
106.8
112.1
104.3
118.5
0.0
114.2
69.1
100.9
Sand
Mid
130.6
119.9
115.3
•163.7
•164.1
134.0
128.0
148.3
•199.5
123.3
147.9
122.1
118.4
•180.6
107.1
110.7
107.0
99.4
114.0
•250.8
96.1
116.2
99.1
100.0
95.9
83.9
93.0
113.3
84.7
97.8
111.7
92.0
90.5
84.3
8270D - 51
High
89.7
90.8
91.9
107.1
105.8
100.4
92.1
94.8
102.7
91.4
103.7
93.3
92.7
96.3
87.0
93.2
92.1
95.3
89.8
114.9
96.8
90.1
93.3
•238.3
89.8
87.9
92.0
87.7
90.9
92.0
99.0
89.0
84.5
87.3
Mean
Rec.
102.0
101.9
101.6
120.6
119.2
112.5
104.7
100.2
110.3
98.1
118.6
100.2
101.7
109.8
96.3
97.2
98.6
104.2
106.1
108.1
104.7
99.7
102.1
75.8
95.9
94.1
96.2
92.6
90.3
101.7
92.9
98.4
75.6
90.7
Revision 4
January 1998
-------
TABLE 12
(continued)
Compound
Dibenzofuran
4-Chlorophenyl phenyl
ether
Fluorene
4-Nitroaniline
N-Nitrosodiphenylamine
4-Bromophenyl phenyl
ether
Hexachlorobenzene
Pentachlorophenol
Phenanthrene
Anthracene
Carbazole
Fluoranthene
Pyrene
3,3'-Dichlorobenzidine
Benzo(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(a)pyrene
lndeno(1 ,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
Mean
Low
89.5
83.0
85.2
77.8
82.6
85.6
95.4
68.2
92.1
101.6
94.4
109.9
106.5
100.0
98.1
100.0
106.6
102.4
107.9
95.1
85.0
98.0
95.1
Clav
Mid
91.7
94.5
94.9
114.8
96.7
92.9
91.7
85.9
93.7
95.0
99.3
101.4
105.8
*492.3
107.0
108.5
109.9
105.2
105.5
105.7
102.6
0.0
94.3
High
109.3
98.7
89.2
94.5
93.8
92.8
92.3
107.7
93.3
93.5
96.6
94.3
107.6
131.4
98.4
100.2
75.6
88.4
80.8
93.8
82.0
81.2
101.0
Low
98.5
95.
102.
129.
92
91
95
,7
,0
,6
.9
.1
.4
53.2
100.0
92.5
105.5
111.6
116.7
100.0
119.3
116.8
121.7
125.5
122
126
118
0
95
.3
.0
.8
.0
.5
Loam
Mid
92.2
94.3
95.5
103.6
93.4
107.6
93.6
89.8
97.8
101.8
96.7
96.6
90.7
*217.6
98.6
93.0
100.7
99.4
97.7
105.2
100.7
33.6
96.5
High
111.4
94.2
93.8
95.4
116.4
89.4
83.7
88.1
113.3
118.4
111.4
109.6
127.5
"167.6
104.0
117.0
93.9
95.1
104.6
90.4
91.9
78.6
104.1
Low
113.8
111.4
121.3
*154.1
97.5
118.0
106.8
96.6
124.4
123.0
115.7
123.2
103.4
100.0
105.0
106.7
106.9
144.7
101.7
133.6
142.3
128.7
113.0
Sand
Mid
92.7
87.7
85.7
89.3
110.9
97.5
94.3
59.8
101.0
94.5
83.2
85.4
95.5
*748.8
93.4
93.6
81.9
89.2
86.2
82.6
71.0
83.0
100.9
High
90.4
90.3
90.9
87.5
86.7
87.1
90.0
81.3
89.9
90.6
88.9
92.7
93.2
100.0
89.3
90.2
93.6
78.1
92.0
91.9
93.1
94.2
92.5
Mean
Rec.
98.8
94.4
95.4
99.1
96.8
95.8
93.7
81.2
100.6
101.2
99.1
102.7
105.2
116.5
101.5
102.9
99.0
103.1
99.9
102.7
98.6
66.4
' Values greater than 150% were not used to determine the averages, but the 0% values were used.
8270D - 52
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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
Naphthalene
Acenaphthylene
Acenaphthene
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(a)pyrene
lndeno(1 ,2,3-cd)pyrene
Benzo(g,h,i)perylene
Dibenz(a,h)anthracene
Certified Value
(mg/kg)
(27.9)b
(0.8)
(0.2)
(15.3)
15.8 ±1.2
(1.3)
23.2 ±2.0
16.7 ±2.0
8.7 ±0.8
(9.2)
7.9 ±0.9
4.4 ± 0.5
5.3 ±0.7
5.7 ±0.6
4.9 ± 0.7
(1.3)
SFE Value8
(mg/kg)
41.3 ±3.6
0.9 ±0.1
0.2 ± 0.01
15.6 ±1.8
16.1 ±1.8
1.1 ±0.2
24.1 ±2.1
17.2 ±1.9
8.8 ±1.0
7.9 ±0.9
8.5 ±1.1
4.1 ±0.5
5.1 ±0.6
5.2 ± 0.6
4.3 ±0.5
1.1 ±0.2
Percent of
Certified Value
(148)
(112)
(100)
(102)
102
(88)
104
103
101
(86)
108
91
96
91
88
(85)
SFE
RSD
8.7
11.1
0.05
11.5
11.2
18.2
8.7
11.0
11.4
11.4
12.9
12.2
11.8
11.5
11.6
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.
8270D - 53
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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
Naphthalene
Acenaphthylene
Acenaphthene
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(a)pyrene
lndeno(1 ,2,3-cd)pyrene
Benzo(g , h , i)perylene
Dibenz(a,h)anthracene
Certified Value
(mg/kg)
9.0 ± 0.7
0.3 ±0.1
4.5 ±1.5
13.6 ±3.1
85.0 ± 20.0
13.4 ±0.5
60.0 ± 9.0
39.0 ± 9.0
14.6 ±2.0
14.1 ±2.0
7.7 ±1.2
2.8 ±2.0
7.4 ± 3.6
5.0 ±2.0
5.4 ±1.3
1.3 ±0.5
SFE Value8 Percent of SFE
(mg/kg) Certified Value RSD
7.4 ± 0.6
0.4 ±0.1
3.3 ±0.3
10.4 ±1.3
86.2 ± 9.5
12.1 ±1.5
54.0 ±6.1
32.7 ± 3.7
12.1 ±1.3
12.0 ±1.3
8.4 ± 0.9
3.2 ± 0.5
6.6 ± 0.8
4.5 ±0.6
4.4 ± 0.6
1.1 ±0.3
82
133
73
77
101
90
90
84
83
85
109
114
89
90
82
85
8.1
25.0
9.0
12.5
11.0
12.4
11.3
11.3
10.7
10.8
10.7
15.6
12.1
13.3
13.6
27.3
Relative standard deviations for the SFE values are based on three replicate extractions.
Data are taken from Reference 10.
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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
Naphthalene
2-Methylnaphthalene
Acenaphthene
Dibenzofuran
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(a)pyrene
Benzo(b)fluoranthene +
Benzo(k)fluoranthene
Certified Value SFE Value3 Percent of SFE
(mg/kg) (mg/kg) Certified Value RSD
32.4 ± 8.2
62.1 ±11.5
632 ± 105
307 ± 49
492 ± 78
161 8 ±340
422 ± 49
1280 ±220
1033 ±285
252 ±8
297 ± 26
97.2 ±17.1
153 ±22
29.55
76.13
577.28
302.25
427.15
1278.03
400.80
1019.13
911.82
225.50
283.00
58.28
130.88
91
122
91
98
87
79
95
80
88
89
95
60
86
10.5
2.0
2.9
4.1
3.0
3.4
2.6
4.5
3.1
4.8
3.8
6.5
10.7
a Relative standard deviations for the SFE values are based on four replicate extractions.
Data are taken from Reference 11.
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TABLE 16
SINGLE LABORATORY RECOVERY DATA FOR SOLID-PHASE EXTRACTION OF
BASE/NEUTRAL/ACID EXTRACTABLES FROM SPIKED TCLP BUFFERS
LOW SPIKE LEVEL
Analyte
1 ,4-Dichlorobenzene
Hexachloroethane
Nitrobenzene
Hexachlorobutadiene
2,4-Dinitrotoluene
Hexachlorobenzene
o-Cresol
m-Cresol*
p-Cresol*
2,4,6-Trichlorophenol
2,4,5-Trichlorophenol
Pentachlorophenol
Spike Level
(M9/L)
3,750
1,500
1,000
250
65
65
100,000
100,000
100,000
1,000
200,000
50,000
Buffer 1 (pH =
Recovery (%)
63
55
82
65
89
98
83
86
*
84
83
82
2.886)
RSD
10
6
10
3
4
5
10
8
*
12
11
9
Buffer 2 (pH =
Recovery (%)
63
77
100
56
101
95
85
85
*
95
88
78
4.937)
RSD
9
4
5
4
5
6
5
3
*
12
3
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
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TABLE 17
SINGLE LABORATORY RECOVERY DATA FOR SOLID-PHASE EXTRACTION OF
BASE/NEUTRAL/ACID EXTRACTABLES FROM SPIKED TCLP BUFFERS
HIGH SPIKE LEVEL
Analyte
1 ,4-Dichlorobenzene
Hexachloroethane
Nitrobenzene
Hexachlorobutadiene
2,4-Dinitrotoluene
Hexachlorobenzene
o-Cresol*
m-Cresol*
p-Cresol*
2,4,6-Trichlorophenol
2,4,5-Trichlorophenol
Pentachlorophenol
Spike Level
(ug/L)
15,000
6,000
4,000
1,000
260
260
400,000
400,000
400,000
4,000
800,000
200,000
Buffer 1 (pH =
Recovery (%)
63
54
81
81
99
89
92
95
82
93
93
84
2.886)
RSD
10
7
4
5
8
8
15
8
14
12
14
9
Buffer 2 (pH =
Recovery (%)
63
46
81
70
98
91
90
82
84
104
97
73
4.937)
RSD
9
7
13
11
3
9
4
6
7
12
23
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
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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 oH = 2.886
Analyte
o-Cresol
m-Cresol**
p-Cresol**
2,4,6-Trichlorophenol
2,4,5-Trichlorophenol
Pentachlorophenol
1 ,4-Dichlorobenzene
Hexachloroethane
Nitrobenzene
Hexachlorobutadiene
2,4-Dinitrotoluene
Hexachlorobenzene
(continued)
Spike Level
(uo/Lr
200,000
—
—
2,000
400,000
100,000
7,500
3,000
2,000
500
130
130
%R
86
77
—
106
93
79
51
50
80
53
89
84
Labi
RSD
8
8
—
6
3
2
5
5
8
8
8
21
n
7
7
—
7
7
7
7
7
7
7
7
7
8270D - 58
%R
35.3
—
—
96.3
80.5
33.8
81.3
66.2
76.3
63.3
35.7
92.3
Lab 2
RSD
0.7
—
—
3.9
4.5
12.2
5.3
2.1
5.3
4.8
2.6
1.6
n
3
—
—
3
3
3
3
3
3
3
3
3
%R
7.6
—
7.7
44.8
63.3
29.2
19.2
12.6
63.9
9.6
58.2
71.7
Lab
3
RSD n
6
—
11
5
11
13
7
11
12
9
17
9
3
—
3
3
3
3
3
3
3
3
3
3
Revision 4
January 1998
-------
TABLE 18
(continued)
Buffer 2 oH = 4.937
Analyte
o-Cresol
m-Cresol**
p-Cresol**
2,4,6-Trichlorophenol
2,4,5-Trichlorophenol
Pentachlorophenol
1 ,4-Dichlorobenzene
Hexachloroethane
Nitrobenzene
Hexachlorobutadiene
2,4-Dinitrotoluene
Hexachlorobenzene
Spike Level
(uolLY
200,00
—
—
2,000
400,000
100,000
7,500
3,000
2,000
500
130
130
%R
97
83
—
104
94
109
50
51
80
57
86
86
Labi
RSD
13
4
—
4
4
11
5
3
4
5
6
7
n
7
7
—
7
7
7
7
7
7
7
7
7
%R
37.8
—
—
91.7
85.2
41.9
79.7
64.9
79.0
60
38.5
91.3
Lab 2
RSD
4.5
—
—
8.0
0.4
28.2
1.0
2.0
2.3
3.3
5.2
0.9
n
3
—
—
3
3
3
3
3
3
3
3
3
%R
6.1
6.0
—
37.7
64.4
36.6
26.5
20.3
59.4
16.6
62.2
75.5
Lab 3
RSD
24
25
—
25
10
32
68
90
6
107
6
5
n
3
3
—
3
3
3
3
3
3
3
3
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
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FIGURE 1
GAS CHROMATOGRAM OF BASE/NEUTRAL AND ACID CALIBRATION STANDARD
R1C
b 8:26:88
StWPLE: BASE ACID STO,2UL/2ttNC-UL
COHDS.:
RrtCEl G 1.2788 UtffELi N 6, 4.6 GUnN: H
CUTn: 5lbHŁ&e«78b kl
CH.1: 51B**S668786 13
J 6
SCANS 2w6 TO 2780
: U 2d- 3
kIC
25:08
•.vvv
33:28
25*
41:48
in
8270D - 60
Revision 4
January 1998
-------
-------
METHOD 8280B
POLYCHLORINATED DIBENZO-P-DIOXINS AND POLYCHLORINATED DIBENZOFURANS 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-trillion concentrations), soil, fly
ash, and chemical waste samples, including stillbottoms, 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)
1,2,3,7,8-Pentachlorodibenzo-p-dioxin (PeCDD)
1,2,3,4,7,8-Hexachlorodibenzo-p-dioxin (HxCDD)
1,2,3,6,7,8-Hexachlorodibenzo-p-dioxin (HxCDD)
1,2,3,7,8,9-Hexachlorodibenzo-p-dioxin (HxCDD)
1,2,3,4,6,7,8-Heptachlorodibenzo-p-dioxin (HpCDD)
1,2,3,4,6,7,8,9-Octachlorodibenzo-p-dioxin (OCDD)
2,3,7,8-Tetrachlorodibenzofuran (TCDF)
1,2,3,7,8-Pentachlorodibenzofuran (PeCDF)
2,3,4,7,8-Pentachlorodibenzofuran (PeCDF)
1,2,3,4,7,8-Hexachlorodibenzofuran (HxCDF)
1,2,3,6,7,8-Hexachlorodibenzofuran (HxCDF)
1,2,3,7,8,9-Hexachlorodibenzofuran (HxCDF)
2,3,4,6,7,8-Hexachlorodibenzofuran (HxCDF)
1,2,3,4,6,7,8-Heptachlorodibenzofuran (HpCDF)
1,2,3,4,7,8,9-Heptachlorodibenzofuran (HpCDF)
1,2,3,4,5,6,7,8-Octachlorodibenzofuran (OCDF)
Total Tetrachlorodibenzo-p-dioxin (TCDD)
Total Pentachlocodibenzo-p-dioxin (PeCDD)
Total Hexachlorodibenzo-p-dioxin (HxCDD)
Total Heptachlorodibenzo-p-dioxin (HpCDD)
Total Tetrachlorodibenzofuran (TCDF)
Total Pentachlorodibenzofuran (PeCDF)
Total Hexachlorodibenzofuran (HxCDF)
Total Heptachlorodibenzofuran (HpCDF)
1746-01-6
40321-76-4
39227-28-6
57653-85-7
19408-74-3
35822-46-9
3268-87-9
51207-31-9
57117-41-6
57117-31-4
70648-26-9
57117-44-9
72918-21-9
60851-34-5
67562-39-4
55673-89-7
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
8280B - 1
Revision 2
January 1998
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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/uL 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 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
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.
8280B-10 Revision 2
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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 spectrometric 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. The 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
8280B -11 Revision 2
January 1998
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concentrations. The percent solids determination does not apply to the extraction of participates
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-extraction of
glassware such as the SDS. Prior to pre-extraction, prepare the thimble by adding 5 g of
70/230 mesh silica gel to the thimble to produce a thin layer in the bottom of the thimble. This
layer will trap 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-extract the SDS 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 2/3 of the sand from the thimble to a clean container, being careful not to disturb
the silica gel layer when transferring the sand. Thoroughly mix the sand with the sample with
a clean 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 extraction, the depression will collapse, and
the sample will be uniformly extracted.
7.2 Chemical waste extraction (including oily sludge/wet fuel oil and stillbottom/oil).
7.2.1 Assemble a flask, a Dean-Stark trap, and a condenser, and pre-extract with
toluene for three hours (see Sec. 4.6). After pre-extraction, allow the apparatus to cool, and
discard the used toluene, or pool 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-
extracted 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-extracted Dean-Stark water separator and condenser
to the flask, and extract the sample by refluxing 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 extract through a rinsed glass fiber filter into a 100-mL round-bottom flask.
Rinse the filter with 10 mL of toluene; combine the extract and rinsate. 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 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.
7.2.3 Prepare an additional two 1-g aliquots of the sample chosen for spiking. After
weighing the sample in a tared pre-extracted 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 extract 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. Cool and filter the toluene extract through a rinsed glass fiber
filter into a 500-mL round-bottom flask. Rinse the filter 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 Dean-Stark 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 x 10Q
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 aliquots 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 aliquots. After allowing the matrix spiking
solution to equilibrate to approximately 1 hour, add the internal standard solution (Sec. 5.10)
and extract the aliquots 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 mark the water
meniscus on the side of the 1-L sample bottle for determination of the exact sample volume.
7.5.2 Add 1 mL of the 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-um 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-um 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 participate
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: Organic-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 bottle(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 into 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 filter 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
participates 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 reaches 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, predean 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 and 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 reaches 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, and 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 37CI4-2,3,7,8-TCDD cleanup
standard (Sec. 5.13). The recovery of this standard is used to monitor the efficiency of the
cleanup procedures. Spike 5 uL 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/uL 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 acid.
Shake for 2 minutes. Remove and discard the acid layer (bottom). Repeat the acid washing
until no color is visible in the acid layer. (Perform acid washing a maximum of 4 times.)
7.8.4 Partition the concentrated extract against 40 mL of 5 percent (w/v) sodium
chloride. (Caution: Acid 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 uL, 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 acid-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 clean 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 acid
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 uL of the continuing
calibration solution (CCS) with 950 uL 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 uL. Proceed
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to Sec. 7.14. If the recovery of any of the analytes 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 analytes 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 uL of the calibration
verification solution to 950 uL 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 uL and proceed with Sec. 7.14. If the recovery
of any of the analytes 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 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 carried 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, filter the eluate, using a 0.7-um 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 rinsates.
7.10.4 While the column is still wet, transfer the concentrated eluate from Sec.
7.9.10 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 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 uL tridecane (or nonane) to the extract and reduce the volume to 100
ML using a gentle stream of clean dry nitrogen (Sec. 7.7). The final extract volume should be
100 uL 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 the 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: 35 - 40 cm/sec at 240°C
Initial Temperature: 170°C
Initial Time: 10 minutes
Temperature Program: increase to 320°C at 8°C/minute
Hold Time: until OCDF elutes
Total Time: 40-45 minutes
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On the DB-5 column, the chromatographic resolution is evaluated using the CCS calibration
standard during both the initial calibration and the calibration verification. The chromatographic
peak separation between the 13C12-2,3,7,8-TCDD peak and the 13C12-1,2,3,4-TCDD peak must
be resolved with a valley of <; 25 percent, where:
Valley = (-) x 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
(Sec. 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-uL 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 separation 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 = (-) x 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 instrument 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 m/z 264 (base peak).
7.13.2 Retention time windows - Prior to the calibration of the target analytes, 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 ^Cl, thus the ion
abundance ratio criterion does not apply to this compound.
7.13.3.1.2 If the laboratory 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 GC column resolution
7.13.3.2 Calculate the relative response factors (RFs) for the seventeen
unlabeled target analytes relative to their appropriate internal standards (RFn) (Table
10), according to the formulae below. For the seven unlabeled analytes and the CI4-
2,3,7,8-TCDD cleanup standard that are found only in the CCS 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 (RfJ (Table 10), in each
calibration standard, according to the following formulae:
(A*1 + Aj) x Qn
-------
The RFn and xRF^ are dimensionless quantities; therefore, the units used to
express the Qn, Qis, 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 (RFJ, where:
F = RF x RF
1 re '" n ' '
This 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 = Standardisation x 1(JO
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 CCS solution on the DB-5 (or equivalent) column, or through
the analysis of the column performance solution on the SP-2331 (or equivalent)
column.
Prepare the CCS 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 CCS
solution in Table 1. Alternatively, use a commercially-prepared solution that contains
the target analytes at the CCS concentrations listed in Table 1.
For the DB-5 (or equivalent) column, begin the 12-hour period by analyzing
the CCS solution. Inject a 2-uL 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-uL 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-uL aliquot of the CCS 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 analytes relative to their
appropriate internal standards (RFn) 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 (RFre), 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 CCS solution shall be
resolved with a valley of <; 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 CCS 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 CCS solution must be within ±30.0% of the mean RFs established
during initial calibration for the analytes in all five calibration standards, and within
± 30.0% of the single-point RFs established during initial calibration for those
analytes present in only the CCS standard (see Sec. 7.13.3.2).
(RF - RFJ
% Difference = -— * * 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 not 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-uL aliquot of the extract to a 0.3-mL vial, and add sufficient
recovery standard solution to yield a concentration of 0.5 ng/uL. Reduce the volume of the
extract back down to 50 uL using a gentle stream of dry nitrogen.
7.14.3 Inject a 2-ul_ 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 uL remain from Sec. 7.14.2. Remove an appropriate size
aliquot from the vial and 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/uL. Reduce the volume of the extract back down to 50 uL 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 clean solvent divided by the volume of the sample aliquot that was diluted.
7.14.4.3 Inject 2 uL 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 time (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 the 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
RT =
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 shirt 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-[COCL]+ 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-
[COCL]* 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 analyte and the internal standard, and flag the result on the report
form.
7.14.5.5 Polychlorinated diphenyl ether (PCDPE) interferences.
The identification of a GC 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-substituted 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:
Q* * (An1 + An2)
cn (ug/kg)
W x (Ai + Aj) x RFn
WATER:
Qis * (An + An2)
Cn (ng/L) =
i + A2) x RFn
where:
An1 and An2 = integrated ion abundances (peak areas) of the quantitation ions of the isomer
of interest (Table 8).
Ais1 and Als2 = 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.
QJS = nanograms of the appropriate internal standard added to the sample prior to
extraction.
RFn = 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 ug/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 Qis.
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
2,3,7,8-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:
* * (An1 + An2) x D
Cn (ug/kg) =
RFre
WATER:
V « (A + A x RFre
Cn (ng/L) =
where:
D = the dilution factor (Sec. 7.14.4.2).
An1, An2, Are1, Are2, Qre, RFre, 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 (ug/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:
100
« RFS x Q,,
where:
A,s1, Ai82, Are1, Are2, (2*, Qrs, and RF^ are defined in Sees. 7.13.3.2 and 7.15.1.
NOTE: When calculating the recovery of the 37CI4-2,3,7,8-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 13C12-1 ,2,3,4-TCDD recovery
standard in the denominator.
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7. 15.5.1 The 13C12-1 ,2,3,4-TCDD is used to quantitate the TCDD and TCDF
internal standards and the cleanup standard, and the 13C12-1,2,3,7,8,9-HxCDD is used
to quantitate the HxCDD, HpCDF and OCDD 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 isomers
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 (Hn) 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:
EDL (ug/kg) =
W x (H + H ) x RFn
WATER:
EDL (ng/L) -
V « (H,l + H*) x RFn
where:
Hn1 and Hn2 = The peak heights of the noise for both of the quantitation ions of the
2,3,7,8-substituted isomer of interest
H^and H^2 = The peak heights of both the quantitation ions of the appropriate
internal standards
D = dilution factor (Sec. 7.14.4.2).
Qis, RFis. W and V are defined in Sees. 7.13.3.2 and 7.15.1.
7. 1 5.6.3 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 the concentration. Calculate the EMPC according to the following formulae:
ALL MATRICES OTHER THAN WATER:
Q,s x (An + An2) x D
EMPCn (ug/kg) =
W x (A(; + A2) x RFn
WATER:
CL x (An1 + An2) x D
EMPCn (ng/L) = —2—1 "
x (Afe + A2) x RFn
where:
Ax1 and A,,2 = Areas of both the quantitation ions.
Ais1, Afe2, Qte, 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
not 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
EMPC 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 greater than
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, and 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 isotopic 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 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, 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.
8280B - 34 Revision 2
January 1998
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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 Revision 2
January 1998
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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-p-dioxins (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, Chemosphere, 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 Revision 2
January 1998
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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 -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 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 - 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. It 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 Revision 2
January 1998
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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 and 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 blood 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 Revision 2
January 1998
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TABLE 1
CALIBRATION SOLUTIONS
Analyte
2,3,7,8-TCDD
2,3,7,8-TCDF
1,2,3,7,8-PeCDF
1,2,3,7,8-PeCDD
* 2,3,4,7,8-PeCDF
* 1,2,3,4,7,8-HxCDF
1,2,3,6,7,8-HxCDF
* 1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
* 1,2,3,7,8,9-HxCDD
* 2,3,4,6,7,8-HxCDF
* 1,2,3,7,8,9-HxCDF
* 1,2,3,4,7,8,9-HpCDF
1,2,3,4,6,7,8-HpCDF
1,2,3,4,6,7,8-HpCDD
OCDD
OCDF
13Cir2,3,7,8-TCDD
13C12-2,3,7,8-TCDF
13C12-1,2l3l6,7,8-HxCDD
13C12-1 ,2,3,4,6,7,8-HpCDF
13C12-OCDD
13C12-1234-TCDD
13C12-123789-HxCDD
37CI4-2378-TCDD
CC1
0.1
0.1
0.1
0.1
0.25
0.25
0.25
0.25
0.5
0.5
0.5
0.5
0.5
1.0
1.0
0.5
0.5
* These compounds are only required in
calculations on these analytes
Concentration
CC2
0.25
0.25
0.25
0.25
0.625
0.625
0.625
0.625
1.25
1.25
0.5
0.5
0.5
1.0
1.0
0.5
0.5
the CC3 solution.
unless they are present in all
8280B - 39
of Standard in ng/uL
CCS
0.5
0.5
0.5
0.5
0.5
1.25
1.25
1.25
1.25
1.25
1.25
1.25
1.25
1.25
1.25
2.5
2.5
0.5
0.5
0.5
1.0
1.0
0.5
0.5
0.25
CC4
1.0
1.0
1.0
1.0
2.5
2.5
2.5
2.5
5.0
5.0
0.5
0.5
0.5
1.0
1.0
0.5
0.5
Therefore, do not
five solutions.
CCS
2.0
2.0
2.0
2.0
5.0
5.0
5.0
5.0
10.0
10.0
0.5
0.5
0.5
1.0
1.0
0.5
0.5
perform % RSD
Revision 2
January 1998
-------
TABLE 2
QUANTITATION LIMITS FOR TARGET COMPOUNDS
Analyte
2,3,7,8-TCDD
2,3,7,8-TCDF
1,2,3,7,8-PeCDF
1,2,3,7,8-PeCDD
2,3,4,7,8-PeCDF
1,2,3,4,7,8-HxCDF
1,2,3,6,7,8-HxCDF
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,8,9-HxCDD
2,3,4,6,7,8-HxCDF
1,2,3,7,8,9-HxCDF
1,2,3,4,6,7,8-HpCDF
1,2,3,4,6,7,8-HpCDD
1,2,3,4,7,8,9-HpCDF
OCDD
OCDF
CAS Number
1746-01-6
51207-31-9
57117-41-6
40321-76-4
57117-31-4
70648-26-9
57117-44-9
39227-28-6
57653-85-7
19408-74-3
60851-34-5
72918-21-9
67562-39-4
35822-46-9
55673-89-7
3268-87-9
39001-02-0
Water
(ng/L)
10
10
25
25
25
25
25
25
25
25
25
25
25
25
25
50
50
Fly
Soil
(M9/kg)
1.0
1.0
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
5.0
5.0
Chemical
Ash
(ug/kg)
1.0
1.0
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
5.0
5.0
Waste*
(ug/kg)
10
10
25
25
25
25
25
25
25
25
25
25
25
25
25
50
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
Revision 2
January 1998
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TABLE 3
INTERNAL STANDARD, RECOVERY STANDARD, AND CLEANUP STANDARD SOLUTIONS
INTERNAL STANDARD SOLUTION
Internal Standards Concentration
5 ng/^L
13CB-2,3,7,8-TCDF 5 ng/pL
13C,2-1,2,3,6,7,8-HxCDD 5 ng/pL
13Cc-1,2,3,4,6,7,8-HpCDF 10 ng/pL
13CC-OCDD 10 ng/pL
RECOVERY STANDARD SOLUTION
Recovery Standards Concentration
13CG-1,2,3,4-TCDD 5 ng/^L
5ng/ML
CLEANUP STANDARD SOLUTION
Cleanup Standard Concentration
37CI4-2,3,7,8-TCDD 5 ng/pL
8280B - 41 Revision 2
January 1998
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TABLE 4
CALIBRATION VERIFICATION SOLUTION
Volume Solution
500 ML CC4 (Table 1)
125 uL Supplemental Calibration solution (below)
50 uL Internal Standard solution (Table 3)
50 uL Recovery Standard solution (Table 3)
50 uL Cleanup Standard solution (Table 3)
225 uL Tridecane (or nonane)
This solution will yield a final volume of 1.0 mL at the concentrations specified for the CCS solution
in Table 1.
Supplemental Calibration Solution Prepared from Commercially-Available Materials
Analyte Concentration (ng/uL)
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 Revision 2
January 1998
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TABLE 5
MATRIX SPIKING SOLUTION
Analyte
Concentration (ng/uL)
2,3,7,8-TCDD
2,3,7,8-TCDF
1,2,3,7,8-PeCDF
1,2,3,7,8-PeCDD
1,2,3,6,7,8-HxCDF
1,2,3,6,7,8-HxCDD
1,2,3,4,6,7,8-HpCDF
1,2,3,4,6,7,8-HpCDD
OCDD
OCDF
2.5
2.5
6.25
6.25
6.25
6.25
6.25
6.25
12.5
12.5
This solution is prepared in tndecane (or nonane) and diluted with acetone prior to use (see Sec.
5.16).
8280B - 43
Revision 2
January 1998
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TABLE 6
PCDD/PCDF ISOMERS IN THE WINDOW DEFINING MIX FOR A 60 m DB-5 COLUMN
Homologue
First
Eluted
Last
Eluted
Approximate
Concentration (ug/mL)
TCDD
TCDF
PeCDD
PeCDF
HxCDD
HxCDF
HpCDD
HpCDF
1,3,6,8-
1,3,6,8-
1,2,4,7,9-
1,3,4,6,8-
1,2,4,6,7,9-
1,2,3,4,6,8-
1,2,3,4,6,7,9-
1,2,3,4,6,7,8-
1,2,8,9-
1,2,8,9-
1,2,3,8,9-
1,2,3,8,9-
1,2,3,4,6,7-
1,2,3,4,8,9-
1,2,3,4,6,7,8-
1,2,3,4,7,8,9-
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
8280B - 44
Revision 2
January 1998
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TABLE 7
RECOMMENDED SELECTED ION MONITORING DESCRIPTORS
Descriptor 1 Descriptor 2 Descriptor 3 Descriptor 4
243
259
277
293
304
306
316
318
320
322
328
332
334
340
342
356
358
376
277
293
311
327
338
340
342
354
356
358
374
376
390
392
402
404
410
446
311
327
345
361
374
376
390
392
402
404
408
410
420
422
424
426
446
480
345
361
379
395
408
410
420
422
424
426
442
444
458
460
470
472
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 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.
8280B - 45 Revision 2
January 1998
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TABLE 8
IONS SPECIFIED FOR SELECTED ION MONITORING FOR PCDDs/PCDFs
Analyte
TCDD
PeCDD
HxCDD
HpCDD
OCDD
TCDF
PeCDF
HxCDF
HpCDF
OCDF
Internal Standards
13C,2-2,3,7,8-TCDD
13Cl2-1,2,3,6,7,8-HxCDD
13CI2-OCDD
13Cl2-2,3,7,8-TCDF
13CI2-1,2,3,4,6,7,8-HPCDF
Recovery Standards
13CB-1,2,3,4-TCDD
13Cl2-1,2,3,7,8,9-HxCDD
Cleanup Standard
37CI4-2,3,7,8-TCDD
Polychlorinated diphenyl ethers
HxCDPE
HpCDPE
OCDPE
NCDPE
DCDPE
Quantitation
320
356
390
424
458
304
340
374
408
442
332
402
470
316
420
332
402
328
376
410
446
480
514
Ions
322
358
392
426
460
306
342
376
410
444
334
404
472
318
422
334
404
(1)
—-
—
—
—
~~~
M-[COCI]+
259
293
327
361
395
243
277
311
345
379
—
—
—
—
—
...
—
265
—
—
—
—
~~~
(1) There is only one quantitation ion monitored for the cleanup standard.
8280B - 46 Revision 2
January 1998
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TABLE 9
CRITERIA FOR ISOTOPIC RATIO MEASUREMENTS FOR PCDDs/PCDFs
Analyte
TCDD
PeCDD
HxCDD
HpCDD
OCDD
TCDF
PeCDF
HxCDF
HpCDF
OCDF
Internal Standards
13CI2-1,2,3,4-TCDD
13Cl2-1,2,3,6,7,8-HxCDD
13CI2-OCDD
13Cl2-2,3,7,8-TCDF
13CB-1,2131416,7,8-HPCDF
Recovery Standards
13CB-,112,3>4-TCDD
13Cl2-1,2,3.7.8,9-HxCDD
Selected
Ions
320/322
356/358
390/392
424/426
458/460
304/306
340/342
374/376
408/410
442/444
332/334
402/404
470/472
316/318
420/422
332/334
402/404
Theoretical
Ion
Abundance
0.77
1.55
1.24
1.04
0.89
0.77
1.55
1.24
1.04
0.89
0.77
1.24
0.89
0.77
1.04
0.77
1.24
Control
Limits
0.65
1.32
1.05
0.88
0.76
0.65
1.32
1.05
0.88
0.76
0.65
1.05
0.76
0.65
0.88
0.65
1.05
-0.89
-1.78
-1.43
-1.20
-1.02
-0.89
-1.78
-1.43
-1.20
-1.02
-0.89
-1.43
-1.01
-0.89
-1.20
-0.89
-1.43
8280B - 47
Revision 2
January 1998
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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
13C12-1,2,3,4-TCDD
13C12-1,2,3,7,8,9-HxCDD
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
13C12-2378-TCDF
37CI4-2378-TCDD
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
13C12-1,2,3,4,6,7,8-HpCDF
13C12-OCDD
8280B - 49
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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
2,3,7,8-tetrachloro-dibenzo-p-dioxin
All other tetrachloro-dibenzo-p-dioxins
1,2,3,7,8-pentachloro-dibenzo-p-dioxin
All other pentachloro-dibenzo-p-dioxins
1,2,3,4,7,8-hexachloro-dibenzo-p-dioxin
1,2,3,6,7,8-hexachloro-dibenzo-p-dioxin
1,2,3,7,8,9-hexachloro-dibenzo-p-dioxin
All other hexachloro-dibenzo-p-dioxins
1,2,3,4,6,7,8-heptachloro-dibenzo-p-dioxin
All other heptachloro-dibenzo-p-dioxins
Octachloro-dibenzo-p-dioxin
All mono-, di-, and trichloro dibenzofurans
2,3,7,8-tetrachlorodibenzofuran
All other tetrachlorodibenzofurans
1,2,3,7,8-pentachlorodibenzofuran
2,3,4,7,8-pentachlorodibenzofuran
All other pentachlorodibenzofurans
1,2,3,4,7,8-hexachlorodibenzofuran
1,2,3,6,7,8-hexachlorodibenzofuran
1,2,3,7,8,9-hexachlorodibenzofuran
2,3,4,6,7,8-hexachlorodibenzofuran
All other hexachlorodibenzofurans
1,2,3,4,6,7,8-heptachlorodibenzofuran
1,2,3,4,7,8,9-heptachlorodibenzofuran
All other heptachlorodibenzofurans
Octachlorodibenzofuran
0.0
1.0
0.0
0.5
0.0
0.1
0.1
0.1
0.0
0.01
0.0
0.001
0.0
0.1
0.0
0.05
0.5
0.0
0.1
0.1
0.1
0.1
0.0
0.01
0.01
0.0
0.001
8280B - 50
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FIGURE 1
GENERAL STRUCTURES OF PCDDs (top) AND PCDFs (bottom)
8
8
8280B - 51
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HO)
.
(D
<-»
(D
0)
co to ro o
o 01 o r»
ooo ro
OOOg
Ol Ol Ol
ooo
3 i i
5 3' 3'
o> u o>
333
Q. Q. Q.
3- 3- 3-
Ł.Ł.<Ł.
Q. Q. Q.
Ol ->l -*
3 .
Relative Intensity
K>
til
O
H
3'
(D
to
*.
o
o
U)
O
to
vl
b'
o
o
o
I
1.3.6.8
1.2.3.4
I
1.2.3.7/1.2.3.8
2.3.7.8
•1.2.8.9
O
C
m
ro
-------
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 - 53 Revision 2
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FIGURE 4
MANUAL DETERMINATION OF S/N
20:00
22:00
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 instrument interface amplifier electronic zero offset be set
high enough so that negative going baseline noise is recorded.
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METHOD 8290A
PQLYCHLORINATED DIBENZODIOXINS (PCDDs^ AND POLYCHLORINATED
DIBENZOFURANS fPCDFs) BY HIGH-RESOLUTION GAS CHROMATOGRAPHY/HIGH-
RESOLUTION MASS SPECTROMETRY (HRGC/HRMS)
1.0 SCOPE AND APPLICATION
1.1 This method provides procedures for the detection and quantitative measurement of
polychlorinated 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)
1,2,3,7,8-Pentachlorodibenzo-p-dioxin (PeCDD)
1,2,3,4,7,8-Hexachlorodibenzo-p-dioxin (HxCDD)
1,2,3,6,7,8-Hexachlorodibenzo-p-dioxin (HxCDD)
1,2,3,7,8,9-Hexachlorodibenzo-p-dioxin (HxCDD)
1,2,3,4,6,7,8-Heptachlorodibenzo-p-dioxin (HpCDD)
1,2,3,4,5,6,7,8-Octachlorodibenzo-p-dioxin (OCDD)
2,3,7,8-Tetrachlorodibenzofuran(TCDF)
1,2,3,7,8-Pentachlorodibenzofuran (PeCDF)
2,3,4,7,8-Pentachlorodibenzofuran(PeCDF)
1,2,3,4,7,8-Hexachlorodibenzofuran (HxCDF)
1,2,3,6,7,8-Hexachlorodibenzofuran (HxCDF)
1,2,3,7,8,9-Hexachlorodibenzofuran (HxCDF)
2,3,4,6,7,8-Hexachlorodibenzofuran (HxCDF)
1,2,3,4,6,7,8-Heptachlorodibenzofuran (HpCDF)
1,2,3,4,7,8,9-Heptachlorodibenzofuran (HpCDF)
1,2,3,4,5,6,7,8-Octachlorodibenzofuran (OCDF)
Total Tetrachlorodibenzo-p-dioxin (TCDD)
Total Pentachlorodibenzo-p-dioxin (PeCDD)
Total Hexachlorodibenzo-/>-dioxin (HxCDD)
Total Heptachlorodibenzo-p-dioxin (HpCDD)
Total Tetrachlorodibenzofuran (TCDF)
Total Pentachlorodibenzofuran (PeCDF)
Total Hexachlorodibenzofuran (HxCDF)
Total Heptachlorodibenzofuran (HpCDF)
1746-01-6
40321-76-4
39227-28-6
57653-85-7
19408-74-3
35822-46-9
3268-87-9
51207-31-9
57117-41-6
57117-31-4
70648-26-9
57117-44-9
72918-21-9
60851-34-5
67562-39-4
55673-89-7
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
8290A - 1
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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/uL) and the highest values for the
octachlorinated congeners (1000 pg/uL). 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 13C12-2,3,7,8-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/uL 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 13C12-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.
13,
5.9 Recovery Standard Solution - This nonane solution contains two recovery standards,
IC12-1,2I3I4-TCDD and 1C12-1,2,3,7,8,9-HxCDD, at a nominal concentration of 50 pg/uL per
compound. 10 to 50 uL 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
8290A - 9 Revision 1
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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.
o/o dry weight = 9 * *Y sample x 1QO
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).
8290A-10 Revision 1
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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 = weight of residue x 2 x 1Q
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.
7.0 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. All samples should be spiked with 100 uL 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
13C12-2,3,7,8-TCDD to give the required 100 ppt fortification level. The fish tissue sample (20
g) must be spiked with 200 uL 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.
8290A-11 Revision 1
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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. The 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.
NOTE: 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.
8290A-12 Revision 1
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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 availability. 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 uL of the sample
fortification solution. Homogenize the mixture for approximately 1 minute with a tissue
homogenizer.
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 mark 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.
8290A-13 Revision 1
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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:
Lipid - „ 100
where:
W,r = weight of the lipid residue to the nearest 0.0001 g calculated from Sec.
7.3.3.3,
V^ = total volume (100 mL) of the extract in ml from Sec. 7.3.2.7,
Wat = weight of the original adipose tissue sample to the nearest 0.01 g from Sec.
7.3.2.1,and
Va, = 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 wool 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.
8290A -14 Revision 1
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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 uL.
NOTE: If the silica gel impregnated with 40 percent sulfuric acid 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.
7.4 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), treat 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
8290A-15 Revision 1
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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 fly ash to two decimal places and transfer to
an extraction jar. Add 100 uL of the sample fortification solution (Sec. 5.8), diluted to
1 mL with acetone, to the sample. Add 150 mL of 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 filter 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 Soxhlet/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 separatory 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-um 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-L 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 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 stirring 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
8290A-17 Revision 1
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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 five 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 filter 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 acid layer (perform a maximum
of four acid 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 filter 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 all traces of
toluene (when applicable) are removed.
7.5.2 Silica/alumina column cleanup
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 gel 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 acid-
impregnated silica gel, and 2 g of silica gel. Tap the column gently after each addition.
A small positive pressure (5 psi) of clean nitrogen may be used if needed. Elute 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 cleaned 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 close 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 Bute the silica 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 it 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 uL of the
calibration verification solution to 950 uL of hexane. Take this solution
through the carbon column cleanup step, concentrate to 50 uL 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 property to ensure that
carbon fines are not carried 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-um filter (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/toluene (75:20:5 v/v)
4 mL of cyclohexane/methylene chloride (50:50 v/v)
The flow rate should be less than 0.5 mL/min. Discard all the column rinsates.
7.5.3.4 While the column is still wet, transfer the concentrated eluate from
Sec. 7.5.2.6 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 and 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 uL using a stream of nitrogen and a sand bath.
Rinse the rotary evaporator flask three times with 300 uL of a solution of 1 percent
toluene in methylene chloride, and add the rinses to the concentrate. Add 10 uL of the
nonane recovery standard solution (Sec. 5.9) for soil, sediment, water, fish, paper pulp
and adipose tissue samples, or 50 uL of the recovery standard solution for sludge, still
bottom and fly ash samples. Store the sample at room temperature in the dark.
7.6 Chromatographic/mass spectrometric conditions and data acquisition parameters
7.6.1 Gas chromatograph operating conditions
Column coating: DB-5
Film thickness: 0.25 urn
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
1st 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 13C-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 instrument 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 5 must be used for the initial calibration.
7.7.1.2 Tune the instrument with PFK, as described in Sec. 7.6.2.2.
7.7.1.3 Inject 2 uL 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-uL 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 the 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 [RF(n); n = 1 to 17]
relative to their appropriate internal standards (Table 5) and the nine RFs for
the 13C12-labeled internal standards [RF(is); is = 18 to 26)] relative to the two
recovery standards (Table 5) according to the following formulae:
_ (An1 + An2) >< Qte
RFis =
(Afe1 + A*) x Qn
(Ate1 + Ais2) x Qn
(Are1 + A3 x Qfe
where:
An1 and An2 = sum of the integrated ion abundances of the quantitation
ions (Tables 6 and 9) for unlabeled PCDDs/PCDFs,
Ais1 and A|82 = sum of the integrated ion abundances of the quantitation
ions (Tables 6 and 9) for the labeled internal standards,
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Are1 and Are2 = sum of the integrated ion abundances of the quantitation
ions (Tables 6 and 9) for the labeled recovery standards,
Qis = quantity of the internal standard injected (pg),
Qre = quantity of the recovery standard injected (pg), and
Qn = quantity of the unlabeled PCDD/PCDF analyte injected (pg).
The RFn and RF* are dimensionless quantities; the units used to express Qit,
Qre and Qn must be the same.
7.7.1.4.5 Calculate the RF values and their respective percent
relative standard deviations (%RSD) for the five calibration solutions:
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+6]+ ion of 13C12-OCDF from the [M+2]+ ion of OCDD
(and [M+4]+ from 13C12-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:
n-1
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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
different. 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 (RFm) to be used for the
determination of the percent recoveries for the nine internal standards are
calculated as follows:
A*"1 * CL
RF = _!• - !»
mffl
where:
m = 18 to 26 (congener type) and j = 1 to 5 (injection number),
Afc"1 = sum of the integrated ion abundances of the quantitation ions
(Tables 6 and 9) for a given internal standard (m = 18 to 26),
Are = sum of the integrated ion abundances of the quantitation ions
(Tables 6 and 9) for the appropriate recovery standard (see
Table 5, footnotes),
Qre, Qfem = 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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R~Fm = 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 and RFJ 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 uL 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 A A 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 carried 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 uL to 50 uL.
NOTE: A final volume of 20 uL or more should be used whenever possible. A 10-uL final
volume is difficult to handle, and injection of 2 uL out of 10 uL leaves little sample for
confirmations and repeat injections, and for archiving.
7.8.2 Inject a 2-uL 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
(SICP) 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 into 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., forTCDDs: 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 verify 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:
cx =
A,. « W « RF.
where:
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Cx = concentration of unlabeled PCDD/PCDF congeners (or group of coeluting isomers
within an homologous series) in pg/g,
AX = 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,
Qi8 = 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
RFn = 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-substituted PCDDs or PCDFs, RFn 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 RT(k) value is the one calculated using the equation in
Sec. 7.7.1.4.6.2. (RFkl 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,, x Q
percent recovery = - - - ^=- x 100
Q x A x RF
n
where:
Ais = sum of the integrated ion abundances of the quantitation ions (Table 6) for the
labeled internal standard,
Are = 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),
Qis = 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
R~F 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/uL 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. Dilute 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 (EDL) 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-substituted 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 =
2.5 x H
X
Hte x W x RFn
where:
EDL = estimated detection limit for homologous 2,3,7,8-substituted PCDDs/PCDFs.
Hx = sum of the 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, RFn, and QK 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 of the 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:
* Sl " 2 ' x 100
where S1 and S2 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 listed 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 of the 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-silica
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 eluters
(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. The concentration calculations are performed as outlined in Sec. 7.9.
The chromatographic separation between the 2,3,7,8-TCDF and its close eluters
(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 are 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 carbon-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 carbon-
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. It 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 uL (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 other 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 = the peak height of 2,3,7,8-TCDD
It is the responsibility of the laboratory to verify 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 letter 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 ion 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 twojiaily 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 uL 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 uL of the recovery standard solution
(Sec. 7.5.3.6) and analyze a 2-uL 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 uL
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 uL 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 ug/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, verify 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
8290A - 37 Revision 1
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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
HCI 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 low 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 Dibenzo-p-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.
8290A - 38 Revision 1
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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-dioxins
in Environmental Samples." Y. Tondeur, W.J. Niederhut, 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-Dioxins and Polychlorinated Dibenzofurans from
Environmental Samples Using Accelerated Solvent Extraction (ASE)," B. E. Richter, J. L.
Ezzell, D. E. Knowles, and F. Hoefler, Chemosphene, 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.
8290A - 39 Revision 1
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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 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 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 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 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.
8290A - 40 Revision 1
January 1998
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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, taking precautions
not to contaminate skin or other articles. Wash exposed skin vigorously and repeatedly until
medical attention is obtained.
8290A - 41 Revision 1
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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 uL 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 uL 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 uL (either in a minivial or
in a capillary tube). Inject 2 uL 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 2V2 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 Revision 1
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TABLE 1
TYPES OF MATRICES, SAMPLE SIZES, AND 2,3,7,8-TCDD-BASED
METHOD CALIBRATION LIMITS (PARTS PER TRILLION)
Lower MCLa
Upper MCLa
Sample
Weight (g)
IS Spiking
Level (ppt)
Final Ext.
Vol. (uL)d
Water
0.01
2
1000
1
10-50
Soil
Sediment
Paper Pulp"
1.0
200
10
100
10-50
Fly Ash
1.0
200
10
100
50
Fish
Tissue0
1.0
200
20
100
10-50
Human
Adipose
Tissue
1.0
200
10
100
10-50
Sludge
Fuel Oil
5.0
1000
2
500
50
Still
Bottom
10
2000
1
1000
50
a 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.
0 One half of the extract from the 20 g sample is used for determination of lipid content (Sec. 7.2.2).
d See Sec. 7.8.1.
NOTE: Chemical reactor residues are treated as still bottoms, if their appearances so suggest.
8290A - 43
Revision 1
January 1998
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TABLE 2
COMPOSITION OF THE SAMPLE FORTIFICATION
AND RECOVERY STANDARD SOLUTIONS8
Sample Fortification Recovery Standard Solution
Analyte Solution Concentration Concentration (pg/uL)
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
13C12-1,2,3,6,7,8-HxCDD 25
13C12-1I2,3,718;9-HXCDD - 50
13C12-1,2,3,4,6,7,8-HpCDD 25
"Ctf-I^.SAej.B-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 Revision 1
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-TCDD(*)
1,2,3,7,8-PeCDDO
1,2,3,6,7,8-HxCDDO
1,2,3,4,7,8-HxCDD
1,2,3,7,8,9-HxCDD(+)
1,2,3,4,6,7,8-HpCDDn
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
1,2,3,4,6,7,8-HpCDFn
1,2,3,4,7,8,9-HpCDF
* The 13C-labeled analogue is used as an internal standard.
+ The ISC-labeled analogue is used as a recovery standard.
8290A - 45
Revision 1
January 1998
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TABLE 4
ISOMERS OF CHLORINATED DIOXINS AND FURANS
AS A FUNCTION OF THE NUMBER OF CHLORINE ATOMS
Number of
Chlorine Atoms
1
2
3
4
5
6
7
8
Total
Number of
Dioxin Isomers
2
10
14
22
14
10
2
1
75
Number of
2,3,7,8-Dioxins
—
—
—
1
1
3
1
1
7
Number of
Furan Isomers
4
16
28
38
28
16
4
1
135
Number of
2,3,7,8-Furans
—
—
—
1
2
4
2
1
10
8290A - 46 Revision 1
January 1998
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TABLE 5
HIGH-RESOLUTION CONCENTRATION CALIBRATION SOLUTIONS
Analyte
Unlabeled Analytes
2,3,7,8-TCDD
2,3,7,8-TCDF
1,2,3,7,8-PeCDD
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCDF
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,8,9-HxCDD
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
1,2,3,4,6,7,8-HpCDD
1,2,3,4,6,7,8-HpCDF
1,2,3,4,7,8,9-HpCDF
OCDD
OCDF
Internal Standards
13C12-2,3,7,8-TCDD
13Cir2,3,7,8-TCDF
13C12-1,2,3,7,8-PeCDD
13C12-1,2,3,7,8-PeCDF
13C,2-1, 2,3,6,7, 8-HxCDD
13C12-1,2,3,4,7,8-HxCDF
13C12-1,2,3,4,6,7,8-HpCDD
13C12-1, 2,3,4,6,7, 8-HpCDF
13C12-OCDD
Recovery Standards
13C12-1,2,3,4-TCDD
13C12-1,2,3,7,8,9-HxCDD
Concentration (pg/pL)
5
200
200
500
500
500
500
500
500
500
500
500
500
500
500
500
1,000
1,000
50
50
50
50
125
125
125
125
250
50
125
4
50
50
125
125
125
125
125
125
125
125
125
125
125
125
125
250
250
50
50
50
50
125
125
125
125
250
50
125
8290A - 47
3
10
10
25
25
25
25
25
25
25
25
25
25
25
25
25
50
50
50
50
50
50
125
125
125
125
250
50
125
2
2.5
2.5
6.25
6.25
6.25
6.25
6.25
6.25
6.25
6.25
6.25
6.25
6.25
6.25
6.25
12.5
12.5
50
50
50
50
125
125
125
125
250
50
125
1
1
1
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2.5
2 5
fc>W
5
5
50
50
50
50
125
125
125
125
250
50
125
Revision 1
January 1998
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TABLE 6
IONS MONITORED FOR HRGC/HRMS ANALYSIS OF PCDDS/PCDFS
Descriptor Accurate Mass*
1 303.9016
305.8987
315.9419
317.9389
319.8965
321.8936
331.9368
333.9338
375.8364
[354.9792]
2 339.8597
341.8567
351.9000
353.8970
355.8546
357.8516
367.8949
369.8919
409.7974
[354.9792]
3 373.8208
375.8178
383.8639
385.8610
389.8156
391.8127
Ion ID Elemental Composition
M C^H^CUO
M+2 C^H^CIa^CIO
M "C^H^CUO
M+2 ^12 ''4 CIs CIO
M Ci2i>4 CI^O2
ft j • o ^ LJ 35/"*i 37^1^^
IVIT^ ^12 4 ^'3 ^-"^2
M "C^H^CIA
i J^O 13^ LJ 35^1 37^1^^
IVl~fc ^/12'*4 ^'3 \s\\Jj
M+2 C12H435CI537CIO
LOCK C9F13
M+2 C^Ha^C^^CIO
M+4 C^Ha^Cla^CljO
ft it • o 13^^ LJ 35^^| 3'^^!^^
IVi^fc ^12 3 4 ^•/'*1^
HJIx. A 13^ LJ 35^1 37^1 ^>
IVI » *r w^2"3 ^'3 wl2v
ft JI^O ^^ LJ 35^^| 37^^|AX
MT^ 0^113 \s\4 V^|VJ2
k •_!_ ji ^% LJ 35 ^^i 37^^i ^^
M+4 ^12"s ^'3 ^'2^2
1 1 • o 13^ LJ 35^1 37^i^%
IVI • fc ^x<9riQ ^1^1 ^^l^^o
KA±A 13/1* LJ 35/^i 37^1 /^
IV|T*fr ^12''3 ^'3 ^'2^2
M+2 C^Ha^Cle^CIO
LOCK C9F13
M+2 ^12^ CIs CIO
jiA±A ^ LJ 35^1 37^1 ^\
M^4 ^12 '"2 ^'4 ^'2^
Ml3o LJ 35/^1 f\
12 2 ^'6
fcJiO 13/** LJ 35/^1 37^1^
M~Z ^'^2*'2 ^5 ^*IV^
M+2 C12H235CI537CI02
M+4 C^Hj^C^C^Oj
8290A - 48
Analyte
TCDF
TCDF
TCDF (S)
TCDF (S)
TCDD
TCDD
TCDD (S)
TCDD (S)
HxCDPE
PFK
PeCDF
PeCDF
PeCDF (S)
PeCDF (S)
PeCDD
PeCDD
PeCDD (S)
PeCDD (S)
HpCDPE
PFK
HxCDF
HxCDF
HxCDF (S)
HxCDF (S)
HxCDD
HxCDD
Revision 1
January 1998
-------
TABLE 6
(continued)
Descriptor Accurate Mass8
401.8559
403.8529
445.7555
[430.9728]
4 407.7818
409.7788
417.8250
419.8220
423.7767
425.7737
435.8169
437.8140
479.7165
[430.9728]
5 441.7428
443.7399
457.7377
459.7348
469.7780
471.7750
513.6775
[442.9728]
S = internal/recovery standard
a The following nuclidic masses were
H = 1.007825
C = 12.000000
13C = 13.003355
F = 18.9984
Ion ID Elemental Composition
M+2 "C^H^CI^CIO,
M+4 "C^H^CI^CIA
M+4 C^H^CI^CIjO
LOCK C9F17
M+2 C^H^CI^CIO
M+4 C^H^CIg^CljO
M "C^H^O
M+2 "C^H^CIe^CIO
M+2 C^H^CIe^CIOj
M+4 C12H35CI537CI2O2
M+2 "C^H^CI^CIOj
M+4 "C^H^CI^CIp,
M+4 C^H^CI^C^O
LOCK C9F17
M+2 C^CI^CIO
KA-4-A O 35f*l 37fM C\
1*1 » Vx<2 ^'6 ^^lo^x
M+2 C1235CI737CI02
M+4 C1235CI637CI2O2
M+2 13C1235CI737CIO2
M+4 13C1235CI637CI2O2
M+4 C^^CIg^CljO
LOCK C10F17
used:
O = 15.994915
^Cl = 34.968853
37CI = 36.965903
8290A - 49
Analyte
HxCDD (S)
HxCDD (S)
OCDPE
PFK
HpCDF
HpCDF
HpCDF (S)
HpCDF
HpCDD
HpCDD
HpCDD (S)
HpCDD (S)
NCDPE
PFK
OCDF
OCDF
OCDD
OCDD
OCDD (S)
OCDD (S)
DCDPE
PFK
Revision 1
January 1998
-------
TABLE 7
PCDD AND PCDF CONGENERS PRESENT IN THE GC
PERFORMANCE EVALUATION SOLUTION AND USED FOR DEFINING
THE HOMOLOGUE GC RETENTION TIME WINDOWS ON A 60-M DB-5 COLUMN
# Chlorine Atoms
4a
5
6
7
8
PCDD Positional
First Eluter
1,3,6,8
1,2,4,6,8/1,2,4,7,9
1 2467,9/1,2,4,6,8,9
1,2,3,4,6,7,9
1,2,3,4,6,7,8,9
Isomer
Last Eluter
1,2,8,9
1,2,3,8,9
1,2,3,4,6,7
1,2,3,4,6,7,8
PCDF Positional Isomer
First Eluter
1,3,6,8
1,3,4,6,8
1,2,3,4,6,8
1,2,3,4,6,7,8
1,2,3,4,6,7,8,9
Last Eluter
1,2,8,9
1,2,3,8,9
1,2,3,4,8,9
1,2,3,4,7,8,9
aln addition to these two TCDD isomers, the 1,2,3,4-, 1,2,3,7-, 1,2,3,8-, 2,3,7,8-, 13C12-2,3,7,8-, and
1,2,3,9-TCDD isomers must also be present as a check of column resolution.
8290A - 50
Revision 1
January 1998
-------
TABLE 8
THEORETICAL ION ABUNDANCE RATIOS AND THEIR CONTROL LIMITS
FOR PCDDS AND PCDFS
# Chlorine
Atoms
4
5
6
6
-------
TABLE 9
RELATIVE RESPONSE FACTOR [RF (NUMBER)] ATTRIBUTIONS
Number
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
Specific Congener Name
2,3,7,8-TCDD (and total TCDDs)
2,3,7,8-TCDF (and total TCDFs)
1,2,3,7,8-PeCDD (and total PeCDDs)
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCDF
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,8,9-HxCDD
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
1,2,3,4,6,7,8-HpCDD (and total HpCDDs)
1,2,3,4,6,7,8-HpCDF
1,2,3,4,7,8,9-HpCDF
OCDD
OCDF
13Cir2,3,7,8-TCDD
13C12-2,3,7,8-TCDF
13C12-1,2,3,7,8-PeCDD
13C12-1,2,3,7,8-PeCDF
13C12-1,2,3,6,7,8-HxCDD
Cio~T |2|O|4. f «O™riXvxUi
13C12-1,2,3,4,6,7,8-HpCDD
13C12-1 ,2,3,4,6,7,8-HpCDF
13C12-OCDD
Total PeCDFs
Total HxCDFs
Total HxCDDs
Total HpCDFs
8290A - 52 Revision 1
January 1998
-------
TABLE 10
2,3,7,8-TCDD TOXICITY EQUIVALENCY FACTORS (TEFS)
FOR THE POLYCHLORINATED DIBENZODIOXINS AND DIBENZOFURANS
Analyte TEFa
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
aTaken from "Interim Procedures for Estimating Risks Associated with Exposures to Mixtures of
Chlorinated Dibenzo-p-Dioxin and -Dibenzofurans (CDDs and CDFs) and 1989 Update", (EPA/625/3-
89/016, March 1989).
8290A - 53 Revision 1
January 1998
-------
TABLE 11
ANALYTE RELATIVE RETENTION TIME REFERENCE ATTRIBUTIONS
Analyte Analyte RRT Reference8
1,2,3,4,7,8-HxCDD 13C12-1,2,3,6,7,8-HxCDD
1,2,3,6,7,8-HxCDF 13C12-1,2,3,4,7,8-HxCDF
1,2,3,7,8,9-HxCDF 13C12-1,2,3,4,7,8-HxCDF
2,3,4,6,7,8-HxCDF 13C12-1,2,3,4,7,8-HxCDF
"The retention time of 2,3,4,7,8-PeCDF on the DB-5 column is measured
relative to 13C12-1,2,3,7,8-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 Revision 1
January 1998
-------
TABLE 12
COMPARISON OF SOXHLET AND PRESSURIZED FLUID EXTRACTION (PFE)
FOR EXTRACTION OF GROUND CHIMNEY BRICK
Analyte
2,3,7,8-TCDD
1,2,3,7,8-PeCDD
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,9,9-HxCDD
1,2,3,4,6,7,8-HpCDD
OCDD
2,3,7,8-TCDF
1, 2,3,7,8 ( + 1,2,3,4,8)-PeCDF
2,3,4,7,9-PeCDF
1,2,3,4,7,8 ( + 1,2,3,4,7,9)-HxCDF
1,2,3,6,7,8-HxCDF
2,3,4,6,7,8-HxCDF
1,2,3,7,8,9-HxCDF
1,2,3,4,6,7,8-HpCDF
1,2,3,4,7,8,9-HpCDF
OCDF
Total TCDD
Total PeCDD
Total HxCDD
Total HpCDD
Total TCDF
Total PeCDF
Total HxCDF
Total HpCDF
* Sum of two extractions of each sample
Data from Reference 8
8290A -
Soxhlet (n=1)
(ng/kg)
6
52
46
120
97
1000
2900
160
430
390
1100
540
400
42
2100
140
2000
440
900
1800
2000
2300
4100
4700
2800
55
PFE (n=2)*
(ng/kg)
6
57
52
130
1000
820
2600
180
470
390
1100
570
360
42
2000
120
2000
530
940
2000
2100
2600
4300
4700
2600
Revision 1
January 1998
-------
TABLE 13
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF URBAN DUST
Analyte
2,3,7,8-TCDD
1,2,3,7,8-PeCDD
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,9,9-HxCDD
1,2,3,4,6,7,8-HpCDD
OCDD
2,3,7,8-TCDF
1, 2,3,7,8 ( + 1,2I3l4,8)-PeCDF
2,3,4,7,9-PeCDF
1,2,3,4,7,8 ( + 1,2,3,4,7,9)-HxCDF
1,2,3,6,7,8-HxCDF
2,3,4,6,7,8-HxCDF
1,2,3,7,8,9-HxCDF
1,2,3,4,6,7,8-HpCDF
1,2,3,4,7,8,9-HpCDF
OCDF
Total TCDD
Total PeCDD
Total HxCDD
Total HpCDD
Total TCDF
Total PeCDF
Total HxCDF
Total HpCDF
ND = Not detected, with detection limit given
* Sum of two extractions of each sample
Data from Reference 8
8290A - 56
Soxhlet (n=1)
(ng/kg)
3.3
11.8
9.8
11.5
ND(8)
113
445
12.5
9.9
13.9
18.7
10.7
3.3
ND(2)
13.2
ND(3)
ND(10)
182
175
86.7
221
333
146
65.9
13.2
in parentheses
PFE (n=2)*
(ng/kg)
3.2
13.1
8.0
9.5
ND(8)
107
314
18.6
12.0
18.1
23.7
15.8
8.7
ND(2)
29.4
ND(3)
ND(10)
325
281
81.7
217
419
179
122
29.4
Revision 1
January 1998
-------
TABLE 14
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF FLY ASH
(with and without HCI pretreatment for PFE)
Analyte
2,3,7,8-TCDD
1,2,3,7,8-PeCDD
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,9,9-HxCDD
1,2,3,4,6,7,8-HpCDD
OCDD
2,3,7,8-TCDF
1, 2,3,7,8 (+1,2,3,4,8)-PeCDF
2,3,4,7,9-PeCDF
1,2,3,4,7,8 ( + 1,2,3,4,7,9)-HxCDF
1,2,3,6,7,8-HxCDF
2,3,4,6,7,8-HxCDF
1,2,3,7,8,9-HxCDF
1,2,3,4,6,7,8-HpCDF
1,2,3,4,7,8,9-HpCDF
OCDF
Total TCDD
Total PeCDD
Total HxCDD
Total HpCDD
Total TCDF
Total PeCDF
Total HxCDF
Total HpCDF
Soxhlet(n=1)
with HCI (pg/kg)t
0.32
1.6
1.2
2.4
2.4
8.2
11.4
3.7
4.2
5.6
7.8
7.2
6.6
0.43
18.0
2.3
13.5
12.0
16.6
38.2
15.0
60.5
83.5
65.2
28.1
PFE (n=2)*
with HCI
(M9/kg)t
0.36
2.1
1.4
2.7
2.3
9.6
12.8
4.3
4.6
6.6
8.7
8.5
7.2
0.56
17.6
2.4
15.8
12.4
20.5
42.4
19.8
67.5
87.3
73.5
32.2
PFE (n=2)*
w/o HCI
(M9/kg)t
0.28
1.7
1.2
2.4
2.2
8.1
10.6
3.4
3.9
5.8
5.4
5.3
4.5
0.30
16.8
2.0
13.9
10.5
16.2
36.7
16.0
56.1
77.4
46.1
26.5
f Fly ash was pretreated with HCI, followed by a water rinse, and extracted with toluene.
J These samples received no HCI pretreatment, and were extracted with a mixture of toluene
and acetic acid.
* Sum of two extractions of each sample
Data from Reference 8
8290A - 57
Revision 1
January 1998
-------
TABLE 15
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF SOIL (EC-2)
Analyte
2,3,7,8-TCDD
1,2,3,7,8-PeCDD
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,9,9-HxCDD
1,2,3,4,6,7,8-HpCDD
OCDD
2,3,7,8-TCDF *
1, 2,3,7,8 ( + 1,2,3,4,8)-PeCDF
2,3,4,7,9-PeCDF
1,2,3,4,7,8 ( + 1,2,3,4,7,9)-HxCDF
1,2,3,6,7,8-HxCDF
2,3,4,6,7,8-HxCDF
1,2,3,7,8,9-HxCDF
1,2,3,4,6,7,8-HpCDF
1,2,3,4,7,8,9-HpCDF
OCDF
Total TCDD
Total PeCDD
Total HxCDD
Total HpCDD
Total TCDF
Total PeCDF
Total HxCDF
Total HpCDF
Soxhlet
ng/kg
270
24
23
83
60
720
4000
100
39
62
740
120
45
4.9
2600
160
7800
430
300
720
1300
620
820
1900
3800
Results (n=10)
%RSD
9.1
12
8.3
3.6
6.2
6.7
6.2
7.3
14
5.5
5.3
6.2
9.0
31
6.7
5.5
8.3
9.7
3.7
5.8
7.0
12
9.4
5.7
8.2
PFE
ng/kg
270
22
24
87
57
720
1 fcU
4200
82
36
60
690
120
60
5.3
2500
160
7000
370
280
690
1300
380
710
1900
3900
Results (n=2)
% RSD
0.0
3.3
3.0
0.8
7.4
1 n
i ,\j
0.0
2.6
3.9
0.0
0.0
0.0
1.2
15
0.0
0.0
3.1
1.9
7.7
2.0
0.0
19
7.0
0.0
3.6
* Single-column analysis only, may include contributions from other isomers that may co-elute.
Data from Reference 8
8290A - 58 Revision 1
January 1998
-------
TABLE 16
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF SEDIMENT (HS-2)
Soxhlet Results (n=10)
Analyte
2,3,7,8-TCDD
1,2,3,7,8-PeCDD
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,9,9-HxCDD
1,2,3,4,6,7,8-HpCDD
OCDD
2,3,7,8-TCDF *
1, 2,3,7,8 (+1,2,3,4,8)-PeCDF
2,3,4,7,9-PeCDF
1,2,3,4,7,8 ( + 1,2,3,4,7,9)-HxCDF
1,2,3,6,7,8-HxCDF
2,3,4,6,7,8-HxCDF
1,2,3,7,8,9-HxCDF
1,2,3,4,6,7,8-HpCDF
1,2,3,4,7,8,9-HpCDF
OCDF
Total TCDD
Total PeCDD
Total HxCDD
Total HpCDD
Total TCDF
Total PeCDF
Total HxCDF
Total HpCDF
ng/kg
ND(1)
1.6
4.5 '
19
24
1200
6500
8.5
1.9
3.7
17
3.7
3.7
ND(1)
91
5.2
300
3.9
17
510
4700
39
33
89
293
% RSD
-
4.6
4.8
4.3
4.3
8.1
4.2
11
17
7.9
7.3
5.6
18
—
1.6
6.7
3.8
14
7.8
5.6
8.3
11
13
3.2
3.3
PFE Results (n=2)
ng/kg
ND(1)
ND(1)
5.2
21
28
1300
7100
6.6
2.0
3.7
17
4.0
4.4
ND(1)
96
5.3
280
2.5
10
570
5100
24
28
87
310
% RSD
—
~
11
0.0
2.6
0.0
0.0
5.4
0.0
3.8
4.3
5.4
3.2
—
3.7
6.7
2.6
34
10
1.3
11
3.0
0.0
12
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 Revision 1
January 1998
-------
TABLE 17
COMPARISON OF SOXHLET AND PFE FOR EXTRACTION OF CONTAMINATED SEDIMENTS
Analyte
2,3,7,8-TCDD
1,2,3,7,8-PeCDD
1,2,3,4,7,8-HxCDD
1,2,3,6,7,8-HxCDD
1,2,3,7,9,9-HxCDD
1,2,3,4,6,7,8-HpCDD
OCDD
2,3,7,8-TCDF *
1,2,3,7,8 ( + 1,2,3,4,8)-PeCDF
2,3,4,7,9-PeCDF
1,2,3,4,7,8 ( + 1,2,3,4,7,9)-HxCDF
1,2,3,6,7,8-HxCDF
2,3,4,6,7,8-HxCDF
1,2,3,7,8,9-HxCDF
1,2,3,4,6,7,8-HpCDF
1,2,3,4,7,8,9-HpCDF
OCDF
Total TCDD
Total PeCDD
Total HxCDD
Total HpCDD
Total TCDF
Total PeCDF
Total HxCDF
Total HpCDF
Hamilton
Soxhlet
3.7
5.1
6.4
27
20
460
3100
61
14
26
27
17
14
ND(2)
130
14
270
50
63
220
850
370
290
240
350
Harbor
PFE
3.1
5.4
7.2
26
28
430
3100
44
14
25
37
16
14
1.6
130
13
210
14
15
180
810
130
110
160
290
Parrots
Soxhlet
19
8.3
8.6
26
24
280
1900
80
ND (20)
22
79
ND (20)
21
4.9
270
17
510
39
87
230
580
400
180
230
400
Bay
PFE
19
6.0
6.7
17
l •
18
250
1600
48
9.8
14
59
15
11
ND(1)
220
12
370
48
66
200
530
270
170
230
360
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 - 60
Revision 1
January 1998
-------
FIGURE 1
GENERAL STRUCTURES OF DIBENZO-p-DIOXIN (TOP) AND DIBENZOFURAN (BOTTOM)
8
8
8290A - 61
Revision 1
January 1998
-------
FIGURE 2
M/AM
5.600
5,600
8,550
Peak profile displays demonstrating the effect of the 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 properly.
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
January 1998
-------
FIGURE 3
TYPICAL 12-HOUR ANALYSIS SEQUENCE OF EVENTS.
8:00 AM
Mass Resolution
Mass Accuracy
Analytical Procedure
Thaw Sample Extract
1
Concentrate to 10 uL
9:00 AM
Initial or
Routine
Calibration
GC Column
Performance
11:00 AM
Samples
Method
Blank
8:00 PM
Mass
Resolution
Routine
Calibration
8290A - 63
Revision 1
January 1998
-------
FIGURE 4
100
22:30
1 LX^A. > *^w«W
25:30
I
27:00
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-silica capillary column under the conditions listed in Sec. 7.6.
8290A - 64
Revision 1
January 1998
-------
FIGURE 5
80-
60-
40-
20-
Ref. mass 304.9824 Peak top
Span. 200 ppm
System file name
Data file name
Resolution
Group number
lonization mode
Switching
Ref. masses
YVES150
A:85Z567
10000
1
EI +
VOLTAGE
304.9824
380.9260
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 MJtiM of 10,500 (10 percent valley definition).
8290A - 65
Revision 1
January 1998
-------
FIGURE 6
MANUAL DETERMINATION OF S/N.
20:00
22:00
24:00
26:00
28:00
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 instrument interface amplifier electronic zero offset be set high
enough so that negative going baseline noise is recorded.
8290A - 66
Revision 1
January 1998
-------
-------
-------
METHOD 8321B
SOLVENT-EXTRACTABLE NONVOLATILE COMPOUNDS BY
HIGH PERFORMANCE LIQUID CHROMATOGRAPHYrTHERMOSPRAY/MASS
SPECTROMETRY (HPLC/TS/MS) OR ULTRAVIOLET (UV) 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-dibromopropyl)
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 Dyes
Disperse Red 1 2872-52-8
Disperse Red 5 3769-57-1
Disperse Red 13 126038-78-6
Disperse Yellow 5 6439-53-8
Disperse Orange 3 730-40-5
Disperse Orange 30 5261-31-4
Disperse Brown 1 17464-91-4
Solvent Red 3 6535-42-8
Solvent Red 23 85-86-9
Anthraquinone Dyes
Disperse Blue 3 2475-46-9
Disperse Blue 14 2475-44-7
Disperse Red 60 17418-58-5
Coumarin Dyes
Fluorescent Brighteners
Fluorescent Brightener 61 8066-05-5
Fluorescent Brightener 236 3333-62-8
Alkaloids
Caffeine 58-08-2
Strychnine 57-24-9
8321B-1 Revision 2
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Analyte CAS No.
Organophosphoms Compounds
Methomyl 16752-77-5
Thiofanox 39196-18-4
Famphur 52-85-7
Asulam 3337-71-1
Dichlorvos 62-73-7
Dimethoate 60-51-5
Disulfoton 298-04-4
Fensulfothion 115-90-2
Merphos 150-50-5
Parathion methyl 298-00-0
Monocrotophos 6923-22-4
Naled 300-76-5
Phorate 298-02-2
Trichlorfon 52-68-6
Tris(2,3-dibromopropyl) phosphate (Tris-BP) 126-72-7
Chlorinated Phenoxvacid Compounds
Dalapon 75-99-0
Dicamba 1918-00-9
2,4-D 94-75-7
MCPA 94-74-6
MCPP 7085-19-0
Dichlorprop 120-36-5
2,4,5-T 93-76-5
Silvex (2,4,5-TP) 93-72-1
Dinoseb 88-85-7
2,4-DB 94-82-6
2,4-D, butoxyethanol ester 1929-73-3
2,4-D, ethylhexyl ester 1928-43-4
2,4,5-T, butyl ester 93-79-8
2,4,5-T, butoxyethanol ester 2545-59-7
Carbamates
Aldicarb* 116-06-3
Aldicarb sulfone 1646-88-4
Aldicarb sulfoxide 1646-87-3
Aminocarb 2032-59-9
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Analyte
Barban
Benomyl
Bromacil
Bendiocarb*
Carbaryl*
Carbendazim*
3-Hydroxycarbofuran
Carbofuran*
Chloroxuron
Chloropropham
Diuron*
Fenuron
Fluometuron
Linuron*
Methiocarb
Methomyl*
Mexacarbate
Monuron
Neburon
Oxamyl*
Propachlor
Propham
Propoxur
Siduron
Tebuthiuron
CAS No.
101-27-9
17804-35-2
314-40-9
22781-23-3
63-25-2
10605-21--7
16655-82-6
1563-66-2
1982-47-4
101-21-3
330-54-1
101-42-8
2164-17-2
330-55-2
2032-65-7
16752-77-5
315-18-4
150-68-5
555-37-3
23135-22-0
1918-16-7
122-42-9
114-26-1
1982-49-6
34014-18-1
a 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 acid
form) and their esters without the use of hydrolysis and esterification in the extraction procedure,
although hydrolysis to the acid 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
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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 an 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 spectrometric (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 pH of a 1-L aqueous sample or 50-g solid sample is adjusted and the sample is
extracted with diethyi 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-BP 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-repeller 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 HCI) 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 acids, react readily with
alkaline substances and may be lost during analysis. Therefore, glassware and glass wool must be
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acid-rinsed, and sodium sulfate must be acidified with sulfuric acid prior to use, to avoid this
possibility.
3.6 Due to the reactivity of the chlorinated herbicides, the standards must be prepared in
acetonitrile. Methylation will occur slowly, if prepared in methanol.
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 will 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-uL 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-uL, 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 single 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-uL injection loop, analytical columns,
purging gases, etc. An automatic injector 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 thermospray 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, analyte 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-
um 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 - C18 reversed-phase column, 100 mm x 2 mm ID, 5-um
particle size of ODS-Hypersil; or C8 reversed phase column, 100 mm x 2 mm ID, 3-um 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 Erlenmeyer 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 - Burrell 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 - 1/fc 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-lined screw-caps or crimp tops.
4.10 Glass wool
4.11 Microsyringes - 100-uL, 50-uL, 10-uL (Hamilton 701 N or equivalent), and 50 uL
(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-mL 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), Na2SO4. Purify by heating at 400°C for 4 hours
in a shallow tray, or by predeaning the sodium sulfate with methylene chloride.
5.4 Ammonium acetate, NH4OOCCH3, solution (0.1 M). Filter through a 0.45-um
membrane filter (Millipore HA or equivalent).
5.5 Acetic acid, CH3CO2H
5.6 Sulfuric acid solution
5.6.1 (1:1, v/v) - Slowly add 50 mL H2SO4 (sp. gr. 1.84) to 50 mL of water.
5.6.2 (1:3, v/v) - Slowly add 25 ml H2SO4 (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, CH2CI2
5.8.2 Toluene, C6H5CH3
5.8.3 Acetone, CH3COCH3
5.8.4 Diethyl Ether, C2H5OC2H5 - 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, CH3CO2C2H5
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 phenoxyacid 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, not the ester form or an ether, as use of these other forms will 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 Tris-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 uL 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/uL 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 of dry samplex10()
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 separator/ funnel. Add 200 uL 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/uL 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 phenoxyacid 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 phenoxyacid moieties
as the acid, 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
Erlenmeyer 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 H2SO4 (1:1) and monitor the pH for 15 minutes, with stirring. If
necessary, add additional H2SO4 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 separatory 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 H2SO4 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
Erlenmeyer flask with a ground-glass stopper. Place 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 some
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 separately 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 Erlenmeyer 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 Erlenmeyer flask. (Rinse the 1000-
mL 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 plasticized 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 acetate (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-
propyhphosphate, 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 analytes 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 all the analytes of interest for a given application respond adequately m a
given ionization mode, a single analysis using that mode may be employed.
7.7.1 Positive ionization mode conditions
Discharge electrode Off
Filament On or off (optional, analyte dependent)
Mass range 150 to 450 amu (analyte dependent, expect 1 to
18 amu higher than molecular weight of the
compound).
Scan time 1.50 sec/scan
Optional repeller wire or plate 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 On
Filament Off
Mass Range 135 to 450 amu
Scan time 1.50 sec/scan
7.7.3 Thermospray temperatures
Vaporizer control 110 to 130°C
Vaporizer tip 200to215°C
jet 210to220°C
Source block 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 uL is normally used. The injection loop must be
overfilled by, minimally, a factor of two (e.g., 20-uL sample used to overfill a 10-uL 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 quadruples) 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 thermospray
interface, circumventing the HPLC.
7.8.1.1 The mass calibration parameters are as follows:
PEG 400 and 600 PEG 800
Mass range 15to765amu Mass range 15to900amu
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
NH4+-H20 36 amu
CH3OH-NrV 50 amu (methanol)
CH3CN-NH4+ 59 amu (acetonitrile)
CH3OOH -MM/ 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)nOH where n=4, gives the
H(OCH2CH2)4OH«NH4+ 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'NHJ* 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 thermospray 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 known that sample interference and/or analyte coelution 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 properly according to the
manufacturer's instructions and that all samples and 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), quadrupole 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 all
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-d5 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 block.
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 the above items will necessitate shutting down the HPLC/TS system, making repairs
and/or replacements, and then restarting the 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 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 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 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 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 and 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, C.A., "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
Spectrometr/1, Anal. Chem.. 1983, 55, 750-754.
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3. Taylor, V., Mickey, 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. EPA: 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, R.A., Cooper, S.D., Pellazari, E.D., Measurements of Organic Pollutants in Water
and Wastewater. ASTM 686. ~"
1 1 . 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 Liquid
Chromatographv in Environmental Analysis. Laurence, J.F., Ed., Humana Press, Clifton, NJ,
13. Voyksner, R.D., "Thermospray HPLC/MS for Monitoring the Environment", In Applications of
New Mass Spectrometrv Techniques in Pesticide Chemistry: Rosen, J.D., Ed., John Wiley and
Sons: New York, 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, ST., 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. 1986.
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 Intertaboratory 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 0"S-LC/MS) Method for Selected Carbamate Pesticides", EPA/600/X-92/102,
August 1992.
24. Markell, C., "3M Data Submission to EPA," letter to B. Lesnik, June 27, 1995.
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TABLE 1
RECOMMENDED HPLC CHROMATOGRAPHIC CONDITIONS
Analytes
Organophosphorus
Compounds
Azo Dyes
Tris(2,3-
dibromopropyl)
Initial Mobile
Phase
(%)
50/50
(water/methanol)
50/50
(Water/CH3CN)
50/50
(water/methanol)
Initial
Time
(min)
0
0
0
Final
Gradient
(linear)
10
5
10
Final Mobile
Phase (%)
100
(methanol)
100
(CH3CN)
100
(methanol)
Time
(min)
5
5
5
phosphate
Chlorinated
phenoxyacid
compounds
75/25
(0.1 M NH4
acetate in 1%
acetic acid/
methanol)
40/60
(0.1 M
Ammonium
acetate in 1 %
acetic acid/
methanol)
15 40/60
(0.1 M NH4
acetate in 1%
acetic acid/
methanol)
5 75/25
(0.1 M Ammonium
acetate in 1 %
acetic acid/
methanol)
10
8321B- 26
Revision 2
January 1998
-------
TABLE 2
RECOMMENDED HPLC CHROMATOGRAPHIC CONDITIONS FOR CARBAMATES
Time (min) Mobile phase A (percent)
Option A 0
30
35
40
45
95
20
0
95
95
Mobile phase B (percent)
5
80
100
5
5
Option B
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)
0 95
30 0
35 0
40 95
45 95
Mobile phase B (percent)
5
100
100
5
5
A = water with 0.1 M ammonium acetate with 1% acetic acid
B = methanol with 0.1 M ammonium acetate with 1% acetic acid, with
optional post-column addition of 0.1 M ammonium acetate.
8321 B-27
Revision 2
January 1998
-------
TABLE 3
COMPOUNDS AMENABLE TO THERMOSPRAY MASS SPECTROMETRY
Disperse Azo Dyes Alkaloids
Methine Dyes Aromatic ureas
Arylmethane Dyes Amides
Coumarin Dyes Amines
Anthraquinone Dyes Amino acids
Xanthene Dyes Organophosphorus Compounds
Flame retardants Chlorinated Phenoxyacid 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
Spike 1
Spike 2
RPD
HPLC/UV
82.2 ± 0.2
87.4 ± 0.6
6.1%
MS
92.5 ± 3.7
90.2 ± 4.7
2.5%
CAD
87.6 ± 4.6
90.4 ± 9.9
3.2%
SRM
95.5117.1
90.0 ± 5.9
5.9%
Data from Reference 16.
8321B- 28 Revision 2
January 1998
-------
TABLE 5
PRECISION AND ACCURACY COMPARISONS OF MS AND MS/MS WITH
HPLC/UV FOR MUNICIPAL WASTEWATER SPIKED WITH DISPERSE RED 1
Spike 1
Spike 2
RPD
HPLC/UV
93.4 ± 0.3
96.2 ±0.1
3.0%
Percent Recovery
MS
102.0 ±31
79.7 ± 15
25%
CAD
82.7 ±13
83.7 ± 5.2
1.2%
Data from Reference 16.
TABLE 6
RESULTS FROM ANALYSES OF ACTIVATED SLUDGE PROCESS WASTEWATER
5 mg/L Spiking Concentration
1
1-D
2
3
RPD
0 mg/L Spiking Concentration
1
1-D
2
3
RPD
Data from Reference 16.
Recovery
HPLC/UV
0.721 ± 0.003
0.731 ± 0.021
0.279 ± 0.000
0.482 ± 0.001
1.3%
0.000
0.000
0.000
0.000
—
8321 B- 29
of Disperse Red 1
MS
0.664 ± 0.030
0.600 ± 0.068
0.253 ± 0.052
0.449 ± 0.016
10.1%
0.005 ± 0.0007
0.006 ± 0.001
0.002 ± 0.0003
0.003 ± 0.0004
18.2%
(mg/L)
CAD
0.796 ± 0.008
0.768 ± 0.093
0.301 ± 0.042
0.510 ±0.091
3.6%
<0.001
<0.001
<0.001
<0.001
—
Revision 2
January 1998
-------
TABLE 7
CALIBRATION MASSES AND % RELATIVE ABUNDANCES OF PEG 400
Mass % Relative Abundance3
18.0
35.06
36.04
50.06
77.04
168.12
212.14
256.17
300.20
344.22
388.25
432.28
476.30
520.33
564.35
608.38
652.41
653.41
696.43
697.44
a Intensities are normalized to mass 432.
8321 B- 30
32.3
13.5
40.5
94.6
27.0
5.4
10.3
17.6
27.0
45.9
64.9
100
94.6
81.1
67.6
32.4
16.2
4.1
8.1
2.7
Revision 2
January 1998
-------
TABLE 8
CALIBRATION MASSES AND % RELATIVE ABUNDANCES OF PEG 600
Mass
18.0
36.04
50.06
77.04
168.12
212.14
256.17
300.20
344.22
388.25
432.28
476.30
520.33
564.35
608.38
652.41
653.41
696.43
a Intensities are normalized to mass 564.
8321 B- 31
% Relative Abundance3
4.7
11.4
64.9
17.5
9.3
43.9
56.1
22.8
28.1
38.6
54.4
64.9
86.0
100
63.2
17.5
5.6
1.8
Revision 2
January 1998
-------
TABLE 9
RETENTION TIMES AND THERMOSPRAY MASS SPECTRA
OF ORGANOPHOSPHORUS COMPOUNDS
Compound Retention Time (min)
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
Mass (% Relative Abundance)"
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 Cl, Br and S, only the base peak of the isotopic cluster is listed.
Data from Reference 17.
8321B-32
Revision 2
January 1998
-------
TABLE 10
SINGLE OPERATOR ACCURACY AND PRECISION FOR LOW CONCENTRATION DRINKING
WATER, LOW CONCENTRATION SOIL, MEDIUM CONCENTRATION DRINKING
WATER, MEDIUM CONCENTRATION SEDIMENT
Matrix
Low cone.
drinking water
(M9/L)
Low cone, soil
(MQ/kO)
Compound
Dimethoate
Dichlorvos
Naled
Fensulfothion
Parathion
methyl
Phorate
Disulfoton
Merphos
Dimethoate
Dichlorvos
Naled
Fensulfothion
Parathion
methyl
Phorate
Disulfoton
Merphos
Mean
Rec. (%)
70
40
0.5
112
50
16
3.5
237
16
ND
ND
45
ND
78
36
118
Std.
Dev.
7.7
12
1.0
3.3
28
35
8
25
4
5
15
7
19
Spike
Cone.
5
5
5
5
10
5
5
5
50
50
50
50
100
50
50
50
Recovery #
Range (%) Analyses
85-54
64-14
2-0
119-106
105-0
86-0
19-0
287-187
24-7
56-34
109-48
49-22
155-81
15
15
15
15
15
15
15
15
15
15
15
15
15
15
15
15
8321B-33
Revision 2
January 1998
-------
TABLE 10
(continued)
Matrix
Medium cone.
drinking water
(M9/L)
Medium cone.
sediment
(mg/kg)
Compound
Dimethoate
Dichlorvos
Naled
Fensulfothion
Parathion
methyl
Phorate
Disulfoton
Merphos
Dimethoate
Dichlorvos
Naled
Fensulfothion
Parathion
methyl
Phorate
Disulfoton
Merphos
Mean Std.
Rec. (%) Dev.
52 4
146 29
4 3
65 7
85 24
10 15
2 1
101 13
74 8.5
166 25
ND
72 8.6
84 9
58 6
56 5
78 4
Spike Recovery #
Cone. Range (%) Analyses
50 61 - 43 12
50 204-89 12
50 9-0 12
50 79 - 51 12
100 133-37 12
50 41-0 12
50 4-0 12
50 126- 75 12
2 91-57 15
2 216-115 15
2 15
2 90-55 15
3 102-66 15
2 70-46 15
2 66-47 15
2 86-70 12
Data from Reference 17.
8321 B- 34
Revision 2
January 1998
-------
TABLE 11
SINGLE OPERATOR ACCURACY AND PRECISION FOR TRIS-BP IN
MUNICIPAL WASTE WATER, DRINKING WATER, CHEMICAL SLUDGE
Mean
Compound Matrix Rec. (%)
Tris-BP Municipal 25
wastewater
Drinking 40
water
Chemical 63
sludge
Spike Recovery #
Std. Dev. Cone. Range (%) Analyses
8.0 2 41-9.0 15
5.0 2 50-30 12
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
Mean
Compound Recovery %
Std. Dev. Spike Cone.
LOW LEVEL DRINKING WATER
Dicamba
2,4-D
MCPA
MCPP
Dichlorprop
2,4,5-T
Silvex
2,4-DB
Dinoseb
Dalapon
2,4-D,ester
HIGH LEVEL DRINKING
Dicamba
2,4-D
MCPA
MCPP
Dichlorprop
2,4,5-T
Silvex
2,4-DB
Dinoseb
Dalapon
2,4-D,ester
63
26
60
78
43
72
62
29
73
ND
73
WATER
54
60
67
66
66
61
74
83
91
43
97
22
13
23
21
18
31
14
24
11
ND
17
30
35
41
33
33
23
35
25
10
9.6
19
8321 B- 36
H9/L
5
5
5
5
5
5
5
5
5
5
5
50
50
50
50
50
50
50
50
50
50
50
Recovery
Range (%)
86-33
37- 0
92-37
116-54
61 -0
138-43
88-46
62- 0
85-49
ND
104-48
103 - 26
119-35
128 - 32
122-35
116-27
99-44
132-45
120 - 52
102-76
56-31
130-76
#
Analyses
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
9
Revision 2
January 1998
-------
TABLE 12
(continued)
Mean
Compound Recovery %
LOW LEVEL SAND
Dicamba
2,4-D
MCPA
MCPP
Dichlorprop
2,4,5-T
Silvex
2,4-DB
Dinoseb
Dalapon
2,4-D,ester
HIGH LEVEL SAND
Dicamba
2,4-D
MCPA
MCPP
Dichlorprop
2,4,5-T
Silvex
2,4-DB
Dinoseb
Dalapon
2,4-D, ester
117
147
167
142
ND
134
121
199
76
ND
180
153
218
143
158
92
160
176
145
114
287
20
Std. Dev.
26
23
79
39
ND
27
23
86
74
ND
58
33
27
30
34
37
29
34
22
28
86
3.6
Recovery
Spike Cone. Range (%)
H9/9
0.1
0.1
0.1
0.1
0.1
0.1
0.1
0.1
0.1
0.1
0.1
pg/g
1
1
1
1
1
1
1
1
1
1
1
147 - 82
180-118
280 - 78
192-81
ND
171 - 99
154-85
245- 0
210-6
ND
239 - 59
209-119
276-187
205-111
226-115
161-51
204-131
225 - 141
192-110
140-65
418- 166
25-17
#
Analyses
10
10
10
10
10
10
10
10
10
10
7
9
9
9
9
9
9
9
9
9
9
7
8321B-37
Revision 2
January 1998
-------
TABLE 12
(continued)
Compound
Mean
Recovery % Std. Dev.
Spike Cone.
Recovery
Range (%)
#
Analyses
LOW LEVEL MUNICIPAL ASH ug/g
Dicamba
2,4-D
MCPA
MCPP
Dichlorprop
2.4,5-T
Silvex
2,4-DB
Dinoseb
Dalapon
2,4-D,ester
83
ND
ND
ND
ND
27
68
ND
44
ND
29
22
ND
ND
ND
ND
25
38
ND
13
ND
23
HIGH LEVEL MUNICIPAL ASH
Dicamba
2,4-D
MCPA
MCPP
Dichlorprop
2,4,5-T
Silvex
2,4-DB
Dinoseb
Dalapon
2,4-D, ester
Source: Reference
All recoveries are in
ND = Not Detected.
66
8.7
3.2
10
ND
2.9
6.0
ND
16
ND
1.9
19.
negative ionization
21
4.8
4.8
4.3
ND
1.2
3.1
ND
6.8
ND
1.7
0.1
0.1
0.1
0.1
0.1
0.1
0.1
0.1
0.1
0.1
0.1
ug/g
1
1
1
1
1
1
1
1
1
1
1
mode, except for 2,4-D,
8321 B-
38
104-48
ND
ND
ND
ND
60- 0
128 - 22
ND
65-26
ND
53-0
96-41
21 - 5
10- 0
16-4.7
ND
3.6- 0
12-2.8
ND
23-0
ND
6.7-0
ester.
9
9
9
9
9
9
9
9
9
9
6
9
9
9
9
9
9
9
9
9
9
6
Revision 2
January 1998
-------
TABLE 13
MULTI-LABORATORY ACCURACY AND PRECISION DATA
FOR THE CHLORINATED PHENOXYACID HERBICIDES
Compound
2,4,5-T
2,4,5-T.butoxy ester
2,4-D
2,4-DB
Dalapon
Dicamba
Dichlorprop
Dinoseb
MCPA
MCPP
Silvex
2,4,5-T
2,4,5-T,butoxy ester
2,4-D
2,4-DB
Dalapon
Dicamba
Dichlorprop
Dinoseb
MCPA
MCPP
Silvex
Spiking Concentration Mean (% Recovery)*
500 mg/L 90
90
86
95
83
77
84
78
89
86
96
50 mg/L 62
85
64
104
121
90
96
86
96
76
65
RSD"
23
29
17
22
13
25
20
15
11
12
27
68
9
80
28
99
23
15
57
20
74
71
8321B- 39
Revision 2
January 1998
-------
TABLE 13
(continued)
Compound
2,4,5-T
2,4,5-T.butoxy ester
2,4-D
2,4-DB
Dalapon
Dicamba
Dichlorprop
Dinoseb
MCPA
MCPP
Silvex
Spiking Concentration Mean (% Recovery)'
5 mg/L 90
99
103
96
150
105
102
108
94
98
87
RSDb
28
17
31
21
4
12
22
30
18
15
15
a Mean of duplicate data from 3 laboratories.
b Relative standard deviation of duplicate data from 3 laboratories.
Data from Reference 20.
8321 B-40 Revision 2
January 1998
-------
TABLE 14
SINGLE-LABORATORY EVALUATION OF AVERAGE RECOVERY
AND PRECISION DATA FOR WATER0
Analyte
Aldicarb sulfoxide
Aldicarb sulfone
Oxamyla
Methomyl
3-Hydroxycarbofurana
Fenuron
Benomyl/Carbendazim
Aldicarb
Aminocarb
Carbofuran
Propoxur
Monuron
Bromacil
Tebuthiuron
Carbaryl
Fluometuron
Propham
Propachlor
Diuron
Siduron
Methiocarb
Barban
Linuron
Chloropropham
Mexacarbate
Chloroxuron
Neburon
Average % Recover/
7.6
56.0
38.9
52.0
22.2
72.5
47.3
81.0
109
85.5
79.1
91.8
87.6
87.1
82.1
84.4
80.7
84.3
90.8
88.0
93.3
88.1
87.1
94.9
79.8
106
85.3
Standard Deviation
2.8
27.1
17.9
19.6
9.3
22.0
14.7
13.7
38.3
10.0
13.7
11.3
12.1
9.0
13.5
8.3
13.8
10.0
14.1
9.5
12.8
11.2
16.8
15.3
12.9
24.9
12.6
%RSD
37.0
48.5
45.9
37.7
41.7
30.3
31.0
16.9
35.1
11.7
17.3
12.3
13.8
10.3
16.5
9.8
17.1
11.9
15.6
10.8
13.8
12.7
19.3
16.1
16.2
23.5
14.8
Values generated from internal response factor calculations.
Nine spikes were performed at three concentrations. The concentrations for Aldicarb sulfoxide,
Barban, Chloropropham, and Mexacarbate spike levels were at 25 ug/L, 50 ug/L, and 100 ug/L.
All other analyte concentrations were 5 ug/L, 10 ug/L, and 20 ug/L One injection was
disregarded as an outlier. The total number of spikes analyzed was 26.
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 % Recovery8
Aldicarb sulfoxide
Aldicarb sulfone
Oxamyl
Methomyl
3-Hydroxycarbofuran
Fenuron
Benomyl/Carbendazim
Aldicarb
Aminocarb
Carbofuran
Propoxur
Monuron
Bromacil
Tebuthiuron
Carbaryl
Fluometuron
Propham
Propachlor
Diuron
Siduron
Methiocarb
Barban
Linuron
Chloropropham
Mexacarbate
Chloroxuron
Neburon
66.9
162
78.9
84.9
105
91.9
95.6
97.9
133
109
104
101
100
104
102
94.5
92.8
94.6
107
100
107
92.3
104
105
77.2
121
92.1
Standard Deviation
31.3
51.4
46.1
25.8
36.3
16.7
18.2
17.0
44.7
14.4
16.5
12.4
9.0
11.9
15.5
15.7
12.0
10.3
17.4
12.0
14.2
15.6
13.6
9.3
9.8
27.3
16.5
%RSD
46.7
31.7
58.5
30.4
34.5
18.1
19.0
17.4
33.6
13.2
15.9
12.3
9.0
11.5
15.2
16.7
12.9
10.9
16.2
12.0
13.2
16.9
13.1
8.9
12.7
22.5
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 pg/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. The 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 OUTLIER REMOVAL)"
Analyte
Aldicarb
Bendiocarb
Carbaryl
Carbendazim
Carbofuran
Diuron
Linuron
Methomyl
Oxamyl
High-Concentration
Samples3
98.7
81.4
92.0
125
87.8
79.9
84.8
93.3
83.8
Percent Recovery
Medium-Concentration
Samples6
110
95.0
108
138
92.3
98.8
93.0
90.8
88.0
Low-Concentration
Samples0
52.0
52.0
62.0
128
72.0
66.0
82.0
90.0
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.
0 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.
d Data from Reference 23.
8321 B-43
Revision 2
January 1998
-------
TABLE 17
MULTI-LABORATORY EVALUATION OF METHOD PRECISION (AFTER OUTLIER REMOVAL)'
High Concentration
Analyte
Aldicarb
Bendiocarfo
Carbaryl
Carbendazim
Carbofuran
Diuron
Linuron
Methomyl
Oxamyl
Average
Std. Dev.
Avg.
88.8
73.3
82.8
28.1
79.0
71.9
76.3
84.0
75.5
sr
11.4
16.1
11.7
5.6
16.7
13.1
8.3
10.8
12.4
SR
34.4
39.3
34.0
15.3
35.2
26.1
32.5
29.4
37.0
%RSDR
12.9
21.9
14.2
19.9
21.2
18.2
10.9
12.9
16.4
16.5
4.0
%RSDR
38.8
53.6
41.1
54.4
44.5
36.3
42.6
35.0
49.1
43.9
7.1
Avg.
44.1
38.0
43.1
13.8
36.9
39.5
37.2
36.3
35.2
Medium Concentration
sr
7.7
6.6
3.0
1.4
5.0
2.6
3.9
2.8
3.7
SR
17.0
16.6
15.7
8.9
16.3
11.8
13.4
15.0
20.8
%RSDr
17.5
17.3
7.0
10.4
13.6
6.5
10.5
7.8
10.4
11.2
4.1
%RSDR
38.5
43.7
36.4
64.2
44.3
29.8
35.9
41.2
59.1
43.7
11.2
Avg
2.6
2.6
3.1
1.6
3.6
3.3
4.1
4.5
4.9
Low Concentration
sr
0.9
0.6
0.7
0.4
0.9
0.5
0.6
0.7
0.5
SR
2.6
1.6
2.3
1.1
3.3
2.6
2.1
4.1
4.6
%RSDr
33.1
21.3
23.3
26.1
25.2
16.2
15.7
15.3
9.7
20.7
7.1
%RSDR
98.2
61.9
75.8
68.2
91.6
77.9
51.4
92.9
93.6
79.1
16.3
sr and SR 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
a Data from Reference 23.
8321B- 44
Revision 2
January 1998
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TABLE 18
SINGLE LABORATORY RECOVERY DATA FOR SOLID-PHASE EXTRACTION OF
CHLORINATED HERBICIDES FROM SPIKED TCLP BUFFERS
Compound
2,4-D
2,4,5-TP
2,4-D
2,4,5-TP
Spike Level (ug/L)
5,000
500
20,000
2000
Buffer 1
Recovery (%)
91
93
100
103*
RSD
2
9
3
2*
Buffer 2
Recovery (%)
79
92*
99*
78
RSD
6
2*
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
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FIGURE 1
SCHEMATIC OF THE THERMOSPRAY PROBE AND ION SOURCE
Flanga •
Sourca
Mounting
Plato
Ion Sampling
Cona
lona
Elactron Vaporizar
Baam
Vapor || Haatar Vaporizar
Tamparatura | Coupling
T4 Block
Tamparatura
T.
X
Vaporizar
Controlar
.
8321B-46
Revision 2
January 1998
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FIGURE 2
THERMOSPRAY SOURCE WITH WIRE-REPELLER
(High sensitivity configuration)
CERAMIC INSULATOR
WIRE REPELLER
8321B-47
Revision 2
January 1998
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FIGURE 3
THERMOSPRAY SOURCE WITH WIRE-REPELLER
(CAD configuration)
CERAMIC INSULATOR
WIRE REPELLER
8321B-48
Revision 2
January 1998
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METHOD 8330A
NITROAROMATICS AND NITRAMINES BY HIGH
PERFORMANCE LIQUID CHROMATOGRAPHY (HPLQ
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:
Analyte
Octahydro-1 ,3,5,7-tetranitro-1 ,3,5,7-tetrazocine
Hexahydro-1 ,3,5-trinitro-1 ,3,5-triazine
1 ,3,5-Trinitrobenzene
1,3-Dinitrobenzene
Methyl-2,4,6-trinitrophenylnitramine
Nitrobenzene
2,4,6-Trinitrotoluene
4-Amino-2,6-dinitrotoluene
2-Amino-4, 6-dinitrotoluene
2,4-Dinitrotoluene
2,6-Dinitrotoluene
2-Nitrotoluene
3-Nitrotoluene
4-Nitrotoluene
Abbreviation
HMX
RDX
1,3,5-TNB
1,3-DNB
Tetryl
NB
2,4,6-TNT
4-Am-DNT
2-Am-DNT
2,4-DNT
2,6-DNT
2-NT
3-NT
4-NT
CAS Number
2691-41-0
121-82-4
99-35-4
99-65-0
479-45-8
98-95-3
118-96-7
1946-51-0
35572-78-2
121-14-2
606-20-2
88-72-2
99-08-1
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.
1.4 The estimated quantitation limits (EQLs) of target analytes determined by Method 8330
in water and soil are presented in Table 1.
8330A -1
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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-um 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
Dry 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 acetonitrile-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-um 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 urn (Supelco LC-18 or equivalent).
Secondary Column: CN reversed-phase HPLC column, 25-cm x 4.6-mm,
5 urn (Supelco LC-CN or equivalent).
8330A - 7 Revision 1
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Mobile Phase: 50/50 (v/v) methanol/organic-free reagent water.
Flow Rate: 1.5mL/min
Injection volume: 100-uL
UV Detector: 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 HPLC 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 analyzable 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.
8330A - 8 Revision 1
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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 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 have procedures for documenting the effect of the matrix on method
performance (precision, accuracy, and quantitation 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
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 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.
8330A - 9 Revision 1
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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 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.M. 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. USAGE Cold Regions
Research and Engineering Laboratory, CRREL Report 89-9.
8330A-10 Revision 1
January 1998
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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.M. 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.M. Miyares (1990). Salting-out solvent extraction for
preconcentration of neutral polar organic solutes from water. Analytical Chemistry, 62:
1355-1356.
9. Miyares, P.M. and T.F. Jenkins (1990). Salting-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
8330A -11 Revision 1
January 1998
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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.
8330A-12 Revision 1
January 1998
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TABLE 1
ESTIMATED QUANTITATION LIMITS
Analytes
HMX
RDX
1,3,5-TNB
1,3-DNB
Tetryl
NB
2,4,6-TNT
4-Am-DNT
2-Am-DNT
2,6-DNT
2,4-DNT
2-NT
4-NT
3-NT
Water
Low-Level
-
0.84
0.26
0.11
-
-
0.11
0.060
0.035
0.31
0.020
-
-
-
(M9/L)
High-Level
13.0
14.0
7.3
4.0
4.0
6.4
6.9
-
-
9.4
5.7
12.0
8.5
7.9
Soil (mg/kg)
2.2
1.0
0.25
0.25
0.65
0.26
0.25
-
-
0.26
0.25
0.25
0.25
0.25
8330A-13
Revision 1
January 1998
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TABLE 2
RETENTION TIMES AND CAPACITY FACTORS ON LC-18 AND LC-CN COLUMNS
Analyte
HMX
RDX
1,3,5-TNB
1,3-DNB
Tetryl
NB
2,4,6-TNT
4-Am-DNT
2-Am-DNT
2,6-DNT
2,4-DNT
2-NT
4-NT
3-NT
Retention time
LC-18
2.44
3.73
5.11
6.16
6.93
7.23
8.42
8.88
9.12
9.82
10.05
12.26
13.26
14.23
(min)
LC-CN
8.35
6.15
4.05
4.18
7.36
3.81
5.00
5.10
5.65
4.61
4.87
4.37
4.41
4.45
Capacity
LC-18
0.49
1.27
2.12
2.76
3.23
3.41
4.13
4.41
4.56
4.99
5.13
6.48
7.09
7.68
factor (k)*
LC-CN
2.52
1.59
0.71
0.76
2.11
0.61
1.11
1.15
1.38
0.95
1.05
0.84
0.86
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.
8330A-14 Revision 1
January 1998
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TABLE 3
SINGLE LABORATORY PRECISION OF METHOD FOR SOIL SAMPLES
Spiked Soils
Analyte
HMX
RDX
1,3,5-TNB
2,4,6-TNT
1,3-DNB
2,4-DNT
Tetryl
Mean Cone.
(mg/kg)
46
60
8.6
46
40
3.5
5.0
17
SD
1.7
1.4
0.4
1.9
1.4
0.14
0.17
3.1
%RSD
3.7
2.3
4.6
4.1
3.5
4.0
3.4
17.9
Field-Contaminated Soils
Mean Cone.
(mg/kg)
14
153
104
877
2.8
72
7.0
669
1.1
1.0
2.3
SD
1.8
21.6
12
29.6
0.2
6.0
0.61
55
0.11
0.44
0.41
%RSD
12.8
14.1
11.5
3.4
7.1
8.3
9.0
8.2
9.8
42.3
18.0
Source: Reference 1.
8330A-15
Revision 1
January 1998
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TABLE 4
MULTILABORATORY ERROR OF METHOD FOR SOIL SAMPLES
Spiked Soils
Mean Cone.
Analyte (mg/kg) SD %RSD
HMX 46 2.6 5.7
RDX 60 2.6 4.4
1,3,5-TNB 8.6 0.61 7.1
46 2.97 6.5
2,4,6-TNT 40 1.88 4.7
1,3-DNB 3.5 0.24 6.9
2,4-DNT 5.0 0.22 4.4
Tetryl 17 5.22 30.7
Source: Reference 1.
TABLE 5
MULTILABORATORY VARIANCE OF METHOD
Analyte Mean Cone. (ug/L)
HMX 203
RDX 274
2,4-DNT 107
2,4,6-TNT 107
3 Nine Laboratories
8330A - 16
Field-Contaminated Soils
Mean Cone.
(mg/kg)
14
153
104
877
2.8
72
7.0
669
1.1
1.0
2.3
FOR WATER
SD
14.8
20.8
7.7
11.1
SD
3.7
37.3
17.4
67.3
0.23
8.8
1.27
63.4
0.16
0.74
0.49
SAMPLES3
%RSD
7.3
7.6
7.2
10.4
%RSD
26.0
24.0
17.0
7.7
8.2
12.2
18.0
9.5
14.5
74.0
21.3
Revision 1
January 1998
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TABLE 6
MULTILABORATORY RECOVERY DATA FOR SPIKED SOIL SAMPLES
Laboratory
1
3
4
5
6
7
8
True Cone
Mean Cone
Std. Dev.
% RSD
% Diff.*
Mean %
Recovery
HMX
44.97
50.25
42.40
46.50
56.20
41.50
52.70
50.35
47.79
5.46
11.42
5.08
95
RDX
48.78
48.50
44.00
48.40
55.00
41.50
52.20
50.20
48.34
4.57
9.45
3.71
96
Concentration (ug/g)
1,3,5-TNB 1,3-DNB Tetryl
48.99
45.85
43.40
46.90
41.60
38.00
48.00
50.15
44.68
3.91
8.75
10.91
89
49.94
45.96
49.50
48.80
46.30
44.50
48.30
50.05
47.67
2.09
4.39
4.76
95
32.48
47.91
31.60
32.10
13.20
2.60
44.80
50.35
29.24
16.24
55.53
41.93
58
2,4,6-TNT
49.73
46.25
53.50
55.80
56.80
36.00
51.30
50.65
49.91
7.11
14.26
1.46
98
2,4-DNT
51.05
48.37
50.90
49.60
45.70
43.50
49.10
50.05
48.32
2.78
5.76
3.46
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)
Recovery (%)
Analyte Soil Method* Aqueous Method**
2,4-DNT 96.0 98.6
2,4,6-TNT 96.8 94.4
RDX 96.8 99.6
HMX 95.4 95.5
* Data from Reference 1.
** Data from Reference 3.
8330A-18 Revision 1
January 1998
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TABLE 8
PRECISION AND ACCURACY DATA FOR THE SALTING-OUT EXTRACTION METHOD
Analyte
HMX
RDX
1,3,5-TNB
1,3-DNB
Tetryl
2,4,6-TNT
2-Am-DNT
2,4-DNT
1,2-NT
1,4-NT
1,3-NT
# Samples
20
20
20
20
20
20
20
20
20
20
20
%RSD
10.5
8.7
7.6
6.6
16.4
7.6
9.1
5.8
9.1
18.1
12.4
Mean Recovery (%)
106
106
119
102
93
105
102
101
102
96
97
Highest
Concentration Tested
1.14
1.04
0.82
1.04
0.93
0.98
1.04
1.01
1.07
1.06
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
Sample
1
2
3
4
5
6
Technique
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
HMX
1.04
1.00
0.93
94
54.2
64.0
57.1
93
85.7
93.1
78.9
45
45.7
48.0
40.8
0.76
1.16
1.19
10.5
11.5
10.3
Analvte Concentration (ua/L)
RDX TNB DNB DNA TNT 24D 4A
2.45 0.47
1.33 0.44
1.35 0.57
79
63.8 0.3 0.33
83.1 0.3 0.34
71.8 0.3 0.29
91
75.3 0.2 0.19
88.8 0.2 0.17
74.7 0.2 0.13
14
16.4 0.17 0.3 0.13
21.6 0.2 0.19
18.9 0.2 0.13
5.77
6.48
6.11
6.17 0.10
7.03 0.10
6.34 0.07
8330A - 20
0.36
0.29
0.28
3.08
3.34
2.89
2.43
2.49
1.99
2.18
2.31
2.07
0.13
0.16
0.16
0.71
0.79
0.82
2A
0.32
0.30
0.56
1.36
2.27
2.05
1.31
1.65
1.89
1.21
1.42
1.64
0.05
0.05
0.14
0.33
0.40
0.70
Revision 1
January 1998
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TABLE 9
(continued)
Sample
7
8
9
10
14
16
18
Technique
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
HMX
134
75.4
115
109
0.61
0.64
0.64
25
30.2
31.2
27.5
0.33
0.62
0.26
0.58
0.77
0.66
165
141
Analvte Concentration (ua/L)
RDX TNB DNB DNA TNT 24D 4A 2A
365
202 0.98 8.12 1.80
308 1.51 11.3 3.44
291 1.41 9.81 3.30
10.9
11.9
11.0
13
12.1 1.14 0.56
12.7 1.50 0.79
11.0 1.34 0.79
7.12
8.23
7.60
13
5.98
12.0
11.6
40
28.7 0.04 0.39 0.13
33.8 0.03 0.43 0.17
32.7 0.03 0.44 0.22
58 97
39.1 0.80 0.96 8.5 5.62
8330A - 21 Revision 1
January 1998
-------
TABLE 9
(continued)
Analvte Concentration (ua/L)
Sample
19
21
22
24
25
27
Technique
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
HMX
152
138
173
172
142
136
252
227
238
226
218
201
203
199
2.15
2.47
2.34
112
82.8
91.0
77.3
RDX TNB DNB DMA TNT
44.4 0.93 0.88
40.9 0.90 0.99
76 17
69.5 2.6 23.1
75.6 0.11 2.5 20.9
72.7 0.11 2.4 20.3
157 5 110
132 6.62 0.30 102
146 6.90 0.33 104
141 6.45 0.31 102
40
35.9
36.5
35.8
7.54
8.91
8.84
0.59
0.63
608 8 180
429 4.45 0.79 137
510 9.53 0.90 149
445 7.37 0.79 128
8330A - 22
24D 4A
9.5
9.3
59
1.20 65.2
1.08 57.7
1.23 55.0
47
42.6
48.0
47.0
2.20
2.74
2.78
10
7.71
8.25
8.16
2A
7.01
6.03
54
56.4
50.5
48.0
65
56.5
63.5
61.8
1.90
2.24
2.08
8
6.20
7.67
6.33
Revision 1
January 1998
-------
TABLE 9
(continued)
Sample
28
29
31
32
Analyte Concentration fua/L)
Technique HMX RDX TNB DNB DMA TNT 24D 4A
Direct 325 102 14 51
Salting-out 290 87.5 0.37 0.10 13.9 42.3
SPE-Cart. 319 109 0.87 0.17 22.0 56.2
SPE-Disk 249 85.7 0.65 0.13 17.2 43.0
Direct
Salting-out
SPE-Cart. 0.43
SPE-Disk 0.28
Direct
Salting-out
SPE-Cart. 0.21
SPE-Disk 0.23
Direct
Salting-out
SPE-Cart.
SPE-Disk 0.38
2A
40
33.5
45.0
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 here. 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 (%)
Sample
4
29
LCS
Technique
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
HMX
0
0
1
3
1
4
6
5
RDX TNB
24
15
12
8
26
7
0 0
4 4
1 7
7 7
DNB DNA TNT 24D 4A
6 100 8 18
0 45 8
0 17 2
1 1
3 3
6 6
13 6
2A
11
5
1
All data are taken from Reference 10.
8330A - 24
Revision 1
January 1998
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TABLE 11
RECOVERY OF ANALYTES FROM SPIKED SAMPLES
Percent Recovery (%)
Sample
LCS1
LCS2
29
4
Technique
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
Direct
Salting-out
SPE-Cart.
SPE-Disk
HMX
99.5
94.2
99.0
92.5
98.8
91.0
93.5
88.0
95.0
107.0
103.0
80.0
105.5
23*
351*
308*
RDX
98.5
91.2
101.0
95.6
98.2
95.0
100.0
102.0
95.5
89.0
107.0
78.0
105.0
191*
95*
49.5*
TNB
95.6
92.9
96.6
89.3
95.9
89.0
83.0
83.0
95.2
85.0
104.0
76.0
103.0
76.0
92.2
87.4
TNT
96.5
83.2
94.1
88.6
97.2
81.0
89.1
78.0
92.8
89.0
05.0
78.0
104.0
83.0
91.1
85.6
24D
98.1
92.1
95.1
86.9
99.2
89.0
89.3
82.0
93.0
65.0
102.0
77.0
105.0
76.0
93.7
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
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FIGURE 1
EXAMPLE CHROMATOGRAMS
EXPLOSIVES ON A
C18 COLUMN
x
X
io
EXPLOSIVES ON A
CN COLUMN
10
1 4
8330A - 26
Revision 1
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METHOD 8330A
NITROAROMATICS AND NITRAMINES BY HIGH
PERFORMANCE LIQUID CHROMATOGRAPHY (HPLC)
>
N^co
r
Salting Out
7.1.1.1 Add 251.3 g of satt
and 770 mL of water
sampla to a 1 L vol. flask.
Mix the contents.
>
t
7.1.1.2 Add 164 ml of
acatonitrila (ACN) and
stir. Allow phasas to
separate.
>
1
7.1.1.3 Transfer ACN layar
to 100 mL vol. flask. Add
10 mL of fresh ACN to 1 L
flask and stir. Transfer 2nd
portion and combina with 1st
100 mL flask.
T
r
7.1.1.4 Add 84 mLof
salt watar to the ACN
extract and stir. Transfer
ACN extract to 10 mL
grad. cylinder.
^
r
7.1. 1.5 Add 1 mLof ACN to
100 mL vol. flask. Stir and
transfer to the 10 mL grad.
cylinder. Record voluma.
Dilute 1:1 with raagant
watar.
1
r
7 1.1.6 Filter if turbid.
Transfer to a vial for
RP-HPLC analysis.
7.1
Is sample in
an aqueous or
soil/sediment
matrix?
7.1.3 Solid Phase
Extraction (SPE).
Go to Mathod 3535
7.1.2 Sample Filtration:
Placa 5 mL sample in
scintilation vial. Add 5 ml
mathanol, shake, and filter.
Discard first 3 mL. Retain
remainder for usa.
—©
8330A - 27
Revision 1
January 1998
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METHOD 8330A
(continued)
7.JL4J. SamEle Homogenjzation
Air dry sample at room Temp.
or below. Avoid exposure to
direct sunlight. Grind sample
in an acetonitrile rinsed mortar.
7.1.4.2 Sample Extraction.
7.1.4.2.1 Place 2 g soil
subsample, 10 mLs
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.
7.2 Set Chromatographic
Conditions.
7.3 Calibration of
HPLC.
7.3.2 Run working stds. in
triplicate. Calculate calibration
factors as described in
Method 8000.
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.
7.4 Sample Analysis.
7.4.1 Analyze samples. Confirm
measurement w/injection onto
CN column.
7.4.3 Refer to Table 2 for
typical anlayte retention
times.
8330A - 28
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-------
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METHOD 4670
TRIAZINE HERBICIDES AS ATRAZINE 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 ug/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 uL 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
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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 ug/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.
4670-5 Revision 0
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NOTE: Spiking at 3 ug/L may require that the sample be diluted to be within the calibration range
for some testing products, however, it provides data regarding the bias (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/B0 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 clean 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
analyte 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 ug/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.
4670-6 Revision 0
January 1998
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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 ug/L). The standards must fall within the B/B0 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 Logit-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/B0 or the Logit 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
4670-7 Revision 0
January 1998
-------
algorithms for competitive binding assays. The Logit B/B0 is calculated according to the
following formula:
Logit (B/Bo) = log.
BO
log.
t _ _B_
^ Bo
where:
loge = Natural log or logarithm base e
B = Response of the standard or sample
BQ = Response of the zero standard
When Logit B/B0 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 (absorbance) 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/BQ 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/B0 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.
4670-8 Revision 0
January 1998
-------
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
limitations 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
4670-9 Revision 0
January 1998
-------
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
EC,
mean concentration = C = ——
n
where Q 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:
C* — C*
RPD = I 1 " 2I x 100
Pi + C2 )
where C, and C2 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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SD =
Ł(CrC)2 SD = ^ x 100
1-1 C
n-1
where C, is the concentration in each replicate, C 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 clean 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 - C
Recovery = %R = -5 H x 100
Cn
where:
Cs = 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 all 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 three graphical representations of the calibration of atrazine
using a competitive binding immunoassay such as those described here.
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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., 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
Compound
Atrazine
Ametryn
Prometryn
Propazine
Prometon
De-ethylated atrazine
Simazine
Terbutryn
Terbuthylazine
Hydroxy atrazine
De-isopropylated atrazine
Cyanazine
CAS#
1912-24-9
834-12-8
7287-19-6
139-40-2
1610-18-0
6190-65-4
122-34-9
886-50-0
5915-41-3
2163-68-0
1007-28-9
21725-46-2
Percent Reactivity
Relative to Atrazine
100
185
113
97
32
22
15
13
5
0.5
0.3
<0.1
TABLE 2
METHOD DETECTION LIMIT (pg/L)
Product
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
Reagent Water
Municipal Tap Water
Well Water
Surface Water
Field-Contaminated
Sample 1
Field-Contaminated
Sample 2
Spike
Cone.
(ug/L)1
0.00
0.00
0.15
1.00
3.00
0.00
0.15
1.00
3.00
0.00
0.15
1.00
3.00
0.00
0.60
2.00
4.00
0.00
0.80
2.00
4.00
Mean Cone.
n (ug/L)
14
14
14
13
14
14
13
14
14
14
14
14
13
14
14
14
14
14
13
14
14
0.02
0.02
0.16
1.13
2.85
0.00
0.15
1.05
2.88
0.02
0.17
1.06
3.44
0.24
0.93
2.19
3.48
0.47
1.28
3.15
4.03
Single
Analyst
RSD
—
—
19.1
9.09
9.08
—
16.6
9.53
9.95
—
27.7
8.22
11.7
29.9
9.27
9.13
8.29
20.1
10.0
7.74
9.52
Overall Recovery
RSD (%)2
—
—
39.6
15.6
9.08
—
43.4
11.5
9.95
—
39.0
16.2
19.1
35.8
17.9
13.8
9.28
30.8
19.5
18.4
12.5
—
—
107
113
95
—
100
105
96
—
113
106
114
—
115
98
81
—
101
134
89
1 Data for the two field-contaminated water samples represent the amount of atrazine added to the
sample and the mean concentration and RSD data represent the amount found in excess of the
background field contamination.
2 Recovery not calculated for unspiked samples.
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FIGURE 1
CALIBRATION DATA FROM A COMPETITIVE IMMUNOASSAY
ti
14
A2
Figure 1a - Generalized plot of immunoassay signal (test response) versus concentration of
calibration standard (ug/L).
B/(Bo-B)
-24 -14 -14 -M 04 OS
locCoooiDtntkn
24
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
24
14
14
ft*
&0
46
•14
i . r
•24 -24 -14 -14 -84 04 «4 14 14 24
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.
0/o_B_ = Response of the standard or sample x 10Q
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 untargeted 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
analytes (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 analytes 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:
Logit (B/B0) = loge
Bn
= log.
Natural Log - The logarithm, base e, of a number. The natural logarithm may also be represented
as "In" or "loge."
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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-um 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 and 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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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 quality 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 of diesel 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
dodecyl 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, DEXSIL Corp., Hamden, CT.
2. Supplementary Validation Data, Additional Analyte and Contaminant Testing Data for the
PetroFLAG'1** Hydrocarbon Analysis System, DEXSIL Corp., Hamden, CT, August 24, 1995.
3. PetroFLAG™ 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 Validation Package IV, Polycyclic Aromatic Hydrocarbon Response data
for Method 9074 Petroleum Hydrocarbons in Soil by Turbimetric Analysis - PetroFLAG'"' 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*
1
2
3
4
5
6
7
Average (ppm)
SD (ppm)
MDL (ppm)
30 ppm diesel fuel
34
24
28
34
36
32
30
31.03
4.12
13.0
30 ppm motor oil
35
41
40
53
46
48
42
43.6
5.91
18.6
Data from Reference 1.
TABLE 2
RELATIVE RESPONSE OF VEGETABLE OILS AS AN INTERFERANT
Analyte Spike
Concentration (ppm)
50
100
200
500
1000
Mineral Oil
Response (ppm)
55
100
189
504
947
Vegetable Oil
Response8 (ppm)
30
45
94
111
208
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 Oil8
0 (0) 100
5 (1) 94
25 (5) 98
50 (10) 95
100 (20) 85
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.
(pg/g)
Diesel Fuel
100
200
300
400
500
600
700
800
900
1000
Corr Coef
Slope
Intercept
Motor Oil
100
200
300
400
500
600
700
800
900
1000
Corr Coef
Slope
Intercept
Trial 1
112
230
312
420
538
626
774
910
1091
1182
Trial 1
121
243
381
428
531
654
717
880
931
1014
PetroFLAG
(pg/g)
Trial 2
116
248
370
455
564
654
790
900
977
1062
0.999
1.126
-2.8
Trial 2
128
292
408
497
554
668
771
883
1052
1098
0.998
1.02
50.9
Trial
73
158
242
299
342
460
509
612
678
646
Trial
123
200
301
341
441
534
609
711
835
887
3550/801 5B
(M9/9)
1 Trial 2
82
156
218
275
344
439
494
607
614
649
0.992
0.679
30.5
1 Trial 2
82
200
275
343
452
528
652
746
881
846
0.997
0.887
20.5
Data from Reference 1.
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TABLE 5
ANOVA RESULTS FOR SPIKED PETROLEUM HYDROCARBON SAMPLES
Analyte/Concentration
Diesel, 200 ppm
Diesel, 1000 ppm
Motor Oil, 200 ppm
Motor Oil, 1000 ppm
n
20
20
20
20
Mean
(x)
1.09
1.00
1.12
0.937
Variance
-------
TABLE 8
RESPONSE OF PetroFLAG SYSTEM WITH VARIOUS SOAP CONCENTRATIONS3
Soap Concentration (ppm)
100 200 500 1000
PetroFLAG Response (ppm) 500 494 488 502 528
Response of the PetroFLAG system for soil containing 500 ppm of diesel 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)
100 200 500 1000
PetroFLAG Response (ppm) 472 474 488 486 496
3 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 ANTHRACENE3
Anthracene Cone, (ppm)
100 200 500 1000 2000
PetroFLAG Response (ppm) 798 1376 1641 1380 1735
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
a 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
Spike Level in ppm PetroFLAG Reading Response Relative to
Compound
Anthracene
Benzo[a]pyrene
Chrysene
Fluoranthene
Pyrene
(Matrix Used)
100 (Soil)
50 (Soil)
16 (Solvent)
200 (Solvent)
200 (Solvent)
in ppm (Rf 10)
798
180
172
101
216
Mineral Oil Calibrator
8
3.6
11
0.5
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 maximum quantifiable concentration.
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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 5050:
Method 901 OB:
Method 9012A:
Method 9013:
Method 9014:
Method
Method
Method
Method
Method
Method
Method
9020B:
9021:
9022:
9023:
9030B:
9031:
9034:
Method 9035:
Method 9036:
Method 9038:
Method 9056:
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 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, AA II)
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
Phenolics (Spectrophotometric, Manual 4-AAP with
Distillation)
Phenolics (Colorimetric, Automated 4-AAP with Distillation)
Phenolics (Spectrophotometric, MBTH with Distillation)
Total Recoverable Oil & Grease (Gravimetric, Separator^
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
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METHOD 9216
POTENTIOMETRIC 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[Cj + EKjj Cjzj]
Where: E = Reference potential
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s = Slope
Cj = Primary ion concentration
KJJ = Selectivity Coefficient
CjZj=lnterfering ion concentration
Zj = charge ratio of interfering ion
Successful analytical conditions depend upon:
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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, (NH^SO^: 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
clean 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 1 0 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.
7.1 1 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 ISE 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. All 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 filling 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, N.A.Chaniotakis, 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 toxirity 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 listed 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
Hydroxide
Fluoride
Chloride
Chlorate
Perchlorate
Bromide
Iodide
Sulfate
Nitrate (N)
Phosphate
Polyphosphate
Bicarbonate
Acetate
Lactate
Phthalate
Ascorbate
Salicylate
logK,
2.8
-3.1
-3.1
-3.4
-3.1
-3.0
-1.2
-4.1
-3.3
-4.0
-4.4
-3.3
-3.2
-4.9
-2.5
-4.2
-0.8
10% Error Ratio (ppm)
-
170
320
1600
830
570
15
1100
200
9500
3400
870
720
Very high
380
Very high
7.0
Source: Reference 1.
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FIGURE 1
NITRITE ELECTRODE pH RESPONSE
200
100
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FIGURE 2
CALIBRATION CURVE FOR STANDARD LEVEL OF NITRITE
Nitrite, mg/L as N
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FIGURE 3
CALIBRATION CURVE FOR LOW LEVEL NITRITE METHOD
200
140
0.01
Nitrite, mg/L as N
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