United States Office of Water EPA-821-B-98-016
Environmental Protection Washington, DC 20460 July 1998
Agency
SEPA Analytical Methods for the Determination
of Pollutants in Pharmaceutical
Manufacturing Industry Wastewater
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Acknowledgments
This methods compendium was prepared under the direction of William A. Telliard of the Engineering and
Analysis Division within EPA's Office of Water. This document was prepared under EPA Contract No.
68-C3-0337 by DynCorp Environmental with the assistance of Interface, Inc.
Disclaimer
This methods compendium has been reviewed by the Engineering and Analysis Division, U.S.
Environmental Protection Agency, and approved for publication. Mention of trade names or commercial
products does not constitute and endorsement or recommendation for use.
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Table of Contents
Introduction
IV
Method 1666
Volatile Organic Compounds Specific to the Pharmaceutical Manufacturing Industry by Isotope
Dilution GC/MS
Revision A, July 1998 1
Method 1667
Formaldehyde, Isobutyraldehyde, and Furfural by Derivatization Followed by High
Performance Liquid Chromatography
Revision A, July 1998 53
Method 1671
Volatile Organic Compounds Specific to the Pharmaceutical
Manufacturing Industry by GC/FID
Revision A, July 1998 75
July 1998
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Introduction
The U.S. Environmental Protection Agency (EPA) has promulgated effluent limitations guidelines and
standards at 40 CFR part 439 for the Pharmaceutical Manufacturing Industry to control the discharge of
pollutants into surface waters of the United States. This compendium of test procedures (methods) supports
the final rule. These methods and methods promulgated at 40 CFR part 136 are used for filing permit
applications and for compliance monitoring under the National Pollutant Discharge Elimination System
(NPDES) program.
This compendium includes only those methods unique to the Pharmaceutical Manufacturing
Industry. Other methods allowed under the proposed rule have been promulgated at 40 CFR part 136.
Questions concerning the methods in this compendium should be directed to:
W.A. Telliard
U.S. EPA
Engineering and Analysis Division
Office of Science and Technology
401 M Street, SW
Washington, DC 20460
July 1998 iv
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Method 1666
Volatile Organic Compounds Specific to the Pharmaceutica
Manufacturing Industry by Isotope Dilution GC/MS
Revision A, July 1998
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Method 1666, Revision A
Volatile Organic Compounds Specific to the Pharmaceutical Manufacturing
Industry by Isotope Dilution GC/MS
1.0 Scope and Application
1.1 This method is for surveying and monitoring under the Clean Water Act. It is used to determine
certain volatile organic pollutants specific to the pharmaceutical manufacturing industry (PMI) that
are amenable to purge-and-trap gas chromatography/mass spectrometry (GC/MS) or direct
aqueous injection GC/MS.
1.2 The PMI analytes listed in Tables 1 and 2 may be determined in waters, soils, and municipal
sludges by this method or the method referenced at the end of that table.
1.3 The detection limits of the method are usually dependent on the level of interferences rather than
instrumental limitations. The minimum levels (MLs) in Tables 3 and 4 are the level that can be
attained with no interferences present.
1.4 The GC/MS portions of this method are for use only by analysts experienced with GC/MS or
under the close supervision of such qualified persons. Laboratories unfamiliar with analysis of
environmental samples by GC/MS should run the performance tests in Reference 1 before
beginning.
1.5 This method is performance-based. The analyst is permitted to modify the method to overcome
interferences or to lower the cost of measurements, provided that all performance criteria in this
method are met. The requirements for establishing method equivalency are given in Section 9.1.2.
2.0 Summary of Method
2.1 Purge-and-trap GC/MS.
Stable, isotopically labeled analogs of the compounds of interest are added to the sample and the
sample is purged with an inert gas at 45 °C in a chamber designed for soil or water samples, as
appropriate. In the purging process, the volatile compounds are transferred from the aqueous
phase into the vapor phase, where they are passed into a sorbent column and trapped. After
purging is completed, the trap is backflushed and heated rapidly to desorb the compounds into a
gas chromatograph (GC). The compounds are separated by the GC and detected by a mass
spectrometer (MS) (References 2 and 3).
2.2 Direct aqueous injection.
Certain volatile, water-soluble organic compounds do not purge well from water and are analyzed
by direct aqueous injection.
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Method 1666, Revision A
2.2.1 The percent solids content of the sample is determined. If the solids content is known or
determined to be less than 1%, stable, isotopically labeled analogs of the compounds of
interest are added to a 5-mL sample. If the solids content of the sample is greater than 1%,
5 mL of reagent water and the labeled compounds are added to a 5-g aliquot of sample.
The mixture is sonicated in a centrifuge vial with little or no headspace for 5 minutes.
During this period the native analytes and labeled analogs will equilibrate between the
solid and aqueous phases. In some cases, additional sonication will be necessary to
establish equilibrium. The resulting suspension is centrifuged and the supernatant liquid
analyzed.
2.2.2 One (iL or more of the aqueous solution (or supernate) is injected into the GC/MS. The
compounds are separated by the GC and detected by the MS (References 2 and 3). The
labeled compounds serve to correct the variability of the analytical technique.
2.3 Identification of a pollutant (qualitative analysis) is performed by calibrating the GC/MS with
authentic standards and storing a mass spectrum and retention time for each compound in a
user-created library. A compound is identified when its retention time and mass spectrum agree
with the library retention time and spectrum.
2.4 Quantitative analysis is performed in one of two ways by using extracted-ion current profile
(EICP) areas. (1) For those compounds listed in Table 1 and Table 2, and for other compounds for
which labeled analogs are available, the GC/MS system is calibrated and the compound
concentration is determined using an isotope dilution technique. (2) For those compounds listed in
Table 1 and Table 2, and for other compounds for which authentic standards but no labeled
compounds are available, the GC/MS system is calibrated and the compound concentration is
determined using an internal standard technique.
2.5 The quality of the analysis is assured through reproducible calibration of the GC/MS system.
3.0 Definitions
There are no definitions unique to this method.
4.0 Interferences
4.1 Impurities in the purge gas, organic compounds outgassing from the plumbing upstream of the
trap, and solvent vapors in the laboratory account for the majority of contamination problems
encountered with this method. The analytical system is demonstrated to be free from interferences
under conditions of the analysis by analyzing reagent water blanks initially and with each sample
batch (samples analyzed on the same 12-hour shift), as described in Section 9.5.
4.2 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 may serve as a check on such
contamination.
July 1998 4
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Method 1666, Revision A
4.3 Contamination by carryover can occur when high-level and low-level samples are analyzed
sequentially. To reduce carryover, the purging device (Figure 1 for samples containing less than
1% solids; Figure 2 for samples containing 1% solids or greater) in purge-and-trap analysis or the
syringe in direct aqueous injection analysis is cleaned or replaced with a clean purging device or
syringe after each sample is analyzed. When an unusually concentrated sample is encountered, it is
followed by analysis of a reagent water blank to check for carryover. Purging devices and syringes
are cleaned by washing with soap solution, rinsing with tap and distilled water, and drying in an
oven at 100-125 °C. The trap and other parts of the system are also subject to contamination;
therefore, frequent bakeout and purging of the entire system may be required.
4.4 Interferences resulting from samples will vary considerably from source to source, depending on
the diversity of the site being sampled.
5.0 Safety
5.1 The toxicity or carcinogenicity 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. Exposure to these compounds should be reduced to the lowest possible level. The
laboratory is responsible for maintaining a current awareness file of OSF£A regulations regarding
the safe handling of the chemicals specified in this method. A reference file of material safety data
sheets should also be made available to all personnel involved in these analyses. Additional
information on laboratory safety can be found in References 5 through 7.
6.0 Equipment and Supplies
Disclaimer: The mention of trade names or commercial products in this Method is for
illustrative purposes only and does not constitute endorsement or recommendation for use by the
Environmental Protection Agency. Equivalent performance may be achievable using apparatus,
materials, or cleaning procedures other than those suggested here. The laboratory is
responsible for demonstrating equivalent performance.
6.1 Sample bottles and septa.
6.1.1 Bottle—25- to 40-mL with screw-cap (Pierce 13075, or equivalent). Detergent wash,
rinse with tap and distilled water, and dry at > 105 °C for a minimum of 1 hour before use.
6.1.2 Septum-Polytetrafluoroethylene (PTFE)-faced silicone (Pierce 12722, or equivalent),
cleaned as above and baked at 100-200 °C for a minimum of 1 hour.
6.2 Purge-and-trap device-Consists of purging device, trap, and desorber.
6.2.1 Purging devices for water and soil samples.
6.2.1.1 Purging device for water samples-Designed to accept 5-mL samples with water
column at least 3 cm deep. The volume of the gaseous headspace between the
5 July 1998
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Method 1666, Revision A
water and trap shall be less than 15 mL. The purge gas shall be introduced less
than 5 mm from the base of the water column and shall pass through the water as
bubbles with a diameter less than 3 mm. The purging device shown in Figure 1
meets these criteria.
6.2.1.2 Purging device for solid samples-Designed to accept 5 g of solids plus 5 mL of
water. The volume of the gaseous head space between the water and trap shall be
less than 25 mL. The purge gas shall be introduced less than 5 mm from the base
of the sample and shall pass through the water as bubbles with a diameter less
than 3 mm. The purging device shall be capable of being controlled at a
temperature of 45±2 °C while the sample is being purged. The purging device
shown in Figure 2 meets these criteria.
6.2.2 Trap-25-30 cm long x 2.5 mm i.d. minimum, containing the following:
6.2.2.1 Methyl silicone packing-l±0.2 cm, 3% OV-1 on 60/80 mesh Chromosorb W, or
equivalent.
6.2.2.2 Porous polymer-15± 1.0 cm, Tenax GC (2,6-diphenylene oxide polymer), 60/80
mesh, chromatographic grade, or equivalent.
6.2.2.3 Silica gel-8± 1.0 cm, Davison Chemical, 35/60 mesh, grade 15, or equivalent. The
trap shown in Figure 3 meets these specifications.
6.2.3 Desorber-Shall heat the trap to 175±5 °C in 45 seconds or less. The polymer section of
the trap shall not exceed a temperature of 180°C and the remaining sections shall not
exceed 220 °C during desorb, and no portion of the trap shall exceed 225 °C during
bakeout. The desorber shown in Figure 3 meets these specifications.
6.2.4 The purge-and-trap device may be a separate unit or coupled to a GC, as shown in Figures
4 and 5.
6.3 Gas chromatograph-Shall be linearly temperature programmable with initial and final holds, and
shall produce results that meet the calibration (Section 10), quality assurance (Section 9), and
performance tests (Section 15) of this method.
6.3.1 Column for purge-and-trap analyses-60 m long x 0.32 mm i.d., fused-silica microbore
column coated with 1.5 (im of phenylmethyl polysiloxane (Restek RTX-Volatiles, or
equivalent).
6.3.2 Column for direct aqueous injection analyses-30 m long x 0.32 mm i.d. fused-silica
microbore column coated with 1.5 (im of 95% dimethyl- 5% diphenyl polysiloxane
specially passivated for chromatography of amines (Restek RTX-5 Amine, or equivalent).
6.3.3 GC operating conditions.
July 1998 6
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Method 1666, Revision A
6.3.3.1 Purge-and-trap-4 minutes at 0 ° C, 8 ° C per minute to 170 ° C. Helium carrier gas
at 1.5 mL per minute.
6.3.3.2 Direct aqueous injection-4 minutes at 40 °C, 8 °C per minute to 100 °C, then 25 °C
to 220°C with a 3-minute hold at 220°C. Helium carrier gas at 1.5 mL per
minute. A pre-column split may be used to achieve acceptable peak shape.
6.4 Mass spectrometer-70 electron volt (eV) electron-impact ionization; shall repetitively scan from 20
to 250 Dalton every 2 to 3 seconds, and produce a unit resolution (valleys between m/z 174 to 176
less than 10% of the height of the m/z 175 peak), background-corrected mass spectrum from 50 ng
4-bromofluorobenzene (BFB) injected into the GC. The BFB spectrum shall meet the
mass-intensity criteria in Table 5. All portions of the GC column, transfer lines, and separator that
connect the GC column to the ion source shall remain at or above the column temperature during
analysis to preclude condensation of less volatile compounds.
6.5 Data system-Shall 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.
6.5.1 Data acquisition-Mass spectra shall be collected continuously throughout the analysis and
stored on a mass-storage device.
6.5.2 Mass spectral libraries-User-created libraries containing mass spectra obtained from
analysis of authentic standards shall be employed to reverse search GC/MS runs for the
compounds of interest (Section 10.2).
6.5.3 Data processing-The data system shall be used to search, locate, identify, and quantify the
compounds of interest in each GC/MS analysis. Software routines shall be employed to
compute retention times and EICP areas. Displays of spectra, mass chromatograms, and
library comparisons are required to verify results.
6.5.4 Response factors and multipoint calibrations-The data system shall be used to record and
maintain lists of response factors (response ratios for isotope dilution) and generate
multi-point calibration curves (Section 10.4). Computations of relative standard deviation
(coefficient of variation) are useful for testing calibration linearity. Statistics on initial and
ongoing performance shall be maintained (Sections 9 and 10).
6.6 Syringes-5-mL glass hypodermic, with Luer-lok tips.
6.7 Micro syringes-10-, 25-, and 100-(iL.
6.8 Syringe valves-2-way, with Luer ends (PTFE).
6.9 Syringe-5 -mL, gas-tight, with shut-off valve.
6.10 Bottles-15-mL, screw-cap with PTFE liner.
7 July 1998
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Method 1666, Revision A
6.11 Balances.
6.11.1 Analytical, capable of weighing 0.1 mg.
6.11.2 Top-loading, capable of weighing 10 mg.
6.12 Equipment for determining percent moisture.
6.12.1 Oven, capable of temperature control at 110±5 °C.
6.12.2 Desiccator.
6.12.3 Beakers-50-to 100-mL.
6.13 Centrifuge apparatus.
6.13.1 Centrifuge capable of rotating 10-mL centrifuge tubes at 5000 rpm.
6.13.2 Centrifuge tubes, 10-mL, with screw-caps to fit centrifuge.
6.14 Sonication apparatus capable of sonicating 10 mL centrifuge tubes and thoroughly agitating
contents.
7.0 Reagents and Standards
7.1 Reagent water-Water in which the compounds of interest and interfering compounds are not
detected by this method. It may be generated by any of the following methods.
7.1.1 Activated carbon-Pass tap water through a carbon bed (Calgon Filtrasorb-300, or
equivalent).
7.1.2 Water purifier-Pass tap water through a purifier (Millipore Super Q, or equivalent).
7.1.3 Boil and purge-Heat tap water to 90-100 °C and bubble contaminant-free inert gas
through it for approximately 1 hour. While still hot, transfer the water to screw-cap
bottles and seal with a PTFE-lined cap.
7.2 Sodium thiosulfate-ACS granular.
7.3 Methanol-Pesticide-quality or equivalent.
7.4 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
compound purity is 96% or greater, the weight may be used without correction to calculate the
concentration of the standard.
July 1998 8
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Method 1666, Revision A
7.5 Preparation of stock solutions for purge-and-trap analysis.
7.5.1 Place approximately 9.5 mL of methanol in a 10-mL ground-glass-stoppered volumetric
flask. Allow the flask to stand unstoppered for approximately 10 minutes or until all
methanol-wetted surfaces have dried. In each case, weigh the stoppered flask, add the
compound, restopper, then immediately reweigh to prevent evaporation losses from
affecting the measurement.
7.5.2 Using a 100-(iL 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 microsyringe.
7.5.3 Fill the flask to volume, stopper, then mix by inverting several times. Calculate the
concentration in milligrams per milliliter (mg/mL; equivalent to micrograms per microliter
[|ig/(iL]) from the weight gain.
7.5.4 Transfer the stock solution to a PTFE-sealed screw-cap bottle. Store, with minimal
headspace, in the dark at -20 to -10 °C.
7.5.5 Replace standards after one month, or sooner if comparison with check standards indicate
a change in concentration. Quality control check standards that can be used to determine
the accuracy of calibration standards may be available from the National Institute of
Standards and Technology, Gaithersburg, Maryland.
7.6 Preparation of stock solutions for direct aqueous injection analysis.
7.6.1 Place approximately 9.0 mL of reagent water in a 10-mL ground-glass-stoppered
volumetric flask. Allow the flask to stand unstoppered for approximately 10 minutes or
until all wetted surfaces have dried. In each case, weigh the stoppered flask, add the
compound, restopper, then immediately reweigh to prevent evaporation losses from
affecting the measurement.
7.6.2 Using a microsyringe, add sufficient liquid (about 100 mg) so that the final solution will
have a concentration of about 10 mg/mL.
7.6.3 Fill the flask to volume, stopper, then mix by inverting several times. Calculate the
concentration in milligrams per milliliter (mg/mL; equivalent to micrograms per microliter
[|ig/(iL]) from the weight gain.
7.6.4 Transfer the stock solution to a PTFE-sealed screw-cap bottle. Store, with minimal
headspace, in the dark at approximately 4°C. Do not freeze.
7.6.5 Replace standards after one month, or sooner if comparison with check standards indicate
a change in concentration. Quality control check standards that can be used to determine
July 1998
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Method 1666, Revision A
the accuracy of calibration standards may be available from the National Institute of
Standards and Technology, Gaithersburg, Maryland.
7.7 Labeled compound spiking solutions.
7.7.1 Purge-and-trap analysis-From stock standard solutions (Section 7.5), or from mixtures,
prepare the spiking solution to contain a concentration of labeled compound such that a 5-
to 10-(iL spike into each 5-mL sample, blank, or aqueous standard analyzed will result in
a concentration of 50 (ig/L of each compound with a minimum level (ML) of 20 (ig/L or
less, a concentration of 500 (ig/L for each compound with an ML of 100 or 200 (ig/L, and
a concentration of 1 mg/L for each compound with an ML of 500 (ig/L (see Table 3).
Include the internal standards (Section 10.4.2) in this solution, if appropriate, so that a
concentration of 50 (ig/L in each sample, blank, or aqueous standard will be produced.
7.7.2 For direct aqueous injection-From stock standard solutions (Section 7.6), or from
mixtures, prepare the spiking solution to contain a concentration such that a 50- to
100-(iL spike into each sample, blank, or aqueous standard analyzed will result in a
concentration of 1 mg/mL of each labeled compound. Include the internal standard in this
solution so that a concentration of 1 mg/mL will be produced.
7.8 Secondary standards-Using stock solutions, prepare a secondary standard in methanol or water, as
appropriate, to contain each pollutant at a concentration of 1 mg/mL, or 2.5 mg/mL for compounds
with higher MLs.
7.8.1 Aqueous calibration standards-Using a microsyringe, add sufficient secondary standard
(Section 7.8) to five reagent water aliquots to produce concentrations in the range of
interest.
7.8.2 Aqueous performance standard-An aqueous standard containing all pollutants, internal
standards, labeled compounds, and BFB is prepared daily, and analyzed each shift to
demonstrate performance (Section 15). This standard shall contain concentrations of
pollutants, labeled compounds, BFB, and internal standards, as appropriate, within a
factor of 1-5 times the MLs of the pollutants listed in Table 3 or 4. It may be one of the
aqueous calibration standards described in Section 7.8.1.
7.8.3 A methanolic standard containing all pollutants specific to this method (Table 1) and
internal standards is prepared to demonstrate recovery of these compounds when syringe
injection and purge-and-trap analyses are compared. This standard shall contain either
100 (ig/mL or 500 (ig/mL of the PMI analytes, and 100 (ig/mL of the internal standards
(consistent with the amounts in the aqueous performance standard in Section 7.8.2).
7.8.4 Other standards that may be needed are those for test of BFB performance (Section 10.1)
and for collection of mass spectra for storage in spectral libraries (Section 10.1.1).
July 1998 10
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Method 1666, Revision A
8.0 Sample Collection, Preservation, and Storage
8.1 Grab samples are collected in glass containers having a total volume greater than 20 mL. For
aqueous samples that pour freely, fill sample bottles so that no air bubbles pass through the sample
as the bottle is filled and seal each bottle so that no air bubbles are entrapped. Maintain the
hermetic seal on the sample bottle until time of analysis.
8.2 Samples are maintained at 0-4°C from the time of collection until analysis. If an aqueous sample
contains residual chlorine, add sodium thiosulfate preservative (10 mg/40 mL) to the empty sample
bottles just prior to shipment to the sample site. EPA Methods 330.4 and 330.5 may be used for
measurement of residual chlorine (Reference 8). If preservative has been added, shake the bottle
vigorously for 1 minute immediately after filling.
8.3 For aqueous samples, experimental evidence indicates that some PMI analytes are susceptible to
rapid biological degradation under certain environmental conditions. Refrigeration alone may not
be adequate to preserve these compounds in wastewaters for more than seven days. For this
reason, a separate sample should be collected, acidified, and analyzed when compounds susceptible
to rapid biological degradation are to be determined. Collect about 500 mL of sample in a clean
container. Adjust the pH of the sample to about 2 by adding hydrochloric acid (1:1) while stirring.
Check pH with narrow range (1.4-2.8) pH paper. Fill a sample bottle as described in Section 8.1.
If residual chlorine is present, add sodium thiosulfate to a separate sample bottle and fill as in
Section 8.1.
8.4 All samples shall be analyzed within 14 days of collection.
9.0 Quality Assurance/Quality Control
9.1 Each laboratory that uses this method is required to operate a formal quality assurance program
(Reference 9). 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.
Laboratory performance is compared to established performance criteria to determine if the results
of analyses meet the performance characteristics of the method.
9.1.1 The analyst shall make an initial demonstration of the ability to generate acceptable
accuracy and precision with this method. This ability is established as described in
Section 9.2.
9.1.2 In recognition of advances that are occurring in analytical technology, and to allow the
analyst to overcome sample matrix interferences, the analyst is permitted certain options to
improve separations or lower the costs of measurements. These options include alternative
concentration and cleanup procedures, and changes in columns and detectors. Alternative
techniques, such as the substitution of spectroscopy or immunoassay, and changes that
degrade method performance, are not allowed. If an analytical technique other than the
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Method 1666, Revision A
techniques specified in this method is used, that technique must have a specificity equal to
or better than the specificity of the techniques in this method for the analytes of interest.
9.1.2.1 Each time a modification is made to this method, the analyst is required to repeat
the procedure in Section 9.2. If the detection limit of the method will be affected
by the change, the laboratory is required to demonstrate that the MDL (40 CFR
part 136, Appendix B) is lower than one-third the regulatory compliance level. If
calibration will be affected by the change, the analyst must recalibrate the
instrument per Section 10.
9.1.2.2 The laboratory is required to maintain records of modifications made to this
method. These records include the information below, at a minimum.
9.1.2.2.1 The names, titles, addresses, and telephone numbers of the
analyst(s) who performed the analyses and modification, and of
the quality control officer who witnessed and will verify the
analyses and modification.
9.1.2.2.2 A listing of pollutant(s) measured, by name and CAS Registry
Number.
9.1.2.2.3 A narrative stating the reason(s) for the modification.
9.1.2.2.4 Results from all quality control (QC) tests comparing the
modified method to this method, including:
(a) Calibration (Section 10)
(b) Calibration verification (Section 15)
(c) Initial precision and accuracy (Section 9.2)
(d) Labeled compound recovery (Section 9.3)
(e) Analysis of blanks (Section 9.5)
(f) Accuracy assessment (Section 9.4)
9.1.2.3 Data that will allow an independent reviewer to validate each
determination by tracing the instrument output (peak height, area, or other
signal) to the final result, including:
(a) Sample numbers and other identifiers
(b) Analysis dates and times
(c) Analysis sequence/run chronology
(d) Injection logs
(e) Sample weight or volume
(f) Sample volume prior to each cleanup step, if applicable
(g) Sample volume after each cleanup step, if applicable
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Method 1666, Revision A
(h) Final sample volume prior to injection (Sections 11 and
12)
(i) Injection volume (Sections 11 and 12)
(j) Dilution data, differentiating between dilution of a sample
or an extract (Section 16.4)
(k) Instrument and operating conditions
(1) Column (dimensions, liquid phase, solid support, film
thickness, etc.)
(m) Operating conditions (temperature, temperature program,
flow rates, etc.)
(n) Detector (type, operating condition, etc.)
(o) Chromatograms, printer tapes, and other recording of raw
data
(p) Quantitation reports, data system outputs, and other data
necessary to link raw data to the results reported.
9.1.3 Analyses of blanks are required to demonstrate freedom from contamination and that the
compounds of interest and interfering compounds have not been carried over from a
previous analysis (Section 4.3). The procedures and criteria for analysis of a blank are
given in Section 9.5.
9.1.4 The laboratory shall spike all samples with labeled compounds to monitor method
performance. This test is described in Section 9.3. When results of these spikes indicate
atypical method performance for samples, the samples are diluted to bring method
performance within acceptable limits (Section 16).
9.1.5 The laboratory shall, on an ongoing basis, demonstrate through the analysis of the aqueous
performance standard (Section 7.8.2) that the analysis system is in control. This
procedure is described in Sections 15.1 and 15.5.
9.1.6 The laboratory shall maintain records to define the quality of data that is generated.
Development of accuracy statements is described in Sections 9.4 and 15.5.2.
9.2 Initial precision and accuracy—To establish the ability to generate acceptable precision and
accuracy, the analyst shall perform the following operations for compounds to be calibrated:
9.2.1 Analyze two sets of four 5-mL aliquots (eight aliquots total) of the aqueous performance
standard (Section 7.8.2) containing Table 1 PMI analytes by purge-and-trap. Or, for
Table 2 PMI analytes, analyze two sets of four aliquots (eight aliquots total) by direct
aqueous injection.
9.2.2 Using results of the first set of four analyses in Section 9.2.1, compute the average
recovery (X) in percent of spike level and the standard deviation of the recovery (s) in
percent of spike level, for each compound, by isotope dilution for pollutants with a labeled
13 July 1998
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Method 1666, Revision A
analog, and by internal standard for labeled compounds and pollutants with no labeled
analog.
9.2.3 For each compound, compare s and X with the corresponding limits for initial precision
and accuracy found in Table 6. If s and X for all compounds meet the acceptance criteria,
system performance is acceptable and analysis of blanks and samples may begin. If,
however, any individual s exceeds the precision limit or any individual X falls outside the
range for accuracy, system performance is unacceptable for that compound
Note: The large number of compounds in Table 6 presents a substantial probability that one or more
will fail one of the acceptance criteria when all compounds are analyzed. To determine if the
analytical system is out of control, or if the failure can be attributed to probability, proceed as
follows.
9.2.4 Using the results of the second set of four analyses, compute s and X for only those
compounds that failed the test of the first set of four analyses (Section 9.2.3). If these
compounds now pass, system performance is acceptable for all compounds, and analysis
of blanks and samples may begin. If, however, any of the same compounds fail again, the
analysis system is not performing properly for the compound(s) in question. In this event,
correct the problem and repeat the entire test (Section 9.2.1).
9.3 The laboratory shall spike all samples with labeled compounds to assess method performance on
the sample matrix.
9.3.1 Spike and analyze each sample according to the appropriate method in Section 11 or 12.
9.3.2 Compute the percent recovery (P) of the labeled compounds using the internal standard
method (Section 10.4.2).
9.3.3 Compare the percent recovery for each compound with the corresponding labeled
compound recovery limit in Table 6. If the recovery of any compound falls outside its
warning limit, method performance is unacceptable for that compound in that sample.
Therefore, the sample matrix is complex and the sample is to be diluted and reanalyzed,
per Section 16.
9.4 As part of the QA program for the laboratory, it is suggested but not required that method
accuracy for wastewater samples be assessed and records maintained. After the analysis of five
wastewater samples for which the labeled compounds pass the tests in Section 9.3.3, compute the
average percent recovery (P) and the standard deviation of the percent recovery (sp) for the labeled
compounds only. Express the accuracy assessment as a percent recovery interval from P - 2sp to P
+ 2sp. For example, if P = 90% and sp = 10%, the accuracy interval is expressed as 70-110%.
Update the accuracy assessment for each compound on a regular basis (e.g., after each five to ten
new accuracy measurements).
July 1998 14
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Method 1666, Revision A
9.5 Blanks-Reagent water blanks are analyzed to demonstrate freedom from carryover and
contamination (Section 4).
9.5.1 The level at which the purge-and-trap system will carry greater than the ML of a pollutant
of interest (Table 1) into a succeeding blank shall be determined by analyzing successively
larger concentrations of these compounds. When a sample contains this concentration or
more, a blank shall be analyzed immediately following this sample to demonstrate no
carryover at the ML.
9.5.2 With each sample batch (samples analyzed on the same 12-hour shift), a blank shall be
analyzed immediately after analysis of the aqueous performance standard (Section 15.1) to
demonstrate freedom from contamination. If any of the compounds of interest (Table 1 or
2) or any potentially interfering compound is found in a blank at greater than the ML
(assuming a response factor of 1 relative to the nearest-eluted internal standard for
compounds not listed in Tables 1 and 2), analysis of samples is halted until the source of
contamination is eliminated and a blank shows no evidence of contamination at this level.
All results must be associated with an uncontaminated method blank.
9.6 The specifications contained in this method can be met if the apparatus used is calibrated properly,
then maintained in a calibrated state. The standards used for calibration (Section 7), calibration
verification (Section 15.5), and initial (Section 9.2) and ongoing (Section 15.5) precision and
accuracy should be identical, so that the most precise results will be obtained. The GC/MS
instrument in particular will provide the most reproducible results if dedicated to the settings and
conditions required for the analyses of volatiles by this method.
9.7 Depending on specific program requirements, field replicates may be collected to determine the
precision of the sampling technique, and spiked samples may be required to determine the accuracy
of the analysis when the internal-standard method is used.
10.0 Calibration and Standardization
Calibration of the GC/MS system is performed by direct aqueous injection (Section 10.3) or purging the
compounds of interest and their labeled analogs from reagent water at the temperature to be used for
analysis of samples (Section 10.2).
10.1 Assemble the GC/MS apparatus and establish the operating conditions to be used for sample
analysis (Section 6.3.3.1 or Section 6.3.3.2). By injecting standards into the GC, demonstrate that
the analytical system meets the minimum levels in Tables 3 or 4 for the compounds for which
calibration is to be performed, and the mass-intensity criteria in Table 5 for 50 ng BFB.
10.1.1 Mass-spectral libraries-Detection and identification of the compounds of interest are
dependent upon the spectra stored in user-created libraries.
10.1.1.1 For the compounds in Tables 1 and 2, and other compounds for which the
GC/MS is to be calibrated, obtain a mass spectrum of each pollutant and
15 July 1998
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Method 1666, Revision A
labeled compound and each internal standard by analyzing an authentic
standard either singly or as part of a mixture in which there is no
interference between closely eluted components. Examine the spectrum to
determine that only a single compound is present. 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 undistorted spectrum at the GC peak
maximum. An undistorted spectrum will usually 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 m/z's or introduce other
distortion.
10.1.1.2 The authentic reference spectrum is obtained under BFB tuning conditions
(Section 10.1 and Table 5) to normalize it to spectra from other
instruments.
10.1.1.3 The spectrum is edited by saving the five most intense mass-spectral
peaks and all other mass-spectral peaks greater than 10% of the base
peak. The spectrum may be further edited to remove common interfering
masses. If five mass-spectral peaks cannot be obtained under the scan
conditions given in Section 6.4, the mass spectrometer may be scanned to
an m/z lower than 20 to gain additional spectral information. The
spectrum obtained is stored for reverse search and for compound
confirmation.
10.2 Assemble the GC/MS apparatus and establish operating conditions given in Section
6.3.3.1. By injecting standards into the GC, demonstrate that the analytical system meets
the minimum levels in Table 3 for the compounds for which calibration is to be performed,
and the mass-intensity criteria in Table 5 for 50 ng BFB.
10.2.1 Assemble the purge-and-trap device. Pack the trap as shown in Figure 3 and
condition overnight at 170-180°C by backflushing with an inert gas at a flow rate
of 20-30 mL/min. Condition traps daily for a minimum of 10 minutes prior to
use.
10.2.1.1 Analyze the aqueous performance standard (Section 7.8.2)
according to the purge-and-trap procedure in Section 11.
Compute the area at the primary m/z (Table 7) for each
compound. Compare these areas to those obtained by injecting 1
(iL of the methanolic standard (Section 7.5.1) to determine
compound recovery. The recovery shall be greater than 50% for
the PMI analytes. Maximum allowable recovery for the PMI
analytes found in Table 1 are shown in Table 8. This recovery is
demonstrated initially for each purge-and-trap GC/MS system.
July 1998 16
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Method 1666, Revision A
The test is repeated only if the purge-and-trap or GC/MS systems
are modified in any way that might result in a change in recovery.
10.2.1.2 Demonstrate that a reliable calibration point can be established at
the ML for each compound (Table 2). If the MLs cannot be met,
adjust the analytical system until this performance is achieved.
10.3 Assemble the GC/MS system for direct aqueous injection and establish the operating conditions to
be used for sample analysis (Section 6.3.3.2). By injecting standards into the GC, demonstrate that
the analytical system meets the minimum levels in Table 4 for the compounds for which calibration
is to be performed, and the mass-intensity criteria in Table 5 for 50 ng BFB.
Demonstrate that 100 ng o-xylene (or o-xylene-d10) produces an area at m/z 106 (or 116)
approximately one-tenth that required to exceed the linear range of the system. The exact value
must be determined by experience for each instrument. It is used to match the calibration range of
the instrument to the analytical range and detection limits required.
10.4 The following calibration steps are to be performed for both the purge-and-trap PMI analytes
found in Table 1 and the direct aqueous injection PMI analytes found in Table 2, as appropriate.
10.4.1 Calibration by isotope dilution-The isotope dilution approach is used for the PMI analytes
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 method (Section 10.4.2) is used. A calibration curve encompassing the
concentration range of interest is prepared for each compound determined. The relative
response (RR) vs. concentration in micrograms per liter is plotted or computed using a
linear regression. An example of a calibration curve for o-xylene using o-xylene-d10 is
given in Figure 6. Also shown are the ±10% error limits (dotted lines). Relative response
is determined according to the procedures described below. A minimum of five data points
are required for calibration.
10.4.1.1 The relative response (RR) of pollutant to labeled compound is determined
from isotope ratio values calculated from acquired data. Three isotope
ratios are used in this process:
Rx = The isotope ratio measured in the pure pollutant (Figure 7A)
Ry = The isotope ratio of pure labeled compound (Figure IB)
Rm = The isotope ratio measured in the analytical mixture of the
pollutant and labeled compounds (Figure 7C)
The correct way to calculate RR is:
17 July 1998
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Method 1666, Revision A
RR =
If Rm is not between 2Ry and 0.5RX, the method does not apply and the
sample is analyzed by the internal standard method (Section 10.4.2).
10.4.1.2 In most cases, the retention times of the pollutant and labeled compound
are similar, and isotope ratios (R's) can be calculated from the EICP
areas, where:
(area at mj ^)
R =
If either of the areas is zero, it is assigned a value of 1 in the calculations;
that is, if area of m/z = 50721, and area of m2/z = 0, then R = 50721/1 =
50720.
The data from these analyses are reported to three significant figures (see
Section 14.6). In order to prevent rounding errors from affecting the
values to be reported, all calculations performed prior to the final
determination of concentrations should be carried out using at least four
significant figures. Therefore, the calculation of R above is rounded to
four significant figures.
The m/z's are always selected such that Rx > Ry. When there is a
difference in retention times (RT) between the pollutant and labeled
compounds, special precautions are required to determine the isotope
ratios.
Rx, Ry, and Rm are defined as follows:
[area mj ^ (at
R =
R =
[area mj ^ (at RTJ]
10.4.1.3 An example of the above calculations can be taken from the data plotted
in Figure 7 for o-xylene and o-xylene-d10. For these data:
July 1998 18
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Method 1666, Revision A
R = = 16890Q
R = —— = 0.00001640
J 60960
R = 96868 =
82508
10.4.1.4 The RR for the above data is then calculated using the equation given in
Section 10.4.1.1. For the example, rounded to four significant figures,
RR = 1.174. Not all labeled compounds elute before their pollutant
analogs.
To calibrate the analytical system by isotope dilution, analyze an aliquot
of each of the aqueous calibration standards (Section 7.8.1) spiked with
an appropriate constant amount of the labeled compound spiking solution
(Section 7.7), using the appropriate procedure in Section 10. Compute
the RR at each concentration.
10.4.1.5 Linearity-If the ratio of relative response to concentration for any
compound is constant (less than 20% 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 shall be used over the five-point calibration
range.
10.4.2 Calibration by internal standard-Used when criteria for isotope dilution (Section 10.4.1)
cannot be met. The method is applied to pollutants having no labeled analog and to the
labeled compounds. The internal standards used for volatiles analyses are
bromochloromethane, 1,4-difluorobenzene, chlorobenzene-d5, and tetrahydrofuran-d8.
Concentrations of the labeled compounds and pollutants without labeled analogs are
computed relative to the nearest eluting internal standard, as shown in Tables 3 and 4.
10.4.2.1 Response factors-Calibration requires the determination of response
factors (RF) which are defined by the following equation:
(A x C.)
RF = -^ ?-
(A x Q
Where:
As = The EICP area at the characteristic m/zfor the
compound in the daily standard
19 July 1998
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Method 1666, Revision A
Ais = The EICP area at the characteristic m/zfor the internal
standard
Cis = The concentration (pg/L) of the internal standard
Cs = The concentration of the pollutant in the daily standard
10.4.2.2 The RF is determined at 10, 20, 50, 100, and 200 (ig/L for the pollutants
(optionally at 5 times or more these concentrations for highly
water-soluble pollutants; see Section 7.8), in a way analogous to that for
calibration by isotope dilution (Section 10.4.1). To produce a calibration
curve, AS*C1S/A1S is plotted against concentration (Cs) for each compound.
10.4.2.3 Linearity-If the relative standard deviation of the RFs for any compound
is constant (less than 35%) over the five-point calibration range, an
averaged RF may be used for that compound; otherwise, the complete
calibration curve for that compound shall be used over the five-point
range.
10.4.3 Combined calibration-By adding the isotopically labeled compounds and internal standards
(Section 7.7) to the aqueous calibration standards (Section 7.8.1), 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 (Section 15.5) by analyzing the aqueous
performance standard (Section 7.8.2). Recalibration is required only if calibration and
ongoing performance (Section 15.5) criteria cannot be met.
11.0 Purge, Trap, and GC/MS Analysis
Samples containing less than 1% solids are analyzed directly as aqueous samples (Section 11.4). Samples
containing 1% solids or greater are analyzed as solid samples utilizing one of two methods, depending on
the levels of pollutants in the sample. Samples containing 1% solids or greater and low to moderate levels
of pollutants are analyzed by purging a known weight of sample added to 5 mL of reagent water (Section
11.5). Samples containing 1% solids or greater and high levels of pollutants are extracted with methanol,
and an aliquot of the methanol extract is added to reagent water and purged (Section 11.6).
11.1 Determination of percent solids.
11.1.1 Weigh 5 to 10 g of sample into a tared beaker.
11.1.2 Dry overnight (12 hours minimum) at 110°C (±5°C), and cool in a desiccator.
11.1.3 Determine percent solids as follows:
,, ,., might of sample dry . _ _
% solids = —2 + £ ^x 100
might of sample wet
11.2 Remove standards and samples from cold storage and bring to 20-25 °C.
July 1998 20
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Method 1666, Revision A
11.3 Adjust the purge gas flow rate to 40 mL/min (±4 mL/min).
11.4 Samples containing less than 1% solids.
11.4.1 Mix the sample by shaking vigorously. Remove the plunger from a 5-mL syringe and
attach a closed syringe valve. Open the sample bottle and carefully pour the sample into
the syringe barrel until it overflows. Replace the plunger and compress the sample. Open
the syringe valve and vent any residual air while adjusting the sample volume to 5±0.1 mL.
Because this process of taking an aliquot destroys the validity of the sample for future
analysis, fill a second syringe at this time to protect against possible loss of data.
11.4.2 Add an appropriate amount of the labeled compound spiking solution (Section 7.7.1)
through the valve bore, then close the valve.
11.4.3 Attach the syringe valve assembly to the syringe valve on the purging device. Open both
syringe valves and inject the sample into the purging chamber. Purge the sample per
Section 11.7.
11.5 Samples containing 1% solids or greater and low to moderate levels of pollutants.
11.5.1 Mix the sample thoroughly using a clean spatula and remove rocks, twigs, sticks, and
other foreign matter.
11.5.2 Weigh 5±1 g of sample into a purging vessel (Figure 2). Record the weight to three
significant figures.
11.5.3 Add 5±0.1 mL of reagent water to the vessel.
11.5.4 Using a metal spatula, break up any lumps of sample to disperse the sample in the water.
11.5.5 Add an appropriate amount of the labeled compound spiking solution (Section 7.7.1) to the
sample in the purge vessel. Place a cap on the purging vessel and shake vigorously to
further disperse the sample. Attach the purge vessel to the purging device, and purge the
sample per Section 11.7.
11.6 Samples containing 1% solids or greater and high levels of pollutants, or samples requiring dilution
by a factor of more than 100 (see Section 16).
11.6.1 Mix the sample thoroughly using a clean spatula and remove rocks, sticks, twigs, and
other foreign matter.
11.6.2 Weigh 5±1 g of sample into a calibrated 15- to 25-mL centrifuge tube. Record the weight
of the sample to three significant figures.
21 July 1998
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Method 1666, Revision A
11.6.3 Add 10 mL of methanol to the centrifuge tube. Cap the tube and shake it vigorously for
15 to 20 seconds to disperse the sample in the methanol. Allow the sample to settle in the
tube. If necessary, centrifuge the sample to settle suspended particles.
11.6.4 Remove approximately 0.1% of the volume of the supernatant methanol using a 15- to
25-(iL syringe. This volume will be in the range of 10-15 (iL.
11.6.5 Add this volume of the methanol extract to 5 mL reagent water in a 5-mL syringe, and
analyze per Section 10.4.1.
11.6.6 For further dilutions, dilute 1 mL of the supernatant methanol (Section 10.6.4) to 10 mL,
100 mL, 1000 mL, etc., in reagent water. Remove a volume of this methanol
extract/reagent water mixture equivalent to the volume in Section 10.6.4, add it to 5 mL
reagent water in a 5-mL syringe, and analyze per Section 11.4.
11.7 Purge the sample for 11±0.1 minute at 45±2°C.
11.8 After the 11-minute purge time, attach the trap to the chromatograph and set the purge-and-trap
apparatus to the desorb mode (Figure 5). Desorb the trapped compounds into the GC column by
heating the trap to 170-180°C while backflushing with carrier gas at 20-60 mL/min for 4 minutes.
Start MS data acquisition upon start of the desorb cycle, and start the GC column temperature
program 3 minutes later. Section 6.3.3.1 provides the recommended operating conditions for the
gas chromatograph. Table 3 provides the retention times and minimum levels that can be achieved
under these conditions. An example of the separations achieved by the column listed is shown in
Figure 8. Other columns may be used provided the requirements in Section 9 are met.
11.9 After desorbing the sample for 4 minutes, recondition the trap by purging with purge gas while
maintaining the trap temperature at 170-180°C. After approximately 7 minutes, turn off the trap
heater and stop the gas flow through the trap. When cool, the trap is ready for the next sample.
11.10 While analysis of the desorbed compounds proceeds, remove and clean the purge device. Rinse
with tap water, clean with detergent and water, rinse with tap and distilled water, and dry for a
minimum of 1 hour in an oven at a temperature greater than 15 0 ° C.
12.0 Direct Aqueous Injection and GC/MS Analysis
Samples containing less than 1% solids are analyzed directly as aqueous samples (Section 12.3). Samples
containing 1% solids or greater are analyzed after equilibration with reagent water containing labeled PMI
analytes and internal standards (Section 12.4).
12.1 Determine percent solids as in Section 11.1.
12.2 Remove standards and samples from cold storage and bring to 20-25 °C.
12.3 Samples containing less than 1% solids.
July 1998 22
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Method 1666, Revision A
12.3.1 Allow solids to settle and remove 5 mL of sample.
12.3.2 Add an appropriate amount of the labeled compound spiking solution (Section 7.7.2).
12.3.3 Inject 1 mL or more directly into the GC injection port. The temperature of the injection
block should be great enough to immediately vaporize the entire sample. An example of
the separations achieved by the column listed is shown in Figure 9.
12.4 Samples containing 1% solids or greater.
12.4.1 Mix the sample thoroughly using a clean spatula and remove rocks, twigs, sticks and other
foreign matter.
12.4.2 Add 5±1 g of sample to a 10-mL centrifuge tube. Using a clean metal spatula, break up
any lumps of sample. Record the sample weight to three significant figures.
12.4.3 Add an appropriate amount of the labeled compound spiking solution (Section 7.7.2) to the
sample in the centrifuge tube.
12 A A Add a measured quantity, Y, (to the nearest 0.1 mL) of reagent water to the tube so as to
minimize headspace.
12.4.5 Place a cap on the centrifuge tube leaving little or no headspace. Place the tube in the
sonicator for a minimum of 5 minutes, turning occasionally. For most samples this should
be sufficient time to distribute labeled and native analytes between the solid and aqueous
phases and to establish equilibrium. Some sample matrices may require more sonication.
12.4.6 On completion of sonication, centrifuge the sample and inject \\\L or more of supernate
directly into the GC injection port. The temperature of the injection block should be great
enough to immediately vaporize the entire sample.
12.5 Liquid samples containing high solids concentrations, such as sludges or mud, may be weighed into
a 10-mL centrifuge tube, have labeled compound spiking solution (Section 7.7.2) added, and be
sonicated as in Section 12.4.5. Centrifugation and injection are to be performed as in Section
12.4.6.
13.0 Qualitative Determination
Identification is accomplished by comparison of data from analysis of a sample or blank with data stored in
the mass-spectral libraries. For compounds for which the relative retention times and mass spectra are
known, identification is confirmed per Sections 13.1 and 13.2.
13.1 A labeled compound or pollutant having no labeled analog (Tables 1 and 2).
23 July 1998
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Method 1666, Revision A
13.1.1 The signals for all characteristic m/z's stored in the spectral library (Section 10.1.1) shall
be present and shall maximize within the same two consecutive scans.
13.1.2 Either (1) the background corrected EICP areas or (2) the corrected relative intensities of
the mass-spectral peaks at the GC peak maximum shall agree within a factor of 2 (0.5-2
times) for all m/z's stored in the library.
13.1.3 The relative retention time (RRT) shall be within ±10% of the RRT in the system
performance standard (Section 15.1)
13.2 Pollutants having a labeled analog (Tables 1 and 2).
13.2.1 The signals for all characteristic m/z's stored in the spectral library (Section 10.1.1) shall
be present and shall maximize within the same two consecutive scans.
13.2.2 Either (1) the background corrected EICP areas or (2) the corrected relative intensities of
the mass-spectral peaks at the GC peak maximum shall agree within a factor of 2 for all
m/z's stored in the spectral library.
13.2.3 The RRT for the pollutant relative to its labeled analog shall be within -2% to +1% of the
RRT in the system performance standard (Section 15.1).
13.3 The m/z's present in the sample mass spectrum that are not present in the reference mass spectrum
shall be accounted for by contaminant or background ions. If the sample mass spectrum is
contaminated, or if identification is ambiguous, an experienced spectrometrist (Section 1.4) is to
determine the presence or absence of the compound.
14.0 Quantitative Determination
14.1 Isotope dilution-Because the pollutant and its labeled analog exhibit the same effects upon purging
and desorption, or equilibration combined with gas chromatography, correction for recovery of the
pollutant can be made by adding a known amount of a labeled compound to every sample prior to
purging or equilibration. Relative response (RR) values for sample mixtures are used in
conjunction with the calibration curves described in Section 10.4.1 to determine concentrations
directly, so long as labeled compound spiking levels are constant. For the o-xylene example given
in Figure 7 (Section 10.4.1.3), RR would be equal to 1.174. For this RR value, the o-xylene
calibration curve given in Figure 6 indicates a concentration of 31.8 (ig/L.
14.2 Internal standard-For the compounds for which the system was calibrated (Table 1 and Table 2)
according to Section 10.4.2, use the response factor determined during the calibration to calculate
the concentration from the equation below, where the terms are as defined in Section 10.4.2.1.
July 1998 24
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Method 1666, Revision A
(As
Concentration = —
14.3 Samples containing >1% solids—The concentration of the pollutant in the solid phase of the
sample is computed using the concentration of the pollutant detected in the aqueous solution, as
follows:
14.3.1 Samples containing low to moderate levels of pollutants (Section 11.5)
r . ,-i
Concentration in solid
sample weight (kg) x percent solids (g) x DF
where:
"percent solids" is from Section 11.1.3 or Section 12.1
Y = volume of water in liters (L) from Section 12.4.4
DF = dilution factor (as a decimal number), where necessary
14.3.2 Methanol extracts (Section 11.6)
Y (L) x p x aqueous cone (Mg/-L)
Concentration in solid (^gl kg) = : :
sample weight (kg) x percent solids (g) x DF
where:
Y = volume of methanol in liters (L) from Section 11.6.3
Fm is the fraction of supernatant removed from the centrifuge tube (e.g., 0.001, Section
11.6.4), and the other terms are as defined in Section 14.3.1
14.3.3 Where the aqueous concentration is in mg/L, the result will be in mg/kg.
14.4 Sample dilution-If the EICP area at the quantitation m/z exceeds the calibration range of the
system, the sample is diluted by successive factors of 10 until the area is within the calibration
range. If dilution of high-solids samples by greater than a factor of 100 is required for
purge-and-trap analysis, then extract the sample with methanol, as described in Section 11.6.
14.5 Dilution of samples containing high concentrations of compounds not in Table 1 or Table 2-When
any peak in the mass spectrum is saturated, dilute the sample per Section 14.4.
14.6 Report results for all pollutants and labeled compounds found in all standards, blanks, and samples
to three significant figures. For samples containing less than 1% solids, the units are micrograms
or milligrams per liter (ng/L or mg/L); and for undiluted samples containing 1% solids or greater,
units are micrograms or milligrams per kilogram ((ig/kg or mg/kg).
25 July 1998
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Method 1666, Revision A
14.6.1 Results for samples that have been diluted are reported at the least dilute level at which the
area at the quantitation m/z is within the calibration range (Section 14.4), or at which no
m/z in the spectrum is saturated (Section 14.5). For compounds having a labeled analog,
results are reported at the least dilute level at which the area at the quantitation m/z is
within the calibration range (Section 14.4) and the labeled compound recovery is within the
normal range for the method (Section 16.2).
15.0 System Performance
15.1 At the beginning of each 12-hour shift during which analyses are performed, system calibration
and performance shall be verified for the pollutants and labeled compounds (Table 1 or Table 2).
For these tests, analysis of the aqueous performance standard (Section 7.8.2) shall be used to
verify all performance criteria. Adjustment and/or recalibration (per Section 10) shall be
performed until all performance criteria are met. Only after all performance criteria are met may
blanks and samples be analyzed.
15.2 BFB spectrum validity-The criteria in Table 5 shall be met.
15.3 Retention times.
15.3.1 Purge-and-trap analysis-The absolute retention times of the internal standards shall fall
within ±30 seconds of the following-bromochloromethane, 954 seconds;
1,4-difluorobenzene, 1052 seconds; chlorobenzene-d5, 1359 seconds. The relative
retention times of all pollutants and labeled compounds shall fall within 5% of the value
given in Table 3.
15.3.2 Direct aqueous injection analysis-The absolute retention time of tetrahydrofuran-ds shall be
263±30 seconds. The relative retention times of all pollutants and labeled compounds
shall fall within 10% of the value given in Table 4.
15.4 GC resolution-The valley height between o-xylene and o-xylene-d10 (at m/z 106 and 116 plotted on
the same graph) shall be less than 10% of the taller of the two peaks.
15.5 Calibration verification and ongoing precision and accuracy-Compute the concentration of each
pollutant (Table 1 or Table 2) by isotope dilution (Section 10.4.1) for those compounds that have
labeled analogs. Compute the concentration of each pollutant that has no labeled analog by the
internal standard method (Section 10.4.2). Compute the concentrations of the labeled compounds
by the internal standard method. These concentrations are computed based on the calibration data
determined in Section 10.
15.5.1 For each pollutant and labeled compound, compare the concentration with the
corresponding limit for ongoing accuracy in Table 6. If all compounds meet the
acceptance criteria, system performance is acceptable and analysis of blanks and samples
may continue. If any individual value falls outside the range given, system performance is
unacceptable for that compound.
July 1998 26
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Method 1666, Revision A
Note: The large number of compounds in Table 6 present a substantial probability that one or more
will fail the acceptance criteria when all compounds are analyzed. To determine if the analytical
system is out of control, or if the failure may be attributed to probability, proceed as follows.
15.5.1.1 Analyze a second aliquot of the aqueous performance standard (Section
7.8.2).
15.5.1.2 Compute the concentration for only those compounds that failed the first
test (Section 15.5.1). If these compounds now pass, system performance
is acceptable for all compounds, and analyses of blanks and samples may
proceed. If, however, any of the compounds fail again, the measurement
system is not performing properly for these compounds. In this event,
locate and correct the problem or recalibrate the system (Section 10), and
repeat the entire test (Section 15.1) for all compounds.
15.5.2 It is suggested but not required that results that pass the specification in Section 15.5.1.2
be added to initial (Section 9.2) and previous ongoing data, that QC charts be updated to
form a graphic representation of laboratory performance (Figure 8), and that a statement
of accuracy be developed for each pollutant and labeled compound by calculating the
average percent recovery (R) and the standard deviation of percent recovery (sr). Express
the accuracy as a recovery interval from R - 2sr to R + 2sr. For example, if R = 95% and
sr = 5%, the accuracy is 85-105%.
16.0 Analysis of Complex Samples
16.1 Some samples may contain high levels (>1000 (ig/kg) of the compounds of interest and of
interfering compounds. Some samples will foam excessively when purged. Others will overload
the trap or the GC column.
16.2 When the recovery of any labeled compound is outside the range given in Table 6, dilute samples
by a factor of 10 with reagent water and analyze this diluted sample. If the recovery remains
outside of the range for this diluted sample, the aqueous performance standard shall be analyzed
(Section 15.1) and calibration verified (Section 15.5). If the recovery for the labeled compound in
the aqueous performance standard is outside the range given in Table 6, the analytical system is
out of control. In this case, the instrument shall be repaired, the performance specifications in
Section 15 shall be met, and the analysis of the undiluted sample shall be repeated. If the recovery
for the aqueous performance standard is within the range given in Table 6, then the method does
not apply to the sample being analyzed, and the result may not be reported for regulatory
compliance purposes.
16.3 When a high level of the pollutant is present, reverse-search computer programs may misinterpret
the spectrum of chromatographically unresolved pollutant and labeled compound pairs with
overlapping spectra. Examine each chromatogram for peaks greater than the height of the internal
standard peaks. These peaks can obscure the compounds of interest.
27 July 1998
-------
Method 1666, Revision A
17.0 Method Performance
17.1 This method was developed and validated in a single laboratory.
17.2 Chromatograms of the aqueous performance standards (Sections 7.8.2 and 15.1) are shown in
Figures 8 and 9.
18.0 Pollution Prevention
18.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity or toxicity
of waste at the point of generation. Many opportunities for pollution prevention exist in laboratory
operation. EPA has established a preferred hierarchy of environmental management techniques
that places pollution prevention as the management option of first choice. Whenever feasible,
laboratory personnel should use pollution prevention techniques to address their waste generation.
When wastes cannot be reduced feasibly at the source, the Agency recommends recycling as the
next best option. The acids used in this Method should be reused as practicable by purifying by
electrochemical techniques. The only other chemicals used in this Method are the neat materials
used in preparing standards. These standards are used in extremely small amounts and pose little
threat to the environment when managed properly. Standards should be prepared in volumes
consistent with laboratory use to minimize the disposal of excess volumes of expired standards.
18.2 For information about pollution prevention that may be applied to laboratories and research
institutions, consult Less is Better: Laboratory Chemical Management for Waste Reduction,
available from the American Chemical Society's Department of Governmental Relations and
Science Policy, 1155 16th Street NW, Washington DC 20036, 202/872-4477.
19.0 Waste Management
19.1 It is the laboratory's responsibility to comply with all Federal, State, and local regulations
governing waste management, particularly the hazardous waste identification rules and
land-disposal restrictions. In addition it is the laboratory's responsibility to protect air, water, and
land resources by minimizing and controlling all releases from fume hoods and bench operations.
Also, compliance is required with any sewage discharge permits and regulations.
19.2 Samples containing acids at a pH of less than 2 are hazardous and must be neutralized before
being poured down a drain or must be handled as hazardous waste.
19.3 For further information on waste management, consult The Waste Management Manual for
Laboratory Personnel and Less is Better: Laboratory Chemical Management for Waste
Reduction, both available from the American Chemical Society's Department of Government
Relations and Science Policy, 1155 16th Street NW, Washington, DC 20036.
July 1998 28
-------
Method 1666, Revision A
20.0 References
1. "Performance Tests for the Evaluation of Computerized Gas Chromatography/Mass Spectrometry
Equipment and Laboratories," U.S. EPA, NERL Cincinnati, OH 45268, EPA-600/4-80-025 (April
1980).
2. Bellar, T. A. and Lichtenberg, J. J., "Journal American Water Works Association," 66, 739
(1974).
3. Bellar, T. A. and Lichtenberg, J. J., "Semi-Automated Headspace Analysis of Drinking Waters and
Industrial Waters for Purgeable Volatile Organic Compounds," in Measurement of Organic
Pollutants in Water and Wastewater, C.E. VanHall, ed., American Society for Testing Materials,
Philadelphia, PA, Special Technical Publication 686 (1978).
4. National Standard Reference Data System, "Mass Spectral Tape Format," U.S. National Bureau
of Standards (1979 and later attachments).
5. "Working with Carcinogens," DREW, PHS, NIOSH, Publication 77-206 (1977).
6. "OSHA Safety and Health Standards, General Industry," 29 CFR 1910, OSHA 2206 (1976).
7. "Safety in Academic Chemistry Laboratories," American Chemical Society Publication, Committee
on Chemical Safety (1979).
8. "Methods 330.4 and 330.5 for Total Residual Chlorine," USEPA, EMSL Cincinnati, OH 45268,
EPA-4-79-020 (March 1979)"Handbook of Analytical Quality Control in Water and Wastewater
Laboratories," U.S. EPA, EMSL Cincinnati, OH 45268, EPA-4-79-019 (March 1979).
9. "Handbook for Analytical Quality Control in Water and Wastewater Laboratories," U.S. EPA,
EMSL Cincinnati, OH 45268, EPA-600/4-79-019 (March 1979).
29 July 1998
-------
Method 1666, Revision A
21.0 Tables
Table 1. Volatile PMI Analytes Amenable to Purge and Trap and Determined by GC/MS Using
Isotope Dilution and Internal Standard Techniques
Pollutant Labeled Compound
PMI Analyte
n-Amyl acetate
n-Amyl alcohol
n-Butyl acetate
n-Butyl alcohol
Tert-butyl alcohol
Cyclohexane
Ethyl acetate
Furfural2
n-Heptane
n-Hexane
Isobutyraldehyde2
Isopropanol
Isopropyl acetate
Isopropylether
Methyl formate
Methylisobutyl ketone
N-pentane3
Tetrahydrofuran
Trichlorofluoromethane
m+p-Xylene
CASRN1
628-63-7
71-41-0
123-86-4
71-36-3
75-65-0
110-82-7
141-78-6
98-01-1
142-82-5
110-54-3
78-84-2
67-63-0
108-21-4
108-20-3
107-31-3
108-10-1
109-66-0
109-99-9
75-69-4
136777-
61-2
EPA- Analog CAS
EGD
977
978
979
1036
.RN1 EPA-
EGD
1343 d10 53001-22-2 1243
1333 d12 1735-17-7 1233
1736 13C 84508-45-2 1636
981
1334 d16 33838-52-7 1234
1335 d14 21666-38-6 1235
982
1044
983
960
991
1341
984
1345 ds 1693-74-9 1245
552
1332 d10 41051-88-1 1232
o-Xylene
95-47-6
1331
•Mo
56004-61-6 1231
July 1998
30
-------
Method 1666, Revision A
1 Chemical Abstracts Service Registry Number.
2 These aldehydes may also be analyzed by Method 1667.
3 n-Pentane is not explicitly a PMI analyte. However, the sum of the concentrations of n-pentane,
n-hexane, and n-heptane are to be used to estimate the concentration of petroleum naphtha in
PMI wastewaters.
31 July 1998
-------
Method 1666, Revision A
Table 2. Volatile PMI Analytes Determined by Direct Aqueous Injection GCMS Using Isotope
Dilution and Internal Standard Techniques
Pollutant Labeled Compound
CASRN1
PMI Analyte
Acetonitrile 75-05-8
Diethylamine 109-89-7
Dimethylamine 124-40-3
Dimethyl sulfoxide 67-68-5
Ethanol 64-17-5
Ethylene glycol 107-21-1
Formamide 75-12-7
Methanol 67-56-1
Methylamine 74-89-5
Methyl Cellosolve® 109-86-4
n-Propanol 71-23-8
Triethylamine 121-44-8
EPA-EGD Analog CAS
RN1 EPA-
EGD
972 d3 2206-26-0 1272
986
987
1037 d6 2206-27-1 1237
1734 d6 1516-08-1 1634
1038
988
1735 d3 1849-29-2 1635
989
1040
755 1-dj not avail. 1255
990
1 Chemical Abstracts Service Registry Number.
July 1998
32
-------
Method 1666, Revision A
Table 3. Gas Chromatographic Retention Times and Minimum Levels for Volatile PMI
Analytes Determined by Purge and Trap GC/MS
EGD
No.1
991
552
984
1344
1243
1343
1235
982
1335
960
1636
1736
181
1245
1345
1233
1333
1234
1336
1334
985
983
1341
978
979
981
207
1232
1332
1231
977
1331
Retention Time
ML2
PMI Analyte
Methyl formate
Trichlorofluoromethane
n-Pentane
Isopropanol
Tert-butyl alcohol-d10
Tert-butyl alcohol
n-Hexane-d14
Isobutyraldehyde
n-Hexane
Isopropylether
Ethyl acetate-13C
Ethyl acetate
Bromochloromethane(I.S.)
Tetrahydrofuran-d8
Tetrahydrofuran
Cyclohexane-d12
Cyclohexane
n-Heptane-d16
n-Butanol
n-Heptane
1,4-Difluorobenzene(I. S.)
Isopropyl acetate
Methylisobutyl ketone
n-Amyl alcohol
n-Butyl acetate
Furfural
Chlorobenzene-d5 (I.S.)
p-Xylene-d10
m,p-Xylene
o-Xylene-d10
n-Amyl acetate
o-Xylene
Mean (sec)
526
613
622
687
730
741
820
823
839
865
925
925
954
956
964
981
996
1013
1015
1033
1052
1128
1157
1202
1268
1354
1359
1368
1379
1413
1417
1424
EGD
181
181
181
181
181
1243
181
181
1235
181
181
1636
181
181
181
181
1233
985
985
1234
985
985
985
985
207
207
207
207
1232
207
207
1231
Relative
0.551
0.642
0.652
0.720
0.765
1.016
0.860
0.863
1.023
0.907
0.970
.000
.000
.002
.012
.028
.015
0.963
0.964
.020
.000
.072
.100
.143
0.933
0.996
.000
.007
.008
.040
.043
.008
(^g/L
100
10
10
200
100
10
10
5
10
20
5
500
10
10
10
500
5
500
10
5
5
1 Three digit EGD numbers beginning with 0, 1, 5, or 9 indicate a pollutant quantified by the internal
standard method; EGD numbers beginning with 2 or 6 indicated a labeled compound quantified by the
internal standard method; EGD numbers beginning with 3 or 7 indicate a pollutant quantified by isotope
dilution. The initial "1" in four digit EGD numbers is to be ignored in applying these rules.
33
July 1998
-------
Method 1666, Revision A
2 This is a minimum level at which the entire analytical system shall give recognizable mass spectra
(background corrected) and acceptable calibration points, taking into account method-specific sample and
injection volumes. The concentration in the aqueous or solid phase is determined using the equations in
Section 14.
July 1998 34
-------
Method 1666, Revision A
Table 4. Gas Chromatographic Retention Times and Minimum Levels for Volatile PMI Analytes by
Direct Aqueous Injection GC/MS
EGD
No-1 PMI Analyte
989 Methylamine
1635 Methyl alcohol-d3
1735 Methyl alcohol
987 Diethylamine
1634 Ethyl alcohol-d5
1734 Ethyl alcohol
1272 Acetonitrile-d3
972 Acetonitrile
1255 n-Propanol-l-d]
755 n-Propanol
986 Diethylamine
1245 Tetrahydrofuran-d8 (I.S.)
1040 Methyl Cellosolve®
(2-Methoxyethanol)
990 Triethylamine
1038 Ethylene glycol
988 Formamide
1237 Dimethyl sulfoxide-d6
1037 Dimethyl sulfoxide
Mean (sec)
81
85
85.5
93
103
104
119
121
170
170.5
188
263
290
EGD Ref
1245
1245
1635
1245
1245
1634
1245
1272
1245
1255
1245
1245
1245
Relative
0.308
0.323
1.006
0.354
0.394
1.010
0.452
1.017
0.464
1.003
0.717
1.000
1.103
1VXAJ
(mg/L)
200
50
200
20
5
20
200
50
372
398
400
639
643
1245
1245
1245
1245
1237
1.414
1.513
1.521
2.431
1.006
200
200
1000
100
1 Three digit EGD numbers beginning with 0, 1, 5, or 9 indicate a pollutant quantified by the internal
standard method; EGD numbers beginning with 2 or 6 indicated a labeled compound quantified by the
internal standard method; EGD numbers beginning with 3 or 7 indicate a pollutant quantified by isotope
dilution. The initial "1" in four digit EGD numbers is to be ignored in applying these rules.
2 This is a minimum level at which the entire analytical system shall give recognizable mass spectra
(background corrected) and acceptable calibration points, taking into account method-specific sample
and injection volumes. The concentration in the aqueous or solid phase is determined using the
equations in Section 14.
35
July 1998
-------
Method 1666, Revision A
Table 5. BFB Mass-Intensity Specifications
m/z Intensity Required
50 15-40% of m/z 95
75 30-60% of m/z 95
95 base peak, 100%
96 5-9% of m/z 95
173 less than 2% of m/z 174
174 greater than 50% of m/z 95
175 5-9% of m/z 174
176 95-101% of m/z 174
177 5-9% of m/z 176
July 1998 36
-------
Method 1666, Revision A
Table 6. Quality Control
EGD
No. PMI Analyte
972 Acetonitrile
977 Amyl acetate
978 Amyl alcohol
979 n-Butyl acetate
1036 n-Butyl alcohol
1343 tert-Butyl alcohol
1333 Cyclohexane
986 Diethylamine
987 Diethylamine
1037 Dimethyl sulfoxide
1734 Ethanol
1736 Ethyl acetate
1038 Ethylene glycol
988 Formamide
981 Furfural
1334 n-Heptane
1335 n-Hexane
982 Isobutyraldehyde
1044 Isopropanol
Acceptance Criteria for PMI Analytes
Acceptance Criteria for Performance Tests (% of Spike
Level)
Spike
Level
50mg/L
10 Mg/L
200 (ig/L
10 Mg/L
200 (ig/L
50 ng/L
10 ng/L
250 mg/L
250 mg/L
250 mg/L
50 mg/L
10 Mg/L
250 mg/L
500 mg/L
100 (ig/L
10 Mg/L
10 ng/L
10 Mg/L
100 (ig/L
Labeled and Native
PMI Analyte
Initial Precision
and Recovery
s
20
20
75
20
108
121
26
31
38
20
15
48
195
113
186
37
34
54
284
Labele
Ana
X Recov
Labeled and
dPMI Native PMI Analyte
ilyte On-going Recovery
ery (P) (R)
70-130 70-130 70-130
70-130
16-166
70-130
d-190
d-202
70-130
10-172
70-130
d-199
d-212
70-134 8-156 70-136
70-132
61-136
68-134
58-139
70-130 59-122 70-130
66-130 70-130 65-130
60-157 58-159 57-160
d-310
60-286
d-282
d-326
51-296
d-297
70-161 14-128 70-164
70-154 5-157 70-157
67-176
d-418
63-180
d-441
37
July 1998
-------
Method 1666, Revision A
983 Isopropyl acetate 10 (ig/L 32 70-147 70-150
960 Isopropyl ether 10 jig/L 21 70-127 70-129
1735 Methanol 50 mg/L 26 57-109 70-130 55-111
989 Methylamine 250 mg/L 36 61-133 59-136
1040 Methyl cello solve 250 mg/L 20 70-130 70-130
991 Methyl formate 50 ng/L 73 20-165 14-171
1341 Methylisobutyl ketone 10 jig/L 42 70-162 70-165
984 n-Pentane 10 (ig/L 52 51-155 47-159
755 n-Propanol 50 mg/L 25 42-130 54-149 40-130
975 Tetrahydrofuran 10 jig/L 89 35-214 42-178 28-221
552 Trichlorofluoromethane 20 (ig/L 20 70-130 70-130
990 Triethylamine 250 mg/L 31 70-133 69-135
1332 m,p-Xylene 20 ng/L 20 70-130 70-130 70-130
1331 o-Xylene 10 jig/L 20 70-130 70-130 70-130
Ju/y 7998 38
-------
Method 1666, Revision A
Table 7. Characteristic
PMI Analyte
Acetonitrile
n-Amyl acetate
n-Amyl alcohol
n-Butyl acetate
n-Butyl alcohol
Tert-butyl alcohol
Cyclohexane
Diethylamine
Diethylamine
Dimethyl sulfoxide
Ethanol
Ethyl acetate
Ethylene glycol
Formamide
Furfural
n-Heptane
n-Hexane
Isobutyraldehyde
Isopropanol
Isopropyl acetate
Isopropyl ether
Methanol
Methylamine
Methyl Cellosolve® (2-
m/z's for Volatile PMI Analytes
Spike Level
I Labeled
(jig/L) (mg/L) Analog
50 d3
10
100
10
200
50 d10
10 d12
250
250
250 d6
50 d5
10 13C
250
500
100
10 d16
10 d14
10
100
10
10
50 d3
250
250
Primary m/z
(Native/Labeled)
41/44
43
70
43
56
59/66
56/96
58
44
47/50
31/33
43/44
31
45
96
71/82
57/66
72
45
43
45
31/33
30
45
Reference
Compound
1272
207
985
207
985
1234
1233
1245
1245
1237
1634
1636
1245
1245
207
1234
1235
181
181
985
181
1635
1245
1245
Methoxyethanol)
39
July 1998
-------
Method 1666, Revision A
Methyl formate
Methylisobutyl ketone
n-Pentane
n-Propanol
Tetrahydrofuran
Trichlorofluoromethane
Triethylamine
m,p-Xylene
o-Xylene
50
10
10
50 1-dj
10 ds
20
250
20 d10
10 d10
60
43
43
31/32
72/80
101
86
106/116
106/16
181
181
1255
1245
181
1245
1232
1231
July 1998
40
-------
Method 1666, Revision A
Table 8. Maximum Recoveries for PMI Analytes by Purge-and-Trap GC/MS
PMI Analyte Maximum Recovery
(%)
n-Amyl acetate 130
n-Amyl alcohol 300
n-Butyl acetate 130
n-Butyl alcohol 440
tert-Butyl alcohol 130
Cyclohexane 130
Ethyl acetate 130
Furfural 170
n-Heptane 140
n-Hexane 140
Isobutyraldehyde 150
Isopropanol 250
Isopropyl acetate 130
Isopropyl ether 130
Methyl formate 130
Methylisobutyl ketone 130
n-Pentane 130
Tetrahydrofuran 150
Trichlorofluoromethane 130
m,p-Xylene 130
o-Xylene 130
41 July 1998
-------
Method 1666, Revision A
Optional
Foam Trap
ExU1/4ln.O.D.
10 mmOass Fril,
Medum Borosi ^
EN 11/4 in. O.D.
UrrmO.D.
Sanrple He I
2-WavS)*inge Valve
17
-------
Method 1666, Revision A
Puge hlelRKng
SampteGulelfiUing
3 in. Long x 6 rrm O.D. Gass Tubfrig
«-rr!Mal
nrnn
Cap
Figuie2. Purging Devbe for Soils or Waters
43
Ju/y 7998
-------
Method 1666, Revision A
Paefctig Detail
Consluciori De&i
iz~ 5 mmQassWool
7.7emSiIea<3el
15 cm Tenax <3C
1— 5 mm Glass Wool
Trsp Inlel
GompfessJon
Rling Nul
and Parties
Besislance Wire
Sdid
Gon Idler
Sensor
Hecfonic
Terrpera yre
Gon*ol and
Pjforreler
TLbing ,25 cm
0.105 in. I.D.
0.125 in. O.D.
Stainless Sled
Fig ure 3. Tra p Co n&truc tion a nd Pac ki ngs.
July 1998
44
-------
Method 1666, Revision A
Carrier-Gas
Row Con rd
Pressure
Regulator
Purge-Gas
RowCotilol
ISxMdeoJar
Sieve Rlter
Liquid Irjecion Porls
Cdurm Oven
Opional iPorlColurm
Seleclbn Valve
Trap hlel
Conlirna Dry Cdurrn
To Detector
Note:
flll lines be Iween lap
and GG should be heated
to 80^0.
Fig ure 4. Sc he mate of Pu ige-a nd -Trap Devbe—Pu rge Mode
45
July 1998
-------
Method 1666, Revision A
Carrier-Gas
RowGorrtol
Pressure
Regulator
Purge-Gas
Row Control
13xMdeeiJar
Sieve Filter
¥
Liquid Injecion Porte
ColurmOven
OpSonal 4-PorlColum
Sdecta Valwe
6-Port
telwe
Van
f
/"Trap hlel
Trap,
200^
Con Irrra lory Cdumn
To DeteeU*
• Analyical Cdurrn
Cfevice
Note:
Al fnesbehfeen irap
and <3C stwdd be heated
Figures. SchernafcofP urge-and -Tra p Devbe—Deso rb iDbde
July 1998
46
-------
Method 1666, Revision A
I I I I
10 —
1JQ —
0.1 —
I I
10 20
ConcenUaion
90
I
100
200
The Dolfed Lines Endosea ±10% Errw Window.
Figu IB 6. Rela trye Response Cal ibratbn C uive b r o-Xybne
47
July 1998
-------
Method 1666, Revision A
Area=1689£Q
MS116
MS 106
M2116
MS 106
M2106
M2116 82508
WZ116
M/Z106
Figure 7. Extrac bd Ion C urren t P rofiles fo r tft) o-Xylene, (B) o -Xytene-d 10and
(P) a flllkture of o-Xybne and o-Xybne-d10
July 1998
48
-------
Method 1666, Revision A
170000 -
160000 -
150000 -
140000 -
130000 -
120000 -
110000 -
100000 -
90000 -
80000 -
70000 -
60000 -
50000 -
40000 -
30000 -
20000 -
10000 -
0
Fib
Operator
Acquired
Instrument
Sample Name
Mix Info
Vial Number
DiXDATAXHOT PURG EiAUG 1994'iJ082694'W PSTD1 ,D
rh
26Aug94 227pm us rg AcqMethod HOT PURGE
HP-1
Holpurge Method SttJ. 080^94
hopurgeanalyiBS (SOppblSBS)
2
UJ
L
UJI
10
15
20
Figu re 8, C hro matogra m of Aq ueous Per fa rma nee
Standard of Arah/bs from Tabte 1
49
July 1998
-------
Method 1666, Revision A
100 H
FS -
Sample
LabRlelD
Analyzed
hsliumenllD
: STD. COlfiOOO
: 4K13UE
: 11/1564
:VQ04
Mn
2.0
4.0
6.0
8.0
10.0
12JO
Figure 9. C hrarnatogram of Aqueous Performance
Standard of Analytes from Table 2
July 1998
50
-------
Method 1666, Revision A
1
•zi m 4 nri nfifi
l~
•3*
A*
Qft f)f"f)
1
J Jfi _
O
s *
S * 1 rcH
f!
3 6
A on —
UilU
1
1 1 1 1 1 1 1 1 1
o-M^sns-d^
*
' * * * * *
1 1 I 1 1 1 1 I 1
£345 6789 10
Anaftjsts
Number
i i i i i i i i i
___, - — --.—
•__._ _•__!___
i i i i i i i i i
5f\ 6/1 6/1 6/1 6£ 6£ 6/3 6G 6/4 6j6
Dale
Analyzed
-^
- -3s
Figu re 10. Quality Con tnol C har te. S how ingArea(bpgraph)andRelali« Response of
o-Xylene to o-Xyb ne -d 10 (bwe r grap h) P lotted as Fu re tb n of 11 me o r A nalysis
Number
51
July 1998
-------
-------
Method 1667
Formaldehyde, Isobutyraldehyde, and Furfural by
Derivatization Followed by High Performance Liquid
Chromatography
Revision A, July 1998
-------
-------
Method 1667, Revision A
Formaldehyde, Isobutyraldehyde, and Furfural by Derivatization Followed by
High Performance Liquid Chromatography
1.0 Scope and Application
1.1 This method is for surveying and monitoring under the Clean Water Act. It is used to determine
certain organic pollutants specific to the pharmaceutical manufacturing industry (PMI) that can be
derivatized and analyzed by high-performance liquid chromatography (HPLC).
1.2 The chemical compounds listed in Table 1 may be determined in waters, soils, and municipal sludges
by this method.
1.3 The detection limits of the method are usually dependent on the level of interferences rather than
instrumental limitations. The limits in Table 2 are the minimum levels that can be reliably quantified
by this method with no interferences present.
Furfural (2-furaldehyde) forms two relatively stable geometric isomers upon derivatization with
2,4-dinitrophenylhydrazine (DNPH). The first isomer (probably anti-) elutes after the formaldehyde
derivative and before the isobutyraldehyde derivative. The second isomer (probably syn-) elutes after
the isobutyraldehyde derivative. Experience with this system has shown that the best quantitative
results (lowest detection limits) are obtained using the area from the first eluted peak rather than that
from the second peak or the sum of the two areas. This method is for use only by analysts experienced
with HPLC or under the close supervision of such qualified persons.
1.4 This method is performance-based. The analyst is permitted to modify the method to overcome
interferences or to lower the cost of measurements, provided that all performance criteria in this
method are met. The requirements for establishing method equivalency are given in Section 9.1.2.
2.0 Summary of the Method
2.1 For solid wastes or for aqueous wastes containing significant amounts of solid material, the aqueous
phase, if any, is separated from the solid phase and stored for later analysis. If necessary, the particle
size of the solids in the waste is reduced. The solid phase is extracted with an amount of extraction
fluid equal to 20 times the weight of the solid phase. The extraction fluid employed is a function of
the alkalinity of the solid phase of the waste. Following extraction, the aqueous extract is separated
from the solid phase by filtration employing 0.6 to 0.9-(im glass-fiber filter.
2.2 If compatible (i.e., multiple phases will not form on combination), the initial aqueous phase of the
waste is added to the aqueous extract, and these liquids are analyzed together. If incompatible, the
liquids are analyzed separately and the results are mathematically combined to yield a
volume-weighted average concentration.
2.3 A measured volume of aqueous sample or an appropriate amount of solids leachate is buffered to
pH=5 and derivatized with 2,4-dinitrophenylhydrazine (DNPH), using either the solid-sorbent or
55 July 1998
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Method 1667, Revision A
methylene chloride derivatization/extraction option. If the solid-sorbent option is used, the derivative
is extracted using solid-sorbent cartridges, followed by elution with ethanol. If the methylene chloride
option is used, the derivative is extracted with methylene chloride. The methylene chloride extracts
are concentrated using the Kuderna-Danish (K-D) procedure and solvent exchanged into methanol
prior to HPLC analysis. Liquid chromatographic conditions are described that permit the separation
and measurement of formaldehyde, isobutyraldehyde, and furfural derivatives in the extract by
absorbance detection at 365 nm.
2.4 The quality of the analysis is assured through reproducible calibration and testing of the
derivatization/extraction procedure and the HPLC system.
3.0 Definitions
There are no specific definitions unique to this method.
4.0 Interferences
4.1 Method interferences may be caused by contaminants in solvents, reagents, glassware, and other
sample processing hardware that lead to discrete artifacts and/or elevated baselines in chromatograms.
All of these materials must be routinely demonstrated to be free from interferences under the conditions
of the analysis by analyzing laboratory reagent blanks as described in Section 9.3.
4.1.1 Glassware must be scrupulously cleaned. Clean all glassware as soon as possible after use
by rinsing with the last solvent used. This should be followed by detergent washing with hot
water, and rinses with tap water and reagent water. It should then be drained, dried, and
heated in a laboratory oven at 130°C for several hours before use. Solvent rinses with
methanol may be substituted for the oven heating. After drying and cooling, glassware should
be stored in a clean environment to prevent any accumulation of dust or other contaminants.
4.1.2 The use of high-purity reagents and solvents helps to minimize interference problems.
Purification of solvents by distillation in all-glass systems may be required.
4.2 Analysis of formaldehyde is complicated by its ubiquitous occurrence in the environment. Acetic acid,
even high-purity acetic acid, is often contaminated with formaldehyde. For this reason, a phthalate
buffer is used in this method instead of an acetate buffer. Wherever acetic acid is used, it must be
demonstrated to be formaldehyde free.
4.3 Matrix interferences may be caused by contaminants that are coextracted from the sample. The extent
of matrix interferences will vary considerably from source to source, depending upon the nature and
diversity of the matrix being sampled. If matrix interferences occur, some additional cleanup may be
necessary.
4.4 The extent of interferences that may be encountered using liquid chromatographic techniques has not
been fully assessed. Although the HPLC conditions described allow for resolution of the specific
compounds covered by this method, other matrix components may interfere.
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Method 1667, Revision A
5.0 Safety
5.1 The toxicity or carcinogenicity 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.
Exposure to these compounds should be reduced to the lowest possible level. The laboratory is
responsible for maintaining a current awareness file of OSHA regulations regarding the safe handling
of the chemicals specified in this method. A reference file of material safety data sheets should also
be made available to all personnel involved in these analyses. Additional information on laboratory
safety can be found in References 1 through 3.
5.2 Formaldehyde has been classified as a potential carcinogen. Primary standards of formaldehyde
should be prepared in a hood, and a NIOSH/MESA approved toxic gas respirator should be worn
when high concentrations are handled.
6.0 Apparatus and Materials
Disclaimer: The mention of trade names or commercial products in this Method is for illustrative
purposes only and does not constitute endorsement or recommendation for use by the Environmental
Protection Agency. Equivalent performance may be achievable using apparatus, materials, or
cleaning procedures other than those suggested here. The laboratory is responsible for
demonstrating equivalent performance.
6.1 Reaction vessel—250-mL Florence flask.
6.2 Separatory funnel—250-mL, with polytetrafluoroethylene (PTFE) stopcock.
6.3 Kuderna-Danish (K-D) apparatus.
6.3.1 Concentrator tube—10-mL, graduated (Kontes K-570050 or equivalent). A ground-glass
stopper is used to prevent evaporation of extracts.
6.3.2 Evaporation flask—500-mL (Kontes K-570001-500 or equivalent). Attach to concentrator
tube with springs, clamps, or equivalent.
6.3.3 Snyder column—Three-ball macro (Kontes K-503000-0121 or equivalent).
6.3.4 Snyder column—Two-ball micro (Kontes K569001-0219 or equivalent).
6.3.5 Springs—!/2", (Kontes K-662750 or equivalent).
6.4 Vials—10- and 25-mL glass, with PTFE-lined screw-caps or crimp-tops.
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Method 1667, Revision A
6.5 Boiling chips—Solvent-extracted with methylene chloride, approximately 10/40 mesh (silicon carbide
or equivalent).
6.6 Balance—Analytical, capable of accurately weighing to the nearest 0.0001 g.
6.7 pH meter—Capable of measuring to the nearest 0.01.
6.8 High-performance liquid chromatograph (modular).
6.8.1 Pumping system—Isocratic, with constant flow control capable of 1.00 mL/min.
6.8.2 High-pressure injection valve with 20-(iL loop.
6.8.3 Column—250 mm long x 4.6 mm inside diameter (i.d.), 5-(im particle size, C18 (or
equivalent).
6.8.4 Absorbance detector—365 nm.
6.8.5 Strip-chart recorder compatible with the detector. Use of a data system is recommended.
6.9 Glass-fiber filter paper, 0.6 to 0.9-(im.
6.10 Solid-sorbent cartridges—Packed with 500 mg C18 (Baker or equivalent).
6.11 Vacuum manifold—Capable of simultaneous extraction of up to 12 samples (Supelco or equivalent).
6.12 Sample reservoirs—50-mL capacity (Supelco or equivalent).
6.13 Pipet—Capable of accurately delivering 0.10 mL of solution (Pipetman or equivalent).
6.14 Water bath—Heated, with concentric ring cover, capable of temperature control of ± 2 °C at 80-90 °C.
The bath should be used in a hood.
7.0 Reagents and Standards
7.1 Reagent grade chemicals shall be used in all tests. Unless otherwise indicated, it is intended that all
reagents shall conform to the specifications of the Committee on Analytical Reagents of the American
Chemical Society, where such specifications are available. Other grades may be used, provided it is
first ascertained that the reagent is of sufficiently high purity to permit its use without lessening the
accuracy of the determinations.
7.2 Reagent water—Water in which the compounds of interest and interfering compounds are not detected
by this method. It may be generated by any of the methods in this subsection.
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Method 1667, Revision A
7.2.1 Activated carbon—Pass tap water through a carbon bed (Calgon Filtrasorb-300, or
equivalent).
7.2.2 Water purifier—Pass tap water through a purifier (Millipore Super Q, or equivalent).
7.2.3 Boil and purge—Heat tap water to 90-100°C and bubble contaminant-free inert gas through
it for approximately 1 hour. While still hot, transfer the water to screw-cap bottles and seal
with a PTFE-lined cap.
7.3 Methylene chloride—HPLC grade or equivalent.
7.4 Methanol—HPLC grade or equivalent.
7.5 Ethanol (absolute)—HPLC grade or equivalent.
7.6 2,4-Dinitrophenylhydrazine (DNPH, 70% w/w) in reagent water.
7.7 Formalin (37.6% w/w) in reagent water.
7.8 Acetic acid (glacial), demonstrated to be formaldehyde-free.
7.9 Potassium acid phthalate.
7.10 Sodium hydroxide solutions, 1 N, and 5 N.
7.11 Sodium chloride.
7.12 Sodium sulfate solution, 0.1 M.
7.13 Hydrochloric acid, 0.1 N.
7.14 Extraction fluid—Dilute 64.3 mL of 1.0 N sodium hydroxide and 5.7 mL of glacial acetic acid to 900
mL with reagent water. Further dilute to 1 L with reagent water. The pH should be 4.93 ± 0.02. If
not, adjust with acid or base.
7.15 Stock standard solutions.
7.15.1 Stock formaldehyde (approximately 1.00 mg/mL)—Prepare by diluting 265 (iL formalin to
100 mL with reagent water.
Standardization of formaldehyde stock solution—Transfer a 25 -mL aliquot of a 0.1 M sodium
sulfite solution to a beaker and record the pH. Add a 25-mL aliquot of the formaldehyde stock
solution (Section 7.15.1) and record the pH. Titrate this mixture back to the original pH
using 0.1 N hydrochloric acid. The formaldehyde concentration is calculated using the
following equation:
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Method 1667, Revision A
Concentration (mg/mL) = 30.03 x (N HCl) x (mL HCl) x 25
where'.
N HCl = Normality of the HCl solution
mL HCl = mL of standardised HCl solution', and
30.03 = Molecular weight of formaldehyde.
7.15.2 Stock formaldehyde, isobutyraldehyde, and furfural—Prepare by adding 265 (iL of formalin,
0.100 g of isobutyraldehyde, and 0.100 g of furfural to 90 mL of reagent water and dilute to
100 mL. The concentrations of isobutyraldehyde and furfural in this solution are 1.00
mg/mL. Calculate the concentration of formaldehyde in this solution using the results of the
assay performed in Section 7.15.1.
7.15.3 Stock standard solutions must be replaced after six months, or sooner if comparison with
check standards indicates a problem.
7.15.4 Aqueous performance standard—An aqueous performance standard containing formaldehyde
(nominally 100 (ig/L), isobutyraldehyde at 100 (ig/L, and furfural at 100 (ig/L shall be
prepared daily and analyzed each shift to demonstrate performance (Section 9).
7.15.5 Preparation of calibration standards.
7.15.5.1 Prepare calibration standard solutions of formaldehyde, isobutyraldehyde,
and furfural in reagent water from stock standard solution (Section 7.15.2).
Prepare these solutions at the following concentrations (in (ig/mL) by serial
dilution of the stock standard solution: 50, 20, 10. Prepare additional
calibration standard solutions at the following concentrations, by dilution of
the appropriate 50, 20, or 10 (ig/mL standard: 5, 0.5, 2, 0.2, 1, 0.1. Make
further dilutions if appropriate.
7.16 Reaction solutions.
7.16.1 DNPH (1.00 mg/mL)—Dissolve 142.9mg70%(w/w)reagentin 100 mL of absolute ethanol.
Slight heating or sonication may be necessary to effect dissolution.
7.16.2 Phthalate buffer (0.1 N)—Prepare by dissolving 20.42 g of potassium acid phthalate in 1 L
of reagent water. Adjust pH to 5 by addition of sodium hydroxide or hydrochloric acid, as
necessary.
7.16.3 Sodium chloride solution (saturated)—Prepare by mixing an excess of the reagent-grade solid
with reagent water.
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8.0 Sample Collection, Preservation, and Storage
8.1 Grab samples are collected in glass containers having a total volume greater than 20 mL. For aqueous
samples that pour freely, fill sample bottles so that no air bubbles pass through the sample as the bottle
is filled and seal each bottle so that no air bubbles are entrapped. Maintain the hermetic seal on the
sample bottle until time of analysis.
8.2 Samples are maintained at 0-4°C from the time of collection until analysis. Samples must be
derivatized within five days of collection and analyzed within three days of derivatization.
9.0 Quality Control
9.1 Each laboratory that uses this method is required to operate a formal quality assurance program
(Reference 4). The minimum requirements of this program consist of an initial demonstration of
laboratory capability and analysis of standards and blanks as tests of continued performance.
Laboratory performance is compared to established performance criteria to determine if the results of
analyses meet the performance characteristics of the method.
9.1.1 The analyst shall make an initial demonstration of the ability to generate acceptable accuracy
and precision with this method. This ability is established as described in Section 9.2.
9.1.2 In recognition of advances that are occurring in analytical technology, and to allow the analyst
to overcome sample matrix interferences, the analyst is permitted certain options to improve
separations or lower the costs of measurements. These options include alternative extraction,
concentration, cleanup procedures, and changes in columns and detectors. Alternative
techniques, such as substitution of immunoassay, and changes that degrade method
performance are not allowed. If an analytical technique other than the techniques specified
in this method is used, that technique must have a specificity equal to or better than the
specificity of the techniques in this method for the analytes of interest.
9.1.2.1 Each time a modification is made to this method, the analyst is required to repeat the
procedure in Section 9.2. If the detection limit of the method will be affected by the
change, the laboratory is required to demonstrate that the method detection limit
(MDL; 40 CFR Part 136, Appendix B) is lower than one-third the regulatory
compliance level. If calibration will be affected by the change, the analyst must
recalibrate the instrument per Section 10.
9.1.2.2 The laboratory is required to maintain records of modifications made to this method.
These records include the information below, at a minimum.
9.1.2.2.1 The names, titles, addresses, and telephone numbers of the
analyst(s) who performed the analyses and modification, and of the
quality control officer who witnessed and will verify the analyses
and modification.
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Method 1667, Revision A
9.1.2.2.2 A list of pollutant(s) measured, including name and CAS Registry
Number.
9.1.2.2.3 A narrative stating the reason(s) for the modification.
9.1.2.2.4 Results from all quality control (QC) tests comparing the modified
method to this method, including:
(a) Calibration (Section 10.1.2)
(b) Calibration verification (Section 10.1.2.2)
(c) Initial precision and accuracy (Section 9.2)
(d) Analysis of blanks (Section 9.3)
(e) Accuracy assessment (Section 9.5)
9.1.2.2.5 Data that will allow an independent reviewer to validate each
determination by tracing the instrument output (peak height, area, or
other signal) to the final result. These data are to include:
(a) Sample numbers and other identifiers
(b) Extraction dates
(c) Analysis dates and times
(d) Analysis sequence/run chronology
(e) Sample weight or volume (Section 11)
(f) Extract volume prior to each cleaning step (Section 11.1.2)
(g) Final extract volume prior to injection (Section 11.3.4.5 or
Section 11.3.5.5)
(h) Injection volume (Section 12)
(I) Dilution data, differentiating between dilution of a sample
or an extract
(j) Instrument and operating conditions
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Method 1667, Revision A
(k) Column and operating conditions (nature of column,
dimensions, flow rates, solvents, etc.)
(1) Detector operating conditions (wavelength, etc.)
(m) Chromatograms, printer tapes, and other recording of raw
data
(n) Quantitation reports, data system outputs, and other data
necessary to link raw data to the results reported
9.1.3 Analyses of blanks are required to demonstrate freedom from contamination and that the
compounds of interest and interfering compounds have not been carried over from a previous
analysis (Section 4). The procedures and criteria for analysis of a blank are described in
Section 9.3.
9.1.4 The laboratory shall, on an ongoing basis, demonstrate through the analysis of the aqueous
performance standard (Section 7.15.4) that the analysis system is in control. This procedure
is described in Section 10.
9.1.5 The laboratory shall maintain records to define the quality of data that is generated.
9.2 Initial precision and accuracy—To establish the ability to generate acceptable precision and accuracy,
the analyst shall perform the following operations for compounds to be calibrated.
9.2.1 Analyze four aliquots of the aqueous performance standard (Section 7.15.4) according to the
method beginning in Section 11. Use the solid-sorbent option or the methylene chloride
option, whichever will be used routinely.
9.2.2 Using results from Section 9.2.1, compute the average percent recovery (X) and the standard
deviation of the recovery (s) for each compound.
9.2.3 For each compound, compare s and X with the corresponding limits for initial precision and
recovery found in Table 3. If s and X for all compounds meet the acceptance criteria, system
performance is acceptable and analysis of blanks and samples may begin. If, however, any
individual s exceeds the precision limit or any individual X falls outside the range for
accuracy, system performance is unacceptable for that compound. This is an indication that
the analytical system is not performing properly for the compound(s) in question. In this
event, correct the problem and repeat the entire test (Section 9.2.1).
9.3 Blanks—Reagent water blanks are analyzed to demonstrate freedom from contamination.
With each sample lot (samples analyzed on the same 12-hour shift), a blank shall be analyzed
immediately after analysis of the aqueous performance standard (Section 9.1.4) to demonstrate
freedom from contamination. If any of the compounds of interest or any potentially interfering
63 July 1998
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Method 1667, Revision A
compound is found in a blank at greater than 10 (ig/L, analysis of samples is halted until the source
of contamination is eliminated and a blank shows no evidence of contamination at this level.
9.4 The specifications contained in this method can be met if the apparatus used is calibrated properly,
then maintained in a calibrated state. The standards used for calibration (Section 7.15.5), calibration
verification (Section 10.1.2.2) and for initial (Section 9.2) and ongoing (Section 9.1.4) precision and
accuracy should be identical, so that the most precise results will be obtained.
9.5 Depending on specific program requirements, field replicates may be collected to determine the
precision of the sampling technique, and spiked samples may be required to determine the accuracy
of the analysis.
10.0 Calibration
10.1 Establish liquid chromatographic operating parameters to produce a retention time equivalent to that
indicated in Table 2 for formaldehyde derivative. Suggested chromatographic conditions are provided
in Section 12.1. Prepare derivatized calibration standards according to the procedure in Section
10.1.1. Calibrate the chromatographic system using the external standard technique (Section 10.1.2).
10.1.1 Process each calibration standard solution through the derivatization option used for sample
processing (Section 11.3.4 or 11.3.5).
10.1.2 External standard calibration procedure.
10.1.2.1 Analyze each derivatized calibration standard using the chromatographic
conditions specified in Section 12.1, and tabulate peak area against
concentration injected. The results may be used to prepare calibration curves
for formaldehyde, isobutyraldehyde, and furfural.
10.1.2.2 The working calibration curve must be verified at the beginning of each 12-
hour shift or every 20 samples, whichever is more frequent, by the
measurement of one or more calibration standards. If the response for any
analyte varies from the previously established responses by more than 10%,
the test must be repeated using a fresh calibration standard after it is verified
that the analytical system is in control. Alternatively, a new calibration
curve may be prepared for that compound. If an autosampler is available,
it is convenient to prepare a calibration curve daily by analyzing standards
along with test samples.
11.0 Sample Extraction, Cleanup, and Derivatization
11.1 Extraction of solid samples.
11.1.1 All solids must be homogeneous. When the sample is not dry, determine the dry weight of the
sample using a representative aliquot.
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Method 1667, Revision A
11.1.1.1 Determination of dry weight—In certain cases, sample results are desired
based on a dry weight basis. When such data is desired, a portion of the
sample is weighed out at the same time as the portion used for the analytical
determination.
Warning: The drying oven should be contained in a hood or vented. Significant laboratory
contamination may result from drying a heavily contaminated hazardous waste sample.
11.1.1.2 Immediately after weighing the sample for extraction, weigh 5-10 g of the
sample into a tared crucible. Determine the percent dry weight of the sample
by drying overnight at 105 °C. Allow to cool in a desiccator before
weighing.
g of dry sample
g of sample
11.1.2 Measure 25 g of solid into a 500-mL bottle with a PTFE-lined screw-cap or crimp-top, and
add 500 mL of extraction fluid (Section 6.13). Extract the solid by rotating the bottle at
approximately 30 rpm for 18 hours. Filter the extract through glass-fiber filter paper and
store in a sealed bottle at 4°C. Each mL of extract represents 0.050 g of solid.
11.2 Cleanup and separation.
11.2.1 Cleanup procedures may not be necessary for a relatively clean sample matrix. The cleanup
procedures recommended in this method have been used for the analysis of various sample
types. If particular circumstances demand the use of an alternative cleanup procedure, the
analyst must meet the specifications in Section 9.1.2.
11.2.2 If the sample is not clean, or the complexity is unknown, the entire sample should be
centrifuged at 2500 rpm for 10 minutes. Decant the supernatant liquid from the centrifuge
bottle and filter through glass-fiber filter paper into a container that can be tightly sealed.
11.3 Derivatization.
11.3.1 For aqueous samples, measure a 50-to 100-mL aliquot of sample. Quantitatively transfer the
sample aliquot to the reaction vessel (Section 6.1).
11.3.2 For solid samples, 1-10 mL of leachate (Section 11.1.2 or Section 11.2.2) will usually be
required. The amount used for a particular sample must be determined through preliminary
experiments.
11.3.3 Derivatization and extraction of the derivative can be accomplished using the solid-sorbent
(Section 11.3.4) or methylene chloride option (Section 11.3.5).
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Method 1667, Revision A
11.3.4 Solid Sorbent Option
11.3.4.1 Add 4 mL of phthalate buffer (Section 7.16.2) and adjust the pH to 5.0±0.1
with sodium hydroxide or hydrochloric acid. Add 10 mL of DNPH reagent,
adjust the total volume to approximately 100 mL with reagent water, seal the
container and place on a wrist-action shaker at room temperature for 1 hour.
Samples or standards containing high analyte concentrations may require
more DNPH reagent for complete reaction.
11.3.4.2 Assemble the vacuum manifold and connect to a water aspirator or vacuum
pump. Assemble solid sorbent cartridges containing a minimum of 1.5 g of
C18 sorbent, using connectors supplied by the manufacturer, and attach the
sorbent train to the vacuum manifold. Condition each cartridge by passing
10 mL dilute phthalate buffer (10 mL 5 N phthalate buffer dissolved in 250
mL of reagent water) through the sorbent cartridge train.
11.3.4.3 Remove the reaction vessel from the shaker and add 10 mL of saturated
sodium chloride solution to the vessel.
11.3.4.4 Add the reaction solution to the sorbent train and apply a vacuum so that the
solution is drawn through the cartridges at a rate of 3 to 5 mL/min. After the
solution has eluted, allow air to be drawn through the cartridge for
approximately 2 minutes to remove all traces of solution, then release the
vacuum.
11.3.4.5 Elute each cartridge train with approximately 9 mL of absolute ethanol,
directly into a 10-mL volumetric flask. Dilute the solution to volume with
absolute ethanol, mix thoroughly, and place in a tightly sealed vial until
analyzed.
11.3.5 Methylene chloride option.
11.3.5.1 Add 5 mL of phthalate buffer (Section 7.16.2) and adjust the pH to 5.0±0.1
with sodium hydroxide or hydrochloric acid. Add 10 mL of DNPH reagent,
adjust the volume to approximately 100 mL with reagent water, seal the
container, and place on a wrist-action shaker at room temperature for 1 hour.
Samples or standards with high analyte concentrations may require more
DNPH reagent for complete reaction.
11.3.5.2 Extract the solution with three 20-mL portions of methylene chloride, using
a 250-mL separatory funnel, and combine the methylene chloride layers. If
an emulsion forms upon extraction, remove the entire emulsion and
centrifuge at 2000 rpm for 10 minutes. Separate the layers and proceed with
the next extraction.
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Method 1667, Revision A
11.3.5.3 Assemble a K-D concentrator by attaching a 10-mL concentrator tube to a
500-mL evaporator flask. Wash the K-D apparatus with 25 mL of
extraction solvent to complete the quantitative transfer.
11.3.5.4 Add one or two clean boiling chips to the evaporation flask and attach a
three-ball Snyder column. Prewet the Snyder column by adding about 1 mL
of methylene chloride to the top. Place the K-D apparatus on a hot water
bath (80-90 °C) so that the concentrator tube is partially immersed in the hot
water and the entire lower rounded surface of the flask is bathed with hot
vapor. Adjust the vertical position of the apparatus and the water
temperature, as required, to complete the concentration in 10-15 minutes. 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 of the liquid reaches 10 mL, remove the K-D apparatus and allow it
to drain and cool for at least 10 minutes.
11.3.5.5 Prior to liquid chromatographic analysis, the solvent must be exchanged to
methanol. The analyst must ensure quantitative transfer of the extract
concentrate. The exchange is performed as described below.
11.4 After cooling and draining as described in Section 11.3.5.4, momentarily remove the Snyder column
and add 5 mL of methanol and a new boiling chip. Attach the micro Snyder column. Concentrate the
extract using 1 mL of methanol to prewet the Snyder column. Place the K-D apparatus on the water
bath so that the concentrator tube is partially immersed in the hot water. Adjust the vertical position
of the apparatus and the water temperature, as required, to complete the concentration. When the
apparent volume of the liquid reaches less than 5 mL, remove the K-D apparatus and allow it to drain
and cool for at least 10 minutes.
Remove the Snyder column and rinse the flask and its lower joint with 1-2 mL of methanol and add
to the concentrator tube. A 5-mL syringe is recommended for this operation. Adjust the extract
volume to 10-mL with methanol. Stopper the concentrator tube and store refrigerated at 4 °C if
further processing will not be performed immediately. If the extract will be stored longer than two
days, it should be transferred to a vial with a PTFE-lined screw-cap or crimp-top. Proceed with the
liquid chromatographic analysis if further cleanup is not required.
12.0 High-Performance Liquid Chromatography
12.1 Chromatographic conditions.
Column: C18, 250 mm long x 4.6 mm i.d., 5-(im particle size (or equivalent).
Mobile Phase: Methanol/water, 75:25 (v/v), isocratic at 30 °C.
Flow Rate: 1.0 mL/min.
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Method 1667, Revision A
UV Detector: 365 nm.
Injection Vol.: 20 (iL.
12.2 Analysis.
12.2.1 Analyze samples by HPLC using conditions described in Section 12.1. Table 2 lists the
retention times and MLs that were obtained under these conditions. Other HPLC columns,
chromatographic conditions, or detectors may be used if the requirements of Section 9 are met.
12.2.2 The width of the retention-time window used to make identifications should be based upon
measurements of actual retention-time variations of standards over the course of a day. Three
times the standard deviation of the retention time for a compound can be used to calculate a
suggested window size; however, the experience of the analyst should weigh heavily in the
interpretation of chromatograms.
12.2.3 If the peak area exceeds the linear range of the calibration curve, a smaller sample volume
should be used. Alternatively, the final solution may be diluted with ethanol or methanol, as
appropriate, and reanalyzed.
12.2.4 If the peak area measurement is prevented by the presence of observed interferences, further
cleanup may be required.
12.3 Calculations.
12.3.1 Calculate the calibration factor (CF) at each concentration and the mean calibration factor
(CFm) as follows (mean value based on 5 points):
area response (A]
CF = =
concentration (C)
± CF
mean CF = CF =
12.3.2 Aqueous samples—Calculate the concentration of each analyte as follows:
A * V * DF
— '-
CF * V
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Method 1667, Revision A
where:
RFm is the mean response factor
As is the area signal from the analyte
Ve is the extract volume
DF is the dilution factor; e.g. 10, if the sample is diluted by a factor of 10
Vs is the sample volume
12.3.3 Solid samples—Calculate the concentration of each analyte using the equation below. A
factor must be included in the equation to account for the weight of the sample used and, if
desired, to correct for dry weight.
A * V * DF
mglkg = —
where:
Ws is the sample weight
%m is the percent moisture of the sample
the other symbols are the same as in Section 12.3.2
13.0 Method Performance
13.1 The MDLs listed in Table 2 were obtained using reagent water and methylene chloride extraction.
Similar results can be obtained using the solid-sorbent method.
13.2 This method has been tested for linearity of recovery from spiked reagent water and has been
demonstrated to be applicable over the range from the ML to 50 times the ML.
13.3 A representative chromatogram is presented as Figure 1.
14.0 Pollution Prevention
14.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity or toxicity of
waste at the point of generation. Many opportunities for pollution prevention exist in laboratory
operation. EPA has established a preferred hierarchy of environmental management techniques that
places pollution prevention as the management option of first choice. Whenever feasible, laboratory
personnel should use pollution prevention techniques to address their waste generation. When wastes
cannot be reduced feasibly at the source, the Agency recommends recycling as the next best option.
The acids used in this Method should be reused as practicable by purifying by electrochemical
techniques. The only other chemicals used in this Method are the neat materials used in preparing
69 July 1998
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Method 1667, Revision A
standards. These standards are used in extremely small amounts and pose little threat to the
environment when managed properly. Standards should be prepared in volumes consistent with
laboratory use to minimize the disposal of excess volumes of expired standards.
14.2 For information about pollution prevention that may be applied to laboratories and research
institutions, consult Less is Better: Laboratory Chemical Management for Waste Reduction,
available from the American Chemical Society's Department of Governmental Relations and Science
Policy, 1155 16th Street NW, Washington DC 20036, 202/872-4477.
15.0 Waste Management
15.1 It is the laboratory's responsibility to comply with all Federal, State, and local regulations governing
waste management, particularly the hazardous waste identification rules and land-disposal restrictions.
In addition it is the laboratory's responsibility to protect air, water, and land resources by minimizing
and controlling all releases from fume hoods and bench operations. Also, compliance is required with
any sewage discharge permits and regulations.
15.2 Samples containing acids at a pH of less than 2 are hazardous and must be neutralized before being
poured down a drain or must be handled as hazardous waste.
15.3 For further information on waste management, consult The Waste Management Manual for
Laboratory Personnel and Less is Better: Laboratory Chemical Management for Waste Reduction,
both available from the American Chemical Society's Department of Government Relations and
Science Policy, 1155 16th Street NW, Washington, DC 20036.
July 1998 70
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Method 1667, Revision A
16.0 References
1. "Working with Carcinogens," DREW, PHS, NIOSH, Publication 77-206 (1977).
2. "OSHA Safety and Health Standards, General Industry," 29 CFR 1910, OSHA 2206, (1976).
3. "Safety in Academic Chemistry Laboratories," American Chemical Society Publication, Committee
on Chemical Safety (1979).
4. "Handbook of Analytical Quality Control in Water and Wastewater Laboratories," U.S. EPA, EMSL
Cincinnati, OH 45268, EPA-600/4-79-019 (March 1979).
71 July 1998
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Method 1667, Revision A
17.0 Tables
Table 1. PMI Analytes to Which This Method Applies
PMI Analyte CASRN1
Formaldehyde 50-00-0
Furfural 98-01-1
Isobutyraldehyde 78-84-2
1 Chemical Abstracts Service Registry Number.
Table 2. Retention Times and Minimum Levels (MLs) for PMI Analytes
PMI Analyte Retention Time1 ML2
(seconds)
Formaldehyde 326 50
Furfural 495 50
Isobutyraldehyde 714 50
1 Retention times are for the DNPH derivative.
2 This is the minimum level at which the entire analytical system shall give a
recognizable signal and an acceptable calibration point, taking into account method-
specific sample and injection volumes.
Table 3. Quality Control Acceptance Criteria for Initial Precision and Recovery
PMI Analyte
Formaldehyde
Furfural
Spike
Level (jig/L)
50
100
Average Percent
Recovery (X)
25-187
70-102
Standard E
(s)
81
16
July 1998 72
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Method 1667, Revision A
Isobutyraldehyde 50 45-121 38
73 July 1998
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250 —
200 —
150 —
100 —
SO
10 PPM STANDARD. Amount: 1JOOO.
Acqured on 17-08-9 4 a I 00:43:45 Reported on 08-17-94 a 110:40:44
I
10
20
I
35
Figure 1. Chiomaliogramofthe2,4-DNPH OerJvafe of Forma tie hyde,
Furlural, and Isobutpaldehyde
75
July 1998
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July 1998
76
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Method 1671
Volatile Organic Compounds Specific to the Pharmaceutica
Manufacturing Industry by GC/FID
Revision A, July 1998
-------
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Method 1671, Revision A
Volatile Organic Compounds Specific to the Pharmaceutical Manufacturing
Industry by GC/FID
1.0 Scope and Application
1.1 This method is for surveying and monitoring under the Clean Water Act. The method is used to
determine certain non-purgeable volatile organic pollutants specific to the pharmaceutical
manufacturing industry (PMI) that are amenable to direct aqueous injection gas chromatography (GC)
and detection by a flame ionization detector (FID).
1.2 The PMI analytes listed in Table 1 may be determined in waters, soils, and municipal sludges by this
method.
1.3 The detection limits of Method 1671 are usually dependent on the level of interferences rather than
instrumental limitations. The minimum levels (MLs) in Table 2 are the level that can be attained with
no interferences present.
1.4 This method is recommended for use by, or under the supervision of, analysts experienced in the
operation of gas chromatographs and in the interpretation of chromatograms.
1.5 This method is performance-based. The analyst is permitted to modify the method to overcome
interferences or to lower the cost of measurements, provided that all performance criteria in this
method are met. The requirements for establishing method equivalency are given in Section 9.2.
2.0 Summary of the Method
2.1 The percent solids content of the sample is determined. If the solids content is less than l%,aninternal
standard(s) is added to a 5-mL sample. If the solids content of the sample is greater than 1%, 5 mL
of reagent water and an internal standard(s) is added to a 5-g aliquot of sample.
The mixture is sonicated in a centrifuge tube with little or no headspace for 5 minutes. During this
period the analytes and the internal standard will equilibrate between the solid and aqueous phases.
In some cases, additional sonication will be necessary to establish equilibrium. The resulting
suspension is centrifuged and the supernatant liquid analyzed.
2.2 An appropriate amount of the aqueous solution (or supernate) is injected into the GC. The compounds
are separated by the GC and detected by the FID.
3.0 Definitions
There are no definitions specific to this method.
79 July 1998
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Method 1671, Revision A
4.0 Interferences
4.1 Impurities in the carrier gas, organic compounds outgassing from the GC plumbing, and solvent
vapors in the laboratory account for the majority of contamination problems encountered with this
method. The analytical system is demonstrated to be free from interferences under conditions of the
analysis by analyzing reagent water blanks initially and with each sample batch (samples analyzed on
the same 12-hour shift), as described in Section 9.4.
4.2 Samples can be contaminated by diffusion of volatile organic compounds through the bottle seal during
shipment and storage. A field blank prepared from reagent water and carried through the sampling
and handling protocol may serve as a check on such contamination.
4.3 Contamination by carryover can occur when high-level and low-level samples are analyzed
sequentially. To reduce carryover, the syringe is cleaned or replaced with a new syringe after each
sample is analyzed. When an unusually concentrated sample is encountered, it is followed by analysis
of a reagent water blank to check for carryover. Syringes are cleaned by washing with soap solution,
rinsing with tap and distilled water, and drying in an oven at 100-125 °C. Other parts of the system
are also subject to contamination; therefore, frequent bakeout and purging of the entire system may
be required.
4.4 Interferences resulting from samples will vary considerably from source to source, depending on the
diversity of the site being sampled.
5.0 Safety
The toxicity or carcinogenicity of each analyte, compound, or reagent used in this method has not been
precisely determined; however, each chemical compound should be treated as apotential health hazard.
Exposure to these compounds should be reduced to the lowest possible level. The laboratory is
responsible for maintaining a current awareness file of OSHA regulations regarding the safe handling
of the chemicals specified in this method. A reference file of material safety data sheets should also
be made available to all personnel involved in these analyses. Additional information on laboratory
safety can be found in References 2-4.
6.0 Equipment and Supplies
6.1 Sample bottles and septa
6.1.1 BottlesS25- to 40-mL with polytetrafluoroethylene (PTFE)-lined screw-cap (Pierce 13075,
or equivalent). Detergent wash, rinse with tap and distilled water, and dry at > 105 °C for a
minimum of 1 hour before use.
6.1.2 SeptaSPTFE-faced silicone (Pierce 12722, or equivalent), cleaned as above and baked at 100-
200 °C for a minimum of 1 hour.
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Method 1671, Revision A
6.2 Gas chromatographS Shall be linearly temperature programmable with initial and final holds, and shall
produce results which meet the calibration (Section 10), quality assurance (Section 9), and
performance tests (Section 13) of this method.
6.2.1 ColumnSSO m long x 0.32 mm i.d. fused-silicamicrobore column coated with 4-(im of bonded
poly(dimethylpolysiloxane) (Supelco SPB-1 Sulfur, or equivalent).
6.2.2 GC operating conditions.
Temperatures:
ColumnS2 minutes at 40°C, 10°C per minute to 180°C.
Injection portS200°C
FIDS300°C
Carrier gasSHydrogen at a head pressure of 10 psig.
An injector split may be used in order to optimize peak shape and repeatability.
6.3 SyringesS5-mL, gas-tight glass hypodermic, with Luer-lok tips.
6.4 Micro syringesSlO-, 25-, and 100-(iL.
6.5 Syringe valvesS2-way with Luer ends, PTFE.
6.6 BottlesS15-mL, screw-cap with PTFE liner.
6.7 Balances.
6.7.1 Analytical, capable of weighing 0.1 mg.
6.7.2 Top-loading, capable of weighing 10 mg.
6.8 Equipment for determining percent moisture.
6.8.1 Oven, capable of being temperature-controlled at 110 ° C (±5 ° C).
6.8.2 Desiccator.
6.8.3 BeakersS50-, 100-mL.
6.9 Centrifuge apparatus.
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Method 1671, Revision A
6.9.1 Centrifuge capable of rotating 10-mL centrifuge tubes at 5000 rpm.
6.9.2 Centrifuge tubes, 10-mL, with screw-caps (PTFE-lined) to fit centrifuge.
6.10 Sonication apparatus capable of sonicating 10-mL centrifuge tubes and thoroughly agitating contents.
7.0 Reagents and Standards
7.1 Reagent water: Water in which the compounds of interest and interfering compounds are not detected
by this method. It may be generated by any of the following methods:
7.1.1 Activated carbonSpass tap water through a carbon bed (Calgon Filtrasorb-3 00, or equivalent).
7.1.2 Water purifierSPass tap water through a purifier (Millipore Super Q, or equivalent).
7.1.3 Boil and purgeSHeat tap water to between 90 and 100 °C and bubble contaminant-free inert
gas through it for approximately 1 hour. While still hot, transfer the water to screw-cap
bottles and seal with a PTFE-lined cap.
7.2 Sodium thiosulfateSACS granular.
7.3 Standard solutionsSPurchased as solutions or mixtures with certification to their purity, concentration,
and authenticity, or prepared from materials of known purity and composition. If compound purity
is 96% or greater, the weight may be used without correction to calculate the concentration of the
standard.
7.3.1 Place approximately 8 mL of reagent water in a 10-mL ground-glass-stoppered volumetric
flask. Allow the flask to stand unstoppered for approximately 10 minutes or until all welted
surfaces have dried. For each analyte, weigh the stoppered flask, add the compound,
restopper, then immediately reweigh to prevent evaporation losses from affecting the
measurement.
7.3.2 LiquidsSUsing a microsyringe, add sufficient liquid (about 100 mg) so that the final solution
will have a concentration of about 10 mg/mL.
7.3.3 GasesSFill a valved 5-mL gas-tight syringe with the compound. Lower the needle to
approximately 5 mm above the meniscus. Slowly introduce the compound above the surface
of the meniscus. The gas will dissolve in the solvent. Repeat if necessary to reach desired
concentration.
7.3.4 Fill the flask to volume, stopper, then mix by inverting several times. Calculate the
concentration in milligrams per milliliter (mg/mL, equivalent to micrograms per microliter
[|ig/(iL]) from the weight gain.
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Method 1671, Revision A
7.3.5 Transfer the stock solution to a PTFE-sealed screw-cap bottle. Store, with minimal
headspace, in the dark at approximately 4°C. Do not freeze.
7.3.6 Replace standards after one month, or sooner if comparison with check standards indicate a
change in concentration. Quality control check standards that can be used to determine the
accuracy of calibration standards may be available from the National Institute of Standards
and Technology, Gaithersburg, MD.
7.4 Secondary standardsSUsing standard solutions (Section 7.3), prepare a secondary standard to contain
each pollutant at a concentration of 100 mg/L or 500 mg/L for compounds with higher MLs. Where
necessary, a concentration of 1000 mg/L may be used.
7.4.1 Aqueous calibration standardsSUsing a syringe or a microsyringe, add sufficient secondary
standard (Section 7.4) to five reagent water aliquots to produce concentrations in the range
of interest.
7.4.2 Aqueous performance standardSAn aqueous standard containing all pollutants and internal
standard(s) is prepared daily, and analyzed each shift to demonstrate performance (Section
13). This standard shall contain concentrations of pollutants and internal standard(s), as
appropriate, within a factor of 1 to 5 times the MLs of the pollutants listed in Table 1. It may
be one of the aqueous calibration standards described in Section 7.4.1.
8.0 Sample Collection, Preservation, and Handling
8.1 Grab samples are collected in glass containers having a total volume greater than 20 mL. For aqueous
samples that pour freely, fill sample bottles so that no air bubbles pass through the sample as the bottle
is filled and seal each bottle so that no air bubbles are entrapped. Maintain the hermetic seal on the
sample bottle until time of analysis.
8.2 Maintain samples at 4 °C from the time of collection until analysis. Do not freeze. If an aqueous
sample contains residual chlorine, add sodium thiosulfate preservative (10 mg/40 mL) to the empty
sample bottles just prior to shipment to the sample site. EPA Methods 330.4 and 330.5 may be used
for measurement of residual chlorine (Reference 5). If preservative has been added, shake the bottle
vigorously for 1 minute immediately after filling.
8.3 For aqueous samples, experimental evidence indicates that some PMI analytes are susceptible to rapid
biological degradation under certain environmental conditions. Refrigeration alone may not be
adequate to preserve these compounds in wastewaters for more than seven days. For this reason, a
separate sample should be collected, acidified, and analyzed when compounds susceptible to rapid
biological degradation are to be determined. Collect about 500 mL of sample in a clean container.
Adjust the pH of the sample to about 2 by adding hydrochloric acid (1:1) while stirring. Check pH
with narrow range (1.4 to 2.8) pH paper. Fill a sample bottle as described in Section 8.1. If residual
chlorine is present, add sodium thiosulfate to a separate sample bottle and fill as in Section 8.1.
8.4 All samples shall be analyzed within 14 days of collection.
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Method 1671, Revision A
9.0 Quality Assurance/Quality Control
9.1 Each laboratory that uses this method is required to operate a formal quality assurance program
(Reference 6). The minimum requirements of this program consist of an initial demonstration of
laboratory capability (Section 9.5) and analysis of standards (Sections 9.6 and 13) and blanks (Section
9.4) as tests of continued performance. Each time a batch of samples is analyzed or there is a change
in reagents or procedures, a method blank must be analyzed as a safeguard against contamination.
9.2 In recognition of advances that are occurring in analytical technology, and to allow the analyst to
overcome sample matrix interferences, the analyst is permitted certain options to improve separations
or lower the costs of measurements. These options include alternative concentration and cleanup
procedures, and changes in columns and detectors. Alternative techniques, such as the substitution
of spectroscopy or immunoassay, and changes that degrade method performance are not allowed. If
an analytical technique other than the techniques specified in this method is used, that technique must
have a specificity equal to or better than the specificity of the techniques in this method for the analytes
of interest.
9.2.1 If the detection limit of the method will be affected by the change, the laboratory is required
to demonstrate that the method detection limit (MDL; 40 CFR 136, Appendix B) is lower than
one-third the regulatory compliance level. If calibration will be affected by the change, the
analyst must recalibrate the instrument per Section 10.
9.2.2 The laboratory is required to maintain records of modifications made to this method. These
records include the information in this subsection, at a minimum.
9.2.2.1 The names, titles, addresses, and telephone numbers of the analyst(s) who performed
the analyses and modification, and of the quality control officer who witnessed and
will verify the analyses and modification.
9.2.2.2 A listing of pollutant(s) measured, by name and CAS Registry Number.
9.2.2.3 A narrative stating the reason(s) for the modification.
9.2.2.4 Results from all quality control (QC) tests comparing the modified method to this
method including:
(a) calibration (Section 10);
(b) calibration verification (Section 13);
(c) initial precision and accuracy (Section 9.5);
(d) analysis of blanks (Section 9.4); and
(e) accuracy assessment (Section 9.6 and 13).
9.2.2.5 Data that will allow an independent reviewer to validate each determination by tracing
the instrument output (peak height, area, or other signal) to the final result. These
data are to include:
July 1998 84
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Method 1671, Revision A
(a) sample numbers and other identifiers;
(b) analysis dates and times;
(c) injection logs;
(d) analysis sequence/run chronology;
(e) sample weight or volume;
(f) sample volume prior to each cleanup step, if applicable;
(g) sample volume after each cleanup step, if applicable;
(h) final sample volume prior to injection;
(I) injection volume;
(j) dilution data, differentiating between dilution of a sample or an extract;
(k) instrument and operating conditions;
(1) column (dimensions, liquid phase, solid support, film thickness, etc.);
(m) operating conditions (temperature, temperature program, flow rates, etc.);
(n) detector (type, operating condition, etc.);
(o) chromatograms, printer tapes, and other recording of raw data; and
(p) quantitation reports, data system outputs, and other data necessary to link
raw data to the results reported.
9.3 With each sample batch, a matrix spike (MS) and matrix spike duplicate (MSB) are analyzed to assess
precision and accuracy of the analysis. The relative percent difference (RPD) between the MS and
MSB shall be less than 30% and compound recoveries shall fall within the limits specified in Table
3. If the recovery of any compound falls outside its warning limit, method performance is
unacceptable for that compound in that sample and the results may not be reported for regulatory
compliance purposes.
9.4 Analyses of blanks are required to demonstrate freedom from contamination and that the compounds
of interest and interfering compounds have not been carried over from a previous analysis (Section
4.3).
9.4.1 With each sample batch (samples analyzed on the same 12-hour shift), a blank shall be analyzed
immediately after analysis of the aqueous performance standard (Sections 9.6 and 13) to demonstrate
freedom from contamination. If any of the compounds of interest or any potentially interfering
compound is found in a blank at greater than the ML (assuming a response factor of 1 relative to the
nearest-eluted internal standard for compounds not listed in Table 1), analysis of samples is halted
until the source of contamination is eliminated and a blank shows no evidence of contamination at this
level.
9.5 Initial precision and recovery—To establish the ability to generate acceptable precision and accuracy,
the analyst shall perform the following operations for compounds to be calibrated.
9.5.1 Analyze two sets of four 5-mL aliquots (eight aliquots total) of the aqueous performance standard
(Section 7.4.2) containing the PMI analytes listed in Table 1.
9.5.2 Using the first set of four analyses, compute the average recovery (X) in percent of spike level and
standard deviation of the recovery (s) in percent of spike level, for each compound.
85 July 1998
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Method 1671, Revision A
9.5.3 For each compound, compare s and X with the corresponding limits for initial precision and accuracy
found in Table 3. If s and X for all compounds meet the acceptance criteria, system performance is
acceptable and analysis of blanks and samples may begin. If, however, any individual s exceeds the
precision limit or any individual X falls outside the range for accuracy, system performance is
unacceptable for that compound.
9.5.4 Using the results of the second set of analyses, compute s and X for only those compounds that failed
the test of the first set of four analyses (Section 9.5.3). If these compounds now pass, the system
performance is acceptable for all compounds, and analysis of blanks and samples may begin. If,
however, any of the same compounds fail again, the analysis system is not performing properly for the
compound(s) in question. In this event, correct the problem and repeat the entire test (Section 9.5).
9.6 The laboratory shall, on an ongoing basis, demonstrate through the analysis of the aqueous
performance standard (Section 7.4.2) that the analysis system is in control. This procedure is
described in Section 13.
9.7 Where available, field replicates may be used to validate the precision of the sampling technique.
9.8 The laboratory shall maintain records to define the quality of data that is generated.
10.0 Calibration
10.1 Inject standards into the GC and adjust the sensitivity to detect an amount of each compound less than
or equal to one-third of the ML listed in Table 2 for the analyte.
10.2 Internal standard calibration procedure. The analyst must select one or more internal standards that
are similar in analytical behavior to the compounds of interest. The analyst must further demonstrate
that the measurement of the internal standard(s) is not affected by method or matrix interferences.
Because of these limitations, no internal standard that would be applicable to all samples can be
required in the method. The method was developed using tetrahydrofuran (THF) as an internal
standard. Where THF is not present in the sample matrix and no interference precludes its use, THF
is to be used as an internal standard for application of this method. If interferences preclude use of
THF and other internal standards, external standard calibration may be used.
10.2.1 Prepare aqueous calibration standards at a minimum of five concentration levels for each
analyte by carefully adding an appropriate amount of secondary standard to reagent water or
to the matrix under study. One of the concentrations should be at or below the ML. The
concentration range should bracket the concentrations expected in the samples and should not
exceed the dynamic range of the GC/FID instrument. These aqueous standards must be
prepared daily.
10.2.2 Prepare a spiking solution containing the internal standard(s) using the procedures described
in Sections 7.3 and 7.4 and add an appropriate amount of internal standard to each aqueous
calibration standard.
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Method 1671, Revision A
10.2.3 Using injections appropriate to optimize system sensitivity and separation of the analytes,
analyze each calibration standard and tabulate peak height or area responses against
concentration for each compound and internal standard. Calculate response factors (RF) for
each compound as follows:
As x cs
where:
As = Response for the analyte to be measured
Ais = Response for the nearest e luting internal standard
Cis = Concentration of the nearest eluting internal standard
Cs = Concentration of the analyte to be measured
If the RF value over the working range is a constant (less than 10% relative standard
deviation), the RF can be assumed to be invariant and the average RF can be used for
calculations. Alternatively, the results can be used to plot a calibration curve of relative
response, AS*C1S/A1S, against analyte concentration (Cs).
11.0 Sample Preparation
Samples containing less than 1% solids are analyzed directly as aqueous samples. Samples containing
1 % solids or greater are analyzed after equilibration with reagent water containing internal standard(s) .
11.1 Determination of percent solids.
11.1.1 Weigh 5 - 1 0 g of sample into a tared beaker.
11.1.2 Dry overnight (12 hours minimum) at 1 10±5 °C, and cool in a desiccator.
11.1.3 Determine the percent solids as follows:
% solids = ^9ht of sample dry x 1QQ
weight of sample wet
11.2 Remove standards and samples from cold storage and bring to 20-25 °C.
11.3 Samples containing less than 1% solids.
11.3.1 Allow solids to settle and remove 5 mL of sample.
11.3.2 Add an appropriate amount of internal standard spiking solution.
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Method 1671, Revision A
11.3.3 Inject a sample directly into the GC. The temperature of the injection block should be great
enough to immediately vaporize the entire sample. An example of the separations achieved
by the column listed is shown in Figure 1.
Note: Use of a 0.2-(iL injection has been found to improve method sensitivity over a larger injection
combined with a split sample. Where possible, splitless injection should be used. All requirements
of this Method must be met regardless of type of injection used.
11.4 Samples containing 1% solids or greater.
11.4.1 Mix the sample thoroughly using a clean spatula and remove rocks, twigs, sticks, and other
foreign matter.
11.4.2 Add 5±1 g of sample to a tared 10-mL centrifuge tube. Using a clean metal spatula, break
up any lumps of sample. Record the sample weight to three significant figures.
11.4.3 Add an appropriate amount of internal standard spiking solution to the sample in the
centrifuge tube.
11.4.4 Add a measured quantity (Y ± 0.1 mL) of reagent water to the tube so as to minimize head
space.
11.4.5 Place a cap on the centrifuge tube leaving little or no headspace. Place the tube in the
sonicator for a minimum of 5 minutes, turning occasionally. For most samples this should be
sufficient time to distribute the analytes and standard(s) between the solid and aqueous phases
and to establish equilibrium. Some sample matrices may require more sonication.
11.4.6 On completion of sonication, centrifuge the sample and inject the same amount of supernate
into the GC that was injected for the calibration standards.
11.5 For liquid samples containing high-solids concentrations, such as sludges or muds, weigh
approximately 5 g (to three significant figures) into a 10-mL centrifuge tube, add an appropriate
amount of internal standard solution, sonicate, centrifuge, and inject as in Section 11.4.6.
12.0 Quantitative Determination
12.1 The calibration curve or averaged response factor determined during calibration is used to calculate
the concentration. For calculation using the averaged RF, the equation below is used, and the terms
are as defined in Section 10.2.3.
As x C,s
Concentration = — -
As x RF
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Method 1671, Revision A
12.2 The concentration of the pollutant in the solid phase of the sample is computed using the concentration
of the pollutant detected in the aqueous solution, as follows:
Concentration in solid(mg/kg) = YL x aclueous conc (m9/L) x percent solids x DF
sample wt (kg)
where:
percent solids is from Section 11.1
Y = Volume of water in liters (L) from 11.4.4
DF = Dilution factor (as a decimal number), if necessary
12.3 Sample dilution—If the calibration range of the system is exceeded, the sample is diluted by successive
factors of 10 until the sample concentration is within the calibration range.
12.4 Report results for all pollutants found in standards, blanks, and samples to three significant figures.
For samples containing less than 1% solids, the units are milligrams per liter (mg/L); and for samples
containing 1% solids or greater, units are milligrams per kilogram (mg/kg).
13.0 System Performance
13.1 At the beginning of each 12-hour shift during which analyses are performed, system calibration and
performance shall be verified. Acceptance criteria for each compound (R) are found in Table 3.
Adjustment and/or recalibration shall be performed until all performance criteria are met. Only after
all performance criteria are met may blanks and samples be analyzed.
13.2 Where THF is used as the internal standard, the absolute retention time of THF shall be 416 seconds
(± 30 seconds). The relative retention times of all pollutants shall fall within 10% of the value given
in Table 2.
14.0 Method Performance
14.1 This method was developed and validated in a single laboratory.
14.2 A chromatogram of the aqueous performance standard is shown in Figure 1.
15.0 Pollution Prevention
15.1 Pollution prevention encompasses any technique that reduces or eliminates the quantity or toxicity of
waste at the point of generation. Many opportunities for pollution prevention exist in laboratory
operation. EPA has established a preferred hierarchy of environmental management techniques that
places pollution prevention as the management option of first choice. Whenever feasible, laboratory
personnel should use pollution prevention techniques to address their waste generation. When wastes
cannot be reduced feasibly at the source, the Agency recommends recycling as the next best option.
89 July 1998
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Method 1671, Revision A
The acids used in this Method should be reused as practicable by purifying by electrochemical
techniques. The only other chemicals used in this Method are the neat materials used in preparing
standards. These standards are used in extremely small amounts and pose little threat to the
environment when managed properly. Standards should be prepared in volumes consistent with
laboratory use to minimize the disposal of excess volumes of expired standards.
15.2 For information about pollution prevention that may be applied to laboratories and research
institutions, consult Less is Better: Laboratory Chemical Management for Waste Reduction,
available from the American Chemical Society's Department of Governmental Relations and Science
Policy, 1155 16th Street NW, Washington DC 20036, 202/872-4477.
16.0 Waste Management
16.1 It is the laboratory's responsibility to comply with all Federal, State, and local regulations governing
waste management, particularly the hazardous waste identification rules and land-disposal restrictions.
In addition, it is the laboratory's responsibility to protect air, water, and land resources by minimizing
and controlling all releases from fume hoods and bench operations. Also, compliance is required with
any sewage discharge permits and regulations.
16.2 Samples containing acids at a pH of less than 2 are hazardous and must be neutralized before being
poured down a drain or must be handled as hazardous waste.
16.3 For further information on waste management, consult The Waste Management Manual for
Laboratory Personnel and Less is Better: Laboratory Chemical Management for Waste Reduction,
both available from the American Chemical Society's Department of Government Relations and
Science Policy, 1155 16th Street NW, Washington, DC 20036.
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Method 1671, Revision A
17.0 References
1. "Standard Test Method for Volatile Alcohols in Water by Direct Aqueous-Injection Gas
Chromatography." 1994 Annual Book of ASTM Standards, Volume 11.02 (Water (II)). ASTM, 1916
Race Street, Philadelphia, PA 19103-1187.
2. "Working with Carcinogens," DREW, PHS, NIOSH, Publication 77-206 (1977).
3. "OSHA Safety and Health Standards, General Industry," 29 CFR 1910, OSHA 2206 (1976).
4. "Safety in Academic Chemistry Laboratories," American Chemical Society Publication, Committee
on Chemical Safety (1979).
5. "Methods 330.4 and 330.5 for Total Residual Chlorine," USEPA, EMSL Cincinnati, OH 45268.
6. "Method of Analytical Quality Control in Water and Wastewater Laboratories," USEPA, EMSL
Cincinnati, OH 45268, EPA-4-79-019 (March, 1979).
7. Technical Report to PhRMA from Tichler & Kocurek by Malcolm Pirnie Laboratory, EPA Water
Docket for Pharmaceutical Manufacturing Industry rule proposed May 2, 1995 (60 FR 21592),
Document Control Number 8166 at Record Section 13.2.4. (February 13, 1997).
91 July 1998
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Method 1671, Revision A
18.0 Tables
Table 1. Non-purgeable Water Soluble PMI Analytes to be Analyzed by Direct Aqueous
Injection GC/FID and Internal Standard Techniques
PMI Analyte
Acetonitrile
Diethylamine
Dimethylamine
Dimethyl sulfoxide
Ethanol
Ethylene glycol
Formamide
Methanol
Methylamine
Methyl Cellosolve® (2-methoxyethanol)
n-Propanol
Triethylamine
CASRN1
75-05-8
109-89-7
124-40-3
67-68-5
64-17-5
107-21-1
75-12-7
67-56-1
74-89-5
109-86-4
71-23-8
121-44-8
EPA-EGD
972
986
987
1037
134
1038
988
135
989
1040
955
990
Chemical Abstracts Service Registry Number
July 1998
92
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Method 1671, Revision A
Table 2.
EGD
No.
989
135
987
134
972
955
986
975
1040
988
1038
990
1037
Gas Chromatographic Retention Times and Minimum Levels for
Soluble PMI Analytes by Direct Aqueous Injection GC/FID
Non-purgeable Water
Retention Time
PMI Analyte
Methylamine
Methanol
Dimethylamine
Ethanol
Acetonitrile
n-Propanol
Diethylamine
Tetrahydrofuran (int std)
Methyl Cellosolve®
Formamide
Ethylene glycol
Triethylamine
Dimethyl sulfoxide
Mean (sec)
128
139
165
188
203
307
341
416
429
473
495
518
676
EGD Ref
975
975
975
975
975
975
975
975
975
975
975
975
975
Relative
0.307
0.334
0.396
0.452
0.488
0.737
0.819
1.000
1.030
1.136
1.189
1.244
1.624
ML1
(mg/L)
50
2(2)
50
2(2)
50
50
50
20
100
100
50
20
1 This is a minimum level at which the entire analytical system shall give an acceptable calibration point,
taking into account method-specific sample and injection volumes. The concentration in the aqueous or solid
phase is determined using the equations in Section 12.
2 The minimum level for this analyte was developed from data provided in Reference 7.
93
July 1998
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Method 1671, Revision A
Table
3. Acceptance Criteria for Performance Tests
Acceptance Criteria (%
Initial Precision and
Accuracy
EGD
No.
972
986
987
1037
134
1038
988
135
989
1040
955
990
PMI Analyte
Acetonitrile
Diethylamine
Dimethylamine
Dimethyl sulfoxide
Ethanol
Ethylene glycol
Formamide
Methanol
Methylamine
Methyl Cellosolve®
n-Propanol
Triethylamine
Spike
Level
50
50
50
50
50
100
200
50
50
50
50
50
s
30
20
27
20
20
22
20
21
20
20
25
47
70
65
70
31
70
70
70
70
70
64
70
70
X
- 146
- 130
- 153
- 130
- 131
- 149
- 130
- 130
- 130
- 130
- 137
- 165
of Spike Level)
On-going
Accuracy
70
70
70
30
70
70
70
70
70
64
70
68
R
- 148
- 130
- 155
- 130
- 132
- 150
- 130
- 130
- 130
- 130
- 139
- 168
July 1998
94
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Method 1671, Revision A
100% -,
FS -
0
Aialysis :4MDLJ,a, 1
Crated a UCMficn Of/DscA4
Sample* :S
hjeciori*: 1
Sample Name: MDL#3
2 3 45 678 9 10 11 12
Figure 1. C h ro matog ra m o f Aqueous Be rtor ma nee
Standard of Anatytes torn Table 1
95
July 1998
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