EMERGENCY RESPONSE ANALYTICAL METHODS
FOR USE ON BOARD MOBILE LABORATORIES
EDITED BY:
David P. Remeta
Michael Gruenfeld
U. S. Environmental Protection Agency
Releases Control Branch
Hazardous Waste Engineering Research Laboratory
Edison, New Jersey 08837
DRAFT DOCUMENT
(SEE DISCLAIMER)
REVISED JUNE, 1987
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DISCLAIMER
This manual of analytical methodology has neither been reviewed nor
cleared by the U.S. Environmental Protection Agency and is provided
solely as a personal communication. The methods contained in this manual
are not offical EPA methods and were developed solely for use on board an
EPA mobile laboratory during emergency response situations in remote
field locations. The mention of trade names or commercial products in
this manual does not constitute endorsement or recommendation for use.
ACKNOWLEDGMENTS
The compilation of the methods contained within this manual necessarily
required the assistance and contributions of many individuals. The
editors gratefully acknowledge the encouragement and support received
from Ira Wilder and Frank Freestone (U.S. EPA, Releases Control Branch)
and Stephen Dorrler and Dr. Joseph Lafornara (U.S. EPA, Environmental
Response Team). The editors also express their gratitude to Dr. Dennis
Stainken and Robyn Montgomery Wagner who assisted in the preparation of
earlier versions of the manual. Members of the RCB Chemistry Staff who
were instrumental in developing and evaluating the methodology contained
herein include: Uwe Frank; Raymond Frederick; Michael Urban; Richard
Losche; Carolyn Esposito; and, Malvina Wilkens. Special appreciation is
also extended to those government and contractor personnel who applied
this methodology both inhouse and in the field on board the EPA mobile
laboratory during numerous emergency response episodes. Members of this
group include: Walter Berger; Daniel Bingham; Janet Cull inane; Bruce
DeMaine; Frank Healy; Lynn Ho; Kevin Hoogerhyde; George Klinger; Michael
Kwiatkowski; Anthony Lombardo; Larry Mall is; Thomas Mancuso; Jeanne-Marie
O'Donnell; Nick Pangaro; Michael Psota; Jay Quimby; Thomas Roberts;
Kathleen Vasile; Kevin Vasilik; and, Bruce Wagner.
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PREFACE
Response to spills of hazardous materials and petroleum oils and discharges from
uncontrolled hazardous waste sites requires rapid and effective ameliorative
measures. During the past decade, the Releases Control Branch (RCB) of EPA's
Hazardous Waste Engineering Research Laboratory (HWERL) has responded to numer-
ous requests for emergency analytical support involving the mitigation and
cleanup of accidental discharges of hazardous substances. On-site acquisition
of analytical data is routinely required to accurately assess the extent of
environmental contamination and efficiently monitor ensuing cleanup efforts.
Consequently, the Branch has designed and deployed mobile laboratories to
furnish the requisite analytical support. Mobile laboratories accomplish this
task by providing multi-parametric analytical capabilities based on the array
of methodology and instrumentation available.
The Chemistry Staff of RCB has focused its method development activities on
generating methodology that is space and time efficient, and thereby suitable
for use within the confines of a mobile analytical laboratory. These methods
are designed to provide real time sample analysis data of known quality to
assist on-scene coordinators (OSC) in developing comprehensive site mitigation
strategies. The methodology has been employed successfully on board mobile
laboratories operating on-site in support of multi-media cleanup efforts. The
accurate qualitative and quantitative multicomponent analyses afforded through
application of such methodology is a practical and cost-effective alternative to
shipping samples off-site for analysis.
The methods, protocols and procedures developed by RCB for specific use aboard
mobile analytical laboratories are compiled in this manual. The manual is
apportioned into several sections primarily according to pollutant type and
sample media analyzed. Sections I, II, and III address the measurement of EPA
designated priority pollutants and hazardous materials in aqueous, sediment,
and air samples, respectively. Rapid extraction techniques coupled with spec-
troscopic or chromatographic analysis permits detection of minute quantities of
hazardous substances in these sample matrices. Section IV is devoted entirely
to the analysis of petroleum oils in aqueous and sediment samples employing
synchronous excitation fluoresence spectroscopy. Section V focuses on protocols
for preparing analytical standards and synthetic samples, while Section VI
presents specialized sample preparation and treatment techniques. Several
miscellaneous, yet important, protocols and procedures appear in Section VII.
Quality Assurance/Quality Control protocols in Section VIII conclude the manual.
This manual is designed to be a comprehensive compilation of analytical methods
and techniques that are applicable for the rapid on-site characterization of
contaminated samples. Users are advised to acquire validation data concurrently
with sample measurements to document the reliability, reproducibility, and
sensitivity of these methods for their specific analysis requirements. The
reader will note that the methods are presented in a consistent format with
extensive cross referencing to other sections of the manual wherever applicable.
The layout and pagination scheme is specifically designed to facilitate updating
existing methods and incorporating additional procedures that will be forwarded
to all individuals receiving copies of this manual.
(ii)
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ERRATA SHEET
Although every effort has been made to assure the accuracy of the narratives
and terminology contained within the methods, inadverent errors and/or omissions
are sometimes unavoidable and often remain unnoticed. Consequently, we would
appreciate receiving your comments, corrections, recommendations, suggestions,
etc., so that the manual can be improved and updated accordingly. For your
convenience, we are including this errata sheet which may be photocopied and
forwarded with your corrections and/or comments to the following address:
U.S. Environmental Protection Agency
Hazardous Waste Engineering Research Laboratory
Releases Control Branch
Woodbridge Avenue
Edison, New Jersey
08837
ATTN: Michael Gruenfeld or David P. Remeta
CORRECTIONS
METHOD . ; PAGE - :
METHOD . ; PAGE - :
METHOD . ; PAGE - :
METHOD . ; PAGE - :
COMMENTS
(iii)
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TABLE OF CONTENTS
DISCLAIMER
ACKNOWLEDGMENTS
PREFACE
ERRATA SHEET
SECTION I: METHODOLOGY FOR THE ANALYSIS OF PRIORITY POLLUTANTS
AND HAZARDOUS MATERIALS IN AQUEOUS SAMPLES
METHOD 1.1 Protocol for Performing Purge and Trap Analyses
of Volatile Organic Compounds
METHOD 1.2 Rapid Extraction and Chromatographic Analysis
of Phenolic Compounds in Aqueous Samples
METHOD 1.3 Rapid Extraction and Chromatographic Analysis
of Base-Neutral Compounds in Aqueous Samples
METHOD 1.4 Rapid Extraction and Chromatographic Analysis
of Polychlorinated Biphenyls in Aqueous Samples
METHOD 1.5 Rapid Extraction and Chromatographic Analysis
of Organochlorine Pesticides in Aqueous Samples
METHOD 1.6 Rapid Extraction and Spectroscopic Analysis
of Fluorescing Organics in Aqueous Samples
METHOD 1.7 Rapid In-Situ Analysis of Hazardous Materials
in Aqueous Samples by Fluorescence Spectroscopy
METHOD 1.8 Rapid In-Situ Analysis of Quinoline in Aqueous
by Fluorescence Spectroscopy
(iv)
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SECTION II: METHODOLOGY FOR THE ANALYSIS OF PRIORITY POLLUTANTS
AND HAZARDOUS MATERIALS IN SEDIMENT SAMPLES
METHOD 2.1 Rapid Extraction and Chromatographic Analysis
of Polychlorinated Biphenyls in Sediment Samples
METHOD 2.2 Rapid Extraction and Spectroscopic Analysis
of Fluorescing Organics in Sediment Samples
SECTION III: METHODOLOGY FOR THE ANALYSIS OF PRIORITY POLLUTANTS
AND HAZARDOUS MATERIALS IN AIR SAMPLES
METHOD 3.1 Protocol for Performing Gas Chromatographic
Analysis of Air Samples
METHOD 3.2 Protocol for Preparing Analytical Standards of
Vaporized Materials
SECTION IV: METHODOLOGY FOR THE ANALYSIS OF PETROLEUM OILS IN
WATER AND SEDIMENT SAMPLES
METHOD 4.1 Rapid Extraction and Spectroscopic Analysis
of Petroleum Oils in Aqueous Samples
METHOD 4.2 Rapid Extraction and Spectroscopic Analysis
of Petroleum Oils in Sediment Samples
SECTION V: PROTOCOLS FOR THE PREPARATION OF ANALYTICAL STANDARDS
AND SYNTHETIC SAMPLES
METHOD 5.1 Protocol for Preparing Analytical Standards
of Hazardous and Carcinogenic Materials
METHOD 5.2 Protocol for Preparing Synthetic Samples of
Hazardous Materials in Water
METHOD 5.3 Protocol for Preparing Oil/Water Dispersions
by Ultrasonification
(v)
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SECTION VI: SPECIALIZED SAMPLE PREPARATION AND TREATMENT TECHNIQUES
METHOD 6.1 Protocol for the Selection and Use of Chromatographic
Cartridges for Environmental Sample Cleanup
METHOD 6.2 Protocol for Removing Elemental Sulfur from
Sediment Extracts
METHOD 6.3 Protocol for Weathering Small Quantities of
Petroleum Oil
METHOD 6.4 Protocol for Purifying Jsopropanol for
Fluorescence Analyses
SECTION VII: MISCELLANEOUS PROTOCOLS AND PROCEDURES
METHOD 7.1 Protocol for the Qualitative Evaluation of
Chromatograms of Polychlorinated Biphenyls
METHOD 7.2 Instructions for Packing and Conditioning Metal
Analytical Columns for Gas Chromatographs
METHOD 7.3 Instructions for Packing and Conditioning Glass
Analytical Columns for Gas Chromatographs
METHOD 7.4 Protocol for Checking the Performance of Glass
Analytical Columns for Gas Chromatographs
SECTION VIII: QUALITY ASSURANCE/QUALITY CONTROL PROTOCOLS
METHOD 8.1 Protocol for Establishing a Calibration Range
for Single Point Analysis
METHOD 8.2 Microcomputer Programs and Sample Analysis
Data Sheets
(vi)
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SECTION I
METHODOLOGY FOR THE ANALYSIS OF PRIORITY POLLUTANTS
AND HAZARDOUS MATERIALS IN AOIJEOUS SAMPLES
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CONTENTS
METHOD 1.1 Protocol for Performing Purge and Trap Analyses
of Volatile Organics
METHOD 1.2 Rapid Extraction and Chromatographic Analysis
of Phenolic Compounds in Aqueous Samples
METHOD 1.3 Rapid Extraction and Chromatographic Analysis
of Base-Neutral Compounds in Aqueous Samples
METHOD 1.4 Rapid Extraction and Chromatographic Analysis
of Polychlorinated Biphenyls in Aquoeus Samples
METHOD 1.5 Rapid Extraction and Chromatographic Analysis
of Pesticides in Aqueous Samples
METHOD 1.6 Rapid Extraction and Spectroscopic Analysis
of Fluorescing Organics in Aqueous Samples
METHOD 1.7 Rapid In-Situ Analysis of Hazardous Materials
in Aqueous Samples by Fluorescence Spectroscopy
METHOD 1.8 Rapid In-Situ Analysis of Quinoline in Aqueous
Samples by Fluorescence Spectroscopy
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METHOD 1.1 PROTOCOL FOR PERFORMING PURGE AND TRAP
ANALYSES OF VOLATILE ORGANIC COMPOUNDS
1. SCOPE AND SUMMARY:
1.1 This Method addresses the determination of volatile organic compounds
in aqueous samples. Specifically, the following classes of volatiles
designated as priority pollutants may be determined by this Method:
purgeable aromatics and purgeable halocarbons. A complete tabulation
of the 31 purgeable compounds is provided in Appendix A of this Method.
1.2 This Method provides a procedure for performing gas chromatographic
(GC) and gas chromatographic/mass spectrometer (GC/MS) analysis of
volatile organics in water samples by the purge and trap technique.
?.. CAUTIONS AND LIMITATIONS:
2.1 Care must be taken to ensure the cleanliness of glassware, syringes
and syringe needles. Rinse all equipment in metHanoi followed by
organic free distilled water. Store the solvent rinsed glassware
and equipment in an oven at approximately 110°C.
2.2 The preparation of standards and samples must be performed inside
a properly ventilated fume hood with appropriate protective apparel
(e.g., laboratory coat, safety glasses, chemical resistant gloves)
to minimize exposure of laboratory personnel to hazardous and/or
carcinogenic substances.
2.3 Standards must be prepared as needed and used immediately to prevent
volatility losses. Refrigerated calibration standards and samples
should be permitted to warm to room temperature before diluting or
purging.
2.4 Due to the volatile nature of the compounds analyzed by this Method,
all samples must be refrigerated at 4°C from the time of collection
to purge and trap analysis. The maximum holding time of the samples
is seven days.
2.5 The analyst should screen samples for unsuspected high concentrations
(i.e., greater than 1 ppm) of volatile organics by using the Procedure
for Screening Samples by Headspace Analysis (refer to Appendix D of
this Method).
2.6 In the event that the sample(s) contain(s) residual chlorine, 10 ug
of sodium thiosulfate (a preservative) must be added to the empty
40 ml sample bottles just prior to shipping to the sampling site.
1.1-1
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2.7 All 5.0 ml calibration standards, samples, and water blanks must be
dosed with a surrogate standard solution (refer to Appendix E of this
Method).
2.8 Purge a standard blank of methanol in organic free distilled water
prior to analysis of the calibration standards.
2.9 Invert the calibration standards and samples repeatedly to ensure
uniform mixing prior to subsampling and purging.
2.10 Purge an organic free distilled water blank (at a GC oven temperature
of 200°C) as necessary between samples to avoid cross contamination.
2.11 Examine the purge seal on the purge and trap unit daily and replace
if damaged (see Figure 3 in Appendix G of this Method).
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Balance, macro-analytical, range of 0 - 150 grams (± 0.1 mg)
2. Gas chromatograph (GC) equipped with a flame ionization detector,
or a gas chromatograph/mass spectrometer (GC/MS)
3. Gas line filters for removal of oil and water (Matheson Gas Products
Co., Model #451) and for removal of oxygen (Supelco Inc., Oxisorb)
4. Graduated cylinders, 100 ml
5. Hand crimpers for serum bottles
6. Headspace vials equipped with teflon lined screw caps
7. Micropipets, adjustable volume (Scientific Manufacturing Industries
Inc. or equivalent)
8. Pasteur pipets, disposable, 6 or 9 inch
9. Purge and trap unit (Hewlett-Packard Model 7675A or equivalent)
10. Purge vials with teflon lined caps
11. Ring stand with clamp
12. Sample bottles with teflon lined screw caps, 40ml
13. Serum bottles, crimp top, teflon septa, 100 ml and 50 ml (Wheaton or
equivalent)
14. Syringes, glass, 30 ml, 10 ml, 500 ul, 50 ul (Hamilton or equivalent)
15. Syringe needles, 2 inch, 1 inch
16. Volumetric flasks, 100 ml
17. Volumetric pipets, 100 ml, 50 ml
3.2 Reagents
1. Oistilled water, organic free
2. Helium gas, ultra-high purity
3. Internal standard of bromochloromethane, 1,4-dichlorobutane and
l-chloro-2-bromopropane (refer to Appendix E of this Method)
4. Methanol, spectrophotometric grade
1.1-2
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5. Pure material for standard preparation
6. Sodium thiosulfate, primary standard grade
4. PROCEniIRE:
4.1 Calibration Standard Preparation
4.1.1 Using a 100 ml graduated cylinder, transfer approximately 90 ml
of methanol into a 100 ml volumetric flask.
4.1.2 Using a macro-analytical balance and a 100 ul syringe, weigh
30 - 40 ul of the pure standard material and dispense this
quantity below the air/methanol interface in the flask.
NOTE 1: The analyst should consult Method 5.1 (Protocol
for Preparing Analytical Standards of Hazardous
and Carcinogenic Materials) for proper standard
preparation techniques.
NOTE 2: Standards must be prepared as needed and analyzed
immediately to prevent volatility losses.
4.1.3 Using a pasteur pipet, add a sufficient volume of methanol to
align the meniscus of the solution with the fiducial mark on
the flask. Stopper the flask and invert repeatedly to ensure
complete dissolution of the standard material.
4.1.4 Prepare calibration standards encompassing the concentration
range of 20 to 200 parts-per-billion in accordance with the
procedure described in Method 8.1 (Protocol for Establishing
a Calibration Range for Single Point Analysis).
1. Using a 50 ml or a 100 ml volumetric pipet, fill a serum
bottle (e.g., 50 ml or 100 ml) with organic free distilled
water and crimp the bottle cap.
2. Using a microliter glass syringe, inject an exact volume of
the stock standard solution below the air/water interface to
yield the desired calibration standard concentration:
Concentration x Injection Volume (ml) = Concentration of
Stock Standard Dilution Volume (ml) Standard (ug/L)
4.2 Analysis of Calibration Standards
4.2.1 Invert each calibration standard solution repeatedly to ensure
complete dissolution of the standard material.
1.1-3
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4.2.2 Secure the calibration solution bottle (e.g., CS/0 to a ring
stand and insert a 10 ml glass syringe (barrel closed) with a
two inch needle below the air/standard solution interface.
4.2.3 Insert a 30 ml glass syringe (barrel open) with a one inch
needle above the air/standard solution interface (see Figure
1 in Appendix G of this Method).
4.2.4 Positively displace 7 ml of the calibration standard solution
by injecting air into the CS/\ bottle with the 30 ml syringe
(i.e., adjust the volume of the calibration standard solution
in the 10 ml syringe to 7 ml).
4.2.5 Carefully withdraw the 10 ml syringe, invert to release trapped
air, and adjust the calibration standard solution volume in the
syringe to exactly 5.0 ml.
4.2.6 Slowly inject the 5.0 ml of standard solution into a clean, dry
purge vial. The analyst should also inject 5 ul of the prepared
surrogate standard stock solution (refer to Appendix E of this
Method) into the purge vial. Immediately screw on the teflon
lined cap to prevent volatility losses.
4.2.7 Repeat steps 4.2.2 - 4.2.6 for the complete set of calibration
standards (e.g., LL, LC, C$B, UC, UL).
4.2.8 Purge the calibration standard solutions prepared in this manner
employing the GC or GC/MS instrument conditions specified in
Appendices B and C, respectively, and the operating procedure
for the purge and trap provided in Appendix 0 of this Method.
4.3 Sample Preparation
4.3.1 In the event that the sample(s) contain(s) residual chlorine,
0.7 g of sodium thiosulfate (a preservative) must be added to
the 40 ml sampling bottles prior to collection of the sample(s).
4.3.2 The samples must be refrigerated at 4°C at all times prior to
purging and must be analyzed within seven days of collection.
4.3.3 The analyst should permit the sample(s) to warm to room temper-
ature and invert the sample bottle(s) repeatedly to ensure
uniform mixing prior to subsampling and purging.
4.4 Analysis of Samples
4.4.1 The analyst should screen the samples for unsuspected high con-
centrations of volatile organics by headspace analysis (refer
to the Procedure for Screening Samples by Headspace Analysis in
Appendix D of this Method).
1.1-4
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4.4.2 Secure the sample bottle to a ring stand and insert a 10 ml
glass syringe (barrel closed) with a two inch needle below the
air/sample solution interface.
4.4.3 Insert a 30 ml glass syringe (barrel open) with a one inch
needle above the air/sample solution interface (see Figure 2
in Appendix G of this Method).
4.4.4 Positively displace 7 ml of the sample by injecting a sufficient
volume of air into the sample bottle with the 30 ml syringe.
4.4.5 Carefully withdraw the 10 ml syringe, invert to release trapped
air and adjust the sample volume in the syringe to 5.0 ml.
4.4.6 Slowly inject the 5.0 ml aqueous sample into a clean, dry purge
vial. The analyst should also inject 5.0 ul of the prepared
surrogate standard stock solution (refer to Appendix E of this
Method) into the purge vial. Immediately screw on a teflon lined
cap to prevent volatility losses.
4.4.7 Repeat steps 4.4.1 - 4.4.6 for each of the aqueous samples.
4.4.8 Purge each of the aqueous samples prepared in this manner employ-
ing the GC or GC/MS instrument conditions specified in Appendices
B and C, respectively, and the operating procedure for the purge
and trap provided in Appendix F of this Method.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %} and the
precision of the analyses (e.g., Cv values < 3 %).
NOTE: The analyst should determine the precision of this Method by
calculating the Cv based on the response values of the surrogate
standards dosed into the LL, C$A, and UL calibration standards.
This precludes the need for replicate analyses of the calibration
solution as specified in Method 8.1.
5.3 Since the calibration range must bracket the'concentration(s) of the
volatile organics measured in the aqueous sample(s), the calibration
standard concentrations may require further adjustment resulting in
either a redefinition or refinement of the calibration range.
1.1-5
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5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
the detection limits for volatile organics, purgeable recoveries, and
potential interferences.
6. REFERENCE: Federal Register, Volume 49, Number 209, October 26, 1984,
Method 624.
1.1-6
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APPENDIX A
PUR6EABLE AROMATIC COMPOUNDS
Quantitative standards for analyzing aqueous samples for purgeable aromatics
are available from several commercial vendors. Supelco, Inc. provides the
following purgeable aromatic compounds Individually packaged in 1.0 ml ampuls
at a concentration of 200 ug/ml in methanol:
Benzene
Chlorobenzene
1,2-Dichlorobenzene
1,3-Dichlorobenzene
1,4-Oichlorobenzene
Ethyl benzene
Toluene
PURGEABLE HALOCARBON COMPOUNDS
Quantitative standards for analyzing aqueous samples for purgeable halocarbons
are available from several commercial vendors. Supelco, Inc. provides the
following purgeable halocarbon compounds individually packaged in 1.0 ml ampuls
at a concentration of 200 ug/ml in methanol:
Bromodichloromethane 1,1-Dichloroethane
Bromoform 1,2-Dichloroethane
Rromomethane 1,1-Dichloroethylene
Carbon tetrachloride trans-l,2-0ichloroethylene
Chlorobenzene 1,2-Dichloropropane
Chloroethane 1,3-Dichloropropene (0.4 mg/ml)
2-Chloroethyl vinyl ether Methylene chloride
Chioroform 1,1.2,2-Tetrachloroethane
Chioromethane Tetrachloroethy1ene
nibromochloromethane 1,1,1-Trichloroethane
1,2-Dichlorobenzene 1,1,2-Trichloroethane
1,3-Di chlorobenzene Tr1chloroethy1ene
1,4-DiChlorobenzene Trichlorofluoromethane
Dichlorodifluoromethane Vinyl chloride
(Source: Supelco, Inc., Catalog 25, 1987)
1.1-7
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Analytical Column: *
Carrier Gas:
Carrier Flow Rate:
Detector:
Detector Temperature:
Injector Temperature:
Oven Temperature:
Program Rate:
Time of Analysis:
APPENDIX B
GAS CHROMATOGRAPHIC CONDITIONS
1% SP-1000 on 60/80 Carbopack B, 6 ft x 2 mm ID,
glass column, 9 inch span
Helium
30 ml/min
Flame lonization (FID)
220°C
200°C
Initial = 60°C ; Final = 220°C
Initial Hold Time = 3 min.
8°C/nrin
Final Hold Time = 15 min.
38 min.
* The column recommended for volatile organic analyses should be packed and
conditioned in accordance with Method 7.3 (Instructions for Packing and
Conditioning Glass Analytical Columns for Gas Chromatography Instruments)
and performance tested according to the procedure and the specifications
described in Appendix D of Method 7.4 (Protocol for Performance Testing
Packed, Conditioned, Glass Analytical Columns for Gas Chromatography
Instruments).
PURGE & TRAP CONDITIONS
Desorb Temperature:
Desorb Time:
Prepurge Time:
200°C
4 min.
0 min.
Purge Time:
Valve Temperatures:
Vent Time:
12 min.
120°C
10 min.
1.1-8
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APPENDIX C
GAS CHROMATOGRAPH / MASS SPECTROMETER CONDITIONS
Analytical Column: * IX SP-1000 on 60/80 Carbopack B, 6 ft x 2 mm ID,
6 inch span
Carrier Flow Rate: 30 ml/min
Injector Temperature: 200°C
Maximum Oven Temperature: 225°C
Oven Temperature: Initial = 60°C ; Final = 220°C
Program Rate: Initial Hold Time = 3 min.
80°C/min
Final Hold Time = 15 min.
Run Time: 30 min.
Solvent Wait: 0 min.
* The column recommended for volatile organic analyses should be packed and
conditioned in accordance with Method 7.3 (Instructions for Packing and
Conditioning Glass Analytical Columns for Gas Chromatography Instruments)
and performance tested according to the procedure and the specifications
described in Appendix D of Method 7.4 (Protocol for Performance Testing
Packed, Conditioned, Glass Analytical Columns for Gas Chromatography
Instruments).
PURGE & TRAP CONDITIONS
Desorb Temperature: 200°C Purge Time: 12 min.
Desorb Time: 4 min. Valve Temperatures: 120°C
Prepurge Time: 0 min. Vent Time: 10 min.
1.1-9
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APPENDIX D
PROCEDURE FOR SCREENING SAMPLES BY HEADSPACE ANALYSIS
1. Standard Preparation and Analysis:
1.1 Prepare a 1.0 mg/L (ppm) standard solution of the suspect compound(s)
according to the procedure specified in Section 4.1 of this Method.
1.2 Establish the following gas chromatographic conditions:
INJECTOR TEMPERATURE 104°C
FID TEMPERATURE 200°C
OVEN TEMPERATURE 200°C
1.3 Withdraw 5.0 ml of the standard solution by positive displacement
(refer to Sections 4.2.2 - 4.2.5 of this Method) and quantitatively
transfer this volume to an 8 ml serum vial equipped with a septum
sealed screw cap. Agitate the contents of this vial vigorously for
approximately one minute.
1.4 Insert a 500 ul syringe above the air/standard solution interface in
the vial and withdraw 200 ul of the headspace. Inject this volume
into the gas chromatograph.
NOTE: Adjust the attenuation on the gas chromatograph to yield
a peak that is approximately 20% - 80% of full scale.
2. Sample Preparation and Analysis:
2.1 withdraw 5.0 ml of the sample solution(s) by positive displacement
(refer to Sections 4.2.2 - 4.2.5 of this Method) and quantitatively
transfer this volume to an 8 ml serum vial equipped with a septum
sealed screw cap. Agitate the contents of this vial vigorously for
approximately one minute.
2.2 Insert a 500 ul syringe above the air/sample solution interface in
the vial and withdraw 200 ul of the headspace. Inject this volume
into the gas chromatograph using the instrument conditions specified
for analysis of the standard solution.
2.3 Compare the GC response values for the 1.0 mg/L standard solution and
the sample(s).
1.1-10
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APPENDIX D (CONTINUED)
2.3.1 Purge the sample If its response value Is less than that of
the 1.0 ppm standard.
2.3.2 Prepare a 1:100 dilution of the sample if its response value
exceeds that of the 1.0 ppm standard. This sample dilution
must be screened in accordance with steps 2.1 - 2.3 outlined
in this Appendix.
NOTE: The analyst should prepare the 1:100 sample dilution
by withdrawing 1.0 ml of the sample via positive
displacement, and quantitatively transferring this
volume to a 100 ml serum bottle containing 99 ml of
organic free distilled water. This bottle should be
inverted repeatedly to ensure uniform mixing.
2.3.3 Purge the sample dilution if its response value is less than
that of the 1.0 ppm standard.
1.1-11
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APPENDIX E
SURROGATE STANDARD DOSING PROCEDURE
1. OVERVIEW: This procedure provides a means for verifying that the purge and
trap unit is functioning properly and may also be employed to
determine the effectiveness of the method in dealing with each
sample matrix encountered.
2. CAUTIONS AND LIMITATIONS:
2.1 The surrogate standard stock solution must be added to each 5 ml
standard, sample and water blank.
2.2 The surrogate standard stock solution must be refrigerated at 4°C.
2.3 This procedure serves as a check only for adverse matrix effects and
malfunctioning of the purge and trap unit.
3. REAGENTS:
3.1 Distilled water, organic free
3.2 Internal standard of bromochloromethane, 1,4-dichlorobutane and
l-chloro-2-bromopropane (20 mg/ml each in 1.0 ml methanol)
4. PROCEDURE:
4.1 Prepare a surrogate standard stock solution with a concentration of
100 mg/L for each of the three components.
4.2 Using a 10 ul syringe, dose 5 ul of the surrogate standard stock
solution into each 5 ml standard, sample and water blank immediately
prior to analysis.
4.3 Compare all standards and samples to a 5 ml water blank dosed with
5 ul of the surrogate stock (e.g., 100% response). The purge and
trap unit is functioning properly when the responses of the standards
and samples are within the 90% - 110% range of the water blank response.
NOTE: If the response of the surrogate standards is not within the
90% - 110% range of the surrogate stock response in the water
blank, repeat the dosed sample or standard analyses.
1.1-12
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APPENDIX F
OPERATING PROCEDURE FOR THE HEWLETT PACKARD 7675A PURGE AND TRAP UNIT
1. Assure that a purgeable column Is Installed in the gas chromatograph (refer
to Appendix B).
2. Examine the purge seal and replace if damaged (see Figure 3 in Appendix G).
3. Press the LINE button and verify that:
3.1 The READY light is on.
3.2 The LINE button is lit.
3.3 Air flow is audible.
4. Assure that the purge flow is 40 ml/min as measured by the bubble flowmeter.
NOTE: Measure flow from the purge tube (see Figure 3 in Appendix G).
5. Swing open the purge unit and establish the following conditions:
DESORR TIME 0 min
PREPURGE TIME 0 min
PURGE TIME 0 min
VENT TIME 1 min
6. Press the START RUN button on the purge and trap unit and assure that the
upper actuator valve rotates (see Figure 4 in Appendix G).
NOTE: If the valve does not rotate, refer to the HP 7675A
Operating and Service Manual (pages 2-15).
7. Press the STOP RUN button on the purge and trap unit.
8. Establish the following purge and trap conditions:
DESORB TEMPERATURE 2008C
DESORB TIME 4 min.
PREPURGE TIME 0 min
PURGE TIME 12 min
VALVE TEMPERATURES 120°C
VENT TIME 10 min
9. Screw a purge vial containing a water blank onto the purge cap.
NOTE: To avoid breakage, do not tighten purge vials excessively.
1.1-13
-------
APPENDIX F (CONTINUED)
10. Press LIST LIST on the GC terminal to verify proper instrument operating
conditions (refer to Appendix B).
11. Assure that the AUXILIARY DEMAND light on the purge and trap unit is flashing.
12. Press the START RUN button on the purge and trap unit; the GC data system is
activated automatically when the PURGE cycle changes to DESORB.
13. The presence of large peaks at ATTN 2^ (other than those of the surrogate
standard) is indicative of contaminants in the water blank. Repeat steps
9-12 again, and if the contamination is serious, refer to the HP 7675A
Operating and Service Manual (pages 4.4 - 5.5).
NOTE; Possible sources of contamination include the carrier gas, line
filters, chemical filter, tenax column, and the GC column. The
analyst should consult Figure 5 in Appendix G for sample chroma-
tograms illustrating potential sources of contamination.
14. Standards and samples must be analyzed under identical instrumental
conditions.
15. The following procedure should be used when operating the purge and trap
unit in conjunction with the GC/MS.
15.1 Follow steps 1 - 7 of this Appendix.
15.2 Establish the following purge and trap conditions:
DESORB TEMPERATURE 200°C
DESORB TIME 4 min.
PREPIIRGE TIME 0 min.
PURGE TIME 12 min.
VALVE TEMPERATURES 120°C
VENT TIME 15 min.
15.3 Screw a purge vial containing a water blank onto the purge cap. To
avoid breakage, do not tighten purge vials excessively.
15.4 Insert the flexible disk labeled "PDATA #1, 10/79" into the disk
drive. Press [get "START" EXECUTE CONTINUE]
1.1-14
-------
APPENDIX 6
Figure 1
Figure 2
1.1-15
-------
APPENDIX 6 (CONTINUED)
PURGE CAP
PURGETUBE
PURGEVESSEL
Figure 4
Figure 3
1.1-16
-------
APPENDIX 6 (CONTINUED)
Good Baseline; Well-conditioned Column.
•- -.
• =
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.-
j ;ijo if a
!) iTO ifO
3. -3
.'
* v'.J
•?.v>? • £ T
i. i.V - '. T-
'. :o •-• s T
nnt»: TttW 1 on tun nm o» W °C.
Contaminants in Tank Gas and Purge Gas Line Contamination in Column FlowlSas
Figure 5
1.1-17
-------
METHOD 1.2 RAPID EXTRACTION AND CHROMATOGRAPHIC ANALYSIS
OF PHENOLIC COMPOUNDS IN AQUEOUS SAMPLES
1. SCOPE AND SUMMARY:
1.1 This Method addresses the determination of phenols in aqueous samples.
Specifically, the following phenolic compounds designated as priority
pollutants may be determined by this method: 4-chloro-3-methylphenol;
2-chlorophenol; 2,4-dichlorophenol; 2,4-dimethylphenol; 2,4-dinitro-
phenol; 2-methyl-4,6-dinitrophenol; 2-nitrophenol; 4-nitrophenol;
pentachlorophenol; phenol; and, 2,4,6-trichlorophenol.
1.2 This Method provides a procedure for rapidly extracting phenolics from
aqueous samples and quantifying the extracts via gas chromatographic
analysis. A 1.0 liter sample of wastewater is pretreated by addition
of sodium chloride and acidification to pH = 2.0 to ensure complete
extraction of the phenolic compounds. A co-solvent mixture of 30:70
methylene chlorideihexane is employed to extract the sample via a
magnetic spinning technique. Flame ionization gas chromatographic
conditions are described which permit direct measurement of phenolic
compounds in t.he extract.
2. CAUTION AND LIMITATIONS:
2.1 Proper pH adjustment of the samples is essential to ensure complete
extraction of phenolic compounds.
?..?. The extraction methodology prescribes a 'salting out1 technique to
reduce the solubility of methylene chloride in the aqueous samples.
The analyst should note that a finite amount of methylene chloride is
still soluble in water under these conditions, resulting in somewhat
higher recoveries of phenolic compounds due to a slight reduction in
the volume of the organic phase.
2.3 All glassware employed for standard and sample preparation must be
rinsed three times with hexane and air dried prior to use.
2.4 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., safety glasses, laboratory coat, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous and/or carcino-
genic substances.
2.5 The spinning extraction procedure must be performed behind a safety
shield in a properly ventilated fume hood.
1.2-1
-------
2.6 The 1.0 liter sample bottles must be vented periodically to release
any pressure buildup during the extraction procedure.
2.7 The gas chromatographic conditions listed in Appendix B of this
Method must be established at least two hours prior to analysis of
the calibration standards and sample extracts to permit sufficient
stabilization of the flame ionization detector.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Autosampler vials equipped with teflon lined septa
2. Beakers, assorted volumes
3. Gas chromatograph equipped with flame ionization detector
4. Graduated cylinders, 1 L and 100 ml
5. Hand crimpers for autosampler vials and serum bottles
6. Magnetic stirring bars, teflon coated, 2 inch
7. Magnetic stirring unit
8. Pasteur pipets, disposable, 6 or 9 inch
9. pH paper, sensitive to pH 2.0
10. Sample bottles equipped with teflon lined screw caps, 1 L (Wheaton
or equivalent)
11. Serum bottles, crimp top, 100 ml and 30 ml (Wheaton or equivalent)
12. Syringes, 500 ul, 100 ul and 10 ul (Hamilton or equivalent)
13. Volumetric flasks, 1.0 L, 100 ml, 50ml, and 25 ml
14. Volumetric pipets, 25ml
3.2 Reagents
1. Phenol Standard Mixture (Supelco, Inc. or equivalent)
2. Hexane, pesticide grade (Burdick ft Jackson or equivalent)
3. Methylene chloride, pesticide grade (Burdick & Jackson or equivalent)
4. Hydrochloric acid, 1.0 Normal solution
5. Distilled, deionized water
6. Sodium Chloride
4. PROCEDURE:
4.1 Standard Preparation
4.1.1 Using a small beaker or pasteur pipet, transfer approximately
4 ml of a 30:70 methylene chloride:hexane solvent mixture to a
5 ml volumetric flask.
1.2-2
-------
NOTE: The analyst should prepare the methylene chloride:hexane
solvent mixture according to the procedure described in
Appendix A of this Method.
4.1.2 Using a. 500 ul syringe, quantitatively withdraw 400.0 ul of the
Phenolic Standard Mixture described in Appendix A and dispense
this volume below the air/solvent interface in the flask.
NOTE: The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
4.1.3 Using a pasteur pipet, add a sufficient volume of the methylene
chloridethexane solvent mixture to align the meniscus of the
solution with the fiducial nark on the flask. Stopper the
flask and invert repeatedly to ensure complete dissolution of
the phenolic compounds.
NOTE: Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and calibration standards by
diluting the stock standard solution appropriately. The cali-
bration range should be established so as to incorporate the
expected concentrations of the samples (refer to Method 8.1 -
Protocol for Establishing a Calibration Range for Single Point
Analysis).
4.2 Sample Preparation
4.2.1 Record individual sample volumes via the following procedure:
1. Prior to extracting the samples, mark the height of the men-
iscus on the exterior of each sample bottle with an indelible
marker or masking tape.
?.. Extract the samples in accordance with Sections 4.2.2 - 4.2.8
of this Method and transfer the supernatants (e.g., methylene
chloride:hexane extracts) to 30 ml crimp-top serum bottles.
3. Empty the aqueous contents of each sample hottle into an
appropriate storage or waste container.
4. Dispense an exact volume of distilled water from a 1.0 L
graduated cylinder into each sample bottle.
5. Record the volume of distilled water required to align the
meniscus with the marker on each sample bottle.
4.2.2 Add 350 grams of sodium chloride to each of the sample bottles.
Secure the screw caps tightly and invert the bottles several
times to ensure complete dissolution of the salt.
1.2-3
-------
NOTE; The addition of sodium chloride to the aqueous samples
prior to extraction serves to reduce the solubility of
methylene chloride in water.
4.2.3 Acidify each of the samples to pH 2.0 by adding an appropriate
volume of 1.0 Normal hydrochloric acid solution to the bottles.
The acidity of the samples should be measured with pH paper.
4.2.4 Place a 2 inch teflon coated magnetic stirring bar into each of
the sample bottles.
4.2.5 Using a 25 ml volumetric pipet, quantitatively transfer 25.0 ml
of the 30:70 methylene chloride:hexane solvent mixture to each
1.0 liter sample bottle and stopper the bottles immediately to
prevent volatility losses of the extraction solvent.
4.2.6 Place the sample bottles on a magnetic stirring unit positioned
behind a safety shield in a fume hood. Adjust the stir bar
spinning rates to achieve a maximum 'coning1 or 'whirlpool'
effect in the sample bottles. The samples should be extracted
in this manner for exactly 30 minutes.
NOTE 1: During the spinning extraction procedure, the analyst
should vent the sample bottles periodically to release
any pressure buildup.
NOTE 2: The analyst should visually inspect the sample bottles
and adjust the spinning rates as necessary to prevent
stir bar "bumping" and to minimize the formation of
emulsions.
4.2.7 After 30 minutes, the magnetic stirring unit should be turned
off and the aqueous and organic layers permitted to separate
for approximately 10 minutes. Using a small beaker or graduated
cylinder, add a sufficient volume of distilled water to each
sample to force the organic phase into the neck of the sample
bottle.
4.2.8 Using pasteur pipets, transfer the solvent extracts containing
the phenolic compounds to 30 ml crimp-top serum bottles.
4.2.9 Measure each sample volume in accordance with Section 4.2.1
(steps 3 - 5) of this Method.
4.3 Gas Chromatographic Analysis
4.3.1 Establish the gas Chromatographic conditions listed in Appendix
B of this Method.
NOTE: The flame ionization detector requires a minimum two hour
stabilization period prior to analysis of the calibration
standards and sample extracts.
1.2-4
-------
4.3.2 Using pasteur pipets, transfer approximately 1 ml of each of
the calibration standards and sample extracts to autosampler
vials. Seal the autosampler vials with a hand crimper and
position the vials in the autoinjector with 30:70 methylene
chloride:hexane blanks inserted at regular intervals.
NOTE: In the event that an autoinjector is unavailable, the
analyst should use a precision quality 10 ul syringe to
manually inject an exact volume (e.g., usually 3 to 5 ul)
of all standard, sample, and solvent solutions.
4.3.3 Replicate injections of all calibration standards and sample
extracts are required. A minimum of three injections of each
solution are necessary to calculate its coefficient of variation
(Cv), which is a measure of precision or reproducibility.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %).
5.3 Since the calibration range must bracket the concentration(s) of the
phenolic compounds measured in the sample extract(s), the calibration
standard concentrations may require further adjustment resulting in
either a redefinition or refinement of the calibration range.
5.5 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
phenol detection limits, extraction recoveries, and potential inter-
ferences.
1.2-5
-------
APPENDIX A
PREPARATION OF A 30:70 (V;V) METHYLENE CHLORIDE:HEXANE MIXTURE
1. Dispense 300 ml of methylene chloride and 700 ml of hexane into separate
1.0 liter graduated cylinders.
NOTE: Due to contraction, expansion, etc., associated with the mixing of
two organic phases, volume addition is not necessarily additive;
therefore, the volumes of methylene chloride and hexane must be
measured separately prior to actual preparation of the mixture.
2. Transfer the 300 ml of methylene chloride from the graduated cylinder to
a 1.0 liter volumetric flask. Carefully dispense the 700 ml of hexane to
the volume of methylene chloride in the volumetric flask.
3. Stopper the flask and invert repeatedly to ensure uniform mixing of the
two organic solvents. The analyst should periodically vent the flask to
release any pressure generated by this mixing process.
NOTE: DO NOT adjust the volume of solution to align the meniscus with the
fiducial mark on th.e volumetric flask. The prepared solution is
30% methylene chloride : 70% hexane (volume:volume).
PHENOLIC STANDARD MIXTURE
Quantitative standards for analyzing aqueous samples for phenolic compounds
are available from several commercial vendors. Supelco, Inc. provides the
following phenolic mixture in a 1.0 ml ampul at the specified concentrations
in methylene chloride:
Component Concentration
4-Chloro-3-methylphenol 2500 ug/ml
2-Chlorophenol 500 ug/ml
?,4-nichlorophenol 500 ug/ml
2,4-Dimethylphenol 500 ug/ml
2,4-Pinitrophenol 1500 ug/ml
2-Methyl-4,6-dinitrophenol 2500 ug/ml
2-Nitrophenol 500 ug/ml
4-Nitrophenol 2500 ug/ml
Pentachlorophenol 2500 ug/ml
Phenol 500 ug/ml
2,4,6-Trichlorophenol 1500 ug/ml
(Source: Supelco, Inc., Catalog 24, 1986)
1.2-6
-------
APPENDIX B
GAS CHROMATOGRAPHIC CONDITIONS
Analytical Column: * 1% SP-1240-DAt on 100/120 Supelcoport, 3 ft x 2 mm
glass column (phosphoric acid treated glass wool
at both ends of the column)
Carrier Gas: Helium
Carrier Flow Rate: 35 ml/min.
Detector: Flame lonization (FID)
Detector Temperature: 260°C
Injector Temperature: 180°C
Oven Temperature: Initial = 85°C ; Final = 200°C
Program Rate: 10°C/min. ; Final Hold Time = 5 min.
Time of Analysis: 16.5 min.
* The column recommended for phenolic analyses should be packed and
conditioned in accordance with Method 7.3 (Instructions for Packing
and Conditioning Glass Analytical Columns for Gas Chromatography
Instruments) and performance tested according to the procedure and
specifications described in Appendix C of Method 7.4 (Protocol for
Performance Testing Packed, Conditioned, Glass Analytical Columns
for Gas Chromatography Instruments).
t NOTE: DA designates that the column packing material is deactivated
Tor the analysis of acidic compounds.
1.2-7
-------
METHOD 1.3 RAPID EXTRACTION AND CHROMATOGRAPHIC ANALYSIS
OF BASE-NEUTRAL COMPOUNDS IN AQUEOUS SAMPLES
1. SCOPE AND SUMMARY:
1.1 This method addresses the determination of Base-Neutral compounds in
aqueous samples. Base-Neutrals comprise a broad array of compound
classes that are extractable in an alkaline media. Appendix A
provides a tabulation of the 46 Base-Neutral extractables that are
designated as priority pollutants and may be determined by this
Method.
1.2 This method provides a procedure for rapidly extracting Base-Neutral
compounds from aqueous samples and quantifying the extracts via gas
chromatographic analysis. A 1.0 liter sample of wastewater is pre-
treated by adjusting its alkalinity to pH 11.0 to ensure complete
extraction of the basic compounds. The alkaline sample is extracted
with hexane using a magnetic spinning technique. Flame ionization
gas chromatographic conditions are described which permit direct
measurement of the Base-Neutral compounds in the extract.
2. CAUTIONS AND LIMITATIONS:
2.1 Proper pH adjustment of the samples is essential to ensure complete
extraction of basic compounds. The analyst should note that alkaline
adjustment causes many inorganic substances (e.g., iron salts) to
precipitate out of solution.
2.2 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., safety glasses, laboratory coat, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous and/or carcino-
genic substances.
2.3 The spinning extraction procedure must be performed behind a safety
shield in a properly ventilated fume hood.
2.4 The 1.0 liter sample bottles must be vented periodically to release
any pressure buildup during the extraction procedure.
2.5 The gas chromatographic conditions listed in the Appendix of this
Method must be established at least two hours prior to analysis of
the calibration standards and sample extracts to permit sufficient
stabilization of the flame ionization detector.
2.6 Sample extracts should be 'screened' by GC flame ionization detection
-------
and subsequently analyzed by GC electron capture detection if enhanced
sensitivity is desired.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Autosampler vials equipped with teflon lined septa
2. Beakers, assorted volumes
3. Distil led water
4. Gas chromatograph equipped with flame ionization and electron
capture detectors
5. Graduated cylinders, 1 L
6. Hand crimpers for autosampler vials and serum bottles
7. Magnetic stirring bars, teflon coated, 2 inch
8. Magnetic stirring unit
9. Pasteur pipets, disposable, 6 or 9 inch
10. pH paper, sensitive to pH 11.0
11. Sample bottles equipped with teflon lined screw caps, 1 L (Wheaton
or equivalent)
12. Serum bottles, crimp top, 30 ml (Wheaton or equivalent)
13. Syringes, 100 ul and 10 ul (Hamilton or equivalent)
14. Volumetric flasks, 100 ml, 50ml, 25 ml, 10 ml and 5 ml
15. Volumetric pipets, 20 ml
3.2 Reagents
1. Base-Neutral Standard Mixtures (Supelco, Inc. or equivalent)
2. Distilled water, organic free
3. Hexane, pesticide grade (Burdick & Jackson or equivalent)
4. Sodium hydroxide, 1.0 Normal solution
4. PROCEDURE
4.1 Standard Preparation
4.1.1 Using a small beaker or pasteur pipet, transfer approximately
4 ml of hexane to a 5 ml volumetric flask.
4.1.2 Using a 1.0 ml syringe, quantitatively withdraw 500.0 ul of one
of the Base-Neutral Standard Mixtures described in Appendix A
of this Method and dispense this volume below the air/hexane
interface in the flask.
NOTE: The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
1.3-2
-------
4.1.3 Using a pasteur pipet, add a sufficient volume of hexane to
align the meniscus of the solution with the fiducial mark on
the flask. Stopper the flask and invert repeatedly to ensure
complete dissolution of the Base-Neutrals. The concentration
of this stock standard solution is 20.0 ug/ml.
NOTE: Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and calibration standards by
diluting the stock standard solution appropriately. The cali-
bration range should be established so as to incorporate the
expected concentrations of the samples (refer to Method 8.1 -
Protocol for Establishing a Calibration Range for Single Point
Analysis).
4.2 Sample Preparation
4.2.1 Record individual sample volumes via the following procedure:
1. Prior to extracting the samples, mark the height of the
meniscus on the exterior of each sample bottle with an
indelible marker or masking tape.
2. Extract the samples in accordance with Sections 4.2.2 -
4.2.7 of this Method and transfer the hexane extracts to
30 ml crimp-top serum bottles.
3. Empty the aqueous contents of each sample bottle into an
appropriate storage or waste container.
4. Dispense an exact volume of distilled water from a 1.0 L
graduated cylinder into each sample bottle.
5. Record the volume of distilled water required to align the
meniscus with the marker on each sample bottle.
4.2.2 Adjust each of the aqueous samples to pH 11.0 by the addition of
an appropriate volume of 1.0 Normal sodium hydroxide solution
to the bottles. Monitor the alkalinity of the samples with pH
paper.
4.2.3 Place a 2 inch teflon coated magnetic stirring bar into each of
the sample bottles.
4.2.4 Using a 20 ml volumetric pipet, quantitatively transfer 20.0 ml
of hexane to each 1.0 liter sample bottle and stopper these
bottles immediately to prevent potential volatility losses of
the extraction solvent.
1.3-3
-------
4.2.5 Place the sample bottles on a magnetic stirring unit positioned
behind a safety shield in a fume hood. Adjust the stir bar
spinning rates to achieve a maximum 'coning' or 'whirlpool'
effect in the sample bottles. The samples should be extracted
in this manner for exactly 30 minutes.
NOTE 1: During the spinning extraction procedure, the analyst
should vent the sample bottles periodically to release
any pressure buildup.
NOTE 2: The analyst should visually inspect the sample bottles
and adjust the spinning rates as necessary to prevent
stir bar "bumping" and to minimize the formation of
emulsions.
4.2.6 After 30 minutes, the magnetic stirring unit should be turned
off and the aqueous and organic layers permitted to separate
for approximately 10 minutes. Using a small beaker or graduated
cylinder, add a sufficient volume of distilled water to each
sample to force the organic phase into the neck of the sample
bottle.
4.2.7 Using pasteur pipets, transfer the hexane extracts containing
the Base-Neutral compounds to 30 ml crimp-top serum bottles.
4.2.8 Measure each sample volume in accordance with Section 4.2.1
(steps 3 - 5) of this Method.
4.3 Gas Chromatographic Analysis
4.3.1 Establish the gas Chromatographic conditions listed in Appendix
B of this Method.
NOTE: The flame ionization detector requires a minimum two hour
stabilization period prior to analysis of the calibration
standards and sample extracts.
4.3.2 Using disposable pasteur pipets, transfer approximately 1 ml of
each of the calibration standards and sample extracts to auto-
sampler vials. Seal the autosampler vials with a hand crimper
and position the vials in the autoinjector with hexane blanks
inserted at regular intervals.
NOTE: In the event that an autoinjector is unavailable, the
analyst should use a precision quality 10 ul syringe to
manually inject an exact volume (e.g., usually 3 to 5 ul)
of all standard, sample, and hexane solutions.
4.3.3 Replicate injections of all calibration standards and sample
extracts are required. A minimum of three injections of each
solution are necessary to calculate its coefficient of variation
(Cv), which is a measure of precision or reproducibility.
1.3-4
-------
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %).
5.3 Since the calibration range must bracket the concentration(s) of the
Base-Neutral compound(s) measured in the sample extract(s), the cali-
bration standard concentrations may require further adjustment result-
ing in either a redefinition or refinement of the calibration range.
5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
Base-Neutral detection limits, extraction recoveries, and potential
interferences.
1.3-5
-------
APPENDIX A
BASE-NEUTRAL STANDARD MIXTURES
Quantitative standards for analyzing aqueous samples for Base-Neutrals
are available from several commercial vendors. Supelco, Inc. provides
the following Base-Neutral Extractables Mixtures in 1.0 ml amber ampuls
at a concentration of 200 ug/ml in methylene chloride:
BASE-NEUTRALS MIX 1
Acenaphthylene
Benzo(b)f1uoranthene
Bis(2-chloroethyl)ether
Bi s(2-ethylhexyl)phthal ate
Bis(2-chloroisopropyl)ether
4-Bromophenylphenyl ether
1,4-Dichlorobenzene
3,3'-Dichlorobenzdine
Dimethyl phthalate
Di-n-butyl phthalate
2,6-Dinitrotoluene
Nitrobenzene
BASE-NEUTRALS MIX 2
Acenaphthene
Anthracene
Benzo(a)anthracene
Bis(2-chloroethoxy)methane
Chrysene
Dibenzo(a,h)anthracene
1,2-Dichlorobenzene
1,3-Dichlorobenzene
Diethyl phthalate
2,4-Dinitrotoluene
Fluorene
Hexachlorobenzene
Hexachlorobutadiene
Naphthalene
Pyrene
BASE-NEUTRALS MIX 3
Azobenzene
Benzyl butyl phthalate
2-r.hloronaphthalene
Fluoranthene
Hexachlorocyclopentadi ene
Hexachloroethane
Isophorone
N-Ni trosodi-n-propylami ne
N-Ni trosodiphenylami ne
Phenanthrene
1,2,4-Trichlorobenzene
BASE-NEUTRALS MIX 4
Benzidine
Benzo(a)pyrene
Benzo(g,h,i)perylene
Benzo(k)f1uoranthene
4-Chlorophenylphenyl ether
Di-n-octyl phthalate
Indeno(l,2,3-cd)pyrene
N-Nitrosodimethylamine
(Source: Supelco, Inc., Catalog 25, 1987)
1.3-6
-------
APPENDIX B
GAS CHROMATOGRAPHIC CONDITIONS
Analytical Column: *
Carrier Gas:
Carrier Flow Rate:
Detector:
Detector Temperature:
Injector Temperature:
Oven Temperature:
Program Rate:
Time of Analysis:
1.5* SP 2250/1.95* SP 2401 on 100/120 Supelcoport
6 ft x 4 mm glass column (mixed phase support)
Helium or Nitrogen
30 ml/min
Flame lonization (FID)
250°C
220°C
Initial = 508C ; Final = 260°C
8°C/min.
Variable (Dependent on Base-Neutrals Analyzed)
The mixed phase column recommended for Base-Neutral analyses should be
packed and conditioned in accordance with Method 7.3 (Instructions for
Packing and Conditioning Glass Analytical Columns for Gas Chromatography
Instruments) and performance tested according to the procedure and the
specifications described in Appendix 8 of Method 7.4 (Protocol for Per-
formance Testing Packed, Conditioned, Glass Analytical Columns for Gas
Chromatography Instruments).
1.3-7
-------
METHOD 1.4 RAPID EXTRACTION AND CHROMATOGRAPHIC ANALYSIS OF
POLYCHLORINATED BIPHENYLS IN AQUEOUS SAMPLES
1. SCOPE AND SUMMARY:
1.1 This method addresses the determination of polychlorinated biphenyl
(PCB) compounds. Specifically, the following PCB Aroclors may be
determined by this method: Aroclor 1016; Aroclor 1221; Aroclor 1232;
Aroclor 1242; Aroclor 1248; Aroclor 1254; and, Aroclor 1260.
1.2 This method provides a procedure for rapidly extracting PCBs from
aqueous samples and quantifying the extracts via gas chromatographic
analysis. A 1.0 liter sample of wastewater 1s extracted with hexane
using a magnetic spinning technique. Electron capture gas chromato-
graphic conditions are described which permit direct measurement of
the PCBs in the extract.
2. CAUTIONS AND LIMITATIONS:
2.1 All glassware employed for standard and sample preparation must be
rinsed three times with hexane and air dried prior to use.
2.2 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., safety glasses, laboratory coat, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous and/or carcino-
genic substances.
2.3 The spinning extraction procedure must be performed behind a safety
shield in a properly ventilated fume hood.
2.4 The 1.0 liter sample bottles must be vented periodically to release
any pressure buildup during the extraction procedure.
2.5 The gas chromatographic conditions listed in Appendix B of this
Method must be established at least two hours prior to analysis of
the calibration standards and sample extracts to permit sufficient
stabilization of the electron capture detector.
2.6 The analyst should refer to Method 7.1 (Protocol for the Qualitative
Evaluation of Chromatograms of Polychlorinated Biphenyls) for proper
interpretation of the PCB chromatograms.
1.4-1
-------
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Autosampler vials equipped with teflon lined septa
2. Beakers, assorted volumes
3. Gas chromatograph equipped with electron capture detector
4. Graduated cylinders, 1 L
5. Hand crimpers for autosampler vials and serum bottles
6. Magnetic stirring bars, teflon coated, 2 inch
7. Magnetic stirring unit
8. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
9. Pasteur pi pets, disposable, 6 or 9 inch
10. Sample bottles equipped with teflon lined screw caps, 1 L (Wheaton
or equivalent)
11. Serum bottles, crimp top, 30 ml (Wheaton or equivalent)
12. Syringes, 200 ul and 10 ul (Hamilton or equivalent)
13. Volumetric flasks, 100 ml, 50ml, and 25 ml
14. Volumetric pi pets, 20 ml
3.2 Reagents
1. PCB Aroclor Standards (Supelco, Inc. or equivalent)
2. Hexane, chromatographic grade (Burdick ft Jackson or equivalent)
3. Distilled water, organic free
4. PROCFDURE:
4.1 Standard Preparation
4.1.1 Using a volumetric pipet or a beaker, transfer approximately
20 ml of hexane to a 25 ml volumetric flask.
4.1.2 Using a micropipet or 200 ul syringe, quantitatively withdraw
150.0 ul of an Aroclor standard containing 200 ug/ml of PCBs and
dispense this volume below the air/hexane interface in the flask
(refer to Appendix A of this Method for a list of PCR Aroclors).
NOTE: The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
4.1.3 Using a pasteur pipet, add a sufficient volume of hexane to align
the meniscus of the solution with the fiducial mark on the flask.
Stopper the flask and invert repeatedly to ensure complete dis-
solution of the PCBs. The concentration of this stock standard
solution is 1200 ng/ml (parts per billion) in PCBs.
1.4-2
-------
NOTE: Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and calibration standards by
diluting the stock standard solution appropriately. The cali-
bration range should be established so as to incorporate the
expected concentrations of the samples (refer to Method 8.1 -
Protocol for Establishing a Calibration Range for Single Point
Analysis).
4.2 Sample Preparation
4.2.1 Record individual sample volumes via the following procedure:
1. Prior to extracting the samples, mark the height of the men-
iscus on the exterior of each sample bottle with an indelible
marker or masking tape.
2. Extract the samples in accordance with Sections 4.2.2 - 4.2.6
of this Method and transfer the supernatants (e.g., hexane
extracts) to 30 ml crimp-top serum bottles.
3. Empty the aqueous contents of each sample bottle into an
appropriate storage or waste container.
4. Dispense an exact volume of distilled water from a 1.0 L
graduated cylinder into each sample bottle.
5. Record the volume of distilled water required to align the
meniscus with the marker on each sample bottle.
4.2.2 Place a 2 inch teflon coated magnetic stirring bar into each of
the sample bottles.
4.2.3 Using a 20 ml volumetric pipet, quantitatively transfer 20.0 ml
of hexane to each 1.0 L sample bottle and stopper the bottles
immediately to prevent volatility losses of the extraction
solvent.
4.2.4 Place the sample bottles on a magnetic stirring unit positioned
behind a safety shield in a fume hood. Adjust the stir bar
spinning rates to achieve a maximum 'coning' or 'whirlpool'
effect in the sample bottles. The samples should be extracted
in this manner for exactly 30 minutes.
NOTE 1: During the spinning extraction procedure, the analyst
should vent the sample bottles periodically to release
any pressure buildup.
NOTE 2: The analyst should visually inspect the sample bottles
and adjust the spinning rates as necessary to prevent
stir bar "bumping" and to minimize the formation of
emulsions.
1.4-3
-------
4.2.5 After 30 minutes, the magnetic stirring unit should be turned
off and the aqueous and organic layers permitted to separate
for approximately 10 minutes. Using a small beaker or graduated
cylinder, add a sufficient volume of distilled water to each
sample to force the organic phase into the neck of the sample
bottle.
4.2.6 Using pasteur pi pets, transfer the hexane extracts containing
the PCBs to 30 ml crimp-top serum bottles.
4.2.7 Measure each sample volume in accordance with Section 4.2.1
(steps 3 - 5) of this Method.
4.3 Gas Chromatographic Analysis
4.3.1 Establish the gas Chromatographic conditions listed in Appendix
B of this Method.
NOTE: The electron capture detector requires a minimum two hour
stabilization period prior to analysis of the calibration
standards and sample extracts.
4.3.2 Using pasteur pipets, transfer approximately 1 ml of each of the
calibration standards and sample extracts to autosampler vials.
Seal the autosampler vials with a hand crimper and position the
vials in the autoinjector with hexane blanks inserted at regular
intervals.
NOTE: In the event that an autoinjector is unavailable, the
analyst should use a precision quality 10 ul syringe to
manually inject an exact volume (e.g., usually 3 to 5 ul)
of all standard, sample, and hexane solutions.
4.3.3 Replicate injections of all calibration standards and sample
extracts are required. A minimum of three injections of each
solution are necessary to calculate its coefficient of variation
(Cv), which is a measure of precision or reproducibility.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %).
1.4-4
-------
5.3 Since the calibration range must bracket the concentration(s) of PCBs
measured in the sample extract(s), the calibration standard concentra-
tions may require further adjustment resulting in either a redefinition
or refinement of the calibration range.
5.4 The analyst should consult Method 7.1 (Protocol for the Qualitative
Evaluation of Chromatograms of Polychlorinated Biphenyls) to assure
proper interpretation of the PCB Chromatograms and identification of
individual PCB Aroclors.
5.5 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
PCB Aroclor detection limits, extraction recoveries, and potential
interferences.
1.4-5
-------
APPENDIX A
PCB AROCLOR STANDARDS
Individual PCB Aroclors are a mixture of numerous components and considerable
overlap in composition occurs among Aroclor Standards. Quantitative standards
for analyzing aqueous samples for PCB content are available from commercial
vendors. Supelco, Inc. provides individual ampuls of the following Aroclors
at the specified PCB concentrations in methanol:
PCB Aroclor Concentration
Aroclor 1016 20(1 ug/ml
Aroclor 1221 200 ug/ml
Aroclor 1232 200 ug/ml
Aroclor 1242 200 ug/ml
Aroclor 1248 200 ug/ml
Aroclor 1254 200 ug/ml
Aroclor 1260 200 ug/ml
(Source: Supelco, Inc., Catalog 25, 19P7)
1.4-6
-------
APPENDIX B
GAS CHROMATOGRAPHIC CONDITIONS
Analytical Column: *
Carrier Ras:
Carrier Flow Rate:
Detector:
Detector Temperature:
Injector Temperature:
Oven Temperature:
Program Rate:
Time of Analysis:
1.5% SP 2250/1.95% SP 2401 on 100/120 Supelcoport
6 ft x 4 mm glass column (mixed phase support)
Argon:Methane (95%:5%)
40 ml/min
Electron Capture (ECD)
260°C
225°C
200°C
Isothermal
Variable (Dependent on PCB Aroclor)
* The mixed phase column recommended for PCB analyses should be packed and
conditioned in accordance with Method 7.3 (Instructions for Packing and
Conditioning Glass Analytical Columns for Gas Chromatography Instruments)
and performance tested according to the procedure and specifications
described in Appendix A of Method 7.4 (Protocol for Performance Testing
Packed, Conditioned, Glass Analytical Columns for Gas Chromatography
Instruments).
1.4-7
-------
METHOD 1.5 RAPID EXTRACTION AND CHROMATOGRAPHIC ANALYSIS OF
ORGANOCHLORINE PESTICIDES IN AOUEOUS SAMPLES
1. SCOPE AND SUMMARY:
1.1 This method addresses the determination of organochlorine pesticide
compounds in aqueous samples. Specifically, the following pesticides
designated as priority pollutants may be determined by this method:
aldrin; a-BHC; b-BHC; g-BHC; d-BHC; 4,4'-DDD; 4,4'-DDE; 4,4'-DDT;
dieldrin; a-endosulfan; b-endosulfan; endosulfan sulfate; endrin;
endrin aldehyde; heptachlor; and, heptachlor epoxide.
1.2 This method provides a procedure for rapidly extracting organochlorine
pesticide compounds from aqueous samples and quantifying the extracts
via gas chromatographic analysis. A 1.0 liter sample of wastewater
is extracted with cyclohexane using a magnetic spinning technique.
Electron capture gas chromatographic conditions are described which
permit direct measurement of the pesticides in the extract.
2. CAUTIONS AND LIMITATIONS:
2.1 Since pesticides are relatively unstable in aqueous solutions, the
analyst should extract the sample(s) immediately to prevent compound
degradation. In the event that extraction into the organic phase
must be delayed, the sample(s) must be refrigerated at 4°C.
2.2 All glassware employed for standard and sample preparation must be
rinsed three times with cyclohexane and air dried prior to use.
2.3 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., safety glasses, laboratory coat, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous and/or carcino-
genic substances.
2.4 The spinning extraction procedure must be performed behind a safety
shield in a properly ventilated fume hood.
2.5 The 1.0 liter sample bottles must be vented periodically to release
any pressure buildup during the extraction procedure.
2.6 The gas chromatographic conditions listed in the Appendix of this
Method must be established at least two hours prior to analysis of
the calibration standards and sample extracts to permit sufficient
stabilization of the electron capture detector.
1.5-1
-------
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Autosampler vials equipped with teflon lined septa
2. Beakers, assorted volumes
3. Gas chromatograph equipped with electron capture detector
4. Graduated cylinders, 1 L
5. Hand crimpers for autosampler vials and serum bottles
fi. Magnetic stirring bars, teflon coated, 2 inch
7. Magnetic stirring unit
8. Pasteur pipets, disposable, 6 or 9 inch
9. Sample bottles equipped with teflon lined screw caps, 1 L (Wheaton
or equivalent)
10. Serum bottles, crimp top, 30 ml (Wheaton or equivalent)
11. Syringes, 100 ul and 10 ul (Hamilton or equivalent)
12. Volumetric flasks, 100 ml, 50ml, and 25 ml
13. Volumetric pipets, 20 ml
3.2 Reagents
1. Cyclohexane, pesticide grade (Rurdick & Jackson or equivalent)
2. Distilled water, organic free
3. Hydrochloric acid, 1.0 Normal solution
4. Pesticide Standard Mixture (Supelco, Inc. or equivalent)
5. Sodium hydroxide, 1.0 Normal solution
4. PROCEDURE
4.1 Standard Preparation
4.1.1 Using a small beaker or pasteur pipet, transfer approximately
9 ml of cyclohexane to a 10 ml volumetric flask.
4.1.2 Using a 200 ul syringe, quantitatively withdraw 100.0 ul of the
Pesticide Standard Mixture described in Appendix A of this Method
and dispense this volume below the air/cyclohexane interface in
the flask.
NOTE; The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
4.1.3 Using a pasteur pipet, add sufficient cyclohexane to align the
meniscus of the solution with the fiducial mark on the flask.
Stopper the flask and invert repeatedly to ensure complete
dissolution of the pesticide compounds. The concentration of
this stock standard solution is 20.0 ug/ml in pesticides.
1.5-2
-------
NOT!:: Long term storage necessitates wrapping parafllm around'
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and calibration standards by
diluting the stock standard solution appropriately. The cali-
bration range should be established so as to incorporate the
expected concentrations of the samples (refer to Method 8.1 -
Protocol for Establishing a Calibration Range for Single Point
Analysis).
4.2 Sample Preparation
4.2.1 Record individual sample volumes via the following procedure:
1. Prior to extracting the samples, mark the height of the men-
iscus on the exterior of each sample bottle with an indelible
marker or masking tape.
2. Extract the samples in accordance with Sections 4.2.2 - 4.2.7
of this Method and transfer the cyclohexane extracts to 30 ml
crimp-top serum bottles.
3. Empty the aqueous contents of each sample bottle into an
appropriate storage or waste container.
4. Dispense an exact volume of distilled water from a 1.0 L
graduated cylinder into each sample bottle.
5. Record the volume of distilled water required to align the
meniscus with the marker on each sample bottle.
4.2.2 Adjust each of the aqueous samples to pH 7.0 by the addition of
an appropriate volume of 1.0 Normal hydrochloric acid solution
or 1.0 Normal sodium hydroxide solution to the bottles. Monitor
the acidity or basicity of the samples with pH paper.
4.2.3 Place a 2 inch teflon coated magnetic stirring bar into, each of
the sample bottles.
4.2.4 Using a 20 ml volumetric pipet, quantitatively transfer 20.0 ml
of cyclohexane to each 1.0 liter sample bottle and stopper the
bottles immediately to prevent potential volatility losses of
the extraction solvent.
4.2.5 Place the sample bottles on a magnetic stirring unit positioned
behind a safety shield in a fume hood. Adjust the stir bar
spinning rates to achieve a maximum 'coning1 or 'whirlpool'
effect in the sample bottles. The samples should be extracted
in this manner for exactly 30 minutes.
NOTE 1: During the spinning extraction procedure, the analyst
should vent the sample bottles periodically to release
any pressure buildup.
1.5-3
-------
NOTE 2: The analyst should visually inspect the sample bottles
and adjust the spinning rates as necessary to prevent
stir bar "bumping" and to minimize the formation of
emulsions.
4.2.6 After 30 minutes, the magnetic stirring unit should be turned
off and the aqueous and organic layers permitted to separate
for approximately 10 minutes. Using a small beaker or graduated
cylinder, add a sufficient volume of distilled water to each
sample to force the organic phase into the neck of the sample
bottle.
4.2.7 Using pasteur pipets, transfer the cyclohexane extracts contain-
ing the pesticide compounds to 30 ml crimp-top serum bottles.
4.2.8 Measure each sample volume in accordance with Section 4.2.1
(steps 3 - 5) of this Method.
4.3 Gas Chromatographic Analysis
4.3.1 Establish the gas Chromatographic conditions listed in Appendix
B of this Method.
NOTE: The electron capture detector requires a minimum two hour
stabilization period prior to analysis of the calibration
standards and sample extracts.
4.3.2 Using disposable pasteur pipets, transfer approximately 1 ml of
each of the calibration standards and sample extracts to auto-
sampler vials. Seal the autosampler vials with a hand crimper
and position the vials in the autoinjector with cyclohexane
blanks inserted at regular intervals.
NOTE: In the event that an autoinjector is unavailable, the
analyst should use a precision quality 10 ul syringe to
manually inject an exact volume (e.g., usually 3 to 5 ul)
of all standard, sample, and cyclohexane solutions.
4.3.3 Replicate injections of all calibration standards and sample
extracts are required. A minimum of three injections of each
solution are necessary to calculate its coefficient of variation
(Cv), which is a measure of precision or reproducibility.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
1.5-4
-------
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %).
5.3 Since the calibration range must bracket the concentration(s) of the
pesticides measured in the sample extract(s), the calibration standard
concentrations may require further adjustment resulting in either a
redefinition or refinement of the calibration range.
5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
organochlorine pesticide detection limits, extraction recoveries, and
potential interferences.
1.5-5
-------
APPENDIX A
PESTICIDE STANDARD MIXTURE
Quantitative standards for analyzing aqueous samples for organochlorine
pesticides are available from several commercial vendors. Supelco, Inc.
provides the following organochlorine pesticide mixture in a 1.0 ml ampul
at the specified concentrations in toluene:hexane (50:50):
Component Concentration
Aldrin 2000 ug/ml
a-BHC 2000 ug/ml
b-BHC 2000 ug/ml
g-BHC 2000 ug/ml
d-RHC 2000 ug/ml
4,4'-ODD 2000 ug/ml
4,4'-DDE 2000 ug/ml
4,4'-DDT 2000 ug/ml
Dieldrin 2000 ug/ml
a-Endosulfan 2000 ug/ml
B-Endosulfan 2000 ug/ml
Endosulfan sulfate 2000 ug/ml
Endrin 2000 ug/ml
Endrin aldehyde 2000 ug/ml
Heptachlor 2000 ug/ml
Heptachlor epoxide 2000 ug/ml
(Source; Supelco, Inc., Catalog 25, 1987)
1.5-6
-------
APPENDIX B
GAS CHROMATOGRAPHIC CONDITIONS
Analytical Column: *
Carrier Gas:
Carrier Flow Rate:
Detector:
Detector Temperature:
Injector Temperature:
Oven Temperature:
Program Rate:
Time of Analysis:
1.5% SP 2250/1.95% SP 2401 on 100/120 Supelcoport
6 ft x 4 mm glass column (mixed phase support)
Argon:Methane (95%:5%)
40 ml/min
Electron Capture (ECD)
250°C
220°C
200°C
Isothermal
25 min:
The column recommended for organochlorine pesticide analyses should be
packed and conditioned in accordance with Method 7.3 (Instructions for
Packing and Conditioning Glass Analytical Columns for Gas Chromatography
Instruments) and performance tested according to the procedure and the
specifications described in Appendix A of Method 7.4 (Protocol for Per-
formance Testing Packed, Conditioned, Glass Analytical Columns for Gas
Chromatography Instruments).
1.5-7
-------
METHOD 1.6 RAPID EXTRACTION AND SPECTROSCOPIC ANALYSIS
OF FLUORESCING ORGANICS IN AQUEOUS SAMPLES
1. SCOPE AND SUMMARY
1.1 This Method addresses the determination of those organic compounds
that are amenable to fluorescence analysis. Specifically, the follow-
ing classes of priority pollutant materials may be determined by this
Method: polynuclear aromatic hydrocarbons (PAHs); non-halogenated
aromatic hydrocarbons (e.g., benzene derivatives); and, nonhalogenated
phenolics. The methodology avoids some of the interferences that are
often encountered when performing direct in-situ quantitative measure-
ment of hazardous materials in water.
1.2 This Method provides a procedure for rapidly extracting fluorescing
organics from aqueous samples and quantifying the extracts via a
sensitive fluorescence spectroscopic technique. A 50 ml sample of
wastewater is extracted with an equivalent volume of cyclohexane using
a magnetic spinning technique. Synchronous excitation fluorescence
spectroscopy permits direct measurement of the compounds in the
extract.
2. CAUTIONS AND LIMITATIONS:
2.1 All glassware employed for standard and sample preparation must be
rinsed three times with cyclohexane and air dried prior to use.
2.? The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., laboratory coat, safety glasses, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous and/or carcino-
genic substances.
2.3 This Method requires an ultrapure grade of cyclohexane; the analyst
should determine solvent purity by monitoring the emission intensity
of cyclohexane under the instrumental conditions described in the
Appendix of this Method.
2.4 The spinning extraction procedure must be performed behind a safety
shield in a properly ventilated fume hood.
2.5 Since the extraction methodology does not utilize sample fractionation
techniques, the cyclohexane extract may contain a complex mixture of
unresolved fluorescing organics.
1.6-1
-------
2.6 Aqueous samples suspected of containing volatile organlcs should be
refrigerated at 4° C prior to extraction by cyclohexane to prevent
sample degradation.
2.7 Measurement of calibration standards, sample extracts, and solvent
blanks must be conducted under identical instrumental conditions of
analysis.
2.8 The spectrophotometer recorder baseline should be deflected at least
ten percent of full scale to avoid erroneous zero measurements.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Balance, macro analytical, range of 0 - 150 grams (± 0.1 mg)
2. Beakers, assorted volumes
3. Erlenmeyer flasks, ground-glass stoppered, 125 ml
4. Fluorescence spectrophotometer equipped with dual scanning
monochromators for synchronous measurement
5. Glass wool, silanized
6. Graduated cylinders, 100 ml, 50 ml
7. Hand crimper for serum bottles
8. Magnetic stirring bars, teflon coated, 1 inch
9. Magnetic stirring unit
10. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
11. Pasteur pipets, disposable, 6 or 9 inch
12. Quartz cuvette, teflon-stoppered, polished on four faces
13. Serum bottles, crimp top, 100 ml (Wheaton or equivalent)
14. Syringe, 50 ul (Hamilton or equivalent)
15. Volumetric flasks, 100 ml
3.2 Reagents
1. Cyclohexane, spectrophotometric grade, ultraviolet cutoff 204 nm
(Burdick A Jackson or equivalent)
2. Distilled water, organic free
3. Test material for standard preparation
4. PROCEDURE:
4.1 Standard Preparation
4.1.1 Using a small beaker or 100 ml graduated cylinder, transfer
approximately 90 ml of cyclohexane to a 100 ml volumetric flask.
1.6-2
-------
4.1.2 Weigh an appropriate amount of the test material required to
prepare a concentrated stock standard solution, and quantita-
tively transfer the material to the volumetric flask.
NOTE: The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
4.1.3 Using a pasteur pipet, add a sufficient volume of cyclohexane
to align the meniscus of the solution with the fiducial mark on
the flask. Stopper the flask and invert repeatedly to ensure
complete dissolution of the test material.
NOTE: Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and set of calibration standards
by diluting the stock standard solution appropriately. The
calibration range should be established so as to incorporate
the expected concentrations of the samples.
NOTE: The analyst should consult Method fi.l (Protocol for
Establishing a Calibration Range for Single Point
Analysis) to ensure the adequacy of the established
calibration range.
4.2 Sample Preparation
4.2.1 Using 50 ml graduated cylinders, quantitatively transfer 50 ml
of each aqueous sample to 125 ml ground-glass stoppered erlen-
meyer flasks.
NOTE: In the event that an indigenous water blank is available
for background correction purposes, this reference sample
should be extracted and analyzed concurrently with the
'contaminated1 aqueous samples.
4.2.2 Place a one inch teflon coated magntic stirring bar into each
of the erlenmeyer flasks.
4.2.3 Using a 50 ml graduated cylinder, quantitatively transfer 50 ml
of cyclohexane into each of the flasks containing the aqueous
samples. Stopper the flasks immediately to prevent potential
volatility losses of the extraction solvent.
4.2.4 Place the flasks on a magnetic stirring unit positioned behind
a safety shield in a fume hood. Adjust the stir bar spinning
rates to achieve a maximum 'coning' or 'whirlpool1 effect in
the erlenmeyer flasks. The samples should be extracted in this
manner for exactly 30 minutes.
1.6-3
-------
NOTE: During the spinning extraction procedure, the analyst
should vent the erlenmeyer flasks periodically to release
excess pressure generated by the spinning process.
4.2.5 After 30 minutes, the magnetic stirring unit should be turned
off and the aqueous and organic layers permitted to separate
for approximately 10 minutes. Using a small beaker or graduated
cylinder, add a sufficient volume of distilled water to each
sample to force the organic phase into the neck of the erlen-
meyer flask.
4.2.6 Using pasteur pipets, transfer the cyclohexane extracts contain-
ing the fluorescing organics to 100 ml crimp-top serum bottles.
NOTE: In the event there is a difficult emulsion in one or
more of the erlenmeyer flasks, filter the cyclohexane
phase through silanized glass wool while transferring
the sample extracts to the serum bottles.
4.3 Fluorescence Analysis
4.3.1 Establish the fluorescence spectrophotometer conditions listed
in the Appendix of this Method.
4.3.2 Using a pasteur pipet, transfer a sufficient volume (e.g., 2 ml)
of the calibration solution to a quartz cuvette for analysis by
synchronous excitation fluorescence spectroscopy.
4.3.3 Observe the synchronous excitation spectra for each set of wave-
length intervals (e.g. A 20, A 30,..., A 100) over the range of
200 nm - 500 nm to establish the optimum wavelength interval for
quantitative measurement.
NOTE: The optimum wavelength interval selected should be a
compromise between maximum fluorescence intensity and
adequate peak resolution.
4.3.4 Using the optimum wavelength interval established in step 4.3.3
and the instrument conditions described in the Appendix, record
the synchronous excitation spectra of the calibration solution
and set of calibration standards in triplicate.
NOTE: Triplicate analyses of all calibration standards and
sample extracts are required to calculate a coefficient
of variation (Cv), which is a measure of the precision
or reproducibility of the analyses.
4.3.5 Record the synchronous excitation spectra of the solvent blank
(cyclohexane), the aqueous sample extracts, and the water blank
(pending availability) under identical instrumental conditions.
1.6-4
-------
4.3.6 Assure instrument stability by scanning the calibration solution
and solvent blank following each fifth sample measurement.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values ' 3 %)
5.3 Since the calibration range must bracket the concentration(s) of
fluorescing organic compounds measured in the sample extract(s), the
calibration standard concentrations may require further adjustment
resulting in either a redefinition or refinement of the initially
established calibration range.
5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
compound detection limits, extraction recoveries, and potential inter-
ferences.
1.6-5
-------
APPENDIX
FLUORESCENCE SPECTROPHOTOMETER CONniTIONS
Initial Excitation Wavelength: 200 nm
* Initial Wavelength Interval (A): 20 nm
Initial Emission Wavelength: 220 nm
Final Excitation Wavelength: 500 nm
Final Emission Wavelength: 500 nm + A
Excitation Slit Width: 10 nm
Emission Slit Width: 10 nm
Wavelength Drive: Excitation
Scanning Mode: Dual Monochromators
Amplifier Dynode Voltage: Ratio
Recorder Power: Servo
The initial wavelength interval (A20) requires an initial emission
wavelength of 220 nm with a starting excitation wavelength of 200 nm.
The starting emission wavelength is sequentially adjusted in 10 nm
increments to values of 230 nm, 240 nm, 250 nm, etc., to record the
synchronous excitation spectra for each set of wavelength intervals
(A30, A40, A 50, etc.), and establish the optimum wavelength interval
for quantitative measurement. The excitation monochromator is scanned
in synchronization with the emission monochromator over the wavelength
range of 200 nm - 500 nm to obtain a complete fluorescence spectrum of
the test material. The optimum wavelength interval selected should
be a compromise between maximum fluorescence intensity and adequate
peak resolution.
1.6-6
-------
METHOD 1.7 RAPID IN-SITU ANALYSIS OF HAZARDOUS MATERIALS IN
AQUEOUS SAMPLES BY FLUORESCENCE SPECTROSCOPY
1. SCOPE AND SUMMARY:
1.1 This Method addresses the determination of those hazardous materials
that are amenable to fluorescence analysis. Specifically, the follow-
ing classes of priority pollutant materials may be determined by this
Method: polynuclear aromatic hydrocarbons (PAHs); non-halogenated
aromatic hydrocarbons (e.g., benzene derivatives); and, nonhalogenated
phenolics.
1.2 This Method provides a procedure for the direct in-situ quantitative
measurement of hazardous materials in water through use of a semi-
automated fluorescence spectroscopic technique. A sample of waste-
water is introduced untreated into a flow-through quartz cuvette and
analyzed via synchronous excitation fluorescence spectroscopy.
2. CAUTIONS AND LIMITATIONS:
2.1 All glassware employed for standard and sample preparation must be
rinsed three times with distilled water and air dried prior to use.
2.2 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., laboratory coat, safety glasses, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous a'nd/or carcino-
genic substances.
2.3 Since the methodology does not utilize sample fractionation techniques
and the wastewater samples may contain a complex mixture of hazardous
materials, the resultant fluorescence spectra may not be resolved into
individual components.
2.4 Aqueous samples suspected of containing volatile organics should be
refrigerated at 4° C prior to fluorescence analysis to prevent sample
degradation
2.5 Measurement of calibration standards, sample extracts, and solvent
blanks must be conducted under identical instrumental conditions of
analysis.
2.6 The spectrophotometer recorder baseline should be deflected at least
ten percent of full scale to avoid erroneous zero measurements.
1.7-1
-------
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Auto analyzer system (peristaltic pump, teflon tubing)
?.. Balance, macro analytical, range of 0 - 150 grams (± 0.1 mg)
3. Beakers, assorted volumes
4. Fluorescence spectrophotometer equipped with dual scanning
monochromators for synchronous measurement
5. Graduated cylinders, 100 ml
6. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
7. Pasteur pipets, disposable, 6 or 9 inch
8. Quartz cuvette, flow-through, polished on four faces
9. Syringe, 50 ul (Hamilton or equivalent)
10. Volumetric flasks, 100 ml
3.2 Reagents
1. Distilled water, organic free
2. Test material for standard preparation
4. PROCEDURE:
4.1 Standard Preparation
4.1.1 Using a small beaker or 100 ml graduated cylinder, transfer
approximately 90 ml of distilled water to a 100 ml volumetric
flask.
4.1.2 Weigh an appropriate amount of the test material required to
prepare a concentrated stock standard solution, and quantita-
tively transfer the material to the volumetric flask.
NOTE 1: The analyst should consult Method 5.1 (Protocol
for Preparing Analytical Standards of Hazardous
and Carcinogenic Materials) for proper standard
preparation techniques.
NOTE 2: In the event that the hazardous material is either
insoluble or slightly soluble in water, the analyst
should consult Method 5.2 (Protocol for Preparing
Synthetic Samples of Hazardous Materials in Water).
4.1.3 Using a pasteur pi pet, add a sufficient volume of distilled water
to align the meniscus of the solution with the fiducial mark on
the flask. Stopper the flask and invert repeatedly to ensure
complete dissolution of the test material.
1.7-2
-------
NOTE: Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and set of calibration standards
by diluting the stock standard solution appropriately. The
calibration range should be established so as to incorporate
the expected concentrations of the samples.
NOTE: The analyst should consult Method 8.1 (Protocol for
Establishing a Calibration Range for Single Point
Analysis) to ensure the adequacy of the established
calibration range.
4.2 Sample Preparation
4.2.1 Direct in-situ measurement of the hazardous materials precludes
the need for extensive sample preparation prior to fluorescence
analysis.
4.2.2 The aqueous samples should be refrigerated in amber glass bottles
at 4° C prior to fluorescence analysis to prevent the possibility
of sample degradation.
4.2.3 In the event that an indigenous water blank is available for
background correction purposes, this reference sample should be
analyzed concurrently with the 'contaminated1 aqueous samples.
4.3 Fluorescence Analysis
4.3.1 Establish the fluorescence spectrophotometer conditions listed
in Appendix A of this Method.
4.3.2 Using a pasteur pipet, transfer a sufficient volume (e.g., 2 ml)
of the calibration solution to a quartz cuvette for analysis by
synchronous excitation fluorescence spectroscopy.
4.3.3 Observe the synchronous excitation spectra for each set of wave-
length intervals (e.g. A 20, A 30,..., A 100) over the range of
200 nm - 500 nm to establish the optimum wavelength interval for
quantitative measurement.
NOTE: The optimum wavelength interval selected should be a
compromise between maximum fluorescence intensity and
adequate peak resolution.
4.3.4 Using the optimum wavelength interval established in step 4.'3.3
and the auto-analyzer mode of operation as described in Appendix
B, record the synchronous excitation spectra of the calibration
solution and set of calibration standards in triplicate.
1.7-3
-------
NOTE; Triplicate analyses of all calibration standards and
sample extracts are required to calculate a coefficient
of variation (Cv), which is a measure of the precision
or reproducibility of the analyses.
4.3.5 Record the synchronous excitation spectra of the solvent blank
(distilled water), the sample solution(s), and the reference
water sample (pending availability) under identical instrumental
and auto-analyzer conditions.
4.3.6 Assure instrument stability by scanning the calibration solution
and solvent blank following each fifth sample measurement.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %}
5.3 Since the calibration range must bracket the concentration(s) of
fluorescing hazardous materials measured in the sample(s), the
calibration standard concentrations may require further adjustment
resulting in either a redefinition or refinement of the initially
established calibration range.
5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
compound detection limits, extraction recoveries, and potential inter-
ferences.
1.7-4
-------
APPENDIX A
FLUORESCENCE SPECTROPHOTOMETER CONDITIONS
Initial Excitation Wavelength: 200 nm
* Initial Wavelength Interval (&): 20 nm
Initial Emission Wavelength: 220 nm
Final Excitation Wavelength: 500 nm
Final Emission Wavelength: 500 nm + A
Excitation Slit Width: 10 nm
Emission Slit Width: 10 nm
Wavelength Drive: Excitation
Scanning Mode: Dual Monochromators
Amplifier Dynorie Voltage: Ratio
Recorder Power: Servo
The initial wavelength interval U20) requires an initial emission
wavelength of 220 nm with a starting excitation wavelength of 200 nm.
The starting emission wavelength is sequentially adjusted in 10 nm
increments to values of 230 nm, 240 nm, 250 nm, etc., to record the
synchronous excitation spectra for each set of wavelength intervals
(A30, A40, A 50, etc.), and establish the optimum wavelength interval
for quantitative measurement. The excitation monochromator is scanned
in synchronization with the emission monochromator over the wavelength
range of 200 nm - 500 nm to obtain a complete fluorescence spectrum of
the test material. The optimum wavelength interval selected should
be a compromise between maximum fluorescence intensity and adequate
peak resolution.
1.7-5
-------
APPENDIX B
AUTO ANALYZER CONDITIONS
1. Properly position the flow-through quartz cuvette in the instrument sample
compartment (i.e., the polished faces of the cuvette must be situated
opposite the exit aperture of the excitation monochromator and the entrance
aperture of the emission monochromator).
2. Ensure that the peristaltic pump tubes are properly connected to the flow-
through cuvette and do not block the excitation or emission light paths.
3. Solvent cleanse the flow-through quartz cuvette by placing the teflon
inlet tube into a beaker of distilled water and activating the peristaltic
pump to pass a total of 25 ml through the cuvette.
4. Record the synchronous excitation spectrum of the distilled water (solvent
blank) using the optimum wavelength interval established in step 4.3.3 of
this Method (refer to the section entitled 'Fluorescence Analysis' on page
1.7-3).
5. Transfer the teflon inlet tube to the beaker containing the Lower Limit (LL)
calibration standard and activate the peristaltic pump, displacing the water
in the flow-through quartz cuvette with the LL standard solution.
6. Record the synchronous excitation spectrum of the LL calibration standard.
7. Transfer the teflon inlet tube to the beaker of distilled water and displace
the LL calibration standard in the cuvette.
8. Fill the cuvette with the next calibration standard of higher concentration
(e.g., LC) and record its synchronous excitation spectrum.
9. Repeat steps 7 and 8, recording the synchronous excitation spectra for the
complete set of calibration standards (e.g., LL, LC, CS/\, C$B, UC, UL), the
sample solution(s), and the reference water sample (pending availability)
under identical instrumental conditions.
1.7-6
-------
METHOD 1.8 RAPID IN-SITU ANALYSIS OF QUINOLINE IN
WATER BY FLUORESCENCE SPECTROSCOPY
1. SCOPE AND SUMMARY:
1.1 This Method specifically addresses the determination of quinoline in
aqueous samples through use of a semi-automated fluorescence spectro-
scopic technique.
1.2 This Method provides a procedure for the direct in-situ quantitative
measurement of quinoline in water. A 50.0 ml sample of wastewater
containing quinoline is acidified to pH 2.8 by the addition of 25 ml
of a buffer solution. The diluted sample is introduced into a flow-
through quartz cuvette and analyzed via synchronous excitation fluor-
escence spectroscopy.
2. CAUTIONS AND LIMITATIONS:
2.1 All glassware employed for standard and sample preparation must be
rinsed three times with distilled water and air dried prior to use.
2.2 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., laboratory coat, safety glasses, chemical resistant gloves) to
minimize exposure of laboratory personnel to quinoline.
2.3 Since the methodology does not utilize sample fractionation techniques
and the wastewater samples may contain fluorescing species other than
quinoline, the resultant fluorescence spectra may not be resolved into
individual components.
2.4 The aqueous samples should be refrigerated in amber glass bottles at
4°C prior to fluorescence analysis to prevent the possibility of sample
degradation.
2.5 Since the fluorescence emission intensity of quinoline is pH dependent,
all calibration standards and sample solutions must be adjusted to pH
2.8 accordingly.
2.6 Measurement of calibration standards, sample extracts, and solvent
blanks must be conducted under identical instrumental conditions of
analysis.
2.7 The spectrophotometer recorder baseline should be deflected at least
ten percent of full scale to avoid erroneous zero measurements.
1.8-1
-------
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Auto analyzer system (peristaltic pump, teflon tubing)
2. Balance, macro analytical, range of 0 - 150 grams (± 0.1 mg)
3. Beakers, 2.0 L, 400 ml, 50 ml
4. Erlenmeyer flasks, ground-glass stoppered, 125 ml
5. Fluorescence spectrophotometer equipped with dual scanning
monochromators for synchronous measurement
6. Graduated cylinders, 1.0 L, 100 ml, 10 ml
7. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
8. Pasteur pipets, disposable, 6 or 9 inch
9. pH meter
10. Quartz cuvette, flow-through, polished on four faces
11. Spatula, stainless steel
12. Syringe, 50 ul (Hamilton or equivalent)
13. Volumetric flasks, 2.0 L, 100 ml
14. Volumetric pipet, 25 ml
3.2 Reagents
1. Distilled water, organic free
2. Hydrochloric acid, concentrated
3. Potassium hydrogen phthalate, primary standard grade
4. Quinoline, primary standard grade
4. PROCEDURE:
4.1 Standard Preparation
4.1.1 Using a small beaker or a 100 ml graduated cylinder, transfer
approximately 90 ml of a 2:1 distilled water:buffer mixture to
a 100 ml volumetric flask.
NOTE: The analyst should prepare the 2:1 distilled water:buffer
mixture according to the procedure described in Appendix
A of this method.
4.1.2 Using a 50 ul syringe and macro-analytical balance, weigh 20 mg
of the quinoline standard and dispense this quantity below the
air/solvent interface in the volumetric flask.
NOTE: The'analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
1.8-2
-------
4.1.3 Using a pasteur pi pet, add a sufficient volume of the 2:1 mixture
to align the meniscus of the solution with the fiducial mark on
the flask. Stopper the flask and invert repeatedly to ensure
complete dissolution of quinoline. The concentration of this
stock standard solution is 200 mg/L (parts-per-million).
NOTE; Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and set of calibration standards
by appropriate dilutions of the stock standard solution in the
2:1 distilled water:buffer mixture. The calibration range must
be established so as to incorporate the expected concentrations
of the samples.
NOTE: The analyst should consult Method 8.1 (Protocol for
Establishing a Calibration Range for Single Point
Analysis) to ensure the adequacy of the established
calibration range.
4.2 Sample Preparation
4.2.1 Using 100 ml graduated cylinders, transfer 50 ml of each sample
to 125 ml ground-glass stoppered erlenmeyer flasks.
NOTE: In the event that an indigenous water blank is available
for background correction, this reference sample should
be analyzed concurrently with the wastewater samples.
4.2.2 Using a 50 ml graduated cylinder or 25 ml volumetric pipet, add
25 ml of a pH 2.8 buffer solution to each of the sample flasks.
NOTE: The analyst should prepare the pH 2.8 buffer solution
according to the procedure described in Appendix A of
this Method.
4.2.3 Stopper the erlenmeyer flasks and invert the samples repeatedly
to ensure uniform mixing.
NOTE: In the event that further dilution of the sample(s) is
(are) required for fluorescence analysis (e.g., sample
response values fall outside the confines of the cali-
bration range), such-sample solutions must he prepared
in the 2:1 distilled water:buffer mixture.
4.3 Fluorescence Analysis
4.3.1 Establish the fluorescence spectrophotometer and auto-analyzer
conditions described in Appendices B and C of this Method.
1.8-3
-------
4.3.2 Employing the optimum wavelength interval of 60 nm and the auto-
analyzer mode of operation, record the synchronous excitation
spectra of the quinoline calibration solution and calibration
standards in triplicate.
NOTE 1; The wavelength interval of 60 nm has been established
as the optimum wavelength interval for quantitative
measurement of quinoline in an aqueous matrix.
NOTE 2: Triplicate analyses of all calibration standards and
sample extracts are required to calculate a coefficient
of variation (Cv), which is a measure of the precision
or reproducibility of the analyses.
4.3.3 Record the synchronous excitation spectra of the solvent blank
(2:1 distilled water:buffer mixture), the sample solution(s),
and the reference water sample (pending availability) under
identical instrumental and auto-analyzer conditions.
4.3.4 Assure instrument stability by scanning the calibration solution
and solvent blank following each fifth sample measurement.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %)
5.3 Since the calibration range must bracket the quinoline concentration(s)
measured in the sample(s), the calibration standard concentrations
may require further adjustment resulting in either a redefinition or
refinement of the initially established calibration range.
5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
compound detection limits, extraction recoveries, and potential inter-
ferences.
1.8-4
-------
APPENDIX A
PREPARATION OF AN 0.1 MOLAR POTASSIUM HYDROGEN PHTHALATE SOLUTION
1. Using a stainless steel spatula and macro-analytical balance, weigh 40.0 gms
of primary standard grade potassium hydrogen phthalate into a 400 ml beaker.
2. Using a 100 ml graduated cylinder, dissolve the weighed material by adding
approximately 100 ml of organic free distilled water to the beaker.
3. Quantitatively transfer the solution from the beaker to a 2.0 L volumetric
flask. Fill the 400 ml beaker with approximately 200 ml of organic free
distilled water and carefully swirl its contents to rinse the walls. Add
this volume of solution to that in the flask and repeat this procedure once
more so that the 2.0 L flask contains a total volume of 500 ml.
4. Using a 1.0 L graduated cylinder, carefully add 1500 ml of organic free
distilled water to the flask. Stopper the flask and invert repeatedly to
ensure uniform mixing of the solution.
5. Using a pasteur pi pet, add a sufficient volume of organic free distilled
water to align the meniscus of the solution with the fiducial mark on the
flask. The concentration of this potassium hydrogen phthalate solution is
0.1 Molar.
PREPARATION OF AN 0.1 MOLAR HYDROCHLORIC ACID SOLUTION
1. Using a 1.0 L graduated cylinder, transfer approximately 950 ml of organic
free distilled water into a 1.0 L volumetric flask.
2. Using a 10 ml graduated cylinder, measure and dispense 8.3 ml of concentrated
hydrochloric acid solution to the distilled water in the flask.
3. Stopper the flask and invert repeatedly to ensure uniform mixing of the acid
solution. Using a 100 ml graduated cylinder, add a sufficient volume of the
organic free distilled water to align the meniscus of the solution with the
fiducial mark on the flask. The concentration of this hydrochloric acid
solution is 0.1 Molar.
1.8-5
-------
APPENDIX A (CONTINUED)
PREPARATION OF A pH 2.8 BUFFER SOLUTION
1. Using a 1.0 L graduated cylinder, measure and dispense 1.0 L of the 0.1 Molar
potassium hydrogen phthalate solution to a 2.0 L beaker.
2. Using a 1.0 L graduated cylinder, measure 600 ml of the 0.1 Molar hydrochloric
acid solution and add this volume to the 1.0 L of solution in the beaker.
3. Using a pH meter, adjust the buffer solution in the beaker to pH 2.8 by adding
a sufficient volume of either 0.1 Molar hydrochloric acid solution or 0.1 Molar
potassium hydrogen phthalate solution.
4. Quantitatively transfer the pH 2.8 buffer solution from the beaker to a 2.0 L
volumetric flask. Using a 400 ml beaker, add organic free distilled water to
the flask until the meniscus of the solution is just below the fiducial mark.
5. Stopper the flask and invert repeatedly to ensure uniform mixing of the buffer
solution. Using a pasteur pipet, add a sufficient volume of organic free
distilled water to align the meniscus of the pH 2.8 buffer solution with the
fiducial mark on the flask. All wastewater samples containing quinoline must
be diluted 2:1 (i.e., 50 ml aqueous sample : 25 ml buffer solution) in this
pH 2.8 buffer solution prior to fluorescence analysis.
PREPARATION OF A 2:1 (V:V) DISTILLED WATER:BUFFER MIXTURE
1. Dispense 1.0 L of organic free distilled water and 500 ml of the pH 2.8 buffer
solution into separate 1.0 L graduated cylinders.
2. Transfer the 1.0 L of organic free distilled water from the graduated cylinder
to a 2.0 L volumetric flask. Carefully dispense the 500 ml of pH 2.8 buffer
solution to the volume of distilled water in the flask.
3. Stopper the flask and invert repeatedly to ensure uniform mixing of the 2:1
distilled water:buffer mixture. All quinoline calibration standards must be
prepared in this 2:1 distilled water:buffer mixture.
1.8-6
-------
APPENDIX B
FLUORESCENCE SPECTROPHOTOMETER CONDITIONS
Initial Excitation Wavelength: 200 nm
* Wavelength Interval (A): 60 nm
Initial Emission Wavelength: 260 nm
Final Excitation Wavelength: 500 nm
Final Emission Wavelength: 560 nm
Excitation Slit Width: 10 nm
Emission Slit Width: 10 nm
Wavelength Drive: Excitation
Scanning Mode: Dual Monochromators
Amplifier Dynode Voltage: Ratio
Recorder Power: Servo
The wavelength interval of 60 nm has been established as the optimum
wavelength interval for quantitative measurement of quinoline in an
aqueous matrix. The maximum fluorescence intensity of quinoline is
observed at an excitation wavelength of 325 nm and an emission wave-
length of 385 nm. The excitation monochromator is scanned in synchro-
nization with the emission monochromator over the excitation wavelength
range of 200 nm - 500 nm to obtain a complete fluorescence spectrum
of quinoline.
1.8-7
-------
APPENDIX B
AUTO ANALYZER CONDITIONS
1. Properly position the flow-through quartz cuvette in the instrument sample
compartment (i.e., the polished faces of the cuvette must be situated
opposite the exit aperture of the excitation monochromator and the entrance
aperture of the emission monochromator).
2. Ensure that the peristaltic pump tubes are properly connected to the flow-
through cuvette and do not block the excitation or emission light paths.
3. Solvent cleanse the flow-through quartz cuvette by placing the teflon inlet
tube into a beaker of the 2:1 distilled water:buffer mixture and activating
the peristaltic pump to pass a total of 25 ml through the cuvette.
4. Record the synchronous excitation spectrum of the 2:1 distilled water:buffer
mixture (solvent blank) using the optimum wavelength interval of 60 nm.
5. Transfer the teflon inlet tube to the beaker containing the Lower Limit (LL)
quinoline calibration standard and activate the peristaltic pump, displacing
the 2:1 distilled water:buffer mixture in the flow-through quartz cuvette
with the LL standard solution.
6. Record the synchronous excitation spectrum of the quinoline LL calibration
standard.
7. Transfer the teflon inlet tube to the beaker containing the 2:1 distilled
watenbuffer mixture and displace the quinoline LL calibration standard in
the cuvette.
8. Fill the cuvette with the next quinoline calibration standard of higher
concentration (e.g., LC) and record its synchronous excitation spectrum.
9. Repeat steps 7 and 8, recording the synchronous excitation spectra for the
complete set of quinoline calibration standards (e.g., LL, LC, CSA, CSg, UC,
and UL), the sample solution(s), and the reference water sample (pending
availability) under identical instrumental conditions.
1.8-8
-------
SECTION II
METHOnOLOfiY FOR THE ANALYSIS OF PRIORITY POLLUTANTS
AND HAZARDOUS MATERIALS IN SEDIMENT SAMPLES
-------
CONTENTS
METHOD 2.1 Rapid Extraction and Chromatographic Analysis of
Polychlorinated Biphenyls 1n Sediment Samples
METHOD 2.2 Rapid Extraction and Spectroscopic Analysis
of Fluorescing Organics in Sediment Samples
-------
METHOD 2.1 RAPID EXTRACTION AND CHROMATOGRAPHIC ANALYSIS OF
POLYCHLORINATED BIPHENYLS IN SEDIMENT SAMPLES
1. SCOPE AND SUMMARY:
1.1 This method addresses the determination of polychlorinated blphenyl
(PCB) compounds. Specifically, the following PCB Aroclors may be
determined by this method: Aroclor 1016; Aroclor 1221; Aroclor 1232;
Aroclor 1242; Aroclor 1248; Aroclor 1254; and, Aroclor 1260.
1.2 This method provides a procedure for rapidly extracting PCBs from
sediment samples and quantifying the extracts via gas chromatographic
analysis. A 10.0 gram sample of contaminated sediment is weighed into
a 250 ml screw top erlenmeyer flask and extracted with hexane using a
gyrotory agitation technique. Electron capture gas chromatographic
conditions are described which permit direct measurement of the PCBs
in the extract.
2. CAUTIONS AND LIMITATIONS:
2.1 All glassware employed for standard and sample preparation must be
rinsed three times with hexane and air dried prior to use.
2.2 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., chemical resistant gloves, safety glasses, etc.) to minimize
exposure of laboratory personnel to hazardous and/or carcinogenic
substances.
2.1 The gyrotory extraction procedure must be performed behind a safety
shield in a properly ventilated fume hood.
2.2 The 250 ml screw top erlenmeyer flasks must be vented periodically to
release any pressure buildup during the extraction procedure.
2.3 The gas chromatographic conditions listed in Appendix B of this
Method must be established at least two hours prior to analysis of
the calibration standards and sample extracts to permit sufficient
stabilization of the electron capture detector.
2.4 The analyst should refer to Method 7.1 (Protocol for the Qualitative
Evaluation of Chromatograms of Polychlorinated Biphenyls) for proper
interpretation of the PCB chromatograms.
2.1-1
-------
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Aluminum foil
2. Aluminum pans, 9 in. x 6 in. x 1 in.
3. Autosampler vials equipped with teflon lined septa
4. Balance, macro analytical, range of 0 - 150 grams (± 0.1 mg)
5. Beakers, assorted volumes
6. Erlenmeyer screw top flasks, 250 ml
7. Gas chromatograph equipped with electron capture detector
8. Graduated cylinder, 100 ml
9. Gyrotory Shaker (New Brunswick Scientific Co., Inc. or equivalent)
10. Hand crimpers for autosampler vials and serum bottles
11. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
12. Pasteur pipets, disposable, 6 or 9 inch
13. Serum bottles, crimp top, 100 ml (Uheaton or equivalent)
14. Spatula, stainless steel
15. Stirring rod, glass, 9 inch by 1/2 inch
16. Syringes, 200 ul and 10 ul (Hamilton or equivalent)
17. Volumetric flasks, 100 ml, 50ml, and 25 ml
18. Volumetric pipets, 20 ml
3:2 Reagents
1. PCB Aroclor Standards (Supelco, Inc. or equivalent)
2. Hexane, chromatographic grade (Burdick & Jackson or'equivalent)
4. PROCEDURE:
4.1 Standard Preparation
4.1.1 Using a volumetric pipet or a beaker, transfer approximately
20 ml of hexane to a 25 ml volumetric flask.
4.1.2 Using a micropipet or 200 ul syringe, quantitatively withdraw
150.0 ul of an Aroclor standard containing 200 ug/ml of PCBs and
dispense this volume below the air/hexane interface in the flask
(refer to Appendix A of this Method for a list of PCB Aroclors).
NOTE: The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
4.1.3 Using a pasteur pipet, add a sufficient volume of hexane to align
the meniscus of the solution with the fiducial mark on the flask.
Stopper the flask and invert repeatedly to ensure complete dis-
2.1-2
-------
solution of the PCBs. The concentration of this stock standard
solution is 1200 ng/ml (parts per billion) in PCBs.
NOTE: Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and calibration standards by
diluting the stock standard solution appropriately. The cali-
bration range should be established so as to incorporate the
expected concentrations of the samples.
NOTE; The analyst should consult Method 8.1 (Protocol for
Establishing a Calibration Range for Single Point
Analysis) to ensure the adequacy of the established
calibration range.
4.2 Sample Preparation
4.2.1 Using a glass stirring rod, mix each of the wet sediment samples
thoroughly to obtain as homogeneous a mixture as possible.
NOTE: In the event that an indigenous sediment blank is avail-
able for background correction, this reference sample
should be extracted and analyzed concurrently with the
'contaminated1 sediment samples.
4.2.1 Place approximately 120 grams of each wet sediment sample into
separate aluminum drying pans positioned on the floor of a fume
hood. Cover each of the trays with a large kimwipe and permit
the samples to air dry in this manner for 24 to 48 hours.
4.2.2 Using a macro analytical balance and a stainless steel spatula,
weigh 10.0 grams of each air-dried sediment sample directly into
tared 250 ml erlenmeyer screw top flasks.
4.2.3 Using a 100 ml graduated cylinder, quantitatively transfer 100 ml
of hexane into each of the flasks containing the sediment samples.
Line the interior of the erlenmeyer caps with aluminum foil and
screw the caps onto the flasks to prevent potential volatility
losses of the extraction solvent.
4.2.4 Place the erlenmeyer flasks on a gyrotory shaker unit positioned
behind a safety shield in a fume hood. Adjust the agitation rate
of the shaker device to 400 RPM and extract the samples under
these conditions for exactly 30 minutes.
NOTE: During the gyrotory extraction procedure, the analyst
should periodically (e.g., at 5 minute intervals) turn
off the gyrotory shaker and vent the erlenmeyer flasks
to release any pressure buildup.
2.1-3
-------
4.2.5 After 30 minutes, the gyrotory shaker should be turned off and
the erlenmeyer flasks removed to a stationary location. The
analyst should permit suspended solids and particulate matter
in the flasks to settle for approximately 30 minutes.
4.2.6 Using pasteur pipets, carefully transfer the hexane extracts
containing the PCBs to 100 ml crimp-top serum bottles. DO NOT
agitate the sediment layer in the flask since particulates may
interfere with gas chromatographic analysis of the extracts.
4.3 Gas Chromatographic Analysis
4.3.1 Establish the gas chromatographic conditions listed in Appendix
B of this Method.
NOTE: The electron capture detector requires a minimum two hour
stabilization period prior to analysis of the calibration
standards and sample extracts.
4.3.2 Using pasteur pipets, transfer approximately 1 ml of each of the
calibration standards and sample extracts to autosampler vials.
Seal the autosampler vials with a hand crimper and position the
vials in the autoinjector with hexane blanks inserted at regular
intervals.
NOTE: In the event that an autoinjector is unavailable, the
analyst should use a precision quality 10 ul syringe to
manually inject an exact volume (e.g., usually 3 to 5 ul)
of all standard, sample, and hexane solutions.
4.3.3 Replicate injections of all calibration standards and sample
extracts are required. A minimum of three injections of each
solution are necessary to calculate a coefficient of variation
(Cv), which is a measure of the precision or reproducibility of
the analyses.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %).
5.3 Since the calibration range must bracket the concentration(s) of PCBs
measured in the sample extract(s), the calibration standard concentra-
2.1-4
-------
tions may require further adjustment resulting in either a redefinition
or refinement of the calibration range.
5.4 The analyst should consult Method 7.1 (Protocol for the Qualitative
Evaluation of Chromatograms of Polychlorinated Biphenyls) to assure
proper interpretation of the PCB chromatograms and identification of
individual PCR Aroclors.
5.5 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
PCB Aroclor detection limits, extraction recoveries, and potential
interferences.
2.1-5
-------
APPENDIX A
PCB AROCLOR STANDARDS
Individual PCB Aroclors are a mixture of numerous components and considerable
overlap In composition occurs among Aroclor Standards. Quantitative standards
for analyzing aqueous samples for PCB content are available from commercial
vendors. Supelco, Inc. provides individual ampuls of the following Aroclors
at the specified PCB concentrations in methanol:
PCB Aroclor Concentration
Aroclor 1016 200 ug/ml
Aroclor 1221 200 ug/ml
Aroclor 1232 200 ug/ml
Aroclor 1242 200 ug/ml
Aroclor 1248 200 ug/ml
Aroclor 1254 200 ug/ml
Aroclor 1260 200 ug/ml
(Source: Supelco, Inc., Catalog 25, 1987)
2.1-6
-------
APPENDIX 8
GAS CHROMATOGRAPHIC CONDITIONS
Analytical Column: *
Carrier Gas:
Carrier Flow Rate:
Detector:
Detector Temperature:
Injector Temperature:
Oven Temperature:
Program Rate:
Time of Analysis:
1.5% SP 2250/1.95% SP 2401 on 100/120 Supelcoport
(mixed phase support); 6 ft x 4 mm I.D. glass column
Argon:Methane (95%:5%)
40 ml/min
Electron Capture (ECD)
260°C
225°C
200°C
Isothermal
Variable (Dependent on PCB Aroclor)
* The mixed phase column recommended for PCR analyses should be packed and
conditioned in accordance with Method 7.3 (Instructions for Packing and
Conditioning Glass Analytical Columns for Gas Chromatography Instruments)
and performance tested according to the procedure and specifications
described in Appendix C of Method 7.4 (Protocol for Performance Testing
Packed, Conditioned, Glass Analytical Columns for Gas Chromatography
Instruments).
2.1-7
-------
METHOD 2.2 RAPID EXTRACTION AND SPECTROSCOPIC ANALYSIS
OF FLUORESCING ORGANICS IN SEDIMENT SAMPLES
1. SCOPE AND SUMMARY
1.1 This Method addresses the determination of those organic compounds
that are amenable to fluorescence analysis. Specifically, the follow-
ing classes of priority pollutant materials may be determined by this
Method: polynuclear aromatic hydrocarbons (PAHs); non-halogenated
aromatic hydrocarbons (e.g., benzene derivatives); and, nonhalogenated
phenolics.
1.2 This Method provides a procedure for rapidly extracting fluorescing
organics from sediment samples and quantifying the extracts via a
sensitive fluorescence spectroscopic technique. A 5.0 gram sample of
contaminated sediment is weighed into a 125 ml ground-glass stoppered
erlenmeyer flask and extracted with isopropanol using a gyrotory
agitation technique. Synchronous excitation fluorescence spectroscopy
permits direct measurement of the compounds in the extract.
2. CAUTIONS AND LIMITATIONS:
2.1 All glassware employed for standard and sample preparation must be
rinsed three times with isopropanol and air dried prior to use.
2.2 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., laboratory coat, safety glasses, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous and/or carcino-
genic substances.
2.3 This Method requires an ultrapure grade of isopropanol; the analyst
should determine solvent purity by monitoring the emission intensity
of isopropanol under the instrumental conditions described in the
Appendix of this Method.
NOTE: Interfering impurities in the isopropanol may be removed by
filtration through granular activated carbon in accordance
with the procedure of Method 6.4 (Protocol for Purifying
Isopropanol for Fluorescence Analyses).
2.4 Measurement of calibration standards, sample extracts, and solvent
blanks must be conducted under identical instrumental conditions of
analysis. The fluorescence spectrophotometer recorder baseline
should be deflected at least ten percent of full scale to avoid
erroneous zero measurements.
2.2-1
-------
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Balance, macro analytical, range of 0 - 150 grams (± 0.1 mg)
2. Beakers, assorted volumes
3. Erlenmeyer flasks, ground-glass stoppered, 125 ml
4. Fluorescence spectrophotometer equipped with dual scanning
monochromators for synchronous measurement
5. Graduated cylinders, 100 ml, 50 ml
6. Gyrotory shaker (New Brunswick Scientific Company or equivalent)
7. Hand crimper for serum bottles
8. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
9. Pasteur pipets, disposable, 6 or 9 inch
10. Quartz cuvette, polished on four faces
11. Serum bottles, crimp top, 30 ml (Wheaton or equivalent)
12. Spatula, stainless steel
13. Stirring rod, glass, 9 inch by 1/2 inch
14. Volumetric flasks, 100 ml
3.2 Reagents
1. Isopropanol, spectrophotometric grade, ultraviolet cutoff 205 nm
(Burdick A Jackson or equivalent)
2. Test material for standard preparation
4. PROCEDURE:
4.1 Standard Preparation
4.1.1 Using a small beaker or 100 ml graduated cylinder, transfer
approximately 90 ml of isopropanol to a 100 ml volumetric flask.
4.1.2 Weigh an appropriate amount of the test material required to
prepare a concentrated stock standard solution, and quantita-
tively transfer the material to the volumetric flask.
NOTE: The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
4.1.3 Using a pasteur pipet, add a sufficient volume of isopropanol
to align the meniscus of the solution with the fiducial mark on
the flask. Stopper the flask and invert repeatedly to ensure
complete dissolution of the test material.
NOTE: Long term storage necessitates wrapping parafilm around
2.2-2
-------
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and set of calibration standards
by diluting the stock standard solution appropriately. The
calibration range should be established so as to incorporate
the expected concentrations of the samples.
NOTE: The analyst should consult Method 8.1 (Protocol for
Establishing a Calibration Range for Single Point
Analysis) to ensure the adequacy of the established
calibration range.
4.2 Sample Preparation
4.2.1 Using a glass stirring rod, mix each of the wet sediment samples
thoroughly to obtain as homogeneous a mixture as possible.
NOTE: In the event that an indigenous sediment blank is avail-
able for background correction, this reference sample
should be extracted and analyzed concurrently with the
'contaminated1 sediment samples.
4.2.2 Using a macro analytical balance and a stainless steel spatula,
weigh 5.0 grams of each sediment sample directly into tared
125 ml ground-glass stoppered erlenmeyer flasks.
4.2.3 Using a 50 ml graduated cylinder, quantitatively transfer 50 ml
of isopropanol into each of the flasks containing the sediment
samples. Stopper the flasks immediately to prevent potential
volatility losses of the extraction solvent.
4.2.4 Place the erlenmeyer flasks on a gyrotory shaker unit positioned
behind a safety shield in a fume hood. Adjust the agitation rate
of the shaker device to 400 RPM and extract the samples under
these conditions for exactly 30 minutes.
NOTE: During the gyrotory extraction procedure, the analyst
should periodically (e.g., at 5 minute intervals) turn
off the gyrotory shaker and vent the erlenmeyer flasks
to release any pressure buildup.
4.2.5 After 30 minutes, the gyrotory shaker should be turned off and
the erlenmeyer flasks removed to a stationary location. The
analyst should permit suspended solids and particulate matter
in the flasks to settle for approximately 30 minutes.
4.2.6 Using pasteur pipets, carefully transfer a sufficient volume
(e.g., 25 ml) of the isopropanol extracts to 30 ml crimp-top
serum bottles. DO NOT agitate the sediment layer in the flask
since particulates will interfere with fluorescence analysis.
2.2-3
-------
4.3 Fluorescence Analysis
4.3.1 Establish the fluorescence spectrophotometer conditions listed
in the Appendix of this Method.
4.3.2 Using a pasteur pipet, transfer a sufficient volume (e.g., 2 nil)
of the calibration solution to a quartz cuvette for analysis by
synchronous excitation fluorescence spectroscopy.
4.3.3 Observe the synchronous excitation spectra for each set of wave-
length intervals (e.g. A 20, A 30 A 100) over the range of
200 nm - 500 nm to establish the optimum wavelength interval for
quantitative measurement.
NOTE; The optimum wavelength interval selected should be a
compromise between maximum fluorescence intensity and
adequate peak resolution.
4.3.4 Using the optimum wavelength interval established in step 4.3.3
and the instrument conditions described in the Appendix, record
the synchronous excitation spectra of the calibration solution
and set of calibration standards in triplicate.
NOTE: Triplicate analyses of all calibration standards and
sample extracts are required to calculate a coefficient
of variation (Cv), which is a measure of the precision
or reproducibility of the analyses.
4.3.5 Record the synchronous excitation spectra of the solvent blank
(isopropanol), the sediment sample extracts, and the reference
sediment extract (if available) under identical instrumental
conditions.
4.3.6 Assure instrument stability by scanning the calibration solution
and solvent blank following each fifth sample measurement.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %)
5.3 Since the calibration range must bracket the concentration(s) of
fluorescing organic compounds measured in the sample extract(s), the
calibration standard concentrations may require further adjustment
2.2-4
-------
resulting in either a redefinition or refinement of the initially
established calibration range.
5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
compound detection limits, extraction recoveries, and potential inter-
ferences.
2.2-5
-------
APPENDIX
FLUORESCENCE SPECTROPHOTOMETER CONDITIONS
Initial Excitation Wavelength: 200 nm
* Initial Wavelength Interval (A): 20 nm
Initial Emission Wavelength: 220 nm
Final Excitation Wavelength: 500 nm
Final Emission Wavelength: 500 nm + A
Excitation Slit Width: 10 nm
Emission Slit Width: 10 nm
Wavelength Drive: Excitation
Scanning Mode: Dual Monochromators
Amplifier Dynode Voltage: Ratio
Recorder Power: Servo
The initial wavelength interval (A 20) requires an initial emission
wavelength of 220 nm with a starting excitation wavelength of 200 nm.
The starting emission wavelength is sequentially adjusted in 10 nm
increments to values of 230 nm, 240 nm, 250 nm, etc., to record the
synchronous excitation spectra for each set of wavelength intervals
(A 30, A 30, A 50, etc.), and establish the optimum wavelength interval
for quantitative measurement. The excitation monochromator is scanned
in synchronization with the emission monochromator over the wavelength
range of 200 nm - 500 nm to obtain a complete fluorescence spectrum of
the test material. The optimum wavelength interval selected should
be a compromise between maximum fluorescence intensity and adequate
peak resolution.
2.2-6
-------
SECTION III
METHOnOLOfiY FOR THE ANALYSIS OF PRIORITY POLLUTANTS
AND HAZARDOUS MATERIALS IN AIR SAMPLES
-------
CONTENTS
METHOD 3.1 Protocol for Performing Gas Chromatographic
Analysis of Air Samples
METHOD 3.2 Protocol for Preparing Analytical Standards
of Vaporized Materials
-------
METHOD 3.1 PROTOCOL FOR PERFORMING GAS CHROMATOGRAPHIC ANALYSIS OF AIR SAMPLES
1. SCOPE AND SUMMARY:
1.1 This Method addresses the determination of purgeable organics commonly
detected in contaminated air samples. Specifically, the following
compounds designated as hazardous materials may be determined by this
Method: acetone, benzene, carbon tetrachloride, chloroform, p-dioxane,
ethylene dichloride, methylene chloride, methyl ethyl ketone, styrene,
tetrachloroethylene, 1,1,1-trichloroethane, trichloroethylene, toluene,
and xylene.
1.2 This Method provides a procedure for rapidly extracting purgeables
from air samples and quantifying the extracts via gas chromatographic
analysis. The charcoal filter of an air sampling tube is extracted
with an appropriate volume of carbon disulfide. Flame ionization gas
chromatographic conditions are described which permit direct measure-
ment of the volatile organics in the extract.
2. CAUTIONS AND LIMITATIONS:
2.1 The extraction solvent (i.e., carbon disulfide) used in this procedure
is extremely flammable and toxic. Consequently, the entire extraction
procedure should be performed within a properly ventilated fume hood
since carbon disulfide readily volatilizes, emitting an odor similar
to that of spoiled eggs.
2.2 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., laboratory coat, safety glasses, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous and/or carcino-
genic substances.
2.3 All standards and samples prepared in carbon disulfide must he sealed
immediately (e.g., volumetric flasks stoppered and serum sample bottle
caps crimped) to prevent volatility losses.
2.4 The gas chromatographic conditions listed in Appendix B of this Method
must be established at least two hours prior to analysis of the cali-
bration standards and sample extracts to permit the flame ionization
detector to stabilize sufficiently.
3.1-1
-------
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Autosampler vials equipped with teflon lined septa
2. Beakers, assorted volumes
3. Decapper pliers for 5 ml crimp top serum bottles
4. Forceps, stainless steel
5. Funnel, glass
6. Gas chromatograph equipped with a flame ionization detector
7. Graduated cylinder, 10 ml
8. Hand crimpers for autosampler vials and serum bottles
9. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
10. Pasteur pipets, disposable, 6 or 9 inch
11. Serum bottles, crimp top, 5 ml (Wheaton or equivalent)
12. Syringes, 500 ul, 100 ul and 10 ul (Hamilton or equivalent)
13. Triangular file
14. Volumetric flasks, 100 ml, 50 ml, 25 ml and 10 ml
3.2 Reagents
1. Carbon disulfide, chromatographic grade (Burdick & Jackson or
equivalent)
2. Purgeable standard(s) (refer to Appendix A for a tabulation of
compounds).
4. PROCEDURE:
4.1 Standard Preparation
4.1.1 Using a small beaker or 10 ml graduated cylinder, transfer 9 ml
of carbon disulfide to a 10 ml volumetric flask. Stopper the
flask immediately to prevent volatility losses.
4.1.2 Using a micropipet or glass syringe, quantitatively withdraw an
appropriate volume of the purgeable standard(s) and dispense
this volume below the air/carbon disulfide interface in the flask
(refer to Appendix A of this Method for a list of purgeables).
NOTE: The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
4.1.3 Using a pasteur pipet, add sufficient volume of carbon disulfide
to align the meniscus of the solution with the fiducial mark on
the flask. Stopper the flask and invert repeatedly to ensure
complete dissolution of the purgeable compound(s).
3.1-2
-------
NOTE; Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and set of calibration standards
by diluting the stock standard solution appropriately. The
calibration range should be established so as to incorporate
the expected concentrations of the samples.
NOTE; The analyst should consult Method 8.1 (Protocol for
Establishing a Calibration Range for Single Point
Analysis) to ensure the adequacy of the established
calibration range.
4.2 Sample Preparation
4.2.1 File the sampling tube near the cotton plug in order to break
the glass without spilling any of the activated charcoal.
4.2.2 Remove the cotton plug with forceps and using a glass funnel,
empty the large section of charcoal into a 5 ml serum bottle
4.2.3 Remove the center urethane foam plug with forceps and empty the
smaller section of charcoal into a second 5 ml serum bottle.
NOTE; The analyst should label the serum sample bottles with
the following information: 1) Sample Number and Date;
2) Charcoal Weight (refer to step 4.2.4, Table I); and,
3) Location of Sample Collection.
4.2.4 According to the charcoal filter tube size, the analyst should
dispense (using micropipets) the appropriate volume of carbon
disulfide into each 5 ml serum sample bottle (refer to Table I
below). The sample bottles should be capped immediately to
prevent volatility losses.
TABLE I
Sampling Tube Size Volume of Carbon
(mg charcoal per section) Disulfide (ml)
100/50 0.5
200/50 1.0
400/200 2.0
600/200 3.0
4.2.5 Shake the sample bottles vigorously for exactly 30 seconds to
extract the purgeable compounds into the carbon disulfide.
Permit the contents of the sample bottles to settle for 30
minutes prior to analyzing the extracts by gas chromatographic
analysis
3.1-3
-------
4.3 Gas Chromatographic Analysis
4.3.1 Establish the gas Chromatographic conditions listed in Appendix
B of this Method.
NOTE; The flame ionization detector requires a minimum two
hour stabilization period prior to analysis of the
calibration standards and sample extracts.
4.3.2 Using pasteur pipets, transfer approximately 1 ml of each of
the calibration standards to autosampler vials. Seal the
autosampler vials with a hand crimper and position the vials
in the autoinjector with carbon disulfide blanks inserted at
regular intervals.
NOTE: In the event that an autoinjector is unavailable, the
analyst should use a precision quality 10 ul syringe to
manually inject an exact volume (e.g., usually 3 to 5 ul)
of all standard, sample, and carbon disulfide solutions.
4.3.3 Using a pair of decapper pliers, remove the crimp tops from the
5 ml serum sample bottles. Transfer approximately 1 ml of each
sample into autosampler vials, seal the vials immediately, and
position the samples in the autoinjector with carbon disulfide
blanks inserted at regular intervals.
4.3.4 -Replicate injections of all calibration standards and sample
extracts are required. A minimum of three injections of each
solution are necessary to calculate its coefficient of variation
(Cv), which is a measure of precision or reproducibility.-
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %).
5.3 Since the calibration range must bracket the concentration(s) of the
purgeable(s) measured in the sample extract(s), the calibration
standard concentrations may require further adjustment resulting in
either a redefinition or refinement of the calibration range.
3.1-4
-------
5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
volatile organic compound detection limits, extraction recoveries, and
potential interferences.
6. REFERENCES: NIOSH Method, Organic Solvents in Air. Method No. P&CAM 127.
3.1-5
-------
APPENDIX A
NIOSH Method Components Commonly Found In Air
Samples and Their Detection Limit (mg/sample)
COMPOUND DETECTION LIMIT
Acetone
Benzene 0.01
Carbon Tetrachloride 0.20
Chloroform 0.10
p-Dioxane 0.05
Ethylene dichloride 0.05
Methylene chloride 0.05
Methyl ethyl ketone 0.01
Styrene 0.10
Tetrachloroethylene 0.06
1,1,1-trichloroethane 0.05
Trichloroethylene 0.05
Toluene 0.01
Xylene 0.02
3.1-6
-------
APPENDIX B
GAS CHROMATOGRAPHIC CONDITIONS
Analytical Column: * 1% SP-1000 on 60/80 Carbopack B, 6 ft x 2 mm ID,
glass column, 9 Inch span
Carrier Gas: Helium
Carrier Flow Rate: 35 ml/min.
Detector: Flame lonization (FID)
Detector Temperature: 200°C
Injector Temperature: 200°C
Oven Temperature: t Initial = 60°C ; Final = 200°C
Program Rate: 8°C/min. ; Final Hold Time = 16 min.
Time of Analysis: 33.5 min.
t The Initial Oven Temperature is adjusted according to the sample
analysis requirements.
* The column recommended for purgeable analyses should be packed and
conditioned in accordance with Method 7.3 (Instructions for Packing
and Conditioning Glass Analytical Columns for Gas Chromatography
Instruments) and performance tested according to the procedure and
specifications described in Appendix D of Method 7.4 (Protocol for
Performance Testing Packed, Conditioned, Glass Analytical Columns
for Gas Chromatography Instruments).
3.1-7
-------
METHOD 3.2 PROTOCOL FOR PREPARING ANALYTICAL STANDARDS OF VAPORIZED MATERIALS
1. OVERVIEW: This method provides a procedure for preparing accurate standards
of vaporized materials in a rigid static chamber for gas chromato-
graphic analysis.
2. CAUTIONS AND LIMITATIONS:
2.1 The analyst should ensure the cleanliness of the serum reaction vials
(refer to step 4.1.1) and the glass syringes (refer to step 4.1.2).
2.2 The preparation of standards should be performed inside a fume hood
with the appropriate protective apparel (e.g., laboratory coat, safety
glasses, chemical resistant gloves) to minimize exposure of laboratory
personnel to hazardous and/or carcinogenic substances.
2.3 The analyst should ensure that all vials are labeled correctly with
compound identification, concentration, and date of preparation.
3. APPARATUS AND REAGENTS
3.1 Apparatus
1. Beakers, 250 ml
2. Hand crimper for 2 ml serum reaction vials
3. Septa discs, teflon or silicon, 22 mm diameter
4. Serum reaction bottles equipped with teflon lined screw caps, 40 ml
5. Serum reaction vials, 2 ml
6. Syringes, glass (1) 10 ul gas tight syringe, (2) 250 ul gas tight
syringe with side port needle, and (2) 500 ul gas tight syringe
with side port needle (Hamilton or equivalent)
7. Tygon tubing, 1/4" in, 2 foot length
8. Vacuum pump, capable of maintaining a pressure drop of 15" Hg
9. Volumetric pi pet, 1 ml
3.2 Reagents
1. Distilled water, organic free
1. Standard test materials, spectroscopic grade (not less than
98% purity)
3.2-1
-------
4. PROCEDURE
4.1 Preliminary Procedure
4.1.1 If visible particles are present in the serum reaction vials,
rinse the vials with distilled water and air dry. Store the
vials in an inverted position to prevent particle contamination.
4.1.2 Ensure the cleanliness of all syringes by drawing ambient air
into the syringe and injecting 3/4 of the total syringe volume
into the gas chromatograph.
NOTE: Ensure that the gas chromatographic conditions reflect
the most sensitive settings (i.e., Attn 2 1).
4.1.3 In the event that a peak occurs in the generated chromatogram,
disassemble the syringe by removing the plunger and connect the
tygon tubing to the air inlet tube adapter of the vacuum pump.
Set the vacuum pump at 15" Hg gauge pressure and place the lip
of the syringe barrel flush against the open end of the tygon
tubing for 20 minutes. Recheck the syringe for cleanliness as
described in step 4.1.2.
4.1.4 Record the ambient temperature (°C) and pressure (mm Hg) at the
time the vial dilutions are made.
4.1.5 The analyst should consult a reference handbook to obtain the
molecular weight and density of each high purity test material
analyzed.
4.2 Standard Preparation
4.2.1 Using a 1 ml volumetric pipet, quantitatively withdraw 1.0 ml of
a high purity (HP) standard material and transfer this volume to
a 2 ml serum reaction vial. Using a hand crimper, seal the vial
and label its contents.
NOTE: The analyst should ensure that a teflon or silicon septum
lines the interior surface of the aluminum cap.
4.2.2 Using a solvent rinsed, air dried, 10 ul glass syringe, quantita-
tively withdraw 5 ul of the HP standard material from the 2 ml
serum reaction vial and inject this volume into a 40 ml serum
reaction bottle.
NOTE 1: The analyst should ensure that a teflon or silicon
septum lines the interior surface of the aluminum cap.
NOTE 2: The 40 ml serum reaction bottles are quantitatively
43.3 mis, based on an average of 4 measurements of
the vial volume (Cv= 0.3%).
3.2-2
-------
4.2.3 Determine the concentration (in ppm) of the stock vapor solution
prepared in 4.2.2 using the microprocessor "Gas Phase Analysis"
program. Using this computer program, determine a dilution
procedure that will satisfy experimental objectives.
4.3 Dilution Sequence
4.3.1 The stock vapor solution in the serum reaction bottle should be
completely volatilized. The analyst should vigorously agitate
the bottle for approximately 15 seconds to ensure complete
volatilization of its contents.
NOTE: DO NOT touch the glass walls of the bottle when shaking.
Grasp the reaction vessel with both thumb and forefinger
securely holding the screw cap.
4.3.2 Insert a 250 ul or 500 ul gas tight syringe (with the plunger
fully depressed) into the serum reaction bottle and flush the
'syringe 5 times with the stock vapor solution without with-
drawing the needle from the vessel. Quantitatively withdraw
the exact volume required to prepare the vaporized standard
solution.
4.3.3 Carefully remove the syringe containing the volume of standard
material without disturbing the position of the syringe plunger.
Insert the syringe into a sealed 40 ml screw cap septum bottle,
depress the plunger, and then carefully withdraw the syringe.
NOTE: The syringe employed for standard preparation must be
cleansed according to step 4.1.3 prior to preparation
of additional standards.
4.3.4 The standard test atmosphere of vaporized material in air may
be analyzed by gas chromatography. In the event that further
dilutions are necessary, the "Gas Phase Analysis" program must
be employed to determine the correct dilution sequence.
3.2-3
-------
SECTION IV
METHOnOLDGY FOR THE ANALYSIS OF PETROLEUM OILS IN WATER AND SEDIMENT
-------
CONTENTS
METHOD 4.1 Rapid Extraction and Spectroscopic Analysis
of Petroleum Oils in Aqueous Samples
METHOD 4.2 Rapid Extraction and Spectroscopic Analysis
of Petroleum Oils in Sediment Samples
-------
METHOD 4.1 RAPID EXTRACTION AND SPECTROSCOPIC ANALYSIS
OF PETROLEUM OIL IN AOIJEODS SAMPLES
1. SCOPE AND SUMMARY
1.1 This Method specifically addresses the determination of petroleum oil
in aqueous samples. Since petroleum oils are comprised of a complex
mixture of organic constituents including polynuclear aromatic hydro-
carbons (PAHs), samples contaminated with petroleum oil are amenable
to fluorescence analysis.
1.2 This Method provides a procedure for rapidly extracting petroleum oil
from aqueous samples and quantifying the extracts via a sensitive
fluorescence spectroscopic technique. A 50.0 ml sample of wastewater
is extracted with an equivalent volume of isopropanol using a gyrotory
agitation technique to co-solubilize the petroleum oil. Synchronous
excitation fluorescence spectroscopy permits direct measurement of
the petroleum oil(s) in the extract.
2. CAUTIONS AND LIMITATIONS:
2.1 All glassware employed for standard and sample preparation must be
rinsed three times with isopropanol and air dried prior to use.
2.2 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., laboratory coat, safety glasses, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous and/or carcino-
genic substances.
2.3 This Method requires an ultrapure grade of isopropanol and cyclohexane;
the analyst should determine solvent purity by monitoring the emission
intensity of both solvents under the instrumental conditions described
in Appendix B of this Method.
NOTE: Interfering impurities in the isopropanol may be removed by
filtration through granular activated carbon in accordance
with the procedure of Method 6.4 (Protocol for Purifying
Isopropanol for Fluorescence Analyses).
2.4 Petroleum oil calibration standards must be prepared as prescribed in
this Method to assure proper solubility of the petroleum oil in water.
2.5 Aqueous samples suspected of containing petroleum oils and/or volatile
organics must be refrigerated at 4°C prior to extraction to prevent
sample degradation.
4.1-1
-------
2.6 Measurement of calibration standards, sample extracts, and solvent
blanks must be conducted under identical instrumental conditions of
analysis.
2.7 The spectrophotometer recorder baseline should be deflected at least
ten percent of full scale to avoid erroneous zero measurements.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Balance, macro analytical, range of 0 - 150 grams (± 0.1 mg)
2. Beakers, assorted volumes
3. Erlenmeyer flasks, ground-glass stoppered, 125 ml
4. Fluorescence spectrophotometer equipped with dual scanning
monochromators for synchronous measurement
5. Graduated cylinders, 250 ml, 100 ml, 50 ml
6. fiyrotory shaker (New Brunswick Scientific Company or equivalent)
7. Hand crimper for serum bottles
8. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
9. Pasteur pipets, disposable, 6 or 9 inch
10. Ouartz cuvette, teflon-stoppered, polished on four faces
11. Serum bottles, crimp top, 100 ml (Wheaton or equivalent)
12. Syringe, 50 ul
13. Volumetric flasks, 500 ml, 100 ml, 50 ml
3.2 Reagents
1. Cyclohexane, spectrophotometric grade, ultraviolet cutoff 204 nm
(Burdick & Jackson or equivalent)
2. Isopropanol, spectrophotometric grade, ultraviolet cutoff 205 nm
(Burdick & Jackson or equivalent)
3. Hi stilled water, organic free
4. Petroleum oil(s) for standard preparation
4. PROCEDURE:
4.1 Standard Preparation
4.1.1 Using a small beaker or 100 ml graduated cylinder, transfer
approximately 90 ml of a 50:50 cyclohexane:isopropanol solvent
mixture to a 100 ml volumetric flask.
4.1-2
-------
NOTE: The analyst should prepare the cyclohexane:isopropanol
solvent mixture according to the procedure described in
Appendix A of this Method.
4.1.2 Using an analytical balance and a 50 ul syringe, weigh 20 tug of
the petroleum oil standard and dispense this quantity below the
air/solvent interface in the flask.
NOTE: The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
4.1.3 Using a pasteur pi pet, add a sufficient volume of the 50:50
cyclohexane:isopropanol solvent mixture to align the meniscus
of the solution with the fiducial mark on the flask. Stopper
the flask and invert repeatedly to ensure complete dissolution
of the petroleum oil. The concentration of this stock solution
is 200 mg/L (parts per million).
NOTE: Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock solution.
4.1.4 Prepare a 20 mg/L stock standard solution of petroleum oil by
diluting 5 ml of the stock solution with 45 ml of isopropanol
in a 50 ml volumetric flask.
4.1.5 Prepare a calibration solution and set of calibration standards
by appropriate dilutions of the stock standard solution in a
50:50 isopropanol:water solvent mixture. The calibration range
should be established so as to incorporate the expected concen-
trations of the samples.
NOTE 1; The analyst should prepare the 50:50 isopropanol:water
solvent mixture according to the procedure described
in Appendix A of this Method.
NOTE 2: The analyst should consult Method 8.1 (Protocol for
Establishing a Calibration Range for Single Point
Analysis) to ensure the adequacy of the established
calibration range.
4.2 Sample Preparation
4.2.1 Using 50 ml graduated cylinders", transfer 50 ml of each aqueous
sample to 125 ml ground-glass stoppered erlenmeyer flasks.
NOTE: In the event that an indigenous water blank is available
for background correction purposes, this reference sample
should be extracted and analyzed concurrently with the
'contaminated1 aqueous samples.
4.1-3
-------
4.2.2 Using a 50 ml graduated cylinder, quantitatively transfer 50 ml
of isopropanol into each of the flasks containing the aqueous
samples. Stopper the flasks immediately to prevent potential
volatility losses of the extraction solvent.
4.2.3 Place the erlenmeyer flasks on a gyrotory shaker unit positioned
behind a safety shield in a fume hood. Adjust the agitation rate
of the shaker device to 400 RPM and extract the samples under
these conditions for exactly 5 minutes.
NOTE; Alternatively, the erlenmeyer flasks may be vigorously
agitated by hand providing that the analyst dons the
appropriate protective apparel (e.g., safety glasses,
laboratory coat, chemical resistant gloves).
4.2.4 After 5 minutes, the gyrotory shaker should be turned off and
the erlenmeyer flasks removed to a stationary location. The
analyst should vent the flasks to release any excess pressure.
4.2.5 Using pasteur pi pets, transfer the isopropanol:water solvent
extracts containing the co-solubilized petroleum oil(s) to 100
ml crimp-top serum bottles.
4.3 Fluorescence Analysis
4.3.1 Establish the fluorescence spectrophotometer conditions listed
in Appendix B of this Method.
4.3.2 Using a pasteur pipet, transfer a sufficient volume (e.g., 2 ml)
of the calibration solution to a quartz cuvette for analysis by
synchronous excitation fluorescence spectroscopy.
4.3.3 Observe the synchronous excitation spectra for each set of wave-
length intervals (e.g. A20, A30,..., A100) over the range of
200 nm - 500 nm to establish the optimum wavelength interval for
quantitative measurement.
NOTE: The optimum wavelength interval selected should be a
compromise between maximum fluorescence intensity and
adequate peak resolution.
4.3.4 Using the optimum wavelength interval established in step 4.3.3
and the instrument conditions described in the Appendix, record
the synchronous excitation spectra of the calibration solution
and set of calibration standards in triplicate.
NOTE: Triplicate analyses of all calibration standards and
sample extracts are required to calculate a coefficient
of variation (Cv), which is a measure of the precision
or reproducibility of the analyses.
4.1-4
-------
4.3.5 Record the synchronous excitation spectra of the solvent blank
(50:50 isopropanol:water), the sediment sample extracts, and
the reference sediment extract (pending availability) under
identical instrumental conditions.
NOTE; If further dilution of the sample extract(s) is required
for fluorescence analyses (i.e., sample responses fall
outside the confines of the calibration range), such
solutions must be prepared using the 50:50 isopropanol:
water solvent mixture to dilute the extract(s).
4.3.6 Assure instrument stability by scanning the calibration solution
and solvent blank following each fifth sample measurement.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %)
5.3 Since the calibration range must bracket the concentration(s) of
petroleum oil(s) measured in the sample extract(s), the calibration
standard concentrations may require further adjustment resulting in
either a redefinition or refinement of the initially established
calibration range.
5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
compound detection limits, extraction recoveries, and potential inter-
ferences.
4.1-5
-------
APPENDIX A
PREPARATION OF A 50:50 (V:V) CYCLOHEXANE;ISOPROPANOL SOLVENT MIXTURE
1. Dispense 250 ml of cyclohexane and 250 ml of isopropanol into separate 250 ml
graduated cylinders.
NOTE: Due to contraction, expansion, etc., associated with the mixing of two
organic phases, volume addition is not necessarily additive; therefore,
the volumes of cyclohexane and isopropanol must be measured separately
prior to actual preparation of the solvent mixture.
2. Transfer the 250 ml of cyclohexane from the graduated cylinder to a 500 ml
volumetric flask. Carefully dispense the 250 ml of isopropanol to the equiv-
alent volume of cyclohexane in the volumetric flask.
3. Stopper the flask and invert repeatedly to ensure uniform mixing of the two
organic solvents. The analyst should periodically vent the flask to release
any pressure generated by this mixing process.
NOTE; DO NOT adjust the volume of solution to align the meniscus with the
fiducial mark on the volumetric flask. The prepared solvent mixture
is 50% cyclohexane : 50% isopropanol (volume:volume).
PREPARATION OF A 50:50 (V:V) ISOPROPANOL:WATER SOLVENT MIXTURE
1. Dispense 250 ml of isopropanol and 250 ml of carbon filtered distilled water
into separate 250 ml graduated cylinders.
NOTE; Due to changes in the total volume of solution upon mixing, the volumes
of isopropanol and distilled water must be measured separately prior
to actual preparation of the solvent mixture.
2. Transfer the 250 ml of isopropanol from the graduated cylinder to a 500 ml
volumetric flask. Carefully dispense the 250 ml of distilled water to the
equivalent volume of isopropanol in the volumetric flask.
3. Stopper the flask and invert repeatedly to ensure uniform mixing of the two
solvents. The analyst should periodically vent the flask to release any
pressure generated by this mixing process.
NOTE; DO NOT adjust the volume of solution to align the meniscus with the
fiducial mark on the volumetric flask. The prepared solvent mixture
is 50% isopropanol : 50% water (volume:volume).
4.1-6
-------
APPENDIX B
FLUORESCENCE SPECTROPHOTOMETER CONDITIONS
Initial Excitation Wavelength: 200 nm
* Initial Wavelength Interval (A): 20 nm
Initial Emission Wavelength: 220 nm
Final Excitation Wavelength: 500 nm
Final Emission Wavelength: 500 nm + A
Excitation Slit Width: 10 nm
Emission Slit Width: 10 nm
Wavelength Drive: Excitation
Scanning Mode: Dual Monochromators
Amplifier Dynode Voltage: Ratio
Recorder Power: Servo
The initial wavelength interval (A 20) requires an initial emission
wavelength of 220 nm with a starting excitation wavelength of 200 nm.
The starting emission wavelength is sequentially adjusted in 10 nm
increments to values of 230 nm, 240 nm, 250 nm, etc., to record the
synchronous excitation spectra for each set of wavelength intervals
(A 30, A 40, A 50, etc.), and establish the optimum wavelength interval
for quantitative measurement. The excitation monochromator is scanned
in synchronization with the emission monochromator over the wavelength
range of 200 nm - 500 nm to obtain a complete fluorescence spectrum of
the petroleum oil. The optimum wavelength interval selected should
be a compromise between maximum fluorescence intensity and adequate
peak resolution. The wavelength interval of 40 nm enables optimum
quantification of the majority of petroleum oils.
4.1-7
-------
METHOD 4.2 RAPID EXTRACTION AND SPECTROSCOPIC ANALYSIS
OF PETROLEUM OIL IN SEDIMENT SAMPLES
1. SCOPE AND SUMMARY
1.1 This Method specifically addresses the determination of petroleum oil
1n sediment samples. Since petroleum oils are comprised of a complex
mixture of organic constituents including polynuclear aromatic hydro-
carbons (PAHs), samples contaminated with petroleum oil are amenable
to fluorescence analysis.
1.2 This Method provides a procedure for rapidly extracting petroleum oil
from sediment samples and quantifying the extracts via a sensitive
fluorescence spectroscopic technique. A 5.0 gram sample of contamin-
ated sediment is weighed into a 125 ml ground-glass stoppered erlen-
meyer flask and extracted with a 50:50 isopropanol: cyclohexane
solvent mixture using a gyrotory agitation technique. Synchronous
excitation fluorescence spectroscopy permits direct measurement of
petroleum oil(s) in the extract.
?.. CAUTIONS AND LIMITATIONS:
2.1 All glassware employed for standard and sample preparation must he
rinsed three times with isopropanol and air dried prior to use.
2.2 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., chemical resistant gloves, safety glasses, etc.) to minimize
exposure of laboratory personnel to hazardous and/or carcinogenic
substances.
2.3 This Method requires an ultrapure grade of isopropanol; the analyst
should determine solvent purity by monitoring the emission intensity
of isopropanol under the instrumental conditions described in the
Appendix of this Method.
NOTE: Interfering impurities in the isopropanol may be removed by
filtration through granular activated carbon in accordance
with the procedure of Method 6.4 (Protocol for Purifying
Isopropanol for Fluorescence Analyses).
2.4 Sediments suspected of containing petroleum oils and/or volatile
organics must be refrigerated at 4°C prior to extraction to prevent
sample degradation.
4.2-1
-------
2.5 Measurement of calibration standards, sample extracts, and solvent
blanks must be conducted under identical instrumental conditions of
analysis.
2.6 The spectrophotometer recorder baseline should be deflected at least
ten percent of full scale to avoid erroneous zero measurements.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Balance, macro analytical, range of 0 - 150 grams (± 0.1 mg)
2. Beakers, assorted volumes
3. Erlenmeyer flasks, ground-glass stoppered, 125 ml
4. Fluorescence spectrophotometer equipped with dual scanning
monochromators for synchronous measurement
5. Graduated cylinders, 250 ml, 100 ml, 50 ml
6. Gyrotory shaker (New Brunswick Scientific Company or equivalent)
7. Hand crimper for serum bottles
8. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
9. Pasteur pi pets, disposable, 6 or 9 inch
10. Quartz cuvette, teflon-stoppered, polished on four faces
11. Serum bottles, crimp top, 30 ml (Wheaton or equivalent)
12. Spatula, stainless steel
13. Stirring rod, glass, 9 inch by 1/2 inch
14. Syringe, 50 ul
15. Volumetric flasks, 500 ml, 100 ml
3.2 Reagents
1. Cyclohexane, spectrophotometric grade, ultraviolet cutoff 204 nm
(Burdick & Jackson or equivalent)
2. Isopropanol, spectrophotometric grade, ultraviolet cutoff 205 nm
(Burdick & Jackson or equivalent)
3. Petroleum oil(s) for standard preparation
4. PROCEDURE:
4.1 Standard Preparation
4.1.1 Using a small beaker or 100 ml graduated cylinder, transfer
approximately 90 ml of a 50:50 cyclohexane:isopropanol solvent
mixture to a 100 ml volumetric flask.
NOTE; The analyst should prepare the cyclohexane:isopropanol
4.2-2
-------
solvent mixture according to the procedure described in
Appendix A of this Method.
4.1.2 Using an analytical balance and a 50 ul syringe, weigh 20 mg of
the petroleum oil standard and dispense this quantity below the
air/solvent interface in the flask.
NOTE: The analyst should consult Method 5.1 (Protocol for Pre-
paring Analytical Standards of Hazardous and Carcinogenic
Materials) for proper standard preparation techniques.
4.1.3 Using a pasteur pipet, add a sufficient volume of the 50:50
cyclohexane:isopropanol solvent mixture to align the meniscus
of the solution with the fiducial mark on the flask. Stopper
the flask and invert repeatedly to ensure complete dissolution
of the petroleum oil. The concentration of this stock standard
solution is 200 mg/L (parts per million).
NOTE: Long term storage necessitates wrapping parafilm around
the stopper and refrigerating the flask at 4°C to prevent
evaporation and/or degradation of the stock standard
solution.
4.1.4 Prepare a calibration solution and set of calibration standards
by diluting the stock standard solution appropriately. The
calibration range should be established so as to incorporate
the expected concentrations of the samples.
NOTE 1: All calibration standards must be prepared in the
50:50 cyclohexane:isopropanol solvent mixture.
NOTE 2: The analyst should consult Method 8.1 (Protocol for
Establishing a Calibration Range for Single Point
Analysis) to ensure the adequacy of the established
calibration range.
4.2 Sample Preparation
4.2.1 Using a glass stirring rod, mix each of the wet sediment samples
thoroughly to obtain as homogeneous a mixture as possible.
NOTE: In the event that an indigenous sediment blank is avail-
able for background correction, this reference sample
should be extracted and analyzed concurrently with the
'contaminated' sediment samples.
4.2.2 Using a macro analytical balance and a stainless steel spatula,
weigh 5.0 (± 0.1) grams of each sediment sample directly into
tared 125 ml ground-glass stoppered erlenmeyer flasks.
4.2.3 Using a 50 ml graduated cylinder, quantitatively transfer 50 ml
of the 50:50 isopropanol:cyclohexane solvent mixture into each
4.2-3
-------
of the flasks containing the sediment samples. Stopper the
flasks immediately to prevent potential volatility losses of
the extraction solvent.
4.2.4 Place the erlenmeyer flasks on a gyrotory shaker unit positioned
behind a safety shield in a fume hood. Adjust the agitation rate
of the shaker device to 400 RPM and extract the samples under
these conditions for exactly 30 minutes.
NOTE; During the gyrotory extraction procedure, the analyst
should periodically (e.g., at 5 minute intervals) turn
off the gyrotory shaker and vent the erlenmeyer flasks
to release any pressure buildup.
4.2.5 After 30 minutes, the gyrotory shaker should be turned off and
the erlenmeyer flasks removed to a stationary location. The
analyst should permit suspended solids and particulate matter
in the flasks to settle for approximately 30 minutes.
4.2.6 Using pasteur pipets, carefully transfer a sufficient volume
(e.g., 25 ml) of the isopropanol extracts to 30 ml crimp-top
serum bottles. 00 NOT agitate the sediment layer in the flask
since particulates will interfere with fluorescence analysis.
4.3 Fluorescence Analysis
4.3.1 Establish the fluorescence spectrophotometer conditions listed
in Appendix R of this Method.
4.3.2 Using a pasteur pipet, transfer a sufficient volume (e.g., 2 ml)
of the calibration solution to a quartz cuvette for analysis by
synchronous excitation fluorescence spectroscopy.
4.3.3 Observe the synchronous excitation spectra for each set of wave-
length intervals (e.g. A 20, A 30,..., A 100) over the range of
200 nm - 500 nm to establish the optimum wavelength interval for
quantitative measurement.
NOTE: The optimum wavelength interval selected should be a
compromise between maximum fluorescence intensity and
adequate peak resolution.
4.3.4 Using the optimum wavelength interval established in step 4.3.3
and the instrument conditions described in the Appendix, record
the synchronous excitation spectra of the calibration solution
and set of calibration standards in triplicate.
NOTE: Triplicate analyses of all calibration standards and
sample extracts are required to calculate a coefficient
of variation (Cv), which is a measure of the precision
or reproducibility of the analyses.
4.2-4
-------
4.3.5 Record the synchronous excitation spectra of the solvent blank
(50:50 cyclohexanerisopropanol), the sediment sample extracts,
and the reference sediment extract (pending availability) under
identical instrumental conditions.
NOTE: If further dilution of the sample extract(s) is required
for fluorescence analyses (i.e., sample responses fall
outside the confines of the calibration range), such
solutions must be prepared using the 50:50 cyclohexane:
isopropanol solvent mixture to dilute the extract(s).
4.3.6 Assure instrument stability by scanning the calibration solution
and solvent blank following each fifth sample measurement.
5. CALCULATIONS:
5.1 All calculations must be performed in accordance with Method 8.1
(Protocol for Establishing a Calibration Range for Single Point
Analysis) employing the appropriate data sheets and microprocessor
programs listed in Method 8.2.
5.2 The analyst must assure the integrity of the established calibration
range (e.g., maximum allowable calibration error = ± 10 %) and the
precision of the analyses (e.g., Cv values < 3 %)
5.3 Since the calibration range must bracket the concentration(s) of
petroleum oil(s) measured in the sample extract(s), the calibration
standard concentrations may require further adjustment resulting in
either a redefinition or refinement of the initially established
calibration range.
5.4 Method validation data (e.g., synthetic samples, reagent blanks, etc.)
should be generated concurrently with sample analysis data to determine
compound detection limits, extraction recoveries, and potential inter-
ferences.
4.2-5
-------
APPENDIX A
PREPARATION OF A 50:50 (V:V) CYCLOHEXANE:ISOPROPANOL SOLVENT MIXTURE
1. Dispense 250 nil of cyclohexane and 250 ml of Isopropanol Into separate 250 ml
graduated cylinders.
NOTE: Due to contraction, expansion, etc., associated with the mixing of two
organic phases, volume addition is not necessarily additive; therefore,
the volumes of cyclohexane and isopropanol must be measured separately
prior to actual preparation of the solvent mixture.
?.. Transfer the 250 ml of cyclohexane from the graduated cylinder to a 500 ml
volumetric flask. Carefully dispense the 250 ml of isopropanol to the equiv-
alent volume of cyclohexane in the volumetric flask.
3. Stopper the flask and invert repeatedly to ensure uniform mixing of the two
organic solvents.- The analyst should periodically vent the flask to release
any pressure generated by this mixing process.
NOTE; DO NOT adjust the volume of solution to align the meniscus with the
fiducial mark on the volumetric flask. The prepared solvent mixture
is 50% cyclohexane : 50% isopropanol (volume:volume).
4.2-6
-------
APPENDIX B
FLUORESCENCE SPECTROPHOTOMETER CONDITIONS
Initial Excitation Wavelength: 200 nm
* Initial Wavelength Interval (A): 20 nm
Initial Emission Wavelength: 220 nm
Final Excitation Wavelength: 500 nm
Final Emission Wavelength: 500 nm + A
Excitation Slit Width: 10 nm
Emission Slit Width: 10 nm
Wavelength Drive: Excitation
Scanning Mode: Dual Monochromators
Amplifier Dynode Voltage: Ratio
Recorder Power: Servo
* The initial wavelength interval (A 20) requires an initial emission
wavelength of 220 nm with a starting excitation wavelength of 200 nm.
The starting emission wavelength is sequentially adjusted in 10 nm
increments to values of 230 nm, 240 nm, 250 nm, etc., to record the
synchronous excitation spectra for each set of wavelength intervals
(A 30, A 40, A 50, etc.), and establish the optimum wavelength interval
for quantitative measurement. The excitation monochromator is scanned
in synchronization with the emission monochromator over the wavelength
range of 200 nm - 500 nm to obtain a complete fluorescence spectrum of
the petroleum oil. The optimum wavelength interval selected should
be a compromise between maximum fluorescence intensity and adequate
peak resolution. The wavelength interval of 40 nm enables optimum
quantification of the majority of petroleum oils.
4.2-7
-------
SECTION V
PROTOCOLS FOR THE PREPARATION OF ANALYTICAL STANPARnS ANO SYNTHETIC SAMPLES
-------
CONTENTS
METHOD 5.1 Protocol for Preparing Analytical Standards
of Hazardous and Carcinogenic Materials
METHOD 5.2 Protocol for Preparing Synthetic Samples
of Hazardous Materials in Water
METHOD 5.3 Protocol for Preparing Oil/Water Dispersions
by Ultrasonification
-------
METHOD 5.1 PROTOCOL FOR PREPARING ANALYTICAL STANDARDS
OF HAZARDOUS AND CARCINOGENIC MATERIALS
1. OVERVIEW: This Method provides a detailed procedure for the proper handling
of hazardous and carcinogenic materials when preparing analytical
standards from pure compounds or highly concentrated solutions.
2. CAUTIONS AND LIMITATIONS:
2.1 Materials must be transported into and out of the glovebox through
the transfer box (airlock). DO NOT open the main side-panel door
of the glovebox.
2.2 The preparation of standards must be performed with the appropriate
protective apparel (e.g., laboratory coat, safety glasses, chemical
resistant gloves) to minimze exposure of laboratory personnel to
hazardous and/or carcinogenic substances.
2.3 In the event that a spill should occur when working in the glovebox
or hood, decontaminate the area before proceeding with the operation.
3. APPARATUS AND REAGENTS
3.1 Apparatus
1. Balance, macro analytical, range of 0 - 150 grams (± 0.1 mg)
2. Glass funnel
3. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
4. Pasteur pipets, disposable, 6 or 9 inch
5. Plastic bags (for disposal)
6. Polyethelene-backed absorbent paper (Benchkote or equivalent)
7. Syringe, glass, 100 ul (Hamilton or equivalent)
8. Spatula, stainless steel
9. Volumetric flasks
10. Weighing vials
3.2 Reagents
1. Test material for standard preparation
2. Hexane, chromatographic grade (Rurdick & Jackson or equivalent)
3. Solvent(s) compatible with analysis
5.1-1
-------
4. PROCEDURE
4.1 Preparation of Glovebox
4.1.1 Decontaminate the glovebox if necessary. Be sure to wear
disposable gloves when performing this procedure.
1. Place plastic bags, hexane, and soap and water into the
glovebox through the transfer box.
a. Open the outer access door of the transfer box. Place
the items into the transfer box, then securely close
the outer access door.
b. Working through the gloves, open the inner access door.
Remove the items from the transfer box, then securely
close the inner access door.
2. Place any contaminated liner which is in the glovebox into
DOUBLE PLASTIC BAGS; seal the bags by knotting at the top.
3. Wipe down the inside of the glovebox with hexane, then with
soap and water. Double plastic bag all clean-up material;
seal the bags by knotting at the top.
4. Remove the contaminated material from the glovebox through
the transfer box using the procedure opposite from that used
to place objects into the glovebox (above).
5. Deposit the contaminated lining and clean-up material into a
5 gallon waste drum. Label the waste drum accordingly (e.g.,
"PCB-Contaminated Solids").
NOTE: All contaminated wastes must be double plastic bagged
before being deposited in the waste drum.
4.1.2 Re-line the glovebox with clean polyethelene-backed absorbent
paper. Again, be sure to place the paper into the glovebox
through the transfer box in the correct manner.
4.1.3 Place all glassware, pipettes, spatulas, solvents, and other
needed equipment into the glovebox through the transfer box.
It is suggested that solvents be contained in automatic
dispensing bottles.
4.1.4 Turn on the glovebox blower, being sure that the exhaust is
vented into a fumehood. Adjust the Differential Pressure
Gauge to a reading of between 0.5 and 1.0.
5.1-2
-------
4.2 Preparation of a Solution from a Pure Solid
4.2.1 Weighing from an Initial Amount > 100 mg
1. For any transfer of a solid material, glass screw-cap vials
are to be used. Rinse the vial and cap with the appropriate
solvent and allow to dry before proceeding.
2. Weigh the empty vial with the cap on the analytical balance.
Record this amount as the tare weight.
3. Place the vial into the glovebox through the transfer box.
4. Working through the gloves, rinse the vial and cap with the
solvent to be used in the analysis. Allow these items to
dry before proceeding.
5. Using a spatula, transfer an approximate amount of the pure
material to be weighed into the vial. Cap the vial, rinse
with solvent, and permit to dry.
6. Remove the vial from the glovehox through the transfer box.
Weigh the vial on the analytical balance. Record this as
the tare + sample weight, the difference between the two
being the actual sample weight.
7. If more sample is needed, repeat steps 5 and 6.
8. Return the vial to the glovebox through the transfer box.
Using the appropriate solvent, rinse the inside of the vial
and pour the contents through a pre-rinsed glass funnel into
a volumetric flask which contains a small amount of solvent.
Repeat this rinsing at least three times. Rinse through the
funnel into the flask to ensure that all material is in
solution.
9. Dilute to volume in the flask as usual.
10. Remove the empty vial from the glovebox through the transfer
box. nispose of by depositing in the appropriate waste
container.
11. Remove contaminated lining from the glovebox as in step 4.1.
4.2.2 Weighing from an Initial Amount > 100 mg
1. Working in a fumehood, remove the plastic seal from the
standard bottle. Rinse the outside of the bottle with the
solvent to be used; let dry.
NOTE: If the standard bottle has a paper label, DO NOT rinse
5.1-3
-------
the outside with solvent, but just wipe thoroughly
with a disposable paper wipe.
2. Weigh the standard bottle on the analytical balance. Record
this weight as the tare + sample.
3. Place the bottle into the glovebox through the transfer box.
Fill the bottle half way with the solvent to be used in the
analysis, cap, swirl, and pour the liquid through a pre-rinsed
funnel into a volumetric flask which contains a small amount
of solvent. Repeat this rinsing at least three times.
Dilute to volume as usual.
NOTE 1; Ensure that the inside of the cap is rinsed during
this process, since some compounds tend to collect
here.
NOTE 2: It is recommended that the total contents of the
standard bottle be used, so that maximum accuracy
will be achieved in weighing such a small amount of
material.
4. Allow the vial to dry, cap it, solvent rinse the outside,
allow to dry. Remove the vial from the glovebox through
the transfer box.
5. Weigh the vial on the analytical balance. Record this weight
as the tare. The difference between this weight and that
recorded in step 2 above is the actual sample weight.
6. Dispose of the empty vial by depositing in the proper waste
container.
5.1-4
-------
METHOD 5.2 PROTOCOL FOR PREPARING SYNTHETIC SAMPLES
OF HAZARDOUS MATERIALS IN WATER
1. OVERVIEW: This method provides a procedure for preparing synthetic samples
of hazardous materials in water to evaluate the efficacy of
extraction methodology.
2. CAUTIONS AND LIMITATIONS:
2.1 The aqueous solubility of the hazardous material must be checked prior
to preparing synthetic samples of the material in water.
2.2 The preparation of standards and synthetic samples must be performed
in a properly ventilated fume hood with the appropriate protective
apparel (e.g., laboratory coat, safety glasses, chemical resistant
gloves) to minimize exposure of laboratory personnel to hazardous
and/or carcinogenic substances.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Beakers, assorted volumes
2. Heating mantle
3. Magnetic stir bars, 2 inch length
4. Magnetic stirring unit
5. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
6. Syringe, glass, 100 ul and 10 ul (Hamilton or equivalent)
7. Volumetric flasks, assorted volumes
3.2 Reagents
1. Distilled water, organic free
2. Hazardous material standards
5.2-1
-------
4. PROCEDURE:
4.1 Water Soluble Hazardous Materials
4.1.1 Using a small beaker, transfer approximately 90 ml of organic
free distilled water to a 100 ml volumetric flask.
4.1.2 Weigh an appropriate amount of the hazardous material required
to prepare the synthetic sample, and quantitatively transfer
the material to the volumetric flask.
NOTE 1: The analyst should consult Method 5.1 (Protocol for
Preparing Analytical Standards of Hazardous and Car-
cinogenic Materials) for proper standard preparation
techniques.
NOTE 2: The analyst should ensure that the quantity of material
is soluble in the volume of water being used.
NOTE 3: If the material is already in an organic solvent (e.g.,
a commercially prepared stock solution), the volume of
the standard solution added should be minimal (e.g.,
< 100 ul per liter) to ensure the aqueous solubility
of the standard and solvent.
4.1.3 Using a pasteur pipet, add a sufficient volume of distilled
water to align the meniscus of the solution with the fiducial
mark on the flask. Stopper the flask and invert repeatedly
to ensure complete dissolution of the hazardous material.
4.1.4 The concentration of the synthetic sample should fall within
the concentration range of the actual samples. In the event
that sample concentrations are unknown, the concentration of
the synthetic sample should be 5 to 15 times greater than the
concentration of the detection limit for the material.
4.1.5 Prepare the synthetic sample by dilution of the stock solution
through use of the appropriate microprocessor program (refer
to Method 8.2 of the Manual).
NOTE 1: The volume of concentrated aqueous solution that is
added should not be more than 10% of the final volume
(e.g., 10 ml of a concentrated solution of hazardous
material in water should be the maximum volume that is
added to 90 ml of water).
NOTE 2: Dilution factors must be included in the calculations.
4.2 Non-Volatile. Partially Water Soluble Hazardous Materials
4.2.1 Fill a beaker with approximately two-thirds of the volume of
5.2-2
-------
water required to prepare the synthetic sample and heat the
water to 60 - 70°C.
4.2.2 Pour the contents of the beaker Into a 100 ml volumetric flask
containing a weighed amount of the hazardous material.
NOTE: Follow the guidelines In step 4.1 to determine the amount
of hazardous material to add to the volumetric flask.
4.2.3 Stopper the flask and invert repeatedly to ensure complete dis-
solution of the hazardous material.
NOTE: It may be necessary to stir the contents of the flask for
30 minutes to one hour using a stir bar and magnetic stir-;
ring unit to achieve complete dissolution of the compound.
4.2.4 Permit the solution to cool to room temperature while ensuring
that no precipitation occurs. In the event that the hazardous
material precipitates out of solution, it may be necessary to
use a co-solvent to dissolve the material in the water (refer
to the Appendix of this Method).
4.2.5 Using a small beaker or pasteur pi pet, add a sufficient volume
of distilled water to align the meniscus of the solution with
the fiducial mark on the flask. Stopper the flask and invert
repeatedly to ensure complete dissolution of the hazardous
material.
5.2.3
-------
APPENDIX
USE OF A CO-SOLVENT TO DISSOLVE RELATIVELY INSOLUBLE HAZARDOUS MATERIALS
1. Dissolve an amount of hazardous material in a solvent in which the material
is soluble. The concentration of this solution should be significantly
greater than that desired for the synthetic sample.
NOTE: In the event that the material is already dissolved in an organic
solvent (e.g., a commercially prepared stock solution), the volume
of the standard added in an organic solvent solution should be kept
minimal (i.e., 100 ul/L or less) so that the solubility of the water
for the material will not be impaired.
2. Prepare the desired concentration of the synthetic sample by dilution (refer
the appropriate microprocessor program of Method 8.2 in the Manual).
5.2-4
-------
METHOD 5.3 PROTOCOL FOR PREPARING OIL/WATER DISPERSIONS BY ULTRASONIFICATION
1. OVERVIEW: This Method provides a procedure for preparing oil-in-water
dispersions to evaluate the efficacy of petroleum oil extraction
methodology.
2. CAUTIONS AND LIMITATIONS:
2.1 This Method is not applicable for preparing dispersions of viscous
oils (i.e., No. 6 Fuel Oil, Bachacquaro Crude Oil).
2.2 Lubricating stock oils containing detergent additives cannot be ultra-
sonified in water effectively due to foaming.
2.3 Ice baths must be used at various stages of the Method to prevent loss
of volatile organics.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Balance, macro analytical, range of 0 - 150 grams (± 0.1 mg)
2. Beaker with spring clamp, 1 L
3. Graduated cylinder, ground-glass stoppered, 50 ml
4. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
5. Styrofoam cooler
6. Syringe, glass, 2 ml (Hamilton or equivalent)
7. Ultrasonifier with standard horn and flat tip probe (Branson or
equivalent)
8. Volumetric flasks, 1.0 L and 250 ml
3.2 Reagents
1. Distilled water, organic free
2. Ice cubes
3. Test oil for standard preparation
5.3-1
-------
4. PROCEDURE:
4.1 Place several 1.0 liter volumetric flasks containing refrigerated
distilled water in a styrofoam cooler with an ice water bath.
4.2 Place a 50 ml graduated cylinder in the 1 liter beaker and secure it
with a triangle spring support. Add 50 ml of refrigerated distilled
water to the graduated cylinder.
4.3 Fill the 1 liter beaker half way with ice cubes and add refrigerated
distilled water until the water level is just above the 50 ml mark
on the graduated cylinder.
4.4 Weigh the 2 ml syringe containing approximately 1 ml of test oil and
quantitatively dispense 100 mg of the test oil into the graduated
cylinder. Reweigh the syringe, and repeat if necessary to obtain a
100 mg transfer.
NOTE: The analyst should consult Method 5.1 (Protocol for Preparing
Analytical Standards of Hazardous and Carcinogenic Materials)
for proper standard preparation techniques.
4.5 Place the graduated cylinder with ice bath into the ultrasonifier
sound box. Drop the probe through the top opening in the sound box
so that the probe tip enters the graduated cylinder without touching
the sides. Adjust the height of the graduated cylinder until the
probe tip is immersed 1/8" into the water.
4.6 Close and lock the sound box door.
4.7 Turn the power output control to position 5 and the timer/energizer
control to 1 minute. After 15 seconds, slowly increase the power
output to maximum for the remaining time.
4.8 When ultrasonfication is complete, open the sound box and remove the
ice bath and graduated cylinder. Unclamp the graduated cylinder and
remove it from the ice bath. Stopper and shake the cylinder a few
times.
4.9 Transfer the dispersion to a 250 ml volumetric flask which contains
100 ml of refrigerated distilled water; place the flask in the
styrofoam cooler ice bath.
4.10 Add another 50 ml of refrigerated distilled water to the graduated
cylinder and reposition it in the 1 liter beaker containing ice water.
Adjust the water level to just above the 50 ml mark on the graduated
cylinder.
4.11 Place the beaker and graduated cylinder back into the sound box as in
steps 4.5 and 4.6. Sonify the solution for 1 minute at maximum power.
4.12 After the second sonification, repeat steps 4.8 and 4.9.
5.3-2
-------
4.13 Add a sufficient volume of refrigerated distilled water to align the
meniscus of the solution with the fiducial mark on the 250 ml flask.
4.14 Invert the flask 20 times, and make dilutions as needed to yield the
required sample concentrations (refer to the Appendix of this Method),
NOTE: Keep the flask containing the dispersion in the ice bath as
much as possible during handling.
5.3-3
-------
APPENDIX
EXAMPLE OF DILUTION SCHEME
1. Concentration of oil in water required 2.5 mg/L
2. 103.4 mg oil was transferred and dispersed in 50 ml of water.
3. 50 ml of dispersion concentrate is diluted to 1 liter with water.
Concentration of oil in 1 liter 0.1 nig/ml
4. 25 ml aliquot from the 1 liter flask is diluted to 1 liter. Concentration
of this dispersion:
25 ml aliquot x 0.1 mg = 2.5 mg in 1 liter (2.5 mg/L)
0.1 mg
ml
5.3-4
-------
SECTION VI
SPECIALIZED SAMPLE PREPARATION AND TREATMENT TECHNIQUES
-------
CONTENTS
METHOD 6.1 Protocol for the Selection and Use of Chromatographic
Cartridges for Environmental Sample Cleanup
METHOD fi.2 Protocol for Removing Elemental Sulfur from
Sediment Extracts
METHOD fi.3 Protocol for Weathering Small Ouantities of
Petroleum Oil
METHOD fi.4 Protocol for Purifying Isopropanol for
Fluorescence Analyses
-------
METHOD 6.1 PROTOCOL FOR THE SELECTION AND USE OF CHROMATOGRAPHIC
CARTRIDGES FOR ENVIRONMENTAL SAMPLE CLEANUP
1. OVERVIEW: This Method provides a rapid procedure for selecting the most
efficient chromatographic cartridge for environmental sample
cleanup.
2. CAUTIONS AND LIMITATIONS:
2.1 The purpose of sample cleanup is to simplify the analysis of one or
more compounds by removing interferences from a complex sample matrix.
2.2 A typical sample extract contains compounds of varying chromatographic
polarity and thus, the choice of a proper cleanup cartridge is crucial
in preventing inadvertent loss of the compound(s) of interest.
2.3 Refer to Appendix A of this Method for a list of organic compounds and
the recommended chromatographic cartridges.
2.4 If any of the compounds of interest are not located within Appendix A
of this Method, the analyst should consult Appendix B for evaluating
chromatographic cartridges.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Alumina Cartridges, CIS Cartridges, Florisil Cartridges, and
Silica Gel cartridges, (Waters Associates Inc., Sep-pak or
equivalent)
2. Conical centrifuge tubes, ground glass stoppered
3. Desktop computer (Hewlett-Packard Model 9825A or equivalent)
4. Microliter syringes, gas-tight, assorted volumes
5. Micropipets, adjustable volume (Scientific Manufacturing Industries
or equivalent)
6. Pasteur pipets, disposable, 6 or 9 inch
7. Pipette bulb
3.2 Reagents
1. Appropriate solvent(s), spectrophotometric grade, as dictated by
the analysis procedure
2. Pure standard(s) of the compound(s) of interest
6.1-1
-------
4. PROCEDURE
4.1 After proper selection of the chromatographic cartridge (refer to
Appendix A), the cartridge should be rinsed with 2 to 3 ml of the
appropriate solvent using a Micro/pettor.
4.2 Using the same Micro/pettor, inject air through the cartridge to
remove any excess solvent.
4.3 Using a gas-tight syringe, withdraw the desired extract volume from
the sample.
4.4 Place the solvent rinsed cartridge at the opening of a conical
centrifuge tube and inject the extract into the longer extension
of the cartridge.
Note: Care should be taken when injecting the sample into the
cartridge extension to avoid overfilling the orifice and
subsequent sample loss.
4.5 With a Micro/pettor set at the desired solvent volume, slowly inject
solvent through the cartridge with sufficient pressure to force the
sample through the cartridge.
Note: The final extract should be colorless. If any colorization
is noticed, the cartridge may have been overloaded. If so,
repeat the entire procedure.
4.6 After the desired amount of solvent has been applied to the cartridge,
inject air through the cartridge to remove any residual sample/solvent.
Note: Check the approximate volume of the sample in the centrifuge
tube to be certain that the desired final volume has been
achieved.
4.7 Place a ground glass stopper on the centrifuge tube and mix thoroughly
for 10 seconds.
4.8 The sample extract is now "cleaned-up" and is ready for further
selective isolation or analysis as dictated by its relative procedure.
5. CALCULATIONS
5.1 The final extract concentration is calculated through use of the
following relation:
cE = (QS/VT)*(VE/VF)
6.1-2
-------
where; CE = Final extract concentration
QS = Quantity of sample; volume (ml), or weight (mg)
Vj = Total volume (extraction solvent + sample)
VE = Volume of sample extract applied to cartridge
Vp = Final volume of sample extract collected after cleanup
5.2 The final surrogate concentration is calculated through use of the
following relation:
cs = (CSS)*(VS/VF)
where; C$ = Final surrogate concentration
C$s= Concentration of surrogate standard
V$ = Volume of surrogate standard applied to cartridge
Vp = Final volume of sample extract collected after cleanup
6.1-3
-------
SELECTION OF CHROMATOGRAPHIC CARTRIDfiES
cr>
COMPOUND
CARTRIDGE
Aflatoxins
Alkaloids
Benzidine
Chlordane
Chlorinated Hydrocarbons
Di phenylhyd razi ne
Haloethers
Herbicides
Lipids
Mycotoxins
Nitrosamines
Florisil
Florisil
Florisil or Silica Gel
Florisil
Florisil
Cl8
Florisil
Florisil
Florisil
Florisil
Florisil
COMPOUND
CARTRIDGE
N-Nitrosodiphenylaminel
Organochlorine Pesticides2
Pesticides
Phenols3
Phthalate Esters
Polychlorinated Biphenyls
Steroids
TCDD4
Toluene^
Toxaphene
Florisil
Florisil
Florisil
Silica Gel
Florisil or Silica Gel
Florisil
Florisil
Silica Gel/Alumina
C18
Florisil
-o
m
1 N-Nitrosodiphenylamine decomposes in the injection port and is detected as diphenylamine.
Hence, all diphenylamine in the sample must he removed by Florisil to prevent interferences.
2 The extract may he cleaned with Florisil to prevent phthalate interferences.
3 Extracted phenols are derivatized with pentafluorobenzylbromide (PFR) prior to silica gel treatment.
4 Due to the extreme toxicity of TCPID (2,3,7,8-Tetrachlorodibenzo-p-dioxin), the analysis method
should be reviewed thoroughly before it is used. The sample extract should undergo a two-
stage cleanup (silica gel followed by alumina).
5 Prerinse C,^ cartridge with distilled water and elute with methanol.
-------
APPENDIX B
1. Evaluating Chromatographic Cartridges
1.1 If the compound of interest is not located within Appendix A then a
preliminary study must be performed before any analysis can proceed.
1.2 Choice of the most efficient cartridge may be narrowed by exmaining
the following guidelines:
1.2.1 Silica and Florisil Cartridges - The nonpolar compounds elute
before the polar compounds. Extraction should be performed in
the least polar solvent possible.
1.2.2 C18 Cartridges - The polar compounds elute before the nonpolar
compounds. Extraction should be performed in water or the
most polar aqueous/organic solvent mixture.
2. Determining the effect of a cartridge on the sample matrix
2.1 Remove an aliquot of an environmental sample, perform the necessary
extraction/dilution sequences and then submit the extract to the
cleanup procedures (Procedure 4) using the chosen cartridge.
NOTE 1: Not more than 100 ul or 100 mg of sample may be loaded onto
the cartridge.
NOTE 2: When determining the dilution sequence achieved through the
cartridge, be certain the final sample extract concentration
will be within the accepted calibration range limits.
2.2 Remove a duplicate aliquot of the same environmental sample and
perform the necessary extraction/dilution sequences, but do not
submit it to any cleanup. The final concentration should be
calculated to be approximately that of the "cleaned-up" sample.
2.3 Analyze both samples according to the prescribed procedures.
2.4 Compare the results of the "cleaned-up" sample vs. the untreated
sample and determine if the sample matrix removal is sufficient
for the analysis requirements.
2.5 If sample matrix removal was not adequate, repeat the procedure for
determining the effect of a cartridge on the sample'matrix using
another class of cartridge. If the removal was sufficient then
proceed to the following procedure for determining the effect of a
cartridge on the compound of interest.
6.1-5
-------
3. Determining the effect of a cartridge on the compound of interest within
a sample matrix
NOTE: The environmental sample used in the procedure for determining
the effect of a cartridge on the sample matrix should not
contain any significant amount of the compound of interest. If
it does, the amount present must be quantified so that its
value may be subtracted from the spiked amount to avoid a
falsely high relative error.
3.1 Remove an aliquot of the same environmental sample and perform the
necessary extraction/dilution sequences (as previously performed in
the procedure for determining the effect of a cartridge on the
smaple matrix).
3.2 Rinse the chosen cartridge with 2 to 3 ml of the appropriate solvent
using the micropipet.
3.3 Remove any excess solvent from the cartridge by injecting air
through the cartridge with the micropipet.
3.4 Inject a known concentration of a surrogate standard of the compound
of interest on the prerinsed cartridge.
NOTE: When determining the dilution sequence achieved through the
cartirdge, assure that the final surrogate extract
concentration will be within the accepted calibration range
limits.
3.5 Submit the sample extract to the cleanup procedure (refer to Sections
4.3 - 4.8) using the cartridge spiked with the surrogate.
3.6 Repeat the above Procedures, 2.1 through 3.5, twice for a total of
three analyses.
3.7 Using a microprocessor, calculate the relative error of each analysis.
3.8 If the average relative error is less than 10%, proceed with the
sample analysis using the newly selected cartridge. If the relative
error is greater than 10%, the entire procedure (1 through 3) may
be repeated or the efficiency of another cartridge may be examined.
6.1-6
-------
METHOD 6.2 PROTOCOL FOR REMOVING ELEMENTAL SULFUR FROM SEDIMENT EXTRACTS
1. OVERVIEW: This method provides a rapid procedure for removing elemental
sulfur from sediment extracts prior to gas chromatographic
analysis for hydrocarbon and/or PCB content.
2. CAUTIONS AND LIMITATIONS: All sample preparation and procedural steps
employed in this method must be performed inside
a fume hood to minimize exposure of laboratory
personnel to hazardous substances.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Beakers, 100 ml, 50 ml
?.. Clamps (for ring stand)
3. Disposable pasteur pipettes, 9 inch length
4. Glass wool
5. Kuderna Danish concentration vials
6. Microspatula, stainless steel
7. Pipette bulb, large
8. Ring stand
3.2 Reagents
1. Copper metal, electrolytic dust (Fisher C-431)
2. Hydrochloric Acid, I Normal (Fisher SO-A-48)
3. Nitrogen gas, prepurified grade (with in-line carbon filter)
4. PROCEDURE:
4.1 The sample extract containing hydrocarbons and/or PCBs must be
concentrated to approximately 2 ml using purified nitrogen gas.
4.2 Solvent rinse a disposable pasteur pipette with organic solvent
(i.e., same solvent as that employed to extract the sediment).
4.3 Place a glass wool plug in the base of the pipette and secure the
pipette in a clamp mounted to a ring stand.
6.2-1
-------
4.4 Using a microspatula, transfer copper metal into the pipette until
the level of the copper is approximately 3 cm above the glass wool.
4.5 Activate the copper metal by passing 5 ml of I Normal HC1 through the
pipette.
Note: Flow may be accelerated by using a pipette bulb to afford
positive displacement.
4.6 Solvent rinse the activated copper column with two 5 ml volumes of
organic solvent (i.e., same solvent as that employed to extract the
sediment).
Note: DO NOT permit the solvent to percolate below the liquid/copper
interface.
4.7 Place a Kuderna Danish concentration vial beneath the mouth of the
pipette to collect the treated sample in steps 4.8 - 4.10.
4.8 Transfer the 2 ml concentrated sample extract to the pipette and
elute the sample to remove any elemental sulfur contained therein.
Mote: The bright copper turns black as the sulfur is removed.
4.9 Rinse the sample container with organic solvent and transfer this
solvent rinse to the copper column.
4.10 Remove any residual sample by passing 5 ml of organic solvent through
the copper column.
4.11 Concentrate the sample extract in the Kuderna Danish concentration
vial to its initial volume using purified nitrogen gas.
4.12 Perform hydrocarbon and/or PCB analysis in accordance with Method 4.2
or Method 2.1.
5. REFERENCES:
5.1 EPA Quality Assurance Newsletter, Volume 1, No. 2, July 1978.
5.2 EPA-600/2-77-lfi3, Correlation of Oils and Oil Products by Gas
Chromatography.
6.2-2
-------
METHOD 6.3 PROTOCOL FOR WEATHERING SMALL QUANTITIES OF PETROLEUM OIL
1. OVERVIEW; This method provides a rapid procedure for weathering small
quantities of petroleum oil to enable comparison of environ-
mentally weathered oil samples with a neat source oil.
2. CAUTIONS AND LIMITATIONS:
2.1 A minimum of 0.5 mg of sample oil and 70 mg of source oil are needed.
2.2 Although Freon is the recommended solvent for this procedure, carbon
tetrachloride may be substituted for oils which are insoluble in
Freon in the small volumes used for transfer and injection.
2.3 This method simulated only the evaporative losses of weathering.
These losses, however, may be the most significant of the short-term
weathering effects.
2.4 Because this identification procedure is intended for small amounts
of oil, preparation of standards for quantification is not feasible.
This method, therefore, applies only to qualitative spill source
identification.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Air supply with in-line carbon filter
2. Reaker, 150 ml
3. Disposable pasteur pipettes
4. Flexible plastic tubing
5. Gas Chromatograph equipped with flame ionization detector
6. Glass tubes
7. Hot plate
8. Steam bath
9. Syringe, 10 ul
10. Water bath
3.2 Reagents
1. Carbon tetrachloride, pesticide grade
2. Crude oil standard
3. Freon 113, or equivalent
6.3-1
-------
4. PROCEDURE:
4.1 Weathering of Sample Extracts
4.1.1 Rinse all glassware with Freon.
4.1.2 Determine the oil content of the sample extract by IR measurement
(refer to Appendix).
4.1.3 If at least 30 mg of oil are present, dilute the sample extract
to yield 30 mg of oil in 100 ml of Freon. If less than 30 mg
'of oil are present, dilute the sample extract to yield 0.5 mg of
oil in 100 ml of Freon. Transfer the dilution to a 150 ml beaker.
4.1.4 Assemble the filtered air stream (refer to Appendix) consisting
of an air source, an activated carbon filter, and a glass tube,
using flexible tubing as the connective interface between the
individual components. Clamp the glass tube to a ringstand.
4.1.5 Place the 150 ml beaker on a 97-99°C steam bath in a hood.
4.1.6 Position the air stream directly above the Freon solution with
enough pressure to create a small dent at the liquid surface.
4.1.7 Evaporate the solvent until only 1-2 ml of the solution remains;
remove the beaker from the steam bath.
4.1.8 Quantitatively transfer the sample to a 10 x 30 mm vial with 3
or 4 ml of Freon. (Use carbon tetrachloride if the solubility
of the oil in Freon is low.)
A.1.9 Replace the glass tube extending from the activated carbon
filter with a disposable Pasteur pipette.
4.1.10 If a 30 mg quantity of oil was used, place the vial containing
the concentrate on a water bath at 40±l°C. Utilize the drav/n-
out tip of the pipette to direct the air flow onto the surface
of solution; adjust thp air pressure to create a small dent at
the surface. Strip to dryness; maintain this condition for an
additional ten minutes.
4.1.11 If a 0.5 mg quantity of oil was used, leave the vial containing
the concentrate at room temperature. Position the filtered air
stream so as to produce a small dent at the solution surface.
Maintain these conditions until only a small amount of solvent
remains. Stop the air flow; let the remaining solvent evaporate
spontaneously. Keep the vial open an additional ten minutes.
4.1.12 Add to the vial 10-20 ul of the same solvent used in step 1-h
(i.e., either Freon or carbon tetrachloride). Swirl to dissolve.
6.3-2
-------
4.1.13 Inject the weathered oil solution into the GC using the
appropriate column and conditions.*
4.2 Weathering of a Neat Reference Oil
4.2.1 Weigh 40 mg of neat oil into a 10 x 30 mm vial.
4.2.2 Place the vial into a water bath at 40±1°C.
4.2.3 Direct the filtered air stream onto the solution surface to
create a small dent.
4.2.4 Maintain these conditions for fifteen minutes.
4.2.5 Remove the vial from the steam bath.
4.2.6 Inject 1/2 - 1 ul of the weathered oil into the GC using the
appropriate column and conditions (refer to Reference).
5. CALCULATIONS; Perform all calculations in accordance with the Quality
Assurance/Quality Control Protocol and the appropriate
Microprocessor Program (refer to Section VIII of the
Methods Manual).
fi. REFERENCE; l ASTM, Part 31, Water D-3328 Method A.
6.3-3
-------
METHOD 6.4 PROTOCOL FOR PURIFYING ISOPROPANOL FOR FLUORESCENCE ANALYSES
1. OVERVIEW: This method provides a procedure for reducing the fluorescence
emission of isopropanol by filtering the solvent through granular
activated carbon to remove fluorescent impurities.
2. CAUTIONS AND LIMITATIONS:
2.1 Before using this procedure, determine whether fluorescing impurities
in the isopropanol interfere at the desired sample sensitivity. It
may not be necessary to perform this purification if the response
at this sample sensitivity does not cause substantial emission
interference with the compound of interest.
2.2 After performing this purification procedure, compare the fluorescence
emission of both the filtered and unfiltered isopropanol to ensure
that no fluorescing substances are leached into the isopropanol from
the activated carbon.
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Beaker, 500 ml
2. Chromatographic column, Kontes, 50 cm x 4.5 cm I.D., equipped with
stopcock
3. Clamp (for ring stand)
4. Fluorescence spectrophotometer equipped with dual monochromators
for synchronous measurement
5. Glass wool
6. Granular activated carbon, 8-12 mesh
7. Powder funnel
8. Ring stand
9. Stop watch
10. Volumetric flask, 1 L
3.2 Reagents
1. Isopropanol, spectrophotometric grade (UV cutoff 205 nm)
6.4-1
-------
4. PROCEDURE:
4.1 Pack 2 cm of glass wool Into a Kontes stopcocked chromatographic column
and attach the column to a ring stand.
4.2 Using a powder funnel, add approximately 20 cm of granular activated
carbon to the chromatographic column.
4.3 Close the stopcock and fill the column with isopropanol (let the filled
column stand for 5 minutes).
4.4 Place a 500 ml beaker beneath the column, open the stopcock, and
establish a flow rate of 15-20 ml/min.
4.5 Let the column drain and discard the filtered isopropanol.
4.6 Close the stopcock and refill the chromatographic column with
isopropanol.
4.7 Place a 1 liter volumetric flask beneath the column, open the stopcock,
and establish a flow rate of 15-20 ml/min.
4.8 Close the stopcock after approximately 40 ml of isopropanol have
drained into the volumetric flask.
4.9 Rinse the volumetric flask with the 40 ml of filtered isopropanol and
discard this solvent rinse.
4.10 Repeat steps 7-9 and then refill the column with isopropanol.
4.11 Slowly open the stopcock and establish a flow rate of 6-10 ml/min.
Note: DO NOT permit the isopropanol to percolate below the liquid/
carbon interface.
4.12 For short term storage of the activated carbon column, ensure that the
level of isopropanol is just above that of the activated carbon;
fit aluminum foil over the top of the column.
4.13 Check the fluorescence emission of the filtered isopropanol to ensure
that the activated carbon removed the fluorescing impurities and
the isopropanol did not leach additional fluorescing substances from
the carbon.
6.4-2
-------
SECTION VII
MISCELLANEOUS PROTOCOLS AND PROCEOIIRES
-------
CONTENTS
METHOD 7.1 Protocol for the Qualitative Evaluation of
Chromatograms of Polychlorinateci Biphenols
METHOD 7.2 Instructions for Packing and Conditioning Metal
Analytical Columns for Gas Chromatographs
METHOD 7.3 Instructions for Packing and Conditioning Glass
Analytical Columns for Gas Chromatographs
METHOD 7.4 Protocol for Checking the Performance of Glass
Analytical Columns for Gas Chromatographs
-------
METHOD 7.1 PROTOCOL FOR THE QUALITATIVE EVALUATION OF GAS
CHROMATOGRAMS OF POLYCHLORIMATED BIPHENYLS
1. SCOPE AND SUMMARY:
1.1 This Method addresses the identification of polychlon'nated biphenyl
(PCB) compounds in sample extracts. Specifically, the following PCB
Aroclors may be identified through application of this protocol:
Aroclor 1016; Aroclor 1221; Aroclor 1232; Aroclor 1242; Aroclor 1248;
Aroclor 1254; and, Aroclor 1260.
1.2 This Method provides a procedure for analyzing gas chromatograms of
PCBs to ensure proper identification and quantification of Aroclors
present either individually or as mixtures in environmental samples.
2. CAUTIONS AND LIMITATIONS:
2.1 The preparation of standards and samples should be performed inside a
properly ventilated fume hood with the appropriate protective apparel
(e.g., safety glasses, laboratory coat, chemical resistant gloves) to
minimize exposure of laboratory personnel to hazardous and/or carcino-
genic substances.
2.2 Accurate interpretation of PCB chromatograms necessitates the use of
uniform instrumental conditions for the analysis and comparison of
the sample chromatograms with those of the Aroclor standards.
2.3 The gas chromatographic conditions listed in Appendix B of this
Method must be established at least two hours prior to analysis of
the calibration standards and sample extracts to permit sufficient
stabilization of the electron capture detector.
2.4 Prior to gas chromatographic analysis of the sample(s), a p.p'-DDE
standard must be analyzed to establish its absolute retention time
which is employed to calculate the relative retention times (RRT^s)
of the sample peaks.
2.5 Due to the complexity of PCB Aroclors, the procedure outlined in
this Method must be followed in its entirety to ensure accurate
identification of the individual PCB Aroclor(s) present in the
sample(s).
2.6 This Method provides a comprehensive protocol for the qualitative
and quantitative gas chromatographic analysis of PCB compounds.
The analyst should refer to Method 1.4 and/or 2.1 for specific
extraction procedures employed for PCB contaminated samples.
7.1-1
-------
3. APPARATUS AND REAGENTS:
3.1 Apparatus
1. Chromatographlc column, 6 ft (1/4 In O.D. x 4 mm I.D.), packed
with 1.5 % SP-2250 and 1.95* SP-2401
2. Gas chromatograph equipped with an electron capture detector
3.2 Reagents
1. PCB Aroclor Standards (Supelco, Inc. or equivalent)
2. p,p'-DDE standard, 0.05 mg/1
4. PROCEDURE:
4.1 Gas Chromatographlc Fingerprint Analysis
4.1.1 Reference standards of individual PCB Aroclors (refer to Appendix
A of this Method for a list of PCB Aroclors) should be prepared
in accordance with Method 5.1 (Protocol for Preparing Analytical
Standards of Hazardous and Carcinogenic Materials). The analyst
may also consult the procedures for standard preparation outlined
in Method 1.4 (Rapid Extraction and Chromatographic Analysis of
Polychlorinated Biphenyls in Aqueous Samples) and/or Method 2.1
(Rapid Extraction and Chromatographic Analysis of Polychlorinated
Biphenyls in Sediment Samples).
4.1.2 Establish the gas Chromatographic conditions listed in Appendix B
of this Method.
NOTE: The electron capture detector requires a minimum two hour
stabilization period prior to analysis of the calibration
standards and sample extracts.
4.1.3 Using an autoinjector or precision quality 10 ul syringe, inject
several microliters of the PCB Aroclor standards into the gas
chromatograph to obtain reference chromatograms. The p,p'-DDE
standard should be analyzed in the same manner to establish its
absolute retention time under these instrumental conditions.
4.1.4 Representative chromatograms of the sample(s) should be generated
by analyzing the sample extract(s) according to step 4.1.3.
4.1.5 Calculate the absolute retention time (ART) of each peak in the
sample chromatogram and its retention time (RRTj) relative to the
absolute retention time of the p,p'-DDE standard. The RRTj of
each sample peak is therefore its elution time relative to that
of the p,p'-DDE standard and is necessarily a unitless parameter.
7.1-2
-------
SAMPLE CALCULATION OF RELATIVE RETENTION TIMES (RRTi):
ART of p.p'-DDE = 31.62 min
ART of Sample Peak = 10.20 min
of p.p'-DDE = 100
of Sample Peak = ART of Sample Peak x RRTi of p.p'-DDE
ART of p.p'-DDE
of Sample Peak = 10.20 min x 100 = 32
31.62 min
4.2 PCB Aroclor Identification
4.2.1 Match the elution or fingerprint pattern of the sample extract
with the reference chromatogram of the suspect PCB Aroclor.
The analyst should refer to Appendix C of this Method for an
illustrative example of determining the elution pattern for
PCB Aroclor 1242.
NOTE: Due to interferences present in the sample matrix, the
sample elution pattern may differ substantially from that
of the PCB Aroclor standard. Therefore, the RRTs should
be used as the major criterion for identification of the
PCB Aroclors in the extract(s).
4.2.2 Match the RRT}S of the sample peaks with those in the suspect
PCB Aroclor standard chromatogram (refer to Appendic C of this
Method). The analyst should note that the calculated RRT^s may
differ within 1-2 RRTi units of the values determined for that
PCB Aroclor standard.
4.2.4 For further verification of the specific PCB Aroclor(s) present
in the sample extract (s), calculate sample peak retention times
(RRT2) relative to the major PCB peaks in the chromatogram (refer
to Appendix C of this Method) and compare these RRT2 values to
those of the PCB Aroclor standard.
SAMPLE CALCULATION OF RELATIVE RETENTION TIMES (RRT2):
RTT2 of sample peak = ART of sample peak x RRT2 of major peak
ART of major peak
RTT2 of sample peak = 12.25 min x 1.00 = 1.20
10.20 min
NOTE; If multiple PCB Aroclors are present in a sample, the
RRT2s must be calculated separately for each PCB Aroclor.
7.1-3
-------
4.3 Quantification of PCB Aroclors
4.3.1 Prepare a PCB calibration solution and calibration standards
by diluting the suspect PCB Aroclor stock standard solution
appropriately. The calibration range should be established so
as to incorporate the expected concentrations of the samples
(refer to Method 8.1 - Protocol for Establishing a Calibration
Range for Single Point Analysis).
4.3.2 Inject several microliters of the PCB calibration solution which
has been identified in the sample extract(s) and compare the
resultant chromatogram with that of the sample chromatogram(s).
4.3.3 Positively identify the PCB Aroclor(s) present in the sample
based upon its gas chromatographic fingerprint, and calculated
and RRT2S for the individual peaks.
4.3.4 Analyze the calibration range of the PCB Aroclor(s) identified
in the sample(s) employing the gas chromatographic conditions
tabulated in Appendix B of this Method.
NOTE; Pending positive identification of the PCB Aroclor(s)
in the sample extract(s), the instrument conditions
may be modified to accelerate the analysis (e.g., the
oven temperature may be increased to 210°C).
4.3.5 The chromatograms of the calibration standards and samples
should be quantitated based on peak heights (as opposed to area
count responses) by employing the appropriate data sheets and
microprocessor programs listed in Method 8.2.
NOTE 1: Baselines must be drawn in. the same manner on all of
the sample and standard chromatograms.
NOTE 2: If multiple PCB Aroclors are present in a sample, the
individual Aroclors must be identified and the concen-
trations of each calculated separately.
7.1-4
-------
APPENDIX A
PCB AROCLOR STANDARDS
Individual PCB Aroclors are a mixture of numerous components and considerable
overlap in composition occurs among Aroclor Standards. Quantitative standards
for analyzing aqueous samples for PCB content are available from commercial
vendors. Supelco, Inc. provides individual ampuls of the following Aroclors
at the specified PCB concentrations in methanol:
PCB Aroclor Concentration
Aroclor 1016 200 ug/ml
Aroclor 1221 200 ug/ml
Aroclor 1232 200 ug/ml
Aroclor 1242 200 ug/ml
Aroclor. 1248 200 ug/ml
Aroclor 1254 200 ug/ml
Aroclor 1260 200 ug/ml
(Source: Supelco, Inc., Catalog 25, 1987)
7.1-5
-------
APPENDIX B
Analytical Column: *
Carrier Gas:
Carrier Flow Rate:
Detector:
Detector Temperature:
Injector Temperature:
Oven Temperature:
Program Rate:
Time of Analysis:
GAS CHROMATOGRAPHIC CONDITIONS
1.5% SP 2250/1.95* SP 2401 on 100/120 Supelcoport
6 ft x 4 mm glass column (mixed phase support)
Argon:Methane (95%:5%)
40 ml/min
Electron Capture (ECD)
260°C
225°C
180°C (Dependent on PCB Aroclor)
Isothermal
Variable (Dependent on PCB Aroclor)
* The mixed phase column recommended for PCB analyses should be packed and
conditioned in accordance with Method 7.3 (Instructions for Packing and
Conditioning Glass Analytical Columns for Gas Chromatography Instruments)
and performance tested according to the procedure and specifications
described in Appendix C of Method 7.4 (Protocol for Performance Testing
Packed, Conditioned, Glass Analytical Columns for Gas Chromatography
Instruments).
7.1-6
-------
APPENDIX C
CHROMATOGRAN OF PCB AROCLOR 1242
CHROMATOGRAM OF p.p'-DDE STANDARD
29.34
3.72
9.42
18.28
29.20
29.27
31.62
TABULATION OF ABSOLUTE AND RELATIVE RETENTION TIMES FOR PCB AROCLOR 1242
ABSOUUTB RETENTION
TIME OP SAMPLE PEAK
4.29 min
4.77 Bin
5.55 min
6.76 min
7.50 min
8.99 min
9.58 min
10.20 min
11.30 min
12.25 min
13.42 Bin
16.22 min
17. SO min
19.71 min
21.13 min
25.34 min
RELATIVE RETENTION TIKE *
USING p.p'-ODE STANDARD
14
15
18
21
24
28
30
32
36
39
42
51
55
62
67
80
RELATIVE RETENTION TIKE t
USING MAJOR PCB PEAK
0.42
0.47
0.54
0.66
0.74
0.88
0.94
1.00
1.11
1.20
1.32
1.59
1.72
1.93
2.07
2.48
• The Relative Retention Tine (RRTi) is calculated from the ratio of the absolute
retention times of the sample peak and the p.p'-ODE standard, multiplied by 100
t The Relative Retention Time (RRty is calculated from the ratio of the absolute
retention times oE the sample peak and the major PCB peak, multiplied by 1.00
7.1-7
-------
METHOD 7.2 INSTRUCTIONS FOR PACKING AND CONDITIONING METAL
ANALYTICAL COLUMNS FOR GAS CHROMATOfiRAPHS
1. OVERVIEW: This method provides a procedure for packing and conditioning
metal analytical columns for gas chromatography instruments.
2. APPARATUS:
1. Clamps
2. Column coiler (or cylindrical object, 6 inch diameter)
3. Column packing material (10 OV-101 on Chromosorb W for analysis
of petroleum oils)
4. Column tubing, stainless steel, 1/8 inch OD
5. Electric vibrator
6. Ferrules, front and back, stainless steel, 1/8 inch ID
7. Gauze
8. Glass wool, silanized
Q. Metal funnel, 1/8 inch threaded tubing connector stem
10. Ring stand, 5 foot heiqht
11. Scissors
I?.. Slide tape ruler, in ft.
13. Swagelok tube nuts, stainless steel 1/8 inch ID
14. Tee connector, stainless steel, 1/8 inch in
15. Thin wire, 2 inch length
16. Tube cutter
17. Tubing reamer
18. Tygon tubing, 1/4 inch ID
19. Vacuum pump
3. PROCEDURE:
3.1 Packing Column
3.1.1 Measure 10 feet of 1/8 inch OD stainless steel tubing to be
used as the GC column.
3.1.2 Cut this measured length of tubing with a tube cutter and widen
the rough openings on both column ends with a tubing reamer.
3.1.3 Insert sufficient silanized glass wool (approximately 1/2
inch square) into one end of the column.
3.1.4 Secure the column (glass wool plug on the bottom end) in a
vertical position by clamping it to a 5 foot ring stand mounted
on a bench top.
7.2-1
-------
3.1.5 Place a 1/8 Inch ID stainless steel Swagelok tube nut and front
and back stainless ferrules on the top end of the column;
hand tighten a metal funnel (equipped with threaded 1/8 inch
tubing connector stem) to the top end of the column.
3.1.6 Place a piece of gauze over the bottom end of the column.
3.1.7 Connect the gauze covered bottom end of the column to a vacuum
pump using a suitable length of 1/4 inch in tygon tubing.
3.1.8 Turn on the vacuum pump and slowly pour the packing material
(10 OV-101 on Chromosorb W) into the funnel. Vibrate the
column from top to bottom using an electric vibrator to assure
a reasonably tight packing. Repeat this procedure (pouring,
then vibrating) until the column is filled.
3.1.9 Turn off the pump and disconnect the vacuum line and metal funnel
3.1.10 Tilt the column and empty out enough packing material from
the top to allow for the insertion of silanized glass wool.
3.1.11 Using a thin wire, insert a glass wool plug (approximately
I/?, inch square) into the injector end of the column until
the glass wool is in contact with the packing material.
3.1.12 Attach a 1/8 inch ID Swagelok tube nut and front and back
stainless steel ferrules (with taper toward column end) to
each end of the column.
3.1.13 Using a stainless steel tee connector, tighten the Swagelok
tube nut to the connector until the ferrules are locked into
place on both ends of the column; remove the tee connector.
3.1.14 Employing a column coiler or suitable substitute, coil the
column to a six inch diameter.
3.2 Conditioning Column
3.2.1 Column conditioning should be performed by placing the column
into the appropriate GC and connecting the injector end of
the column to the instrument injection port.
Note: HO NOT connect the other end of the column to the
instrument detector during column conditioning; irrepar-
able damage to the detector may otherwise occur.
3.2.2 Condition the column isothermally by setting the GC oven
temperature 25-50°C below the upper temperature limit of the
packing material.
Note: The conditioning temperature for 10 OV-101 on Chromosorb
W packing material is 325°C; the upper temperature
limit is 350°C.
7.2-2
-------
3.2.3 Set the carrier gas flow to 30 ml/min during conditioning,
and condition the column for 1-3 days.
3.2.4 The conditioned column is connected to the instrument detector
for performance evaluation.
3.3 Column Performance Evaluation
3.3.1 Refer to the procedures of Method 7.4 to assure proper perform-
ance of the conditioned column based on theoretical plate and
resolution criteria.
3.3.2 Once it is established that the column meets the minimum
performance criteria, the column must be entered into the GC
column inventory system.
7.2-3
-------
METHOD 7.3 INSTRUCTIONS FOR PACKING AND CONDITIONING GLASS
ANALYTICAL COLUMNS FOR GAS CHROMATOGRAPHS
1. OVERVIEW; This method provides a procedure for packing and conditioning
glass analytical columns for gas chromatography instruments.
2. APPARATUS:
1. Chromatographic column, glass, 1/4 inch OD stem
2. Clamps (for ring stand)
3. Column packing material (1% SP-1240 DA for phenols; 1.5% SP-2250/1.95%
SP-2401 for base neutrals, pesticides, and PCBs)
4. Ferrules, Supelco (Supeltex M-2) Vespels, 1/4 inch ID
5. Funnel, metal or glass, 1/4 inch OD stem
6. Gauze
7. Glasrench, Supelco (Model B)
8. Glass wool (silanized for general use and for the anlaysis of neutral
and basic compounds; phosphoric acid treated for the analysis of acidic
compounds such as EPA Consent Decree phenolic compounds and for columns
to be filled with 1% SP-1240 DA packing material)
9. 0-rings
10. Ring stand, 3 foot height
11. Scissors
12. Swagelok tuhe nuts, 1/4 inch in
13. Thin wire, 6 inch length
14. Tygon tubing, 1/4 inch ID
15. Vacuum pump
3. PROCEDURE:
3.1 Packing Column
3.1.1 Determine the appropriate span and glass column needed.
1. For a Hewlett-Packard GC Model 5830 or 5840, an "ON
COLUMN" injection type glass column, of six or nine inch
span (i.e., six inch for ECD, nine inch for FID and NPD)
is required (refer to figure in Appendix).
2. For a Perkin-Elmer GC Model 900, an "OFF COLUMN" injection
type glass column with a nine inch span is required (refer
to figure in Appendix).
7.3-1
-------
3.1.2 Determine the correct length of glass column needed.
1. For packing with 1% SP-1240 DA material, use a three foot
(1/4 in OD x 2 mm ID) column.
2. For packing with 1.5% SP-2250/1.95% SP-2401 material,
use a six foot (1/4 in OD x 4 mm ID) column.
3.1.3 Insert sufficient pre-treated glass wool (approximately 1/2
inch square) to extend 1/2 to 1 inch into the detector end of
the empty glass column.
Note 1;'Use phosphoric acid treated glass wool for columns packed
with 1% SP-1240 DA, and silanized glass wool for
columns packed with 1.5* SP-2250/1.95% SP-2401.'
Note 2: The detector end is 4 1/2 inches below the injector end
for "ON COLUMN" injection type columns; the injection
ends are flush with one another for "OFF COLUMN"
injection type columns (refer to figure in Appendix).
3.1.4 Place a piece of gauze over the detector end of the column.
3.1.5 Using a suitable length of 1/4 inch ID tygon tubing, slip one
end over the gauze covered column end, and connect the other
end to a vacuum pump.
3.1.6 Using a 1 inch length of 1/4 inch ID tygon tubing, connect
the injector end of the column to a glass or metal funnel.
3.1.7 Secure the column by clamping it to a ring stand, turn on the
vacuum pump, and slowly pour the packing material into the
funnel. Tap the column lightly as it is being filled to
assure a reasonably tight packing. Repeat this procedure
(pouring, then tapping) until the column is filled.
Note 1: For "ON COLUMN" injection type columns, the packing
material is filled to approximately 4 1/2 inches
below the injector end of the column. The 4 1/2
inches of empty space is the injection port for this
type of column.
Note 2: For "OFF COLUMN" injection type columns, the column
is filled until the level of packing material in the
injector end is even with the level of packing material
in the detector end.
3.1.8 Using a thin wire, insert a glass wool plug (approximately
1/2 inch square) into the injector end of the column until
the glass wool is in contact with the packing material.
7.3-2
-------
3.1.9 Slide an 0-ring and a 1/4 inch ID Swagelok tube nut onto the
injector end of the column; install the ferrule with taper
toward the column end.
3.2 Conditioning Column
3.2.1 Column conditioning should be performed by placing the column
into the appropriate GC and connecting the injector end of
the column to the instrument injection port using a glasrench.
Note: DO NOT connect the other end of the column to the
instrument detector during column conditioning;
irreparable damage to the detector may occur.
3.2.2 Condition the column isothermally by setting the GC oven
temperature 25-50°C below the upper temperature limit of the
packing material (refer to table in Appendix).
3.2.3 Set the carrier gas flow to 30 ml/min during conditioning,
and condition the column for 1-3 days.
3.2.4 After the column is conditioned, slide an 0-ring and a 1/4
inch ID Swagelok tube nut onto the detector end of the column;
install the ferrule with taper toward the column end.
3.2.5 Connect the conditioned column to the instrument detector
(using a glasrench) for performance evaluation.
3.3 Column Performance Evlauation
3.2.1 Refer to the procedures of Method 7.4 to assure proper perform-
ance of the conditioned column based on theoretical plate and
resolution criteria.
3.2.2 Once it is established that the column meets the minimum
performance criteria, the column must be entered into the GC
column inventory system.
7.3-3
-------
APPENDIX
FIGURES ILLUSTRATING TWO DIFFERENT TYPES OF COLUMN CONFIGURATIONS
"ON COLUMN"
Injection Type Glass Column
(Hewlett-Packard, Models 5830/5840)
"OFF COLUMN"
Injection Type Glass Column
(Perkin-Elmer, Model 900)
TABLE OF TEMPERATURE LIMITS FOR SELECTED COLUMN PACKING MATERIALS
COLUMN PACKING MATERIAL MAXIMUM TEMP (°C) CONDITIONING TEMP (°C)
1% SP-1240 DA
1.5% SP-2250/1.95% SP-2401
200
260
175
230
7.3-4
-------
METHOD 7-4: PROTOCOL FOR CHECKING THE PERFORMANCE OF GLASS
ANALYTICAL COLUMNS FOR GAS CHROMATOGRAPHS
1. OVERVIEW; This method provides procedures for checking the performance
of packed, conditioned, glass analytical columns for gas
chromatography instruments. The column performance evaluations
are based on theoretical plate (N) and resolution (R) criteria.
2. APPARATUS AND REAGENTS:
2.1 Apparatus
1. Gas chromatograph equipped with electron capture and flame
iom'zation detectors
2. Glass syringes, 10 ul
3. Packed, conditioned, glass GC columns:
a. Mixed Phase Columns - packed with 1.5% SP-2250/1.95% 2401 on
100/120 Supelcoport; 6 ft (1/4 in OD x 4 mm ID); 6 inch span for
ECD, 9 inch span for FID; conditioned according to Method 7.3.
b. Phenolic Columns - packed with 1% 1240 DA on 100/120 Supelcoport;
3 ft (1/4 in OD x 2 mm ID); 6 inch span for ECD, 9 inch span for
FID; conditioned according to Method 7.3.
c. Purgeable Columns - packed with 1% SP 1000 on Carbowax; 6 inch
span for ECD, 9 inch span for FID; fi ft (1/4 in On x 2 mm ID);
conditioned according to Method 7.3.
2.2 Reagents: Refer to the appropriate Appendix for specific reagents.
3. PROCEDURE:
3.1 Refer to Appendix A: Procedure For Performance Checking Mixed Phase
Columns For GC-ECD Analyses.
3.2 Refer to Appendix B: Procedure For Performance Checking Mixed Phase
Columns For GC-FID"Analyses.
3.3 Refer to Appendix C: Procedure For Performance Checking Phenolic
Columns For GC-FID Analyses.
3.4 Refer to Appendix D: Procedure For Performance Checking Purgeable
Columns For GC-FID Analyses.
7.4-1
-------
4. CALCULATIONS:
4.1 The number of Theoretical Plates (N) is calculated using the
following relation:
N = (5.545H(TrHC)/W1/2)]2
where; Tr = Retention time (minutes)
C = Chart speed (mm/min)
h/1/2 = Width of peak at half height (mm)
Note; The numerical constant 5.545 is required when measuring the
width of peaks at half height rather than at the baseline.
4.2 The Resolution (R) between two peaks is calculated using the
following relation:
R =
W1/2)
where; T = Difference in Retention Times for peaks 1 and 2
C = Chart Speed (mm/mi n)
Wlj/2 - Width of peak 1 at half height (mm)
= Width of peak 2 at half height (mm)
Note: The numerical constant 1.177 is required when measuring the
width of peaks at half height rather than at the baseline.
7.4-2
-------
APPENDIX A
PROCEDURE FOR PERFORMANCE CHECKING A MIXED PHASE COLUMN FOR GC-ECD ANALYSES
1. OBJECTIVE: This method provides a procedure for checking the performance of
a packed, conditioned, mixed phase column, on a gas chromatograph
equipped with an electron capture detector. The column performance
evaluation is based on theoretical plate (N) and resolution (R)
criteria.
2. APPARATUS AND REAGENTS:
2.1 Apparatus
1. Glass column; 6 ft (1/4 in OD X 4 mm in), 6 inch span; packed
with 1.5% SP-2250/1.95% SP-2401 on 100/120 Supelcoport and
conditioned according to Method 7.3.
2. Gas chromatograph equipped with an electron capture detector
3. Syringe, 10 ul
2.2 Reagents
1. Argon/Methane gas (95%/5%)
2. Hexane, spectrophotometric grade
3. Pure standards of p.p'-DDE, Dieldrin and p,p'-DDT
3. PROCEDURE:
3.1 Prepare a solution of pesticides containing approximately 550 ppb
p,p'-nnE, 600 ppb Dieldrin, and 750 ppb p,p'-DDT in Hexane.
3.2 Set gas chromatographic conditions as follows:
TEMPERATURE 1 200°C
TIME 1 25.0 min
INJECTOR TEMP ?15°C
DETECTOR TEMP 150°C
CHART SPEED 1.00 cm/min
ZERO 10.0%
ATTENUATION 2 8
SLOPE SENSITIVITY 0.50
AREA REJECTION 1
ARGON/METHANE FLOW 40.0 ml/min
7.4-3
-------
4. CALCULATIONS:
4.1 Calculate theoretical plates (N) using the p,p'-DDT peak and the
following relation:
N = (5.545)-[(Tr)-(C)/(W1/2)]2
where; Tr = Retention time (minutes)
C = Chart speed (mm/mi n)
Wi/2 = Width of p.p'-DDT peak at half height (mm)
Note 1: Peak height of p.p'-DDT should be 50-60% of full scale
with a retention time of 16-20 minutes.
Note 2; Theoretical plates should have a value of at least 3000.
Note 3: The numerical constant 5.545 is required when measuring the
width of peaks at half height rather than at the baseline.
4.2 Calculate resolution (R) using the p.p'-OO-E and Dieldrin peaks and
the following relation:
R = (1.177)'( T)'(C)/(Wl1/? + W21/2)
where; T = Difference in retention times for peaks 1 and 2
C = Chart speed ( mm/mi n)
Wli/2 = Width of p.p'-ODE peak at half height (mm)
W2l/2 = wic|th of Dieldrin peak at half height (mm)
Note 1: Resolution should be close to 1.0 or greater.
Note 2: The numerical constant 1.177 is required when measuring the
width of peaks at half height rather than at the baseline.
7.4-4
-------
APPENDIX B
PROCEDURE FOR PERFORMANCE CHECKING A MIXED PHASE COLUMN FOR GC-FID ANALYSES
1. OBJECTIVE: This method provides a procedure for checking the performance of
a packed, conditioned, mixed-phase column, on a gas chromatograph
equipped with a flame ionization detector. The column performance
evaluation is based on theoretical plate (N) and resolution (R)
criteria.
2. APPARATUS AND REAGENTS:
2.1 Apparatus
1. Glass column; 6 ft (1/4 in OD X 4 mm ID), 9 inch span; packed
with 1.5% SP-2250/1.95% SP-2401 on 100/120 Supelcoport and
conditioned according to Method 7.3.
2. Gas chromatograph equipped with a flame ionization detector
3. Syringe, 10 ul
2.2 Reagents
1. High purity Helium gas
2. Methanol, spectrophotometric grade
3. Pure standards of N-Butylphthalate, Dimethylphthalate and
Acenaphthene
3. PROCEDURE:
3.1 Prepare a solution containing approximately 500 ppm N-Butlyphthalate,
160 ppm Dimethylphthalate and 60 ppm Acenaphthene in Methanol.
3.2 Set gas chromatographic conditions as follows:
TEMPERATURE 1 180°C
TIME 1 18.0 min
INJECTOR TEMP 225°C
DETECTOR TEMP 220°C
CHART SPEED 1.00 cm/min
ZERO 10.0%
ATTENUATION 2 6
SLOPE SENSITIVITY 0.50
AREA REJECTION 1000
HELIUM GAS FLOW 40.0 ml/min
7.4-5
-------
4. CALCULATIONS:
4.1 Calculate theoretical plates (N) using the N-Butylphthalate peak
and the following relation:
N = (5.545H(Tr)-(C)/(W1/2)]2
where; Tr = Retention time (minutes)
C = Chart speed (mm/min)
Wj/2 = Width of N-Butylphthalate peak at half height (mm)
Note 1: The peak height of N-Butylphthalate should be at least
50-60% of full scale with a retention time of 10-12 minutes.
Note 2: Theoretical plates should have a value of at least 2500.
Note 3; The numerical constant 5.545 is required when measuring the
width of peaks at half height rather than at the baseline.
4.2 Calculate resolution (R) using the Acenaphthene and Dimethylphthalate
peaks and the following relation:
R = (1.177H T)-(C)/(Wl1/2 + W21/2)
where; T = Difference in retention times for peaks 1 and 2
C = Chart speed (mm/min)
Wlj/2 = Width of Acenaphthene peak at half height (mm)
W2l/2 = W1dtn of Dimethylphthalate peak at half height (mm)
Note 1; Resolution should be close to 1.0 or greater.
Note 2: The numerical constant 1.177 is required when measuring the
width of peaks at half height rather than at the baseline.
7.4-6
-------
APPENDIX C
PROCEDURE FOR PERFORMANCE CHECKING A PHENOLIC COLUMN FOR GC-FID ANALYSES
1. OBJECTIVE:
This method provides a procedure for checking the performance of
a packed, conditioned, phenolic column, on a gas chromatograph
equipped with a flame ionization detector. The column performance
evaluation is based on theoretical plate (N) and resolution (R)
criteria.
2. APPARATUS AND REAGENTS:
2.1 Apparatus
1. Glass column; 3 ft (1/4 in OD X 2 mm ID), 9 inch span; packed
with 1.0* SP-1240 DA on 100/120 Supelcoport and conditioned
according to Method 7.3.
?.. Gas chromatograph equipped with a flame ionization detector
3. Syringe, 10 ul
2.2 Reagents
1. High purity Helium gas
2. Methanol, spectrophotometric grade
3. Pure standards of 2,4-Dimethylphenol, 2,4-Dichlorophenol and
2,4,6-Trichlorophenol
3. PROCEDURE:
3.1 Prepare a solution containing approximately 225 ppm 2,4-Dimet.hylphenol ,
420 ppm 2,4-Dichlorophenol, and 180 ppm 2,4,6-Trichlorophenol in
Methanol
3.2 Set gas chromatographic conditions as follows:
TEMPERATURE 1
TIME 1
INJECTOR TEMP
DETECTOR TEMP
CHART SPEF.n
ZERO
ATTENUATION
SLOPE SENSITIVITY
AREA REJECTION
HELIUM GAS FLOW
100°C
15.0 min
220°C
260°C
1.00 cm/min
10.0%
2 7
0.50
1000
30.0 ml/min
7.4-7
-------
4. CALCULATIONS:
4.1 Calculate theoretical plates (N) using the 2,4,6-Trichlorophenol
peak and the following relation:
N = (5.545)'[(Tr)-(C)/(W1/2)]2
where; Tr = Retention time (minutes)
C = Chart speed ( mm/mi n)
wl/2 = Width of 2,4,6-Trichlorophenol peak at half height (mm)
Note 1; The peak height of 2,4,6-Trichlorophenol should be at least
50-60% of full scale with a retention time of 9-11 minutes.
Note 2: Theoretical plates should have a value of at least 2000.
Note 3: The numerical constant 5.545 is required when measuring the
width of peaks at half height rather than at the baseline.
4.2 Calculate resolution (R) using the 2, 4-nimethyl phenol and 2,4-Dichloro-
phenol peaks and the following relation:
R = (1.177H T)'(C)/(Wl1/2 + W21/2)
where; T = Difference in retention time for peaks 1 and 2
C = Chart speed (mm/mi n)
= Width °f 2, 4-Dimethyl phenol peak at half height (mm)
= Width of 2,4-Dichlorophenol peak at half height (mm)
Note 1: Resolution should be close to 1.0 or greater.
Note 2: The numerical constant 1.177 is required when measuring the
width of peaks at half height rather than at the baseline.
7.4-8
-------
APPENDIX D
PROCEOURE FOR PERFORMANCE CHECKING A PURGEABLE COLUMN FOR GC-FID ANALYSES
1. OBJECTIVE; This method provides a procedure for checking the performance of
a packed, conditioned, purgeable column, on a gas chromatograph
equipped with a flame ionization detector. The column performance
evaluation is based on theoretical plate (N) and resolution (R)
criteria.
2. APPARATUS AND REAGENTS:
2.1 Apparatus
1. Glass column; 6 ft (1/4 in OD X 2 mm ID), 9 inch span; packed
with 1% SP-1000 on Carbopack B and conditioned according to
Method 7.3.
2. Gas chromatograph equipped with a flame ionization detector
3. Syringe, 10 ul
2.2 Reagents
1. Methanol
2. Pure standards of Trichloroethylene, Benzene, and Toluene
3. Ultra-high purity Helium
3. PROCEDURE:
3.1 Prepare a solution containing approximately 525 ppm Trichloroethylene,
160 ppm Renzene, and 490 ppm Toluene in Methanol.
3.2 Set gas chromatographic conditions as follows:
TEMPERATURE 1 140°C
TIME 1 30.0 min
INJECTOR TEMP 200°C
DETECTOR TEMP 220°C
CHART SPEED 1.0 cm/min
ZERO 10.0%
ATTENUATION 2 8
SLOPE SENSITIVITY 0.5
AREA REJECTION 100
HELIUM GAS FLOW 30.0 ml/min
7.4-9
-------
4. CALCULATIONS:
4.1 Calculate theoretical plates (N) using the Toluene peak and the
following relation:
N = (5.545)-[(Tr)-(C)/(W1/2)]2
where; Tr = retention time in minutes
C = Chart speed (mm/min)
Wj/2 = Width of Toluene peak at half height (mm)
Note 1: The peak height of Toluene should be at least 50-60% of
full scale with a retention time of 15-17 minutes.
Note 2: Theoretical plates should have a value of at least 3000.
Note 3: The numerical constant 5.545 is required when measuring the
width of peaks at half height rather than at the baseline.
4.? Calculate resolution (R) using the Trichloroethylene and Benzene
peaks and the following relation:
R = (1.177)-( T)-(C)/(Wl1/2 + W21/2)
where; T = Difference in retention times for peaks 1 and ?.
C = Chart speed (mm/mi n)
= Width of Trichloroethylene peak at half height (mm)
= Width of Benzene peak at half height (mm)
Note 1: Resolution should be close to 1.0 or greater.
Note 2: The numerical constant 1.177 is required when measuring the
width of peaks at half height rather than at the baseline.
7.4-10
-------
SECTION VIII
DUALITY ASSURANCE/QUALITY CONTROL PROTOCOLS
-------
CONTENTS
METHOD fi.l Protocol for Establishing a Calibration Range
for Single Point Analysis
METHOD 8.2 Microcomputer Programs and Sample Analysis
Data Sheets
-------
METHOD 8.1 PROTOCOL FOR ESTABLISHING A CALIBRATION
RANGE FOR SINGLE POINT ANALYSES
1. OVERVIEW: Successful implementation of single point analysis requires that
an instrument be calibrated for a particular compound using a
standard solution of known concentration. This "calibration
solution" yields an instrumental response value (e.g., area, peak
height) which defines the midpoint of a restricted measurement
range, hereafter designated the "calibration range". Demarcation
of the "calibration range" is accomplished by calculation of
"calibration errors", with the criterion that the individual
"calibration errors" not exceed a predetermined value (e.g.,
Ec<10%). An acceptable range for quantisation is thereby derived
and the advantage of single point analysis realized through
utilization of this method.
2. PROCEDURE:
2.1 Propose a likely "calibration solution" and "calibration range" for
the specific compound of interest. This may be achieved through
the use of a plot (i.e., response versus concentration), previously
acquired analytical data, or a desirable quantitative measurement
range.
2.2 Prepare two stock solutions of the compound of interest and label
these as Stock A and Stock B.
Note: If the stock solutions are purchased as commercially
prepared standards, Stock A and Stock B should be
obtained from different commercial sources.
2.3 From Stock A, prepare a matching pair of "calibration solutions"
having identical concentrations. From Stock B, prepare a single
solution similar in concentration to that of the "calibration
solutions".
Note; The dilution scheme used to prepare the "calibration
solutions" from Stock A should differ significantly
from that used to prepare the pseudo-calibration
solution from Stock B.
2.4 Using chromatographic and/or spectroscopic techniques, analyze both
the Stock A "calibration solutions" and Stock B pseudo-calibration
solution in triplicate. Record the concentrations and instrumental
response values for the three solutions.
8.1-1
-------
2.5 Calculate the "normalized response" of the Stock B pseudo-
calibration solution through the use of the following relation:
Rn = R'(CA/CB)
where C/\ is the concentration of the Stock A "calibration
solutions", and CB, R, and Rn are the concentration, response,
and "normalized response" of the Stock B pseudo-calibration
solution, respectively.
2.6 Utilizing the "normalized response" of the Stock B pseudo-
calibration solution and the actual response values for the
Stock A "calibration solutions", calculate the mean "calibration
solution" response (Rc) and its coefficient of variation (Cv).
Note; The value of Rc will be used as the "calibration solution"
response for all subsequent calculations; the value of Cv
should not exceed 3% (i.e., Cv < 3%).
2.7 From Stock A, prepare four additional solutions; two of which yield
response values that define the lower (LL) and upper (UL) limits of
the "calibration range"; a solution (LC) that is intermediate between
the "calibration solution" and the lower limit of the range; and, a
solution (DC) that is intermediate between the "calibration solution"
and the upper limit of the range.
2.8 Using chromatographic and/or spectroscopic techniques, analyze the
LL, LC, UC, and UL solutions in triplicate. Calculate the mean
response and coefficient of variation for each of the four solutions.
Note: The individual values of Cv should not exceed 3%.
2.9 Utilizing the concentrations and the mean response values for the
LL, LC, UC, and UL solutions, calculate each "calibration error"
(Ec) as follows:
2.9.1 Calculate the "concentration factors" (F values) for the LL,
LC, UC, and UL solutions by normalizing the concentration of
each of the solutions to the concentration of the "calibration
solution", as denoted in the following example:
F = Concentration of upper range limit solution
UL Concentration of calibration solution"
2.9.2 Multiply the mean response (Rc) of the "calibration solution"
by each F value, thereby determining the expected response (Re)
for the LL, LC, UC, and UL solutions, as projected from the
response of the "calibration solution":
Re • (FUL)'(RC)
8.1-2
-------
2.9.3 Calculate the individual (Ec) as follows:
EC - (Ro - Re)
«e
where R0 is the mean of the observed response values for the
LL, LC, UC, and UL solutions.
2.10 Decisions regarding the adequacy of the proposed "calibration range"
and associated "calibration solution" require that the individual
"calibration errors" not exceed 10% (i.e., Ec < 10%).
2.11 When Ec > 10%, either constrict the proposed "calibration range", or
select a different "calibration solution". Changing the "calibration
solution" necessitates redefining the "calibration range".
Note: The "calibration errors" calculated for the LC and UC solutions
may assist in the demarcation of the new "calibration range",
providing they satisfy the Ec acceptance criterion (Ec < 10%).
2.12 The "calibration range" must be checked on a periodic (e.g., daily)
basis during which the analyst should:
1. Analyze the Stock A "calibration solutions" in triplicate.
2. Analyze the LL and UL "calibration range" solutions in triplicate.
3. Prepare a "fresh" pseudo-calibration solution from Stock R and
analyze in accordance with sections 2.4 - 2.5 of this protocol.
4. Calculate the mean LL, UL, and "calibration solution" responses,
ensuring that their Cv values do not exceed 3%.
5. Determine the adequacy of the "calibration range" (Ec < 10%).
2.13 When measurements outside of the proposed "calibration range" must
be performed, the analyst may select one of the following options:
1. Concentrate or dilute the sample(s).
2. Utilize the method of least squares analysis.
3. Prepare closely matching solutions of reference standards.
4. Use a more sensitive analytical technique.
5. Vary the injection size for GC or HPLC analyses.
3. CALCULATIONS: All calculations pertaining to establishment of the calibra-
tion range and subsequent sample analyses should be performed
through use of the appropriate microprocessor programs and
data sheets (refer to Method 8.2). A sample calculation
employing the programs and data sheets for the gas chromato-
graphic analysis of o-dichlorobenzene appears in Appendix A
of this Method.
8.1-3
-------
APPENDIX A
EVALUATION OF CALIBRATION RANGE - SAMPLE CALCULATION
The following sample calculation employing the Evaluation of Calibration
Range and Quantitative Analysis data sheets of Method 8.2 should serve as an
Illustrative example In the proper application of Single Point Analysis for
gas chromatographic analysis of o-dichlorobenzene. Several liters of ground
water from a sampling well suspected of being contaminated with o-dichloro-
benzene were collected and shipped to the RGB laboratory for analysis. The
protocol employed by the analyst for establishing a GC-FID calibration range
and extracting the sample was as follows:
1. PREPARATION OF STOCK SOLUTIONS AND CALIBRATION STANDARDS
1.1 A quantity of 100.5 mg of pure o-dichlorobenzene was diluted to a total
volume of 10.0 ml and labeled as Stock Solution A. The pure standard
used to prepare Stock Solution A was obtained from Research Triangle
Park.
1.2 A quantity of 75.8 mg of pure o-dichlorobenzene was diluted to a total
volume of 5.0 ml and labeled as Stock Solution B. The pure standard
used to prepare Stock Solution B was purchased from the Aldrich Chemical
Company.
1.3 A matching pair of "calibration solutions" (labeled C/AI and C/A2) were
prepared by diluting 2.5 ml of Stock Solution A to a final volume of
5.0 ml
1.4 A "pseudo-calibration solution" (labeled C/B) was prepared by diluting
2.0 ml of Stock Solution B to a final volume of 6.0 ml.
1.5 The lower limit (LL) of the calibration range was defined by diluting
1.25 ml of Stock Solution A to a final volume of 5.0 ml
1.6 An intermediate (LC) solution between the lower limit of the range and
the "calibration solution" was prepared by diluting 2.0 ml of Stock
Solution A to a final volume of 5.0 ml.
1.7 An intermediate (CU) solution between the "calibration solution" and
the upper limit of the range was prepared by diluting 3.0 ml of Stock
Solution A to a final volume of 5.0 ml.
1.8 The upper limit (UL) of the calibration range was defined by diluting
3.75 ml of Stock Solution A to a final volume of 5.0 ml.
8.1-4
-------
APPENDIX A (CONTINUED)
2. GAS CHROMATOGRAPHIC ANALYSIS OF CALIBRATION STANDARDS
Each of the standards comprising the calibration range were measured in
triplicate by flame ionization detection gas chromatographic analysis:
Calibration Standard
C/AI
C/A2
C/B
LL
LC
CU
UL
GC-FID Response (Area Counts)
3726000 3643000 3598000
3690000 3654000 3622000
3710000
1847000
2920000
4455000
5567000
3642000
1853000
2990000
4491000
5593000
3675000
1859000
2963000
4475000
5628000
3. PREPARATION AND ANALYSIS OF SAMPLES
Three one-liter (1.0 L) samples of the ground water were extracted with
20.0 ml of hexane and analyzed in triplicate:
Sample No.
1
2
3
GC-FID Response (Area Counts)
3346000 3275000 3446000
3354000 3290000 3406000
3262000 3255000 3339000
4. CALCULATIONS
The completed Evaluation of Calibration Range and Quantitative Analysis data
sheets comprise the following three pages. The microcomputer program output
sheets summarizing the input data and sample analysis results appear on pages
8.1-9 and 8.1-10 of Appendix A.
8.1-5
-------
APPENDIX A (CONTINUED)
EVALUATION OF CALIBRATION RANGE - CHROMATOGRAPHIC MEASUREMENTS
Page
of
Chromatogram Reference Nos;
Analyst; D.f, REMETA Date; Slllff? Compound(s): Q-
Approval; M. GROEMFE& Date; sMsi Sol vent (s);
QUANTITY OF STOCK MATERIALS A AND B
UNITS: g mg X ug
Tare + Stock A
- Tare Weight -
Weight Stock A « ICONS' (1)
Tare + Stock B
- Tare Weight -
Weight Stock B » "7£".g (2)
VOLUME OF INITIAL STOCK DILUTIONS
UNITS: ml X ul
Volume Stock A = IQ.Q (3)
Volume Stock B
(4)
DILUTION SEQUENCE OF CALIBRATION SOLUTIONS
LL
CS (A)
LC
TTTB)
UNITS: ml /ml
ul/ul
HI
CU
HI
UL
(TOT
RESPONSE OF CALIBRATION SOLUTIONS
Peak N»
-Baseline
Peak size
TIT
UNITS: PEAK HEIGHT PEAK AREA
* INJECTION VOLUME
CS (A2
(14)
Peak Max
-Baseline
t Peak Size
LC
(12)
CS (B)
UH
UL (17)
3*iW/c~H*
Peak Max
-Baseline
Peak Size
CS
Al)
_QH
t Response Values for Mult1component Peak Standards
and Samples Must Be Reported as the Total Response
(e.g.. Summation of Individual Peaks) Using the
'Supplemental GC Calibration Range Data Sheet1
Injection Volumes of Calibration Solutions and Samples Must be Identical.
8.1-6
-------
APPENDIX A (CONTINUED)
QUANTITATIVE ANALYSES - CHROMATOGRAPHIC MEASUREMENTS
Page *]j of 3 Are Measurements In Calibration Range? Yes
Analyst ; D. P. GEMETA Date: 5/2/g7 Compound ( s );
Approval :M. GJ&J0Jf&AO*te:S'M87 Sol vent (s):
Uncertain
t Mean (Corrected) Calibration Solution Response: 3.635'SS (1)
t Mean Calibration Solution Concentration: SZOZS" (2) UNITS: mg/ml^£_ ug/ml _
Volume of Sample Extract/Initial Dilution: 1O.O ml (3)
SAMPLE NO. /_
CHROMATOGRAM NOS.
fJ/A
* INJECTION VOLUME = 3.Qul
SAMPLE RESPONSE - UNITS: PEAK HEIGHT PEAK AREA X
Peak Max
-Baseline
Peak Size
SAMPLE (4)
33frff £3
3Z75" E3
3W6" £3
QUANTITY/VOLUME OF SAMPLE
SAMPLE DILUTION SEQUENCE
Tare + Blank
- Tare Weight -
Wt. of Sample «
Volume of Sample = /Q
g UNITS:
9
g (5) SAMPLE
00ml (5) N/fl
ml /ml ul/ul
(6)
SAMPLE NO.
CHROMATOGRAM NOS.
* INJECTION VOLUME
SAMPLE RESPONSE
UNITS: PEAK HEIGHT PEAK AREA X
Peak Max
-Baseline
Peak Size
SAMPLE (4)
3asy-£:3
32.9O £"3
3^0££3
QUANTITY/VOLUME OF SAMPLE
Tare + Blank g
- Tare Weight - g
Wt. of Sample » g (5)
Volume of Sample = IOOQm\ (5)
SAMPLE DILUTION SEQUENCE
. UNITS: ml/ml ul/ul
SAMPLE
(6)
t Refer to Calculated Output from Evaluation of Calibration Range Data Sheet.
* Injection Volumes of Calibration Solutions and Samples Must be Identical.
8.1-7
-------
APPENDIX A (CONTINUED)
QUANTITATIVE ANALYSES - CHROMATOGRAPHIC MEASUREMENTS (CONTINUATION SHEET)
Page ^ of
Analyst: O.R
Are Measurements In Calibration Range? Yes X Uncertain_
Date: 512107 Compound!s):j
Sol vent(s):
SAMPLE NO. 3 CHROMATOGRAM NOS. A//9
INJECTION VOLUME = 3.Qul
SAMPLE RESPONSE - UNITS: PEAK HEIGHT _ PEAK AREA _X_
Peak Max
-Baseline
Peak Size
SAMPLE (4)
32.62. £13
"3^ ^^^7?
333V £3
QUANTITY/VOLUME OF SAMPLE
Tare + Blank g
- Tare Weight -
SAMPLE DILUTION SEQUENCE
UNITS: ml/ml ul/ul
wt. or
Volume of
Sample
Sampl e
a q
" /000ml
(5)
(5)
SAMPLE
A///9
(6)
SAMPLE NO.
CHROMATOGRAM NOS.
SAMPLE RESPONSE
UNITS: PEAK HEIGHT
INJECTION VOLUME
PEAK AREA
ul
SAMPLE (4)
Peak Max
-Baseline
Peak Size
QUANTITY/VOLUME OF SAMPLE
Tare + Blank g
- Tare Weight - g
Wt. of Sample = g (5)
SAMPLE DILUTION SEQUENCE
UNITS: ml /ml ul/ul
Volume of Sample » ml (5)
SAMPLE
(6)
Injection Volumes of Calibration Solutions and Samples Must be Identical,
8.1-8
-------
APPENDIX A (CONTINUED)
CALIBRATION RANGE PROGRAM OUTPUT
CHROMATOGRAPHY CALIBRATION RANGE PROGRAM
»(1)WEIGHT OF CALIBRATION MATERIAL A =» 100.5 mg
*(2)WEIGHT OF CALIBRATION MATERIAL B = 75.B mg
*(3)VOLUME OF FIRST STOCK (A) DILUTION » 10 ml
»(4)VOLUME OF FIRST STOCK (B) DILUTION a 5 ml
»<5)LL DILUTION FACTOR = .25
»(6)LC DILUTION FACTOR = .4
»(7)C (STOCK A) DILUTION FACTOR • .5
*(B)C (STOCK B) DILUTION FACTOR » .333333
»(9)UC DILUTION FACTOR = .6
*(10)UL DILUTION FACTOR = .75
*(ll)MEAN LL SOLUTION RESPONSE - 1.853E+06
*(12)MEAN LC SOLUTION RESPONSE = 2.95767E+06
*(13)MEAN STOCK A/CALIBRATION SOLUTION (1) RESPONSE = 3.65567E+06
*(14)MEAN STOCK, A/CALIBRATION SOLUTION (2) RESPONSE = 3.65533E+06
*(15>M£AN STOCK'S/CALIBRATION SOLUTION RESPONSE = 3.67567E+06
*( 16) MEAN UC SOLUTION RESPONSE • 4.47367E-»-06
*(17)MEAN UL SOLUTION RESPONSE » 5.596E*06
** STOCK A/CALIBRATION SOLUTION (1) RESPONSE = 3.656E-»-06
*» STOCK A/CALIBRATION SOLUTION (2) RESPONSE » 3.655E+06
*» NORMALIZED STOCK B/CALIBRATION SOLUTION RESPONSE =» 3.655E-«-06
** MEAN CORRECTED CALIBRATION SOLUTION RESPONSE *> 3.655E-K56
«* STOCK A/CALIBRATION SOLUTION CONCENTRATION - 5.025E+40 mg/ml
*« Cv OF CORRECTED CALIBRATION SOLUTION RESPONSE = 0.0%
*» EC (LL) - 1.4X
** EC (LC) = 1. I*
** EC (UC) = 2.0%
** EC (UL) = 2. 1%
CALUCLAT ION/CHECK BY (SIGN BELOW)
8.1-9
-------
APPENDIX A (CONTINUED)
QUANTIFICATION PROGRAM OUTPUT
CHROMATOGRAPHY QUANTIFICATION PROGRAM
»(1>MEAN (CORRECTED) CALIBRATION SOLUTION RESPONSE = 3.655E+06
* (2) STOCK A/CALIBRATION SOLUTION CONCENTRATION = 5.025 mg/rnl
»(3)VOLUME OF FIRST SAMPLE DILUTION/EXTRACT - 20 ml
•SAMPLE *1
»<4>SAMPLE SOLUTION RESPONSE = 3.35567E+O6
*(5)SAMPLE VOLUME = 1000 ml
••CONCENTRATION OF SAMPLE «1 = 9.227E+01 PPM (w/w>
•SAMPLE <*2
»(4>SAMPLE SOLUTION RESPONSE = 3.35E+06
•(5)SAMPLE VOLUME = 1000 ml
••CONCENTRATION OF SAMPLE #2 = 9.211E+01 PPM (w/w)
•SAMPLE 4*3
*(4)SAMPLE SOLUTION RESPONSE - 3.28533E+06
•(5)SAMPLE VOLUME = 10OO ml
••CONCENTRATION OF SAMPLE #3 = 9.034E+01 PPM Cw/w)
CALCULATION/CHECK BY: (SIGN BELOW)
8.1-10
-------
APPENDIX B
PROTOCOL FOR USE OF SIGNIFICANT FIGURES
1. OVERVIEW: The following set of fundamental rules prescribes specific guide-
lines for retaining the proper number of significant figures when
reporting analytical results. This protocol was developed by the
Chemistry Staff of the Releases Control Branch to assure infra-
laboratory compliance with its QA/QC Program.
2. DEFINITION:
A number is said to contain only SIGNIFICANT FIGURES if all
digits in the reported result are known definitely, except the
last digit, which may be in doubt.
3. SIGNIFICANCE OF ZEROES:
3.1 Final zeroes on the right side of the decimal point are always
significant (e.g., 12.3400 has six significant figures).
3.2 Zeroes on the left side of the decimal point that are preceded by
non-zero digits are significant (e.g., 1200.34 contains six sig-
nificant figures).
3.3 Zeroes directly to the right of the decimal point that precede non-
zero digits are not significant, providing there are no significant
digits on the left side of the decimal point (e.g., 0.0024 contains
only two significant figures, whereas 10.0024 has six significant
figures).
3.4 Final zeroes in a whole number may or may not be significant, and
should be specified to avoid ambiguTty (e.g., 1000 has one significant
figure; reporting the result as 1000. implies a measurement accuracy
of + 1 and hence, four significant figures).
4. ARITHMETIC OPERATIONS
4.1 TRUNCATION
All instrument response values should be truncated (NOT ROUNDED OFF)
after the fourth digit to ensure consistency in reporting analytical
8.1-11
-------
APPENDIX B (CONTINUED)
results (e.g., 123499 is reported as 123400; 123411 is also reported
as 123400).
4.2 EXPONENTIAL NOTATION
All numbers that are either preceded by, or that terminate in, non-
significant zeroes are to be expressed in exponential notation. Use
of this format is restricted to numbers that contain more than two
non-significant zeroes (e.g., 1234000 should be expressed as 1234 E+3;
0.0000000135 should be expressed as 0.135 E-7; 1000 is expressed as
1 E+3, whereas 100 is written as 100; 0.0003 is expressed as 0.3 E-3,
whereas 0.003 is written as 0.003).
4.3 SERIES OF OPERATIONS
Retain all figures throughout the calculation and round off the final
reported result to the number of significant figures contained in the
least accurate physical parameter (refer to Section 5.3).
5. APPLICATION TO EXISTING QA/QC PROGRAM:
5.1 On the Evaluation of Calibration Range, Spectroscopic and Chromato-
graphic Data Sheets:
1. All dilution sequences must incorporate the proper number of sig-
nificant figures (e.g., a 1:10 dilution employing a 10 ml volu-
metric pipet and a 100 ml volumetric flask is correctly recorded
as 10.0/100.0, NOT 1/10);
2. Response values are entered in truncated form where applicable
(refer to Section 4.1);
3. Concentration values are entered in terms of exponential notation
where appropriate (refer to Section 4.2).
5.2 The Computer Programs DO NOT ROUND OFF the final value to the proper
number of significant figures. The analyst must therefore complete
this operation through visual examination of certain physical values
that enter into the computation. These include:
1. The gravimetric/volumetric quantities of stock and sample material;
8.1-12
-------
APPENDIX B (CONTINUED)
2. The dilution sequences employed in standard and sample preparation;
3. The sample and standard response values;
4. The injection sizes (chromatographic analyses only).
The final value must be rounded off to the proper number of signifi-
cant figures based on the number of significant figures contained in
the least accurate of these four physical parameters.
6. SAMPLE CALCULATION:
The following tabulation provides a summary of the four physical parameters
examined in a typical chromatographic analysis of a single component sample.
In this particular example, the injection sizes are the limiting physical
parameter, restricting the accuracy of the final value to TWO significant
figures. This value is recorded on the Chromatographic Data Sheet in the
appropriate box to indicate that this is the reported result.
PHYSICAL PARAMETER SCHEMATIC SEQUENCE PARAMETER ACCURACY
OR PHYSICAL VALUE (I SIGNIFICANT FIGURES)
s
T
A
N
D
A
R
D
S
A
M
P
L
E
QUANTITY OF STOCK
DILUTION SEQUENCE
RESPONSE (AREA)
INJECTION SIZE
QUANTITY/EXTRACT
DILUTION SEQUENCE
RESPONSE (AREA)
INJECTION SIZE
0.0120 g / 100.0 ml
10.0 ml / 100.0 ml
331900
3.0 ul
25.0 g / 50.0 ml
10.0 ml / 50.0 ml
234800
3.0 ul
3
3
4
2
3
3
4
2
SAMPLE CONCENTRATION (Computer Printout): 84.89303 mg/L
SAMPLE CONCENTRATION (Reported): 85 mg/L
8.1-13
-------
METHOD 8.2 MICROCOMPUTER PROGRAMS AND SAMPLE ANALYSIS DATA SHEETS
1. OVERVIEW: This particular method is comprised of several sample analysis
data sheets and their respective microcomputer programs. The
programs and data sheets were designed specifically to apply
Method 8.1 to the chromatographic and/or spectroscopic analysis
of environmental samples containing hazardous materials. Each
of the appendices in this Method addresses a specific application
of Single Point Analysis. Sample hardcopy outputs of the micro-
computer programs are available upon request.
2. CONTENTS:
2.1 APPENDIX A: CHROMATOGRAPHIC ANALYSIS
2.1.1 Calibration Range Prediction Program
This program is employed by the analyst to establish a chroma-
tographic calibration range amenable to Single Point Analysis
as outlined in Method 8.1. The criterion used to define the
acceptability of the proposed calibration range is that the
calibration errors (denoted as Ec) for the LL, LC, DC, and UL
calibration standards not exceed a critical value of 10%.
2.1.2 Quantitative Analyses Program
This program enables the analyst to determine the concentration
of environmental contaminants in the sample(s) providing that a
suitable chromatographic calibration range has been established
through use of the Calibration Range Prediction Program.
2.1.3 Chromatographic Data Sheets
There are three types of chromatographic data sheets presented
in Appendix A of this Method. The 'Evaluation of Calibration
Range1 and 'Quantitative Analyses' data sheets are employed to
transcribe analytical data from the chromatograms of calibration
standards and samples. Consequently, these data sheets furnish
the input parameters for the Calibration Range Prediction and
Quantitative Analyses Programs, respectively. The analyst is
advised to refer to the 'Sample Calculation1 in Appendix A of '
Method 8.1 for guidance in completing these two types of data
sheets. The 'Supplemental GC Calibration Range' data sheets
are used exclusively for muHi component peak standards and
samples (e.g., PCBs) in which the response values are reported
as the total response (e.g., summation of individual peaks) to
ensure accurate quantification.
8.2-1
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2.2 APPENDIX B: SPECTROSCOPIC ANALYSIS
2.2.1 Calibration Range Prediction Program
This program is employed by the analyst to establish a spectro-
scopic calibration range amenable to Single Point Analysis as
outlined in Method 8.1. The criterion used to define the
acceptability of the proposed calibration range is that the
calibration errors (denoted as Ec) for the LL, LC, UC, and UL
calibration standards not exceed a critical value of 10%.
2.2.2 Quantitative Analyses Program
This program enables the analyst to determine the concentration
of environmental contaminants in the sample(s) providing that a
suitable spectroscopic calibration range has been established
through use of the Calibration Range Prediction Program.
2.2.3 Quantitative Analyses Data Sheets
There are two types of spectroscopic data sheets presented in
Appendix B of this Method. The 'Evaluation of Calibration
Range* and 'Quantitative Analyses' data sheets are employed
to transcribe analytical data from the spectra of calibration
standards and samples. Consequently, these data sheets furnish
the input parameters for the Calibration Range Prediction and
Quantitative Analyses Programs, respectively. The analyst is
advised to refer to the 'Sample Calculation' in Appendix A of
Method 8.1 for guidance in completing these data sheets since
these are quite similar to the chromatographic data sheets.
8.2-2
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APPENDIX A
EVALUATION OF CALIBRATION RANGE - CHROMATOGRAPHIC MEASUREMENTS
Page of
Analyst:
Approval:
Chromatogram Reference Nos:
Dat e: Compound (s):_
Date: Sol vent (s):
QUANTITY OF STOCK MATERIALS A AND B
UNITS: g mg ug
VOLUME OF INITIAL STOCK DILUTIONS
UNITS: ml ul
Tare + Stock A
- Tare Weight -
Weight Stock A =•
Tare + Stock B
- Tare Weight -
Weight Stock B °
(1)
(2)
Volume Stock A = (3)
Volume Stock B « (4)
DILUTION SEQUENCE OF CALIBRATION SOLUTIONS
UNITS: ml/ml
ul/ul
LL
J5I
CS (A)
ALL
121
J5E
(TOT
RESPONSE OF CALIBRATION SOLUTIONS
UNITS: PEAK HEIGHT PEAK AREA
* INJECTION VOLUME
Peak MM
-Baseline
t Peak Size
Peak Ma«
-Baseline
t Peak Size
Peak Ma>
-Baseline
t Peak Size
LL (11)
LC (12)
CS
Al) (13)
1
CS (A2
(14)
CS (B) (15)
UC (16)
UL (17)
Response Values for Multl component Peak Standards
and Samples Must Be Reported as the Total Respons
(e.g., Summation of Individual Peaks) Using the
'Supplemental GC Calibration Range Data Sheet1
* Injection Volumes of Calibration Solutions and Samples Must be Identical.
8.2-3
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APPENDIX A (CONTINUED)
QUANTITATIVE ANALYSES - CHROMATOGRAPHIC MEASUREMENTS
Page
Analyst:
of
Approval:
Are Measurements In Calibration Range? Yes_
Date: Compound(s):
Sol vent(s):
Uncertain
Date:
t Mean (Corrected) Calibration Solution Response:
t Mean Calibration Solution Concentration:
(1)
Volume of Sample Extract/Initial Dilution:
(2) UNITS: mg/ml ug/ml
ml (3)
SAMPLE NO.
CHROMATOGRAM NOS.
SAMPLE RESPONSE
UNITS: PEAK HEIGHT
INJECTION VOLUME
PEAK AREA
ul
Peak Max
-Baseline
Peak Size
SAMPLE (4)
QUANTITY/VOLUME OF SAMPLE
SAMPLE DILUTION SEQUENCE
Tare + Blank
- Tare Weight -
Ut. of Sample =
Volume of Sample =
g
9
g (5)
ml (5)
UNITS: ml/ml ul/ul
SAMPLE (6)
SAMPLE NO.
CHROMATOGRAM NOS.
SAMPLE RESPONSE
UNITS: PEAK HEIGHT
* INJECTION VOLUME
PEAK AREA
ul
Peak Max
-Baseline
Peak Size
SAMPLE (4)
QUANTITY/VOLUME OF SAMPLE
SAMPLE DILUTION SEQUENCE
Tare + Blank
- Tare Weight -
Wt. of Sample =
Volume of Sample =
g
. g
9 (5)
ml (5)
UNITS: ml/ml ul/ul
SAMPLE (6)
t Refer to Calculated Output from Evaluation of Calibration Range Data Sheet.
* Injection Volumes of Calibration Solutions and Samples Must be Identical.
8.2-4
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APPENDIX A (CONTINUED)
QUANTITATIVE ANALYSES - CHROHATOGRAPHIC MEASUREMENTS (CONTINUATION SHEET)
Page of
Analyst:
Approval:
Are Measurements 1n Calibration Range? Yes_
Date: Compound(s):
Date: Solvent(s):
Uncertain
SAMPLE NO.
CHROMATOGRAM NOS.
SAMPLE RESPONSE
UNITS: PEAK HEIGHT
* INJECTION VOLUME
PEAK AREA
QUANTITY/VOLUME OF SAMPLE
Tare + Blank g
- Tare Weight - g
Wt. of Sample = g (5)
Volume of Sample
jnl (5)
SAMPLE DILUTION SEQUENCE
UNITS: ml/ml ul/ul
SAMPLE
ul
Peak Max
-Baseline
Peak Size
SAMPLE (4)
*
SAMPLE NO.
CHROMATOGRAM NOS.
SAMPLE RESPONSE
UNITS: PEAK HEIGHT
INJECTION VOLUME
PEAK AREA
QUANTITY/VOLUME OF SAMPLE
Tare + Blank g
- Tare Weight - g
Wt. of Sample = g (5)
Volume of Sample
ml (5)
SAMPLE DILUTION SEQUENCE
UNITS: ml/ml ul/ul
ul
Peak Max
-Baseline
Peak Size
SAMPLE (4)
SAMPLE
T6T
Injection Volumes of Calibration Solutions and Samples Must be Identical.
8.2-5
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APPENDIX A (CONTINUED)
SUPPLEMENTAL GC CALIBRATION RANGE DATA SHEET
Page of Chromatogram Reference Nos.
Analyst: Date: Competing s):
Approval: Date: Solvent(s):
RESPONSE OF CALIBRATION SOLUTIONS
UNITS: PEAK HEIGHT PEAK AREA * INJECTION VOLUME - ul
TT (ID
Peak Max
-Baseline
Peak Size.
LL UIL
Peak Max
-Baseline
Peak Size
"LIT"
Peak Max
-Baseline
Peak Size
LC
Peak Max
-Baseline
Peak Size
C/A1
Peak Max
-Baseline
Peak Size
T7AT
Peak Max
-Baseline
Peak Size
* Injection Volumes of Calibration Solutions and Samples Must be Identical
8.2-6
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APPENDIX A (CONTINUED)
SUPPLEMENTAL GC CALIBRATION RANGE DATA (CONTINUATION SHEET)
Peak Max
-Baseline
Peak Size
C/A2 (14)
Peak Max
-Baseline
Peak Size
C/AZ (14)
Peak Max
-Baseline
Peak Size
C/B (15)
Peak Max
-Baseline
Peak Size
C/B (15)
Peak Max
-Baseline
Peak Size
CU (16)
Peak Max
-Baseline
Peak Size
CU (16)
Peak Max
-Baseline
Peak Size
UL (17)
Peak Max
-Baseline
Peak Size
UL (17)
8.2-7
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APPENDIX B
EVALUATION OF CALIBRATION RANGE - SPECTROSCOPIC MEASUREMENTS
Page of
Analyst:
Approval:
Spectra Reference Nos:
Date: Compound (s):_
Date: Sol vent (s):
QUANTITY OF STOCK MATERIALS A AND B
UNITS: g mg ug
Tare + Stock A
- Tare Weight
Weight Stock A
Tare + Stock B
- Tare Weight
Weight Stock B
(1)
(2)
VOLUME OF INITIAL STOCK DILUTIONS
UNITS: ml ul
Volume Stock A
Volume Stock B
(3)
(4)
DILUTION SEQUENCE OF CALIBRATION SOLUTIONS - UNITS: ml/ml ul/ul
TT
CS (A)
cu
LC
CSTTBT
UL
(10)
RESPONSE OF CALIBRATION SOLUTIONS - UNITS: PEAK HEIGHT PEAK AREA
Pe*k Max
-Baseline
Peak Size
SOLVENT * (11)
CS (Al
(14)
Peak Max
-Baiellne
Peak Stze
LL (12)
Peak Hi*
-Baseline
Peak Size
LC (13)
UC (17)
CS (A2
(15)
UL (18)
CS (B) (16)
* Set Solvent Response Equal to 0 for Dual Beam Spectrophotometers (e.g., UV, IR)
8.2-8
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APPENDIX B (CONTINUED)
QUANTITATIVE ANALYSES - SPECTROSCOPIC MEASUREMENTS
Page of
Analyst:
Approval:
Are Measurements In Calibration Range? Yes Uncertain^
Date: Compound(s):
Solvent(s):
Date:
Mean Solvent Response:
(1) (Item # 11 on Calibration Range Data Sheet)
t QUANTITY/VOLUME OF SAMPLE BLANK
Tare + Blank g
- Tare Weight - g
Wt. of Blank = g (2)
Volume of Blank
ml (2)
t VOLUME OF EXTRACT/INITIAL DILUTION
Volume = ml (3)
(Must be Identical to Sample Volumes)
RESPONSE OF SAMPLE BLANK (Spectrum No. )
Peak Max
-Baseline
Peak Size
SAMPLE BLANK (4)
BLANK DILUTION SEQUENCE - UNITS: ml/ml
SAMPLE BLANK
Mean Calibration Solution Concentration:
Mean (Corrected) Calibration Solution Response:
(6) UNITS: mg/ml ug/ml
(7)
SAMPLE NO.
SPECTRA NOS.
SAMPLE DILUTION SEQUENCE - UNITS: ml/ml
SAMPLE
T9T
RESPONSE OF SAMPLE
QUANTITY/VOLUME OF SAMPLE
Tare + Blank g
- Tare Weight - g
Wt. of Sample = g (8)
Volume of Sample = ml (8)
* Set Solvent Response Equal to 0 for Dual Beam Spectrophotometers (e.g., UV, IR)
t Set Sample Blank and Sample Volumes Equal to 1 for In-SItu Aqueous Measurements
§ Refer to Calculated Output from Evaluation of Calibration Range Data Sheet
Peak Max
-Baseline
Peak Size
SAMPLE (10)
8-. 2-9
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APPENDIX B (CONTINUED)
QUANTITATIVE ANALYSES - SPECTROSCOPIC MEASUREMENTS (CONTINUATION SHEET)
Page of
Analyst:
Approval:_
Are Measurements 1n Calibration Range? Yes Uncerta1n_
Date: Compound (s):
Solvent(s):
Date:
SAMPLE NO.
SPECTRA NOS.
t QUANTITY/VOLUME OF SAMPLE
Tare + Blank g
- Tare Weight - g
Wt. of Sample =» g (8)
Volume of Sample
ml (8)
SAMPLE DILUTION SEQUENCE - UNITS: ml/ml
J2L
RESPONSE OF SAMPLE
Peak Max
-Baseline
Peak Size
SAMPLE (10)
SAMPLE NO.
SPECTRA NOS.
t QUANTITY/VOLUME OF SAMPLE
Tare + Blank g
- Tare Weight - g
Wt. of Sample = g (8)
Volume of Sample
_ml (8)
SAMPLE DILUTION SEQUENCE - UNITS: ml/ml
SAMPLE
RESPONSE OF SAMPLE
Peak Max
-Baseline
Peak Size
SAMPLE (10)
SAMPLE NO.
SPECTRA NOS.
t QUANTITY/VOLUME OF SAMPLE
Tare + Blank g
- Tare Weight - g
Wt. of Sample = g (8)
Volume of Sample
ml (8)
SAMPLE DILUTION SEQUENCE - UNITS: ml/ml
SAMPLE
RESPONSE OF SAMPLE
Peak Max
-Baseline
Peak Size
SAMPLE (10)
t Set Sample Volumes Equal to 1 for In-SItu Aqueous Measurements
8.2-10
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