May 1986
ANALYTICAL METHODS FOR
U.S. EPA PRIORITY
POLLUTANTS AND 301 (h)
PESTICIDES IN ESTUARINE
AND MARINE SEDIMENTS
Prepared by:
Tetra Tech, Inc.
11820 Northup Way, Suite 100
Bellevue, Washington 98005
Prepared for:
Marine Operations Division: 301 (h) Program
Office of Marine and Estuarine Protection
U.S. Environmental Protection Agency
401 M Street SW
Washington, D.C. 20460
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CONTENTS
Page
1.0 SCOPE AND APPLICATION I-i
2.0 SUMMARY OF METHOD 1-4
3.0 INTERFERENCES 1-5
4.0 SAFETY 1-6
5.0 APPARATUS AND EQUIPMENT 1-7
6.0 REAGENTS AND CONSUMABLE MATERIALS I-11
7.0 SAMPLE COLLECTION, PREPARATION, AND STORAGE 1-15
8.0 CALIBRATION AND STANDARDIZATION 1-16
9.0 QUALITY ASSURANCE/QUALITY CONTROL 1-21
10.0 PROCEDURE 1-24
11.0 QUANTITATIVE DETERMINATION (CALCULATIONS) 1-37
12.0 PRECISION AND ACCURACY 1-44
13.0 REFERENCES 1-45
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CONTENTS
LIST OF FIGURES iv
LIST OF TABLES v
ACKNOWLEDGEMENTS vi
INTRODUCTION viii
SECTION I. ANALYSIS OF EXTRACTABLE ORGANIC COMPOUNDS
IN ESTUARINE AND MARINE SEDIMENTS
1.0 SCOPE AND APPLICATION 1-1
2.0 SUMMARY OF METHOD 1-4
3.0 INTERFERENCES 1-5
4.0 SAFETY 1-6
5.0 APPARATUS AND EQUIPMENT 1-7
6.0 REAGENTS AND CONSUMABLE MATERIALS l-ll
7.0 SAMPLE COLLECTION, PREPARATION, AND STORAGE 1-15
8.0 CALIBRATION AND STANDARDIZATION 1-16
9.0 QUALITY ASSURANCE/QUALITY CONTROL 1-21
10.0 PROCEDURE 1-24
11.0 QUANTITATIVE DETERMINATION (CALCULATIONS) 1-37
12.0 PRECISION AND ACCURACY 1-44
13.0 REFERENCES 1-45
SECTION II. ANALYSIS OF VOLATILE ORGANIC COMPOUNDS
IN ESTUARINE AND MARINE SEDIMENTS
1.0 SCOPE AND APPLICATION II-l
2.0 SUMMARY OF METHOD 11-2
3.0 INTERFERENCES II-3
11
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4.0 SAFETY II-4
5.0 APPARATUS AND EQUIPMENT II-5
6.0 REAGENTS AND CONSUMABLE MATERIALS 11-9
7.0 SAMPLE COLLECTION, PREPARATION, AND STORAGE 11-12
8.0 CALIBRATION AND STANDARDIZATION 11-13
9.0 QUALITY CONTROL 11-18
10.0 PROCEDURE 11-20
11.0 QUANTITATIVE DETERMINATION (CALCULATIONS) 11-25
12.0 PRECISION AND ACCURACY 11-27
13.0 REFERENCES 11-27
SECTION III. ANALYSIS OF METALS AND METALLOIDS
IN ESTUARINE AND MARINE SEDIMENTS
1.0 SCOPE AND APPLICATION III-l
2.0 SUMMARY OF METHOD 111-2
3.0 DEFINITIONS III-2
4.0 INTERFERENCES III-3
5.0 SAFETY III-4
6.0 APPARATUS AND EQUIPMENT III-5
7.0 REAGENTS AND CONSUMABLE MATERIALS III-7
8.0 SAMPLE COLLECTION, PREPARATION, AND STORAGE 111-8
9.0 CALIBRATION AND STANDARDIZATION II1-9
10.0 QUALITY CONTROL 111-11
11.0 PROCEDURE 111-18
12.0 CALCULATIONS 111-21
13.0 PRECISION AND ACCURACY 111-21
14.0 REFERENCES 111-21
111
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FIGURES
Number Page
1-1 Relative response calibration curve 1-47
1-2 Extracted ion current profiles for chromatographically
resolved labeled (^2/2) and unlabeled (mi/z) pairs 1-47
1-3 Extracted ion current profiles for (3A) unlabeled compound,
(38) labeled compound, and (3C) equal mixture of unlabeled
and labeled compounds 1-47
1-4 Flow chart for sample preparation 1-48
II-l Apparatus for vacuum distillation and cryogenic concen-
tration H-28
II-2 Relative response calibration curve 11-29
11-3 Extracted ion current profiles for (A) the unlabeled pol-
lutant, (3) the labeled analog, and (C) a mixture of the
labeled and unlabeled compounds 11-29
III-l Quality control chart 111-24
1v
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TABLES
Number Page
1-1 Gas chromatography of extractable compounds 1-49
1-2 OFTPP mass-intensity specification 1-52
1-3 Summary of available precision and recovery data 1-53
1-4 Precision and accuracy of method blanks 1-54
II-l Volatile organic analytes 11-30
II-2 BFB mass-intensity specification 11-31
II-3 Percent spike recoveries for volatile priority pollutants
using vacuum distillation 11-32
III-l General information for each priority pollutant metal 111-25
III-2 Typical data obtained on a certified reference material 111-26
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ACKNOWLEDGEMENTS
This document has been reviewed by the 301(h) Task Force of the Environ-
mental Protection Agency, which includes representatives from the Water
Management Divisions of U.S. EPA Regions I, II, III, IV, IX, and X; the
Office of Research and Development Environmental Research Laboratory -
Narragansett (located in Narragansett, RI and Newport, OR), and the Marine
Operations Division in the Office of Marine and Estuarine Protection, Office
of Water.
This technical guidance document was produced for the U.S. Environmental
Protection Agency under the 301(h} post-decision technical support contract
No. 68-01-6938, Allison J. Duryee, Project Officer. This report was prepared
^y Tetra Tech, Inc., under the direction of Or. Thomas C. Ginn.
SECTION I
The primary authors were Mr. Robert C. Barrick and Mr. Harry R. Seller.
The assistance of Mr. Raleigh C. Farlow is appreciated.
Existing U.S. EPA analytical methods were incorporated into Section I
whenever possible. Specifically, many sections were adapted from the Contract
Laboratory Program for Organlcs Analysis (Section I, reference 2) and U.S. EPA
Method 1625 Revision B (Section It reference 3), which was developed by the
Industrial Technology Division of the Office of Water Regulation and Standards.
Validation data presented in Section I (Precision and Accuracy) were
generated by California Analytical Laboratories and Weyerhaeuser Technology
Center.
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SECTION II
The primary authors were Mr. Robert C. 3amck and Mr. Harry R. Seller.
The assistance of Mr. Raleigh C. Farlow is appreciated.
The procedure described in Section II is largely a compilation of
methods developed by U.S. EPA. Specifically, the methods were developed
by the Environmental Monitoring Systems Laboratory (EMSL) in Las Vegas
(Section II, references 1 and 2) and the Industrial Technology Division
of the Office of Water Regulation and Standards (Section II, reference 3).
Dr. M. Hiatt (Analytical Technologies, Inc., National City, CA, previously
at EMSL Las Vegas) was a valuable source of technical information presented
in th-is document.
SECTION III
The primary authors were Mr. Robert C. Barrick, Mr. Harry R. Seller,
and Mr. Robert W. Deverall. The assistance of Or. Charles R. Lytle is
appreciated.
Validation data presented in Section III (Precision and Accuracy)
were generated by Analytical Service Laboratories, Ltd.
Mention of trade names or commercial products herein does not constitute
endorsement for use by U.S. EPA or Tetra Tech, Inc.
vil
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INTRODUCTION
The three analytical methods in this document have been designed to be
consistent with probable uses of 301(h) monitoring data. Comparison of
sediment contaminant concentrations from contaminated and relatively uncon-
taminated areas often require sensitive analytical techniques for a wide
range of chemically diverse pollutants. The recommended 301(h) procedures
allow for sensitive analyses of the target compounds with a reasonable
amount of laboratory effort. Organophosphate 301(h) pesticides have not yet
been tested with the recommended techniques (i.e., Malathion, Parathion,
Oemeton, Guthion). Analyses for 2.3,7,8-TCOO with appropriate detection
limits will require the dedicated U.S. EPA Contract Laboratory Program
procedure for dioxin analysis (9/15/83), which involves selected ion monitoring
(SIM) GC/MS analysis.
There are currently no formally approved U.S. EPA procedures for analyzing
priority pollutants and 301(h) pesticides in sediments at trace levels
(e.g., at the low part per billion level for organic pollutant analysis).
However, various U.S. EPA procedures were reviewed during development of
this report [e.g.. Interim Methods for the Sampling and Analysis of Priority
Pollutants in Sediments and Fish Tissue (1977. revised 1980); Contract
Laboratory Program procedures for organics analysis and inorganics analysis].
Consequently, the recommended 301(h) procedures include portions of U.S. EPA
analytical and quality assurance procedures that were considered appropriate
for sensitive, full-scan analyses. The 301(h) methods have been assembled
according to guidelines for EMSL (Environmental Monitoring and Support
Laboratory, Cincinnati) analytical methods (as specified in EPA-600/8-83-020).
viii
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SECTION I
ANALYSIS OF EXTRACTABLE ORGANIC COMPOUNDS
IN ESTUARINE AND MARINE SEDIMENTS
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ANALYSIS OF 6XTRACTABLE ORGANIC COMPOUNDS
IN ESTUARINE AND MARINE SEDIMENTS
1.0 SCOPE AND APPLICATION
1.1 This method is designed to determine the semivolatlle priority pollutants
(Table 1-1) associated with the Clean Water Act Section 301(h) regulation
[40 CFR 125.58(lc} and (v)]. Additional compounds amenable to extraction and
analysis by capillary column gas chromatography-mass spectrometry (GC/MS)
and/or gas chromatography-electron capture detection (GC/ECD) may be suitable
for analysis, subject to testing.
These procedures are applicable when low part per billion analyses are
required to monitor differences between sediments from relatively uncon-
"aminated reference areas and those from contaminated estuarine and marine
environments.
Two GC/MS options included 1n the method are analyses by isotope dilution
GC/MS (strongly recommended) or by a GC/MS internal standard technique
(minimum required). In both cases, the laboratory procedures for sample
extraction and concentration of the resulting extract are identical. Compound-
specific recovery corrections used in the isotope dilution technique are
designed to increase the accuracy of the analysis and the comparability of
results among laboratories. In addition, use of the multiple recovery
standards in each analysis increases confidence in the validity of detection
limits reported for undetected target compounds. By forcing a search for
every recovery standard 1n the sample extract (more than 50 are available),
the technique also increases the efficiency of detection and reporting
frequency of target compounds that otherwise may be overlooked in complex
extracts.
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1.2 The compounds listed in Table 1-1 include pesticides subject to regulation
under Section 301(h) of the dean Water Act. However, the applicability of
this .Tiethod to non-chlorinated organoohosohorous pesticides (Malathion,
Parathion, Oemeton, and Guthion) has not been demonstrated. Chemists at the
Food and Dr-jg Administration recently published a technique for determining
organophosphate pesticides of wide ranging polarity in matrices including
fatty animal tissue (J.J. Blaha and P.J. Jackson, J. Assoc. Anal. Chem.,
Vol. 68, pp. 1095-1099, 1985): The technique involves liquid-liquid parti-
tioning and gel permeation chromatography [both are included in this recommended
301(h) procedure] as well as N/P alkali thermionic or flame photometric
detection. Further work is required to determine the suitability of the
recommended 301(h) procedure for organophosphorous pesticides in sediments.
1.3 The detection limit of this method is usually dependent upon the level
of interferences rather than instrumental limitations. The limits listed in
Table 1-1 represent the minimum quantity that can be detected with no
interferences present.
Lower limits of detection (LLD) are established by analysts based on
their experience with the instrumentation and with interferences in the
sample matrix being analyzed. LLD are greater than the instrumental detection
limits in Table 1-1 because they take into account sample interferences. To
estimate LLD, the noise level should be determined in the retention window
for the quantitation mass of representative analytes. These determinations
should be made for at least three field samples in the sample set under
analysis. The signal required to attain a signal/noise ratio of at least
two should then be estimated. This signal is the minimum response required
to identify a potential signal for quantification. The LLD is the concentration
corresponding to the level of this signal based on calibrated response
factors. Based on best professional judgment, this LLD would then be applied
to samples in the set with comparable or lower interference. Samples with
much higher interferences (e.g., at least a factor of two higher) should be
assigned LLO at a multiple of the original LLD.
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These LLD values may be less than the rigorously defined method detection
imits specified in the revised "Guidelines Establishing Test Procedures for
the Analysis of Pollutants" (40 CFR Part 136, 10/26/84). This latter procedure
requires the analysis of seven replicate samples and a statistical determina-
tion of the method detection limit with 99 percent confidence. Data quantified
between the LLO and the rigorous method detection limit are valid and useful
in environmental investigations of low-level contamination, but have a lower
statistical confidence associated with them than data quantified above the
method detection limit.
LLD for the described analytical method on a dry-weight basis are 10-25
ug/kg for aromatic hydrocarbons, phthalates, chlorinated hydrocarbons, and
halogenated ethers (GC/MS analysis). LLD for GC/MS analyses of pesticides
are 50 ug/kg (dry weight). The corresponding GC/ECO detection limits for
pesticides are 0.1-5 ug/kg. An LLD of 10 ug/kg is attainable for GC/ECO
analysis of total PCBs.
*.4 The GC/MS portions of this method are for use only by analysts experienced
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2.0 SUMMARY OF METHOD
2.1 A homogenized sediment samole is Soxhlet-extracted with methylene
chonde/nethanol (2/1, //v). The resulting extract is subjected to liauid-
liquid partitioning with water and is dried by elution through a sodium
sulfate column. Elemental sulfur, a common interferent in estuarine and
marine sediments, is removed from the extract with metal 1 ic mercury. Biological
macromolecules are then removed from the extract by gel permeation chromato-
graphy (GPC) (reference 2), A portion of the extract (20%) is subjected to
alumina chromatography to separate polar compounds from pesticides and PCBs
prior to capillary GC/ECO analysis (reference 2). The remaining 80. of the
extract is subjected to reverse phase column chromatography (bonded C^g
solid phase) to reduce interferences from unresolved paraffinic hydrocarbons
prior to capillary GC/MS analysis for acid, base, and neutral compounds. An
isotope dilution technique (EPA Method 1625 Revision B, reference 3) is
highly recommended but not required for all compounds analyzed by GC/MS.
This technique involves spiking the homogenized sediment sample with the
stable isotope-labeled analogs of most of the pollutants to be analyzed by
GC/MS. The advantage of isotope dilution is that reliable recovery corrections
can be made for each analyte with a labeled analog or a chemically similar
analog.
2.1.1 Much of the text of EPA Method 1625 Revision 3 has been incorporated
into this method in modified form. The modifications were necessary because,
in relation to Method 1625 Revision 8, the present method involves different
sample matrices (sediments), different calibration requirements, and additional
analytes (pesticides and PCBs, both requiring GC/ECD analysis).
2.2 Identification of compounds is performed by comparing the GC retention
times and background-corrected characteristic spectral masses with those of
authentic standards. Tentative identifications of low levels of pesticides
and PCBs are made by comparing GC retention times to standards. The identities
of pesticides and PCBs are confirmed by GC/ECO analysis on an alternative
column phase or by GC/MS when sufficient concentrations occur.
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\3 Quantitative analysis is performed by GC/MS using extracted ion current
profile (EICP) areas. Isotope dilution, with labeled analogs of pollutants
acting as recovery standards, is the method of quantification when labeled
compounds are available. When the isotope dilution technique is used but
certain labeled compounds are unavailable [e.g., labeled indeno(l,2,3-c,d)
pyrene], the nearest eluting, most chemically similar labeled compound is
used as a recovery standard. Pesticides and PCBs are quantified by an
internal standard method. Concentrations of compounds quantified by GC/MS
are reported after correcting for method recoveries when the isotope dilution
technique is used. Recoveries of Isotope labeled standards are determined
with the internal standard technique.
3.0 INTERFERENCES
3.1 Solvents, reagents, glassware, and other sample processing hardware may
yield contamination artifacts and/or elevated baselines, causing misinter-
oretation of chromatograms and spectra. All materials should be demonstrated
o be free from Interferences under the conditions of the analysis by running
method blanks Initially and with each sample lot (Sect. 9.4). Specific
selection of reagents and purification of solvents by distillation in all-
glass systems are required. H1gh-pur1ty, dist1lled-in-glass solvents are
commercially available (e.g., Burdlck and Jackson Laboratories, Muskegon, MI).
An effective way of cleaning laboratory glassware is to cover it with aluminum
foil, heat it at 450° C for several hours, and rinse it with polar and non-
polar solvents before use. Note that heating without subsequent solvent
rinsing may not eliminate laboratory residues of PCBs and other chlorinated
hydrocarbons.
3.2 Phthalates are common laboratory contaminants that are used widely as
plasticizers. Phthalates can derive from plastic labware, plastic tubing,
plastic gloves, plastic coated glassware clamps, and have been found as a
contaminant In Na-SQ^. Polytetrafluoroethylene (PTFE) can be used instead
of polypropylene or polyethylene to minimize this potential source of
ontamination. However, use of PTFE labware will not necessarily preclude
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all ohthalate contamination. Also ?TFE has been shown to be caoable of
adsoroing certain oriority pollutants, so careful rinsing must be emoloyed.
3.3 Interferences coextracted from sediment samples affect the lower limits
of detection (ILQ) and quantitation limits. For this reason, sample extract
cleanup is necessary to yield reproducible and reliable analyses of contaminants
present at low concentrations in sediment samples.
3.3.1 Elemental sulfur, often prevalent in poorly oxygenated sediments,
is coextracted with organic pollutants and can interfere significantly with
both GC/ECD and GC/MS analyses. Sulfur removal is an integral step in this
method to alleviate this interference (Sect. 10.1.11).
3.3.2 Paraffinic, chromatographical ly unresolvable hydrocarbons,
derived from petroleum contamination, can interfere with GC/MS analyses of a
broad range of compounds. Reverse phase column chromatography (Sect.
10.1.16) is included in the procedure to reduce this interference.
4.0 SAFETY
4.1 The toxicity or carcinogerncity of each compound or reagent used in
this method has not been precisely determined. However, each chemical
compound should be treated as a potential health hazard and exposure should
be reduced as much as possible. The laboratory is responsible for maintaining
a current awareness file of OSHA regulations regarding the safe handling of
the chemicals specified in this method. These procedures for the safe
handling of chemicals should be made available to and followed by all personnel
involved in these analyses. Additional information on laboratory safety can
be found in references 4-6.
4.2 The following compounds covered by this method have been tentatively
classified as known or suspected human or mammalian carcinogens: benzene,
benzo(a)anthracene, 3t3'-d1chlorobenzidine, benzo(a)pyrene, dibenzo(a.h)anthra-
cene, N-nitrosod1methylamine, 4,4'-OOT, alpha-, beta-, delta-, and gamma-
hexachlorocyclohexane, and PCBs. Standards of these compounds should be
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prepared in a hood and a NIOSH/MESA-approved toxic gas respirator should be
orn when high concentrations are handled. All people working with toxic
chemicals should receive adequate instruction and training on when and how
to use respirators. See OSHA regulations for further guidance.
5.0 APPARATUS AND EQUIPMENT
5.1 Soxhlet Extractor - 50-ml extractor (Corning 3740-S or equivalent), or
85-mL extractor (Corning 3740-M or equivalent), with 250-mL flask (Corning
4320-250 or equivalent) and condenser with 34/45 joint. Cellulose thimbles
of the appropriate size should be cleaned with the extraction solvent mixture
for at least 30 cycles.
5.2 Drying Column - 30 cm x 2 cm borosillcate glass chromatography column
with glass wool plug. Glass wool should be extracted with the appropriate
solvents and allowed to dry before use.
~.3 Kuderna-Oanish (K-0) Apparatus
5.3.L Concentrator Tube - 10 ml, graduated (Kontes K-570050-1025 or
equivalent). A ground glass stopper (19/22 joint) is used to prevent
evaporation of extracts.
5.3.2 Evaporation Flask - 500 ml (Kontes K-570050-0500 or equivalent),
attached to concentrator tube with springs (Kontes K-662750-0012).
5.3.3 Snyder Column - three-ball macro (Kontes K-503000-0232 or equiva-
lent).
5.3.4 Snyder Column - two-ball micro (Kontes K-469002-0219 or equivalent).
5.3.5 Silicon Carbide Boiling Chips - approximately 10/40 mesh, extracted
with methylene chloride and heated at 450° C for 1 h minimum. Uncleaned
boiling chips can be a significant source of contamination.
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5.4 Separator/ Funnel 500 ml, borosilicate glass with PTFE stoocock.
5.5 Borosilicate Glass Beaker - 400 -""L and 100 it.
5.6 Water Bath - seated, with concentric ring cover, caoable of teiiperature
control (^_20 C), installed in a fume hood.
5.7 Samole Vials - amber glass, 2-5 ml with PTFE-lined screw cao.
5.8 Analytical Balance - capable of weighing 0.1 mg.
5.9 Nitrogen evaporation device - equipped with a water bath that can be
maintained at 35-40° C. The N-Evao by Organomation Associates, Inc., South
Berlin, MA is suitable.
5.10 Balance - capable of 100 g to the nearest 0.01 g.
5.11 Disposable Pasteur Pipets - sealed with aluminum foil and annealed at
450° C for several h, and rinsed with solvents before use.
5.12 Drying Oven.
5.13 Annealing Oven - capable of reaching 450 C.
5.14 Dessicator.
5.15 Chromatography Column for Alumina - 5-mL, disposable, borosilicate
glass serological pipet with borosilicate glass wool plug. (Glass wool must
be extracted with the appropriate solvents and allowed to dry before use).
5.16 Reverse Phase Cleanup Columns - 3-mL, solid phase extraction (SPE)
columns containing Octadecyl (Baker-10 SPE, #7020-3, or equivalent). Column
cleaning/conditioning is discussed in Sect. 10.1.16.
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5.17 Gel Permeation Chromatography Cleanup Device -
5.17.1 Automated system: gel permeation chromatograph (GPC), Analytical
Biochemical Labs, Inc. GPC Autoprep 1002, including:
25 mm ID x 600 700 mm glass column packed with 70 g of Bio
Beads S-X3
Syringe - 10 ml with Luer Lok fitting
t Syringe Filter Holder and Fitters - stainless steel and PTFE,
Gelman 4310 or equivalent.
5.17.2 Manual system assembled from parts (Wise, R.H., D.F. Bishop,
R.I. Williams, and B.M. Austern. Gel permeation Chromatography in the GC/MS
analysis of organics in sludges. U.S. EPA, Municipal Environmental Research
Laboratory, Cincinnati, OH. 45268). (See reference 2, p. 0-35).
5.18 Gas Chromatograph 1) one equipped with electron capture detector
(ECD) and 2) one interfaced to the mass spectrometer (Sect. 5.19). Both
should have spHtless injection ports for capillary column, temperature
programs with 30° C hold, and should meet all the performance specifications
in Sect. 9.9.
5.18.1 Column - 30*5 m x 0.25*0.02 mm 1.0. 51 phenyl, 94% methyl, IS
vinyl sllicone bonded phase (0.25 urn film thickness) fused silica capillary
column (J & W OB-5 or equivalent).
5.19 Mass Spectrometer - 70 eV electron impact ionization, should repeatedly
scan from 35 to 450 amu in 0.95 to 1.00 second and should produce a unit
resolution (valleys between m/z 441-442 less than 101 of the height of the
441 peak), background-corrected mass spectrum from 20 ng decafluorotri-
phenylphosphine (DFTPP) introduced through the GC inlet. The spectrum
should meet the mass-intensity criteria 1n Table 1-2 (reference 7). The use
of a conversion dynode to enhance high mass sensitivity is recommended. The
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mass spectrometer should be coupled with the GC such that the end of the
capillary column terminates within 1 CT of the ion source out does not
intercept the electron or ion beams. All portions of the column that connect
the GC to the ion source should remain at or above the column temperature
during analysis to areclude condensation of less volatile compounds.
5.20 Data System - should collect and record MS data, store mass intensity
data in spectral libraries, process GC/MS data, generate reports, and compute
and record response factors.
5.20.1 Data Acquisition - mass spectra should be collected continuously
throughout the analysis and stored on a mass storage device.
5.20.2 Mass Spectral Libraries - user-created libraries containing
mass spectra obtained from analysis of authentic standards should be employed
to reverse search GC/MS runs for the compounds of interest (Sect. 8.2).
5.20.3 Data Processing - the data system should be used to search,
locate, identify, and quantify the compounds of interest in each GC/MS
analysis. Software routines should be employed to compute retention times
and peak areas. Displays of spectra, mass chromatograms, and library compari-
sons are required to verify results.
5.20.4 Response Factors and Multipoint Calibrations - the data system
should be used to record and maintain lists of response factors (response
ratios for the isotope dilution technique) and multipoint calibration curves
(Sect. 8). Computations of relative standard deviation (coefficient of
variation) are useful for testing calibration linearity.
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«,0 REAGENTS AND CONSUMABLE MATERIALS (partially adapted from references 2
d 3).
6.1 Reagents
6.1.1 Acetone, benzene, n_-hexane, isooctane, methanol, and methylene
chloride (CH2C12) (pesticide quality, distilled-in-glass).
6.1.2 Alumina neutral, super Woelm or equivalent (Universal Scientific,
Atlanta, GA). Extract alumina with methylene chloride for 30-40 cycles in a
Soxhlet extractor to remove contamination. Allow solvent to evaporate.
Prepare activity III alumina by adding 7 percent (v/w) reagent water to
neutral alumina that has been activated at approximately 225° C for at least
2 h or preferably overnight. Store in tightly sealed, clean glass container.
6.1.3 Hydrochloric acid - concentrated, make 2N HC1 with reagent
water. Solvent clean in a separatory funnel with methylene chloride.
6.1.4 Metallic mercury - reagent mercury cleaned with pesticide quality
CH-Cl- or equivalent.
6.1.5 Potassium hydroxide - reagent grade, 6N in reagent water.
Solvent clean in a separatory funnel with methylene chloride.
6.1.6 Sodium sulfate - reagent grade, granular anhydrous, rinsed with
CH2C12 (20 ml/g) and conditioned at 450° C for 1 h minimum.
6.1.7 Reagent water - water in which the compounds of interest and
interfering compounds are not detected by this method.
6.2 GPC Calibration Solutions:
6.2.1 Corn oil - 200 mg/mL in CHC1.
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6.2.2 Bis(2-ethylhexyl)phthalate and pentachlorophenol - 4 mg/ml in
6.3 Stock Standard Solutions purchased as solutions or mixtures with
certification to their purity, concentration, and authenticity, or prepared
from materials of known purity and composition. If the compound purity is
96 percent or greater, the weight may be used without correction to compute
the concentration of the standard. When not being used, standards are
stored in the dark at -20 to -10° C in screwcapped vials with PTFE-lined
lids. A mark is placed on the vial at the level of the solution so that
solvent evaporation loss can be detected. The vials are brought to room
temperature prior to use. Any precipitate is redissolved and solvent is
added if solvent loss has occurred.
6.3.1 Preparation of stock solutions prepare in methylene chloride,
benzene, isooctane, or a mixture of these solvents according to the steps
below. Observe the safety precautions given in Sect. 4. The large number
of labeled and unlabeled acid and base/neutral compounds used for combined
calibration (Sect. 8) and calibration verification (Sect. 9.9.1.3) require
high concentrations (approximately 40 mg/mL) when individual stock solutions
are prepared, so that dilutions of mixtures will permit calibration with all
compounds in a single set of solutions. The working range for most compounds
is 1-50 ug/mL. Compounds with a reduced MS response may be prepared at
higher concentrations.
Standards for GC/ECD have lower working ranges (e.g., 0.04 to 2.0 ug/mL
for single component pesticides) than GC/MS standards. However, GC/ECD
stock solutions should be prepared with at least 10 mg of the pure material
(e.g., in 10 rrt. of solvent) to reduce potential weighing error.
6.3.2 Dissolve an appropriate amount of assayed reference material in
a suitable solvent. For example, weigh 400 mg naphthalene in a 10-mL ground
glass stoppered volumetric flask and fill to the mark with benzene. After
the naphthalene is completely dissolved, transfer the solution to a 15 ml
vial with PTFE-lined cap.
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6.3.3 Stock standard solutions should be checked for signs of degradation
prior to the preparation of calibration or performance test standards.
Quality control check samples that can be used to determine the accuracy of
calibration standards are available from the U.S. Environmental Protection
Agency, Environmental Monitoring and Support Laboratory, Cincinnati, Ohio
45268.
6.3.4 Stock standard solutions should be replaced after 6 mo, or
sooner if comparison with quality control check samples indicates a change
in concentration.
6.4 Injection Internal Standard Solutions
6.4.1 GC/MS internal standard solution - prepare 2,2'-difluorobiphenyl
(DFB) at a concentration of 2 mg/mL in benzene.
6.4.2 GC/ECO Internal standard solution - prepare decafluorobenzo-
phenone (OFBP) at a concentration of 2.5 ug/ml in isooctane.
6.5 GC/MS Secondary Dilution Standards using stock solutions (Sect. 6.3),
prepare a secondary standard containing each of the unlabeled priority
pollutants 1n Table 1-1 at a concentration of 100 ug/mL, or at a higher
concentration appropriate to the MS response of the compounds.
6.6 Labeled Compound Spiking Solution - prepare a spiking solution from
stock standard solutions prepared as in Sect. 6.3, or from mixtures, at a
concentration of 100 ug/mL or at a concentration appropriate to the MS
response of each compound. The deuterium and C-labeled compounds listed
in Table 1-1 are commercially available individually or as mixtures (e.g.,
Merck Sharp & Dohme/Isotopes, Montreal, Canada].
6.7 Solutions for obtaining authentic mass spectra (Sect. 8.2) - prepare
mixtures of labeled and unlabeled compounds at concentrations that will
assure that authentic spectra are obtained for storage in libraries.
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6.8 Calibration Solutions - the concentrations of calibration solutions
suggested in the following sections are intended to bracket concentrations
that will be encountered during samole analysis without overloading GC
columns or saturating detection systems.
6.8.1 GC/MS calibration solutions - combine O.I ml of the spiking
solution (Sect. 6.6) with 10, 50, 100, 200, and 500 uL of the secondary
dilution solution (Sect. 6.5) and bring to 1.00 ml total volume each. This
will produce calibration solutions of nominal I, 5, 10, 20, and 50 ug/nt of
the pollutants and a constant nominal 10 ug/mL of the labeled compounds.
Spike each solution with 10 uL of the GC/MS internal standard solution,
yielding 20 ug/mL.
6.8.2 PCB calibration solutions -
6.8.2.1 Aroclor stock solution for GC/MS - prepare a solution in
hexane with 250 ng/uL of each of three PCB mixtures, Aroclor 1016, Aroclor
1254, and Aroclor 1260.
6.8.2.2 Aroclor standard solution for GC/ECO - dilute the stock
solution (Sect. 6.8.2.1) to one-tenth its original concentration. It is
essential that this solution be prepared directly from the batch used for
Sect. 6.8.2.1. Combine 20, 50, 250, 500, and 1,000 uL of the diluted standard
with 100 uL of the GC/ECO internal standard solution (Sect. 6.4.2) and bring
each solution to a final volume of 5.0 mL.
This will produce calibration solutions of nominal concentrations of
100, 250, 1,250, 2.500. and 5.000 ng/mL of the 1:1:1 Aroclor mixture and a
constant nominal concentration of 50 ng/mL of internal standard.
6.8.3 Pesticide calibration solution - combine 20 uL of the GC/ECD
internal standard solution with 2. 5, 10. 50, and 100 uL of a 20 ug/mL stock
solution of all chlorinated pesticides listed in Table 1-1 (except toxaphene)
and bring to a 1.0 ml total volume. This will produce calibration solutions
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of 40, 100, 200, 1,000, and 2,000 ng/ml of each pesticide and a constant
nternal standard concentration of 50 ng/mL.
6.8.4 Toxaphene calibration solution - prepare toxaphene solutions of
100, 250, 1,250, 2,500, and 5,000 ng/mL with constant internal standard
concentration of 50 ng/mL.
6.8.5 OFTPP solution - prepare at 20 ug/mL in acetone from a stock
solution at I mg/mL. The dilute (20 ug/mL) solution is susceptible to
adsorption to vial walls and reaction with solvent impurities and may require
weekly replacement. The stock solution is likely to be stable for 6 mo to
several years (reference 1).
6.9 Stability of Solutions - al'I standard solutions (Sect. 6.4-6.8.4)
should be analyzed within 48 h of preparation and on a monthly basis thereafter
for signs of degradation. Standards will remain acceptable if response
factors relative to the internal standard correspond within +15 percent to
hose obtained in the initial analysis of the standard.
7.0 SAMPLE COLLECTION. PREPARATION. AND STORAGE
7.1 Minimize handling and avoid possible sources of contamination during
collection (e.g., sampling gear, grease from ship winches or cables, ship
engine exhaust, improper subsampling procedures).
7.2 Collection of a minimum of 100 g (wet weight) should be sufficient for
analysis. Sediment samples are stored in 240-fflL (8-oz) or larger, wide-mouth
jars with PTFE-Hned screw Hds. The container, lid, and liner should be
detergent washed, rinsed twice with tap water, once with distilled water,
once with methanol or acetone, and once with high-purity methylene chloride.
Firing of the glass jar at 450° C for 1 h may be substituted for the solvent
rinses.
7.3 Samples should be stored in the dark and frozen at -20° C until
.'xtraction. Care should be taken to prevent container breakage during
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freezing. Leave sufficient headsoace for the water to expand and freeze the
containers at an angle.
7.4 U.S. EPA gives no official guidance on sediment holding times but
recommends t.nat water samoles stored at 4° C be extracted within 10 days of
samole receiot (reference 2). Because sediments can be frozen at -20° C,
longer holding times (e.g., up to 6 mo) are appropriate. Extracts should be
analyzed within 40 days of extraction (reference 2). Effort should be made
to analyze the samples as soon as possible after extraction because some of the
more labile analytes may degrade in solution. Degradation may occur even in
the dark under refrigeration, possibly as the result of free radical formation.
3.0 CALIBRATION AND STANDARDIZATION (adapted from reference 3)
8.1 Establish the GC/MS operating conditions in Table 1-1. Analyze standards
per the procedure in Sect. 10.2 to demonstrate that the analytical system
meets the detection limits in Table 1-1 and the mass-intensity criteria in
Table 1-2 for 20 ng OFTPP.
8.2 Mass Spectral Libraries - detection and identification of compounds of
interest are dependent upon spectra stored 1n user-created libraries.
8.2.1 Obtain a mass spectrum of each pollutant, labeled compound, and
the internal standard by analyzing an authentic standard either singly or as
part of a mixture in which no interference exists between closely eluting
components. Confirmation that only a single compound 1s present is attained
by examination of the spectrum. Fragments not attributable to the compound
under study indicate the presence of an interfering compound.
8.2.2 Adjust the analytical conditions and scan rate (for this test
only) to produce an undlstorted spectrum at GC peak maximum. An undistorted
spectrum will be obtained if five complete spectra are collected across the
upper half of the GC peak. Software algorithms designed to "enhance" the
spectrum may eliminate distortion, but may also eliminate authentic masses
or introduce other distortion.
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8.2.3 The authentic reference spectrum is obtained under OFTPP tuning
conditions (Sect. 8.1 and Table 1-2) to normalize it to spectra from other
instruments.
8.2.4 The spectrum is edited for entry in the library by saving the
five most intense mass spectral peaks and all other mass spectral peaks
greater than 10 percent of the base peak. This edited spectrum is stored
for reverse search and for compound confirmation.
8.3 Polar Compound Detection - demonstrate that unlabeled pentachlorophenol
and benzidine are detectable at the 10 ug/mL level (per all criteria in
Sect. 10.4). The 10 ug/mL calibration standard (Sect. 6.8.1) can be used to
demonstrate this performance.
8.4 Calibration with the Isotope dilution technique - the isotope dilution
technique is used when labeled compounds are available and interferences do
not preclude Its use. If either of these conditions precludes isotope
dilution, the internal standard method, .(Sect. 8.5) is used and noted as such
in the report.
8.4.1 A calibration curve encompassing the concentration range is
prepared for each compound determined. The relative response (pollutant to
labeled) versus concentration 1n standard solutions is plotted or computed
using a linear regression. The example 1n Figure 1-1 shows a calibration
curve for an unlabeled compound and its labeled analog. Also shown are the
+10 percent error limits (dotted lines). Relative Response (RR) is determined
according to the procedures described below. A minimum of five data points
are employed for calibration.
8.4.2 The relative response of a pollutant to its labeled analog is
determined from isotope ratio values computed from acquired data. Three
isotope ratios are used in this process:
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R = the isotooe ratio measured for the pure pollutant
3 = the isotooe ratio measured for the labeled comoound
P = the isotooe ratio of an analytical mixture of pollutant and labeled
compounds.
The m/zs are selected such that Rx>Ry. If &, is not between 2 R and 0.5RX,
the method does not apply and the sample is analyzed by the internal standard
method (Sect. 3.5).
3.4.3 Capillary columns usually separate the pollutant-labeled pair,
with the labeled compound eluting first (Figure 1-2). For this case,
R = (area m./z)/l
at the retention time of the pollutant (RT2) and
R = l/(area m-/z)
y 2
at the retention time of the labeled compound (RTj). Also,
R = [area m./z (at RT-)]/[area rru/z (at RT^)]
as measured in the mixture of the pollutant and labeled compounds (Figure 1-2),
and RR » R .
8.4.4 Special precautions are taken when the pollutant and its labeled
analog are not chromatographlcally separated and have overlapping spectra,
or when another labeled compound with interfering spectral masses overlaps
the pollutant (which can occur with isomeric compounds). In such cases, it
is necessary to determine the respective contributions of the pollutant and
labeled compounds to the respective EICP areas. If the peaks are separated
well enough to permit the data system or operator to remove the contributions
of the compounds to each other, the equations in Sect. 8.4.3 apply. This
usually occurs when the height of the valley between the two GC peaks at the
same m/z is less than 10 percent of the height of the shorter of the two
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peaks. If significant chromatographic and spectral overlap occur, RR is
omputed using the following equation:
where R is measured as shown in Figure I-3A, R is measured as shown in
x y
Figure I-3B, and R is measured as shown in Figure I-3C. For the exairple,
R * 46100/4780 * 9.644
R - 2650/43600 = 0.0608
R 49200/48300 - 1.019
m
8.4.5 To calibrate the analytical system by isotope dilution, analyze
a 1.0 uL aliquot of each of the GC/HS calibration standards (Sect. 6.8.1}
using the procedure in Sect. 10.2. Compute the RR at each concentration.
8.4.6 Linearity - if the ratio of relative response to concentration
for any compound is constant (less than 20 percent coefficient of variation]
over the five-point calibration range, an averaged relative response/concen-
tration ratio may be used for that compound; otherwise, the complete cal ibration
curve for that compound should be used over the five-point calibration
range.
8.5 Calibration by Internal Standard - used when criteria for isotope
dilution (Sect. 8.4) cannot be met. The internal standard used for both
acid and base/neutral analyses is 2,2'-d1fluorob1phenyl. The internal
standard for pesticide and PCB analysis by GC/ECO is decaf luorobenzophenone.
The internal standard method 1s used to measure labeled compounds for intra-
laboratory statistics (Sect. 9.5.1).
8.5.1 Response factors - calibration requires the determination of
response factors (RF) which are defined by the following equation:
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= (As
A = the area of the target peak in the daily standard
A.s = the area of the internal standard peak
C. = the concentration of the internal standard (ug/ml)
C - the concentration of the compound in the daily standard (ug/mL).
8.5.1.1 The response factor is determined over the range of
concentrations described in Sect. 6.8.1, 6.8.2, 6.8.3, and 6.8.4. The
amount of internal standard added to each extract is the same so that C-s
remains constant. The RF is plotted versus concentration for each compound
(or class of compounds in the case of toxaphene) in the standard (Cs) to
produce a calibration curve.
8.5.1.2 Linearity - if the response factor (RF) for any compound
is constant (less than 35 percent coefficient of variation) over the calibration
range, an averaged response factor may be used for that compound; otherwise,
the complete calibration curve for that compound should be used over the
range.
8.6 Combined Calibration - by using calibration solutions (Sect. 6.8.1)
containing the pollutants, labeled compounds, and the internal standard, a
single set of analyses can be used to produce calibration curves for the
isotope dilution and internal standard methods. These curves are verified
each shift by analyzing the 10 ug/mL calibration standard (Sect. 9.9.1).
Pesticide and PCS calibration standards must be analyzed separately by
GC/ECD. Recalibration 1s required only if calibration verification (Sect.
9.9.1.3) criteria cannot be met.
8.7 Ongoing Calibration (see Sect. 9.9)
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9.0 QUALITY ASSURANCE/QUALITY CONTROL [For further guidance, see Quality
Assurance/Quality Control (QA/QC) for 301(h) Monitoring Programs: Guidance
on Field and Laboratory Methods (Tetra Tech 1986).]
9.1 Laboratories that use this method are required to operate formal quality
assurance programs. The requirements of the programs are an initial demon-
stration of laboratory capability, analyses of replicates and matrix spikes
used to evaluate and document data quality, and analysis of standards and
blanks used to test continued performance.
9.2 Initial Demonstration of GC/MS Capability - the analyst should make an
initial demonstration of the ability to generate acceptable accuracy and
precision with the GC/MS component of this method. This ability is established
as described in reference I.
9.3 The analyst is permitted to modify this method to improve separations or
lower the costs of measurements, provided that the new method is demonstrated
to perform comparably to the present method (I.e., with comparable spike
recoveries and precision).
9.4 Blanks method blanks should be analyzed by GC/MS and GC/ECD to demon-
strate freedom from contamination.
9.4.1 At least one method blank must be included with each batch of
samples; method blanks must constitute at least 5 percent of all samples
analyzed.
9.4.2 Method blank concentrations of compounds of interest and of
potentially interfering compounds should be less than 5 percent of the
expected values for the corresponding analytes in samples and below the LLD,
if possible. It 1s recommended that if blank concentrations of compounds of
interest (except phthalates) are greater than 30 percent of the corresponding
analyte concentrations in samples, sample analysis should be halted until
the contamination source is eliminated.
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9.5 Spiked samoles are required to assess method performance on the sample
matrix.
9.5.1 cor samples analyzed by the isotooe dilution technique, the
percent recovery (?) of labeled comoounds can be computed by the internal
standard method (Sect. 8.5) and serves as an indication of analytical accuracy
(but not necessarily of extraction efficiency). After the analysis of five
samples, comnute the average percent recovery (P) and the standard deviation
of the percent recovery (s ) for the labeled compounds only. Express the
accuracy assessment as a percent recovery interval from P - 2s to P * 2s .
For example, if P = 90 percent and s = 10 percent, the accuracy interval is
expressed as 70-110 percent. Update the accuracy assessment for each compound
on a regular basis (e.g., after each 5-10 new accuracy measurements).
9.5.2 Laboratories unable to use isotope dilution must analyze matrix
spikes of pollutants (other than pesticides and PCBs) at a frequency of
5 percent of all samples analyzed or once with each sample set, whichever is
more frequent. Compounds should be added at concentrations 1 to 5 times
those in the sample.
9.5.3 All laboratories are required to spike samples with PCBs or
pesticides at a frequency of 5 percent of all samples analyzed or once per
sample set, whichever is more frequent. The spike can be a standard pesticide
mixture or an Aroc lor mixture, wh ichever is cons idered to be more representative
of the sample. The mixture should be added at one to five times the sample
concentration of these compounds.
9.6 Replicates (i.e., analyses of at least two separate aliquots from a
sediment homogenate) must be analyzed by GC/MS and GC/ECO to monitor the
precision of laboratory analyses. At a minimum, 5 percent of the analyses
should be laboratory replicates. A triplicate analysis should be performed
with each sample batch of over 40 samples.
9.7 The laboratory should maintain records to define the quality of data
that are generated. These records include documentation of blanks and
reports of labeled compound recovery (Sect. 9.5.1), if the latter is applicable.
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9.8 The laboratory should, on an ongoing basis, demonstrate through calibration
verification that the analysis system is in control (Sect. 9.9.1.3).
9.9 System and Laboratory Performance
9.9.1 At the beginning and end of each 12-h shift during which analyses
are performed, GC/MS system performance and calibration are verified for all
pollutants and labeled compounds. For these tests, analysis of the 10 ug/ml
calibration standards (Sect. 6.8.1) should be used to verify all performance
criteria. The GC/ECD performance is checked at the beginning and end of
each shift or at least every 6 h by analyses of 250 and 100 ug/mL solutions
of the PCS and pesticide standards (Sect. 6.8.2.2 and 6.8.3).
9.9.1.1 Retention times - the absolute GC/MS retention time of
2,2'-difluorobiphenyl should be within the range of 1078 to 1248 sec. The
absolute GC/ECO retention time of 4§4'-DOT should be within the range of
1050 and 1200 sec.
9.9.1.2 GC resolution for GC/MS analysis - the valley height
between anthracene and phenanthrene at m/z 178 (or the analogs at m/z 188)
should not exceed 10 percent of the taller of the two peaks.
GC resolution for GC/ECO analysis - the valley height between two peaks
should not exceed 25 percent of the taller of the two peaks for the following
pairs: beta- and delta-HCH, dieldrln and 4,4'-OOE, 4,4'-000 and endrin
aldehyde, and endosulfan sulfate and 4,4'-ODT.
9.9.1.3 Ongoing calibration verification compute the concentration
of each pollutant (Table 1-1) by isotope dilution (Sect. 8.4) for those com-
pounds that have labeled analogs. Compute the concentration of each pollutant
that has no labeled analog with the nearest elutlng labeled standard.
Compute the concentration of the labeled compounds by the internal standard
method. Also compute individual pesticide concentrations and total PCB and
toxaphene concentrations by the internal standard method (GC/ECD). These
1-23
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concentrations are computed based on the calibration data determined in
Sect. 3. ''reparations of new cal ibration standards or revisions of cal ibration
curves are reauired if ooserved resoonses of analytes vary from predicted
responses by more t^an *_20 percent. Samoles and blanks may be run only
after calibration cerforTiance meets this control limit.
9.9.1.4 Multiple peaks - each comoound injected should give a
single, distinct GC peak.
9.9.2 DFTPP spectrum validity - inject 1 uL of the DFTPP solution
(Sect. 6.8.5) either separately or within a few seconds of injection of the
standard (Sect. 9.9.1) analyzed at the beginning of each shift. The criteria
in Table 1-2 should be met.
10.0 PROCEDURE (see Figure 1-4)
10.1 Samole Extraction and Concentration
10.1.1 Homogenize samoles prior to analysis to ensure that representative
aliquots are taken. Mix any water that has separated from the sediment back
into the sample. Remove and make note of nonrepresentative material (e.g.,
twigs, leaves, shells, rocks, and any material larger than 1/4 in). It is
recommended that any removal of material be performed in the field by the
sampling personnel if sampling conditions permit (e.g., if contamination can
be avoided on board ship).
10.1.2 Add a 100-g (wet wt) sediment aliquot (weighed to the nearest
0.1 g) to a precleaned Soxhlet thimble for extraction. Spike with 10 ug of
each neutral stable isotope-labled compound and 15 ug of each acid stable
isotope-labeled compound if using the isotope dilution technique. Use a
separate aliquot for a dry-wt to wet-wt ratio determination.
10.1.2.1 To determine the sediment dry weight, transfer an aliquot
of approximately 3 g (weighed to the nearest 0.1 g) to a preweighed dish.
Allow the sample to dry in a 105° C oven overnight and determine the solid
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residue weight (to the nearest 0.1 g). Calculate and report the percent
.olids (T J as:
TS = [dry residue wt (g)]/[wet sample wt (g)] x 1001
10.1.3 Soxhlet-extract the sediment with CH2Cl2/MeOH (2/1) for 24 h
(60-90 cycles). Before extraction, fill the thimble (containing sediment)
with pure MeOH and stir the sediment-methanol mixture to enhance removal of
water. Cover the sample with a thin layer of solvent-cleaned glass wool.
Add CH2Cl2/MeOH to the 250-mL flask such that the combined total of solvent
in the thimble and flask is at least 210 ml. Stir the sample in the thimble
at least twice (after the second cycle and after approximately 12 h) to
prevent solvent channeling. (The glass wool should be removed during stirring
and then replaced.) The Soxhlet apparatus should be wrapped up to the
condenser with aluminum foil to ensure even heating during cycling.
10.1.4 Re-extraction of sediments at a pH below the pK s (i.e., the
Q
negative logarithm of an acid dissociation constant) of target acidic compounds
may enhance extraction recoveries for these compounds. For example, an
extraction pH of 2 would be well below the pK s of the acidic analytes
a
(e.g., the pK of- pentachlorophenol is approximately 4.7). This additional
a
extraction step is optional and has not been tested with this 301(h) protocol.
It is not acceptable to acidify the extract before the initial Soxhlet
extraction because acidification at the temperatures required for the Soxhlet
extraction can degrade some potential analytes.
10.1.5 Alternative methods of sediment extraction may be used if
evidence of acceptable performance (i.e., equivalent or better apparent
extraction efficiency) 1s provided.
10.1.6 Liquid-Liquid Extraction
10.1.6.1 After Soxhlet extraction, transfer the extract to a 500-
itl separatory funnel. Rinse the Soxhlet flask twice with clean extraction
>olvent and add this rinse to the extract 1n the separatory funnel. Wash
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the solvent extract with approximately 100 ml of pH 2, SO* Na2$04 saturated
organic-free *ater. The oH of the water should be adjusted with solvent-
cleaned HC1. Oxidizing acids (e.g., H.SOJ must not be used because they
can cause losses of target compounds. Collect and store the CH^C^ layer.
The Duroase of wasmng the extract «nth an acidic aqueous solution is to
remove water and nethanol from the CH.C1- and to enhance the partitioning of
acidic organic comoounds into the CH-CU ^yer. Re-extract the acidic
aqueous phase twice with 60 ml of dean CH.CU and add both extracts to the
initial CH-Cl, fraction.
10.1.6.2 Adjust the pH of the aqueous phase to >_12 with solvent-
cleaned, 6 N KOH. Back-extract the base compounds three times with 60 ml
CH-CU. The pH adjustment to alkaline conditions enhances the partitioning
of basic compounds into the CH.CU layer. Combine all CH2C12 layers from
Sect. 10.1.6.1 and 10.1.6.2.
10.1.6.3 Formations of emulsions or precipitates during liquid-
liquid extraction should be noted and considered when reviewing results.
The addition of Na^SO^ may reduce emulsions. However, if the emulsion
interface between layers is more than one-third the volume of the solvent
layer, the analyst must emoloy mechanical techniques to complete the phase
separation. The optimum technique depends on the sample and may include
stirring, filtration of the emulsion through pre-deaned glass wool, centri-
fugation, or other physical methods (reference 2).
10.1.7 Dry the total combined solvent extract by pouring it through an
anhydrous Na-SO. drying column (approximately 30 cm x 2 cm). Use approxi-
mately 30 mL of CH-Cl- to rinse the drying column and combine this with the
dried extract. Collect the extract in a Kuderna-Oanish (K-0) 500-mL evaporation
flask containing I to 2 clean boiling chips.
10.1.8 Attach a 3-ball macro Snyder column to the K-0 evaporation
flask and concentrate the extract on an 80° C water bath. Pre-wet the
Snyder column by adding about 1 ml of CH2C12 to the top of the column.
Place the K-D apparatus on the hot water bath so that the concentrator tube
1-26
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is partially immersed in the hot water and the entire lower rounded surface
af the flask is bathed with hot vapor. At the proper rate of distillation,
the balls of the column will actively chatter but the chambers will not
flood with condensed solvent. When the apparent volume reaches 5 ml, remove
the K-D apparatus from the water bath and rinse the flask with 3 ml CH^C^
draining into the concentrator tube. Reduce the contents of the concentrator
tube to 3 ml using a stream of purified NZ gas, never allowing the extract
to go to dryness.
10.1.9 Alternative methods of extract concentration may be used if
evidence of acceptable performance [I.e., retention of more volatile compounds
(e.g., naphthalene) comparable to that of K-0 concentration] is provided.
10.1.11 Elemental sulfur removal - Mercury cleanup is required to
remove elemental sulfur, which interferes with GC/MS and GC/ECO analyses,
from the extract. Some losses of benzidine and endrin aldehyde may occur in
this step.
10.1.11.1 Transfer the extract to a clean, screw-capped test tube
and shake vigorously with approximately 0.5 mL precleaned mercury for at
least 4 h. Filter the desulfurized extract to remove metallic mercury and
its salts. Shake the test tube and mercury with 1-2 mL of CH2C12, rinse the
filter with this solvent, and combine the rinsing with the desulfurized
extract. This process may have to be repeated for samples with high sulfur
content.
Alternatively, mercury treatment could be performed more quickly by
vigorous agitation for several minutes on a vortex mixer. A potential
problem with this technique is that the mercury could become so finely
dispersed that 1t would pass through the filter. Vortex mixers may be used
only if appropriate method performance can be demonstrated (e.g., mercury
should not pass through the filter).
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10.1.11.2 Another commonly used method of sulfur removal, elution
through an activated cooper column, nay not be a suitable substitute for
mercury treatment because coooer fay strongly retain some oolar analytes.
10.1.12 Extract cleanup - GPC cleanuo is reauired to seoarate biological
macromolecules from the analytes.
10.1.12.1 GPC setuo and calibration (reference 2).
10.1.12.1.1 Place 70 g of Bio Beads S-X3 in a 400-mL beaker.
Cover the beads with CH.CU. Allow the beads to swell overnight. Transfer
the swelled beads to the column and start pumping solvent through the column,
from bottom to top, at 5 ml/mm. After 1 h, adjust the pressure on the
column to 7-10 psi and pump for an additional 4 h to remove air from the
column. Adjust the column pressure as required to maintain 7-10 psi.
10.1.12.1.2 Calibration of the column - Load 5 mL of the
corn oil solution into sample loop No. 1 and 5 ml of the phthalate-PCP
solution into loop No. 2. Inject the corn oil solution and collect 10 ml
fractions for 36 mm. Determine the corn oil elution pattern by evaporation
of each fraction to dryness followed by a gravimetric determination of the
residue. Analyze the phthalate-PCP fractions by GC/FIO, a UV-spectroohotometer
at 254 nm, or a GC/MS system. Plot the concentration of each component in
each fraction versus total eluant volume. Choose a "dump time" that allows
>_85X removal of the corn oil and ^851 recovery of the phthalate. Select the
"collect time" to extend at least 10 min after the elution of
pentachlorophenol. Wash the column for at least 15 mm between samples.
Typical parameters are: dump time, 30 mm (150 ml); collect time, 36 mm
(180 ml); and wash time, 15 mm (75 ml). The S-X3 Bio Beads column may be
reused for several months, but should be checked by system recalibration for
every 20 extracts loaded onto the GPC.
10.1.12.2 Extract cleanup - Prefilter or load all extracts via
the filter holder to avoid partlculates that might cause system blockage.
Load the extract (approximately 3 ml) onto the GPC column. Do not apply
1-28
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excessive pressure when loading the GPC. Purge the sample loading tubing
horoughly with solvent between extracts. Process the extracts using the
dump, collect, and wash parameters as selected from the calibration and
collect the cleaned extracts in 400-mL beakers.
10.1.12.3 Rerun the phthalate-PCP calibration solution to check
and recalibrate the system once for every 20 extracts loaded onto the GPC.
The recoveries and elution profiles are reported as deliverables.
10.1.13 Transfer the extract to a Kuderna-Oanish (K-0) concentrator
consisting of a 10-mL concentrator tube, a 500-mL evaporation flask, boiling
chips, and a Snyder column. Carefully concentrate the extract to 2.5 ml
using methods previously described and the Nj 9as blowdown technique.
Nitrogen blowdown should be performed at approximately 35° C. A gentle
stream of clean, dry NZ (filtered through a column of activated carbon)
should be used. The inside walls of the tube containing the extract should
be rinsed down with the appropriate solvent several times during concentration.
The extract must not be allowed to go to dryness.
10.1.14 Use a 201 aliquot (500 uL) of the extract for alumina column
cleanup (Sect. 10.1.15) and subsequent GC/ECO analysis for pesticides and
PCBs. Use the remaining 801 (2 ml) for GC/MS analysis. If the sediment
sample appears contaminated with petroleum or was collected from an area of
known or suspected petroleum contamination (e.g., most nearshore environments
near urban or industrial centers) a further extract cleanup is required
prior to GC/MS analysis. SPE column cleanup (Sect. 10.1.16) removes some of
the paraffinic hydrocarbon constituents that contribute to the unresolved
complex mixture (UCH) typically observed in gas chromatograms of petroleum
extracts.
10.1.14.1 Solvent exchange of extract for alumina cleanup (reference
2) - transfer 0.5 mL of the extract to a separate concentrator tube. Add 5
ml of hexane and a boiling chip and mix using a vortex mixer. Attach a
two-ball micro-Snyder column. Pre-wet the Snyder column by adding 0.5 mL of
lexane to the top of the column. Place the K-0 apparatus on a hot water
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oath (80-90° C) so that the concentrator tube is partially immersed in the
not rfater. Adjust the vertical position of the apparatus and the water
temoerature as required to complete trie concentration in 5 to 10 min.
Concentrate the extract to an aooarent volume of approximately 1 TiL. Use N^
slowdown to reduce the /olume to 0.5 ml. Dilute to 1 ml by adding 0.5 ml of
acetone. Proceed with alumina column cleanup.
10.1.15 Alumina column setup and use - The alumina column cleanuo is
required to remove polar interferences prior to GC/ECO analysis of oesticides
and PC3s (reference 2).
10.1.15.1 Add 3 g of activity III neutral alumina to the clean,
5-mL disposable serological pipet (with glass wool dug). Tap the column to
settle the alumina, do not prewet the alumina with solvent.
10.1.15.2 Transfer the 1.0 ml hexane/acetone extract (Sect.
10.1.14.1) to the top of the alumina column with a disposable Pasteur pipet.
Collect the eluate in a 10 ml K-0 concentration tube. Add 1 ml of hexane to
the original extract concentrator tube to rinse it. Transfer these rinsings
to the alumina column. Elute the column with an additional 9 ml of hexane.
Do not allow the column to go dry during the addition and elution of the
'ample.
10.1.15.3 Note that batches of alumina may differ and storage may
alter the water content of deactivated alumina. Thus, column performance
must be checked regularly and for each batch of alumina. PCS and pesticide
standards (e.g., from Sect. 6.8.2.2, 6.8.3) and a suitable model poTar
compound (e.g., trlbromophenol) should be used to determine the appropriate
elution volumes for these pollutants. Recovery of single PCB or pesticide
components should be greater than 855 and the trlbromophenol should not be
detected.
10.1.15.4 Concentrate the eluate to a final volume of 500 uL
using a micro-Snyder column and the N- gas blowdown technique.
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10.1.15.5 Care must be taken to allow the N2 gas to create only a
small dimple on the surface of the solvent and prevent blowdown to dryness.
Submit extract for GC/ECD analyses.
10.1.16 SPE or reverse phase column cleanup used to reduce or eliminate
the interferences caused by chromatographically unresolvable, nonpolar
petroleum constituents. While this procedure enhances the sensitivity of
GC/MS analyses for many pollutants, it can also result in only partial
recovery of some potential analytes (e.g., dichlorobenzenes, chlorinated
butadienes, certain PCS congeners) from the extract because the interferents
removed in this step partially co-elute with these compounds. These partial
recoveries should be assessed with standards prior to routine use of SPE
columns. Elution volumes may be adjusted to optimize recoveries while
reducing the UCM as much as possible. The use of the isotope dilution
technique will enable correction for losses of compounds with labeled analogs.
10.1.16.1 Exchange the 2 mL extract into MeOH as follows: Use
the N2 blowdown technique to reduce the extract volume to approximately 0.5
ml. Add 3 ml of MeOH to the extract and carefully reduce the volume to I ml
using the N2 gas blowdown technique. Repeat this procedure to ensure that
the extract is adequately exchanged from CH2C12 to MeOH. The presence of
even a small amount of CH2C12 will reduce the polarity differences between
liquid and solid phases that control the chromatographic process and will
thus allow carryover of Interferents into the final extract.
10.1.16.2 Condition the SPE column with three column volumes of
methanol prior to applying the extract. It 1s also recommended that 10 ml
of 0.51 HC1 (pH 2-3) be eluted through the column after methanol elution.
Although not discussed by the manufacturer, this cleanup step has been
reported to be effective at reducing residual contamination (R.J. Ozretich
and W.P. Schroeder. Submitted for publication. "Determination of Priority
Organic Pollutants in Marine Sediment, Tissue, and Reference Materials
Utilizing Bonded-phase Sorbents". Analytical Chemistry).
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10.1.16.3 Place a K-Q concentrator tube beneath the column onor
to applying the extract, ^ooly and draw the l-mL extract (in MeOH) to the
top of the column, eollowed by a 0.5-mL rinse of MeOH from the concentrator
tube. Elute additional MeOH until a total of 7 nL is collected in the K-0
concentrator tjbe.
10.1.16.4 Exchange and concentrate the eluate to a final volume
of 0.4 ill (in CH.C1-). Care must be taken to avoid the loss of volatile
pollutants during concentration. Use CH-CU and the N^ gas blowdown technique
to adjust the final volume.
Metfianol is not an ideal solvent for solvent exchange because of its
low volatility relative to many organic analytes. It is possible that
acetone (because of its polarity and relatively high volatility) would be a
favorable substitute for methanol for the reverse phase column cleanup
step. However, acetone has not been tested and no validation data are
available.
10.1.16.5 Submit the extract for GC/MS analysis.
10.2 GC/MS Analysis
10.2.1 Establish the following operating conditions for the GC (Table
1-1): 5 mm at 30° C; 30-280° C at 8° C/min; isothermal at 280° C until
benzo(g,h,i)perylene elutes. Make certain that the concentrated extract or
standard is at room temperature and make note of any precipitate that does
not redissolve.
10.2.2 Add 5 uL of the GC/MS internal standard solution to the 400 uL
extract to yield a 10 ug spike. Add the solution immediately prior to
injection to minimize the possibility of loss by evaporation, adsorption, or
reaction. Mix thoroughly.
10.2.2.1 It is advised that a late eluting internal injection
standard (e.g., 5-alpha-cholestane) be used in addition to DFB. The use of
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early eluting (DFB) and late eluting injection standards will allow the
jnalyst to detect and compensate for problems in the GC injection port related
to differential loading of analytes onto the GC column.
10.2.3 Inject 1.0-1.5 uL and start the GC column initial isothermal
hold. Start MS data collection after the solvent peak elutes. Stop data
collection after the benzo(g,h,iJperylene elutes.
10.2.3.1 Dilution and re-injection are required for samples that
exceed the upper concentration limit of the calibration standards. Data for
compounds within the calibration range should be acquired in the initial
run. Data for compounds exceeding the calibration range should be acquired
after dilution. Respike the sample with labeled compounds and assume 100
percent recovery. This assumption is not unreasonable considering the high
concentrations of native compounds involved when dilution is necessary.
10.3 GC/ECO Analysis
10.3.1 The recommended GC conditions are modified from those specified
in reference 2:
Helium carrier gas: 4 ml/min at 280° C and 25 psi
Septum purge: 15 mL/m1n
Split vent: none
Initial temperature: 60° C, Initial hold 2 min
Program at 25° C/m1n to 160° C
Program at 5° C/m1n from 160° C
Final temperature: 270° C;
hold until decachlorobiphenyl elutes
Injection port temperature: 225° C
10.3.2 Add 10 uL of the GC/ECO internal standard solution to the 500
uL extract to yield a 25 ng spike. Add the solution immediately prior to
injection to minimize the possibility of loss by evaporation, adsorption, or
eaction. Mix thoroughly. Inject 1.0-1.5 uL.
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10.3.3 Dilution and r»-in;ection are reauired for samples that exceed
the upper concentration limit of the calibration standards. Data for compounds
within the calibration range should 5e retained from the initial run. Data
for compounds exceeding the cal i brat ion range should be acauired after dilution.
Column overloading can result in abnormal peak shape, which can reduce
the accuracy of quantification. It may also result in a marked increase in
the retention time of the peak maximum, which can displace an analyte from
the retention time window established with standards at lower concentra-
tions.
10.4 Qualitative Determination
10.4.1 Qualitative determination is accomplished by comparison of data
from analysis of a sample or blank with data from analysis of the shift
standard (Sect. 9.9.1) and, for GC/MS analyses, with data stored in the
spectral libraries (Sect. 8.2.4). Identification is confirmed when spectra
and retention times agree per the criteria below.
10.4.2 Labeled compounds and pollutants having no labeled analog:
10.4.2.1 The-signals for all characteristic masses stored in the
spectral library (Sect. 8.2.4) should be present and should maximize within
the same two consecutive scans.
10.4.2.2 Either 1) the background corrected extracted ion current
profile (EICP) areas, or 2) the corrected relative intensities of the mass
spectral peaks at the GC peak maximum should agree within a factor of two
(0.5 to 2 times) for all masses stored in the spectral library.
10.4.2.3 The retention time difference between an analyte and the
nearest eluting internal standard during sample analysis should be within +5
scans or +_5 sec (whichever is greater) of this difference in the shift standard
(Sect. 9.9.1).
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10.5 Pollutants Having a Labeled Analog:
10.5.1 The signals for all characteristic masses stored in the spectral
library (Sect. 8.2.4) should be present and should maximize within the same
two consecutive scans.
10.5.2 Either 1) the background corrected EICP areas, or 2) the corrected
relative intensities of the mass spectral peaks at the GC peak maximum should
agree within a factor of two for all masses stored in the spectral library.
10.5.3 The retention time difference between the pollutant and its
labeled analog should agree within +2 scans or +2 sec (whichever is greater)
of this difference in the shift standard (Sect. 9.9.1).
10.6 If an experimental mass spectrum contains masses that are not present
in the reference mass spectrum, an experienced spectrometrist is to determine
the presence or absence of the compound.
10.7 Chlorinated Pesticides and PC8s
10.7.1 Single component chlorinated pesticides are tentatively identified
by comparison of sample peak relative retention times to those of authentic
standards (Sect. 6.8.3). Three times the standard deviation of .relative
retention times established from calibration standards (Sect. 6.8.3) can be
used to calculate relative retention time window boundaries. Confirm the
identities of pesticides by comparing the relative retention times of sample
and standard peaks on another column phase (e.g., 86Xdimethyl[141]-cyanopropyl
phenyl polysiloxane or J&U 08-1701). Confirmation by GC/MS is required when
concentrations are sufficient.
10.7.2 Peaks of multi-component mixtures (PCBs and toxaphene) are
tentatively identified in samples by comparison of relative retention times
to those of authentic standards (Sect. 6.8.2.2 and 6.8.4). Three times the
standard deviation of relative retention times established from standards
1-35
-------
can be used to calculate relative retention time window boundaries. Choose
as many peaks as possible *mle avoiding those with potential interferences
(e.g., ?CBs co-eluting with DOT and DOE isomers). Label on all sample chroma-
tograms the peaks identified as PC3 and toxaphene congeners. All GC/ECD
chromatograms are oart of the deliverables. Interpretation of chromatograms
requires the attention of an experienced analyst. Chromatograms of individual
Aroclors (e.g., 1242, 1254, 1260 in three separate standards) may facilitate
interpretation. Confirm the identities of all selected congeners by injection
on an alternative column phase (e.g., J&W 08-1701). Confirmation by GC/MS
is required if concentrations are sufficient.
10.8 Tentatively Identified Compounds (GC/MS Analysis) - The ten non-target
peaks of greatest area in the RIC (reconstructed ion chromatogram) should be
identified and quantified, if possible.
10.8.1 Guidelines for making tentative identification (reference 2):
1) Tentative identifications should be based on a forward search
of the EPA/NIH mass spectral library. Sample spectra should
be visually compared with the most similar library match.
2) Relative intensities of major ions in the reference spectrum
(ions greater than 10 percent of the most abundant ion) should
be present in the sample spectrum.
3) The relative intensities of the major ions should agree within
^_20 percent. (Example: For an ion with an abundance of 50
percent in the standard spectra, the corresponding sample ion
abundance must be between 30 and 70 percent.)
4) Molecular ions present in reference spectrum should be present
in sample spectrum.
1-36
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5) Ions present in the reference spectrum but not In the sample
spectrum should be reviewed for possible subtraction from the
sample spectrum because of background contamination or co-eluting
compounds. Data system library reduction programs can sometimes
create these discrepancies.
10.8.1.1 If, in the opinion of the mass spectral specialist, no
valid tentative identification can be made, the compound should be reported
as unknown. The mass spectral specialist should give additional classification
of the unknown compound if possible (e.g., unknown phthalate, unknown hydro-
carbon, unknown aromatic compound, unknown chlorinated compound). If probable
molecular weights can be distinguished, include them.
10.8.2 Tentative quantification - quantification of TIOs will be based
on the internal standard technique and an assumed response factor of one (in
the absence of data from authentic standards). The uncertain nature of this
quantification should be clearly noted in the data report.
11.0 QUANTITATIVE DETERMINATION (CALCULATIONS)
11.1 Isotope Dilution Technique - by adding a known amount of a labeled
compound to every sample prior to extraction, correction for losses of the
pollutant during the analysis can be made because the pollutant and its
labeled analog exhibit similar behavior during extraction, concentration,
and gas chromatography. Note that pollutants and their labeled analogs are
not always retained Identically by complex matrices, so their behavior during
the extraction step may differ. It 1s not the intention of this technique
to account for matrix recovery efficiency, only for subsequent analytical
recovery efficiency.
11.1.1 Relative response (RR) values for sample mixtures are used in
conjunction with calibration curves described 1n Sect. 8.4 to determine
concentrations directly, so long as labeled compound spiking levels are
constant.
1-37
-------
11.1.2 Specifically, tie concentration, C(in ug/kg), can be determined
as:
(ug/' (Ax x Z1s x 1.25 x 103)/(S x A-s x RF)
where:
A = the area at the characteristic mass for the compound in the sample
A
A. = the area of the characteristic mass for the internal standard
Z. = the absolute amount, in ug, of the GC/MS internal standard added
to the final extract prior to instrumental analysis
S = sample dry weight (g) that was extracted.
1-38
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This technique is not preferred and is unnecessary if the nearest eluting,
most chemically similar labeled compound is used as a recovery standard for
pollutants without available labeled analogs.
11.2.1.1 The stable isotope labeled compound recovery, X, is
determined and reported for each sample in the following manner:
X » [C(ug/kg)/CA(ug/kg]] x 100*
where C and C. are as defined in Sections 11.2.1 and 11.1.2, respectively.
11.2.2 GC/ECO internal standard method
11.2.2.1 Pesticides - compute the concentration in the sample, C
(in ug/kg), using the response factor (RF, determined from calibration data)
and the following equation:
C(ug/kg) - (A^ x Z1$ x 5 x 103)/(S x A1s x RF)
where:
A * the area of the integrated GC peak for the compound in the sample
(A represents the summation of areas for a group of GC peaks
if toxaphene is being quantified)
A. ' the area of the integrated GC peak for the internal standard
Z1s 3 the absolute amount, in ug, of the GC/ECO internal standard
added to the final extract prior to Instrumental analysis
S * the sample dry weight (g) that was extracted.
11.2.2.2 PCBs - accurate PCB quantification is difficult to
achieve in routine full-scan analyses. It has been common practice to quantify
PCBs with packed-column GC/ECO by comparing several selected peaks in samples
to corresponding peaks in commercial Aroclor formulations that most closely
resemble the sample. Shortcomings of this technique have been described
elsewhere (e.g., references 8 and 9). The critical difficulties with this
1-39
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procedure relate to two factors: (1) environmental PCS assemblages often
differ considerably from comercial Aroclor mixtures because of the variable
properties of PC8 congeners (e.g., aqueous solubility, volatility, suscepti-
bility to biodegradation) and (2) the ECD has a markedly variable resoonse
to the 209 PC3 congeners depending on the number and position of chlorine
atoms on the biphenyl nucleus (e.g., reference 10).
It has been suggested that "the least systematic error [in PCB quantifi-
cation] -vi 11 be given by the summation of all or at least nearly all areas
of PC3 peaks corrected by their individual ECO-response factor and their
biphenyl content" (reference 11). Another alternative is to use GC/MS instead
of GC/ECD. However, GC/MS analysis is relatively insensitive unless selected
ion monitoring (SIM) is used, which can involve considerable effort and expense.
The Quantification technique recommended in this 301(h) document relies
on high resolution (capillary column) GC/ECD and a determination of response
factors for resolvable PCB peaks (as suggested in the previous paragraph).
The technique is modified from the Webb and McCall technique (reference 12),
which has been widely used for packed-column PCB quantification. Briefly,
the resolved peaks in a PCB standard (Sect. 6.8.2.1, 6.8.2.2) are quantified
by GC/MS and GC/ECD. The GC/MS results are used to correct for the variability
of ECD response. Samples are analyzed and quantified by GC/ECD. Total PCBs
are calculated as the sum of all resolved, response factor-corrected PCS peaks.
11.2.2.2.1 GC/MS analysis of PCB standard - each resolvable
peak in a PCB calibration standard (Sect. 6.8.2.1) is quantified by GC/MS,
which can identify the chlorine content of biphenyls in each peak and can
quantify PCBs based on their chlorine content. This quantification does not
require that the analyst know the exact identity of the congeners consti-
tuting a peak, only the chlorine content must be determined.
Another GC/MS calibration standard is necessary to perform this quanti-
fication. An MS response factor standard consisting of representatives of
all the congener groups (mono- through deca-chloroblphenyl) is necessary to
convert areas of peaks in ion plots to the appropriate masses of chloro-
1-40
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biphenyls. A standard solution should be made with approximately 10 ng/uL
of each of the following congeners (see reference 13 for an explanation of
these choices):
2
2.3
2,4,5
2,2',4,6
2,2r,3.4,5'
2,2-,4,4-.5,6'
2.2',3.4',5.6,6'
2,2',3.3-,4,5',6,6-
2,2',3,3',4,4',5,5',6,6' - (used for nona- and deca-congeners).
These congeners are available from Ultra Scientific, Inc. (Hope, RI) except
the heptachloro-congener, which is available from Wellington Environmental
Consultants, Inc. (Guelph, Ontario, Canada). The primary quantification
ions used for mono- through deca-chlorobiphenyl are: 138, 222, 256, 292,
326, 360, 394, 430, 464, and 498. The spectrum for each peak should be
manually confirmed at least once for the Aroclor standard.
Relative amounts of co-elut1ng congeners of different chlorine content
in a given peak in the standard can be determined during GC/MS analysis.
Co-elution can be accounted for with appropriate response factors. For
example, if a peak is composed of tetrachloro- and pentachloro-isomers as
determined by ion plots of m/z 292 and 326, the 2,2',4,6 response factor is
used for the m/z 292 area and the 2,2',3,4,5' response factor for the m/z
326 area. Care must be taken to ensure that M-70 Ions are not interpreted
as M+ ions if congeners differing by two chlorine atoms co-elute.
11.2.2.2.2 GC/ECD analysis of PCB standard (Sect. 6.8.2.2) -
each resolvable peak in the PCB calibration standard is .quantified by GC/ECO
according to the internal standard technique (Sect. 8.5). The GC/ECO analysis
1s performed with the same GC column phase and temperature program used for
GC/MS analysis of the standard. An ECO response factor is established for
1-41
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GC/MS analysis of the standard. An ECO response factor (RF) is established
for e*cn oeak based on the GC/MS analysis of the PCS standard using the
eauat'on defined in Section 3.5.1
cs = the mean concentration of the peak in the PC3 standard as
determined by GC/MS (determined with at least three replicate
analyses).
11.2.2.2.3 GC/ECO .PC3 quantification in samples - total PCBs
are calculated as the sum of all PCS peaks identified in a sample
(Sect. 10.7.2):
C (ug/kg, dry wt) *
£ [(Ax x Zis x 5 x 500)/(S x Ais x RF)]]
where:
i » each identified PCB peak, with n total peaks
Ax = area of the integrated GC peak for the compound in the sample
AIS = area of the integrated GC peak for the internal standard
I- » the absolute amount, in ug, of the GC/ECD internal standard added
to the final extract prior to instrumental analysis
S » the sample dry weight (g) extracted
RF = calibration response factor (Sect. 8.5.1).
1-42
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11.2.2.2.4 This quantification method involves two noteworthy
imitations:
(1) Interferences can be a significant problem in ECO analyses.
PCS peaks co-eluting with interferents may be neglected or
quantified, in either case resulting in a decrease in accuracy.
It 1s essential that experienced analysts evaluate chromatograms
to determine the presence of suspected interferents.
Interferents suspected of overwhelming PCS peaks should be
neglected. The mercury and alumina column cleanup steps are
designed to preclude major Interferences. High resolution
capillary columns also reduce the potential for co-eluting
interferences.
(2) When two or more congeners have identical retention times on
a given column phase, it 1s Impossible to determine their
relative concentrations in a peak when using GC/ECO. Thus,
it is not possible to determine whether sample peaks are
composed of the same relative combination of congeners as
corresponding standard peaks. Thus, the response factor for
a peak may be different during calibration and sample analysis.
The potential error in assigning appropriate reponse factors
has been minimized in this technique by the use of high
resolution capillary columns.
11.2.2.2.5 Alternative techniques of detection [e.g., Hall
electrolytic conductivity detector (HECO) or MS (with selected 1on monitoring)]
can provide comparable or superior PCB Identification and quantification
relative to ECO (e.g.. references 13 and 14) and are acceptable substitutes
for ECO detection. Although ECO Is widely available and 1s more sensitive
for PCBs than HECO or MS, HECO has a linear response to chlorine content and
1s more specific to chlorinated compounds, and MS offers more definitive
compound identification than ECO.
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11.2.2.3 Quantify PCBs by summing the resoonse factor-corrected
areas of the characteristic PCB peaks identified in Sect. 10.7.2. Report
the results as total PCBs.
11.3 Seoort results for all pollutants and labeled compounds found in all
standards and samples, in ug/kg, to two significant figures. Note in the
report all compounds that have not been recovery corrected. Report results
for blanks as total ng/sample.
12.0 PRECISION AND ACCURACY
Laboratory intercomoarison studies of the precision and accuracy attainable
with this technique will be required. Available precision data for single
laboratory analyses of a series of contaminated estuarine sediments are
given in Table 1-3 (data from reference 15). In these analyses, spiking
levels for base/neutral compounds were lower than those recommended in this
protocol (i.e., 5 ug spike instead of 10 ug), and methanol was not added
directly to the extraction thimble as now recommended in Sect. 10. PC8
results were generated by packed column analysis and comparison of sample
chromatograms to Aroclor standards. The accuracy of these PCB analyses was
not assessed. Validation data using the PCS quantification procedure in
this 301(h) document has not yet been generated.
In a recent method test using the same procedure, replicate blanks were
spiked with known amounts of labeled and unlabeled compounds. The blanks
were then taken through the entire procedure and the amounts of the unlabeled
compounds were calculated using the isotope dilution technique (i.e., the
calculated amounts of the unlabeled compounds were adjusted for the recovery
of the labeled compounds). The ratio of the calculated amount of the unlabeled
compounds relative to their actual spiked amount (expressed as percent) is
given in Table 1-4. The precision and accuracy results for blanks (Table
1-4) compare favorably with acceptance criteria in Method 1625 Revision B.
Compounds outside acceptance criteria for either precision or accuracy include:
benzidine, bis(2-chloroisopropy1) ether, bis(Z-chloroethoxy) methane,
hexachlorocyclopentadiene, N-nitrosodiphenylamine, N-nitrosodipropylamine,
1-44
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N-nitrosodimethylamine, and butylbenzyl phthalate. The spike level used in
hese preliminary tests was less than 0.1 that in Method 1625 and the analytical
procedure requires several stages of sample cleanup (none is required in
Method 1625 Revision B).
13.0 REFERENCES
1. "Performance Tests for the Evaluation of Computerized Gas Chromatography/-
Mass Spectrometry Equipment and Laboratories," USEPA, EMSL/Cincinnati, OH
45268, EPA-600/4-80-025 (April 1980).
2. U.S. Environmental Protection Agency. 1984 (revised January, 1985).
U.S. EPA Contract Laboratory Program statement of work for organics
analysis, multi-media, multi-concentration. IFB WA 85-J176, J177,
JUS.
3. Fed. Register, Vol. 49, No. 209, October 26. 1984, pp. 43416-43429.
4. "Carcinogens - Working with Carcinogens," OHEW, PHS, CDC, NIOSH, Publica-
tion 77-206 (Aug 1977).
5. "OSHA Safety and Health Standards, General Industry," OSHA 2206, 29 CFR
1910 (revised Jan 1976).
6. "Safety in Academic Chemistry Laboratories," ACS Publications, Committee
on Chemical Safety, 3rd Edition (1979).
7. Eichelberger, J.W., L.E. Harris, and W.L. Budde, "Reference compound to
calibrate ion abundance measurement in gas chromatography-mass
Spectrometry," Anal. Chem. Vol. 47, 1975, pp. 995-1000.
8. Ouinker, J.C., M.T.J. Hlllebrand, K.H. Palmork, and S. Wilhelmsen, "An
evaluation of existing methods for quantitation of PCBs in environ-
mental samples and suggestions for an improved method based on measurement
of individual components," Bull. Environm. Contain. Toxicol. Vol. 25,
1980, pp. 956-964.
9. Alford-Stevens, A.L., W.L. Budde, and T.A. Bellar, "Interlaboratory
study on determination of PCBs in environmentally contaminated sediments,"
Anal. Chem. Vol. 57, 1985, pp. 2452-2457.
10. Mullln, M.D., C.M. Pochini, S. McCrindle, M. Romkes, S.H. Safe, and
L.M. Safe, "High-resolution PCB analysis: synthesis and chromatographic
properties of all 209 PCB congeners," Environ. Sd. Techno!. Vol. 18,
1984, pp. 468-476.
11. Ballschmlter, K., and M. Zell, "Analysis of PCB by glass capillary .gas
chromatography," Fresenius Z. Anal. Chem. Vol. 302, 1980, pp. 20-31.
1-45
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12. Webb, R.G., and A.C. McCall, "Quantitative PCB standards for electron
capture gas chromatograpny," J. Chromatographic Science, Vol. 11, 1973,
pp. 366-373.
13. Gebhart, J.E., T.L. Hayes, A.L. Alford-Stevens, and W.L. Budde, "Mass
spectrometric determination of polychlorinated biphenyls as isomer
grouos," Anal. Chem. Vol. 57, 1985, pp. 2458-2463.
14. Sonchik, S., 0. Madeleine, P. Macek, and J. Longbottom, "Evaluation of
sample preparation techniques for the analysis of PCBs in oil,"
J. Chromatographic Science, Vol. 22, 1984, pp. 265-271.
15. Tetra Tech, Inc. Commencement Bay nearshore/tideflats remedial investi-
gation. Vol. 1. Final report prepared for the Washington State Department
of Ecology and U.S. Environmental Protection Agency. 1985.
1-46
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(38)
(3C)
M,/Z
M./Z
AREA AT
AREA AT
AOAt>UO fhUM KIMMIHll J
46100
4780
AREA - 43600
AREA - 2650
AREA - 48300
AREA - 49200
AOAP11D
ftirtN(NC( 3
Figure 1-1. Relative response calibration curve.
Figure 1-2. Extracted ion current profiles for chromatographical ly resolved labeled
(m?/z) and unlabeled (HK/Z) pairs.
Figure 1-3. Extracted ion current profiles for (3A) unlabeled compound, (3B) labeled
compound, and (3C) equal mixture of unlabeled and labeled compounds.
-------
SAMPLE
5P'KE 'C
SO TOPS
3CXHLET EXTRACT
WASH EXTRACT
AfPHtZ
»CUE
SLLRlfl REMOVAL IHg)
GPC
CONCENTRATE TO 2J rr«.
8O%
20%
EXCHANGE NTO kMOH
SP COLUMN CLEANUP
CONCENTRATE AND
EXCHANGE
AOOGC-MS
NTERNALSTANOAflO
INJECT ON
GCMS
PCR ISOTOPE OLUTCN TECHNOUE - RECOMMENOEO BUT NOT REOUIREO
E
-------
TABLE L-l. GAS CHROMATOGRAPHY OF EXTRACTABLE COMPOUNDS
Compound
Instrumental
Quantitation Senji-
Hetention Time (Primary) twity (ng)
Sec Relative^ m/z SC/NS
DOES
CASRN
2,2'-difluorpoipneny1 (OFB)
N-nitrosodjnietnylaflMne
phenol-djf1'
pnenol
pis(2-chlon>ethyl)ether-dg
pi$(2-cf»loroetnyljether
I ,3-d1chloroBenzene-d4
1,3-dlchloroDenzene
1 , 4-d i c n 1 orooenzene-d*
1,4-dicnlorooenzene
l,2-dicnlorooenzene-d4
1 ,2-dlcnl orooenzene
Ou(2-cnloronopropyl)ether-di2
tm(2-cnloroisopropy] letter
nexachloroetnane-lJCl2'
hexacnloroetftane
N-mtrosodi-n-prnpylamine
nitrooenzene-d5
nttrooenzene
isopnorone-dg
nopnorone
2, 4-dimetny 1 pfienol -dj
2, 4-d Imetnyl pnenol
Dis(2-cnloroetnoxy)metnane
1.2.4-trichlorooenzene-d3
i.2,4-tMchlorooenzene
naphthalene-tig
naphthalene . -
nexacnlorooutadlene- <
hexacn 1 oroputad iene
n ex ac n I oroc yc 1 open tad 1 en e- ^C 4
nexacnlorocydopcntadiene
2-cnloronapnthalene-d7
2 -chloronapntnal ene
Diphenyl-dio
Oipnenyl
acenapntnylene-dg
acenapnthylene
d imethy 1 ptttna 1 ate- 04
dimetnylpntnalate
2,6-dimtrotolutne-d3
2.6-dinUroto)u»ne
acenaphthene-diQ
acenapnthene
dioenzofuran-dg
dibenzofuran
fluorer e-d^o
f luorene
4-chloropnenylpnenyl ether-ds
4-cnlpropnenylphenyl ether
diethyl phtnalate-d4
dietnyl pfttftalate
2,4-dinitrotoluene-d3
2,4-dlmtrotoluene
1,2-diphenylhydrazinerdo
I ,2-dipnenylhydrazinelJ'
1163
385
696
700
696
704
722
724
737
740
7S8
760
788
799
819
823
830
84S
849
881
889
921
924
939
955
958
963
967
1005
1006
1147
1142
1185
1200
1205
1211
1245
1247
1269
1273
1283
1300
1298
1304
1331
1335
139S
1401
1406
1409
1409
1414
1344
1359
1433
1439
1.000
0.330
0.600
1.003
0.596
1.012
0.621
1.003
0.634
1.004
0.652
1.003
0.678
1.014
0.704
1.005
0.714
0.726
1.005
0.757
1.009
0.792
1.003
0.807
0.821
1.003
0.828
1.004
0.864
1.000
0.986
0.996
1.019
.013
.036
.005
.071
.002
.091
.003
.103
.013
.116
1.005
1.144
1.003
1.119
1.004
1.209
1.002
1.211
1.004
1.156
I. Oil
1.232
1.004
190
74
71
94
101
93
152
146
152
146
152
146
131
121
204
201
70
128
123
83
82
125
122
93
183
180
136
123
231
225
241
237
169
162
164
154
160
152
167
163
167
165
164
154
176
168
176
166
209
204
153
149
168
165
82
77
I
5
1
1
I
1
I
I
1
1
1
1
1
1
I
1
2
1
1
1
1
1
I
1
1
1
1
1
1
1
3
3
1
1
2
2
1
1
1
1
1
1
I
1
I
1
2
2
1
1
NNOMA
PHENOL
3CEE
13-2CIBNZ
14-2CLBNZ
12-2CLBNZ
B2CIE
6CLETH
NNONPRA
N8N2
ISOPHORONE
24-2HPHN
3CEOM
124-3CLBNZ
NAPHTHAIEFC
6CLBUTAO
6CLCYCPEN
2-CLNAP
B1PHENYL
ACENAPTYLE
OHP
26-2NTOL
ACENAPE
DIBNZFURAN
FLUORENE
4CPPE
OEP
24-2NTOL
12-2PHHYZ
62-75-9
108-95-2
111-44-4
541-73-1
106-46-7
95-50-1
108-60-1
67-72-1
621-64-7
98-95-3
78-59-1
105-67-9
111-91-1
120-82-1
91-20-3
87-68-3
77-47-4
91-58-7
92-52-4
208-96-8
131-11-3
606-20-2
33-32-9
132-64-9
86-73-7
7005-72-3
84-66-2
121-14-2
122-66-7
1-49
-------
TABLE
Sionenylannne-diQ
:ipnen/iaimne
1-1 i trosod ipne^y ' aimne-Jc
4-oromopneny i pnenyl e'.ner
lexacnl oropenzene-* JC*
ie*acn!orooeiiene
3nenantnrene-ai3
snenantnrene
intnracene-ajQ
irtnr scene
aioenzotniopnene-cg
siaenzotmopnene
carpazole
31-n-oulyl pntnalate-34
ai-n-outyl pntnalate
f luorantnene-dio
f luorantnene
3yrene-di n
uyrene
aenzidine-Cg
Denziaine
DutylQenzyl sntnalate
cnrysene-di2
cnrysene
aenzo( a| dntnracene-dj 2
oenzolaiantnracene
J , j' -aicnlorooenzidine-a§
J.3'-aienlorooenzidme
3is(2-etnylhe*yl)pntnalate-a4
D i s( 2 -etnylheiyi) pntnalate
di-n-octyl pntnalate-d4
di-n-octyl pntnalate
aenzo( D) f Iuorantnene-di2
oenzo(Dj f luorantnene
Den zoi is) f Iuorantnene-di2
Denzo(ic) fluorantRene
aenzoi ajpyrene-di2
Denzo( ajpyrene
aenzoig,n,i)perylene-d|2
aenzoi g ,n , i ) pery 1 ene
mdenol 1 .2.3-c.d)oyrene
d iDenzoi a,n) antnracene
2-cnioropnenol-d4
Z -cnl oropnenol
2-nitropnenol-a4
2 -nitropnenol
2 ,4-dicnlorppnenol-d3
2,4-dichloropnenol
4 -cM oro-3 -metny 1 pneno 1 -dj
4-cnloro-3-metnylpnenol
2,4,6-tr icnloropnenol-d2
2,4,6-trichloropnenol
2,4,5-trichlorcpnenol
2 .3.6- tricnl oropnenol
2 ,4-dimtropnenol-d3
2.4-dinitropnenol
4-nitropnenol*d4
4-nitropnenol
2 -metny 1 -4 , 6-d tn i tropfienol -to
2-metnyl-4,6-dinitroptienol
pentacnloropnenol*13Cg
pentacnl oropnenol
1437
1439
1433
1439
1498
1521
1522
1578
1533
1538
1592
1559
1564
1650
1719
1723
1813
1817
1844
1852
1854
1355
2060
2081
2083
2082
2090
2086
2088
2123
2124
2239
2240
2281
2286
2287
2293
2350
23S2
2741
2750
2650
2660
701
705
398
900
944
947
1086
1091
1162
1165
1170
1195
1323
1325
1349
1354
1433
1435
1559
1561
1.236
1.001
1.236
1. 001
1.283
1.308
1. 000
1.357
1.003
1.365
1.003
.1.340
1.003
1.419
1.478
1.002
1.559
1.002
1.586
1.004
1.594
1.000
1.771
1.789
1.001
1.790
1.004
1.794
1.001
1.325
1.000
1.925
1.000
1.961
1.002
1.966
1.003
2.021
1.001
2.357
1.003
0.967
0.970
0.603
1.006
0.772
1.002
0.812
1.003
0.934
1.005
0.999
1.003
1.007
1.028
1.138
1.002
1.160
1.004
1.232
1.001
1.340
1.001
179
169
175
169
248
292
234
138
178
188
178
192
134
167
153
[49
212
202
212
202
192
134
149
240
223
240
223
258
252
153
149
153
149
264
252
264
252
264
252
288
276
276
273
132
128
143
139
167
162
109
107
200
196
196
196
137
184
143
139
200
198
272
266
1
1
1
1
1
1
I
1
4
1
1
1
I
3
1
I
I
t
1
1
5
5
.1
1
1
I
1
5
5
I
1
1
2
2
2
2
1
1
2
2
1
1
2
2
2
2
2
2
20
20
6
6
13
13
5
5
3P*
NNP
4-8PPE
5CL3NZ
PNENANTHRN
ANTHRACENE
OIBNZTHIO
CARBAZQLE
OINBP
FUIOMNTHN
PYRESE
azio
3UTBNZPHT
CHRYSENE
3AA
J3-2CL3ZIO
82ETHXPHTH
2NOCTP
BBF
BICF
SAP
3GHIP
1NOENO-PYR
08AHA
2-CLPHN
2-NPHN
24-2CLPHN
4-CL2-MPHN
246-3CLPHN
236-3CLPHN
24-2HPHN
4-NPHN
46-2NOCRES
5CLPHN
122-39-4
36-30-6
101-55-3
118-74-1
35-01-3
120-12-7
132-75-0
36-74-2
34-74-2
206-44-0
129-00-0
92-87-5
35-68-7
218-01-9
56-55-3
91-94-1
117-81-7
117-34-0
2Q5-99-2
207-08-9
50-32-8
191-24-2
193-39-5
53-70-3
95-57-8
88-75-5
120-83-2
59-50-7
38-06-2
93-37-55
51-28-5
100-02-7
534-52-1
87-86-5
A Relative retention times for labeled compounds are referenced to DFB. Relative retention tines
for unlabeled compounds are referenced to their labeled analogs or to tne most cnemically similar,
most closely eluting labeled compounds if labeled analogs are not listed.
-------
TABLE I-l. (Continued)
BtUtiv*
totantion
TIM
Compouno to OF 8
aacafluorooenropnanona (OFBP) 0.736
toiapnana (mntura) 1.2*1.9
Arodor 1242 (PCB «uturt)
Aroelor 12S4 [PCB «iiiurt|
Arcclor 1260 PCS «i«turtj
a-MCM 1.32
f-HCH 1.36
y-MCH 1.41
5-MCH 1.43
»»drm
naptacnlor
napticnlor cooiidt
y-cnloratna
d-»ndoiulf«n
duldrin
4.4--OOE
3-tndOiulfan
endrin
tnonn aldahyda
4. 41 -000
amtoiwlfan lulfata
X- en lord ana
4,4'-OOT
,3.7.8-TCDO*6'
.64
.70
.83
.85
.91
.98
.00
.02
.02
.07
.10
.13
.13
.17
.01
Quantisation
(Prmary)
m/z
231.
183.
183.
183.
183.
263.
100.
3U.
373.
195.
373.
241.
246.
207.
263.
239.
272.
336.
239.
320.
233
iai
181
181
265
272
3SS
379
207
375
263
2*8
(95
277
165
387
338
237
322
GC/NS
(no,)
10
10
10
10
2
3
2
3
2
2
2
2
2
2
2
2
2
2
2
5
2
2
3
GC/ECO
5
100
100
100
100
20
5
5
20
5
5
IS
10
5
5
5
30
5
S
30
JO
JO
10 _
.('J
40
ODES
TOMPHENE
peas
ocas
peas
6U-CHX-A
6CL-CHX-8
UNOA*
6CL-CHX-0
ALORIN
MfPTACHLOR
MEPCL EPOI
ENOOSULFAN
OIELORIN
ODE
ENOHIN
ENOtlN-ALO
000
ENOOSLFN-S
DOT
OI011N
CASRN
aooi-35-2
S3469-21-9
11097 -69-1
11096-32-5
319-84 -6
J19-84-7
319-86-8
M-89-9
309-00-2
76-44-8
1024-S7-3
115-29*7
U-57-1
72-55-9
115-29-7
72-20-8
7421-93-4
72-54*8
1031-07-8
50-29-3
1746-01-6
Additional 301 (h) P*iticidtt:
Ot«ttan
Gutnion
Mil«thion
Piritnion
Nttnoiycnlor
Mir«
1
1
1
1
.19
.19
.51
.52
127.
291.
23ft.
272.
99.
109.
227.
237.
174
139
274
274
STSTOX
6UTHION
NAUntlOM
PARATMION
SO MCTNOnCL
100 NIREX
8065-48-3
86-50-0
121-75-5
S6-38-Z
72-43-5
2385-85-5
ODES COM*.
2 U
Uealad racovarj (ivrrogtca) ittndartf; tjotoptetl'- laaalad turrogatas do not
I; liotoplcilly liDllafl lurrggiut do not rv*»a DOES codai.
C-liDtl*> rKO»«ry [iurroq«U|
0« ttc tad it «ioe«
4 0«ttcttU » dlpAtnylwina.
5 LOM l»«tl Mounts (<2 ng) of DOT irt dtfly«rofialag«i«ttd and convirtM to DOE tt v«rt«Dlt ritit
on th« GC tjtt«.
6 AcctptiDli (Jttietlon Haiti will B« *tt*«1n«6)« «Hh tnt U.S. EPA Contrict L«Bor«tory Prog raw
Oloaift A«i«l7«i* prKMurt {So11/S«dlwnt Mtrii. Niltl-Conccncncloii, S*ltcti« Ion Monitoring (SIM)
GC/NS Analyiit; 9/15/83].
Colunn: 30 0.2S m !.«.. 941 mtwj\, 41 pnwyl, II vinyl bondad pft«t« fultd Ullct capillary
(JW 08-5. or fojulvalMit)
Twiptratun progra (GC/NS): 5 «1n «t 30« C; 30-2800 C *t 80 C par mln; itotncrvtl tt ZBOO c until
wio(a. n.tjporyltni alutit.
C«rrur gat Hn««r ««locUy: 30 OB/UC.
1-51
-------
ABLE 1-2. DFTPP MASS-INTENSITY SPECIFICATION
'ass Intensity
51 30-605 of mass 198
53 Less t.nan 2% of mass 69
70 Less tnan l\ of mass 69
127 40-60* of mass 198
197 Less than 1*. of mass 198
198 3ase pea*, 100* relative abundance
199 5-91 of mass 198
275 10-301 of mass 198
365 U of mass 198
141 Less than mass 4-13
442 Greater than 401 of mass 198
443 17-231 of mass 442
1-52
-------
r-SLE M. SUMMARY OF AVAILABLE PRECISION WO RECOVERY DATA*
Phenols
Aromatic hydrocarbons
Chlorinated hydrocarbons
Total PCBs
Phthalates
Miscellaneous compounds
Benzyl alcohol
Olbenzofuran
Phenols
Aromatic hydrocarbons
Chlorinated hydrocarbons
Phthalates
Surface
Sediments
Subsurface
Sediments
Enqlish Sole
Muscle
o
Lwers
Precision*3 (Mean Coefficient of Variation)
1 l7
I61
* 54
47 (59)
80 (99)
31 (120)
71
M8
^41
+ 15
* 44
100
Percent Recovery^
69 (67) 17 (67)
60 (94) 33 (98)
17 (140) 11 (124)
59 44
15
* 34
a Source: Tetra Tech, Inc. 1985. Commencement Bay Nearshore/Tideflats
Remedial Investigation. Vol. 1. Final report prepared for the Washington
State Department of Ecology and U.S. Environmental Protection Agency.
15 Precision determined by multiple sets of replicate analyses. Value
shown is mean coefficient of variation in sets of replicates with detected
values (recovery corrected).
c Values shown are mean percent recoveries of isotopically labeled compounds
added in quantities within a factor of ten of the lower limit of detection.
The values in parentheses are the mean percent recoveries obtained from
multiple matrix spike samples. The matrix spike compounds were added at
levels several times higher than the isotope recovery standards.
1-53
-------
:=EC:S::N AND ACCURACY OF METHOD BLANKS
Percent Recovery*
EPA Priority Pollutants
Phenols
phenol
2, 4-dimethyl phenol
2-chlorophenol
2,4-dichlorophenol
4-chloro-3-methyl phenol
2,4, 6-tr ichl orophenol
pentachlorophenol
2-nitrophenol
4-mtrophenol
2,4-dinitropnenol
4, 6-dimtro-2 -methyl phenol
Aromatic Hydrocarbons
naphthalene
acenaphthene
acenaphthyl'ene
fluorene
phenanthrene
anthracene
fluoranthene
pyrene
benz(a)anthracene
chrysene
benzo(b) fluoranthene
ben zo(k) fluoranthene
benzo( |a) pyrene
indeno( 1 ,2, 3-cd) pyrene
d ibenzo ( a ,h ) anthracene
benzo(ghi)perylene
Chlorinated Hydrocarbons
1,2-dichlorobenzene
1,3-dichlorobenzene
1,4-dichlorobenzene
1,2,4-trichlorobenzene
2-chl oronaphthal ene
hexachlorobenzene
hexachloroethane
hexachl orobutad 1 ene
hexachlorocycl open tad 1 ene
Blank
1
96
92
100
98
99
98
110
96
110
99
110
110
120
110
120
120
120
120
120
120
120
100
110
130
160
150
120
100
87
110
93
110
160
73
98
23
Blank
2
102
120
102
101
97
100
110
99
110
100
100
no
110
110
120
120
120
120
110
110
120
102
113
125
161
189
120
110
120
120
98
110
110
104
105
25
Blank
3
91
110
110
100
110
140
120
120
110
98
110
120
120
120
130
120
110
130
150
95
150
110
120
130
170
180
120
110
94
120
130
120
120
69
130
24
Blank
4
96
97
104
110
100
100
110
110
110
110
110
120
120
120
120
130
130
130
130
110
130
110
110
130
160
190
120
110
120
130
100
120
110
77
110
25
Mean
96
105
104
102
102
110
113
106
110
102
108
115
118
115
123
123
120
125
129
109
130
106
113
129
163
177
120
108
105
120
105
115
125
81
111
24
Coeff. of
Variation
4.7
5.0
4.2
5.2
5.7
18.6
4.4
10.3
0.0
5.5
4.7
5.0
4.3
5.0
4.1
4.1
6.8
4.6
13.4
9.5
10.9
5.0
4.2
1.9
3.0
10.6
0.0
4.7
16.4
6.8
15.9
5.0
19.0
19.6
12.4
3.9
1-54
-------
TABLE 1-4. (Continued)
Phthalates
bis(2-ethylhexyl)phtha1ate 120 130 120 130 125 4.6
benzylbutylphthalateb 270 298 120 310 250 35.2
di-n-butylphthalate 120 120 130 120 123 4.1
di-n-octylphthalate 120 120 120 120 120 0.0
diethylphthalate 120 120 130 120 123 4.1
dimethylphthalate 110 120 140 120 123 10.3
Malogenated Ethers
bis(2-chloroethyl)ether 91 93 91 100 94 4.6
bis(2-chloroisopropylJether c c c 61 ---
bis(2-chloroethoxy)methane 200 110 190 140 160 26.5
4-chlorophenylphenylether 120 120 140 120 125 8.0
4-bromophenylphenylether 150 140 120 150 140 10.1
Organonitrogen Compounds
nitrobenzene 37 21 19 c 19 38.4
N-nitrosodipropylamine 26 135 150 73 96 59.7
N-nitrosodimethylamine 110 120 110 120 115 5.0
N-nitrosodiphenylamine 76 89 173 77 104 44.9
2,4-dimtrotoluene 120 120 120 100 115 8.7
2,6-dinitrotoluene 130 74 130 100 109 24.9
benzidine 0000
3,3'-dichlorobenzidine 120 170 140 120 138 17.2
1,2-diphenylhydrazine 120 110 140 100 118 14.5
Miscellaneous
isophorone 130 120 72 120 111 23.6
a Method blanks were processed after spiking with known amounts of unlabeled
and labeled compounds. Recovery-corrected concentration of unlabeled priority
pollutants was calculated using the recovery of labeled analogs for each
compound. The final percent recovery for the unlabeled compounds was then
computed as the ratio of the calculated concentration to the known spike
level of each compound.
b Benzylbutylphthalate results are anomolously high because of laboratory
contamination traced to mercury used in the sulfur-removal step of the
procedure. The contamination was subsequently eliminated.
c Spectral interferences precluded quantification.
1-55
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SECTION II
ANALYSIS OF VOLATILE ORGANIC COMPOUNDS
IN ESTUARINE AND MARINE SEDIMENTS
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CONTENTS
Page
1.0 SCOPE AND APPLICATION H'l
2.0 SUMMARY OF METHOD II-2
3.0 INTERFERENCES ll'*
4.0 SAFETY II-4
5.0 APPARATUS AND EQUIPMENT H-5
6.0 REAGENTS AND CONSUMABLE MATERIALS 11-9
7.0 SAMPLE COLLECTION, PREPARATION, AND STORAGE 11-12
8.0 CALIBRATION AND STANDARDIZATION H-13
9.0 QUALITY CONTROL U-18
10.0 PROCEDURE H-20
11.0 QUANTITATIVE DETERMINATION (CALCULATIONS) 11-25
12.0 PRECISION AND ACCURACY H-27
13.0 REFERENCES H-27
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ANALYSIS OF VOLATILE ORGANIC COMPOUNDS
IN ESTUARINE AND MARINE SEDIMENTS
1.0 SCOPE AND APPLICATION
1.1 This method is designed to determine the volatile priority pollutants
(Table II-l) associated with Clean Water Act Section 301(h) regulation [40
CFR 125.58(k) and (v)]. Additional compounds amenable to purge-and-trap gas
chromatography-mass spectrometry (GC/MS) may be suitable for analysis,
subject to testing.
1.2 The chemical compounds listed in Table II-l can be determined in sediment
samples collected from estuarine and marine environments by this method.
1.3 The detection limit of this method is usually dependent upon the level
of interferences rather than instrumental limitations.
Lower limits of detection (LLD) are established by analysts based on
their experience with the instrumentation and with interferences in the
sample matrix being analyzed. LLD are greater than instrumental detection
limits because they take Into account sample interferences. To estimate
LLD, the noise level should be determined in the retention window for the
quantitation mass of representative analytes. These determinations should
be made for at least three field samples in the sample set under analysis.
The signal required to exceed the average noise level by at least a factor
of two should then be estimated. This signal 1s the minimum response required
to identify a potential signal for quantification. The LLD 1s the concentration
corresponding to the level of this signal based on calibrated response
factors. Based on best professional judgment, this LLO would then be applied
to samples 1n the set with comparable or lower Interference. Samples with
much higher interferences (e.g., at least a factor of two higher) should be
assigned LLD at a multiple of the original LLD.
II-l
-------
These LLO values may be less than the rigorously defined method detection
limits specified in the revised "Guidelines Establishing Test Procedures for
the Analysis of Pollutants" (40 CFR Part 136, 10/26/84). This latter procedure
requires the analysis of seven replicate samples and a statistical determination
of the method detection limit with 99 percent confidence. Data quantified
between the LLD and the rigorous method detection limit are valid and useful
in environmental investigations of low-level contamination, but have a lower
statistical confidence associated with them than data quantified above the
method detection limit.
The LLD are roughly 5-10 ppb (wet weight) with the exception of acrolein,
which has not been thoroughly tested on sediment matrices with this method.
2.0 SUMMARY OF METHOD
2.1 Volatile organic compounds are vacuum extracted from a 5-g (wet wt)
sediment sample and concentrated in a cryogenic trap cooled with" liquid
nitrogen (references 1 and 2). The cryogenic trap is then transferred to a
conventional purge-and-trap device. The extract is diluted to 5 mL with
water and treated as an aqueous sample. In the purge-and-trap device, the
volatile organic compounds are purged from the aqueous phase into a gaseous
phase with an inert carrier gas. The volatile compounds are passed into a
sorbent column and trapped. After purging 1s completed, the trap 1s backflushed
and heated rapidly to desorb the compounds Into a gas chromatograph (GC).
The compounds are separated by GC and detected with a mass spectrometer (MS).
Analysis is carried out by GC/MS either according to the Isotope dilution
technique (U.S. EPA Method 1624 Revision B; reference 3) or U.S. EPA Method
624 (reference 4). Both of these methods were developed for water/wastewater
sample matrices. The Isotope dilution technique, which requires spiking the
sample with a mixture of stable isotope labeled analogs of the analytes, is
preferred because 1t provides reliable recovery data for each analyte.
Method 624 requires spiking samples with only three surrogate compounds and
does not allow for recovery corrections. If uniformly high recoveries can
II-2
-------
"De attained with Method 624, then addition of numerous labeled compounds
(Method 1624 Revision 3) and recovery corrections are unnecessary. However,
until such performance can be demonstrated, Method 1624 Revision B provides
a Jetailed and valuable assessment of analytical performance.
Hiatt (reference 5) proposed another vacuum distillation procedure (for
tissue matrices) that did not include a purge-and-trap device. In this
technique, volatile organic compounds are transferred directly from a
cryogenically cooled trap to a fused-silica capillary column for GC/MS
analysis. This capillary column technique allows for optimum resolution and
rapid conditioning between samples. However, the performance of the technique
has not been thoroughly tested (reference 6). A potential problem is that
water can enter the capillary column and cause chromatographic problems or
it can freeze, effectively plugging the column. Thus, Hiatt's original
procedure (references 1 and 2), which has been tested more thoroughly, is
recommended here.
Vacuum distillation is recommended rather than direct or heated purge-
and-trap extraction (i.e., without vacuum distillation) because the former
technique has been demonstrated to allow for better recoveries of spiked
compounds than the latter techniques (reference 1).
2.2 Laboratories may use alternative analytical procedures if evidence of
performance comparable to the recommended procedure is provided.
3.0 INTERFERENCES
3.1 Impurities in the purge gas, organic compounds out-gassing from the
plumbing upstream of the trap, and solvent vapors in the laboratory account
for the majority of contamination problems. The analytical system is demon-
strated to be free from Interferences under conditions of the analysis by
analyzing blanks initially and with each sample lot (samples analyzed on the
same 8-h shift), as described In Sect. 9.3. Common laboratory solvents
(e.g., methylene chloride) are often contaminants 1n volatlles analyses.
II-3
-------
3.1.1 Newly packed traps should be conditioned overnight at 170°-180° C
by backflushing with an inert gas at a flow rate of 20-30 mL/min. Traps
rcust oe conditioned daily for a minimum of 10 min before use.
3.2 There is potential for ambient contamination of samples and extracts
when using vacuum and cryogenic concentration techniques. Care must be
taken to eliminate any leaks in the vacuum extraction and concentration
device: A critical source of potential contamination is pump oil vapor and
exhaust from the vacuum pump; this should not be a problem if the system is
properly sealed. A cold trap is placed between the vacuum pump and
concentration trap to prevent condensation of pump oil vapors in the concen-
tration trap (Figure II-I). All materials in the vacuum extraction and
concentration device that contact the sample and its vapors must be made of
stainless steel and/or borosilicate glass. All connections and seals must
be free of elastomers or grease that either outgas or allow penetration of
ambient contaminant vapors.
3.3 Samples can be contaminated by diffusion of volatile organic compounds
(particularly methylene chloride) through the bottle seal during shipment
and storage. A field blank prepared from reagent water and carried through
the sampling and handling protocol serves as a check on such contamination.
3.4 Contamination by carry-over can occur when high level and low level
samples are analyzed sequentially. When an unusually contaminated sample is
analyzed, it should be followed by analysis of a reagent water blank to
check for carry-over. Because the transfer lines, trap, and other parts of
the system can retain contaminants and interferences, frequent bakeout and
purging of the entire system may be required.
4.0 SAFETY
4.1 The toxidty or cardnogenicity of each compound or reagent used in
this method has not been precisely determined. However, each chemical
compound should be treated as a potential health hazard and exposure should
be reduced as much as possible. The laboratory is responsible for maintaining
II-4
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a current awareness file of OSHA regulations regarding the safe handling of
the chemicals specified in this method. A reference file of data handling
sheets should also be made available to all personnel involved in these
analyses. Additional information on laboratory safety can be found in
references 7-9.
4.2 The following compounds covered by this method have been tentatively
classified as known or suspected human or mammalian carcinogens: benzene,
carbon tetrachloride, chloroform, and vinyl chloride. Primary standards of
these toxic compounds should be prepared in a hood, and a NIOSH/MESA-approved
toxic gas respirator should be worn when high concentrations are handled.
4.3 The following safety measures must be employed when handling cryogenic
and vacuum systems:
4.3.1 Liquid nitrogen (LN^) must not be allowed to contact flesh since
it will cause extreme frostbite and deaden, (kill) tissues.
4.3.2 The concentrator and cold traps must never be closed off or
sealed after allowing any concentration of liquid air. The liquid air will
vaporize, resulting in tremendous pressure buildup and explosive damage to
the vacuum system. Always vent any vessel immediately after removing the
cryogenic or LN. bath. Wear safety goggles when working with cryogenic and
vacuum systems.
5.0 APPARATUS AND EQUIPMENT
5.1 Sample Handling Equipment
5.1.1 Stainless steel spatula.
5.1.2 Sample vessel - Pyrex flask with 15 mm 0-ring connector, washed
with detergent and rinsed with distilled water and oven-dried at 450° C.
II-5
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5.1.3 0-ring, Buna N - sonicated with 50 percent methanol/water then
dried by vacuum at 60° C. 0-rings made of TFE (tetrafluorothylene) are not
recommended because they do not produce adequate seals under vacuum.
5.2 Apparatus for Vacuum Distillation and Cryogenic Concentration (Figure
II-l).
5.2.1 Vacuum pump - capable of achieving 10 Torr and 25 L/min.
5.2.2 Vaccum/pressure gauge - with a range of subatmospheric pressure
to 10 psi.
5.2.3 Concentrator trap or purge flask - 25 ml capacity (Tekmar Part
No. 14-0957-024 or equivalent) modified with 9-fim 0-ring connectors.
5.2.4 Cold trap - glass trap (easily produced by glassblowing, Figure
II-l) with 0-ring fittings (e.g., Kontes 671750-009).
5.2.5 Transfer line - 1/4 in o.d. glass-lined stainless steel tubing.
Lines should be kept as short as possible to minimize sample carryover.
5.2.6 Vacuum valves - Nupro B-48KT or equivalent.
5.2.7 Oewar flasks - 665 ml or 1,000 mL, for liquid nitrogen bath.
5.2.8 Assorted compression fittings and graphite ferrules (Figure II-l).
5.2.9 Ultrasonic bath - Branisonic 12 or equivalent.
5.2.10 Heater tape - to wrap around stainless steel lines and valve
bodies to maintain a temperature of 60° C.
5.2.11 Pinch clamps, Thomas - to secure 0-r1ng connections.
II-6
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5.3 Purge-and-Trap Device - capable of meeting specifications listed in
U.S. EPA Method 1624 Revision B (see below). Complete devices consisting of
a purging device (the concentrator trap, Sect. 5.2.3), a Tenax/silica trap,
and a desorber are commercially available (e.g., Tekmar Model LSC-2, Tekmar
Co., Cincinnati, OH).
5.3.1 Trap - 25 to 30 cm x 2.5 mm i.d. minimum, containing the following:
5.3.1.1 Methyl silicone packing - one ^0.2 cm, 3 percent OV-1 on
60/80 mesh Chromosorb W, or equivalent.
5.3.1.2 Porous polymer - 15 +1.0 cm, Tenax GC (2,6-diphenylene
oxide polymer), 60/80 mesh, chromatographic grade, or equivalent.
5.3.1.3 Silica gel 8 *l.O cm, Oavison Chemical, 35/60 mesh,
grade 15, or equivalent.
5.3.2 Oesorber - should heat the trap to 175 +5° C in 45 sec or less.
The polymer section of the trap should not exceed 180° C, and the remaining
sections should not exceed 220° C.
5.3.3 Commercial purge and trap devices are easily coupled to GC
systems.
5.4 GC/MS (Gas Chromatograph-Mass Spectrometer) System.
5.4.1 GC - should be linearly temperature programmable with initial
and final temperature holds.
5.4.2 GC column - 6 ft long x 0.1 in i.d. (stainless steel or glass)
packed with 1 percent SP-1000 on Carbopak B, 60/80 mesh or equivalent.
5.4.3 MS - 70 eV electron impact ionization; capable of repeatedly
scanning from 20 to 250 amu every 2 to 3 sec.
II-7
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5.4.4 GC/MS interface GC to MS interfaces constructed of all-glass
or glass-lined materials are recommended. Glass can be deactivated by
silanizing with dichloro-dimethyl silane.
5.5 Data System - should collect and record MS data, store mass intensity
data in spectral libraries, process GC/MS data and generate reports, and
calculate and record response factors.
5.5.1 Data acquisition mass spectra should be collected continuously
throughout the analysis and stored on a mass storage device.
5.5.2 Mass spectral libraries user-created libraries containing mass
spectra obtained from analysis of authentic standards should be employed to
reverse search GC/MS runs for the compounds of interest.
5.5.3 Data processing - the data system should be used to search,
locate, identify, and quantify the compounds of Interest in each GC/MS
analysis. Software routines should be employed to compute retention times
and extracted ion current profile (EICP) areas. Displays of spectra, mass
chromatograms, and library comparisons are required to verify results.
5.5.4 Response factors and multipoint calibrations - the data system
should be used to record and maintain lists of response factors (response
ratios for isotope dilution) and generate multi-point calibration curves.
Computations of relative standard deviation (coefficient of variation) are
useful for testing calibration linearity.
5.6 Other Materials
5.6.1 Syringe, 10 uL +_ I percent of volume.
5.6.2 Syringe, 50 ul +_ I percent of volume.
II-8
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5.6.3 Syringe, 5 ml ^ 1 percent of volume, gas-tight with shut-off.
5.6.4 Bubble flowmeter.
6.0 REAGENTS AND CONSUMABLE MATERIALS
6.1 Reagent Water
6.1.1 Reagent water Is defined as water free of interferences (i.e.,
interferents are not observed at the detection limits of the compounds of
interest).
6.1.2 Prepare water by boiling 1 I of freshly distilled water down to
900 ml and transferring the water to a 1-L volumetric flask that has been
modified by replacing the ground glass joint with a 15-mm i.d., Buna-N
0-ring connector.
6.1.3 Connect the flask to the distillation apparatus at the sample
chamber site and evacuate for 15 min while continuously agitating the flask
in an ultrasonic cleaner.
6.1.4 After evacuation, release an inert gas (N2 or He can be used)
into the flask until equilibrium is obtained, then seal with a cap made from
a Buna-N 0-ring connector.
6.2 Methanol - pesticide quality or equivalent.
6.3 Standard Solutions - purchased as solutions or mixtures with certification
of their purity, concentration, and authenticity, or prepared from materials
of known purity and composition. If compound purity is 96 percent or greater,
the weight may be used without correction to calculate the concentration of
the standard.
II-9
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6.4 Preparation of Stock Solutions prepare in methanol using liquid or
gaseous standards per the steps below. Observe the safety precautions given
in Sect. 4.
6.4.1 Place approximately 9.8 mL of methanol in a 10-mL ground glass
stoppered volumetric flask. Allow the flask to stand unstoppered for approxi-
mately 10 min or until all methanol-wetted surfaces have dried. In each
case, weigh the flask, immediately add the compound, then immediately reweigh
to prevent evaporation losses from affecting the measurement.
6.4.1.1 Liquids - using a 100 uL syringe, permit two drops of
liquid to fall into the methanol without contacting the neck of the flask.
Alternatively, inject a known volume of the compound into the methanol in
the flask using a micro-syringe. With the exception of 2-chloroethylvinyl
ether, stock standards of compounds that boll above room temperature are
generally stable for at least 4 wk when stored at 4 C.
6.4.1.2 Gases (chloromethane, bromomethane, chloroethane, vinyl
chloride) - fill a valved 5-mL gas-tight syringe with the compound. Lower
the needle to approximately 5 mm above the methanol meniscus. Slowly introduce
the compound above the surface of the meniscus. The gas will dissolve
rapidly in the methanol.
6.4.2 Fill the flask to volume, stopper, then mix by inverting several
times. Calculate the concentration In mg/mL (ug/uL) from the weight gain
(or density if a known volume was injected).
6.4.3 Transfer the stock solution to a Teflon sealed screw-cap bottle.
Store, with minimal headspace, 1n the dark at -10 to -20° C.
6.4.4 Prepare fresh standards weekly for the gases and 2-chloroethylvinyl
ether. All other standards are replaced after 1 mo, or sooner if comparison
with check standards indicates a change 1n concentration of over 10 percent.
Quality control check standards that can be used to determine the accuracy
11-10
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of calibration standards are available from the U.S. Environmental Protection
Agency, Environmental Monitoring and Support Laboratory, Cincinnati, Ohio.
6.5 Labeled Compound Spiking Solution - from stock standard solutions
prepared as above, or from mixtures, prepare the spiking solution to contain
a concentration such that a 5-10 uL spike into each 5-mL sample "extract",
blank, or aqueous standard analyzed will result in a concentration of 10 ng/mL
of each labeled compound. For the gases and for the water soluble compounds
(acrolein, acrylonitrile), a concentration of 50 ng/mL may be used. Include
the internal standards (Sect. 8.1.2) 1n this solution so that a concentration
of 10 ng/mL in each sample, blank, or aqueous standard will be produced.
6.6 Secondary Standards - using stock solutions, prepare a secondary standard
in methanol to contain each pollutant at a concentration of 250 ug/mL. For
the gases and water soluble compounds (Sect. 6.5), a concentration of 1.25
mg/mL may be used.
5.7 Aqueous Calibration Standards the concentrations of calibration
solutions suggested in this section are intended to bracket concentrations
that will be encountered during sample analysis that will not overload the
analytical system. Use sufficient amounts of the secondary standard (Sect.
6.6) and reagent water to produce concentrations of 5, 10, 20, 50, and 100 ug/L
in the aqueous calibration standards. The concentrations of gases and water
soluble compounds will be higher (I.e., 25, 50, 100, 250, and 500 ug/L).
Analysts may use a wider range of standard concentrations if linearity can
be demonstrated.
6.8 Aqueous Performance Standard - an aqueous standard containing all
pollutants. Internal standards, labeled compounds, and BFB (4-bromofluoro-
benzene) 1s prepared dally and 1s analyzed each shift to demonstrate performance
(Sect. 8.2). This standard should contain either 10 or 50 ug/L of the
labeled and pollutant gases and water soluble compounds, 5 ug/L of BFB, and
10 ug/L of all other pollutants, labeled compounds, and Internal standards.
It may be the nominal 10 ug/L aqueous calibration standard (Sect. 6.7).
11-11
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6.9 A methanolic standard containing all pollutants and internal standards
is prepared to demonstrate recovery of these compounds when syringe injection
and purge-and-trap analyses are compared. This standard should contain
either 10 ug/mL or 50 ug/mL of the gases and water soluble compounds, and 10
ijg/mL of the remaining pollutants and internal standards (consistent with
the amounts in the aqueous performance standard in Sect. 6.8).
6.10 Other standards that may be needed are those for testing of BFB per-
formance (Sect. 8.2.1) and for collecting mass spectra for storage in spectral
libraries (Sect. 8.1.4).
6.11 High Purity Helium - 99.999 percent.
6.12 Liquid Nitrogen (LN2).
7.0 SAMPLE COLLECTION. PREPARATION, AND STORAGE
7.1 In the field, sources of contamination include sampling gear, grease
from ship winches or cables, ship engine exhaust, dust, and ice used for
cooling. Efforts should be made to minimize handling and to avoid sources
of contamination.
7.2 Fill two separate 40-ml, screw cap glass vials with sediment, leaving
no headspace. The vials and tetrafluoroethylene (TFE)-backed silicon septa
used for sealing them should be cleaned with detergent, rinsed once with tap
water, rinsed with distilled water, and dried at >105° C. Solvent cannot be
used as it will interfere with the analysis. To obtain a sample with no
headspace, fill the vial to overflowing so that a convex (upward) meniscus
forms at the top (1f there 1s adquate water 1n the sediment). Place the cap
(TFE side down) carefully on the opening of the vial, displacing excess
water. Screw on the vial cap and verify the seal by inverting the vial. If
the vial 1s properly sealed, air bubbles will not appear when it is inverted.
Samples should be taken from single grab samples, as volatile compounds can
be lost during compositing of grab samples.
.11-12
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7.3 To avoid cross-contamination, equipment used in sample handling (e.g.,
spatulas) should be thoroughly cleaned before each sample is processed.
Instruments must be of a material that can be easily cleaned (e.g., stainless
steel, anodized aluminum, or borosilicate glass). Before the next sample is
processed, instruments should be washed with a detergent solution, rinsed
with tap water, soaked in high-purity acetone and methylene chloride, and
finally rinsed with reagent water.
7.4 U.S. EPA recommends that sediment samples be stored in the dark at 4 C
and analyzed within ten days of sample receipt (reference 10). Freezing is
not recommended because no headspace will be left to compensate for the
expansion of water during freezing.
8.0 CALIBRATION ANO STANDARDIZATION
8.1 Initial Calibration
8.1.1 Calibration by the Isotope dilution technique the isotope
dilution technique is used for the purgeable organic compounds when appropriate
labeled compounds are available and when Interferences do not preclude the
analysis. If labeled compounds are not available or Interferences are
present, the internal standard technique (Sect. 8.1.2) 1s used. A calibration
curve encompassing the concentration range of Interest 1s prepared for each'
compound determined. The relative response (RR) versus concentration (ug/L)
1s plotted or computed using a linear regression. An example of a calibration
curve for a pollutant and Us labeled analog 1s given 1n Figure II-2. Also
shown are the ^10 percent error limits (dotted lines). Relative response is
determined according to the procedures described below. A minimum of five
data points 1s required for calibration (Sect. 6.7).
8.1.1.1 The relative response (RR) of pollutant to labeled compound
is determined from Isotope ratio values calculated from acqufred data.
Three Isotope ratios are used 1n this process:
11-13
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R = the isotope ratio measured in the pure pollutant (Figure II-3A)
R = the isotope ratio of pure labeled compound (Figure II-3B)
R = the isotope ratio measured in the analytical mixture of the pollutant
m
and labeled compounds (Figure II-3C).
The correct way to calculate RR is:
RR » (Ry - Rm)(Rx « l)/(Rm - Rx)(R^ « 1).
If R is not between 2R and 0.5R , the method does not apply and the sample
is analyzed by the internal standard technique (Sect. 8.1.2).
8.1.1.2 In most cases, the retention times of the pollutant and
labeled compound are the same and isotope ratios (R's) can be calculated
from the EICP areas, where:
R » (area at m./z]/(area at m2/z)
If either of the areas is zero, it Is assigned a value of one in the calcu-
lations; that is, 1f: area of m./z«50721, and area of nyz'O, then R=50721/i3
50721. The m/z's are always selected such that RX>R . When -there is a
difference in retention times (RT) between the pollutant and labeled compounds,
specidl precautions are required to determine the Isotope ratios.
R , R , and R are defined as follows:
RX [area m^i (at RT2)]/1
R I/ [area m2/z (at RT^]
Rm » [area m^/z (at RT2)]/[area m2/z (at RTj)].
8.1.1.3 An example of the above calculations can be taken from
the data plotted in Figure II-3 for a pollutant and its labeled analog. For
these data. RX » 168920/1 - 168920, R - 1/60960 » 0.00001640, and Rffl -
96868/82508 - 1.174. The RR for the above data 1s then calculated using the
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equation given In Sect. 8.1.1.1. For the example, RR=1.174. Note: Not all
labeled compounds elute before their pollutant analogs.
8.1.1.4 To calibrate the analytical system by isotope dilution,
analyze a 5-ml aliquot of each of the aqueous calibration standards (Sect. 6.7)
spiked with an appropriate constant amount of the labeled compound spiking
solution (Sect. 6.5), using the purge-and-trap procedure in Sect. 10.
Compute the RR at each concentration.
8.1.1.5 Linearity if the ratio of relative response to concen-
tration for any compound 1s constant (less than 20 percent coefficient of
variation) over the five-point calibration range, an averaged relative
response/concentration ratio may be used for that compound; otherwise, the
complete calibration curve for that compound should be used over the five-point
calibration range.
8.1.2 Calibration by internal standard - used when criteria for isotope
jllutlon (Sect. 8.1.1) cannot be met. The method 1s applied to pollutants
having no labeled analog and to the labeled compounds themselves. The
internal standards used for volatHes analyses are bromochloromethane,
2-bromo-l-chloropropane, and l,4-d1chlorobutane. Concentrations of the
labeled compounds and pollutants without labeled analogs are computed relative
to the nearest eluted Internal standard.
8.1.2.1 Response factors - calibration requires the determination
of response factors (RF), which are defined by the following equation:
RF - (As x C1s)/(A1s x C )
where:
AS * the EICP area at the characteristic m/z for the compound in the
dally standard
A. » the EICP area at the characteristic m/z for the internal standard
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C. = the concentration (ug/l) of the internal standard
C = the concentration of the pollutant in the daily standard.
8.1.2.2 The response factor is determined at 5, 10, 20, 50, and
100 ug/L for the pollutants (optionally at five times these concentrations
for gases and water soluble pollutants see Sect. 6.6 and 6.7), in a way
analogous to that for calibration by isotope dilution (Sect. 8.1.1.4). The
RF is plotted against concentration for each compound in the standard (C )
to produce a calibration curve.
3.1.2.3 Linearity if the response factor (RF) for any compound
is constant (less than 35 percent coefficient of variation) over the five-
point calibration range, an averaged response factor may be used for that
compound; otherwise, the complete calibration curve for that compound should
be used over the five point range.
8.1.3 Combined calibration - by adding the isotopically labeled compounds
and internal standards (Sect. 6.5) to the aqueous calibration standards
(Sect. 6.7), a single set of analyses can be used to produce calibration
curves for the isotope dilution and Internal standard methods.
8.1.4 Mass spectral libraries detection and identification of the
compound of Interest during calibration and sample analysis are dependent
upon the spectra stored 1n user-created libraries.
8.1.4.1 Obtain a mass spectrum of each pollutant and labeled
compound and each Internal standard by analyzing an authentic standard
either singly or as part of a mixture 1n which there is no interference
between closely eluted components. That only a single compound is present
is determined by examination of the spectrum. Fragments not attributable to
the compound under study Indicate the presence of an interfering compound.
"Adjust the analytical conditions and scan rate (for this test only) to
produce an undlstorted spectrum at the SC peak maximum. An undlstorted
spectrum will usually be obtained 1f five complete spectra are collected
across the upper half of the GC peak. Software algorithms designed to
11-16
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"enhance" the spectrum may eliminate distortion, but may also eliminate
authentic ions or introduce other distortion.
8.1.4.2 Obtain the authentic reference spectrum under BFB tuning
conditions (Table II-2) to normalize it to spectra from other instruments.
8.1.4.3 The spectrum is edited by saving the five most intense
mass spectral peaks and all other mass spectral peaks greater than 10 percent
of the base peak. This spectrum is stored for reverse search and for compound
confirmation.
8.2 Ongoing Calibration
8.2.1 The BFB standard must be analyzed at the beginning of each 8-h
shift. The tuning criteria in Table II-2 must be met before blanks and
samples may be analyzed.
8.2.2 At the beginning and end of each 8-h shift, system calibration
should be verified by purging the aqueous performance standard (Sect. 6.8).
8.2.2.1 Calibration is tested by computing the concentration of
unlabeled compounds by the isotope dilution technique (Sect. 8.1.1) for
compounds with labeled analogs. Concentrations of unlabeled compounds
without labeled analogs are calculated according to the internal standard
technique (Sect. 8.1.2).
A complete (five-point) recallbration should be performed when results
vary from predicted concentrations by more than +25 percent. The last
sample analyzed before falling criteria should then be reanalyzed. If the
results differ by more than +20 percent (I.e., at least twice the mean
reproduciblllty for replicate analysis of sediment samples, Table II-3), it
is assumed that the Instrument was out of control during the original analysis
and the earlier data should be rejected. Reanalysls of samples should
orogress 1n reverse order until it is determined that there 1s <20 percent
difference between Initial and reanalysis results.
11-17
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9..0 QUALITY CONTROL [For further guidance, see Quality Assurance/Quality
Control (QA/QC) for 301{h) Monitoring Programs: Guidance on Field and
Laboratory Methods (Tetra Tech 1986).]
9.1 Each laboratory that uses this method is required to operate a formal
quality assurance program. The minimum requirements of this program consist
of an. initial demonstration of laboratory capability, analysis of samples
spiked with labeled compounds to evaluate and document data quality, and
analysis of standards and blanks as tests of continued performance.
9.2 Initial Demonstration of Laboratory Capability
9.2.1 Analyze the aqueous performance standard (Sect. 6.8) according
to the purge-and-trap procedure in Sect. 10. Compute the area at the primary
m/z (Table II-l) for each compound. Compare these areas to those obtained
by injecting one uL of the methanolic standard (Sect. 6.9) to determine
compound recovery. The recovery should be greater than 20 percent for the
water soluble compounds (acrolein and acrylonitrile), and 60-110 percent for
all other compounds. This recovery should be demonstrated initially for
each purge-and-trap GC/MS system. The test should be repeated only if the
purge and trap or GC/MS systems are modified in any way that might result in
a change in recovery.
9.3 Blanks
9.3.1 Reagent water blanks must be analyzed to demonstrate freedom
from carry-over (Sect. 3) and contamination.
9.3.1.1 The level at which the purge-and-trap system will carry
greater than 5 ug/L of a pollutant of interest into a succeeding blank
should be determined by analyzing successively larger concentrations of
these compounds. When a sample contains this concentration or more, a blank
should be analyzed immediately following this sample to demonstrate no
carry-over at the 5 ug/L level.
11-18
-------
9.3.1.2 With each sample lot (samples analyzed on the same 8-h
sh'ift), a blank should be analyzed immediately after analysis of the aqueous
performance standard (Sect. 8.2.2) to demonstrate freedom from contamination.
If any of the compounds of interest, except common laboratory contaminants
(e.g., -nethylene chloride and toluene), or any potentially interfering
compound is found in a blank at greater than 10 ug/L (assuming a response
factor of 1 relative to the nearest eluted internal standard for compounds
not listed in Table II-l), analysis of samples is halted until the source of
contamination is eliminated and a blank shows no evidence of contamination
at this level. This control action also applies if methylene chloride or
toluene is detected in a blank at greater than 50 ug/L.
9.4 Sample Spiking
9.4.1 The laboratory should spike all samples with labeled compounds
to assess method performance on the sample matrix.
9.4.2 Spike and analyze each sample according to the method beginning
in Sect. 10.
9.4.3 Compute the percent recovery (P) of the labeled compounds using
the internal standard technique (Sect. 8.1.2).
9.5 Replicates
9.5.1 Replicate analyses (i.e., analyses of two subsamples from the
same sediment homogenate) must be performed to monitor laboratory precision.
9.5.2 At a minimum, 5 percent of the analyses should be laboratory
replicates. A triplicate analysis should be performed with each sample
batch of over 40 samples.
11-19
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10.0 PROCEDURE
10.1 Sample Processing
10.1.1 Homogenize (stir) samples with a spatula prior to analysis to
ensure that representative aliquots are taken. Mix any water that has
separated from the sediment back Into the sample. Remove and make note of
nonrepresentative material (e.g., twigs, leaves, shells, rocks, and any
material larger than 1/4 in). It is recommended that removal of material be
performed in the field by sampling personnel.
10.1.2 Dry weight determination dry weight determinations may be
performed -as follows: transfer an aliquot of approximately 3 g (weighed to
the nearest 0.1 g) to a preweighed dish. Allow the sample to dry in an oven
at 105° C overnight and determine the solid residue weight to the nearest
0.1 g. The percent total solids 1s calculated as:
T [dry residue wt (g)]/[wet sample wt (g)]
10.1.3 Immediately after homogenlzation, use a stainless steel spatula
to transfer a 5-g aliquot to a preweighed sample vessel (Sect. 5.1.2).
Weigh the transferred portion to the nearest 0.1 g.
10.1.4 Spike 50 ng of each labeled compound (or 250 ng of gaseous and
water soluble compounds) Into 2 ml "of reagent water and add to the sample
matrix. Seal the sample vessel with an 0-ring connector and clamp and
sonicate for 10 m1n. After sonlcatlon, store the sample contained in the
sample vessel overnight 1n a refrigerator/freezer and analyze the next day.
10.2 Vacuum Distillation and Concentration (Reference 2)
10.2.1 The vacuum extractor must be airtight and free of moisture
before extraction can be started.
11-20
-------
10.2.2 A clean 100-mL pyrex flask is connected to the vacuum distillation
apparatus at the sample vessel site (see Figure II-P, the vacuum pump
started, and V?"'* °Pened t° evacuate the apparatus. Line condensation is
prevented by warming the transfer lines while evacuating the system. Heating
tape is effective in creating even transfer line temperatures and can be
used continuously during the procedure.
10.2.3 The vacuum apparatus is pressurized with helium by closing V,
and opening V.. The apparatus is tested for leaks with a helium leak detector
or a liquid leak detector (e.g., Snoop), and appropriate adjustments are
made as necessary. When the apparatus has been found to be airtight, close
Vlt open V3 and then heat the transfer lines and concentrator trap to 100° C
for 5 min to eliminate any residual contamination.
10.2.4 The flask containing the sample should be immersed in liquid
nitrogen before the flask 1s uncapped. To begin the distillation, close V2
(with V, and V. remaining open), cool the concentrator trap with a liquid
iltrogen bath, and replace the empty sample vessel with the cooled sample
flask. Disconnect the vacuum source by closing V^. Open V^ to permit
vapors from the sample vessel to reach the concentrator trap. Immerse the
sample vessel in a 50° C water bath and sonicate for 5 min.
10.2.5 Connect the vacuum source to the sample vessel by opening V^.
The lower pressure hastens the transfer of volatile compounds from the
sample to the cooled concentrator trap. After 15 min of vacuum, close V^
and open V. to fill the system with helium to atmospheric pressure. Close
V. and V- to isolate the condensate. The distillation is now completed and
the condensate is ready for transfer to a purge-and-trap device. The condensate
can be held 1n the liquid nitrogen bath for up to 1 h prior to analysis.
Care should be taken to ensure that moisture does not freeze in the narrow
glass tubing in the concentrator trap. Careful drying of the system prior
to analysis and maintenance of an airtight system will preclude this problem.
11-21
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10.2.6 Disconnect the sample concentrator trap from the vacuum apparatus
and connect it to the purge-and-trap device. Some outgassing is observed
when the sample condensate is melted; therefore, the condensate should be
kept frozen until the concentrator trap is attached to the purge-and-trap
device. After attachment, warm the concentrator trap walls to loosen the
condensate and allow the ring of ice formed during condensation to drop to
the bottom of the trap. To this partially melted extract, add 3 mL of
reagent water containing 50 ng of each of the internal standards (bromo-
chloromethane, 2-bromo-l-chloropropane, and 1,4-dichlorobutane). The internal
standards are added after vacuum extraction to allow the analyst to assess
analytical losses of labeled compounds during the extraction/concentration
procedure.
10.3 Purge-and-Trap Procedure
10.3.1 Because commercial purge flasks must be slightly modified (with
0-ring fittings) to be attached to the vacuum distillation apparatus, a
simple 0-ring adapter 1s necessary to connect the purge flask to the commercial
device for which it was designed. The modified purge flask (Sect. 5.2.3)
used in this procedure can be fitted to a commercial purgeand-trap device
(e.g.. a Tekmar ALS interfaced with a Tekmar LSC-2) with 9-mm 0-ring fittings
fused to short sections of glass tubing. Commercial purge-and-trap devices
are almost entirely automated and are easy to operate with manufacturer's
instructions.
10.3.2 Purge the extract solution with the concentrator trap immersed
in an ice-water bath for 5 m1n followed by Immersion in a 55° C-water bath
for an additional 7 m1n. This provides conditions for reproducibly melting
the frozen extracts 1n order to obtain reproducible purging efficiencies.
11-22
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10.3.3 The GC conditions for analysis are as follows:
Injector zone temp. 225 C
Initial GC oven temp. 60° C
Final GC temp. 175° C
Initial hold time 3 min
Ramp rate 8° C/min
Final hold time 24 min
Jet separator oven temp. 225 C
10.4 Qualitative Determination - accomplished by comparison of data from
analysis of a sample or blank with data from analysis of the shift standard
(Sect. 8.2.2). Identification is .confirmed when spectra and retention times
agree according to the criteria below.
10.4.1 Labeled compounds and pollutants having no labeled analog:
10.4.1.1 The signals for all characteristic masses stored in the
spectral library (Sect. 8.1.4.3) should be present and should maximize
within the same two consecutive scans.
10.4.1.2 Either I) the background-corrected EICP areas or 2) the
corrected relative Intensities of the mass spectral peaks at the GC peak
maximum should agree within a factor of two (0.5 to 2 times) for all masses
stored in the library.
10.4.1.3 The retention time relative to the nearest eluted internal
standard should be within +7 scans or +20 sec, whichever is greater, of this
difference 1n the shift standard.
10.4.2 Pollutants with a labeled analog:
10.4.2.1 The signals for all characteristic masses stored in the
spectral library should be present and should maximize within the same two
consecutive scans.
11-23
-------
10.4.2.2 Either 1) the background corrected EICP areas or 2) the
corrected relative intensities of the mass spectral peaks at the GC peak
maximum should agree within a factor of two for all masses stored in the
spectral library.
10.4.2.3 The retention time difference between the pollutant and
its labeled analog should agree within +2 scans or ^6 sec, whichever is
greater, of this difference in the shift standard.
10.4.2.4 If the experimental mass spectrum contains masses that
are not present in the reference spectrum, an experienced spectrometrist
must determine the presence or absence of the compound.
10.5 Tentatively Identified Compounds (GC/MS Analysis) - The ten non-target
peaks of greatest area in the RIC (reconstructed ion chromatogram) should be
identified and quantified, if possible.
10.5.1 Guidelines for making tentative identification (reference 10):
(L) Tentative identifications should be based on a forward
search of the EPA/NIH mass spectral library. Sample spectra
should be visually compared with the most similar library match.
(2) Relative intensities of major Ions in the reference spectrum
(ions greater than 10 percent of the most abundant ion) should be
present in the sample spectrum.
(3) The relative Intensities of the major ions should agree
within +20 percent. (Example: For an ion with an abundance of 50
percent 1n the standard spectra, the corresponding sample ion
abundance must be between 30 and 70 percent.)
(4) Molecular Ions present 1n reference spectrum should be
present in sample spectrum.
11-24
-------
(5) Ions present in the reference spectrum but not in the
sample spectrum should be reviewed for possible subtraction from
the sample spectrum because of background contamination or co-eluting
compounds. Data system library reduction programs can sometimes
create these discrepancies.
10.5.1.1 If, in the opinion of the mass spectral specialist, no
valid tentative identification can be made, the compound should be reported
as unknown. The mass spectral specialist should give additional classification
of the unknown compound if possible (e.g., unknown hydrocarbon, unknown
aromatic compound, unknown chlorinated compound). If probable molecular
weights can be distinguished. Include them.
10.5.2 Tentative quantification - quantification of TIOs will be based
on the internal standard technique and an assumed response factor of one (in
the absence of data from authentic standards). The uncertain nature of this
quantification should be clearly noted In the data report.
11.0 QUANTITATIVE DETERMINATION (CALCULATIONS)
11.1 Isotope Dilution - by adding a known amount of a labeled compound to
every sample prior to vacuum distillation, correction for recovery of the
pollutant can be made because the pollutant and its labeled analog exhibit
similar behavior during purging, desorptlon, and gas chromatography. Note
that pollutants and their labeled analogs are not always retained identically
by complex matrices, so their behavior during the extraction step may differ.
Use of this technique 1s to enable correction for analytical losses after
extraction, not for matrix recovery.
11.1.1 Relative response (RR) values for sample mixtures are used in
conjunction with calibration curves described in Sect. -8.1.1 to determine
concentrations directly, so long as labeled compound spiking levels are
constant.
11-25
-------
11.1.2 For the isotope dilution technique, concentration is calculated
as follows:
C (ug/kg, dry wt sediment) a
C (ug/kg) x RR x n
where
C. = the concentration of the stable Isotope labeled compound as
spiked into the sample
RR » relative response of unlabeled pollutant to isotope labeled
surrogate in the sample
RR< » relative response at 1 point in calibration
1 hu
Z. » absolute amount of unlabeled compound at i point of calibration
th
ZA1 3 absolute amount of labeled compound at 1 point in calibration
n » number of calibration points.
11.2 Internal Standard calculate the concentration using the response
factor determined from calibration data (Sect. 8.1.2) and the following
equation:
Concentration » (A$ x C1s)/(*1s x RF) where the terms are as defined in
Sect. 8.1.2.1. except that C1s is 1n ug/kg (dry sediment) and A$ is the EICP
area at the characteristic m/z for the analyte 1n the sample.
11.3 If the EICP area at the quantltatlon mass for any compound exceeds the
calibration range of the system, a smaller sample aliquot should be analyzed
if possible. However, sample sizes of less than 0.5 g are not recommended
because such small samples may not be representative.
11-26
-------
11.4 Report results for all pollutants and labeled compounds found in
samples, in ug/kg (dry weight) to three significant figures. Pollutants and
labeled compounds in blanks should be reported in ng/sample.
12.0 PRECISION AND ACCURACY
12.1 Recoveries from replicate spiked water and sediment analyses are
presented in Table II-3 (references 1 and 2). These analyses were not
performed with the isotope dilution technique and recovery results are
uncorrected.
13.0 REFERENCES
1. H1att, M.H., "Analysis of F1sh and Sediment for Volatile Priority
Pollutants." Anal. Chem. Vol. 53. 1981. pp. 1541-1543.
2. H1att, M.H., and T.I. Jones. Isolation of Purgeable Organics from
Solid Matrices by Vacuum Distillation. U.S. Environmental Protection
Agency, Region IX, Las Vegas Laboratory, 1984.
3. Fed7 Register, Volume 49, No. 209. October 26, 1984, pp. 43407-43415.
4. Fed. Register, Volume 49, No. 209, October 26, 1984, pp. 43373-43384.
5. H1att, M.H., "Determination of Volatile Organic Compounds in Fish
Samples by Vacuum Distillation and Fused SIHca Capillary Gas Chroma-
tography/Mass Spectrometry," Anal. Chem. Vol. 55, 1983, pp. 506-516.
6. H1att, M.H. 4 November 1985. Personal Communication (phone by Mr. Harry
Seller). Analytical Technologies, Incorporated, National City, CA.
7. "Working with Carcinogens.' OHEW, PHS, NIOSH, Publication 77-206 (1977).
8. "OSHA Safety and Health Standards. General Industry," 29 CFR 1910, OSHA
2206, (1976).
9. "Safety 1n Academic Chemistry Laboratories," American Chemical Society
Publication, Committee on Chemical Safety (1979).
10. U.S. Environmental Protection Agency. 1984 (revised January, 1985).
U.S. EPA Contract Laboratory Program - statement of work for organics
analysis, multi-media, multi-concentration. IFB WA 85-J176, J177,
J178.
11-27
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VACUUM
GAUGE
'/, OD GLASS-LINED
S.S. TUBING
4' S.S.
TEE UNION
^-O-AINO CONNECTOR
25mL
PURGE
FLASK
(CONCENTRATOR
TRAP)
v,
db
NUPRO B 4BKT
LIQUID
NITROGEN
BATH
V.
C^3=
COLO TRAP
LIQUID
NITROGEN
BATH
* VACUUM
PUMP
AUAMtU I ktui HI It KIHt I .'
Null HuKU »M) IMAf UlVUt I1, till Imiuuili IN IIL.UKI
Figure 11-1. Apparatus fur vacuum dislillatiun and tryoyemt contenli at ion
-------
10 -
IU
10 -
IX)
01 -
I I i
I I
I I 1^ I I
5 10 20 50 100
CONCENTRATION (Hfl/L)
AOAPUD fMM (UdiMCi 1
(A)
(B)
(C)
ADAHIO
AREA 168920
M./Z
M./Z
AREA 60960
MJZ
M./2
M,/Z 96868
M,/Z 82508
.MJZ
M./Z
Figure 11-2.
Figure 11-3.
Relative response calibration curve.
Extracted ion current profiles for (A) the unlabeled pollutant, fb) the
labeled analog, and (C) a mixture of the labeled and the unlabeled compounds
-------
TABLE 11-1. VOLATILE ORGANIC ANALYTES
I
CJ
O
Analyte
Acrolein
Acrylomtrile
Benzene
Bromod ichloromethane
Bromoform
Bromonethane
Carbon tetrachlorlde
Chlorobenzene
Chloroethane
2-chloroethylvinyl ether
Chloroform
Chloromethane
Dlbromochloromethane
1,1-dichloroethane
1 ,2-dichloroethane
1.1-dtchloroethene
trans-1 ,2-dichloroethene
1 ,2-dichloropropane
cis-1 ,3-dichloropropene
trans-1 ,3-dichloropropene
Ethylbenzene
Hethylene chloride
1 ,1,2,2-tetrachloroethane
Tetrachloroethene
Toluene
1 ,1,1-trichloroethane
1 ,1,2-tr ichloroethane
Trichloroethene
Vinyl chloride
CASRN
107-02-8
107-13-1
71-43-2
75-27-4
75-25-2
74-83-9
56-23-5
108-90-7
75-00-3
110-75-8
67-66-1
74-87-3
124-48-1
75-34-3
107-06-2
75-35-4
156-60-5
78-87-5
10061-01-5
10061-02-6
100-41-4
75-09-2
79-34-5
127-18-4
108-88-3
71-55-6
79-00-5
79-01-6
75-01-4
ODES
ACROLEIN
ACRYLNTRLE
BENZENE
2CLBRHETHA
BROMOFORM
METIIYLBR
CARBON TET
CLBNZ
ETHYL CL
2-CLEVE
CHLOROFORM
METHYL CL
2BRCLMETH
11-2CLETH
12-2CLEIH
11-2CLETHE
12-2CLETHE
12-2CLPRP
C13-2CLPRE
T13-2CLPRP
ETHYLBEN2
METHYLE CL
4CLETHAN
4CLETHE
TOLUENE
111-3CLETH
112-3CLETH
3CLETHE
VINYL CL
Quantitation
Ion (m/z)
56
53
78
83
173
94
117
112
64
63
83
50
129
63
62
96
96
63
75
75
106
84
83
164
92
97
97
130
62
Secondary
lon(s)
55
51, 52
--
85. 129
171. 175
%
119, 121
114
66
65. 106
85
52
206. 208. 127
65, 83
64. 98
61. 98
61, 98
65. 114
71
11
91
86
85. 168
129. 131. 166
91
99. 117. 119
83, 85. 99
95, 9/, 132
64
-------
TABLE II-2. BFB MASS-INTENSITY SPECIFICATION
Mass Intensity Required
SO 15-401 of mass 95
75 30-601 of mass 95
95 Base peak,
1001 relative abundance
96 5-9% of mass 95
173 <2X of mass 174
174 >50I of mass 95
175 5-9% of mass 174
176 >95X but <10U of mass 174
177 5-9% of mass 176.
11-31
-------
TABLE 11-3. PERCENT SPIKE RECOVERIES FOR VOLATILE
PRIORITY POLLUTANTS USING VACUUM DISTILLATION*
Average Percent Average Percent
Spiking Compound Recovery (Water)b Recovery (Sediment)c
Chloromethane
Bromomethane
Vinyl chloride
Chloroetnane
Methylene chloride
1,1-dichloroethene
1,1-dichloro ethane
trans-l.Z-dichloroethene
Chloroform
1,2-dichloroethane
1, ltl-tricnloroethane
Carbon tetrachloride
Acrylomtrile
Bromodichloromethane
1,2-dichloropropane
trans-1.3-dichloropropene
Tnchloroethene
Benzene
Oibromochlorome thane
1,1,2-tr ichloroethane
ci_s-l,3-dichloropropene
Bromoform
Tetrachloroethene
1,1,2.2-tetrachloroethane
Toluene
Chlorobenzene
Ethylbenzene
2-chloroethyl vinyl ether
Acrolein
105 * 22
UO " 23
83 " 12
103 " 16
126 " 22
98 * 5
96 * 5
98 * 5
93 * 8
93 " 10
104 * 9
102 ~ 10
85 * 13
108 " 10
104 7 7
109 " 9
105 * 9
106 * 7
102 * il
95 * 8
109 * 9
104 + 14
105 " 9
90 7 9
106 * 7
101 7 7
103 * 5
94 * 50
113 7 76
98 * 22
86 ~ 24
108 * 35
106 7 27
LCd~
82 * 9
101 « 7
92 7 10
102 * 11
96 * 17
106 * 11
100 " 13
89 * 3
96 < 8
96 ~ 4
91 " 6
98 " 6
94 T 4
98 ~ 10
98 ~ 5
92 ~ 7
90 ~ 9
104 " 13
98 7 8
102 * 4
101 " 5
97 7 5
--
NAe
Average compound recovery 102 ^ 8 96 ^ 7
a From references 1 and 2.
*> Reagent water was spiked with 25 ug/L of each compound except acrolein
and acrylonitrile. which were added at 100 ug/L. The recoveries are averaged
from 9 analyses and were calculated by comparing vacuum extracted determinations
to determinations for which spikes were added directly to a purge-and-trap
device.
c Ten-gram sediment samples were spiked at 25 ppb. The recoveries were
averaged from 9 analyses witn three matrix types.
4 Laboratory contamination prevented the generation of valid data.
e Compound was not added to this matrix.
11-32
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SECTION III
ANALYSIS OF METALS AND METALLOIDS
IN ESTUARIHE AND MARINE SEDIMENTS
-------
CONTENTS
1.0 SCOPE AND APPLICATION III-l
2.0 SUMMARY OF METHOD HI-Z
3.0 DEFINITIONS 1 1 1 -2
4.0 INTERFERENCES II 1-3
5.0 SAFETY HI -4
6.0 APPARATUS AND EQUIPMENT UI-5
7.0 REAGENTS AND CONSUMABLE MATERIALS I II -7
8.0 SAMPLE COLLECTION, PREPARATION, AND STORAGE III-8
9.0 CALIBRATION AND STANDARDIZATION I II -9
10.0 QUALITY CONTROL III-ll
11.0 PROCEDURE 111-18
12.0 CALCULATIONS II 1-21
13.0 PRECISION AND ACCURACY HI-21
14.0 REFERENCES HI-21
-------
ANALYSIS OF METALS AND METALLOIDS IN
ESTUARINE AND MARINE SEDIMENTS
1.0 SCOPE AND APPLICATION
1.1 This method is designed to determine antimony, arsenic, beryllium,
cadmium, chromium, copper, lead, mercury, nickel, selenium, silver, thallium,
and zinc in sediments and dredged materials. These procedures are applicable
when sensitive analyses are required to monitor concentration differences
between relatively uncontaminated reference areas and contaminated estuarine
and marine environments.
1.2 A universal wet oxidation procedure (acid digestion) is recommended
that is capable of providing a clean extract suitable for analysis by atomic
absorption spectrophotometry (AAS). This digestion has proven effective
when determining most of the priority pollutant metals listed above (with
the possible exception of beryllium and thallium) (e.g., Table III-2).
Because of a lack of reference materials certified for beryllium and thallium,
little is known regarding method suitability for these elements.
1.3 The proposed method involves a rigorous acid digestion that most probably
extracts metal phases not available to biota (in addition to biologically
available phases). However, the silicate matrix of the sediment will not be
decomposed. Because of this, any element tightly bound as a naturally
occurring silicate may not be fully recovered (a total metals digestion
would include hydrofluoric add).
1.4 Typical limits of detection (LOD) are presented in Table III-l. These
vary depending upon the element measured, method of detection, and instrument
sensitivity.
III-l
-------
2.0 SUMMARY OF METHOD
2.1 A representative sample of sediment is homogenized wet, subsampled, and
digested using a *et oxidation method. The resulting extract is analyzed
for the metals of interest using various atomic absorption (AA) techniques
such as:
direct aspiration (DFAA) = for higher concentration metals
graphite furnace (GFAA) » for lower concentration metals
hydride generation (HYOAA) * for hydride forming elements (antimony,
arsenic, selenium)
cold vapor (CVAA) » for mercury.
Descriptions of these techniques may be found in references 1 through 5.
2.2 Alternative methods of detection may be used providing their performance,
limitations, and applicability have been established and approved by U.S. EPA.
Inductively coupled plasma (ICP) emission spectrometry may be used for
routine metal analyses not requiring the generally lower detection limits
attainable by graphite furnace atomic absorption.
3.0 DEFINITIONS
Certified Reference Materials (CRM): A homogeneous sample that has
been analyzed a sufficient number of times by numerous qualified laboratories.
The data are compiled and certified values are determined through statistical
analysis. A number of CRM are commercially available in a wide range of
matrices for metals analyses. For sediments, representative examples include
National Bureau of Standards river sediment (SRM 11645) and estuarine sediment
(SRM 11646) (references 6 and 7) and National Research Council of Canada
marine sediments MESS-1 and BCSS-1 (reference 8).
Control Standard: A solution, independent of the calibration standards,
whose analyte concentration is known. These are often analyzed as an external
check after calibration.
III-2
-------
Limit of Detection (LOO): The LOO is the lowest concentration level
that can be determined to be statistically different from a blank. The
recommended value for LOO is 3o , *here o is the standard deviation of the
blank in replicate analyses (reference 9).
Matrix Modifier: A reagent added to a sample that alters some asoect
of its composition (references 10-12).
4.0 INTERFERENCES
4.1 Interferences should be considered to be any chemical or physical
phenomenon that can influence the accuracy of the data during an analytical
operation. These can have either a positive or a negative effect on the
result depending on their nature.
4.2 Contamination of the sample can occur during any stage of collection,
handling, storage, or analysis. Potential contaminant sources must be known
and steps should be taken to minimize or eliminate them. Some of the most
common sources of contamination include prolonged exposure of the sediment
to fumes and dust containing metals; insufficiently clean sample containers,
storage facilities, and testing apparatus; as well as the use of contaminated
reagents during analysis (reference 13).
4.3 Most instrumental methods are prone to matrix interferences, which can
either suppress or enhance the analyte signal. If a matrix interference is
suspected. Its effect should be determined and corrective action taken. A
common first course of corrective action is the method of standard addition
(MSA). Details of the technique are provided in Sect. 10.4.2 (adapted from
reference 14). Some common matrix Interferences are listed below, along
with suggested corrective measures (references 15, 16).
4.3.1 Matrix products - spectral interferences can occur due to light
scattering by products of the atomlzatlon process (e.g., refractory oxides).
Flame temperature or fuel-to-ox1dant ratio can be varied to minimize the
III-3
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effect. Alternately, if the source of the interference is known, an excess
of the interferent (radiation buffer) can be added to the sample and standards
(reference 15).
4.3.2 Non-specific absorption (light scatter) - usually due to dissolved
solids or suspended particulates present in the sample prior to atomization,
which absorb analyte radiation. Background correction (e.g., continuous
source deuterium lamp, Zeeman effect) should be used whenever this occurs.
4.3.3 Interelement interference sediments contain elements in widely
varying concentration ranges. In some cases, a trace component being sought
may have its primary absorption line close to the absorption or emission
line of a major component. If this occurs, an interference is observed
proportional to the concentration of the interfering element. A secondary
absorption line that is not affected may be used to overcome this problem.
4.4 Chemical interferences - some of which are poorly understood, can occur
during instrumental analysis of the sample extracts and are a particular
problem for GFAA. A great many of these interferences have been addressed
in the literature and in most cases a sample pretreatment or instrumental
modification has been proposed as a remedy. A review of the recent literature
is provided by reference 17.
5.0 SAFETY
Laboratory personnel should be well versed in standard laboratory
safety practices. It is the responsibility of all staff and management to
ensure that safety training 1s mandatory. The laboratory is resonsible for
maintaining a current awareness file of OSHA regulations regarding the safe
handling of the chemicals specified in this method. A reference file of
data on handling chemicals safely should also be made available to all
personnel involved in these analyses. Additional information on laboratory
safety can be found in references 18-20.
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5.1 Chemicals and reagents should be properly labeled and stored in an area
appropriate to their properties. Any reagents whose composition or properties
may change with time must 5e dated and properly disposed of on or before the
expiration date.
5.2 Areas where strong oxidizing agents and flammable or explosive materials
are used should be well labeled and the necessary restrictions should be
imposed.
5.3 Where laboratory apparatus and instrumentation are used, the manu-
facturer's safety precautions should be strictly followed.
5.4 Wearing of safety clothing such as lab coats, gloves, and eye protection
should be mandatory when working with or around potentially dangerous equipment.
5.5 Contaminated sediments (Including dredged materials) can contain levels
of substances that may be hazardous. Anyone handling these samples should
be aware of this and take the necessary precautions.
6.0 APPARATUS ANO EQUIPMENT
6.1 Sample Containers wide-mouth, screw-cap jars made of either borosi 1 icate
glass or noncontaminatlng plastic (Mnear or high-density polyethylene, or
equivalent). Quartz or tetrafluoroethylene (TFE) containers are preferred
but may be prohibitively expensive. All containers should be prerinsed with
dilute acid and distilled deionlzed water (DOW) as described in Sect. 10.7.
6.2 Homogenizing Vessel - a plastic or glass container large enough to mix
the entire sample. A plastic spatula or glass stirring rod will be used to
homogenize the sediment. All sampling and subsampllng tools should be
rinsed with dilute add and DOW, as described in Sect. 10.7, between each
sample and subsample.
6.3 Digestion Vessels - 125-mL borosllIcate glass Erlenmeyer flasks equipped
with all glass reflux caps (Tuttle covers). Tuttle covers or equivalent
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reflux caps are essential for preventing evaporative loss of volatile compounds
or elements during high-temperature digestion. They are commercial ly available
(e.g.. Fisher Scientific) and are easily produced from borosilicate test
tubes.
6.4 Hot Plate - a thermostatically controlled plate with a range of 75 to
400° C.
6.5 Fumehood - a properly constructed hood capable of withstanding acid
fumes. It must be equipped with an exhaust fan having sufficient capacity
to remove all fumes and should be constructed of noncontaminating materials
(e.g., PVC), if possible.
6.6 Atomic Absorption Spectrophotometer (AAS).
6.6.1 The AAS must have sufficient sensitivity and stability to perform
within the specifications required by the method (Sect. 11). The instrument
should have automatic background correction, direct aspiration flame, as
well as flameless capabilities. The Instrument must have a routine maintenance
program to ensure proper performance and trouble-free operation. All source
lamps should be handled with care and the exit windows kept free of dust and
fingerprints. Periodic intensity and stability checks of the lamps should
be made. Any lamp showing signs of deterioration should be replaced (refer-
ence 4).
6.6.2 A graphite furnace (also called carbon rod) attachment for the
AAS is recommended when determining most elements in the low concentration
ranges. Most, 1f not all, AAS manufacturers offer this equipment as an
accessory. The stability and sensitivity afforded by the furnace is typically
one to two orders of magnitude better than direct aspiration (reference 21).
6.6.3 In addition to the graphite furnace, another flameless attachment
can be used in conjunction with the AAS to determine the hydride-forming
elements (arsenic, antimony, and selenium). Most such attachments may also
be used to analyze for mercury using the cold vapor technique. These methods
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are preferable to the graphite furnace since they vaporize the analyte from
the samole matrix prior to detection.
6.6.4 If available, a dedicated mercury monitor may be used for the
determination of mercury using the cold vapor technique. These units are
typically designed to give maximum sensitivity required for low-level
determinations.
6.7 ICP Emission Spectrometer (optional) - ICP emission spectroscopy enables
one to make simultaneous multielement analyses. The ICP instrument must
have sufficient sensitivity and stability to perform within the specifications
required by the method. Certain elements of interest are not amenable to
ICP analysis of sediments due to volatility or spectral interferences (i.e., As,
Hg, Pb, Se. and Tl).
7.0 REAGENTS AND CONSUMABLE MATERIALS
The purity of all reagents used for trace metal determinatians is
extremely important. Reagents should be checked for purity prior to use to
confirm the absence of contamination (reference 13). American Chemical
Society (ACS) reagent grad'e acids are suitable for routine analyses. Low
level analyses may require Instra-analyzed grade acids (J.T. Baker Chemical
Company) or equivalent.
7.1 Distilled Oe1on1zed Water (DOW) - a water purified by distillation (or
equivalent) followed by conditioning with a mixed bed Ion exchanger. Such
units are commercially available and yield a water with a typical resistivity
of IS megohms/cm.
7.2 Hydrochloric Add - concentrated (351).
7.3 Hydroxylamine Hydrochloride [201 (w/v)]: - dissolve 20 g of American
Chemical Society (ACS) grade NHjOH'HCl 1n 100 ml of DOW. Store in a precleaned
glass or plastic bottle. Prepare weekly.
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7.4 Nitric Acid - concentrated (70%).
7.5 Sodium Borohydride, ACS Grade - granular or powder.
7.6 Sodium Hydroxide, ACS Grade - pellets or flakes.
7.7 Stannous Chloride [201 (w/v)] dissolve 20 g of ACS grade SnCl2 in 20
mL of concentrated hydrochloric acid. Warm gently until solution clears,
cool, and add DOW until the solution reaches a 100 ml volume. Store in a
precleaned glass or plastic bottle. Prepare fresh daily.
7.8 Stock Standard Solutions These standards (typically 1,000 ppm) can be
purchased as certified solutions or prepared from ACS-grade metal salts and
pure compounds. Suitable procedures for preparing stock solutions are well
documented (e.g., reference 22) and include the steps below.
7.8.1 Accurately weigh 1,000 mg of pure metal or metal equivalent of
the salt and dissolve in a minimum amount (usually about 20 ml) of an
appropriate acid. Once the reagent is dissolved, dilute the solution to
1,000 ml with OOU and store in a precleaned plastic bottle. The solution is
usually stable for at least a year but must be checked periodically against
an in-house control standard (Sect. 10).
8.0 SAMPLE COLLECTION, PREPARATION. AND STORAGE
8.1 Possible problems during sample collection include contamination from
the sampling device, airborne dust, engine exhaust, winches or steel cables,
cross-contamination from previous samples, or improper subsampling procedures.
Avoid using metal during sample collection, if possible. If metal must be
used, high-grade stainless steel is preferred.
8.2 A minimum sample size of 5 g (wet wt) is required for the analysis of
all priority pollutant metals. To allow for duplicates, spikes, and required
reanalysis, a minimum sample size of 50 g (wet wt) 1s recommended. To allow
III-8
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for mixing of the samole and possible nonrepresentative material, a 240 rnL
(8 oz) jar is recommended for collection.
8.3 Store samples in clean containers after collection and, if possible,
pack them in ice. Samples should be stored at -20° C. Although freezing is
not required for all U.S. EPA procedures (e.g., reference 23), it is recommended
to minimize potential alteration of analytes by microbes. Care should be
taken to prevent container breakage during freezing. Leave sufficient
headspace for water to expand and freeze the containers at an angle.
8.4 No recommended holding time for sediments has been established by
U.S. EPA. A holding time of 6 months (except for mercury samples, which
should be held a maximum of 30 days) is consistent with the holding time
required by U.S. EPA for water samples (reference 14).
9.0 CALIBRATION AND STANDARDIZATION
9.1 Calibration standards are prepared by serial dilutions of the stock
solutions. The acid matrix of the standards should be as closely matched to
the samples as possible. Mixed standards of more than one element may be
prepared only after their compatibility has been determined. Some common
mixed standards include, but may not be limited to, the following:
- Cd, Cu, Pb, N1, and In
As, Se, and Sb
9.1.1 Do not add an incompatible anion to a mixed or single element
standard. For example, adding chloride to a silver standard could form a
precipitate of silver chloride (AgCl).
9.1.2 Do not mix metals that are Incompatible 1n solution. For example,
lead and chromium may form a precipitate of lead chrornate (PbCrOd).
9.2 Concentration ranges of the standards should bracket those for the
samples to be analyzed. At least four analyses (one blank and three standards
III-9
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of increasing concentration) should be used to calibrate the instrument at
the beginning of each shift.
9.3 Stability of a calibration standard varies with element, acid matrix,
concentration, and presence of other elements. As a general rule, standards
should be continuously monitored and replaced when necessary. As a matter
of protocol, the following can be used as a guideline:
less than 0.1 ppm - prepare daily
0.1 to 1 ppm - prepare weekly
1.0 to 10 ppm - prepare monthly
10 to 100 ppm - prepare quarterly
100* ppm prepare yearly (at a minimum)
9.4 Initial Standardization - follow manufacturer's suggestions for standard-
izing instrument and check sensitivity performance with specifications. If
performance is acceptable, proceed with analysis; if not, refer to manu-
facturer's troubleshooting guide.
9.5 After standardizing the instrument, analyze an independent control
standard as a check. If the result is acceptable, proceed; otherwise,
troubleshoot calibration standards, control standard, or instrument.
9.6 Ongoing Calibration (reference 14) - the instrument should be tested
with a single point calibration every 2 h during an analysis run or at a
frequeny of 10 percent of the analyses, whichever is more frequent. A
calibration check must also be run after the last sample in a laboratory
shift. A standard concentration in the middle of the initial calibration
range should be used.
If the difference between the ongoing calibration result and the known
standard concentration is greater than +10 percent (or +20 percent for
mercury analysis), the instrument must be recalibrated and the preceding 10
samples reanalyzed for the analytes affected.
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9.7 In the event that a sample is outside of the linear response of the
instrument, it must be diluted to within range or reanalyzed using a less
sensitive setup. This is commonly accomplished by calibrating the instrument
with higher concentration standards using a secondary or tertiary wavelength
with less sensitivity.
10.0 QUALITY CONTROL [see reference U and Quality Assurance/Quality
Control (QA/QC) for 301(h) Monitoring Programs: Guidance on Field and
Laboratory Methods (Tetra Tech 1986).]
A quality control program enables the assessment of the precision and
accuracy of data. Precision is estimated by analysis of replicates. Accuracy
is estimated by the analysis of blanks, spiked samples,.and laboratory
control samples (reference 24).
10.1 Replicates can be chosen to reflect the precision of most stages of
the overall analytical method. Replicates can consist of different subsamples
of a sediment homogenate or replicate instrumental analyses of the same
digestion extract.
10.1.1 Replicate analyses of sediment subsamples are important because
"the greatest potential for sample deterioration and/or contamination occurs
during preanalysis steps of sample collection, handling, preservation, and
storage" (reference 23).
10.1.2 Replicate analyses of a digestate focus only on the bench
chemistry and instrumental variability of the method. Together with replicate
analysis of sediment subsamples, they can be used to assess the impact of
each stage on the overall precision of the analytical result.
10.1.3 At least one replicate (a subsample of a sediment homogenate)
must be analyzed from each group of samples of a similar matrix type and
concentration for each batch of samples or for each 20 samples, whichever is
more frequent. If two analytical methods are used for the same element in a
batch of samples, duplicates must be run by each method used.
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The relative oercent differences (RPO) for each component are calculated
as follows:
' °
where
DI » first sample value.
03 3 second sample value.
10.2 As in the case of replicates, clanks can be chosen to address most
stages of the overall analytical method. They include transportation,
cross-contamination, reagent, and calibration blanks.
10.2.1 Transportation blanks are derived from empty containers that
have been stored with samples in the field and carried with them to the
laboratory. A small amount of 5 percent (v/v) HN03 is used to rinse the
inside of the container. The acid rinse is then retained for analysis.
Transportation blanks serve as estimates of contamination during preanalysis
steps (Sect. 10.1.1).
10.2.2 Cross-contamination blanks are used to estimate concentration
from sampling and homogenizing utensils that may carry over from one sample
to the next. They are prepared by collecting a final rinse after cleaning
utensils. The final rinse should be performed with a known volume of 5
percent (v/v) HN03. One cross-contamination blank should be analyzed for
each batch of samples.
10.2.3 Reagent (preparation) blanks are aliquots of 5 percent (v/v)
HN03 that are processed through each sample preparation step (e.g., reagent
addition, digestion, dilution). At least one reagent blank must be prepared
for each batch of samples or for every 20 samples, whichever is more frequent.
Reagent blanks serve as estimators of contamination resulting from the
chemical analysis steps.
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All samples *ith at least one analyte concentration that is less than
10 times the corresponding concentration in the associated reagent blank
roust be redigested and reanalyzed.
10.2.4 Calibration blanks consist of 5 percent (v/v) HN03 and are
analyzed each time the instrument is calibrated, at the beginning of each
analysis run, and at a frequency of 10 percent during a run. Calibration
blanks are used to ensure that the analytical instrument is not introducing
false positive results during analysis. (Ongoing calibration quality assurance
is discussed in Sect. 9.6.)
10.3 The results obtained from the reagent blanks can be used to calculate
the LOO (Sect. 3) for the method. This is the assigned minimum value above
which reliable data can be reported. Results for at least the reagent blank
should be reported with the final data set.
10.4 Spiked samples are samples to which small volumes of standard solutions
of the elements of Interest have been added. Spiked samples provide a means
of assessing losses during digestion, distillation, or other pretreatment
steps. The spike is added before the pretreatment steps and should be 0.5
to 2.0 times the concentrations of the elements in the sample. At least one
spiked sample must be analyzed for each batch of samples of a similar matrix
type and concentration or for each 20 samples, whichever is more frequent.
10.4.1 The percent recovery for each element is calculated as follows:
I Rpcoverv - fsp1ke * sample result) - (sample result) IQ(J
* cugve J (spike added)
Spike percent recoveries should not be used to determine a correction factor
to compensate for losses.
10.4.2 If graphite furnace AA 1s used, a single analytical spike is
required after any digestion steps to determine if the method of standard
additions (MSA) 1s required (reference 14 was used to develop this section).
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The spike should be added at a concentration (in the sample) that is
twice the LOO. The unsoiked sample aliquot must be compensated for any
volume change in the spiked samples by addition of DOW to the unspiked
sample aliquot. The percent recovery of the spike should be calculated as
in Sect. 10.4.1
10.4.2.1 If the sample absorbance or concentration is >50 percent
of the spike^ and the spike recovery is between 85 and 115 percent, the
sample should be quantified directly from the calibration curve.
10.4.2.2 If the spike recovery is less than 40 percent, the
sample must be diluted and rerun with another spike. Dilute the sample by a
factor of 5 to 10 and rerun. This step must only be performed once. If
after dilution the spike recovery is still <40 percent, there are inter-
ferences associated with the instrumental technique that prevent GFAA analysis
of the sample.
10.4.2.3 If the spike recovery is >40 percent and the sample
absorbance or concentration is <50 percent of the spike , report the analyte
as less than the LOO or less than the LOO times the dilution factor if the
sample was diluted.
10.4.2.4 If the sample absorbance or concentration is >50 percent
of the spike and the spike recovery is <85 or >115 percent, the sample must
be quantified by MSA.
10.4.2.5 The following procedures should be incorporated into MSA
analyses.
that spike1 is defined throughout Sect. 10.4.2 as (absorbance or
concentration of spike sample) minus (absorbance or concentration of the
sample.]
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a) Data from MSA calculations must be within the linear range as
determined by the calibration curve generated at the beginning
of the analytical r-jn.
b) The samole and three spikes must be analyzed consecutively
for MSA quantitation (the "initial" spike run data is speci-
fically excluded from use in the MSA quantitation).
c) Spikes (post-digestion, as for the "initial" spike in Sect.
10.4.2) should be prepared such that:
Spike 1 is approximately 50 percent of the sample
absorbance.
Spike 2 is approximately 100 percent of the sample
absorbance.
Spike 3 1s approximately 150 percent of the sample
absorbance.
d) The data for each MSA analysis should be clearly identified
in the raw data documentation along with the slope, intercept
and correlation coefficient (r) for tire least squares fit of
the data.
10.5 Laboratory control samples are certified reference materials (CRM)
submitted blind to the laboratory. CRM provide an estimate of the accuracy
of the overall method. A CRM must be chosen that has a similar matrix to
samples and contains all the analytes. CRM can be purchased from a number
of agencies and are available for a variety of sediments (see Sect. 3.0). A
catalog of CRM (reference 7) 1s available from the National Bureau of Standards,
Office of Standard Reference Materials, Room B311, Chemistry Building,
National Bureau of Standards, Washington, DC 20234 (301/921-2045). Information
on the National Research Council Canada CRMs (reference 8) 1s available from
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Marine Analytical Chemistry Standards Program, Division of Chemistry, National
Research Council, Montreal Soad, Ottawa, Canada, K1A-OR9 (613/993-9101).
10.5.1 Unlike an analyte spike (Sect. 10.4), a CRM tests the dissolution
technique as rfell as instrument calibration and matrix interferences.
10.5.2 A minimum of one CRM should be analyzed for each survey or 2
percent of the total number of samples (i.e., 1 per 50 samples), whichever
is more frequent).
10.5.3 The percent recovery for each element for the overall method is
calculated as follows:
I Recovery » x 100
where
x » the analytical result for the element
REF » the certified result for the element.
The data obtained for each reference material should be used to troubleshoot
the method if results fall outside the acceptable range (i.e., the 95 percent
confidence interval). Percent recovery values should not be used to determine
a correction factor to compensate for apparent procedural losses.
10.6 Maintenance of Records the data obtained from any QC work should be
recorded in an organized manner to allow for easy retrieval and reviewing.
If sufficient data have been collected, it 1s recommended that these be
plotted on a control chart for a quick visual assessment. A typical control
chart for CRM results 1s presented 1n Figure III-l.
10.6.1 The quality control chart can be used to determine if the
following recommended guidelines are met:
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10,6.1.1 Not more than 5 percent of the results lie outside two
standard deviations (warning 1 irm't). A result outside three standard deviations
requires action.
10.6.1.2 There are no regular periodic variations.
10.7 Cleaning and preparation of labware is an integral part of a quality
assurance/quality control (QA/QC) program. Many cleaning procedures have
been proposed in the literature that are suitable for decontaminating equip-
ment. The main concerns with cleaning are removing elements of interest
from labware while maintaining an inactive surface. Some cleaning procedures
tend to be too harsh, producing an surface with an ion exchange capacity.
In this case a solution could partially or completely 'lose" an analyte to
the container walls (references 25, 26).
10.7.1 When analyte concentrations vary by orders of magnitude, it is
best to use dedicated labware; I.e.. relatively high-concentration samples
-hould have their own labware that 1s never used for low-concentration
samples. This helps avoid cross-contamination (carryover).
10.7.2 A good universal cleaning procedure for glass and plasticware
is outlined below.
10.7.2.1 Wash labware with a metal-free detergent and warm water.
10.7.2.2 Rinse at least three times with tap water followed by
distilled deIonized water (DOW).
10.7.2.3 Soak equipment or labware in a dilute acid (25 percent
HNO]) bath for 24 h. If possible, the bath should be maintained at an
elevated temperature (70° C).
10.7.2.4 Rinse labware with large volumes of DDW and use imme-
diately. If a time lapse must exist, the apparatus should be stored under
ust-free conditions and rinsed further with DOW prior to use.
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NOTES: Change the acid batch periodically such that no significant
buildup of metals occurs.
At no time should a metal-containing reagent such as chromic
acid be used.
10.8 Round Robin or Interlaboratory Check Programs - In addition to the
quality control measures discussed above, all laboratories should participate
in interlaboratory check programs (see Part II of Exhibit E in reference U).
11.0 PROCEDURE
II.1 Homogenize samples prior to analysis to ensure that representative
aliquots are taken. Place the entire sample into the homogenizing vessel
and blend with a plastic spatula or glass rod. Mix any water that has
separated from the sediment back into the sample. Remove and make note of
nonrepresentative material (e.g., twigs, leaves, shells, rocks, and any
material larger than 0.25 in).
11.2 Analyze a separate aliquot of sediment for moisture content.
11.2.1 Weigh a small aluminum drying dish to the nearest 0.1 mg (0).
11.2.2 Add approximately 2-3 g of homogenized sediment to the dish and
reweigh (A).
11.2.3 Dry sediment at 103° C overnight, cool in a dessicator, and
reweigh (B).
11.2.4 Calculate percent moisture as follows:
I H20 - A_-£ x 100
A-D
11.3 Accurately weigh a 5-g (wet) aliquot of homogenized sediment to the
nearest 0.1 mg. Transfer the weighed sediment to a precleaned 125-mL Erlenmeyer
111-18
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flask equipped with an all-glass reflux cap. Analyze a sufficient number of
reagent blanks, sample duplicates, analyte spikes, and certified reference
materials concurrently (Sect. 10).
11.4 Slowly add 5 ml of concentrated nitric acid followed by 10 ;rl of
concentrated hydrochloric acid. If foaming occurs during acid addition,
swirl the flasks while adding 2- to 3-mL increments. Allow flasks to stand
at room temperature for approximately 15 hours in a dust-free ventilated
environment. Periodically swirl the flasks to ensure adequate mixing of the
sediment and acid.
11.5 After 15 hours, gently heat the flask to approximately 100° C and hold
at this temperature for one hour. Gradually increase the temperature in
50° C increments to a maximum of 250° C. Continue heating until all reddish
brown fumes have disappeared and organic matter has been digested. This
usually takes about 4 hours. If large amounts of organic matter remain,
additional nitric acid should be added in 2- to 3-mL increments and heating
should be continued until the organic matter has been consumed. Do not rush
the initial digestion as losses of volatile elements will likely occur.
Once digestion is complete, cool flasks to room temperature.
NOTE: Most hotplates do*not have a uniform temperature over the entire
surface. Rotate flasks as required to ensure that all samples digest in
approximately the same time.
11.6 When the digestion 1s complete, rinse the reflux caps with OOW and
combine the rinse with the extract 1n the flask. Transfer the extract to a
precleaned 100-mL volumetric flask. Rinse the Erlenmeyer flask three times
with OOU and combine with the extract 1n the volumetric flask. Adjust the
volume to 100 ml with OOW and transfer to a precleaned plastic bottle.
NOTE: Some elements are not as stable as others 1n solution and therefore
should be analyzed first. Stability can be determined by dally analysis of
the extracts. However, the following can be used as a guideline:
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Sb, Pb, Hg, Se and Ag - analyze within 1 day
As and Cd - analyze within 2 days
Cr, Cu, Ni and Zn analyze within I week
3e and T1 - to be determined.
11.7 Instrumental analysis The extracts will be analyzed using various
techniques of atomic absorption spectrophotometry (AAS) or atomic emission
spectrophotometry. The method of choice depends on instrument availability,
analyte concentration, and sample matrix. In some instances it may be
useful to use more than one method to confirm a result.
11.7.1 Follow the manufacturer's instructions for initial setup and
calibrate as outlined in Sect. 9 of this method. As every instrument responds
uniquely to a given set of conditions, it is the analyst's responsibility to
develop the optimum set of parameters. Use calibration standards and CRM to
ensure that optimum conditions exist.
11.7.2 Table III-l lists some general information for each of the
priority pollutant metals.
11.7.3 It is possible to use alternate methods of detection providing
they have been validated using a sufficient number of previously analyzed
samples or CRM.
11.8 All data generated must be clearly recorded on a strip chart or printer,
or manually logged in prepared tables. The order 1n which the extracts are
analyzed should be the same as 1t appears in the records. The data, when
assembled, should be reported 1n consistent units (i.e., mg/L) to avoid
errors when calculating the final results (ug/g). The final report should
contain all necessary methods, results, quality control data (e.g., reagent
blank values), and limits of detection for each element. The report must
clearly state if any data were blank-corrected.
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12.0 CALCULATIONS
12.1 All results are reported as micrograms of element per dry gram of
sediment:
ug/g ELEMENT » C x Y
(dry weight basis) W (1-M)
where:
C * concentration (may be blank corrected) of element in final extract
(ug/mL)
V » volume of final extract (ml)
W * weight of wet sediment (g)
M » sediment moisture expressed as a decimal.
Reagent blank corrections may be made and blank values must always be reported.
13.0 PRECISION AND ACCURACY
In order to estimate precision and accuracy (single lab, multi-operator),
a number of CRM and analyte spikes were analyzed using this method. Table
III-2 summarizes typical data obtained. No data are currently available for
either beryllium or thallium.
14.0 REFERENCES
1. Ebdon, L. 1982. An introduction to atomic absorption spectroscopy: a
self-teaching approach. Heyden, London.
2. Oittrlch, K. 1982. Atomic absorption spectrometry. Scientific Pocket-
books, Vol. 276: Chemistry series. Akad-Verlag, Berlin.
3. Cresser, M.S. and B.L. Sharp (eds). 1981. Annual reports on analytical
spectroscopy. The Royal Society of Chemistry, London.
4. Cantle, J.E. (ed). Techniques and Instrumentation in analytical chemistry,
Volume 5: Atomic absorption spectrometry. Elsevier, Amsterdam.
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5. Slavin, W. 1984. Graphite furnce AAS a source book. Perkin-Elmer
Corp, Ridgefield, CT.
6. Taylor, J.K. 1985. Standard reference materials: handbook for SRM
users. National Bureau of Standards Special Publication 260-100.
National Bureau of Standards, Washington, DC.
7. Hudson, C.H. (ed). 1984. NBS standard reference materials catalog.
1984-1985. National Bureau of Standards Special Publication 260.
National Bureau of Standards, Washington, OC.
8. National Research Council Canada. 1981. Marine sediment reference
materials. National Research Council Canada, Division of Chemistry,
Marine Analytical Chemistry Standards Program, Ottawa, Canada.
9. Keith, L.J., W. Crummet, J. Oeegan, Jr., R.A. Libby, J.K. Taylor, and
G. Wentler. 1983. Principles of environmental analysis. Anal.
Chem. 55:2210-2218.
10. Manning, O.C., and W. Slavin. 1983. The determination of trace elements
in natural waters using the stabilized temperature platform furnace.
Applied Spectroscopy 37:1-11.
11. Hinderberger, E.J., M.L. Kaiser, and S.R. Kolrtyohann. 1981. Furnace
atomic absorption analysis of biological samples using the L'vov platform
and matrix modification. Atomic Spectroscopy 2:1-7.
12. Sturgeon, R.E., S.N. WIlHe, and S.S. Berman. 1985. Preconcentration
of selenium and antimony from seawater for determination by graphite
furance atomic absorption spectrometry. Anal. Chem. 57:6-9.
13. Murphy, T.J. 1976. The role of the analytical blank in accurate trace
analysis, pp. 509-539. In: Accuracy in Trace Analysis: Sampling,
Sample Handling, and Analysis. National Bureau of Standards Special
Publication 422. National Bureau of Standards, Washington, OC.
14. U.S. Environmental Protection Agency. 1985. U.S. EPA Contract Laboratory
Program - statement of work for inorganic analyses, multi-media multi-
concentration.
15. Skoog, D.A. 1985. Principles of Instrumental Analysis. Saunders,
Philadelphia, PA. pp. 270-279.
16. Veil Ion, C. 1976. Optical atomic spectroscopic methods, pp. 123-181.
In: Trace Analysis: Spectroscopic Methods for Elements. 0. Winefordner
(ed). Wiley. New York.
17. Slavin, W., and D.C. Manning. 1982. Graphite furnace interferences, a
guide to the literature. Prog. Anal. Atomic Spectroscopy 5:243-340.
18. Carcinogens - working with carcinogens. OHEW, PHS, CDC, NIOSH. Publica-
tion 77-206 (Aug. 1977).
111-22
-------
19. OSHA safety and health standards, general industry. OSHA 2206, 29 CFR
1910 (revised Jan. 1976).
20. Safety in academic chemistry laboratories. ACS Publications, Committee
on Chemical Safety, 3rd Edition (1979).
21. Fuller, C.W. 1978. Electrothermal atomization for atomic absorption
spectroscopy. The Chemical Society, London.
22. U.S. Environmental Protection Agency. 1979. Methods for chemical
analysis of water and wastes, pp. 202.1-289.2. U.S. Environmental
Protection Agency Environmental Monitoring and Support Laboratory.
Cincinnati, OH.
23. Plumb, R.H., Jr. 1981. Procedures for handling and chemical analysis
of sediment and water samples. Technical Report EPA/CE-81-1. Environ-
mental Protection Agency/Corps of Engineers Technical Committee on
Criteria for Dredged and Fill Material, U.S. Army Waterways Experiment
Station, Vlclcsburg. MS. 471 pp.
24. U.S. Environmental Protection Agency. 1983. Guidance for preparation
of combined work/quality assurance project plans for water monitoring.
Office of Water Regulations and Standards, U.S. EPA, Washington. OC.
33 pp.
25.- Batley, G.E., and 0. Gardner. 1977. Sampling and storage of natural
waters for trace metal analysis. Water Res. 44:745-756.
26. Laxen, O.P.H., and R.M. Harrison. 1981. Cleaning methods for polythene
containers prior to the determination of trace metals in freshwater
samples. Anal. Chem. 53:345-350.
111-23
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o
UJ
ffi
o
0]
U
TIME SCALE
X * 3S
x * 2S
CERTIFIED MEAN (x)
X - 2S
K - 3S
8 ± 2S = WARNING LIMIT
(95% CONFIDENCE)
X i 3S = ACTION LIMIT
Figure III-l. Quality control chart.
-------
TABLE III-l. GENERAL INFORMATION FOR EACH PRIORITY POLLUTANT METAL
E 1 e*nei :
Antimony
Arsenic
Beryl 1 mm
Cadmium
Chromium
Copper
Lead
Mercury
Nickel
Selenium
Silver
Thai 1 lum
Zinc
1
HYOAA
GFAA
ICP
HYOAA
GFAA
GFAA
OFAA
GFAA
OFAA
GFAA
ICP
OFAA
GFAA
ICP
OFAA
GFAA
CVAA
OFAA
GFAA
ICP
HYOAA
GFAA
OFAA
GFAA
ICP
GFAA
OFAA
ICP
Wavelength
(nm)
217.6
217.6
193. 7
193.7
234.9
228.8
228.3
357.9
357.9
324.7
324.7
383.3
383.3
253.6
232.0
232.0
197.3
197.3
328.1
328.1
276.8
213.9
1.3.0.2
O.I
0.1
3.0
O.I
0.1
0.05
O.I
0.1
0.2
0.02
6.0
0.1
0.01
0.6
1.0
0.1
0.01
0.5
0.02
1.5
0.01
0.1
0.1
0.1
0.7
0.1
1.0
0.2
Signal
Peak Area
Peak Height
Peak Area
Peak Height
Peak Height
Direct
Peak Height
Direct
Peak Height
Direct
Peak Height
Direct
Peak Height
Peak Height
Direct
Peak Height
Peak Area
Peak Height
Direct
Peak Height
Peak Height
Direct
Notes3
Requires a Matrix Modifier
[e.g.. Ni(N03,2]
Requires a Matrix Modifier
(e.g.. NMdM2p04)
Requires a Matrix Modifier
[e.g.. N1(NO])2]
I HYOAA Hydride generation atomic adsorption.
GFAA Graphite furnace atonic adsorption.
OFAA Direct flame atomic adsorption.
CVAA Cold vapor atonic adsorption.
ICP Inductively coupled plasma.
2 I.0.0. limit of detection - mlcrogram of element per dry gran of sediment (ppm) dased
on 5 g (wet) to 100 me. The Unit was determined as twice the standard deviation of a
repeated series of Blanks (n-5 to 10; 95X confidence level).
3 For example, see reference 10 for discussion of these matrix modifiers.
DOES codes for all elements are tht element n
111-25
-------
TABLE ::;-z. TYPICAL DATA OBTAINED ON A CERTIFIED REFERENCE MATERIAL
.NATIONAL RESEARCH COUNCIL OF CANADA MARINE SEDIMENT (MESS -I)]
Element
Antimony
Arsenic
Beryl 1 im
Cadmium
Chromium
Copper
Lead
Mercury
Nickel
Selenium
Si 1 ver
Thai 1 ium
Zinc
Certified/Spued
(x *_ S.O.-j
5b
As
3e
Cd
Cr
Cu
Pb
Hg
Ni
Se
Ag
Tl
Zn
0.73
10.6
1.9-
0.59
71.
25.1
34.0
0.171
29.5
> 0.08
1 1-2
* 0.2
* 0.10
1 ll-
1 3.8
1 6.1
* 0.014
± 2-7
(0.4)
50.0
N.
191.
(spike)
C.3
1 l7-
n
5
20
--
20
10
20
20
20
10
5
5
20
Found
(x ^S.D.)
0.61
9.43
No
0.58
30.
22.5
29.7
0.185
25.5
0.33
55.0
No
176.
* 0.
* 0.
Data
±°-
* 2.
1 °-
09
32
07
0
71
1 1.8
* 0.
015
1 2.1
1 °-
02
^ 6.0
Data
*_ 4.5
Detection
Met nod 2
HYOAA
HYDAA
--
GFAA
DFAA
DFAA
DFAA/GFAA4
CVAA
OFAA
HYDAA
OFAA
--
DFAA
1 All results expressed as micrograms of element per gram of sediment.
2 HYOAA = Hydride generation atomic absorption.
GFAA = Graphite furnace atomic absorption.
DFAA = Direct flame atomic absorption.
CVAA = Cold vapor atomic absorption.
3 N.C. = Not Certified.
4 GFAA may require dilution.
111-26
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