Jimmie W. Hodgeson J. Collins (Technology Applications Inc.) R.E. Barth (Technology Applications Inc.)
Jimmie W. Hodgeson J. Collins (Technology Applications Inc.) R.E. Barth (Technology Applications Inc.)
July 1990
Jimmie W. Hodgeson
552-1
METHOD 552
1.2 This method is applicable to the determination of these analytes over the
concentration ranges typically found in drinking water1,2,4, subject to the
method detection limits (MDL) listed in Table 2. The detection limits observed
may vary according to the particular matrix analyzed and the specific
instrumentation employed. The haloacetic acids are observed ubiquitously in
chlorinated supplies at concentrations normally within the spiking level ranges
in Tables 2-5.
1.3 Tribromoacetic acid has not been included because of problems with extraction
and chromatography by this method. The mixed bromochloroacetic acids have
recently been synthesized. The bromochloroacetic acid is present in
chlorinated supplies and method validation data are provided herein.
However, neat material for this compound is not readily available. The mixed
trihalogenated acids may also be present. These are not included because of
current problems with sample purity and the chromatography for these two
compounds.
1.4 The 2-chlorophenol has not been included as a method analyte in the above
list, primarily because its realistic detection limit in environmental samples is
likely to be above the odor threshold. Poor precision is usually obtained for
this compound at even higher levels. In addition, this analyte displays
552-2
instability under the dechlorination/preservation conditions described herein.
Nevertheless, some method validation data are given in Tables 2-7.
1.5 This method is designed for analysts skilled in liquid-liquid extractions, extract
concentration techniques, derivatization procedures and the use of GC and
interpretation of gas chromatograms.
1.6 When this method is used for the analyses of waters from unfamiliar sources,
analyte identifications must be confirmed by at least one additional qualitative
technique, such as GC/mass spectroscopy (MS) or by GC using dissimilar
columns.
2.1 A 100 mL volume of sample is adjusted to pH 11.5 and extracted with methyl-
tert-butyl ether (MTBE) to remove neutral and basic organic compounds. The
aqueous sample is then acidified to pH 0.5 and the acids are extracted into
MTBE. After the extract is dried and concentrated, the acids are converted to
their methyl esters with diazomethane (DAM). Excess DAM is removed and
the methyl esters are determined by capillary GC using an electron capture
detector (ECD). An alternative microextraction procedure is also offered in
which a 30 mL sample is extracted without cleanup with a single 3 mL aliquot
of MTBE for direct analysis by GC-ECD after methylation. Samples containing
high concentrations of haloacetic acids and other disinfection byproducts, or
other potentially interfering organic compounds, may require the sample
cleanup.
3.0 DEFINITIONS
3.3 Laboratory Duplicates (LD1 and LD2) -- Two sample aliquots taken in the
analytical laboratory and analyzed separately with identical procedures.
Analyses of LD1 and LD2 give a measure of the precision associated with
laboratory procedures, but not with sample collection, preservation, or storage
procedures.
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3.4 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.
3.5 Laboratory Reagent Blank (LRB) -- An aliquot of reagent water that is treated
exactly as a sample including exposure to all glassware, equipment, solvents,
reagents, internal standards, and surrogates that are used with other samples.
The LRB is used to determine if method analytes or other interferences are
present in the labora-tory environment, the reagents, or the apparatus.
3.6 Field Reagent Blank (FRB) -- Reagent water placed in a sample container in the
laboratory and treated as a sample in all respects, including exposure to
sampling site conditions, storage, preservation and all analytical procedures.
The purpose of the FRB is to determine if method analytes or other
interferences are present in the field environment.
3.11 Calibration Standard (CAL) -- A solution prepared from the primary dilution
standard solution and stock standard solutions of the internal standards and
surrogate analytes. The CAL solutions are used to calibrate the instrument
response with respect to analyte concentration.
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3.12 Quality Control Sample (QCS) -- A sample matrix containing method analytes
or a solution of method analytes in a water miscible solvent which is used to
fortify reagent water or environmental samples. The QCS is obtained from a
source external to the laboratory, and is used to check laboratory performance
with externally prepared test materials.
4.0 INTERFERENCES
4.1.2 The use of high purity reagents and solvents helps to minimize
interference problems. Purification of solvents by distillation in all-
glass systems may be required. The extraction solvent, MTBE, may
need to be redistilled.
4.3 The acid forms of the analytes are strong organic acids which react readily
with alkaline substances and can be lost during sample preparation.
Glassware and glass wool must be acid-rinsed with (1+9) hydrochloric acid,
552-5
and the sodium sulfate must be acidified (see Section 7.6) with sulfuric acid
prior to use to avoid analyte losses due to adsorption.
4.4 Organic acids and phenols, especially chlorinated compounds, cause the most
direct interference with the determination. The addition of base and
subsequent extraction of the basic sample removes many neutral and basic
chlorinated hydrocarbons and phthalate esters that might otherwise interfere
with the electron capture analysis.
4.6 Matrix interferences may be caused by contaminants that are coextracted from
the sample. The extent of matrix interferences will vary considerably from
source to source, depending upon the water sampled. Positive identifications
should be confirmed using the confirmation column specified in Table 1 or by
the use of gas chromatography with mass spectrometric detection.
5.0 SAFETY
5.1 The toxicity or carcinogenicity of each reagent used in this method has not
been precisely defined; however, each chemical compound must be treated as
a potential health hazard. From this viewpoint, exposure to these chemicals
must be reduced to the lowest possible level by whatever means available.
The laboratory is responsible for maintaining a current awareness file of OSHA
regulations regarding the safe handling of the chemicals specified in this
method. A reference file of material data handling sheets should also be made
available to all personnel involved in the chemical analysis. Additional
references to laboratory safety are available and have been identified6-8 for the
information of the analyst.
5.2 Diazomethane is a toxic carcinogen and can explode under certain conditions,
when produced in a purified or highly concentrated form. In this form, the
following safety precautions must be followed.
5.2.2 Use a safety screen. Wear protective clothing and a shielded safety
hood.
552-6
5.2.5 Avoid grinding surfaces, ground glass joints, sleeve bearings, glass
stirrers.
5.3 For the above reasons, the diazomethane generation apparatus used in the
esterification procedure specified in this method3 produces only micromolar
amounts of diazomethane in very dilute solution (Section 11.3) to minimize
safety hazards. In this form, the solution is not explosive. Nevertheless, the
following precautions should be followed.
5.3.2 When handling the diazomethane solution, avoid contact with skin. If
contact is made, immediately wash the exposed area with warm water.
5.4 The toxicity of the extraction solvent, MTBE, has not been well defined.
Susceptible individuals may experience adverse affects upon skin contact or
inhalation of vapors. For such individuals a mask may be required. Protective
clothing and gloves should be used and MTBE should be used only in a
chemical fume hood or glove box.
6.0 APPARATUS AND EQUIPMENT (All specifications in Sections 6.0 and 7.0 are
suggested. Catalog numbers are provided for illustration only.)
6.1 Separatory Funnels -- 250 mL, with TFE fluorocarbon stopcocks, ground glass
or TFE fluorocarbon stoppers.
6.2 Screw Cap -- 40 mL vials (Pierce #13219 or equivalent). Screw caps should
have TFE fluorocarbon liners.
552-7
6.6 Gas Chromatograph -- Analytical system complete with gas chromatograph
equipped for electron capture detection, split/splitless capillary injection,
temperature programming, differential flow control, and with all required
accessories including syringes, analytical columns, gases and strip-chart
recorder. A data system is recommended for measuring peak areas. An
autoinjector is recommended for improved precision of analyses. The gases
flowing through the election capture detector should be vented through the
laboratory fume hood system.
6.7 Vials -- Amber glass, 7-10 mL capacity with TFE-fluorocarbon lined screw cap.
6.10 Pasteur Pipets -- Glass disposable, 5¾" length wide bore diameter. (Baxter
Scientific Products Giant-Pette-Pipets, Cat. No. P5240-1 or equivalent)
7.1 Glass Wool -- Acid washed, Heat to 400°C for one hour.
7.4 Ethyl Ether -- Nanograde, redistilled in glass if necessary. Ethers must be free
of peroxides as indicated by EM Quant test strips, available from EM Science,
Gibbstown, NJ. Procedures recommended for removal of peroxides are
provided with the test strips. Ethers must be periodically tested (monthly) for
peroxide formation during use.
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7.5 Methyl-Tert-Butyl Ether -- Nanograde, redistilled in glass if necessary. The
same peroxide precautions as in Section 7.4 apply to this ether.
7.6 Sodium Sulfate -- ACS, granular, acidified, anhydrous. Heat in a shallow tray
at 400°C for a minimum of four hours to remove phthalates and other
interfering organic substances. Alternatively, extract with methylene chloride
in a Soxhlet apparatus for 48 hours. Acidify by slurrying 100 g sodium sulfate
with just enough ethyl ether to cover the solid. Add 0.1 mL concentrated
sulfuric acid and mix thoroughly. Remove the ether under vacuum or allow to
evaporate in a loosely covered beaker in a hood. Mix 1 g of the resulting solid
with 5 mL of reagent water and measure the pH of the mixture. It must be
below pH 4. Store at 130°C.
7.7 Sulfuric Acid Solution (1+1) -- Slowly add 50 mL H2SO4 (sp. gr. 1.84) to 50 mL
of reagent water.
7.10 Carbitol -- (Diethylene glycol monoethyl ether), ACS. Available from Aldrich
Chemical Co.
7.13 Silica Gel -- Chromatographic grade, nominal 100 mesh. Heat to 400°C for
four hours. Store at 130°C.
552-9
8.0 SAMPLE COLLECTION, PRESERVATION AND STORAGE
8.1.2 After collecting the sample in the bottle containing the dechlorination
reagent, seal the bottle and agitate for one minute.
9.0 CALIBRATION
9.2 Internal Standard Calibration Procedure -- This approach requires the analyst
to select one or more internal standards which are compatible in analytical
behavior with the method analytes. For the single laboratory precision and
accuracy data reported in Tables 2-7, one internal standard
(1,2,3-trichloropropane) was employed. The concentration of the internal
standard used in obtaining these data was 0.4 µg/mL in the final 5.0 mL
concentrate (Section 11.3.3).
9.2.1 Prepare separate stock standard solutions for each compound of interest
at a concentration of 1-5 mg/mL in MTBE solvent. Method analytes
may be obtained as neat materials or ampulized solutions (>99% purity)
from a number of commercial suppliers.
552-10
9.2.2 Prepare primary dilution standard solutions by combining and diluting
stock calibration standards with MTBE. The primary dilution standards
are used to prepare calibration standards, which comprise at least three
concentration levels (optimally five) of each analyte with the lowest
standard being at or near the method detection limit of each analyte.
The concentrations of the other standards should define a range
containing the expected sample concentrations or the working range of
the detector.
9.2.4 Inject 2 µL of each standard and calculate the relative response for each
analyte (RRa) using the equation:
RRa = Aa /Ais
552-11
check must be repeated using a freshly prepared calibration standard.
Should the retest fail, a new calibration curve must be generated.
10.2 Laboratory Reagent Blanks (LRB) -- Before processing any samples, the analyst
must analyze at least one LRB to demonstrate that all glassware and reagent
interferences are under control. In addition, each time a set of samples is
extracted or reagents are changed, a LRB must be analyzed. If within the
retention time window (Section 11.4.4) 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.
10.3.2 Calculate the mean percent recovery (R) and the standard deviation of
the recovery (SR). For each analyte, the mean recovery values for all
must fall in the range of R ±30% (or within R ±3SR if broader) using the
values for R and SR for reagent water in Table 4. The standard
deviation should be less than ±30% or 3SR, whichever is larger. For
those compounds that meet these criteria, performance is considered
acceptable and sample analysis may begin. For those compounds that
fail these criteria, this procedure must be repeated using a minimum of
five fresh samples until satisfactory performance has been
demonstrated.
552-12
10.3.4 The analyst is permitted to modify GC columns, GC conditions,
detectors, extraction techniques, concentration techniques (i.e.,
evaporation techniques), internal standard or surrogate compounds.
Each time such method modifications are made, the analyst must repeat
the procedures in Section 10.3.1.
10.4.2 If the extract reanalysis fails the 70-130% recovery criterion, the problem
must be identified and corrected before continuing.
10.4.3 If the extract reanalysis meets the surrogate recovery criterion, report
only data for the analyzed extract. If sample extract continues to fail
the recovery criterion, report all data for that sample as suspect.
10.5.1 When using the internal standard calibration procedure, the analyst is
expected to monitor the IS response (peak area or peak height) of all
samples during each analysis day. The IS response for any sample
chromatogram should not deviate from daily calibration standard's IS
response by more than 30%.
552-13
10.5.3 If consecutive samples fail the IS response acceptance criterion,
immediately analyze a calibration check standard.
10.6.1 The laboratory must analyze at least one LFB sample with every
20 samples or one per sample set (all samples extracted within a
24-hour period), whichever is greater. Fortified concentrations near
Level 3 (Table 4) are recommended. Calculate accuracy as percent
recovery (R). If the recovery of any analyte falls outside the control
limits (see Section 10.6.2), that analyte is judged out of control, and the
source of the problem should be identified and resolved before
continuing analyses.
10.6.2 Prepare control charts based on mean upper and lower control limits,
R ±3 SR. The initial demonstration of capability (Section 10.3)
establishes the initial limits. After each four to six new recovery
measurements, recalculate R and SR using all the data, and construct
new control limits. When the total number of data points reach 20,
update the control limits by calculating R and SR using only the most
recent 20 data points. At least quarterly, replicates of LFBs should be
analyzed to determine the precision of the laboratory measurements.
Add these results to the ongoing control charts to document data
quality.
10.7.2 Calculate the mean percent recovery, R, of the concentration for each
analyte, after correcting the total mean measured concentration, A, from
the fortified sample for the back-ground concentration, B, measured in
the unfortified sample, i.e.:
552-14
R = 100 (A - B) / C,
R = 100 (A - B)/C
R* ±3Sc,
or Sc = (Sa 2 + Sb 2 )1/2 ,
552-15
10.7.5 If the recovery of any such analyte falls outside the designated range,
and the laboratory performance for that analyte is shown to be in
control (Section 10.6), the recovery 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.
10.8 Quality Control Sample (QCS) -- At least quarterly, analyze a QCS from an
external source. If measured analyte concentrations are not of acceptable
accuracy, check the entire analytical procedure to locate and correct the
problem source.
10.9 The laboratory may adapt additional quality control practices for use with this
method. The specific practices that are most productive depend upon the
needs of the laboratory and the nature of the samples. For example, field or
laboratory duplicates may be analyzed to assess the precision of the
environmental measurements or field reagent blanks may be used to assess
contamination of samples under site conditions, transportation and storage.
11.0 PROCEDURE
11.1.1 Remove the samples from storage (Section 8.1.3) and allow to
equilibrate to room temperature.
552-16
from the water phase for a minimum of 10 minutes. If the emulsion
interface between layers is more than one-third the volume of the
solvent layer, the analyst must employ mechanical techniques to
complete the phase separation. The optimum technique depends upon
the sample, but may include stirring, filtration of the emulsion through
glass wool, centrifugation, or other physical methods. Discard the
organic phase and return the aqueous phase to the 250 mL separatory
funnel.
11.1.5 Add sufficient 1:1 H2SO4 in reagent water (ca. 15-20 mL) to adjust the
pH to pH ≤0.5. Add 15 mL of MTBE and extract for two minutes as in
Section 11.1.3. Allow the phases to separate for 10 minutes. If an
emulsion persists employ the same procedures for separation as in
Section 11.1.3. Separate the phases and collect the MTBE phase in a
40 mL screw cap vial (Section 6.2). Add 15 mL of MTBE to the sample
and repeat the extraction a second time. Combine the extracts in the
40 mL vial.
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11.1.7.4 Rinse sides of 40 mL sample tube with approximately
0.7 mL of clean MTBE. Transfer this MTBE into the
drying tube using the same pipet as in Step 3.
11.2.1 Remove the samples from storage and allow them to equilibrate to
room temperature.
11.2.4 Add 1.5-3.0 mL concentrated sulfuric acid to lower the pH to less than
0.5. The analyst must verify that the pH is less than 0.5.
11.2.5 Add accurately 3.0 mL methyl tertiary butyl ether (MTBE) using a
pipet.
11.2.7 Cap all vials immediately, and shake by hand to break up clumps.
Vent, recap, and lay vials on their sides until all vials have been
shaken. Clumps of undissolved salt will cause loss of analytes.
11.2.9 Remove vials from shaker and allow to stand for five minutes for phase
separation.
11.2.10 Transfer exactly 2.0 mL of the ether extract (top layer) using a pipet
into a 2.0 mL volumetric flask.
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Be careful to not include any water.
11.3.2 Add a sufficient amount of ethyl ether to Tube 1 to cover the first
impinger. Add 5 mL of MTBE to the collection vial. Set the nitrogen
flow at 5-10 cm3/min. Add 2 mL Diazald solution and 1.5 mL of 37%
KOH solution to the second impinger. Connect the tubing as shown
and allow the N2 flow to purge the diazomethane from the reaction
vessel into the collection vial for 30 minutes. Cap the vial when
collection is complete and maintain at 0-5°C. When stored at 0-5°C this
diazomethane solution may be used over a period of 48 hours.
552-19
11.3.4 Esterification of 30 mL extract (from Section 11.2.11).
11.3.5 Allow the sample from Section 11.3.3.2 or 11.3.4.2 to remain in contact
with dizaoamethane for 30 minutes. Remove any unreacted
diazomethane by addition of 0.2 g silica gel. Effervescence due to
nitrogen evolution is a further indication that excess diazomethane is
present. Mix gently by inverting once.
11.3.6 After a contact time of 15-20 minutes, transfer a portion of the extract
solution to an appropriate vial for injection into the GC. A duplicate
GC vial may be filled from excess sample extract, if desired. Analyze
the samples as soon as possible. Alternatively, the sample extract, after
removal from the silica gel, may be stored for 48 hours at 0-4°C away
from light in glass vials with TFE-lined caps.
11.4.1 Table 1 summarizes the recommended operating conditions for the GC.
Included in Table 1 are the retention times observed using this method.
An example of the separation achieved using these conditions is shown
in Figure 2. Other GC columns, chromatographic conditions, or
detectors may be used if the requirements of Section 10.3 are met.
11.4.2 Calibrate the system daily as described in Section 9.0. The standards
and extracts must be in MTBE.
11.4.3 Inject 2 µL of the sample extract. Record the resulting peak size in area
units.
11.4.4 The width of the retention time window used to make identifications
should be based upon measurements of actual retention time variations
of standards over the course of a day. Three times the standard
deviation of 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 chromatograms.
552-20
11.4.5 If the response for the peak exceeds the working range of the system,
dilute the extract and reanalyze.
12.0 CALCULATIONS
12.1 Calculate analyte concentrations in the sample from the response for the
analyte relative to the internal standard (RRa) using the equation in
Section 9.2.4.
12.2 For samples processed as part of a set where recoveries falls outside of the
control limits established in Section 10.0, results for the affected analytes must
be labeled as suspect.
14.0 REFERENCES
1. Quimby, B.D., Delaney, M.F., Uden. P.C., and Barnes, R.M. Anal. Chem. 52,
1980, pp. 259-263.
2. Uden, P.C. and Miller, J.W. J. Am. Water Works Assoc. 75, 1983, pp. 524-527.
3. Hodgeson, J.W., Cohen, A.L., and Collins, J.D. "Analytical Methods for
Measuring Organic Chlorination Byproducts", Proceedings Water Quality
Technology Conference (WQTC-16), St. Louis, MO, Nov. 13-17, 1988, American
Water Works Association, Denver, CO, pp. 981-1001.
552-21
4. Fair. P.S., Barth, R.C., Flesch, J. J., and Brass, H. "Measurement of Disinfection
Byproducts in Chlorinated Drinking Water," Proceedings Water Quality
Technology Conference (WQTC-15), Baltimore, Maryland, November 15-20,
1987, American Water Works Association, Denver, CO, pp. 339-353.
5. ASTM Annual Book of Standards, Part 31, D3694, "Standard Practice for
Preparation of Sample Containers and for Preservation", American Society for
Testing and Materials, Philadelphia, PA, p. 679, 1980.
7. "OSHA Safety and Health Standards, General Industry", (29 CFR 1910), OSHA
2206, Occupational Safety and Health Administration, Washington, D.C.
Revised January 1976.
9. ASTM Annual Book of Standards, Part 31, D3370, "Standard Practice for
Sampling Water", American Society for Testing and Materials, Philadelphia,
PA, p. 76, 1980.
10. Glaser, J.A., Foerst, D.L., McKee, G.D., Quave, S.A., and Budde, W.L.
Environ. Sci. Technol. 15, 1981, pp. 1426-1435.
11. Chinn, R. and Krasner, S. " A Simplified Technique for the Measurement of
Halogenated Organic Acids in Drinking Water by Electron Capture Gas
Chromatography". Presented at the 28th Pacific Conference on Chemistry and
Spectroscopy, Pasadena, CA, October, 1989.
552-22
TABLE 1. RETENTION DATA AND CHROMATOGRAPHIC CONDITIONS
Retention Time
(min)
Analyte Column A Column B
Monochloroacetic Acid 5.77 10.97
Monobromoacetic Acid 8.70 13.03
Dichloroacetic Acid 9.40 12.72
Trichloroacetic Acid 12.20 14.37
1,2,3-Trichloropropanea 13.28 13.87
Bromochloroacetic Acid 13.52 15.11
Dibromoacetic Acid 16.00 16.83
2-Chlorophenol 16.65 18.32
2,4-Dichlorophenol 20.70 19.27
2,4,6-Trichlorophenol 21.94 22.08
3,5-Dichlorobenzoic Acidb 23.06 23.95
Column A: DB-1701, 30 m x 0.32 mm i.d., 0.25 µm film thickness, Injector Temp. =
200°C, Detector Temp. = 290°C, Helium Linear Velocity = 27 cm/sec,
Splitless injection with 30 second delay
Program: Hold at 50°C for 10 minutes, to 210°C at 10°C/min. and hold 10 minutes.
Column B: DB-210, 30 m x 0.32 mm i.d., 0.50 µm film thickness, Injector Temp. = 200°C,
Detector Temp. = 290°C, Linear Helium Flow = 25 cm/sec, splitless injection
with 30 second delay.
Program: Hold at 50°C for 10 minutes, to 210°< at 10°C/min and hold 10 minutes.
a
Internal Standard.
b
Surrogate Acid.
552-23
TABLE 2. ANALYTE RECOVERY AND PRECISION DATA AND METHOD
DETECTION LIMITSa
552-24
TABLE 3. ANALYTE RECOVERY AND PRECISION DATAa
552-25
TABLE 4. ANALYTE RECOVERY AND PRECISION DATAa
552-26
TABLE 5. ANALYTE RECOVERY AND PRECISION DATAa
Mean Rel.
Fortified Meas. Std. Std. Mean
Conc. Conc. Dev. Dev. Recovery
Analyte (µg/L) (µg/L) (µg/L) (%) (%)
Monochloroacetic Acid 10.0 7.08 0.16 2.3 71
Monobromoacetic Acid 10.0 7.62 0.18 2.4 76
Dichloroacetic Acid 25.0 24.1 0.41 1.7 96
Trichloroacetic Acid 5.00 5.70 0.11 1.9 114
Bromochloroacetic Acid 5.00 4.66 0.22 4.7 93
Dibromoacetic Acid 5.00 5.35 0.096 1.8 107
2-Chlorophenol 12.50 12.7 0.66 5.2 102
2,4-Dichlorophenol 10.00 11.0 0.57 5.2 110
2,4,6-Trichlorophenol 5.00 5.18 0.072 1.4 104
a
Produced by the analysis of seven aliquots of fortified reagent water.
552-27
TABLE 6. ANALYTE RECOVERY AND PRECISION DATAa
552-28
TABLE 7. ANALYTE RECOVERY AND PRECISION DATAa
552-29
FIGURE 2A
FIGURE 2B
552-30
552-31
552-32