EPA Document #: EPA/600/R-05/052
METHOD 529 DETERMINATION OF EXPLOSIVES AND RELATED
COMPOUNDS IN DRINKING WATER BY SOLID PHASE
EXTRACTION AND CAPILLARY COLUMN GAS
CHROMATOGRAPHY/MASS SPECTROMETRY (GC/MS)
Revision 1.0
September, 2002
J.W. Munch
NATIONAL EXPOSURE RESEARCH LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
CINCINNATI, OHIO 45268
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METHOD 529
DETERMINATION OF EXPLOSIVES AND RELATED COMPOUNDS IN DRINKING
WATER BY SOLID PHASE EXTRACTION AND CAPILLARY COLUMN
GAS CHROMATOGRAPHY/MASS SPECTROMETRY(GC/MS)
1. SCOPE AND APPLICATION
1.1 This method provides procedures for the determination of explosives and related
compounds in finished drinking water. The method may be applicable to untreated
source waters and other types of water samples, but it has not been evaluated for
these uses. The method is applicable to a variety of explosives that are efficiently
partitioned from the water sample onto a solid phase extraction (SPE) sorbent, and
sufficiently volatile and thermally stable for gas chromatography. The method
includes the following compounds:
ANALYTE
2-amino-4,6-dinitrotoluene
4-amino-2,6-dinitrotoluene
3,5-dinitroanaline
1 ,3-dinitrobenzene
2 ,4-dinitrotoluene
2, 6-dinitro toluene
hexahydro- 1 ,3,5-trinitro- 1 ,3,5-triazine (RDX)
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrophenylmethylnitramine (Tetryl)
2,4,6-trinitrotoluene (TNT)
CAS NUMBER
35572-78-2
19406-51-0
618-87-1
99-65-0
121-14-2
606-20-2
121-82-4
98-95-3
88-72-2
99-08-1
99-99-0
99-35-4
479-45-8
118-96-7
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1.2 Detection limit (DL) is defined as the statistically calculated minimum concentration
that can be measured with 99% confidence that the reported value is greater than
zero (1). The DL is compound dependent and is particularly dependent on
extraction efficiency, sample matrix, and instrument performance. DLs for method
analytes are listed in Tables 3, 9, and 15.
1.3 This method should be performed only by analysts with experience in solid phase
extractions and GC/MS analyses.
2. SUMMARY OF METHOD
Analytes and surrogates are extracted by passing a 1 L water sample through a solid phase
extraction (SPE) cartridge containing 500 mg of a divinylbenzene/vinylpyrrolidone
copolymer or a 47 mm disk containing a reverse phase sulfonated polystyrene divinyl-
benzene phase. The organic compounds are eluted from the solid phase with a small
quantity of ethyl acetate. The sample components are separated, identified, and measured
by injecting an aliquot of the concentrated extract into a high resolution fused silica
capillary column of a GC/MS system with a programmed temperature vaporizing (PTV)
injector. Compounds eluting from the GC column are identified by comparing their
measured mass spectra and retention times to reference spectra and retention times in a
data base. Analysis may be performed using either full scan or selected ion monitoring
(SIM) mass spectrometry. Reference spectra and retention times for analytes are obtained
by the measurement of calibration standards under the same conditions used for samples.
The concentration of each identified component is measured by relating the MS response
of the quantitation ion(s) produced by that compound to the MS response of the
quantitation ion(s) produced by a compound that is used as an internal standard. Surrogate
analytes, whose concentrations are known in every sample, are measured with the same
internal standard calibration procedure.
The method provides instructions and demonstration data for both the SPE disk and
cartridge extraction techniques and for three different GC/MS techniques: PTV splitless
injection with full scan mass spectrometry, cold on-column injection with full scan mass
spectrometry, and PTV splitless injection with SIM mass spectrometry.
3. DEFINITIONS
3.1 ANALYSIS BATCH -- A set of samples analyzed on the same instrument during a
24 hour period that begins and ends with the analysis of the appropriate Continuing
Calibration Check (CCC) standards. Additional CCCs may be required depending
on the length of the analysis batch and/or the number of Field Samples.
3.2 CALIBRATION STANDARD (CAL) -- A solution prepared from the primary
dilution standard solution or stock standard solutions and the internal standards and
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surrogate analytes. The CAL solutions are used to calibrate the instrument response
with respect to analyte concentration.
3.3 CONTINUING CALIBRATION CHECK (CCC) -- A calibration standard
containing one or more method analytes, which is analyzed periodically to verify the
accuracy of the existing calibration for those analytes.
3.4 DETECTION LIMIT (DL) ~ The minimum concentration of an analyte that can be
identified, measured and reported with 99% confidence that the analyte
concentration is greater than zero. This is a statistical determination (Sect. 9.2.4),
and accurate quantitation is not expected at this level (1).
3.5 EXTRACTION BATCH -- A set of up to 20 field samples (not including QC
samples) extracted together by the same person(s) during a work day using the same
lot of solid phase extraction devices and solvents, surrogate solution, and fortifying
solutions. Required QC samples for each extraction batch include: Laboratory
Reagent Blank, Laboratory Fortified Blank, Laboratory Fortified Matrix, and either a
Field Duplicate or Laboratory Fortified Matrix Duplicate.
3.6 FIELD DUPLICATES (FD1 and FD2) -- Two separate samples collected at the
same time and place under identical circumstances, and treated exactly the same
throughout field and laboratory procedures. Analyses of FD1 and FD2 give a
measure of the precision associated with sample collection, preservation, and
storage, as well as with laboratory procedures.
3.7 INTERNAL STANDARD (IS) -- A pure analyte(s) added to a sample, extract, or
standard solution in known amount(s) and used to measure the relative responses of
other method analytes and surrogates that are components of the same solution. The
internal standard must be an analyte that is not a sample component.
3.8 LABORATORY FORTIFIED BLANK (LFB) -- An aliquot of reagent water or other
blank matrix to which known quantities of the method analytes are added in the
laboratory. The LFB is analyzed exactly like a sample, including the use of sample
preservatives, and its purpose is to determine whether the methodology is in control,
and whether the laboratory is capable of making accurate and precise measurements.
3.9 LABORATORY FORTIFIED SAMPLE MATRIX (LFM) -- An aliquot of an
environmental sample to which known quantities of the method analytes are added
in the laboratory. The LFM is analyzed exactly like a sample, and its purpose is to
determine whether the sample matrix contributes bias to the analytical results. The
background concentrations of the analytes in the sample matrix must be determined
in a separate aliquot and the measured values in the LFM corrected for background
concentrations.
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3.10 LABORATORY FORTIFIED SAMPLE MATRIX DUPLICATE (LFMD) -- A
second aliquot of the Field Sample, or duplicate Field Sample, that is used to prepare
the LFM. The LFMD is fortified, extracted and analyzed identically to the LFM.
The LFMD is used instead of the Laboratory Duplicate to assess method precision
when the occurrence of target analytes are low.
3.11 LABORATORY REAGENT BLANK (LRB) -- An aliquot of reagent water or other
blank matrix that is treated exactly as a sample including exposure to all glassware,
equipment, solvents, reagents, internal standards, surrogates, and sample
preservatives that are used with other samples. The LRB is used to determine if
method analytes or other interferences are present in the laboratory environment, the
reagents, or the apparatus.
3.12 MATERIAL SAFETY DATA SHEET (MSDS) -- Written information provided by
vendors concerning a chemical's toxicity, health hazards, physical properties, fire,
and reactivity data including storage, spill, and handling precautions.
3.13 MINIMUM REPORTING LEVEL (MRL) -- The minimum concentration that can
be reported as a quantitated value for a target analyte in a sample following analysis.
This defined concentration can be no lower than the concentration of the lowest
calibration standard for that analyte, and can only be used if acceptable quality
control criteria for the analyte at this concentration are met.
3.14 PRIMARY DILUTION STANDARD SOLUTION (PDS) -- A solution of several
analytes prepared in the laboratory from stock standard solutions and diluted as
needed to prepare calibration solutions and other needed analyte solutions.
3.15 PROGRAMMED TEMPERATURE VAPORIZING INJECTOR (PTV) -- A GC
injector capable of rapid heating. Typical use of a PTV injector involves introducing
the sample with the injector cool, then rapidly heating it at 100-200 degrees per
minute to volatilize the analytes onto the GC column. One advantage of this type of
injection is that thermally labile analytes in a mixture can be transferred to the GC
column at a lower temperature than in conventional hot injections.
3.16 QUALITY CONTROL SAMPLE (QCS) -- A solution of method analytes of known
concentrations that is obtained from a source external to the laboratory and different
from the source of calibration standards. It is used to check laboratory performance
with externally prepared test materials.
3.17 SELECTED ION MONITORING (SIM) -- A chromatographic/mass spectrometric
technique where only one or a few ions are monitored. When used with gas
chromatography, the set of ions monitored is usually changed periodically
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throughout the chromatographic run, to correlate with the characteristic ions of each
analyte or group of analytes as they elute from the chromatographic column.
3.18 STOCK STANDARD SOLUTION (SSS) -- A concentrated solution containing one
or more method analytes prepared in the laboratory using assayed reference materials
or purchased from a reputable commercial source.
3.19 SURROGATE ANALYTE (SUR) -- A pure analyte, which is extremely unlikely to
be found in any sample, and which is added to a sample aliquot in a known amount
before extraction or other processing, and is measured with the same procedures
used to measure other sample components. The purpose of the SUR is to monitor
method performance with each sample.
4. INTERFERENCES
4.1 During analysis, major contaminant sources are reagents and SPE devices. Analyses
of laboratory reagent blanks provide information about the presence of
contaminants. Solid phase extraction devices described in this method have two
potential sources of contamination, both the solid phase sorbent and for cartridge
extractions, the polypropylene cartridge that it is packed in. Manufacturers' brands
and lot numbers of these devices should be monitored and tracked to ensure that
contamination will not preclude analyte identification and quantitation.
4.2 Interfering contamination may occur when a sample containing low concentrations
of compounds is analyzed immediately after a sample containing relatively high
concentrations of compounds. Injection port liners must be replaced as needed
(cleaning and deactivation by the analyst is not recommended). After analysis of a
sample containing high concentrations of compounds, a laboratory reagent blank
should be analyzed to ensure that accurate values are obtained for the next sample.
In the case of automated analysis, the analyst may not be aware of high concentration
samples until after an entire batch is analyzed. In this situation, the analyst should
carefully review data from samples analyzed immediately after high concentration
samples, and reanalyze them if necessary.
5. SAFETY
5.1 The toxicity or carcinogenicity of many of the chemicals used in this method has not
been precisely defined; each chemical should be treated as a potential health hazard,
and exposure to these chemicals should be minimized. Each laboratory is
responsible for maintaining awareness of OSHA regulations regarding safe handling
of chemicals used in this method. Each laboratory should maintain a file of
applicable MSDSs.
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5.2 Some method analytes including 2,4- and 2,6-dinitrotoluene, RDX, and TNT have
been classified as known or suspected human or mammalian carcinogens. Pure
standard materials and stock standard solutions of these compounds should be
handled with suitable protection to skin, eyes, etc. (2-5).
5.3 RDX, Tetryl and TNT are explosives and the neat materials require special handling to
be used safely. It is highly recommended that only dilute solutions of these
materials, available as standards from commercial suppliers be used for this
method (6).
6. EQUIPMENT AND SUPPLIES (References to specific brands or catalog numbers are
included for illustration only, and do not imply endorsement of the product.)
6.1 GLASSWARE ~ All glassware must be meticulously cleaned. This may be
accomplished by washing with detergent and water, rinsing with water, distilled
water, and solvent rinsing or heating (where appropriate) at 400° C for 2 h in a
muffle furnace. Volumetric glassware should never be heated to the temperatures
obtained in a muffle furnace.
6.2 SAMPLE CONTAINERS - 1 L or 1 qt amber glass bottles fitted with
polytetrafluoroethylene (PTFE) lined polypropylene screw caps. Amber bottles are
highly recommended since some of the method analytes are sensitive to light and
may degrade upon exposure. Clear glass bottles may be used if they are wrapped in
foil, or samples are stored in boxes that prevent exposure to light.
6.3 VOLUMETRIC FLASKS - various sizes.
6.4 LABORATORY OR ASPIRATOR VACUUM SYSTEM - Sufficient capacity to
maintain a vacuum of approximately 25 cm (10 in.) of mercury.
6.5 MICRO SYRINGES ~ various sizes.
6.6 VIALS ~ Various sizes of amber vials with PTFE lined screw caps for storing
standard solutions and extracts.
6.7 DRYING COLUMN ~ The drying tube should contain about 5 to 7 grams of
anhydrous sodium sulfate to remove residual water from the extract. Any small tube
may be used, such as a syringe barrel, a glass dropper, etc. as long as no particulate
sodium sulfate passes through the column into the extract.
6.8 ANALYTICAL BALANCE - Capable of weighing 0.0001 g accurately.
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6.9 FUSED SILICA CAPILLARY GAS CHROMATOGRAPHY COLUMN -- Any
capillary column that provides adequate resolution, capacity, accuracy, and precision
can be used. Medium polarity, low bleed columns are recommended for use with
this method to provide adequate chromatography and minimize column bleed.
Deactivated injection port liners are highly recommended. Example retention times
are presented in Table 2.
6.9.1. Column - 15 m x 0.25 mm i.d. fused silica capillary column coated with a
0.25 (unbonded film of polyphenylmethylsilicone, (Agilent DB-5ms or
equivalent).
NOTE: Although other columns may be used, longer or thicker film
columns are not recommended because increased residence time in the
column has been shown to contribute to analyte degradation (7).
6.10 GAS CHROMATOGRAPH/MASS SPECTROMETER/DATA SYSTEM
(GC/MS/DS)-
6.10.1 The GC must be capable of temperature programming and should be
equipped for PTV splitless or cold on-column injection. The injection
system should not allow the analytes to contact hot stainless steel or other
metal surfaces that promote decomposition. Satisfactory data could not be
demonstrated for this method using hot splitless injections.
A large volume injector may be used to increase method sensitivity, or to
avoid extract evaporation, if all criteria in Sections 9 and 10 are met.
6.10.2 Deactivated injection port liners should be used, and it is highly
recommended that they be replaced when necessary with a new deactivated
liner. Cleaning and deactivation of injection port liners by the analyst is not
recommended. The injection port liner should be replaced as necessary to
meet all QC requirements. The frequency will vary by the type of
instrument used and the type and number of samples analyzed.
In general, packed injection port liners should not be used with this method.
The use of liner packings such as glass wool and fused silica wool during
development of this method caused significant breakdown of some method
analytes, particularly RDX. Carbofrits (Restek) did show acceptable
performance and may be used with PTV splitless injection. (The data
shown in this method in Tables 3-8 were generated using a Carbofrit.)
6.10.3 The mass spectrometer must be capable of electron ionization at a nominal
electron energy of 70 eV to produce positive ions. The spectrometer must
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be capable of scanning at a minimum from 45 to 250 amu with a complete
scan cycle time (including scan overhead) of 0.7 sec or less. (Scan cycle
time = total MS data acquisition time in sec divided by number of scans in
the chromatogram). The spectrometer must produce a mass spectrum that
meets all criteria in Table 1 when an injection of approximately 5 ng of
BFB is introduced into the GC. A single spectrum at the apex of the
chromatographic peak, or an average of the three spectra at the apex of the
peak, or an average spectrum across the entire GC peak may be used to
evaluate the performance of the system. Background subtraction is
permitted. The scan time must be set so that all analytes have a minimum
of 5 scans across the chromatographic peak. Seven to ten scans across
chromatographic peaks are strongly recommended.
NOTE: Although BFB is most commonly used as the tune check for
volatile compound analysis, it was selected for use in this method for
semivolatile analytes because the ions used for the BFB tune check are
similar to the analyte quantitation ions in this method.
6.10.4 If the SIM option is selected for use with this method, the mass
spectrometer must be capable of this technique. Some ion trap mass
spectrometers have an option similar to SIM known as Selected Ion Storage
(SIS). Selected ion storage may be used with this method, however a
preliminary assessment during method development showed little if any
increase in method sensitivity using SIS.
6.10.5 An interfaced data system is required to acquire, store, reduce, and output
mass spectral data. The computer software should have the capability of
processing stored GC/MS data by recognizing a GC peak within any given
retention time window. The software must also allow integration of the ion
abundance of any specific ion between specified time or scan number
limits, calculation of response factors as defined in Sect. 10.2.5 or
construction of a linear regression calibration curve, and calculation of
analyte concentrations.
6.11 VACUUM MANIFOLD -- A vacuum manifold (Supelco # 57030 and #57275 for
extraction cartridges or Varian #12146001 for disks, or equivalent) is required for
processing samples through the extraction/elution procedure. An automatic or
robotic sample preparation system designed for use with solid phase extraction
cartridges/disks may be utilized in this method if all quality control requirements
discussed in Sect. 9 are met. Automated systems may use either vacuum or positive
pressure to process samples and solvents through the disk or cartridge. All
extraction and elution steps must be the same as in the manual procedure.
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Extraction and/or elution steps may not be changed or omitted to accommodate the
use of an automated system.
7. REAGENTS AND STANDARDS
7.1 HELIUM ~ carrier gas, purity as recommended by the GC/MS manufacturer.
7.2 SOLID PHASE EXTRACTION CARTRIDGES -- Waters Porapak RDX.
Cartridges are inert non-leaching plastic, for example polypropylene, or glass, and
must not contain plasticizers that leach into the ethyl acetate eluant and prevent the
identification and quantitation of method analytes. The polypropylene cartridges (6
mL volume) are packed with 500 mg of a divinylbenzene/vinylpyrrolidone
copolymer. The packing must have a narrow size distribution and must not leach
interfering organic compounds into the eluting solvent.
7.3 SOLID PHASE EXTRACTION DISKS - 3M Empore SDB-RPS, 47mm.
NOTE: This SPE material is a chemically modified polystyrene divinyl benzene
material. Use of conventional polystyrene divinyl benzene materials will not result
in acceptable data.
7.4. COPPER SULFATE -- ACS grade. Used as a sample preservative for inhibiting
microbial activity in stored samples.
7.5 TRIZMA PRE-SET CRYSTALS (pH 7.0 BUFFER) -- Tris (hydroxymethyl)
aminomethane and Tris hydrochloride (Sigma Chemical Co.). Used as a binding
agent for free chlorine, and to keep the copper in solution. Alternatively, a two
component buffer of tris(hydroxymethyl)aminomethane [CAS# 77-86-1] and
tris(hydroxymethyl)aminomethane hydrochloride [CAS# 1185-53-1] maybe used.
Proportions of each to be used are listed in Sect. 8.2.1.
7.6 SOLVENTS -
7.6.1 ETHYL ACETATE AND METHANOL - High purity pesticide quality or
equivalent.
7.6.2 REAGENT WATER ~ Water in which an interference is not observed at
>l/3 the MRL of any of the compounds of interest. Prepare reagent water
by passing tap water through a filter bed containing about 0.5 kg of
activated carbon or by using a water purification system. Store in clean,
narrow-mouth bottles with PTFE lined septa and screw caps.
7.7 SODIUM SULFATE, ANHYDROUS - ACS grade. Material should be Soxhlet
extracted with methylene chloride for a minimum of 4 h or heated to 400° C for 2 h
in a muffle furnace.
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7.8 ANALYTE STOCK STANDARD SOLUTIONS -- It is recommended that
individual solutions of analytes, or mixtures of analytes, be purchased from
commercial suppliers. All work done during development of this method was done
with commercial mixtures. RDX, Tetryl and TNT are explosives and require special
handling to be used safely (6). They are not readily available as neat materials.
Stock standard solutions of explosives are frequently provided in acetonitrile. This
is acceptable because the acetonitrile will be sufficiently diluted in calibration
standards and spiking solutions. Store the amber vials at 0° C or less, or according
to the supplier's recommendations.
Stock standards of method analytes that are not explosive may be prepared from neat
materials. Typical concentrations of stock standards that can be used conveniently
are 1-10 mg/mL.
7.9 ANALYTE PRIMARY DILUTION STANDARD SOLUTION(S) -- The stock
standard solutions may be used to prepare a primary dilution standard (PDS)
solution that contains multiple method analytes in ethyl acetate. Aliquots of each of
the stock standard solutions are combined to produce the primary dilution in which
the concentration of the analytes is at least equal to the concentration of the most
concentrated calibration solution, typically, 10 ng/|j,L. Store the primary dilution
standard solution in an amber vial at 0° C or less, and check regularly for signs of
degradation or evaporation, especially just before preparing calibration solutions.
Alternatively, mixtures of method analytes to be used as primary dilution standards
may also be purchased from commercial suppliers. Typically these will be in
acetonitrile. This is acceptable because the acetonitrile will be sufficiently diluted in
calibration standards and spiking solutions. Store the primary dilution standard
solution in an amber vial at 0° C or less, or according to the supplier's
recommendations.
7.10 CALIBRATION SOLUTIONS -- These standards are prepared by combining
appropriate aliquots of a primary dilution standard solution (Sect. 7.9) and the
fortification solution of internal standards and surrogates (Sect. 7.11). All
calibration solutions should contain at least 80% ethyl acetate to avoid gas
chromatographic problems due to mixed solvents. The following calibration ranges
are suggested, but the range maybe modified depending on instrument sensitivity.
For full scan mass spectral analysis, prepare a series of six calibration solutions in
ethyl acetate which contain analytes of interest at suggested concentrations of 10,
5.0, 2.0, 1.0, 0.50, and 0.10 ng/jj,L. For SIM analysis, prepare a series of seven
calibration solutions in ethyl acetate which contain analytes of interest at suggested
concentrations of 2.0, 1.0, 0.50, 0.25, 0.10, 0.050, and 0.025 ng/jaL. Add the
internal standard to each CAL standard at a constant concentration (5-7 ng/p,L is
recommended). Surrogate analytes are also added to each CAL solution, and maybe
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added at a constant concentration or varied concentrations (similar to those for
method analytes), at the discretion of the analyst. If the surrogate analytes are added
at a constant concentration, 1-5 ng/uL is recommended for full scan analysis and
lng/p,L is recommended for SIM analysis. Store these solutions in amber vials at 0°
C or less. Check these solutions regularly for signs of evaporation and/or
degradation. During method development these solutions were stable for at least 60
days.
7.11 INTERNAL STANDARD SOLUTION AND SURROGATE STOCK SOLUTIONS --
7.11.1 INTERNAL STANDARD STO CK - Prepare a stock standard of the internal
standard, 3,4-dinitrotoluene [CAS# 610-39-9], at approximately 2.0 mg/mLin
ethyl acetate. Store at 0° C or less. During method development, this solution
was stable for at least 1 year.
7.11.2. SURROGATE STANDARD STOCK SOLUTIONS - Prepare or purchase
individual stock standards of the following surrogate analytes: 1,3,5-
trimethyl-2-nitrobenzene commonly known as 2-nitromesitylene [CAS#
603-71-4], l,2,4-trimethyl-5-nitrobenzene commonly known as 2-
nitropseudocumene [CAS# 610-91-3], and nitrobenzene-c/5 [CAS# 4165-
60-0]. Standards of 2-nitromesitylene and 2-nitropseudocumene should be
prepared in methanol at concentrations of 1-2 mg/mL. Nitrobenzene-c/5
should be prepared in methylene chloride at a concentration of 1-2 mg/mL.
Store at 0° C or less. During method development, these solutions were
stable for at least 1 year.
7.12 SAMPLE FORTIFICATION SOLUTIONS -
7.12.1 INTERNAL STANDARD FORTIFICATION SOLUTION - Prepare a
dilution of the stock standard in ethyl acetate such that the final
concentration is 0.20 mg/mL. This solution is used to add the internal
standard to each sample, LRB, LFB, and LFM extract. Add 5-25 \iL of this
solution to each 1 mL extract to obtain the recommended final
concentration of 1-5 ng/uL. Check these solutions regularly for signs of
evaporation and/or degradation. During method development these
solutions were stable for at least 60 days.
7.12.2 ANALYTE FORTIFICATION SOLUTION(S) - This solution contains all
method analytes of interest in methanol. These solutions are used to fortify
LFBs and LFMs with method analytes. It is recommended that more than
one concentration of this solution be prepared. During the method
development, two solutions were used. One containing 100 fj,g/mL of each
analyte, was used for higher concentration fortifications, and the other
containing 10 ng/mL of each analyte in methanol was used for lower level
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fortifications. Check these solutions regularly for signs of evaporation
and/or degradation. During method development these solutions were
stable for at least 60 days.
7.12.3 SURROGATE ANALYTE FORTIFICATION SOLUTION - Prepare a
solution of the three surrogate standards listed in Sect. 7.11.2 in methanol.
If a single concentration of each was used to prepare calibration standards,
then prepare this solution such that a convenient fortification volume will
correspond to an equivalent concentration in the extracts. If the surrogates
have been calibrated over a concentration range, add surrogate to the
samples, LRBs, LFBs, LFMs at a single concentration in the range of 1-5
Hg/L. Check these solutions regularly for signs of evaporation and/or
degradation. During method development these solutions were stable for at
least 60 days.
7.13 GC/MS TUNE CHECK SOLUTION ~ Bromofluorobenzene (BFB) [CAS# 460-00-4]
in ethyl acetate; suggested concentration, 5 fj,g/mL. Store this solution in an amber vial
at 0° C or less.
8. SAMPLE COLLECTION, PRESERVATION, AND STORAGE
8.1 SAMPLE COLLECTION ~ When sampling from a water tap, open the tap and
allow the system to flush until the water temperature has stabilized (usually about 2
min). Adjust the flow to about 500 mL/min and collect samples from the flowing
stream. The sample should nearly fill the 1 L or 1 qt bottle, but does not need to be
headspace free. If preservatives have been added to the bottle prior to sampling, be
careful not to rinse them out during sample collection. Keep samples sealed from
collection time until analysis. When sampling from an open body of water, fill the
sample container with water from a representative area. Sampling equipment,
including automatic samplers, must be free of plastic tubing, gaskets, and other parts
that may leach interfering analytes into the water sample.
8.2 SAMPLE DECHLORINATION AND PRESERVATION ~ All samples must be
dechlorinated and preserved with a microbial inhibitor at the time of collection.
Jenkins, et al. (8) have clearly shown the need for microbial preservation of several
of the method analytes. Copper sulfate pentahydrate is added as a microbial
inhibitor at a concentration of 0.5 g/L of water (9). The Trizma Pre-Set Crystals
pH7 buffer (Sect. 7.5) is added at a concentration of 5.0 g/L. The buffer acts to tie
up free chlorine, and also keeps the copper in solution (9). The buffer should be
added to all samples, even those that are unchlorinated. The copper sulfate and
Trizma may be added to the sample bottles as dry salts prior to transporting the
sample bottles to the sampling site.
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8.2.1 As an alternative to the Trizma Pre-Set Crystals pH7 buffer, the following
may be used : 0.3 g of tris(hydroxymethyl)aminomethane plus 4.7 g of
tris(hydroxymethyl)aminomethane hydrochloride for each 1 L sample.
8.3 SAMPLE TRANSPORT AND STORAGE -- All samples should be iced during
shipment and must not exceed 10° C during the first 48 hours. Samples should be
confirmed to be at or below 10° C when they are received at the laboratory. Samples
stored in the lab must be held at or below 6° C until extraction, but should not be
frozen.
Samples that are significantly above 10° C at the time of collection, may need to be
iced or refrigerated for a period of time, in order to chill them prior to shipping. This
will allow them to be shipped with sufficient ice to meet the above requirements.
8.4 HOLDING TIME — Samples must be extracted within 14 days of collection.
Sample extracts may be stored for up to 30 days after sample extraction, when stored
in amber vials at 0° C or less.
9. QUALITY CONTROL
9.1 Quality control (QC) requirements include: the initial demonstration of laboratory
capability (summarized in Table 23) followed by regular analyses of continuing
calibration checks, laboratory performance check standards, laboratory reagent
blanks, laboratory fortified blanks, and laboratory fortified matrix samples. In
addition, a DL must be determined for each analyte of interest. These criteria are
considered the minimum acceptable QC criteria, and laboratories are encouraged to
institute additional QC practices to meet their specific needs. The laboratory must
maintain records to document the quality of the data generated. A complete
summary of QC requirements is summarized in Table 24.
9.2 INITIAL DEMONSTRATION OF CAPABILITY (IDC) -- Requirements for the
initial demonstration of laboratory capability are described in the following sections
and summarized in Table 23.
9.2.1 INITIAL DEMONSTRATION OF LOW CARTRIDGE/DISK
EXTRACTION BACKGROUND AND SYSTEM BACKGROUND --
Before any samples are analyzed, or any time a new lot or brand of solid
phase extraction disks or cartridges is received from a supplier, it must be
demonstrated that a laboratory reagent blank (LRB) is reasonably free of
any contamination that would prevent the determination of any analyte of
concern (Sect. 9.2.1.2 and 9.4).
9.2.1.1 A source of potential contamination is the solid phase extraction
media which may contain phthalate esters, silicon compounds,
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and other contaminants that could interfere with the
determination of method analytes. Although extraction media
are generally made of inert materials, they may still contain
extractable organic material. If the background contamination is
sufficient to prevent accurate and precise measurements, the
condition must be corrected before proceeding with the initial
demonstration.
9.2.1.2 Other sources of background contamination are solvents,
reagents, and glassware. Background contamination must be
reduced to an acceptable level before proceeding with the next
section. Background from method analytes and interferences
should be < 1/3 the MRL.
9.2.2 INITIAL DEMONSTRATION OF PRECISION (IDP) -- Prepare 4-7
replicate LFBs fortified at 2-5 \igfL, or other mid-range concentration, using
solutions described in Sect. 7.12.2 and 7.12.3. This concentration maybe
lower if SIM is being used. Sample preservatives described in Sect. 8.2
must be added to these samples. Extract and analyze these replicates
according to the procedure described in Sect. 11. The relative standard
deviation (RSD) of the results of the replicate analyses must be less than or
equal to 20% for all method analytes and surrogates.
9.2.3 INITIAL DEMONSTRATION OF ACCURACY (IDA) - Using the same
set of replicate data generated for Sect. 9.2.2, calculate average recovery.
The average recovery of the replicate values for all analytes and surrogates
must be within 70-130% of the true value.
9.2.4 DL DETERMINATION - Replicate analyses for this procedure should be
done over at least 3 days (both the sample extraction and the GC analyses
should be done over at least 3 days). Prepare at least 7 replicate LFBs using
solutions described in Sect. 7.12.2 and 7.12.3, at a concentration estimated
to be near the DL. This concentration may be estimated by selecting a
concentration at 2-5 times the noise level. Concentrations shown in the
example data in Tables 3, 9, and 15 may be used as a guide, however the
appropriate concentration will be dependent upon the injection technique
and the sensitivity of the GC/MS system used. Sample preservatives
described in Sect. 8.2 must be added to these samples. Analyze the seven
replicates through all steps of Sect. 11. Calculate the DL using the
following equation:
DL St^.^ j_ alpha = 0 9
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where:
t( n -11 - alpha = o 99) = Student's t value for the 99% confidence level with
n-1 degrees of freedom
n = number of replicates
S = standard deviation of replicate analyses.
NOTE: Do not subtract blank values when performing DL calculations.
9.2.5 The analyst is permitted to modify GC columns or GC conditions. Each
time such method modifications are made, the analyst must repeat the
procedures in Sect. 9.2.1 through 9.2.4.
9.3 MINIMUM REPORTING LEVEL (MRL) - The MRL is the threshold
concentration of an analyte that a laboratory can expect to accurately quantitate in an
unknown sample. The MRL should be established at an analyte concentration either
greater than three times the DL or at a concentration which would yield a response
greater than a signal-to-noise ratio of five. Although the lowest calibration
standard for an analyte may be below the MRL, the MRL for an analyte must
never be established at a concentration lower than the lowest calibration
standard for that analyte.
9.4 LABORATORY REAGENT BLANKS (LRB) - With each extraction batch,
analyze a laboratory reagent blank to determine the background system
contamination. If, within the retention time window of any analyte, the LRB
produces a peak that would prevent the determination of that analyte, determine the
source of contamination and eliminate the interference before processing samples.
Background contamination must be reduced to an acceptable level before
proceeding. Background from method analytes or contaminants that interfere with
the measurement of method analyses should be < 1/3 the MRL. Any time a new
batch of SPE cartridges is received, or new supplies of other reagents are used,
repeat the demonstration of low background described in Sect. 9.2.1.
9.5 CONTINUING CALIBRATION CHECK (CCC) - This calibration check is
required at the beginning of each day that samples are analyzed, after every ten field
samples, and at the end of any group of sample analyses. See Sect. 10.3 for
concentration requirements and acceptance criteria.
9.6 MS TUNE CHECK ~ This performance check consists of verifying the MS tune
using the mass spectrum of BFB. A complete description of the check is in Sect.
10.2.1. This check must be performed each time a major change is made to the mass
spectrometer, and each time analyte calibration is performed (i.e., average RFs are
calculated, or first or second order calibration curves are developed).
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9.7 LABORATORY FORTIFIED BLANK (LFB) -- With each extraction batch, extract
and analyze an LFB containing each analyte of concern. If more than 20 field
samples are included in a batch, analyze an LFB for every 20 samples. The fortified
concentration of the LFB should be rotated between low, medium, and high
concentrations from day to day. The low concentration LFB must be as near as
practical to the MRL. Results of LFB analyses corresponding to the low fortification
concentration for an analyte must be within 50-150% of the true value. Results of
LFB analysis from medium and high level concentrations must be 70-130% of the
true value for all analytes.
9.8 INTERNAL STANDARD (IS) -The analyst must monitor the peak area of the IS in
all injections during each analysis day. The IS response (peak area) in any
chromatographic run should not deviate from the response in the most recent CCC
by more than 30%, and must not deviate by more than 50% from the area measured
during initial analyte calibration. If the IS area in a chromatographic run does not
meet these criteria inject a second aliquot of that extract.
9.8.1 If the reinjected aliquot produces an acceptable internal standard response,
report results for that aliquot.
9.8.2 If the reinjected extract fails again, the analyst should check the calibration
by reanalyzing the most recently acceptable calibration standard. If the
calibration standard fails the criteria of Sect. 10.3.3, recalibration is in order
per Sect. 10.2. If the calibration standard is acceptable, extraction of the
sample may need to be repeated provided the sample is still within the
holding time. Otherwise, report results obtained from the reinjected extract,
but annotate as suspect. Alternatively, collect a new sample and re-analyze.
9.9 SURROGATE RECOVERY - The surrogate standards are fortified into all
calibration standards, samples, LFBs, LFMs, FDs, FRBs and LRBs. The surrogate is
a means of assessing method performance from extraction to final chromatographic
measurement.
9.9.1 Surrogate recovery criteria are 70-130% of the fortified amount for all
method surrogates. When surrogate recovery from a sample, blank, or CCC
does not meet these criteria, check : (1) calculations to locate possible
errors, (2) standard solutions for degradation, (3) contamination, and (4)
instrument performance. Correct any problems that are identified. If these
steps do not reveal the cause of the problem, reanalyze the extract.
9.9.2 If the extract reanalysis meets the surrogate recovery criterion, report only
data for the reanalyzed extract.
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9.9.3 If the extract reanalysis fails the recovery criterion, the analyst should check
the calibration by reanalyzing the most recently acceptable calibration
standard. If the calibration standard fails the criteria of Sect. 10.3.3,
recalibration is in order per Sect. 10.2. If the calibration standard is
acceptable, it may be necessary to extract another aliquot of sample if
sample holding time has not been exceeded. If the sample re-extract also
fails the recovery criterion, report all data for that sample as suspect, or
analyze a duplicate sample.
9.10 LABORATORY FORTIFIED SAMPLE MATRIX (LFM) -- Determine that the
sample matrix does not contain materials that adversely affect method performance.
This is accomplished by analyzing replicates of laboratory fortified matrix samples
and ascertaining that the precision, accuracy, and method detection limits of analytes
are in the same range as obtained with laboratory fortified blanks. If a variety of
different sample matrices are analyzed regularly, for example, drinking water from
groundwater and surface water sources, LFM data should be collected for each
matrix. Over time, LFM data should be documented for all routine sample sources
for the laboratory. A laboratory fortified sample matrix should be extracted and
analyzed for each extraction batch. If more than 20 samples are processed in a
batch, extract and analyze an LFM for every 20 samples. If the recovery data for an
LFM does not meet the recovery criteria in Sect. 9.7, and LFBs show the laboratory
to be in control, then the samples from that matrix (sample location) are documented
as suspect due to matrix effects.
9.10.1 Within each extraction batch, a minimum of one field sample is fortified as
an LFM for every 20 samples analyzed. The LFM is prepared by spiking a
sample with an appropriate amount of the fortification solutions described
in Sect. 7.12.2 and 7.12.3. Select the spiking concentration that is at least
twice the matrix background concentration. Selecting a duplicate bottle of a
sample that has already been analyzed, aids in the selection of appropriate
spiking levels. If this is not possible, use historical data or rotate through
low, medium and high calibration concentrations to select a fortifying
concentration.
9.10.2 Calculate the percent recovery (R) for each analyte, after correcting the
measured fortified sample concentration, A, for the background
concentration, B, measured in the unfortified sample, i.e.,
C
where C is the fortified concentration. Compare these values to
control limits for LFBs (Sect. 9.7).
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9.10.3 Recoveries may exhibit a matrix dependence. For samples fortified at or
above their native concentration, recoveries should range between 70 and
130% for all method analytes. If the accuracy (percent recovery) of any
analyte falls outside the designated range, and the laboratory performance
for that analyte is shown to be in control, the accuracy problem encountered
with the fortified sample is judged to be matrix related, not system related.
The result for that analyte in the unfortified sample is labeled
suspect/matrix to inform the data user that the results are suspect due to
matrix effects.
9. 1 1 FIELD DUPLICATES (FD) -- Within each extraction batch, a minimum of one field
sample should be analyzed in duplicate. If more than 20 samples are extracted in a
batch, analyze a FD for each 20 samples. Duplicate sample analyses serve as a
check on sampling and laboratory precision. If analytes are not routinely observed in
field samples, duplicate LFMs should be analyzed to substitute for this requirement.
Refer to Sect. 9.10.1 for guidance on spiking concentrations.
9.1 1.1 Calculate the relative percent difference (RPD) for duplicate measurements
(FD1 and FD2) as shown below.
RPD- FD1~FD2 .(100)
9. 1 1 .2 Relative percent differences for laboratory duplicates and duplicate LFMs
should fall in the range of ± 30 %.
NOTE: Greater variability may be observed for target analytes with
concentrations at the low end of the calibration range.
9.12 QUALITY CONTROL SAMPLE (QCS) -- Each time that new PDSs are prepared,
analyze a QCS from an external source. If standards are prepared infrequently,
analyze a QCS at least quarterly. The QCS may be injected as a calibration standard,
or fortified into reagent water and analyzed as an LFB. If the QCS is analyzed as a
calibration check standard, then the acceptance criteria are the same as for the CCC
(Sect. 10.3.3). If the QCS is analyzed as an LFB, then the acceptance criteria are the
same as for an LFB (Sect. 9.7). If measured analyte concentrations are not of
acceptable accuracy, check the entire analytical procedure to locate and correct the
problem.
10. CALIBRATION AND STANDARDIZATION
10.1 Demonstration and documentation of acceptable mass spectrometer tune and initial
calibration is required before any samples are analyzed. After initial calibration is
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successful, a continuing calibration check is required at the beginning and end of
each period in which analyses are performed, and after every tenth sample.
Verification of mass spectrometer tune must be repeated each time a major
instrument modification or maintenance is performed, and prior to analyte
recalibration.
10.2 INITIAL CALIBRATION
10.2.1 MS TUNE ~ Calibrate the mass and abundance scales of the MS with
calibration compounds and procedures prescribed by the manufacturer with
any modifications necessary to meet tuning requirements. Inject 5 ng or less
of BFB into the GC/MS system. Acquire a mass spectrum that includes
data for m/z 45-250. If the BFB mass spectrum does not meet all criteria in
Table 1, the MS must be retuned and adjusted to meet all criteria before
proceeding with calibration. A single spectrum at the apex of the
chromatographic peak, or an average of the three spectra at the apex of the
peak, or an average spectrum across the entire GC peak may be used to
evaluate the performance of the system. Background subtraction is
permitted.
NOTE: Although BFB is most commonly used as the tune check for
volatile compound analysis, it was selected for use in this method for
semivolatile analytes because the ions used for the BFB tune check are
similar to the analyte quantitation ions in this method.
10.2.2 ANALYTE CALIBRATION FOR FULL SCAN MASS
SPECTROMETRY -- Inject an aliquot, usually 1-2 uL (unless a large
volume injector is used), of a medium to high concentration calibration
solution. For example, 2-10 |j,g/mL, and acquire and store data from m/z
45-250 with a total cycle time (including scan overhead time) of 0.7 sec or
less. Cycle time must be adjusted to measure at least five or more scans
during the elution of each GC peak. Seven to ten scans across each GC peak
are recommended.
NOTE: On some instruments that cannot scan quickly, e.g. ion traps, the
scan time and the chromatographic peak width may need to be carefully
evaluated to make sure that this requirement is met.
Chromatographic conditions used during method development are outlined
below. These conditions were found to work well on the instrumentation
used. Since some of the method analytes are vulnerable to active sites and
thermal decomposition, optimum chromatographic conditions may vary
with individual instrument design. Although the following conditions are
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recommended, GC conditions may be modified, if all performance criteria
in Sections 9 and 10 are met.
10.2.2.1 PTV Splitless Injection, Full Scan Mass Spectrometry -
Inject with the injector temperature at 60° C, program the
injector at 200° C per min to 250° C. Hold in the Splitless mode
for 1.0 min. Use an initial GC oven temperature of 45° C for 3.3
minutes. Program the oven temperature at 7° C per min to 210°
C. Then program at 20° C per min to 250° C. An example
chromatogram obtained under these conditions is shown in
Figure 1.
10.2.2.2 Cold On-Column Injection, Full Scan Mass Spectrometry-
Inject with the injector temperature at 50° C, program the
injector at 150° C per min to 220° C. Hold in the Splitless mode
for 2.0 min. Use an initial GC oven temperature of 50° C for 3
minutes. Program the oven temperature at 7° C per min to 210°
C. Then program at 20° C per min to 250° C.
10.2.3 Performance criteria for the calibration standards. Examine the stored
GC/MS data with the data system software. The GC/MS/DS peak
identification software should be able to recognize a GC peak in the
appropriate retention time window for each of the compounds in the
calibration solution, and make correct identifications (Sect. 11.8, 11.9).
10.2.4 If all performance criteria are met, inject an appropriate volume (usually 1-2
|j,L unless a large volume injector is used) of each of the other CAL
solutions using the same GC/MS conditions.
10.2.4.1 Some GC/MS systems may not be sensitive enough to detect
some of the analytes in the two lowest concentration CAL
solutions. If this is the case, it is acceptable to calibrate using
the remaining higher concentration points, as long as a minimum
of 5 calibration points are used to generate the calibration curve
or average response factor (RF) for each analyte. In addition,
some GC/MS systems might reach signal saturation at the
highest calibration concentration. If this is the case, it is
acceptable to drop the highest point and calibrate on the
remaining points, as long as at least 5 calibration concentrations
are used to generate the calibration curve or average RF for each
analyte. Points in the middle of the calibration range may not be
dropped. Use a minimum of 3 standards for a calibration range
of 1 order of magnitude, and at least 5 standards for 2 orders of
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magnitude. Data outside of the established calibration range
should never be reported.
10.2.5 Concentrations maybe calculated through the use of average response
factor (RF) or through the use of a calibration curve. Average RF
calibrations may only be used if the RF values over the calibration range are
relatively constant (<30% RSD).
Average RF is determined by calculating the mean RF of each calibration
point.
„„ _
(
where:
Ax = integrated abundance (peak area) of the quantitation ion
of the analyte.
Ais = integrated abundance (peak area) of the quantitation ion
internal standard.
Qx = quantity of analyte injected in ng or concentration units.
Qis = quantity of internal standard injected in ng or
concentration units.
10.2.6 As an alternative to calculating average RFs and applying the RSD test, use
the GC/MS data system software to generate a linear regression or quadratic
calibration curve. The analyst may choose whether or not to force zero, to
obtain a curve that best fits the data. Examples of common GC/MS system
calibration curve options are: 1) Ax /Ais vs Qx /Qis and 2) RF vs Ax /Ais.
10.2.7 Acceptance criteria for the calibration of each analyte is determined by
calculating the concentration of each analyte and surrogate in each of the
analyses used to generate the calibration curve or average RF. Each
calibration standard, except the lowest point, for each analyte must calculate
to be 70-130% of its true value. The lowest point must calculate to be 50-
150% of its true value. If this criteria cannot be met, reanalyze the
calibration standards, or select an alternate method of calibration. The data
in this method were generated using both linear and quadratic fits,
depending upon the analyte. Quadratic fit calibrations should be used with
caution, because the non-linear area of the curve may not be reproducible.
1 0.2.8 ANALYTE CALIBRATION FOR SIM MASS SPECTROMETRY ~~
Collect data for a mid to high level standard in the full scan mode as
directed in Sect. 10.2.2 above.
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Chromatographic conditions used during method development are outlined
below. These conditions were found to work well on the instrumentation
used. Since some of the method analytes are vulnerable to active sites and
thermal decomposition, optimum chromatographic conditions may vary
with individual instrument design. Although the following conditions are
recommended, GC conditions may be modified, if all performance criteria
in Sections 9 and 10 are met.
10.2.8.1 PTV Splitless Injection, SIM Mass Spectrometry -
Inject with the injector temperature at 60° C, program the
injector at 150° C per min to 220° C. Hold in the Splitless mode
for 2.0 min. Use an initial GC oven temperature of 45° C for 3
minutes. Program the oven temperature at 7° C per min to 210°
C. Then program at 20° C per min to 250° C.
10.2.9 Divide the chromatogram into retention time windows, each of which
contains one or more chromatographic peaks. During SIM analysis a small
number of selected masses corresponding to dominant ions for the analytes
in that window will be scanned within each window. The number of
windows used and the selection of the retention times for each window will
depend upon the type of instrumentation used. Highest sensitivity can be
expected when a minimum number of ions are scanned in a window, i.e.
one analyte per window. However, if analytes are closely eluting, it may be
necessary to include more than one analyte per window.
CAUTION: When acquiring SIM data, GC operating conditions must be
carefully reproduced for each analysis to provide reproducible retention
times. If this is not done, the correct ions will not be monitored at the
appropriate times. As a precautionary measure, the chromatographic peaks
in each window must not elute too close to the edge of the time window.
As a minimum, there should be at least 5 sec between the edge of the time
window and the beginning or end of an analyte peak.
10.2.10 Using the GC/MS software, create an acquisition method such that 2 or 3
identifying ions are scanned for each chromatographic peak. Three ions are
recommended, but in some cases, only 2 ions may be used if only 2
abundant ions are present in the mass spectrum, e.g. RDX. Suggested
quantitation ions and secondary confirmation ions are listed in Table 2. Re-
analyze the calibration standard using the SIM acquisition method.
Continue with calibration as described in Sect. 10.2.3 through 10.2.7.
10.3 CONTINUING CALIBRATION CHECK (CCC) -- The minimum daily calibration
verification is as follows. Verify the initial calibration at the beginning and end of
each group of analyses, and after every tenth sample during analyses. (In this
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context, a "sample" is considered to be a field sample. LRBs, LFMs, LFBs and
CCCs are not counted as samples.) The beginning CCC each day should be at or
near the MRL in order to verify instrument sensitivity prior to any analyses. If
standards have been prepared such that all low CAL points are not in the same CAL
solution, it may be necessary to analyze two CAL solutions to meet this requirement.
Alternatively, it may be cost effective to obtain a customized standard to meet this
criteria. Subsequent CCCs can alternate between a medium and high concentration
standard.
10.3.1 Inject an aliquot of the appropriate concentration calibration solution and
analyze with the same conditions used during the initial calibration.
10.3.2 Determine that the absolute areas of the quantitation ions of the internal
standard have not changed by more than 30% from the areas measured in
the most recent continuing calibration check, or by more than 50% from the
areas measured during initial calibration. If this area has changed by more
than these amounts, adjustments must be made to restore system sensitivity.
These adjustments may include cleaning of the MS ion source, or other
maintenance as indicated in Sect. 10.3.4. Major instrument maintenance
requires recalibration. Control charts are useful aids in documenting system
sensitivity changes.
10.3.3 Calculate the concentration of each analyte and surrogate in the check
standard. The calculated amount for each analyte for medium and high level
CCCs must be within 70-130% of the true value. The calculated amount
for the lowest calibration point for each analyte must be within 50-150% of
the true value. If these conditions do not exist, remedial action should be
taken which may require recalibration. Any field sample extracts that have
been analyzed since the last acceptable calibration verification should be
reanalyzed after adequate calibration has been restored, with the following
exception. If the continuing calibration check in the middle or at the
end of an analysis batch fails because the calculated concentration is
>130% of the true value, and field sample extracts showed no detection
of method analytes, non-detects may be reported without re-analysis.
10.3.4 Some possible remedial actions are listed below. This list is not meant to
be all inclusive. Major maintenance such as cleaning an ion source,
cleaning quadrupole rods, replacing filament assemblies, replacing the
electron multiplier, changing the GC column, etc. require returning to the
initial calibration step (Sect. 10.2).
10.3.4.1 Check and adjust GC and/or MS operating conditions; check the
MS resolution, and calibrate the mass scale.
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10.3.4.2 Replace the GC injection port liner; cleaning and deactivating
used liners is not recommended.
10.3.4.3 Flush the GC column with solvent according to manufacturer's
instructions.
10.3.4.4 Break off a short portion (about 1 meter) of the column from the
end near the injector, or replace GC column. This action will
cause a change in retention times. A change in retention times
may require a change in the SIM scan time windows.
10.3.4.5 Prepare fresh CAL solutions, and repeat the initial calibration
step.
10.3.4.6 Clean the MS ion source and rods (if a quadrupole).
10.3.4.7 Replace any components that allow analytes to come into
contact with hot metal surfaces.
10.3.4.8 Replace the MS electron multiplier, or any other faulty
components.
11. PROCEDURE
11.1 SOLE) PHASE EXTRACTION
11.1.1 This procedure may be performed manually or in an automated mode (Sect.
6.11) using a robotic or automatic sample preparation device. If an
automatic system is used to prepare samples, follow the manufacturer's
operating instructions, but all extraction and elution steps must be the same
as in the manual procedure. Extraction and/or elution steps may not be
changed or omitted to accommodate the use of an automated system.
11.1.2 Mark the level of the sample on the outside of the sample bottle for later
sample volume determination (Sect. 11.5). Verify that the sample is
between pH 5 and 7, and is free of residual chlorine. If the sample is an
LRB or LFB, add sample preservatives specified in Sect. 8.2. Add an
aliquot of the surrogate fortification solution(s), and mix immediately until
homogeneous. The resulting concentration of these compounds in the water
should be l-5|j,g/L. Generally, SIM analyses should be performed with
surrogate concentrations at the low end of this range. If the sample is an
LFB or LFM, add the desired amount of analyte fortification solution.
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11.2. CARTRIDGE EXTRACTION
11.2.1. CARTRIDGE CLEAN-UP AND CONDITIONING -- Rinse each cartridge
with three, 5 mL aliquots of ethyl acetate. Let the cartridge drain dry after
each flush. Then rinse the cartridge with three, 5 mL aliquots of methanol,
but DO NOT allow the methanol to elute below the top of the cartridge
packing. From this point, do not allow the cartridge packing to go dry.
Rinse the cartridge with two, 10 mL aliquots of reagent water, stopping the
flow when there is approximately 4 cm of reagent water in the column.
Attach the transfer tube, and turn on the vacuum, and begin adding sample
to the cartridge.
11.2.2. Adjust the vacuum so that the approximate flow rate is 10-15 mL/min.
After all of the sample has passed through the SPE cartridge, draw air or
nitrogen through the cartridge for 10-15 min at high vacuum (10-15 in. Hg).
The drying time may vary, depending upon the strength of the vacuum
source, and the number of cartridges being processed simultaneously. Do
not dry the cartridge for more than 10-15 min. Additional time is not likely
to increase dryness, and excess drying times have been shown to decrease
recoveries of nitrobenzene (10).
NOTE: Samples with a high level of hardness and/or high TOC may exhibit
a lower flow rate than "cleaner" samples at the same vacuum setting. This
may be due to partial plugging of the solid phase. Fortified sample matrices
of these types showed no loss of method performance.
11.2.3. Place a collection tube in the vacuum chamber underneath each extraction
cartridge. Rinse the inside of each sample bottle with 5 mL ethyl acetate
and use vacuum to pull the rinse solvent through the transfer tube and
through the cartridge, collecting the solvent in a collection tube. Remove
the transfer tubing from the top of the cartridge. Add another 5 mL of ethyl
acetate to the top of the cartridge with a disposable pipette. Pull this solvent
through the cartridge at low vacuum, such that the solvent exits the
cartridge in a dropwise fashion. Combine this second eluate with the first
one. Dry and concentrate the combined eluate as directed in Sect. 11.4.
11.3. DISK EXTRACTION
11.3.1 DISK CLEAN-UP AND CONDITIONING - Rinse each disk with three, 5
mL aliquots of ethyl acetate. Let the disk dry after each rinse. Then rinse
the disk with three, 5 mL aliquots of methanol, but DO NOT allow the
methanol to elute below the top of the disk. From this point, do not allow
the disk to go dry. Rinse the disk with two, 10 mL aliquots of reagent
water, stopping the flow when there is approximately 3-4mm of reagent
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water on top of the disk. Turn on the vacuum, and begin adding sample to
the disk.
11.3.2. The sample can be passed through the disk at high vacuum (10-15 in. Hg).
After the entire sample has been extracted, dry the disk at full vacuum for 5-
10 min. Do not dry the disk for more than 10 min. Additional time is not
likely to increase dryness, and excess drying times have been shown to
decrease recoveries of nitrobenzene (10).
11.3.3. Place a collection tube in the vacuum manifold beneath the filtration
glassware setup for each extraction disk. Rinse the inside of each sample
bottle with 5 mL ethyl acetate and use this rinse solvent to elute each disk,
collecting the solvent in a collection tube. Add another 5 mL of ethyl
acetate to the top of the disk with a disposable pipette, using the solvent to
wash down the sides of the filtration glassware. Pull this solvent through
the disk. Add another 3 mL of ethyl acetate to the top of the disk with a
disposable pipette, using the solvent to wash down the sides of the filtration
glassware. Pull this solvent through the disk. Dry and concentrate the
combined eluates as directed in Sect. 11.4.
11.4. Small amounts of residual water from the sample container and the SPE cartridge
may form an immiscible layer with the eluate. To eliminate residual water, pass the
eluate through the drying column (Sect. 6.7). The drying column is packed with
approximately 5 to 7 grams of anhydrous sodium sulfate, and is pre-wetted with a
small volume of ethyl acetate prior to passing the eluate through it. Collect the dried
eluate in a clean collection tube. After passing the eluate through the drying tube,
wash the sodium sulfate with at least 3 mL ethyl acetate and collect it in the same
tube. Concentrate the extract to approximately 0.9 mL in a warm (40° C) water bath
under a gentle stream of nitrogen. Do not concentrate the extract to less than 0.5
mL, as this will result in loss of analytes. Add the internal standard (Sect. 7.12.1).
Adjust final volume to 1 mL. Make any volume adjustments with ethyl acetate.
11.5 Fill the sample bottle to the volume mark noted in Sect. 11.1.2. with tap water.
Transfer the tap water to a 1000 mL graduated cylinder, and measure the sample
volume to the nearest 10 mL. Record this volume for later analyte concentration
calculations. As an alternative to this process, the sample volume maybe
determined by the difference in weight between the full bottle (before extraction)
and the empty bottle (after extraction). Assume a sample density of 1.0 g/mL.
11.6 Analyze an aliquot of the sample extract with the GC/MS system under the same
conditions used for the initial and continuing calibrations.
11.7 At the conclusion of data acquisition, use the same software that was used in the
calibration procedure to identify peaks in predetermined retention time windows of
529-27
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interest. Use the data system software to examine the ion abundances of
components of the chromatogram.
11.8 IDENTIFICATION OF ANALYTES FOR FULL SCAN MASS SPECTROMETRY --
Identify a sample component by comparison of its mass spectrum (after background
subtraction) to a reference spectrum in the user-created data base. The GC retention
time of the sample component should be within 1-2 sec of the retention time observed
for that same compound in the most recently analyzed continuing calibration check
standard. Ideally, the width of the retention time window should be based upon
measurements of actual retention time variations of standards over the course of a day.
Three times the standard deviation of a retention time can be used to calculate a
suggested window size for a compound. However, the experience of the analyst should
weigh heavily in the interpretation of the chromatogram.
11.8.1 In general, all ions that are present above 10% relative abundance in the mass
spectrum of the standard should be present in the mass spectrum of the sample
component and should agree within absolute 20%. For example, if an ion has
a relative abundance of 30% in the standard spectrum, its abundance in the
sample spectrum should be in the range of 10 to 50%.
11.9 IDENTIFICATION OF ANALYTES FOR SIM MASS SPECTROMETRY --
Identify a sample component by comparison of its selected ions to a reference
spectrum in the user-created data base. The GC retention time of the sample
component should be within 1-2 sec of the retention time observed for that same
compound in the most recently analyzed continuing calibration check standard.
Ideally, the width of the retention time window should be based upon measurements
of actual retention time variations of standards over the course of a day. Three times
the standard deviation of a retention time can be used to calculate a suggested
identification window size for a compound. However, the experience of the analyst
should weigh heavily in the interpretation of the chromatogram.
NOTE: Because less spectral information is being acquired for analyte identification
when SIM is used, retention times of the analyte peaks are an even more important
factor in analyte identification than in full scan data analysis.
11.9.1 The apex of the chromatographic peak for the quantiation ion and
confirmation ions for each analyte must be within ± 1 scan of one another.
11.9.2 The integrated ion current for each quantitation and confirmation ion must
be at least three times background noise, and must not have saturated the
detector.
11.9.3 All ions monitored in the standard should be present in the SIM mass
spectrum of the sample component and should agree within absolute 20%.
529-28
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For example, if an ion has a relative abundance of 30% in the standard
spectrum, its abundance in the sample spectrum should be in the range of 10
to 50%.
12. DATA ANALYSIS AND CALCULATIONS
12.1 Complete chromatographic resolution is not necessary for accurate and precise
measurements of analyte concentrations if unique ions with adequate intensities are
available for quantitation. Identification is hampered when sample components are
not resolved chromatographically and produce mass spectra containing the same ions
contributed by more than one analyte. When GC peaks obviously represent more
than one sample component (i.e., broadened peak with shoulder(s) or valley between
two or more maxima), appropriate analyte spectra and background spectra can be
selected by examining plots of characteristic ions for each tentatively identified
component. When analytes coelute (i.e., only one GC peak is apparent), the
identification criteria can be met but each analyte spectrum will contain extraneous
ions contributed by the coeluting compound. In validating this method,
concentrations were calculated by measuring the quantitation ions listed in Table 2.
Other ions may be selected at the discretion of the analyst. If the response of any
analyte exceeds the calibration range established in Sect. 10, dilute the extract, add
additional internal standard, and reanalyze. The resulting data should be
documented as a dilution, with an increased MRL.
12.1.1 Calculate analyte and surrogate concentrations, using the multipoint
calibration established in Sect. 10. Do not use daily calibration verification
data to quantitate analytes in samples. Adjust final analyte concentrations
to reflect the actual sample volume determined in Sect. 11.5.
12.1.2 Calculations should utilize all available digits of precision, but final
reported concentrations should be rounded to an appropriate number of
significant figures (one digit of uncertainty).
13. METHOD PERFORMANCE
13.1 PRECISION, ACCURACY AND DLs- Single laboratory accuracy and precision
data from both fortified reagent water and fortified matrices are presented in Tables
3-20. Detection limits (DLs) are presented in Tables 3, 9 and 15 for the three
techniques demonstrated.
13.1.1 Data presented in Tables 3-8 and 21-22 were obtained on a Varian Saturn 4
GC/MS (ion trap) system, using PTV splitless injection and full scan mass
spectrometry. Gas chromatographic conditions are given in Sect. 10.2.2.1.
529-29
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NOTE: Selected ion storage was evaluated on this instrument. Sensitivity
was not significantly improved relative to full scan monitoring.
13.1.2 Data presented in Tables 9-14 were obtained on a Schimadzu QP5050A
GC/MS (quadrupole) system, using PTV splitless injection and SIM mass
spectrometry. A high pressure injection (14.5 psi) was employed on this
instrument. Other gas chromatographic conditions are given in Sect.
10.2.8.1.
13.1.3 Data presented in Tables 15-20 were obtained on a Varian Saturn 4 GC/MS
(ion trap) system, using cold on-column injection and full scan mass
spectrometry. Gas chromatographic conditions are given in Sect. 10.2.2.2.
13.2 EVALUATION OF ADDITIONAL EXPLOSIVES - Nitroglycerin (NG), octahdro-
l,3,5,7-tetranitro-l,3,5,7-tetrazocine (HMX ) and pentaerythritoltetra-nitrate (PETN)
were evaluated for inclusion into this method. HMX was never observed in any of
the chromatograms. Precision and accuracy for NG and PETN were not sufficient to
reliably meet the QC criteria in the method. The method may be suitable for
qualitative identification of these two compounds.
13.3 Data were not obtained for 2,4-dinitrotoluene with the on-column technique. The
mass spectrum of this compound was observed to change with concentration. At
low concentrations, an unusual spectrum was observed. It was determined that this
resulted from reactions of 2,4-dinitrotoluene with residual solvent in the ion trap.
This is more likely to happen with on-column techniques than splitless techniques
because more solvent is transferred to the column. Although the on-column
technique was not demonstrated with a quadrupole instrument, it is suspected that
this observation was an ion trap phenomenon.
13.4 Data were not obtained for 3,5-dinitroanaline with the on-column technique.
Because a commercial standard mixture was used, where the concentration of this
analyte was five times lower than the other analytes, many of the demonstration
samples contained it at a concentration too low to be accurately quantified.
13.5 Some matrix effects may be observed. Some method analytes extracted from some
water matrices may exhibit "matrix induced chromatographic response
enhancement" (11-13). This phenomenon has been widely reported for the analysis
of thermally labile pesticides in water matrices, and maybe applicable to some of
these method analytes as well. Compounds that exhibit this phenomenon may give
analytical results that exceed 100% recovery in fortified matrix samples, especially
at low concentrations. It has been proposed that thermally labile compounds are
susceptible to decomposition in the GC injection port and/or while in the GC
column. Co-extracted matrix components help shield and protect these compounds
through the GC system, thus resulting in a higher response than when clean
529-30
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standards are analyzed. This effect can usually be minimized by reducing the
residence time of method analytes in the GC system by using a relatively high linear
velocity and a short column. The use of clean, deactivated injection port liners is
also advised.
13,6 ANALYTE STABILITY STUDIES--
13.6.1 SAMPLES - Chlorinated surface water samples, fortified with method
analytes at 5.0 \igfL, were preserved and stored as required in Sect. 8. The
average of replicate analyses (n=5) conducted on days 0, 7, 11, 14, and 19
are presented in Table 21. These data document the 14 day holding time.
13.6.2 EXTRACTS - Extracts from the day 0 holding time study described above
were stored at 0 °C, and analyzed in replicate (n=5) on days 0, 17, 15, 21,
28, and 35. The results of these analyses are presented in Table 22, and
document the 30 day holding time.
14. POLLUTION PREVENTION
14.1 This method utilizes SPE technology to remove the analytes from water. It requires
the use of very small volumes of organic solvent and very small quantities of pure
analytes, thereby minimizing the potential hazards to both the analyst and the
environment when compared with the use of large volumes of organic solvents in
conventional liquid-liquid extractions.
14.2 For information about pollution prevention that may be applicable to laboratory
operations, consult "Less Is Better: Laboratory Chemical Management for Waste
Reduction" available from the American Chemical Society's Department of
Government Relations and Science Policy, 1155 16th Street N.W., Washington,
D.C., 20036, or on-line at http://membership.acs.org/c/ccs/pub_9.htm.
15. WASTE MANAGEMENT
15.1 The analytical procedures described in this method generate relatively small amounts
of waste since only small amounts of reagents and solvents are used. The matrices
of concern are finished drinking water or source water. However, the Agency
requires that laboratory waste management practices be conducted consistent with all
applicable rules and regulations, and that laboratories protect the air, water, and land
by minimizing and controlling all releases from fume hoods and bench operations.
Also, compliance is required with any sewage discharge permits and regulations,
particularly the hazardous waste identification rules and land disposal restrictions.
For further information on waste management, consult "The Waste Management
Manual for Laboratory Personnel" available on-line from the American Chemical
529-31
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Society at http://chemistry.org/portal/Chemistry?PID=acsdisplay.html&DOC=
govemment%5Cpublications%5Ceip Jiazwaste.html.
16. REFERENCES
1. Glaser, J.A., D.L. Foerst, G.D. McKee, S.A. Quave, and W.L. Budde, "Trace Analyses for
Wastewaters," Environ. Sci. Technol.. 15 (1981) 1426-1435.
2. "Carcinogens - Working With Carcinogens," Department of Health, Education, and
Welfare, Public Health Service, Center for Disease Control, National Institute for
Occupational Safety and Health, Publication No. 77-206, Aug. 1977.
3. "OSHA Safety and Health Standards, General Industry," (29CFR1910), Occupational
Safety and Health Administration, OSHA, (Revised, July 1, 2001).
4. "Safety in Academic Chemistry Laboratories," American Chemical Society Publication,
Committee on Chemical Safety, 6th Edition, available from the ACS Office of Society
Services by e-mail at OSS@acs.org.
5. "Drinking Water Standards and Health Advisories," United States Environmental
Protection Agency, Office of Water, EPA 822-B-00-001, 2000.
6. "Method 8095, Explosives by Gas Chromatography," United States Environmental
Protection Agency, Office of Solid Waste, 2000.
7. Walsh, M.E. and T.A. Ranney, "Determination of Nitroaromatic, Nitramine, and Nitrate
Ester Explosives in Water Using Spe and GC-ECD,", U.S. Army Corps of Engineers, Cold
Regions Research and Engineering Laboratory, Special Report 98-2.
8. Jenkins, T.F., P.G. Thorne, E.F. McCormick, and K.F. Myers, "Preservation of Water
Samples Containing Nitroaromatics and Nitramines," U.S. Army Corps of Engineers, Cold
Regions Research and Engineering Laboratory, Special Report 95-16, 1995.
9. Winslow, S.D., B.V. Pepich, M.V. Bassett, S.C. Wendelken, D.J. Munch, and J.L.
Sinclair, "Microbial Inhibitors for U.S. EPA Drinking Water Methods for the
Determination of Organic Compounds," Environ. Sci. Technol., 35 (2001) 4103-4110.
10. Winslow, S.D., B. Prakash, MM Domino, B.V. Pepich and D.J. Munch, "Method 526,
Determination of Selected Semivolatile Organic Compounds in Drinking Water by Solid
Phase Extraction and Capillary Column Gas Chromatography/ Mass Spectrometry
(GC/MS)," In Methods for the Determination of Organic and Inorganic Compounds in
Drinking Water. Vol. 1, EPA 815-R-00-014, 2000.
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11. Erney, D.R., T.M. Pawlowski, C.F. Poole, "Matrix Induced Peak Enhancement of
Pesticides in Gas Chromatographv." J. High Resol. Chromatogr.. 20 (1997) 375-378.
12. Hajslova, J., k. Holadova, V. Kocourek, J. Poustka, M. Godula, P. Cuhra, M. Kempny,
"Matrix Induced Effects:A Critical Point in the Gas Chromatographic Analysis of Pesticide
Residues." J. Chromatogr.. 800 (1998) 283-295.
13. Erney, D.R., A.M. Gillespie, D.M. Gilvydis, and C.F. Poole, "Explanation of the Matrix-
Induced Chromatographic Response Enhancement of Organophosphorous Pesticides
During Open Tubular Column Gas Chromatography with Splitless or Hot On-column
Injection and Flame Photometric Detection," J. Chromatogr.. 638 (1993) 57-63.
529-33
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17. TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
Table 1. Ion Abundance Criteria for BFB
Mass
(m/z)
50
75
95
96
173
174
175
176
177
Relative Abundance Criteria
15-40% of mass 95
30-80% of mass 95
Base peak, 100% relative abundance
5-9% of mass 95
< 2% of mass 174
> 50% of mass 95
5-9% of mass 174
> 95% but < 101% of mass 174
5-9% of mass 176
Purpose of Checkpoint a
Low-mass sensitivity
Low-mass sensitivity
Low-mass resolution and sensitivity
Low-mass resolution and isotope ratio
Mid-mass resolution
Mid mass resolution and sensitivity
Mid-mass resolution
Mid-mass resolution and isotope ratio
Mid-mass resolution
a All ions are used primarily to check the mass measuring accuracy of the mass spectrometer and
data system, and this is the most important part of the performance test. The resolution checks,
which include natural abundance isotope ratios, constitute the next most important part of the
performance test. Finally, the ion abundance ranges are designed to encourage some
standardization to fragmentation patterns.
529-34
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Table 2. Retention Times and Suggested Quantitation and Confirmation Ions for
Explosive Analytes
a,b
Analyte
nitrobenzene (NB)
2-nitrotoluene (2NT)
3-nitrotoluene (3 NT)
4-nitrotoluene (4NT)
1 ,3-dinitrobenzene (DNB)
2,6-dinitrotoluene (2,6DNT)
2,4-dinitrotoluene (2,4DNT)
1,3,5-trinitrobenzene (TNB)
2,4,6-trinitrotoluene (TNT)
RDX
4-amino-2,6-dinitrotoluene (4AmDNT)
3,5-dinitroanaline (DNA)
2-amino-4,6-dinitrotoluene (2AmDNT)
Tetryl
3,4-dinitrotoluene (IS)
nitrobenzene- ds (SUR)
1 ,3,5-trimethyl-2-nitrobenzene (SUR)
1 ,2,4-trimethyl-5-nitrobenzene (SUR)
Retention
Time
(min)
7.72
9.40
10.18
10.57
15.13
15.32
16.60
19.35
19.43
21.97
22.81
23,00
23,47
24.77
17.42
7.66
12.33
14.72
Quantitation
Ion
(m/z)
51 or 77
65
91
65 or 91
50 or 75
165
165
75 or 213
210
46
180
183 or 64
180 or 78
194 or 77
63 or 182
82
148 or 91
148 or 91
Confirmation
Ions
(m/z)
51,77, 123
91, 120
65, 137
65,91, 137
50, 75, 76
63,89
63,78
74,75,213
63,89
75, 128
78, 104
63,64,183
52,78, 180
51,77, 194
63,78, 182
54, 128
91,93, 148
77,91, 148
a Even when instruments are tuned to BFB criteria, differences in instrumentation can cause
slight variations in mass spectra, e.g. quadrupole vs. ion trap. The ions listed here are suggested
ions because they were successfully used during method development.
Two or three ions from the Quantitation Ion and Confirmation Ion columns were used for SIM
monitoring.
529-35
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Table 3. Detection Limits Obtained Using Programmed Temperature Vaporizing (PTV)
Splitless Injection and Full Scan Mass Spectrometry
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene (TNT)
RDX
4- amino-2 ,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
DLOig/L)a
[fortified cone, for DL
determination]
0.039 [0.10]
0.11 [0.50]
0.083 [0.50]
0.13 [0.50]
0.021 [0.10]
0.029 [0.10]
0.11 [0.50]
0.037 [0.50]
0.084 [0.50]
0.082 [0.50]
0.065 [0.50]
0.052 [0.20]
0.075 [0.50]
0.18 [1.0]
DL Gig/L) b
[fortified cone, for DL
determination]
0.18 [0.50]
0.041 [0.10]
0.091 [0.50]
0.029 [0.10]
0.076 [0.50]
0.059 [0.50]
0.11 [0.50]
0.042 [0.50]
0.046 [0.50]
0.12 [0.50]
0.072 [0.50]
0.011 [0.10]
0.053 [0.50]
0.058 [1.0]
a Data from replicate disk extractions, n=8.
Data from replicate cartridge extractions, n=8.
529-36
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Table 4. Precision and Accuracy Data Obtained from Reagent Water Samples Fortified
with Method Analytes at Low Concentrations, Using SPE Disk Extraction, PTV
Splitless Injection and Full Scan Mass Spectrometry (n=8)
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 , 3 , 5 -trinitrobenzene
2,4,6-trinitrotoluene
RDX
4- amino-2 ,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
Fortified
Concentration
(Hi/L)
0.10
0.50
0.50
0.50
0.10
0.10
0.50
1.0
1.0
0.50
0.50
0.20
0.50
1.0
Accuracy
(% recovery)
70.6
102
114
81.4
126
83.6
101
111
136
103
130
130
127
134
Precision
(%RSD)
18
7.1
4.9
11
5.4
12
7.5
4.5
4.5
5.3
3.4
6.3
3.9
4.2
529-37
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Table 5. Precision and Accuracy Data Obtained from Reagent Water Samples Fortified
with Method Analytes at Low Concentrations, Using SPE Cartridge
Extraction, PTV Splitless Injection and Full Scan Mass Spectrometry (n=8)
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 , 3 , 5 -trinitrobenzene
2,4,6-trinitrotoluene
RDX
4- amino-2 ,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
Fortified
Concentration
(Hi/L)
0.5
0.5
0.5
0.1
0.5
0.5
0.5
0.5
0.5
0.5
0.5
0.1
0.5
1.0
Accuracy
(% recovery)
79.2
82.4
82.0
110
99.1
109
99.3
122
85.2
131
108
133
108
124
Precision
(%RSD)
16
15
7.4
9.0
5.1
3.6
7.1
2.3
3.6
6.3
4.4
2.6
3.3
1.6
529-38
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Table 6. Precision and Accuracy Data Obtained from Reagent Water Samples Fortified
with Method Analytes at 5.0 ng/L, Using PTV Splitless Injection and Full Scan
Mass Spectrometry
Analyte
nitrobenzene
2 -nitro toluene
3 -nitro toluene
4-nitrotoluene
1 ,3-dinitrobenzene
2 ,6-dinitrotoluene
2 ,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3 ,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
nitrobenzene-c/5 (SUR)
l,3,5-trimethyl-2-
nitrobenzene (SUR)
l,2,4-trimethyl-5-
nitrobenzene (SUR)
Disk Extraction (n=5)
Accuracy
(% recovery)
87.8
87.0
85.9
90.4
99.7
92.2
93.8
102
111
97.1
100
105
109
104
NDa
94.7
106
Precision
(%RSD)
7.8
5.6
7.9
1.4
8.5
7.5
3.7
4.4
3.2
2.5
7.5
2.8
3.5
5.4
ND
3.8
9.3
Cartridge Extraction (n=5)
Accuracy
(% recovery)
97.3
95.0
95.8
99.6
92.3
88.1
97.5
98.5
101
102
101
97.5
102
82.7
90.2
96.0
102
Precision
(%RSD)
9.3
12
5.9
5.0
6.3
5.1
4.8
4.9
6.0
8.1
4.8
4.4
3,1
5.4
8.3
2.7
6.0
ND- Not determined; These data were collected before this surrogate was added to the method.
529-39
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Table 7. Precision and Accuracy Data Obtained from Tap Water (Chlorinated Water
from a Surface Source) Fortified with Method Analytes at 5.0 ng/L, Using PTV
Splitless Injection and Full Scan Mass Spectrometrya
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2 ,6-dinitrotoluene
2 ,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3 ,5 -dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
nitrobenzene-c/5 (SUR)
l,3,5-trimethyl-2-
nitrobenzene (SUR)
l,2,4-trimethyl-5-
nitrobenzene (SUR)
Disk Extraction (n=5)
Accuracy
(% recovery)
96.3
84.5
81.2
98.1
103
96.1
101
111
109
111
108
114
115
116
NDb
109
112
Precision
(%RSD)
5.1
6.0
8.3
6.3
8.7
2.3
5.8
5.5
8.2
4.8
4.8
1.3
1.7
4.5
ND
9.9
4.0
Cartridge Extraction (n=5)
Accuracy
(% recovery)
88.0
85.6
91.1
87.5
87.1
82.8
94.3
101
97.9
107
100
97
101
87.6
87.5
90.6
86.6
Precision
(%RSD)
11
11
3,8
7.1
11
7.1
11
9.0
7.1
6.2
6.0
6.1
6.2
8.9
2.2
10
5.0
a The sample matrix was dechlorinated and preserved as described in Section 8 prior to
fortification with method analytes.
ND- Not determined; These data were collected before this surrogate was added to the method.
529-40
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Table 8. Precision and Accuracy Data Obtained from Tap Water (Chlorinated Water
from a Ground Source with a High Hardness Level a) Fortified with method
Analytes at 5.0 i-ig/L, Using PTV Splitless Injection and Full Scan Mass
Spectrometry
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2 ,6-dinitrotoluene
2 ,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3,5-dinitroanaline b
2-amino-4,6-dinitrotoluene
Tetryl
nitrobenzene- ds (SUR)
l,3,5-trimethyl-2-
nitrobenzene (SUR)
l,2,4-trimethyl-5-
nitrobenzene (SUR)
Disk Extraction (n=5)
Accuracy
(% recovery)
92.1
85.9
87.8
95.1
102
96.3
103
106
118
93.1
106
115
112
107
NDC
96.0
107
Precision
(%RSD)
17
12
5.4
9.7
3.4
4.3
6.9
8.2
8.0
9.7
6.8
8.3
7.4
16
ND
8.4
7.8
Cartridge Extraction (n=5)
Accuracy
(% recovery)
92.5
88.4
93.7
91.8
90.3
89.2
97.7
105
106
114
103
103
110
92.9
84.7
95.0
96.8
Precision
(%RSD)
12
4.1
7.7
7.7
9.0
4.3
4.7
6.6
4.8
11
4.1
4.5
6.4
7.9
15
10
6.2
a Hardness level measured at 310 and 360 mg/L as calcium carbonate, for disk and cartridge
samples, respectively. The sample matrix was dechlorinated and preserved as described in
Section 8 prior to fortification with method analytes.
3,5-Dinitroanaline was fortified at 0.2
ND - Not determined; These data were collected before this surrogate was added to the method.
529-41
-------
Table 9. Detection Limits Obtained Using Programmed Temperature Vaporizing (PTV)
Splitless Injection and Selected Ion Monitoring Mass Spectrometry
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene (TNT)
RDX
4- amino-2 ,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
DLOig/L)a
[fortified cone, for DL
determination]
0.009 [0.05]
0.008 [0.05]
0.005 [0.05]
0.005 [0.05]
0.006 [0.05]
0.004 [0.05]
0.004 [0.05]
0.01 8 [0.05]
0.004 [0.05]
0.006 [0.05]
0.005 [0.05]
0.008 [0.01]
0.020 [0.05]
0.090 [0.10]
DL Gig/L) b
[fortified cone, for DL
determination]
0.010 [0.05]
0.003 [0.05]
0.004 [0.05]
0.006 [0.05]
0.003 [0.05]
0.004 [0.05]
0.008 [0.05]
0.012 [0.05]
0.008 [0.05]
0.010 [0.05]
0.019 [0.05]
0.029 [0.01]
0.030 [0.05]
0.14 [0.25]
a Data from replicate disk extractions, n=8.
Data from replicate cartridge extractions, n=7.
529-42
-------
Table 10. Precision and Accuracy Data Obtained from Reagent Water Samples Fortified
with Method Analytes at Low Concentrations, Using SPE Disk Extraction, PTV
Splitless Injection and Selected Ion Monitoring Mass Spectrometry (n=8)
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 , 3 , 5 -trinitrobenzene
2,4,6-trinitrotoluene
RDX
4- amino-2 ,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
Fortified
Concentration
(Hi/L)
0.05
0.05
0.05
0.05
0.05
0.10
0.10
0.05
0.10
0.10
0.25
0.02
0.10
0.25
Accuracy
(% recovery)
87.1
79.7
112
123
124
89.2
95.7
120
104
133
98.4
107
119
113
Precision
(%RSD)
6.5
6.6
2.8
2.5
3.3
2.2
3.2
9.4
1.9
3.2
3.2
10
3.1
10
529-43
-------
Table 11. Precision and Accuracy Data Obtained from Reagent Water Samples Fortified
with Method Analytes at Low Concentrations, Using SPE Cartridge
Extraction, PTV Splitless Injection and Selected Ion Monitoring Mass
Spectrometry (n=7)
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4- amino-2 ,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
Fortified
Concentration
(H8/L)
0.05
0.05
0.05
0.05
0.05
0.10
0.10
0.10
0.10
0.10
0.25
0.05
0.10
0.25
Accuracy
(% recovery)
107
110
114
119
128
102
109
108
110
114
103
95.6
134
102
Precision
(%RSD)
5.7
1.8
2.0
3.2
1.6
3.1
5.4
8.9
4.3
8.9
3.7
9.4
5.6
17
529-44
-------
Table 12. Precision and Accuracy Data Obtained from Reagent Water Samples Fortified
with Method Analytes at 1.0 ng/L, Using PTV Splitless Injection and Selected
Ion Monitoring Mass Spectrometry
Analyte
nitrobenzene
2 -nitro toluene
3 -nitro toluene
4-nitrotoluene
1 ,3-dinitrobenzene
2 ,6-dinitrotoluene
2 ,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3,5-dinitroanaline a
2-amino-4,6-dinitrotoluene
Tetryl
nitrobenzene-c/5 (SUR)
l,3,5-trimethyl-2-
nitrobenzene (SUR)
l,2,4-trimethyl-5-
nitrobenzene (SUR)
Disk Extraction (n=4)
Accuracy
(% recovery)
91.3
89.7
91.3
90.5
99.0
98.1
102
106
110
100
116
104
113
122
88.5
84.9
87.4
Precision
(%RSD)
5.8
1.8
1.7
2.0
2.8
2.7
3.3
2.3
2.6
2.8
2.8
2.6
1.9
3.2
2.3
2.3
3.3
Cartridge Extraction (n=4)
Accuracy
(% recovery)
86.5
91.9
89.7
89.8
96.2
98.8
100
96.9
104
108
117
107
111
111
86.5
84.4
88.2
Precision
(%RSD)
3,4
3,0
4.8
4.1
5.9
5.1
4.9
6.2
5.1
6.4
4.7
8.6
5.7
8.9
2.2
1.6
2.4
3,5-Dinitroanaline was fortified at 0.2 i-ig/L.
529-45
-------
Table 13. Precision and Accuracy Data Obtained from Tap Water (Chlorinated Water
from a Surface Source) Fortified with Method Analytes at 1.0 ng/L, Using PTV
Splitless Injection and Selected Ion Monitoring Mass Spectrometrya
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2 ,6-dinitrotoluene
2 ,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3 ,5 -dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
nitrobenzene-c/5 (SUR)
l,3,5-trimethyl-2-
nitrobenzene (SUR)
l,2,4-trimethyl-5-
nitrobenzene (SUR)
Disk Extraction (n=4)
Accuracy
(% recovery)
71.5
79.8
78.9
74.9
80.7
81.2
88.7
82.3
91.1
85.2
103
96.5
97.4
99.5
87.4
93.3
100
Precision
(%RSD)
3.4
2.7
2.6
2.6
2.1
2.3
2.6
1.6
2.5
1.3
2.0
2.3
0.8
3.2
3.4
2.8
3.1
Cartridge Extraction (n=4)
Accuracy
(% recovery)
85.8
93.2
91.2
89.8
102
101
108
104
116
111
127
119
123
137
88.4
83.2
89.7
Precision
(%RSD)
4.8
4.2
3,6
4.1
2.4
2.2
2.4
3,3
3,8
3,8
2.5
1.8
2.5
3,8
5.1
4.6
4.1
a The sample matrix was dechlorinated and preserved as described in Section 8 prior to
fortification with method analytes.
3,5-Dinitroanaline was fortified at 0.2 ng/L.
529-46
-------
Table 14. Precision and Accuracy Data Obtained from Tap Water (Chlorinated Water
from a Ground Source with a High Hardness Level a ) Fortified with Method
Analytes at 1.0 ng/L, Using PTV Splitless Injection and Selected Ion
Monitoring Mass Spectrometry
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2 ,6-dinitrotoluene
2 ,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3,5-dinitroanaline b
2-amino-4,6-dinitrotoluene
Tetryl
nitrobenzene- d5 (SUR)
l,3,5-trimethyl-2-
nitrobenzene (SUR)
l,2,4-trimethyl-5-
nitrobenzene (SUR)
Disk Extraction (n=4)
Accuracy
(% recovery)
95.6
102
94.6
94.6
86.1
91.7
90.1
88.1
94.1
102
105
106
106
121
97.1
89.2
86.0
Precision
(%RSD)
8.6
4.6
3.3
5.1
5.1
2.4
5.9
4.2
9.4
3.4
7.1
5.6
4.6
4.1
5.5
6.5
3.1
Cartridge Extraction (n=4)
Accuracy
(% recovery)
84.2
85.1
85.0
85.5
104
104
109
109
113
124
117
98.5
113
120
87.6
84.7
92.2
Precision
(%RSD)
2.4
3,8
2.4
3,6
1.0
0.8
1.5
2.1
1.2
6.9
1.2
3,3
2.8
5.9
4.5
4.3
4.5
a Hardness level measured at 380 and 390 mg/L as calcium carbonate, for disk and cartridge
samples, respectively. The sample matrix was dechlorinated and preserved as described in
Section 8 prior to fortification with method analytes.
3,5-Dinitroanaline was fortified at 0.2 \igfL.
529-47
-------
Table 15. Detection Limits Obtained Using On-Column Programmed Temperature
Vaporizing (PTV) Injection and Full Scan Mass Spectrometry
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene (TNT)
Hexahydro-l,3,5-trinitro- 1,3,5-
triazine (RDX)
4- amino-2 ,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
2,4,6-trinitrophenylmethylnitramine
(Tetryl)
DLOig/L)a
[fortified cone, for DL
determination]
0.031 [0.10]
0.013 [0.10]
0.019 [0.05]
0.010 [0.05]
0.012 [0.05]
0.005 [0.05]
NDC
0.026 [0.05]
0.010 [0.05]
0.040 [0.10]
0.027 [0.10]
ND
0.05 [0.25]
0.28[1.0]d
DL Gig/L) b
[fortified cone, for DL
determination]
0.024 [0.05]
0.010 [0.05]
0.007 [0.05]
0.012 [0.05]
0.016 [0.05]
0.005 [0.05]
ND
0.050 [0.05]
0.007 [0.05]
0.024 [0.05]
0.007 [0.05]
ND
0.039 [0.25]
ND
a Data from replicate disk extractions, n=8.
b Data from replicate cartridge extractions, n=7.
c ND - Not determined; see Sect. 13.3 and 13.4.
dn=6
529-48
-------
Table 16. Precision and Accuracy Data Obtained from Reagent Water Samples Fortified
with Method Analytes at Low Concentrations, Using SPE Disk Extraction, On
Column PTV Injection and Full Scan Mass Spectrometry (n=8)
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 , 3 , 5 -trinitrobenzene
2,4,6-trinitrotoluene
RDX
4- amino-2 ,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
Fortified
Concentration
(Hi/L)
0.25
0.25
0.10
0.10
0.10
0.10
NDa
0.10
0.25
0.10
0.25
ND
1.0
1.0
Accuracy
(% recovery)
68.5
64.4
85.1
81.0
77.6
104
114
88.6
102
92.8
107
111
Precision
(%RSD)
12
5.9
6.7
5.5
5.6
3.8
16
4.3
13
5.8
7.1
5.2
a ND - Not determined; See Sect. 13.3 and 13.4.
529-49
-------
Table 17. Precision and Accuracy Data Obtained from Reagent Water Samples Fortified
with Method Analytes at Low Concentrations, Using SPE Cartridge
Extraction, On-Column PTV Injection and Full Scan Mass Spectrometry (n=8)
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 , 3 , 5 -trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3 ,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
Fortified
Concentration
(Hi/L)
0.10
0.10
0.05
0.05
0.25
0.10
NDa
0.25
0.25
0.10
0.25
ND
1.0
1.0
Accuracy
(% recovery)
77.0
74.9
114
116
92.1
114
126
116
117
101
106
108
Precision
(%RSD)
11
10
4.0
6.6
2.8
3.7
11.5
19
10
7.4
9.6
7.1
a ND - Not determined; see Sect. 13.3 and 13.4.
529-50
-------
Table 18. Precision and Accuracy Data Obtained from Reagent Water Samples Fortified
with Method Analytes at 1.0 ng/L, Using On-Column PTV Splitless Injection
and Full Scan Mass Spectrometry
Analyte
nitrobenzene
2 -nitro toluene
3 -nitro toluene
4-nitrotoluene
1 ,3-dinitrobenzene
2 ,6-dinitrotoluene
2 ,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
nitrobenzene-c/5 (SUR)
l,3,5-trimethyl-2-
nitrobenzene (SUR)
l,2,4-trimethyl-5-
nitrobenzene (SUR)
Disk Extraction (n=4)
Accuracy
(% recovery)
95.3
92.3
93.3
91.2
92.8
101
NDa
110
123
112
125
ND
107
111
90.4
92.8
96.8
Precision
(%RSD)
2.6
2.5
1.8
3.6
4.0
2.8
6.5
2.0
1.6
4.0
7.1
5.2
3.1
1.4
1.5
Cartridge Extraction (n=4)
Accuracy
(% recovery)
95.0
91.5
91.1
88.7
98.7
104
ND
105
117
122
123
ND
106
108
90.7
88.5
98.0
Precision
(%RSD)
1.7
3,4
1.3
1.7
8.0
4.1
14
6.3
5.9
6.8
9.6
7.1
2.6
2.2
3,0
ND - Not determined; See Sect. 13.3 and 13.4.
529-51
-------
Table 19. Precision and Accuracy Data Obtained from Tap Water (Chlorinated Water
from a Surface Source) Fortified with Method Analytes at 1.0 ng/L, Using On-
Column PTV Injection and Full Scan Mass Spectrometry a
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2,6-dinitrotoluene
2,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4- amino-2 ,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
nitrobenzene-c/5 (SUR)
l,3,5-trimethyl-2-
nitrobenzene (SUR)
l,2,4-trimethyl-5-
nitrobenzene (SUR)
Disk Extraction (n=4)
Accuracy
(% recovery)
78.6
77.1
78.8
76.9
83.6
86.7
NDb
107
112
99.3
107
ND
95.4
103
90.0
97.2
104
Precision
(%RSD)
4.1
3.2
2.8
3.3
2.8
2.8
6.7
2.8
11
2.7
5.5
2.7
5.0
3.8
3.6
Cartridge Extraction (n=4)
Accuracy
(% recovery)
95.1
90.0
91.7
88.5
95.9
105
110
131
127
128
107
123
93.8
86.2
96.7
Precision
(%RSD)
2.8
2.2
2.5
3,1
1.8
2.1
2.2
1.8
7.8
3,8
3,1
6.5
2.8
3,3
1.8
a The sample matrix was dechlorinated and preserved as described in Section 8 prior to
fortification with method analytes.
ND - Not determined; see Sect. 13.3 and 13.4.
529-52
-------
Table 20. Precision and Accuracy Data Obtained from Tap Water (Chlorinated Water
from a Ground Source with a High Hardness Level a) Fortified with Method
Analytes at 1.0 ng/L, Using On-Column PTV Injection and Full Scan Mass
Spectrometry
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dinitrobenzene
2 ,6-dinitro toluene
2 ,4-dinitro toluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
nitrobenzene- d5 (SUR)
l,3,5-trimethyl-2-
nitrobenzene (SUR)
l,2,4-trimethyl-5-
nitrobenzene (SUR)
Disk Extraction (n=4)
Accuracy
(% recovery)
94.1
88.5
92.2
89.0
96.4
101
NDb
129
133
126
128
ND
110
132
89.1
91.4
96.8
Precision
(%RSD)
2.4
2.7
3.0
2.5
4.0
2.4
5.3
3.0
5.6
4.4
2.4
5.6
4.4
4.3
4.6
Cartridge Extraction (n=4)
Accuracy
(% recovery)
91.4
90.0
89.9
89.6
107
105
123
136
150
140
121
135
88.2
92.1
97.3
Precision
(%RSD)
3,5
2.4
2.6
2.6
6.3
1.2
8.8
1.7
3,7
3,9
6.5
5.3
3,2
6.0
5.3
a Hardness level measured at 380 and 390 mg/L as calcium carbonate, for disk and cartridge
samples respectively. The sample matrix was dechlorinated and preserved as described in
Section 8 prior to fortification with method analytes.
b ND - Not determined; See Sect. 13.3 and 13.4.
529-53
-------
Table 21. Aqueous Holding Time Data for Surface Water Samples Fortified with Method
Analytes at 5 fig/L and Preserved and Stored as Directed in Section 8.
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dhiitrobenzene
2 ,6-dinitro toluene
2 ,4-dinitro toluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-dinitrotoluene
3 ,5-dinitroanaline
2-amino-4,6-dinitrotoluene
Tetryl
DayO
Amount a
fcg/L)
4.2
3.9
3.8
4.0
4.1
3.8
3.9
4.8
4.4
4.7
4.8
1.0
4.6
4.5
Day?
Amount
(jig/L)
4.4
4.1
3.9
4.1
4.3
3.7
4.0
5.4
4.5
4.8
5.1
1.1
4.8
4.9
Day 11
Amount
WL)
4.3
4.0
3.7
4.0
4.2
3,6
3,9
5.3
4.2
4.4
4.9
1.1
4.5
4.9
Day 14
Amount
(Hg/L)
4.1
3.8
3.6
3.9
4.1
3.6
3.6
5.2
3.9
4.4
4.7
1.1
4.5
4.4
Day 19
Amount
«L)
4.2
3.6
3.6
3.6
4.0
3.5
3.6
5.1
4.0
4.4
4.8
1.1
4.6
5.1
a Each concentration represents the mean of 5 replicate measurements. The RSD of each set of
replicates is less than 5.2%.
b 3,5-Dinitroanaline was fortified at lp,g/L.
529-54
-------
Table 22. Extract Holding Time Data for Surface Water Samples Fortified with Method
Analytes at 5 fig/L, Preserved and Stored as Directed in Section 8.
Analyte
nitrobenzene
2-nitrotoluene
3-nitrotoluene
4-nitrotoluene
1 ,3-dhiitrobenzene
2 ,6-dinitro toluene
2,4-dinitrotoluene
1 ,3,5-trinitrobenzene
2,4,6-trinitrotoluene
RDX
4-amino-2,6-
dinitrotoluene
3,5-dinitroanaline b
2-amino-4,6-
dinitrotoluene
Tetryl
DayO
Amount a
fcg/L)
4.4
4.1
4.0
4.2
4.2
3,9
4.0
4.7
4.4
4.6
4.7
1.0
4.6
4.4
Day?
Amount
fcg/L)
4.6
4.2
4.0
4.2
4.2
4.0
4.2
5.2
4.6
5.1
5.1
1.1
4.8
4.7
Day 15
Amount
fog/L)
4.4
4.1
3.8
4.1
4.2
3.8
3.7
5.0
4.2
4.6
4.8
1.1
4.4
4.7
Day 21
Amount
fcg/L)
4.2
3.8
3.6
3.7
4.0
3.5
3.6
5.0
4.0
4.5
4.7
1.1
4.5
4.6
Day 28
Amount
WL)
4.2
3.9
3.8
3.7
4.9
3.6
3.7
5.2
4.1
4.5
4.9
1.1
4.5
4.7
Day 35
Amount
fog/L)
4.3
4.2
4.0
3.9
4.3
3.7
3.8
5.2
4.2
4.4
4.9
1.1
4.7
4.7
a Each concentration represents the mean of 5 replicate measurements. The RSD of each set of
replicates is less than 15% except for 2-nitrotoluene on day 15 which was 23%.
b 3,5-Dinitroanaline was fortified at lp,g/L.
529-55
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Table 23. Initial Demonstration of Capability (IDC) Requirements (Summary)
Method Ref.
Requirement
Specification and Frequency
Acceptance Criteria
Sect. 9.2.1
Initial Demonstration of
Low Method
Background
Analyze LRB prior to any other IDC
steps-
Demonstrate that all target analytes are
< 1/3 the MRL, and that possible
interferences from extraction media do
not prevent the identification and
quantification of method analytes.
Sect. 9.2.2
Initial Demonstration of
Precision (IDP)
Analyze 4-7 replicate LFBs fortified at 2-
RSD must be <20% for all analytes.
Sect. 9.2.3
Initial Demonstration of
Accuracy (IDA)
Calculate average recovery for replicates
used in IDP
Mean recovery 70-130% of true value.
Sect. 9.2.4
Detection Limit
(DL) Determination
Over a period of three days, prepare a
minimum of 7 replicate LFBs fortified at
a concentration estimated to be near the
DL. Analyze the replicates through all
steps of the analysis. Calculate the DL
using the equation in Section 9.2.4.
NOTE: Data from DL replicates are not
required to meet method precision and
accuracy criteria. If the DL replicates are
fortified at a low enough concentration, it
is likely that they will not meet precision
and accuracy criteria.
529-56
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Table 24. Quality Control Requirements (Summary)
Method Ref.
Sect. 8.1 -
Sect 8.4
Sect. 8.4
Sect. 9.10
Sect. 9.11
Sect. 9.12
Sect 9.4
Requirement
Sample Collection,
Preservation, and
Holding Time
Extract Holding Time
Laboratory Fortified
Sample Matrix (LFM)
Field Duplicates
Quality Control Sample
(QCS)
Laboratory Reagent
Blank (LRB)
Specification and Frequency
14 days, with addition of Trizma buffer
and copper sulfate
30 days
Analyze one LFM per extraction batch
(20 samples or less) fortified with method
analytes at approximately 2 times the
native concentration.
Analyze 1 FD for each 20 samples, or 1
per extraction batch, whichever is greater.
Analyze QCS whenever new standards
are prepared, or at least quarterly.
One with each extraction batch of up to
20 samples.
Acceptance Criteria
Iced or refrigerated at 10° C or less for up
to 48 hours to allow time for shipping;
refrigerated at 6° C or less after arrival at
the laboratory.
Stored at 0° C or less in amber vials.
Recoveries not within 70-130% of the
fortified amount may indicate a matrix
effect.
Suggested RPD±30%.
If analyzed as a calibration sample, CCC
criteria apply. If analyzed as an LFB,
those criteria apply.
Demonstrate that all target analytes are
< 1/3 the MRL, and that possible
interference from extraction media do not
prevent the identification and
quantification of method analytes.
529-57
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Method Ref.
Requirement
Specification and Frequency
Acceptance Criteria
Sect. 9.7
Laboratory Fortified
Blanks (LFB)
Analyze at least one LFB daily or one for
each extraction batch of up to 20 field
samples. Rotate the fortified concentration
between low, medium and high amounts.
Results of LFB analyses must be 70-
130% of the true value for each analyte
and surrogate for all fortified
concentrations greater than the lowest
CAL point. Results of LFBs
corresponding to the lowest CAL point
must be 50-150% of the true value.
Sect. 9.8
Internal Standard
The internal standard, 3,4-dinitrotoluene,
is added to all calibration standards and
extracts.
Peak area counts for the IS in LFBs,
LRBs and sample extracts must be within
70-130% of the peak area in the most
recent CCC, and 50-150% of average area
in the initial calibration.
Sect. 9.9
Surrogate Standards
Surrogate standards (nitrobenzene-J5;
1,3,5-trimethyl-2-nitro-benzene; and
1,2,4-trimethyl-5-nitro-benzene) are
added to all calibration standards,
samples, LFBs, LFMs, FDs, and LRBs.
Recovery for all surrogates in all
calibration standards, LRB, LFB, LFM,
FD and sample extracts must be 70-130%
of the true value.
Sect. 10.2.1
MS Tune Check
Analyze BFB to verify MS tune before
initial calibration and before every
recalibration.
Criteria are given in Table 1.
529-58
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Method Ref.
Requirement
Specification and Frequency
Acceptance Criteria
Sect. 10.2.2-
Sect. 10.2.10
Initial Calibration
Use internal standard calibration
technique to generate an average RF or
first or second order calibration curve.
Use a minimum of 3 standards for a
calibration range of 1 order of magnitude,
and at least 5 standards for 2 orders of
magnitude.
When each calibration standard is
calculated as an unknown using the
calibration curve, the result must be 70-
130% of the true value for all but the
lowest standard. The lowest standard
must be 50-150% of the true value.
Sect. 10.3
Continuing Calibration
Check
Verify initial calibration by analyzing a
calibration standard prior to analyzing
samples, after every 10 samples, and after
the last sample. Always analyze a low
concentration (near the MRL) CCC at the
beginning of the analysis period. Rotate
through low, medium, and high
concentration calibration standards to
meet every 10 sample requirement.
The result for each analyte and surrogate
must be 70-130% of the true value for all
concentrations except the lowest CAL
point for each analyte. The lowest CAL
point for each analyte must be 50-150%
of the true value.
The peak area of the IS must be within
70-130% of the peak area in the most
recent CCC, and 50-150% of the average
peak area calculated during initial
calibration.
529-59
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Figure 1
Chromatogram of Explosives and Related Compounds
100% -
1
•
TOT-
•
•
SUR
SUR+ 2NT 3 T4NT
N
IB
4
k
SUR
NG
2,
DNB
6DNT
800 1000 1200 1400
8.40 10.50 12.60 14.69
100%
TOT
2.4DNT
IS
1600
16.79
TNT
PETN
—4—
RDX
1
Jl
4AmDNT
2AmDNT
DMA
Tetiyl
(
JV
1800
18.90
2000
21.00
2200
23.10
Total ion Chromatogram of 16 explosives and related compounds, using a J&W
DBS-MS column and the conditions described in Sect. 10.2.2.1; 5ng each analyte,
surrogate and internal standard, except DMA which is 1 ng. See Table 2 and Sect.
13.2 for compound abbreviations used to label peaks.
* Candidate internal standard that was eventually dropped from the method.
529-60
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