METHOD 528 DETERMINATION OF PHENOLS IN DRINKING WATER
BY SOLID PHASE EXTRACTION AND CAPILLARY COLUMN
GAS CHROMATOGRAPHY/MASS SPECTROMETRY (GC/MS)
J.W. Munch - April 2000
NATIONAL EXPOSURE RESEARCH LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
CINCINNATI, OHIO 45268
DETERMINATION OF PHENOLS 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 phenols 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 phenols that are efficiently partitioned from the water sample onto a modified
polystyrene divinylbenzene solid phase sorbent, and sufficiently volatile and thermally
stable for gas chromatography. The method includes the following compounds:
ANALYTE CAS NUMBER
2-methylphenol (o-cresol) 95-48-7
1.2 Method detection limit (MDL) is defined as the statistically calculated minimum concentra-
tion that can be measured with 99% confidence that the reported value is greater than zero
(1). The MDL is compound dependent and is particularly dependent on extraction
efficiency, sample matrix, and instrument performance. MDLs for method analytes range
from 0.02-0.58 :g/L, and are listed in Table 1. The concentration calibration range
demonstrated by this method is 0.1 :g/L to 15 :g/L for most analytes, and approximately
1.0 :g/L to 15 :g/L for 2,4-dinitrophenol, 4-nitrophenol, 2-methyl-4,6-dinitrophenol, and
1.3 This method should be performed only by analysts with experience in solid phase extrac-
tions 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 0.5 g of a modified polystyrene divinyl benzene copolymer.
The organic compounds are eluted from the solid phase with a small quantity of methylene
chloride. 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. 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.
Reference spectra and retention times for analytes are obtained by the measurement of calibra-
tion 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.
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 Calibra-
tion 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 surrogate
analytes. The CAL solutions are used to calibrate the instrument response with respect to
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 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.
3.5 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
3.6 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.
3.7 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.
3.8 LABORATORY FORTIFIED SAMPLE MATRIX (LFM) -- An aliquot of an environ-
mental 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.
3.9 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.
3.10 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, and 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
3.11 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.
3.12 METHOD DETECTION LIMIT (MDL) -- 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 (Section 9.2.4), and
accurate quantitation is not expected at this level. (1)
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 PEAK TAILING FACTOR (PTF) -- A calculated value that indicates the amount of
peak tailing exhibited by a chromatographic peak. The value is calculated by dividing the
peak width of the back half of the peak (at 10% peak height), by the peak width of the
front half of the peak (at 10% peak height). The calculation is demonstrated in Figure 4.
3.15 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.
3.16 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 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.
3.18 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.
4.1 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 the polypropylene cartridge that it is
packed in. Brands and manufacturers lot numbers of these devices should be monitored
and tracked to ensure that contamination will not preclude analyte identification and
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. Syringes and splitless injection port liners must be cleaned
carefully or replaced as needed. 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.
4.3 Silicone compounds may be leached from autosampler vial septa by methylene chloride.
This contamination of the extract will be enhanced if particles of the septa are introduced
into standards and sample extracts by the needle used for injection. These silicone
compounds should, in most cases, have no effect on the analysis. However, the analyst
should be aware of this potential problem.
4.4 Airborne phenol may be a source of phenol contamination in samples and sample extracts.
Samples should not be stored or extracted in areas where phenol is used for other
4.5 2,3,4,5-Tetrachlorophenol is used as one of the internal standards for the quantitation of
reactive and thermally labile phenols. Tetrachlorophenol isomers may be present at low
levels (less than 4% total tetrachlorophenol) in pentachlorophenol used as a pesticide and
wood preservative. However, occurrence of pentachlorophenol in U.S. drinking waters is
rare, and measured concentrations are typically 1 :g/L or less. If a matrix interference
with the internal standard is suspected, an alternate internal standard may be selected.
5.1 The toxicity or carcinogenicity of 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. Additional references to
laboratory safety are cited (2-4).
5.2 Some method analytes and solvents, including 2,4,6-trichlorophenol, pentachloro- phenol,
and methylene chloride 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.
6. EQUIPMENT AND SUPPLIES (All specifications are suggested. References to specific
brands or catalog numbers are included for illustration only.)
6.1 GLASSWARE -- All glassware must be meticulously cleaned. This may be
accomplished by washing with detergent and water, rinsing with water, distilled water, or
solvents, air-drying, and heating (where appropriate) 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. Although specific contamination
problems from bottle caps were not observed during method development, phenolic resin
bottle caps should be avoided.
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.
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. During the course of the development of this method,
two columns were used. Although these are both polyphenylmethylsilicone columns, the
exact phase is slightly different. Information on the exact composition of each phase is
available from the manufacturers. Most of the work was performed with column 1. Any
column which provides analyte separations equivalent to or better than these columns may
be used. Example chromatograms are shown in Figs 1-3. Retention times are presented
in Table 2.
6.9.1. Column 1- 30 m ?0.25 mm id fused silica capillary column coated with a 0.25
:m bonded film of polyphenylmethylsilicone, (J&W DB-5ms).
6.9.2 Column 2- 30 m ?0.25 mm id fused silica capillary column coated with a 0.25
:m bonded film of polyphenylmethylsilicone, (SGE BPX5).
6.10 GAS CHROMATOGRAPH/MASS SPECTROMETER/DATA SYSTEM
6.10.1 The GC must be capable of temperature programming and should be equipped
for split/splitless injection. The injection system must not allow the analytes to
contact hot stainless steel or other metal surfaces that promote decomposition.
Other injection techniques such as temperature programmed injections, cold on-
column injections and large volume injections may be used if QC criteria in
Section 9 and 10 are met. If an alternate injection technique is performed, the
analyst will need to select an instrument configuration which has been specifically
designed for that application. Performance data in Section 17 include data
obtained both by hot, splitless injection and temperature programmed splitless
6.10.2 The GC/MS interface should allow the capillary column or transfer line exit to be
placed within a few mm of the ion source. Other interfaces, for example the
open split interface, are acceptable if the system has adequate sensitivity.
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 be
capable of scanning at a minimum from 45 to 450 amu with a complete scan
cycle time (including scan overhead) of 1.0 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 3 when an injection of approximately 5 ng of DFTPP 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 recommended.
6.10.4 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) 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 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 cartridge. 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.
7. REAGENTS AND STANDARDS
7.1 HELIUM -- carrier gas, purity as recommended by the GC/MS manufacturer.
7.2 SOLID PHASE EXTRACTION CARTRIDGES -- Varian Bond Elut PPL or equivalent.
Cartridges are inert non-leaching plastic, for example polypropylene, or glass, and must
not contain plasticizers that leach into the methylene chloride eluant and prevent the
identification and quantitation of method analytes. The polypropylene cartridges (6 mL
volume) are packed with 0.5 g highly cross-linked, and chemically modified styrene divinyl
benzene copolymer. The packing must have a narrow size distribution and must not leach
interfering organic compounds into the eluting solvent.
7.3 SOLVENTS --
7.3.1 Methylene chloride, acetone, and methanol. High purity pesticide quality or
7.3.2 Reagent water. Water in which an interference is not observed at >1/3 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.
7.4 HYDROCHLORIC ACID -- 6 N and 0.05 N.
7.5 SODIUM SULFATE, ANHYDROUS -- (Soxhlet extracted with methylene chloride for
a minimum of 4 h or heated to 400癈 for 2 h in a muffle furnace.)
7.6 STOCK STANDARD SOLUTIONS -- Individual solutions of surrogates, internal
standards, and analytes, or mixtures of analytes, may be purchased from commercial
suppliers or prepared from pure materials. To prepare stocks from neat materials, add 10
mg (weighed on an analytical balance to within 0.1 mg) of the pure material to 1.9 mL of
methanol, methylene chloride, or acetone in a 2 mL volumetric flask, dilute to the mark,
and transfer the solution to an amber glass vial. The solvent to be used is dependent upon
the final use of the standard. In general, calibration standards and internal standards are
prepared in methylene chloride, sample fortification solutions are prepared in methanol or
acetone. Follow any specific instructions for each standard or standard mixture. If
compound purity is confirmed by the supplier at >96%, the weighed amount can be used
without correction to calculate the concentration of the solution (5 :g/:L). Store the
amber vials at 0癈 or less.
7.7 PRIMARY DILUTION STANDARD SOLUTION -- The stock standard solutions are
used to prepare a primary dilution standard solution that contains multiple method analytes
in methylene chloride. 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, that is, 15 ng/:L. Store
the primary dilution standard solution in an amber vial at 0癈 or less, and check regularly
for signs of degradation or evaporation, especially just before preparing calibration
solutions. Mixtures of method analytes to be used as primary dilution standards may also
be purchased from commercial suppliers.
7.8 CALIBRATION SOLUTIONS (CAL1 through CAL7) -- Prepare a series of seven
calibration solutions in methylene chloride which contain analytes of interest at suggested
concentrations of 15,10, 5, 2, 1, 0.5, and 0.1 ng/:L, with a constant concentration of
each internal standard in each CAL solution (2-5 ng/:L is recommended). Surrogate
analytes are also added to each CAL solution, and may be added at a constant
concentration or varied concentrations (similar to those for method analytes), at the
discretion of the analyst. CAL1 through CAL7 are prepared by combining appropriate
aliquots of a primary dilution standard solution (Sect. 7.7) and the fortification solution of
internal standards and surrogates (Sect. 7.10). All calibration solutions should contain at
least 80% methylene chloride to avoid gas chromatographic problems due to mixed
solvents. Store these solutions in amber vials at 0癈 or less. Check these solutions
regularly for signs of evaporation and/or degradation.
NOTE: Because the MS sensitivity to analytes 9-12 (Table 2) is significantly less than
compounds 1-8, it may be more convenient to prepare calibration solutions in which the
concentrations of analytes 9-12 (Table 2) are higher than the concentrations of analytes 1-
8. Use of this option is at the discretion of the analyst. Calibration requirements are
specified in Sect. 10.
7.9 INTERNAL STANDARD SOLUTION(S) -- This method uses two internal standards:
1,2-dimethyl-3-nitrobenzene (IS#1) and 2,3,4,5-tetrachlorophenol (IS#2). The first
internal standard, 1,2-dimethyl-3-nitrobenzene is used to monitor instrument sensitivity and
is used to quantify analytes 1-8 in Table 2. The second internal standard, 2,3,4,5-
tetrachlorophenol is used to quantify analytes 9-12 (Table 2). IS#2 was selected for its
chemical similarity to these compounds which are susceptible to adsorption and/or thermal
decomposition in the GC inlet. A full explanation of the use of 2,3,4,5-tetrachlorophenol
to quantify these compounds is given in Section 13. If cold, on-column or temperature
programmed injection techniques are used, acceptable performance may be obtained
using only one internal standard (IS#1).
1,2-Dimethyl-3-nitrobenzene (Aldrich) -- 100 :g/mL in methylene chloride.
Use 25 :L of this solution per 1 mL of sample extract for a final concentration of
2,3,4,5-Tetrachlorophenol (Chem Service Inc.) -- 200 :g/mL in methylene
chloride. Use 25 :L of this solution per 1 mL of sample extract for a final
concentration of 5 :g/mL.
7.9.3 The internal standard solutions listed above can be made individually or together
in one solution.
7.10 SAMPLE FORTIFICATION SOLUTIONS --
7.10.1 Surrogate fortification solutions --
2-Chlorophenol-3,4,5,6-d4 (Chem Service Inc.) -- 100 :g/mL in
methanol. Use 20 :L of this solution per 1 L of water sample for a
final concentration of 2 :g/L.
2,4-Dimethylphenol-3,5,6-d3 (CDN Isotopes) -- 100 :g/mL in
acetone. Use 20 :L of this solution per 1 L of water sample for a
final concentration of 2 :g/L.
2,4,6-Tribromophenol -- 200 :g/mL in methanol. Use 25 :L of this
solution per 1 L water sample for a final concentration of 5 :g/L.
7.10.2 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 :g/mL of each analyte, was used for higher concentration
fortifications, and the other containing10 :g/mL of each analyte in methanol was
used for lower level fortifications.
NOTE: Because the MS sensitivity to analytes 9-12 (Table 2) is significantly
less than analytes 1-8, it may be more convenient to prepare analyte fortification
solutions in which the concentrations of analytes 9-12 are higher than the
concentrations of analytes 1-8. Use of this option is at the discretion of the
7.11 GC/MS TUNE CHECK SOLUTION -- Decafluorotriphenylphosphine (DFTPP), 5
:g/mL in methylene chloride. Store this solution in an amber vial at 0癈 or less.
7.12 SODIUM SULFITE, ANHYDROUS -- Reducing agent used to reduce residual chlorine
at the time of sample collection.
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. 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 acidified at the time of collection. Residual chlorine is reduced by
addition of 40-50 mg of sodium sulfite. It may be added as a solid to the sample bottles
before the bottles are transported to the field. It is very important that the sample be
dechlorinated prior to acidification. Wait until sodium sulfite is dissolved before
acidification. Adding sodium sulfite and HCl (together) to the sample bottles prior to
shipping bottles to the sampling site is not permitted. After dechlorination, samples are
acidified to less than pH 2 with 6 N hydrochloric acid. The acid serves as a chemical and
biological preservative. This pH is the same that is used in the sample extraction, and is
required to support the recovery of several method analytes.
8.3 SAMPLE TRANSPORT AND STORAGE -- All samples should be iced during
shipment and must not exceed 10o C during the first 48 hours. Samples should be
confirmed to be at or below 10o C when they are received at the laboratory. Samples
stored in the lab must be held at or below 6o C until extraction, but should not be frozen.
8.4 HOLDING TIME -- Results of holding time studies of all method analytes showed that all
compounds are stable for 14 days in water samples when the samples are dechlorinated,
preserved, and stored as described in Sect. 8.2 and 8.3. Therefore, samples must be
extracted within 14 days of collection. Sample extracts may be stored at 0癈 or less for
up to 30 days after sample extraction. Data from holding time studies are shown in Tables
7 and 8.
9. QUALITY CONTROL
9.1 Quality control (QC) requirements include: the initial demonstration of laboratory
capability (summarized in Table 9) followed by regular analyses of continuing calibration
checks, laboratory performance check standards, laboratory reagent blanks, laboratory
fortified blanks, and laboratory fortified matrix samples. An MDL 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 10.
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 9.
9.2.1 INITIAL DEMONSTRATION OF LOW CARTRIDGE EXTRACTION
BACKGROUND AND SYSTEM BACKGROUND -- Before any samples
are analyzed, or any time a new supply of solid phase extraction 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.
188.8.131.52 A source of potential contamination is the solid phase extraction
cartridge which may contain phthalate esters, silicon compounds, and
other contaminants that could interfere with the determination of
method analytes. Although extraction cartridges 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.
184.108.40.206 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
9.2.2 INITIAL DEMONSTRATION OF PRECISION (IDP) -- Prepare 4-7
replicate LFBs fortified at 5-10 :g/L, or other mid-range concentration. Sample
preservatives described in Sect. 8.2 must be added to these samples. Extract
and analyze these replicates according to the procedure described in Section 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 with the exception of
phenol. The RSD for replicate analyses for phenol must be less than or equal to
9.2.3 INITIAL DEMONSTRATION OF ACCURACY (IDA) -- Using the same
set of replicate data generated for Section 9.2.2, calculate average recovery.
The average recovery of the replicate values must be within 70-130% of the true
value, except for phenol. Phenol will typically be recovered less effectively than
other method analytes. Because of its higher water solubility some breakthrough
from the extraction cartridge does occur. The recovery limits for phenol are 50-150%.
9.2.4 MDL 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 at a
concentration estimated to be near the MDL. This concentration may be
estimated by selecting a concentration at 2-5 times the noise level.
Concentrations shown in the example data in Table 1 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 Section 11. Calculate the MDL using the following
MDL = St( n - 1, 1 - alpha = 0.99)
t( n - 1,1 - alpha = 0.99) = 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 MDL calculations.
9.2.5 The analyst is permitted to modify GC columns, GC conditions, extract
evaporation techniques, internal standards or surrogate compounds. 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 MDL 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 DFTPP. 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).
9.7 PEAK TAILING FACTOR (PTF) -- This check consists of calculating the PTF as
described in Sect. 10.2.3.1. and in Figure 4. This check must be performed once every
24 hr of instrument operation.
9.8 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 a 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 lowest CAL point for an analyte must be 50-150% of
the true value for all analytes. Results of LFB analysis from medium and high level
concentrations must be 70-130% of the true value for all analytes except phenol. The
acceptance limit for phenol is 50-150% of the true value.
9.9 INTERNAL STANDARD (IS) --The analyst must monitor the peak area of the 1,2-
dimethyl-3-nitrobenzene (IS#1) in all injections during each analysis day. The IS#1
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#1 area in a chromatographic
run does not meet these criteria inject a second aliquot of that extract.
NOTE: The peak area of 2,3,4,5-tetrachlorophenol may not be consistent. It may vary
depending upon the composition of the extract or standard being analyzed. See Section
13.2.1 for a detailed explanation.
9.9.1 If the reinjected aliquot produces an acceptable internal standard response,
report results for that aliquot.
9.9.2 If a deviation of greater than 30% is obtained for the reinjected extract, when
compared to the most recent CCC, the analyst should check the calibration by
reanalyzing the most recently acceptable calibration standard. If the calibration
standard fails the criteria of Section 10.3.3, recalibration is in order per Section
10. 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.
9.10 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.
9.10.1 Surrogate recovery criteria are 70-130% of the fortified amount for 2-
chlorophenol-3,4,5,6-d4 and 2,4-dimethylphenol-3,5,6-d3. The criteria for
2,4,6-tribromophenol is 60-130% of the fortified amount. 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
9.10.2 If the extract reanalysis meets the surrogate recovery criterion, report only data
for the reanalyzed extract.
9.10.3 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 Section 10.3.3, recalibration is in
order per Section 10. 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 reextract also fails the recovery criterion, report all
data for that sample as suspect.
9.11 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, matrix independence should be established for each. 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 a LFM for
every 20 samples. If the recovery data for an LFM does not meet the recovery criteria in
Sect. 9.8, 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.
9.11.1 Within each extraction batch, a minimum of one field sample is fortified as a
LFM for every 20 samples analyzed. The LFM is prepared by spiking a sample
with an appropriate amount of the fortification solution. The concentrations 5,
10, and 15 :g/L are suggested spiking concentrations. Select the spiking
concentration that is closest to, and at least twice the matrix background
concentration. Use historical data or rotate through the designated
concentrations to select a fortifying concentration. Selecting a duplicate bottle of
a sample that has already been analyzed, aids in the selection of appropriate
9.11.2 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.,
where C is the fortified concentration. Compare these values to control
limits for LFBs (Sect. 9.8).
9.11.3 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 except phenol which should be recovered at 50-150%. If the
accuracy 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
NOTE: Matrix effects are expected to be more likely with compounds 9-12
(Table 2) than other method analytes.
9.12 FIELD DUPLICATES (FD) -- Within each extraction batch, a minimum of one field
sample should be analyzed in duplicate. Duplicate sample analyses serve as a check on
sampling and laboratory precision. If analytes are not routinely observed in field samples,
duplicate LFMDs should be analyzed to substitute for this requirement.
9.12.1 Calculate the relative percent difference (RPD) for duplicate measurements
(FD1 and FD2) as shown below.
9.12.2 Relative percent differences for laboratory duplicates and LFMDs 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.
9.13 QUALITY CONTROL SAMPLE (QCS) -- Each time that new standards 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 a LFB, then the acceptance criteria are the same as for an LFB (Sect.
9.8). 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
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 calibration. A peak tailing factor check is
required every day that samples are analyzed. In periods of continuous operation, the
peak tailing factor must be performed every 24 hr.
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
DFTPP solution into the GC/MS system. Acquire a mass spectrum that
includes data for m/z 45-450. If the DFTPP mass spectrum does not meet all
criteria in Table 3, 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.
The tune check may be performed as a separate analysis, or for routine MS tune
verification, DFTPP may be added to one or more of the CAL standards used
for calibration verification, so that the tune check and calibration verification can
be performed in a single analysis. DFTPP elutes shortly after pentachlorophenol
on both of the columns cited in Sect.6.9.
10.2.2 ANALYTE CALIBRATION -- Inject an aliquot of a medium concentration
calibration solution. For example, 2-10 :g/mL, and acquire and store data from
m/z 45-350 with a total cycle time (including scan overhead time) of 1.0 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
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
10.2.2.1 The following parameters are suggested GC conditions for hot,
splitless injection: injector temperature 200o C, carrier gas head
pressure 12-15 psi. Inject at an oven temperature of 35o C and hold
in splitless mode for 0.2 min. After 6 min, temperature program the
GC oven at 8o C per min to 250o C. Start data acquisition at
approximately 10 min. Example chromatograms are shown in
Figures 1 and 2.
10.2.2.2 The following parameters are suggested injection conditions for
temperature programmed splitless injection. Inject with the injector
temperature at 25?C, program the injector at 200?C per min to
200?C. Hold in the splitless mode for 1.0 min. Use the same
column temperature program as listed in Sect. 10.2.2.1. An
example chromatogram is shown in Figure 3.
10.2.3 Performance criteria for the calibration standards. Examine the stored GC/MS
data with the data system software.
10.2.3.1 PEAK TAILING FACTOR (PTF) -- Peak tailing can be a
problem associated with phenols. The phenols most likely to tail are
those with low acidity constants: 2,4-dinitrophenol, 4-nitro-phenol,
pentachlorophenol and 2-methyl-4,6-dinitrophenol. These
compounds must exhibit a peak tailing factor of 5 or less at a
concentration equivalent to 5-10 :g/L in a water sample (5-10
:g/mL in an extract or calibration standard). For example peak
tailing factor calculations, see Fig.4. Peak tailing factors must be
evaluated for the four analytes listed above each day that samples
are analyzed. In periods of continuous instrument operation, verify
acceptable PTFs every 24 hr. Peak tailing factors may be evaluated
in either a CAL standard, LFB or LFM.
10.2.3.2 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. 11.5).
10.2.4 If all performance criteria are met, inject an aliquot of an appropriate volume
(usually 1-2 :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 (0.1
and 0.5 :g/mL). 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. Data outside of the
established calibration range should never be reported.
10.2.5 Concentrations may be 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
Average RF is determined by calculating the mean RF of each calibration point,
with a minimum of five calibration concentrations.
( Ax)( Qis)
( Ais)( Qx)
Ax = integrated abundance (peak area) of the quantitation ion of
Ais = integrated abundance (peak area) of the quantitation ion
Qx = quantity of analyte injected in ng or concentration units.
Qis = quantity of internal standard injected in ng or concentration
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 /Q is 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
point, 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 presented in this method were
obtained using linear regression (RF vs Ax /Ais). Quadratic fit calibrations
should be used with caution, because the non-linear area of the curve may not be
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 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. 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
1,3-dimethyl-2-nitrobenzene has 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
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, 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.
10.3.4.2 Clean or replace the splitless injection liner; silanize a new injection
10.3.4.3 Flush the GC column with solvent according to manufacturer's
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.
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.
11.1 CARTRIDGE 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. 11.2). Verify that the sample is at pH 2 or less and
is free of residual chlorine. If the sample is a LRB or LFB, add sodium sulfite
and acidify following procedures 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 2-5 :g/L. If the
sample is a LFB or LFM, add the desired amount of analyte fortification
11.1.3 CARTRIDGE CLEAN-UP AND CONDITIONING -- Rinse each cartridge
with three, 3 mL aliquots of methylene chloride. Let the cartridge drain dry after
each flush. Then rinse the cartridge with three, 3mL 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 with three,
3mL aliquots of 0.05 N hydrochloric acid, but before the dilute acid level drops
below the top edge of the packing, turn off the vacuum. Add approximately 3
mL additional 0.05 N hydrochloric acid to the cartridge, attach the transfer tube,
and turn on the vacuum, and begin adding sample to the cartridge.
11.1.4 Adjust the vacuum so that the approximate flow rate is 20 mL/min (50 min for a
1 L sample). After all of the sample has passed through the SPE cartridge, draw
air or nitrogen through the cartridge for 15-30 min at high vacuum (10-15 in Hg).
The cartridge packing should appear dry (light tan color) before continuing with
the elution steps. It is important that the cartridge packing be dry, in order to
obtain good recoveries. The drying time may vary, depending upon the strength
of the vacuum source, and the number of cartridges being processed
simultaneously. The color and appearance of the packing is the most reliable
indicator of dryness. During the method development, drying for more than 60
minutes was not observed to have any negative effect upon the sample data.
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.
11.1.5 Rinse the inside of each sample bottle with 8-10 mL methylene chloride and use
vacuum to pull the 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 2-3 mL methylene chloride 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. Small amounts of
residual water from the sample container and the SPE cartridge may form an
immiscible layer with the eluate. Pass the eluate through the drying column (Sect.
6.7), which is packed with approximately 5 to 7 grams of anhydrous sodium
sulfate, and collect in a clean collection tube. Wash the sodium sulfate with at
least 2 mL methylene chloride and collect in the same tube. Concentrate the
extract to approximately 0.9 mL in a warm (40癈) water bath under a gentle
stream of nitrogen. Do not concentrate the extract to less than 0.5 mL, as this
will result in losses of analytes. Add the internal standards (Sect 7.9). Adjust
final volume to 1 mL. Make any volume adjustments with methylene chloride.
11.2 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 may be 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.
11.3 Analyze an aliquot of the sample extract with the GC/MS system under the same
conditions used for the initial and continuing calibrations (Sect. 10.2.2 and 10.3).
11.4 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
interest. Use the data system software to examine the ion abundances of components of
11.5 Identification of analytes. 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.
11.5.1 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%. Some ions, particularly the
molecular ion, are of special importance, and should be evaluated even if they
are below 10% relative abundance.
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
characteristic 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 Section
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 Section 11.2.
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). Experience indicates that three significant figures may
be used for concentrations above 99 :g/L, two significant figures for
concentrations between 1.0-99 :g/L, and one significant figure for lower
13. METHOD PERFORMANCE
13.1 PRECISION, ACCURACY AND MDLs-- Single laboratory accuracy and precision
data from both fortified reagent water and fortified matrices using hot, splitless injection
are presented in Tables 4 and 5. Table 6 includes data from 2 matrices using temperature
programmed splitless injection. Method detection limits (MDLs) are presented in Table 1
for both types of injectors used. MDLs were calculated using the formula in Section
9.2.4. Although the calculated MDLs using the two different types are not dramatically
different, for compounds 9-12 (Table 2) the peak shapes are significantly better using
temperature programmed injection, and the peak heights and areas are greater.
13.2 POTENTIAL PROBLEM COMPOUNDS ?br>
13.2.1 2,4-Dinitrophenol, 4-nitrophenol, 2-methyl-4,6-dinitrophenol, pentachlorophenol
and 2,4,6-tribromophenol have a tendency to exhibit a chromatographic
phenomenon known as "matrix-induced chromatographic response
enhancement" (5-8). Compounds that exhibit this phenomenon often give
analytical results that exceed 100% recovery. The theory behind this
phenomenon is that these compounds are susceptible to adsorption and/or
thermal degradation in the GC inlet. The "cleaner" the matrix they are injected
in, e.g. clean solvent, the more they degrade. When they are injected in a sample
extract, matrix components in the sample extract "protect" these compounds
from decomposition and a relatively greater response is observed. While most
of the literature references to this phenomenon refer to organophosphate
pesticides in river water and food samples, the effect seen during development of
this method suggests the same type of problem occurs with these acidic phenols.
This method uses 2,3,4,5-tetrachlorophenol as the internal standard for
quantifying these analytes. The chromatographic behavior of 2,3,4,5-
tetrachlorophenol mimics these particular method analytes. Therefore its use as
an internal standard helps maintain accurate measurement of these analytes. It
should be noted however that these particular analytes will probably not be
measured with the same level of precision and accuracy as other method
analytes, but the precision and accuracy requirements should still be achievable.
13.2.2 The same compounds listed in sect. 13.2.1. also have a tendency to tail. QC
criteria for peak tailing factors have been given in Sect. 10.2.3.1. During method
development, significantly less peak tailing was observed using temperature
programmed injection. Other measures shown to minimize peak tailing and
improve peak shape are pressure pulsed injection, and increasing the GC oven
temperature program rate. Pulsed injection is recommended on GCs which have
that option available. A faster GC oven temperature program is recommended if
there are no interferences, and if the minimum number of scans across all
chromatographic peaks can be obtained. This is a function of how fast the MS
13.2.3 Phenol is very water soluble compared to other method analytes. Breakthrough
experiments performed during method development indicate that some
breakthrough from the SPE cartridge can be expected. Breakthrough can be
minimized by monitoring the flow of the sample through the cartridge. In general,
slower flow rates will minimize breakthrough. Precision and accuracy
requirements in Sect. 10 should be achievable.
13.3 HOLDING TIME STUDY RESULTS ?br>
13.3.1 Holding time studies for aqueous samples were conducted for a period of 35
days. Chlorinated surface water samples fortified with method analytes and
preserved and stored according to requirements in Section 8, were analyzed on
days 0, 7, 10, 15, 23, 28, and 35. Small, but statistically significant losses of 2-
chlorophenol, o-cresol, and 2,4-dimethylphenol were observed beginning
between day 15 and 23. Therefore the aqueous holding time was determined to
be 14 days. Data from these studies are in Table 7.
13.3.2 Holding time studies for sample extracts were conducted for a period of 35
days. A single set of extracts were stored at 0癈, and analyzed on days 0, 14,
23, and 35. No significant losses were observed within this time frame.
Therefore the extract holding time was established at 30 days. Data from these
studies are in Table 8.
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
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.
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" also
available from the American Chemical Society at the address in Section 14.2.
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 2206, (Revised, January 1976).
4. "Safety in Academic Chemistry Laboratories," American Chemical Society Publication,
Committee on Chemical Safety, 3rd Edition, 1979.
5. 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.
6. Mol, H.G.J., M. Althuizen, H. Janssen, and C.A. Cramers, "Environmental Applications of
Large Volume Injection in Capillary GC Using PTV Injectors," J. High Resol. Chromatogr., 19
7. Erney, D.R., T.M. Pawlowski, C.F. Poole, "Matrix Induced Peak Enhancement of Pesticides in
Gas Chromatography," J. High Resol. Chromatogr., 20 (1997) 375-378.
8. 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.
17. TABLES, DIAGRAMS, FLOWCHARTS, AND VALIDATION DATA
TABLE 1. METHOD DETECTION LIMITS a
Hot Splitless Injection b Temperature Programmed
Splitless Injection c
Spiking MDL Spiking MDL
: : : :
Conc. (:g/L) (:g/L) Conc. (:g/L) (:g/L)
phenol 1.0 0.58 0.1 0.025
2-chlorophenol 0.1 0.020 0.1 0.041
2-methylphenol (o-cresol) 0.1 0.026 0.1 0.028
2-nitrophenol 0.1 0.026 0.1 0.044
2,4-dimethylphenol 0.1 0.026 0.1 0.034
2,4-dichlorophenol 0.1 0.027 0.1 0.046
4-chloro-3-methylphenol 0.1 0.036 0.1 0.042
2,4,6-trichlorophenol 0.1 0.046 0.1 0.024
2,4-dinitrophenol 1.0 0.31 0.5 0.22
4-nitrophenol 1.0 0.42 0.5 0.18
2-methyl-4,6-dinitrophenol 1.0 0.26 0.5 0.092
pentachlorophenol 1.0 0.25 0.5 0.081
a- data obtained using Column 1
TABLE 2. RETENTION TIMES (RTs) AND SUGGESTED QUANTITATION IONS (QIs)
Cmpd Analyte RTs (min) RTs (min) QIs IS
#a (m/z) Ref
column 1 b column 2 c
1 phenol 11:00 12:38 94 1
2 2-chlorophenol 11:07 12:50 128 1
3 2-methylphenol (o-cresol) 12:52 14:27 107 1
4 2-nitrophenol 14:36 16:22 139 1
5 2,4-dimethylphenol 15:00 16:35 107 1
6 2,4-dichlorophenol 15:22 17:04 162 1
7 4-chloro-3-methylphenol 17:46 19:27 142 1
8 2,4,6-trichlorophenol 18:56 20:42 97 1
9 2,4-dinitrophenol 21:30 23:30 2
10 4-nitrophenol 21:53 23:44 139 2
11 2-methyl-4,6-dinitrophenol 23:09 25:09 2
12 pentachlorophenol 25:14 27:12 266 2
13 1,2-dimethyl-3-nitrobenzene (IS#1) 17:43 19:29 134
14 2,3,4,5-tetrachlorophenol (IS#2) 22:09 24:02 232
15 2-chlorophenol-3,4,5,6-d4 (SURR) 11:04 12:47 132 1
16 2,4-dimethylphenol-3,5,6-d3 (SURR) 14:59 16:34 110 1
17 2,4,6-tribromophenol (SURR) 23:37 25:37 330 2
a- Number refers to peak number in Figures 1-3.
b- Column 1- 30 m ?0.25 mm id DB-5ms (J&W), 0.25 :m film thickness.
c- Column 2- 30 m ?0.25mm id BPX5 (SGE), 0.25 :m film thickness.
d- Because the MS response to these compounds is low, and both the listed ions are near 100% ion
abundance, the signal from both ions may be added together to increase sensitivity.
TABLE 3. ION ABUNDANCE CRITERIA FOR DECAFLUOROTRIPHENYLPHOSPHINE (DFTPP)
Purpose of Checkpoint a
Mass (m/z) Relative Abundance Criteria
51 10-80% of the base peak low mass sensitivity
68 <2% of mass 69 low mass resolution
70 <2% of mass 69 low mass resolution
127 10-80% of the base peak low-mid mass sensitivity
197 <2% of mass 198 mid-mass resolution
198 base peak or >50% of 442 mid-mass resolution and sensitivity
199 5-9% of mass 198 mid-mass resolution and isotope ratio
275 10-60% of the base peak mid-high mass sensitivity
365 >1% of the base peak baseline threshold
441 Present and < mass 443 high mass resolution
442 base peak or >50% of 198 high mass resolution and sensitivity
443 15-24% of mass 442 high mass resolution and isotope ratio
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 three resolution checks, which include natural abundance isotope ratios, constitute the next most important
part of the performance test. The correct setting of the baseline threshold, as indicated by the presence of low intensity ions, is the next most
important part of the performance test. Finally, the ion abundance ranges are designed to encourage some standardization to fragmentation
TABLE 4. RESULTS FROM REPLICATE ANALYSES OF REAGENT WATER FORTIFIED WITH METHOD ANALYTES
AT 1-10 : g/L a USING HOT SPLITLESS INJECTION
Concentration= 10 : g/L b, n=4 Concentration= 5 : g/L c , n=4 Concentration= 1 : g/L b, n=7
Mean % RSD (%) Mean % RSD (%) Mean % RSD (%)
Recovery Recovery Recovery
phenol 75.9 4.3 103 18 90.8 20
2-chlorophenol 91.3 2.6 86.7 4.3 89.7 7.9
2-methylphenol (o-cresol) 93.3 2.2 86.1 3.6 84.9 7.3
2-nitrophenol 97.4 3.6 97.8 4.8 75.0 11
2,4-dimethylphenol 91.0 1.7 74.7 8.5 76.3 8.0
2,4-dichlorophenol 94.3 2.9 94.4 1.8 88.2 4.8
4-chloro-3-methylphenol 96.5 1.9 92.3 4.2 90.8 8.4
2,4,6-trichlorophenol 89.9 3.4 91.5 4.7 92.9 11
2,4-dinitrophenol 107 3.6 104 6.0 61.8 16
4-nitrophenol 107 3.0 90.5 11 96.3 14
2-methyl-4,6-dinitrophenol 105 3.2 83.0 1.6 91.5 8.9
pentachlorophenol 103 1.9 86.2 9.8 103 7.6
2-chlorophenol-3,4,5,6-d4 (SURR) 93.4 1.8 84.9 6.6 95.9 11
2,4-dimethylphenol-3,5,6-d3 (SURR) 94.0 2.0 82.0 7.2 95.1 13
2,4,6-tribromophenol (SURR) 82.2 1.8 77.1 6.1 98.0 8.6
a- Surrogate concentrations in all sample are 5 :g/L for tribromophenol and 2 :g/L for the deuterated phenols.
b- Data obtained using Column 1.
c- Data obtained using Column 2
TABLE 5. ACCURACY a AND PRECISION DATA FOR METHOD ANALYTES FORTIFIED AT 10 : g/L IN THREE MATRICES
HOT SPLITLESS INJECTION
ANALYTE HARD GROUND WATER CHLORINATED SURFACE SIMULATED HIGH TOC WATER
WATER c n=4 c, d
% Rec RSD (%) %Rec RSD (%) %Rec RSD (%)
phenol 77.6 5.1 73.9 4.5 73.7 5.1
2-chlorophenol 91.2 2.6 88.6 3.0 85.8 5.2
2-methylphenol (o-cresol) 93.2 3.4 91.3 2.3 89.2 4.3
2-nitrophenol 102 2.5 99.4 2.1 99.0 2.8
2,4-dimethylphenol 86.3 2.0 86.6 1.9 83.2 5.3
2,4-dichlorophenol 94.8 1.2 93.3 1.4 90.0 5.1
4-chloro-3-methylphenol 98.5 1.6 97.5 1.4 94.6 3.9
2,4,6-trichlorophenol 95.5 3.9 92.7 3.1 89.1 4.1
2,4-dinitrophenol 117 4.2 118 2.3 121 1.0
4-nitrophenol 102 2.4 102 3.8 95.9 2.3
2-methyl-4,6-dinitrophenol 115 2.2 113 1.5 115 0.71
pentachlorophenol 110 4.0 108 5.1 105 1.8
2-chlorophenol-3,4,5,6-d4 (SURR) 93.0 2.4 91.5 1.6 89.1 5.3
2,4-dimethylphenol-3,5,6-d3 (SURR) 88.9 3.0 87.2 2.4 85.4 4.8
2,4,6-tribromophenol (SURR) 74.7 2.6 76.3 5.1 71.0 4.5
a- Accuracy is presented as % recovery
b- Hard municipal chlorinated ground water, 450mg/L hardness measured as calcium carbonate.
c- Data obtained using Column 1.
d- Simulated high organic content (high Total Organic Carbon) sample prepared by adding 10 mg/L humic acid (Fluka Chemical Corp., Milwaukee, WI) to reagent
TABLE 6. RESULTS OF REPLICATE ANALYSES IN TWO MATRICES USING TEMPERATURE PROGRAMMABLE
ANALYTE Reagent Water Reagent Water Chlorinated Surface Water
Concentration= 0.5 : g/L, n=7 Concentration= 5 : g/L, n=4 Concentration= 10 : g/L, n=5
% Recovery RSD (%) % Recovery RSD (%) % Recovery RSD (%)
phenol 74.5 13 107 21 74.1 7.5
2-chlorophenol 88.3 11 95.5 7.7 94.8 4.3
2-methylphenol (o-cresol) 96.2 9.4 93.0 4.5 103 8.1
2-nitrophenol 89.1 8.9 102 6.8 94.5 7.4
2,4-dimethylphenol 82.0 8.2 85.5 8.8 93.8 9.8
2,4-dichlorophenol 83.0 8.3 100 7.3 93.7 2.9
4-chloro-3-methylphenol 79.0 6.6 101 7.5 96.6 2.9
2,4,6-trichlorophenol 78.9 8.1 102 10 90.7 3.3
2,4-dinitrophenol 63.8 22 95.4 6.6 113 4.7
4-nitrophenol 84.1 14 103 8.1 102 6.8
2-methyl-4,6-dinitrophenol 62.4 9.4 103 5.1 107 4.9
pentachlorophenol 72.0 7.2 88.0 7.2 93.8 6.8
2-chlorophenol-3,4,5,6-d4 (SURR) 79.1 7.6 87.6 3.9 84.7 10
2,4-dimethylphenol-3,5,6-d3 (SURR) 79.9 8.5 91.9 7.8 97.0 8.3
2,4,6-tribromophenol (SURR) 79.6 7.8 86.1 7.8 79.5 12
a- Data obtained using Column 1.
TABLE 7. RESULTS OF AQUEOUS HOLDING TIME STUDIES FOR METHOD 528 ANALYTES a
ANALYTE DAY 0 DAY 7 DAY 10 DAY 15
% Rec RSD (%) % Rec RSD (%) % Rec RSD (%) % Rec RSD (%)
phenol 74.5 1.9 77.0 3.8 74.3 3.4 74.3 3.5
2-chlorophenol 91.5 1.9 88.8 1.3 88.0 2.9 85.8 4.1
2-methylphenol (o-cresol) 99.9 2.8 96.0 1.2 94.3 2.6 93.0 2.8
2-nitrophenol 99.1 4.3 100 4.0 98.8 3.9 101 3.1
2,4-dimethylphenol 85.5 4.2 80.7 3.9 79.1 8.1 75.2 4.7
2,4-dichlorophenol 94.2 2.2 93.0 2.5 92.3 1.0 91.8 3.3
4-chloro-3-methylphenol 96.5 2.1 95.5 3.5 96.5 2.3 96.5 3.9
2,4,6-trichlorophenol 94.8 3.3 97.1 2.9 96.0 2.8 95.0 3.3
2,4-dinitrophenol 102.8 5.1 102 3.4 107 8.0 112 2.9
4-nitrophenol 100.1 3.2 100 4.2 101 4.9 96.2 3.3
2-methyl-4,6-dinitrophenol 96.1 5.1 109 8.7 113 5.8 115 5.0
pentachlorophenol 94.5 4.1 102 2.1 103 3.5 102 2.3
a- All analytes fortified into a chlorinated surface water at a concentration of 10 :g/L, dechlorinated and acidified according to Section 8, stored
for 48 hr at 10?C, followed by storage at 6?C. For each time point, n=5.
TABLE 8. RESULTS OF EXTRACT HOLDING TIME STUDIES FOR METHOD 528 ANALYTES a
ANALYTE DAY 0 DAY 14 DAY 23 DAY 35
% Rec RSD (%) % Rec RSD (%) % Rec RSD (%) % Rec RSD (%)
phenol 74.5 1.9 74.2 3.8 72.7 3.4 74.9 3.5
2-chlorophenol 91.5 1.9 89.8 1.3 88.6 2.9 89.1 4.1
2-methylphenol (o-cresol) 99.9 2.8 97.5 1.2 98.4 2.6 96.6 2.8
2-nitrophenol 99.1 4.3 100 4.0 96.4 3.9 101 3.1
2,4-dimethylphenol 85.5 4.2 86.9 3.9 88.0 8.1 88.7 4.7
2,4-dichlorophenol 94.2 2.2 95.4 2.5 96.1 1.0 98.6 3.3
4-chloro-3-methylphenol 96.5 2.1 98.9 3.5 98.1 2.3 102 3.9
2,4,6-trichlorophenol 94.8 3.3 97.1 2.9 97.6 2.8 102 3.3
2,4-dinitrophenol 102.8 5.1 104 3.4 90.9 8.0 99.1 2.9
4-nitrophenol 100.1 3.2 91.1 4.2 89.3 4.9 96.3 3.3
2-methyl-4,6-dinitrophenol 96.1 5.1 108 8.7 98.2 5.8 110 5.0
pentachlorophenol 94.5 4.1 99.1 2.1 100 3.5 105 2.3
a- All extracts were from the Day 0 aqueous holding time samples, and were stored in amber vials at 0?C. For each time point, n=5.
TABLE 9. INITIAL DEMONSTRATION OF CAPABILITY (IDC) REQUIREMENTS
Method Requirement Specification and Frequency Acceptance Criteria
Sect. 9.2.1 Initial Demonstration of Low Analyze LRB prior to any other IDC Demonstrate that all target analytes are below
Method Background steps. 1/3 the MRL, and that possible interferences
from extraction media do not prevent the
identification and quantification of method
RSD must be #20% for all analytes except
Sect. 9.2.2 Initial Demonstration of Analyze 4-7 replicate LFBs fortified
phenol which must be # 30%.
Precision (IDP) at 5-10:g/L
Sect. 9.2.3 Initial Demonstration of Calculate average recovery for Mean recovery 70-130% of true value, except
Accuracy (IDA) replicates used in IDP phenol which is 50-150%
Sect. 9.2.4 Method Detection Limit Over a period of three days, Note: Data from MDL replicates are not
(MDL) Determination prepare a minimum of 7 replicate required to meet method precision and
LFBs fortified at a concentration accuracy criteria. If the MDL replicates are
estimated to be near the MDL. fortified at a low enough concentration, it is
Analyze the replicates through all likely that they will not meet precision and
steps of the analysis. Calculate the accuracy criteria.
MDL using the equation in Section
TABLE 10. QUALITY CONTROL REQUIREMENTS (SUMMARY)
Method Requirement Specification and Frequency Acceptance Criteria
Sect. 8.4 Sample Holding Time 14 days, dechlorinated and acidified Iced or refrigerated at 10癈 or less for up to
to pH#2 48 hours, 6癈 thereafter.
Sect. 8.4 Extract Holding Time 30 days Stored at 0癈 or less in amber vials.
Sect.9.11 Laboratory Fortified Sample Analyze one LFM per extraction Recoveries not within 70-130% of the fortified
Matrix (LFM) batch (20 samples or less) fortified amount may indicate a matrix effect.
with method analytes at a
concentration close to the native
Sect. 9.12 Field Duplicates Analyze 1 FD for each 20 samples, Suggested RPD?0%.
or 1 per extraction batch, whichever
Sect. 9.13 Quality Control Sample Analyze QCS whenever new If analyzed as a calibration sample, CCC
(QCS) standards are prepared, or at least criteria apply. If analyzed as a LFB, those
quarterly. criteria apply.
Sect 9.2.1 Laboratory Reagent Blank Daily, or with each extraction batch Demonstrate that all target analytes are below
(LRB) of up to 20 samples, whichever is 1/3 the MRL, and that possible interference
more frequent. from extraction media do not prevent the
identification and quantification of method
Sect. 9.8 Laboratory Fortified Blanks Analyze at least one LFB daily or for Results of LFB analyses must be 70-130% of
(LFB) each extraction batch of up to 20 the true value (except phenol) for each analyte
field samples. Rotate the fortified and surrogate for all fortified concentrations
concentration between low, medium greater than the lowest CAL point. Results of
and high amounts. LFBs corresponding to the lowest CAL point
must be 50-150% of the true value.
Sect. 9.9 Internal Standard 1,2-dimethyl-3-nitrobenzene (IS#1) Peak area counts for IS#1 in LFBs, LRBs and
and 2,3,4,5-tetrachloro-phenol sample extracts must be within 70-130% of the
(IS#2) are added to all standards peak area in the most recent CCC, and 50-
and sample extracts. 150% of average area in the initial calibration.
Sect 9.10 Surrogate Standards Surrogate standards are added to all Recovery for 2-chlorophenol-3,4,5,6-d4 and
standards, samples, LFBs, LFMs, 2,4-dimethylphenol-3,5,6-d3 in all standards,
FDs, LRBs, and LFBs. LRB, LFB, LFM, FD and sample extracts
must be 70-130% of the true value. Recovery
for 2,4,6-tribromophenol must be 60-130%.
Sect. 10.2.1 MS Tune Check Analyze DFTPP to verify MS tune Criteria are given in Table 3.
before initial calibration and before
Sect.10.2.2 Initial Calibration Use internal standard calibration When each calibration standard is calculated as
technique to generate an average RF an unknown using the calibration curve, the
or first or second order calibration result must be 70-130% of the true value for all
curve. Use at least 5 standard but the lowest standard. The lowest standard
concentrations that span the must be 50-150% of the true value.
approximate range of 0.1- 15 :g/L.
Sect. 10.2.3.1 GC Performance-Peak Tailing Calculate the peak tailing factor for Peak tailing factor of 5 or less. (See Fig.4 for
Check compounds listed in the referenced calculation of PTF.)
section, at the beginning of each day
during which samples are analyzed.
In cases of continuous instrument
operation, check peak tailing factors
every 24 hr.
Sect. 10.3 Continuing Calibration Check Verify initial calibration by analyzing a The result for each analyte and surrogate must
calibration standard prior to analyzing be 70-130% of the true value for all
samples, after every 10 samples, and concentrations except the lowest CAL point
after the last sample. Always analyze for each analyte. The lowest CAL point for
a low concentration (near the MRL) each analyte must be 50-150% of the true
CCC at the beginning of the analysis value.
period. The peak area of IS#1 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.
FIGURE 1.CHROMATOGRAM OF PHENOL STANDARD MIX (5 ng/ : L EACH ANALYTE) ON A DB-5ms COLUMN WITH
HOT SPLITLESS INJECTION a.
numbers refer to compounds as listed in Table 2.
FIGURE 2.CHROMATOGRAM OF PHENOL STANDARD MIX (5 ng/ : L EACH ANALYTE) ON A BPX5 COLUMN WITH
HOT SPLITLESS INJECTION. a
a- peak numbers refer to compounds as listed in Table 2..
CHROMATOGRAM OF PHENOL STANDARD MIX (5 ng/ : L EACH ANALYTE) ON A DB-5ms COLUMN
WITH TEMPERATURE PROGRAMMED SPLITLESS INJECTION. a
numbers refer to compounds as listed in Table 2.
FIGURE 4. PEAK TAILING FACTOR (PTF) CALCULATION.
Peak Tailing Factor =
BD = 10 % peak height
Note: the PTF should be calculated from the single ion chromatogram of the quantitation ion.
This example is for the pentachlorophenol peak. The PTF = 2.5.