November 2003
Environmental Technology
Verification Report
Strategic Diagnostics Inc.
Deltatox®
Rapid Toxicity Testing System
Prepared by
Battel le
Batteiie
Putting Technology To Work
Under a cooperative agreement with
SEPA U.S. Environmental Protection Agency
ETV ElV ElV
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November 2003
Environmental Technology Verification
Report
ETV Advanced Monitoring Systems Center
Strategic Diagnostics Inc.
Deltatox®
Rapid Toxicity Testing System
by
Ryan James
Amy Dindal
Zachary Willenberg
Karen Riggs
Battelle
Columbus, Ohio 43201
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Notice
The U.S. Environmental Protection Agency (EPA), through its Office of Research and
Development, has financially supported and collaborated in the extramural program described
here. This document has been peer reviewed by the Agency. Mention of trade names or
commercial products does not constitute endorsement or recommendation by the EPA for use.
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Foreword
The U.S. Environmental Protection Agency (EPA) is charged by Congress with protecting the
nation's air, water, and land resources. Under a mandate of national environmental laws, the
Agency strives to formulate and implement actions leading to a compatible balance between
human activities and the ability of natural systems to support and nurture life. To meet this
mandate, the EPA's Office of Research and Development provides data and science support that
can be used to solve environmental problems and to build the scientific knowledge base needed
to manage our ecological resources wisely, to understand how pollutants affect our health, and to
prevent or reduce environmental risks.
The Environmental Technology Verification (ETV) Program has been established by the EPA to
verify the performance characteristics of innovative environmental technology across all media
and to report this objective information to permitters, buyers, and users of the technology, thus
substantially accelerating the entrance of new environmental technologies into the marketplace.
Verification organizations oversee and report verification activities based on testing and quality
assurance protocols developed with input from major stakeholders and customer groups
associated with the technology area. ETV consists of seven environmental technology centers.
Information about each of these centers can be found on the Internet at http://www.epa.gov/etv/.
Effective verifications of monitoring technologies are needed to assess environmental quality
and to supply cost and performance data to select the most appropriate technology for that
assessment. Under a cooperative agreement, Battelle has received EPA funding to plan,
coordinate, and conduct such verification tests for "Advanced Monitoring Systems for Air,
Water, and Soil" and report the results to the community at large. Information concerning this
specific environmental technology area can be found on the Internet at
http ://www. ep a. go v/et v/centers/center 1. html.
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Acknowledgments
The authors wish to acknowledge the support of all those who helped plan and conduct the
verification test, analyze the data, and prepare this report. Many thanks go to Battelle's Medical
Research and Evaluation Facility for providing the facilities for and personnel capable of
working with chemical warfare agents and biotoxins. We would also like to thank
Karen Bradham, U.S. EPA National Exposure Research Laboratory; Steve Allgeier, U.S. EPA
Office of Water; Ricardo DeLeon, Metropolitan Water District of Southern California; Yves
Mikol, New York City Department of Environmental Protection; and Stanley States, Pittsburgh
Water and Sewer Authority, for their careful review of the test/QA plan and this verification
report.
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Contents
Page
Notice ii
Foreword iii
Acknowledgments iv
List of Abbreviations viii
1 Background 1
2 Technology Description 2
3 Test Design and Procedures 4
3.1 Introduction 4
3.2 Test Design 5
3.3 Test Samples 6
3.3.1 Quality Control Samples 6
3.3.2 Drinking Water Fortified with Contaminants 8
3.3.3 Drinking Water Fortified with Potential Interferences 8
3.4 Test Procedure 8
3.4.1 Test Sample Preparation and Storage 8
3.4.2 Test Sample Analysis Procedure 9
3.4.3 Stock Solution Confirmation Analysis 9
4 Quality Assurance/Quality Control 12
4.1 Quality Control of Stock Solution Confirmation Methods 12
4.2 Quality Control of Drinking Water Samples 12
4.3 Audits 13
4.3.1 Performance Evaluation Audit 13
4.3.2 Technical Systems Audit 14
4.3.3 Audit of Data Quality 14
4.4 QA/QC Reporting 15
4.5 Data Review 15
5 Statistical Methods and Reported Parameters 16
5.1 Endpoints and Precision 16
5.2 Toxicity Threshold 17
5.3 False Positive/Negative Responses 17
5.4 Field Portability 17
5.5 Other Performance Factors 18
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6 Test Results 19
6.1 Endpoints and Precision 19
6.1.1 Contaminants 19
6.1.2 Potential Interferences 19
6.1.3 Precision 24
6.2 Toxicity Threshold 25
6.3 False Positive/Negative Responses 26
6.4 Field Portability 26
6.5 Other Performance Factors 27
7 Performance Summary 28
8 References 29
Figures
Figure 2-1. Deltatox® Rapid Toxicity Testing System 2
Tables
Table 3-1. Contaminants and Potential Interferences 5
Table 3-2. Summary of Quality Control and Contaminant Test Samples 7
Table 3-3. Dose Confirmation Results 10
Table 3-4. Water Quality Parameters 11
Table 4-1. Summary of Performance Evaluation Audit 14
Table 4-2. Summary of Data Recording Process 15
Table 6-la. Aldicarb Percent Inhibition Results 20
Table 6-lb. Colchicine Percent Inhibition Results 20
Table 6-lc. Cyanide Percent Inhibition Results 21
Table 6-Id. Dicrotophos Percent Inhibition Results 21
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Table 6-le. Thallium Sulfate Percent Inhibition Results 22
Table 6-If. Botulinum Toxin Percent Inhibition Results 22
Table 6-lg. Ricin Percent Inhibition Results 23
Table 6-lh. Soman Percent Inhibition Results 23
Table 6-li. VX Percent Inhibition Results 24
Table 6-2. Potential Interferences Results 25
Table 6-3. Toxicity Thresholds 26
Table 6-4. False Negative Responses 27
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List of Abbreviations
AMS
Advanced Monitoring Systems
ASTM
American Society for Testing and Materials
ATEL
Aqua Tech Environmental Laboratories
DI
deionized water
DDW
dechlorinated drinking water from Columbus, Ohio
ec50
effective concentration causing 50% inhibition
EPA
U.S. Environmental Protection Agency
ETV
Environmental Technology Verification
HDPE
high-density polyethylene
ID
identification
LD
lethal dose
[iL
microliter
mg
milligram
mL
milliliter
NSDWR
National Secondary Drinking Water Regulations
%D
percent difference
PE
performance evaluation
QA
quality assurance
QC
quality control
QMP
quality management plan
SOP
standard operating procedure
TSA
technical systems audit
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Chapter 1
Background
The U.S. Environmental Protection Agency (EPA) supports the Environmental Technology
Verification (ETV) Program to facilitate the deployment of innovative environmental tech-
nologies through performance verification and dissemination of information. The goal of the
ETV Program is to further environmental protection by accelerating the acceptance and use of
improved and cost-effective technologies. ETV seeks to achieve this goal by providing high-
quality, peer-reviewed data on technology performance to those involved in the design,
distribution, financing, permitting, purchase, and use of environmental technologies.
ETV works in partnership with recognized testing organizations; with stakeholder groups
consisting of buyers, vendor organizations, and permitters; and with the full participation of
individual technology developers. The program evaluates the performance of innovative tech-
nologies by developing test plans that are responsive to the needs of stakeholders, conducting
field or laboratory tests (as appropriate), collecting and analyzing data, and preparing peer-
reviewed reports. All evaluations are conducted in accordance with rigorous quality assurance
(QA) protocols to ensure that data of known and adequate quality are generated and that the
results are defensible.
The EPA's National Exposure Research Laboratory and its verification organization partner,
Battelle, operate the Advanced Monitoring Systems (AMS) Center under ETV. The AMS Center
recently evaluated the performance of the Strategic Diagnostics Inc. Deltatox® rapid toxicity
testing system. Rapid toxicity testing systems were identified as a priority technology
verification category through the AMS Center stakeholder process.
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Chapter 2
Technology Description
The objective of the ETV AMS Center is to verify the performance characteristics of environ-
mental monitoring technologies for air, water, and soil. This verification report provides results
for the verification testing of Deltatox®. Following is a description of Deltatox®, based on
information provided by the vendor. The information provided below was not subjected to
verification in this test.
Deltatox® is an in vitro testing system that uses bioluminescent bacteria to detect toxins in air,
water, soil, and sediment. Deltatox® is a metabolic inhibition test that provides both acute
toxicity and genotoxic analyses. Deltatox® uses a strain of naturally occurring luminescent
bacteria, Vibrio fischeri. Vibrio fischeri are non-pathogenic, marine, luminescent bacteria that
are sensitive to a wide range of toxicants. When properly grown, luminescent bacteria produce
light as a by-product of their cellular respiration. Cell respiration is fundamental to cellular
metabolism and all associated life processes. Bacterial bioluminescence is tied directly to cell
respiration, and any inhibition of cellular activity (toxicity) results in a decreased rate of
respiration and a corresponding decrease in the rate of luminescence. The more toxic the sample,
the greater the percent light loss from the test suspension of luminescent bacteria.
The Vibrio fischeri are supplied in a standard freeze-dried (lyophilized) state, which maintains
their sensitivity and stability. Deltatox® was tested as a stand-alone instrument along with the
Deltatox® reagent. Each test uses approximately one million organisms, and each organism is
less than one micrometer in diameter, providing a veiy high surface-to-volume ratio, increasing
sensitivity and statistical significance. To analyze
water samples, the Vibrio fischeri are reconsti-
tuted in a salt solution, 2.5 milliliters (mL) of the
water sample are diluted with 250 microliters
(jjL) of a Deltatox® reagent, then approximately
1 mL of water sample is added to 100 pL of the
reconstituted bacteria. Luminescence readings are
taken prior to adding the drinking water and then
at 5 minutes after the addition. Results are
displayed as percent inhibition.
Deltatox® is a self-calibrating photometer that
incorporates a photomultiplier tube, a data
collection and reduction system, and software.
Figure 2-1. Deltatox® Rapid Toxicity
Testing System
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Deltatox® can be battery operated and is field-portable, but it does not have temperature control
capabilities. It detects light intensity at 490 nanometers, the wavelength emitted by the bacteria.
Deltatox® can store up to 200 data points. These data can be downloaded to a personal computer
with Windows® 95, 98, or subsequent operating system, running HyperTerminal/Terminal or a
similar program. The data are downloaded as a standard ASCII text file, which can be viewed
and edited in any standard ASCII text editor. Deltatox® is 10 inches x 6 inches x 4.5 inches and
weighs 5.3 pounds (6 pounds with batteries). It operates on five standard "C" type batteries or a
Universal Power Adapter (5.0 volts, direct current at four amps). Deltatox costs $5,900, and the
consumables cost $370 for 100 to 150 tests.
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Chapter 3
Test Design and Procedures
3.1 Introduction
The objective of this verification test of rapid toxicity technologies was to evaluate their ability
to detect certain toxins and to determine their susceptibility to interfering chemicals in a
controlled experimental matrix. Rapid toxicity technologies do not identify or determine the
concentration of specific contaminants, but serve as a screening tool to quickly determine
whether water is potentially toxic. Rapid toxicity technologies use bacteria (e.g., Vibrio
fischeri), enzymes (e.g., luciferase), or small crustaceans (e.g., Daphnia magna) that either
directly, or in combination with reagents, produce a background level of light or use dissolved
oxygen at a steady rate in the absence of toxic contaminants. Toxic contaminants in water are
indicated by a change in the color or intensity of light produced or by a decrease in the dissolved
oxygen uptake rate in the presence of the contaminants.
As part of this verification test, Deltatox® was subjected to various concentrations of
contaminants such as industrial chemicals, pesticides, rodenticides, pharmaceuticals, nerve
agents, and biological toxins. Each contaminant was added to separate drinking water samples
and analyzed. In addition to determining whether Deltatox® can detect the toxicity caused by
each contaminant, its response to interfering compounds in clean drinking water, such as water
treatment chemicals and by-products, was evaluated. Table 3-1 shows the contaminants and
potential interferences that were evaluated during this verification test.
This verification test was conducted according to procedures specified in the Test/QA Plan for
Verification of Rapid Toxicity Technologies.(1) Deltatox® was verified by analyzing a
dechlorinated drinking water (DDW) sample from Columbus, Ohio, fortified with various
concentrations of the contaminants and interferences shown in Table 3-1. Hereafter in this
report, DDW will refer to dechlorinated drinking water from Columbus, Ohio. Where possible,
the concentration of each contaminant or potential interference was confirmed independently by
Aqua Tech Environmental Laboratories (ATEL), Marion, Ohio, or by Battelle, depending on the
analyte.
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Table 3-1. Contaminants and Potential Interferences
Category
Contaminant
Carbamate pesticide
aldicarb
Pharmaceutical
colchicine
Industrial chemical
cyanide
Organophosphate pesticide
dicrotophos
Rodenticide
thallium sulfate
Biological toxins
botulinum toxin, ricin
Nerve agents
soman, VX
Potential interferences
aluminum, copper, iron, manganese, zinc,
chloramination by-products, and chlorination
by-products
Deltatox® was evaluated by
• Endpoint and precision—percent inhibition for all concentration levels of contaminants and
potential interfering compounds and precision of replicate analyses
• Toxicity threshold for each contaminant
• False negative responses—contaminants that were reported as producing inhibition results
similar to the negative control when the contaminant was present at lethal concentrations
• False positive responses—occurrence of inhibition significantly greater than the inhibition
reported for unspiked American Society for Testing and Materials (ASTM) Type II deionized
(DI) water samples (zero inhibition)
• Field portability
• Ease of use
• Throughput.
3.2 Test Design
Deltatox® was used to analyze the DDW sample fortified with contaminants at concentrations
ranging from lethal levels to concentrations 1,000 times less than the lethal dose. The lethal dose
of each contaminant was determined by calculating the concentration at which 250 mL of water
would probably cause the death of a 154-pound person. These calculations were based on
toxicological data available for each contaminant. For soman, the stock solution confirmation
showed degradation in the water; therefore, the concentrations analyzed were less than
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anticipated. Whether the concentration is still a lethal dose, as is the case for all contaminants,
depends on the characteristics of the individual person and the amount of contaminant ingested.
Inhibition results (endpoints) from four replicates of each contaminant at each concentration
level were evaluated to assess the ability of Deltatox® to detect toxicity at various concentrations
of contaminants, as well as to measure the precision of Deltatox® results.
The response of Deltatox® to compounds used during the water treatment process (identified as
potential interferences in Table 3-1) was evaluated by analyzing separate aliquots of DDW
fortified with each potential interference at approximately one-half of the concentration limit
recommended by the EPA's National Secondary Drinking Water Regulations (NSDWR)(2)
guidance. For analysis of by-products of the chlorination process, the unspiked DDW was
analyzed because Columbus, Ohio, uses chlorination as its disinfectant procedure. For the
analysis of by-products of the chloramination process, a separate drinking water sample from
St. Petersburg, Florida, which uses chloramination as its disinfection process, was obtained. The
samples were analyzed after residual chlorine was removed using sodium thiosulfate.
Sample throughput was measured based on the number of samples analyzed per hour. Ease of
use and reliability were determined based on documented observations of the operators and the
verification test coordinator. In addition to comprehensive testing in Battelle laboratories,
Deltatox® was operated in the basement of a Columbus, Ohio, home to test its ability to be
transported and operated in a non-laboratory setting.
3.3 Test Samples
Test samples used in the verification test included drinking water and quality control (QC)
samples. Table 3-2 shows the number and type of samples analyzed. QC samples included
method blanks and positive and negative control samples. The fortified drinking water samples
were prepared from a single drinking water sample collected from the Columbus, Ohio, system.
The water was dechlorinated using sodium thiosulfate and then fortified with various concen-
trations of contaminants and interferences. Using this DDW (Columbus, Ohio, dechlorinated
drinking water), individual solutions containing each contaminant and potential interference
were prepared and analyzed. The DDW containing the potential interferences was analyzed at a
single concentration level, while four dilutions (made using the DDW) were analyzed for each
contaminant using Deltatox®. Mixtures of contaminants and interfering compounds were not
analyzed. One concentration level of cyanide was analyzed in the field setting.
3.3.1 Quality Control Samples
QC samples included method blank samples, which consisted of ASTM Type IIDI water;
positive control samples, which consisted of ASTM Type II DI water or DDW (depending on
vendor preference) fortified with a contaminant and concentration selected by the vendor; and
negative control samples, which consisted of the unspiked DDW. The method blank samples
were used to help ensure that no sources of contamination were introduced in the sample
handling and analysis procedures. Either zinc sulfate or phenol were suggested by the vendor for
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Table 3-2. Summary of Quality Control and Contaminant Test Samples
Concentration
Type of Sample
Sample Characteristics
Levels (mg/L)
No. of Sample Analyses
Method blank
NS(a)
9
Positive control
115 (Phenol)
10
Quality control
25 (Zinc sulfate)
14
Negative control (unspiked
NS
44
DDW)
Aldicarb
280; 28; 2.8; 0.28
4 per concentration level
Colchicine
240; 24; 2.4; 0.24
4 per concentration level
Cyanide
250; 25; 2.5; 0.25
4 per concentration level
Dicrotophos
1,400; 140; 14; 1.4
4 per concentration level
Thallium sulfate
2,400; 240; 24; 2.4
4 per concentration level
DDW fortified
with contaminants
Botulinum toxin®
0.30; 0.030; 0.0030;
0.0030
4 per concentration level
Ricin(c)
15; 1.5; 0.15; 0.015
4 per concentration level
Soman
0.18(d); 0.018;
0.0018; 0.00018
4 per concentration level
VX
0.22; 0.022; 0.0022;
0.00022
4 per concentration level
Field location
Cyanide
2.5
4
Aluminum
0.36
4
DDW fortified
Copper
0.65
4
with potential
interferences
Iron
0.069
4
Manganese
0.26
4
Zinc
3.5
4
Disinfectant
by-products
Chloramination by-
products
Chlorination by-products
NS
NS
4
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3.3.2 Drinking Water Fortified with Contaminants
Approximately 150 liters of Columbus, Ohio, tap water were collected in a high-density
polyethylene (HDPE) container. The sample was dechlorinated with 0.5 mL of 0.4 M sodium
thiosulfate for every liter of water. All subsequent test samples were prepared from this DDW
and stored in glass containers to avoid chlorine leaching from HDPE containers.
A stock solution of each contaminant was prepared in ASTM Type IIDI water at concentrations
above the lethal dose level. The stock solution was diluted in DDW to obtain one sample
containing the lethal dose concentration for each contaminant and three additional samples with
concentrations 10, 100, and 1,000 times less than the lethal dose. Table 3-2 lists each concentra-
tion level and the number of samples analyzed at each level.
3.3.3 Drinking Water Fortified with Potential Interferences
Individual aliquots of the DDW were fortified with one-half the concentration specified by the
EPA's NSDWR for each potential interference. Table 3-2 lists the interferences, along with the
concentrations at which they were tested. Four replicates of each of these samples were
analyzed. To test the sensitivity of Deltatox® to by-products of the chlorination process as
potential interferences, the unspiked DDW (same as the negative control) was used since the
water sample originated from a utility that uses chlorination as its disinfectant procedure. In a
similar test involving the by-products of the chloramination process, an additional water sample
was obtained from St. Petersburg, Florida, a city that uses chloramination as its disinfectant
procedure. The residual chlorine in both of these samples was removed using sodium thiosulfate,
and then the samples were analyzed in replicate with no additional fortification of contaminants.
3.4 Test Procedure
3.4.1 Test Sample Preparation and Storage
A drinking water sample was collected as described in Section 3.3.2 and, because free chlorine
kills the bacteria within the Deltatox® reagent and can degrade the contaminants during storage,
was immediately dechlorinated with sodium thiosulfate. Prior to preparing each stock solution,
dechlorination of the water sample was qualitatively confirmed by adding an n,n-diethyl-p-
phenylenediamine tablet to a 25-mL aliquot of the DDW. Once dechlorination was confirmed,
all the contaminant samples, potential interference samples, and negative control QC samples
were made from this DDW, while the method blank sample was prepared from ASTM Type II
DI water. The positive control samples were made using ASTM Type II DI water in Class A
volumetric glassware. All QC samples were prepared prior to the start of the testing and stored at
room temperature for a maximum of 60 days. The aliquots of DDW containing the contaminants
were prepared within seven days of testing and stored in the dark at room temperature without
chemical preservation. Aliquots to be analyzed by each technology were placed in uniquely
labeled sample containers. The sample containers were assigned an identification (ID) number.
A master log of the samples and sample ID numbers for each technology was kept by Battelle.
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3.4.2 Test Sample Analysis Procedure
To analyze the test samples, the Vibrio fischeri were reconstituted in a salt solution, and an
aliquot of drinking water was added to a small amount of the reconstituted bacteria. The sample
cuvettes were inserted into the Deltatox® for a luminescence reading prior to adding the drinking
water and then at 5 minutes after the addition. Software within the Deltatox® automatically
calculates the result (percent inhibition) for each sample.
For each contaminant, Deltatox® analyzed the lethal dose concentration and three additional
concentration levels four times. Only one concentration of potential interference was analyzed.
Deltatox® reports the percent inhibition for each sample. When Deltatox® produced percent
inhibitions greater than 50% for a contaminant, EC50 (effective concentration causing 50%
inhibition) values were also calculated and reported. To test the field portability of Deltatox®, a
single concentration level of cyanide, prepared in the same way as the other DDW samples, was
analyzed in replicate by Deltatox® in the basement of a Columbus, Ohio, home. Sample analysis
procedures were performed in the same way as during testing in the laboratory. Two operators
performed all the analyses using Deltatox®. Both held bachelor's degrees in the sciences and
were trained by the vendor to operate Deltatox®.
3.4.3 Stock Solution Confirmation Analysis
The concentrations of the contaminant and interfering compound stock solutions were verified
with standard analytical methods, with the exception of colchicine, ricin, and botulinum toxin—
contaminants without standard analytical methods. Aliquots to be analyzed by standard methods
were preserved as prescribed by the method. In addition, the same standard methods were used
to measure the concentrations of each contaminant/potential interference in the unspiked DDW
so that background concentrations of contaminants or potential interferences were accounted for
within the displayed concentration of each contaminant/potential interference sample. Table 3-3
lists the standard methods used to measure each analyte; the results from the stock solution
confirmation analyses (obtained by reporting the lethal dose concentration for the contaminants
and the single concentration that was analyzed for the potential interferences); and the
background levels of the contaminants and potential interferences measured in the DDW
sample, which were all non-detect or negligible.
Standard methods were also used to characterize several water quality parameters such as the
concentration of trihalomethanes, haloacetic acids, and total organic halides; turbidity; dissolved
organic carbon content; pH; alkalinity; specific conductivity; and hardness. Table 3-4 lists these
measured water quality parameters for both the water sample collected in Columbus, Ohio,
representing a water system using chlorination as the disinfecting process, and the water sample
collected in St. Petersburg, Florida, representing a water system using chloramination as the
disinfecting process.
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Table 3-3. Dose Confirmation Results
Method
Average Concentration
± Standard Deviation N
= 4 (mg/L)
Background in
DDW Sample
(mg/L)
Contaminant
Aldicarb
EPA 531.1(3)
280 ± 28
<0.0007
Colchicine
(a)
NA03'
NA
Cyanide
EPA 335.1(4)
250 ± 15
0.008
Dicrotophos
EPA SW846 (8141A)®
1,400 ± 140
<0.002
Thallium sulfate
EPA 200.8(6)
2,400 ± 24
<0.001
Botulinum toxin
(a)
NA
NA
Ricin
(a)
NA
NA
Soman
(<0
0.18(d)± 0.001
<0.05
VX
(<0
0.20 ± 0.02
<0.05
Potential Interference
Aluminum
EPA 200.8
0.36 ±0.01
<0.10
Copper
EPA 200.8
0.65 ±0.01
0.011
Iron
EPA 200.8
0.069 ± 0.008
<0.04
Manganese
EPA 200.8
0.26 ±0.01
<0.01
Zinc
EPA 200.8
3.5 ±0.35
0.30
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Table 3-4. Water Quality Parameters
Dechlorinated
Dechlorinated Columbus, St. Petersburg, Florida,
Ohio, Tap Water (disinfected Tap Water (disinfected by
Parameter
Method
by chlorination)
chloramination)
Turbidity
EPA 180.1(7)
0.1 NTU(a)
0.3 NTU
Organic carbon
SM 5310(8)
2.5 mg/L
2.9 mg/L
Specific conductivity
SM 2510(S)
364 |imho
460 |imho
Alkalinity
SM 2320(S)
42 mg/L
97 mg/L
pH
EPA 150.1(9)
7.65
7.95
Hardness
EPA 130.2(9)
112 mg/L
160 mg/L
Total organic halides
SM 5320B(8)
190 |ig/L
83 |ig/L
Total trihalomethanes
EPA 524.2(10)
52.8 |ig/L
2.4 |ig/L
Total haloacetic acids
EPA 552.2(11)
75.7 |ig/L
13.5 |ig/L
(a) NTU = nephelometric turbidity unit.
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Chapter 4
Quality Assurance/Quality Control
QA/QC procedures were performed in accordance with the quality management plan (QMP) for
the AMS Center(12) and the test/QA plan for this verification test.(1)
4.1 Quality Control of Stock Solution Confirmation Methods
The stock solutions for aldicarb, cyanide, dicrotophos, and thallium sulfate were analyzed using
a standard reference method at ATEL. As part of ATEL's standard operating procedures (SOPs)
various QC samples were analyzed with each sample set. These included matrix spike,
laboratory control spike, and method blank samples. According to the standard methods used for
the analyses, recoveries of the QC spike samples analyzed with samples from this verification
test were within acceptable limits of 75% to 125%, and the method blank samples were below
the detectable levels for each analyte. For VX and soman, the confirmation analyses were
performed at Battelle using a Battelle SOP. Calibration standard recoveries of VX and soman
were always between 69% and 130%, and most of the time were between 90% and 100%.
Standard analytical methods for colchicine, ricin, and botulinum toxin were not available and,
therefore, not performed. QA audits and balance calibrations assured that solutions for these
compounds were accurately prepared.
4.2 Quality Control of Drinking Water Samples
A method blank sample consisting of ASTM Type IIDI water was analyzed once by Deltatox®
for approximately every 20 drinking water samples that were analyzed. According to the
Deltatox® procedure, the first sample of each analysis set was treated as the zero control sample
to correct the response of the instrument with respect to a clean water sample. For the majority
of this verification test, this sample was the method blank. When the method blank sample
(ASTM Type n DI water) was added to the bacteria and the five-minute reaction period had
ended, the operators placed the cuvette into the Deltatox®; but, according to its protocol,
Deltatox® did not report a measurement of luminescence and prompted the insertion of the first
sample cuvette. After testing, it was ascertained that, to obtain inhibition data about the method
blank samples, ASTM Type II DI water should have been analyzed as a sample in some position
other than the first in the analysis set. This was not done. Therefore, the Deltatox® data set is
lacking method blank data. However, a negative control sample (unspiked DDW) was analyzed
with approximately every four samples. The absolute inhibitions of the negative controls were
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small, indicating that they caused inhibition similar to the ASTM Type IIDI water, which was
used as the zero control sample (i.e., set to zero inhibition). Results from samples fortified with
contaminants were compared with the results from the negative control to determine if inhibition
was caused by the contaminant. A positive control sample also was analyzed once for approxi-
mately every 20 drinking water samples. While performance limits were not placed on the
results of the positive control sample, the vendor informed Battelle that, if the positive control
samples did not cause greater than approximately 50% inhibition, it would indicate to the
operator that Deltatox® was operating incorrectly. More than 50% inhibition was observed in
each analysis of the positive control sample, indicating the proper functioning of Deltatox®. For
10 positive control samples of phenol, inhibitions of 73% ± 5% were measured. For 14 samples
of zinc sulfate, inhibitions of 94% ±5% were measured.
4.3 Audits
4.3.1 Performance Evaluation Audit
The concentration of the standards used to prepare the contaminant and potential interferences
was confirmed by analyzing solutions of each analyte prepared in ASTM Type II DI water from
two separate commercial vendors using the confirmation methods. The standards from one
source were used to prepare the stock solutions during the verification test, while the standards
from a second source were used exclusively to confirm the accuracy of the measured concentra-
tion of the first source. The percent difference (%D) between the measured concentration of the
performance evaluation (PE) sample and the prepared concentration of that sample was
calculated using the following equation:
M
%D = — x 100%
A (1)
where M is the absolute value of the difference between the measured and the prepared concen-
tration and A is the prepared concentration. The %D between the measured concentration of the
PE standard and the prepared concentration had to be less than 25 for the measurements to be
considered acceptable. Table 4-1 shows the results of the PE audit for each compound. All %D
values were less than 25.
Given the lack of confirmation methodology for some of the contaminants in this verification
test, PE audits were not performed for all of the contaminants. PE audits were performed when
more than one source of the contaminant or potential interference was commercially available
and when methods were available to perform the confirmation. To assure the purity of the other
standards, documentation, such as certificates of analysis, was obtained for colchicine,
botulinum toxin, and ricin. In the case of VX and soman, which were obtained from the U.S.
Army, the reputation of the source, combined with the confirmation analysis data, provided
assurance of the concentration analyzed.
13
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Table 4-1. Summary of Performance Evaluation Audit
Average Measured
Concentration ±
Standard Deviation
Actual Concentration
Percent
(mg/L)
(mg/L)
Difference
Aldicarb
0.00448 ± 0.000320
0.00500
11
Cyanide
0.207 ± 0.026
0.200
4
Contaminant
Dicrotophos
0.00728 ± 0.000699
0.00748
3
Thallium
0.090 ± 0.004
0.100
10
sulfate
Aluminum
0.512 ±0.013
0.500
2
Copper
0.106 ±0.002
0.100
6
Potential
interference
Iron
0.399 ± 0.004
0.400
0.30
Manganese
0.079 ± 0.003
0.100
21
Zinc
0.106 ±0.016
0.100
6
4.3.2 Technical Systems Audit
The Battelle Quality Manager conducted a technical systems audit (TSA) to ensure that the
verification test was performed in accordance with the test/QA plan(1) and the AMS Center
QMP.(12) As part of the audit, the Battelle Quality Manager reviewed the contaminant standard
and stock solution confirmation methods, compared actual test procedures with those specified
in the test/QA plan, and reviewed data acquisition and handling procedures. Observations and
findings from this audit were documented and submitted to the Battelle verification test
coordinator for response. No findings were documented that required any significant action. The
records concerning the TSA are permanently stored with the Battelle Quality Manager.
The EPA Quality Manager also conducted a TSA to ensure that the verification test was
performed in accordance with the test/QA plan(1) and the AMS Center QMP.(12) As part of the
audit, the EPA Quality Manager compared actual test procedures with those specified in the
test/QA plan and reviewed data acquisition and sample preparation records and procedures. No
significant findings were observed during the EPA TSA. The records concerning the TSA are
permanently stored with the EPA Quality Manager.
4.3.3 Audit of Data Quality
At least 10% of the data acquired during the verification test were audited. Battelle's Quality
Manager traced the data from the initial acquisition, through reduction and statistical analysis,
to final reporting, to ensure the integrity of the reported results. All calculations performed on
the data undergoing the audit were checked.
14
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4.4 QA/QC Reporting
Each internal assessment and audit was documented in accordance with Sections 3.3.4 and 3.3.5
of the QMP for the ETV AMS Center.(12) Once the assessment report was prepared, the Battelle
verification test coordinator ensured that a response was provided for each adverse finding or
potential problem and implemented any necessary follow-up corrective action. The Battelle
Quality Manager ensured that follow-up corrective action was taken. The results of the TSA
were sent to the EPA.
4.5 Data Review
Records generated in the verification test were reviewed before these records were used to
calculate, evaluate, or report verification results. Table 4-2 summarizes the types of data
recorded. The review was performed by a technical staff member involved in the verification
test, but not the staff member who originally generated the record. The person performing the
review added his/her initials and the date to a hard copy of the record being reviewed.
Table 4-2. Summary of Data Recording Process
Data to be
Recorded
Responsible
Party
Where
Recorded
How Often
Recorded
Disposition of Data(a)
Dates, times of test
events
Battelle
Laboratory
record books
Start/end of test, and
at each change of a
test parameter
Used to organize/check
test results; manually
incorporated in data
spreadsheets as
necessary
Sample preparation
(dates, procedures,
concentrations)
Battelle
Laboratory
record books
When each sample
was prepared
Used to confirm the
concentration and
integrity of the samples
analyzed, procedures
entered into laboratory
record books
Test parameters
(contaminant
concentrations,
location, etc.)
Battelle
Laboratory
record books
When set or
changed
Used to organize/check
test results, manually
incorporated in data
spreadsheets as
necessary
Stock solution
confirmation
analysis, sample
analysis, chain of
custody, and
results
Battelle or
contracted
laboratory
Laboratory
record books,
data sheets, or
data acquisition
system, as
appropriate
Throughout sample
handling and
analysis process
Transferred to
spreadsheets/agreed
upon report
(a) All activities subsequent to data recording were carried out by Battelle.
15
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Chapter 5
Statistical Methods and Reported Parameters
The statistical methods presented in this chapter were used to verify the performance parameters
listed in Section 3.1.
5.1 Endpoints and Precision
Deltatox® reports the percent inhibition for each sample analyzed. Each DDW sample containing
contaminants was compared with a negative control sample that, for this verification test, was
unspiked DDW. This comparison was made by subtracting the percent inhibition of the negative
control within a sample set from the inhibition produced by each sample in the sample set.
Therefore, the percent inhibition of the negative control sample within each sample set was zero
percent.
For contaminants that induced inhibition of greater that 50%, the concentration of contaminant
that affects 50% of the bacteria in the Deltatox® reagent (EC50) was estimated from the linear
regression of the log of each concentration level of the contaminant versus the percent
inhibition. For contaminants that did not induce inhibition of greater than 50%, this calculation
was not appropriate.
The standard deviation (S) of the results for the replicate samples was calculated, as follows, and
used as a measure of technology precision at each concentration.
5 =
In, -s 2 ~>1'2
—j Z '» -I)
: —1 k=r '
(2)
where n is the number of replicate samples, Ik is the percent inhibition measured for the kth
sample, and I is the average percent inhibition of the replicate samples. Because the average
inhibitions were frequently near zero for this data set, relative standard deviations often would
have greatly exceeded 100%, making the results difficult to interpret. Therefore, the precision
results were left in the form of standard deviations so the reader could easily view the
uncertainty around the average for results that were both near zero and significantly larger than
zero.
16
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5.2 Toxicity Threshold
The toxicity threshold was defined as the lowest concentration of contaminant to exhibit a
percent inhibition significantly greater than the negative control. Also, the inhibition of the
toxicity threshold had to be significantly different than the inhibition of the other concentrations
analyzed. Since the inhibition of the test samples was calculated with respect to the inhibition of
each negative control sample, the percent inhibition of the negative control was always zero. An
inhibition was significantly greater than the negative control if the average inhibition plus or
minus the standard deviation did not include zero.
5.3 False Positive/Negative Responses
A response would be considered false positive if an unspiked drinking water sample produced
an inhibition significantly greater than zero when determined with respect to ASTM Type IIDI
water. Depending on the degree of inhibition in the sample, toxicity due to subsequent
contamination of that sample may not be detectable or could be exaggerated as a result of the
baseline inhibition. To test for this possibility, the percent inhibition of the unspiked drinking
water was determined with respect to ASTM Type II DI water. Drinking water samples collected
from water systems using chlorination and chloramination as the disinfecting process were
analyzed in this manner. An inhibition was considered significantly different from zero if the
average inhibition, plus or minus the standard deviation, did not include zero.
A response was considered false negative when Deltatox® was subjected to a lethal concentra-
tion of some contaminant in the DDW and did not indicate inhibition significantly greater than
the negative control (zero inhibition) and the other concentration levels analyzed. Requiring the
inhibition of the lethal dose sample to be significantly greater than zero and the other concentra-
tion levels more thoroughly incorporated the uncertainty of all the measurements made by
Deltatox® in determining a false negative result. A difference was considered significant if the
average inhibition plus or minus the standard deviation did not encompass the value or range of
values that were being compared.
5.4 Field Portability
The results obtained from the measurements made on drinking water samples in the laboratory
and field setting were compiled independently and compared to assess the performance of the
Deltatox® under different analysis conditions. Means and standard deviations of the endpoints
generated in both locations were used to make the comparison. Also, qualitative observations of
Deltatox® in a non-laboratory setting were made by the verification test coordinator and
operators. Factors such as the ease of transport and set-up, demand for electrical power, and
space requirement were documented.
17
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5.5 Other Performance Factors
Ease of use (including clarity of the instruction manual, user-friendliness of software, and
overall convenience) was qualitatively assessed throughout the verification test through
observations of the operators and verification test coordinator. Sample throughput was evaluated
quantitatively based on the number of samples that could be analyzed per hour.
18
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Chapter 6
Test Results
6.1 Endpoints and Precision
Tables 6-la-i present the percent inhibition data for nine contaminants, and Table 6-2 presents
data for five potential interferences and the drinking water samples disinfected by both
chlorination and chloramination. Given in each table are the concentrations analyzed, the
percent inhibition results for each replicate at each concentration, and the average and standard
deviation of the inhibition of the four replicates at each concentration. EC50 values also are given
when applicable. Samples that produced negative percent inhibition values indicated an increase
in light production by the bacteria relative to the negative control.
6.1.1 Contaminants
The contaminants that were analyzed by Deltatox® during this verification test produced one of
two trends apparent from Tables 6-la-i. Contaminants caused percent inhibitions that, starting
from the lowest concentration that produced inhibitions near zero, either increased in proportion
to the concentration in the sample, resulting in the two highest concentration levels exhibiting
higher inhibitions than the other concentration levels, or did not change considerably regardless
of what concentration was analyzed. Aldicarb, dicrotophos, and thallium sulfate fall into the
former category, while colchicine, botulinum toxin, ricin, VX, and soman fall into the latter
category. The one exception was cyanide, for which the inhibitions of all four concentration
levels were significantly different from one another and the inhibitions increased with
concentration.
6.1.2 Potential Interferences
Table 6-2 presents the results from the samples that were analyzed to test the effect of potential
interferences on Deltatox®. Aluminum, iron, and manganese exhibited percent inhibitions near
zero, indicating little or no response to these compounds, while copper and zinc exhibited higher
inhibitions of 38% and 22%, respectively, indicating a slightly elevated response.
19
-------
Table 6-la. Aldicarb Percent Inhibition Results
Concentration Inhibition Average Standard Deviation EC50
(mg/L) (%) (%) (%) (mg/L)
-3
0.28 J -1 2
-2
70.5
14
2.8 ^ 6 5
6
24
28 H 26 1
26
73
28° 74
(Lethal Dose) 74
66
Table 6-lb. Colchicine Percent Inhibition Results
Concentration Inhibition Average Standard Deviation EC50
(mg/L) (%) (%) (%) (mg/L)
0
0.24 ^ 2 2
4
0
4
2.4 ^ 3 2
4
2
24 "4 0 4
0
JNAV 7
9
24° 5
(Lethal Dose) 25
8
(a) NA = Not applicable.
20
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Table 6-lc. Cyanide Percent Inhibition Results
Concentration Inhibition Average Standard Deviation EC50
(mg/L) (%) (%) (%) (mg/L)
4
5 5 1
0.25 4
5
7.6
15
12
2.5 14 2
16
14
85
25 I6. 81 4
O 1
80
106
250 101
(Lethal Dose) 104
102
28
2.5 35
(Field Location) 32
29
NA(a)
(a) NA = Not applicable.
Table 6-Id. Dicrotophos Percent Inhibition Results
Concentration Inhibition Average Standard Deviation EC50
(mg/L) (%) (%) (%) (mg/L)
-3
1.4 -2 2
1
540
7
14 2 5
3
42
140 25 12
24
68
1,400 54 65 8
(Lethal Dose) 65
73
21
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Table 6-le. Thallium Sulfate Percent Inhibition Results
Concentration Inhibition Average Standard Deviation EC50
(mg/L) (%) (%) (%) (mg/L)
5
2,4 4 5 1
5
NA(a)
4
24 3 2 3
4
13
1 O
240 14 4
17
24
2,400 24
(Lethal Dose) 23
27
(a) NA = Not applicable.
Table 6-If. Botulinum Toxin Percent Inhibition Results
Concentration Inhibition Average Standard Deviation EC50
(mg/L) (%) (%) (%) (mg/L)
-8
0.00030 -4 3
-4
NA(a)
-3
0.003 4 -5 1
-6
-4
0.030 -3 2
-5
-6
0.30 0
(Lethal Dose) 1
-1
(a) NA = Not applicable.
22
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Table 6-lg. Ricin Percent Inhibition Results
Concentration Inhibition Average Standard Deviation EC50
(mg/L) (%) (%) (%) (mg/L)
1
0.015 J 3 2
2
NA(a)
1
0.15 * 3 1
4
1.5 -9
1
-2
7
15 ° 2 4
(Lethal Dose) 2
-2
(a) NA = Not applicable.
Table 6-lh. Soman Percent Inhibition Results
Concentration Inhibition Average Standard Deviation EC50
(mg/L) (%) (%) (%) (mg/L)
1
0
0.00018 -4 1 5
8
NA(a)
5
0.0018 10 8 3
10
-10
0.018 ~2 -6 3
-6
4
0.18® 0
(Lethal Dose) 4
0
(a) NA = Not applicable.
(b) Due to the degradation of soman in water, the stock solution confirmation analysis confirmed that the
concentration of the lethal dose was 61% of the expected concentration of 0.30 mg/L.
23
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Table 6-li. VX Percent Inhibition Results
Concentration
Inhibition
Average
Standard Deviation
EC-50
(mg/L)
(%)
(%)
(%)
(mg/L)
-6
0.00022
-3
0
3
-2
4
0
0.0022
-6
-1
1
6
9
NA(a)
2
0.022
2
2
3
2
1
0.22
6
8
f.
(Lethal Dose)
5
3
0
z
(a) NA = Not applicable.
All of the contaminant and potential interference samples were prepared in the DDW and
compared with unspiked DDW. Therefore, any background inhibition in the DDW was
corrected by subtracting the inhibition caused by the negative control sample. To investigate
whether Deltatox® is sensitive to by-products of disinfecting processes, dechlorinated drinking
water samples from water systems that use chlorination and chloramination were analyzed and
compared with ASTM Type IIDI water as the control sample. This determination is crucial
because the ability of Deltatox® to detect toxicity is dependent on the bacteria's background
light production in a clean drinking water matrix. If clean drinking water produces 100%
inhibition of light, inhibition caused by contaminants could not be detected. On average, the
chlorinated sample exhibited no detectable inhibition, indicating no toxicity, while the
chloraminated sample exhibited nearly complete inhibition (average 88% inhibition). This
suggests that samples that have been disinfected by using a chloramination process are likely to
produce false positive results because the background water sample would completely inhibit the
Deltatox® reagent. For aldicarb, cyanide, and dicrotophos, whose inhibitions increased with
concentration and spanned the range from approximately no inhibition to greater than 50%
inhibition, EC50 values were calculated and reported in Tables 6-la, 6-lc, and 6-Id. Because
inhibitions did not reach 50% for the other contaminants, EC50 values could not be calculated.
6.1.3 Precision
Across all the contaminants and potential interferences, the standard deviation was measured
and reported for each set of four replicates to evaluate the Deltatox® precision. The standard
deviation of the four replicate measurements was greater than 10% for only one sample and, in
most cases, it was less than 5%.
24
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Table 6-2. Potential Interferences Results
Potential
Concentration
Inhibition
Average
Interferences
(mg/L)
(%)
(%)
Standard Deviation (%)
-2
Aluminum
0.36
7
6
2
3
4
36
Copper
0.65
40
42
34
38
4
-10
Iron
0.069
1
3
-5
-3
6
6
Manganese
0.26
-6
-1
-6
-2
6
24
Zinc
3.5
26
23
13
22
6
Chlorination
by-products
NA(a)
(b)
-4
9
89
Chloramination
NA
87
1
by-products
OO 00
00 00
66
(a) NA = Not applicable.
(b) Chlorination by-product data averaged over negative control data compared to ASTM Type IIDI water.
6.2 Toxicity Threshold
Table 6-3 gives the toxicity thresholds, as described in Section 5.2, for each contaminant. The
lowest toxicity threshold concentration was for cyanide at 0.25 mg/L, indicating that Deltatox®
was most sensitive to cyanide. For colchicine, botulinum toxin, ricin, soman, and VX, no
inhibition greater than the negative control was detected, regardless of the concentration level,
indicating that the technology was not highly responsive to these contaminants.
25
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Table 6-3. Toxicity Thresholds
Contaminant Concentration (mg/L)
Aldicarb 28
Colchicine ND(a)
Cyanide 0.25
Dicrotophos 140
Thallium sulfate 240
Botulinum toxin ND
Ricin ND
Soman ND
VX ND
(a) ND = Significant inhibition was not detected.
6.3 False Positive/Negative Responses
False positive responses were observed for unspiked chloraminated tap water. As described in
Section 6.1.2, for a clean tap water sample that had been disinfected using a chloramination
process, Deltatox® reported almost complete inhibition (-88%). By-products of this chloramina-
tion process apparently inhibited the Deltatox® reagent. The water sample treated by chlorination
and then subsequently dechlorinated caused no detectable inhibition. A false negative response is
when a lethal dose of contaminant is present in the water sample and the inhibition is not sig-
nificantly different from either the negative control or the other lower concentration levels.
Table 6-4 gives these results. The inhibition induced by lethal doses of aldicarb, cyanide,
dicrotophos, and thallium sulfate was detectable by Deltatox®, while colchicine, botulinum toxin,
ricin, soman, and VX did not indicate inhibition greater than the negative control, indicating false
negative responses.
6.4 Field Portability
A single concentration of cyanide was prepared and analyzed in replicate at a field location to
examine its ability to be used in a non-laboratory setting. Deltatox® and necessary accessories
were conveniently transported to the field in the hard plastic carrying case provided by the
vendor. Fully loaded, the case weighed about 15 pounds. At the field location, Deltatox® was
operated with five "C" batteries on a small table in the basement of a house. Table 6-lc shows
the results of the cyanide samples analyzed in the field, along with the results of the cyanide
samples analyzed in the laboratory. The concentration of the solution analyzed in the field was
2.5 mg/L. The inhibition produced in the field was 31% ± 3%, and the inhibition produced in the
laboratory at the same concentration was 14% ± 2%. While these inhibitions are not the same,
the field measurements were made on freshly prepared solutions with a newly reconstituted batch
of bacteria. The precision of the results and the fact that the absolute percent inhibition was
26
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Table 6-4. False Negative Responses
Contaminant
Lethal Dose
Concentration
(mg/L)
False Negative
Response
Aldicarb
280
no
Colchicine
240
yes
Cyanide
250
no
Dicrotophos
1,400
no
Thallium sulfate
2,400
no
Botulinum toxin
0.30
yes
Ricin
15
yes
Soman
0.18(a)
yes
VX
0.22
yes
(a) Due to the degradation of soman in water, the stock solution confirmation
analysis confirmed that the concentration of the lethal dose was 61% of the
expected concentration of 0.30 mg/L.
within 20% of that in the laboratory indicate that Deltatox® functioned properly at the field
location. In addition, the positive control samples analyzed at the field location produced
inhibitions of 86% and 73% for phenol and zinc sulfate, respectively. These inhibitions are very
similar to the overall average inhibitions for those controls, as shown in Table 4-1.
The Deltatox® reagent must be kept at approximately -20°C prior to reconstitution and, once
reconstituted, needs to be consumed within two hours. These factors could be problematic in a
long-term field deployment.
6.5 Other Performance Factors
The step-by-step pictorial instruction manual for Deltatox® was easy to understand, which
enabled operators to become quickly adept at analyzing multiple sample sets. Deltatox® was very
straightforward to operate. The operators analyzed 20 samples per hour. Although the operators
had scientific backgrounds, based on observations of the verification test coordinator, an operator
with little technical training would probably be able to follow the manual instructions to analyze
samples successfully.
27
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Chapter 7
Performance Summary
Parameter
Compound
Lethal
Dose (LD)
Cone.
Average Inhibitions at Concentrations
Relative to the LD Concentration (%)
Range of
Standard
Deviations (%)
Toxicity
Thresh.
(mg/L/'"
LD
LD/10
LD/100
LD/1,000
Contaminants in
DDW
Aldicarb
280
72
26
6
-1
1-5
28
Colchicine
240
12
0
3
2
2-9
ND(b)
Cyanide
250
103
81
14
5
1^
0.25
Dicrotophos
1,400
65
25
2
-2
2-12
140
Thallium
sulfate
2,400
25
14
2
5
1^
240
Botulinum
toxin(c)
0.30
-2
-3
-5
-4
1-3
ND
Ricin(d)
15.0
2
-4
3
3
1-5
ND
Soman
0.18
-------
Chapter 8
References
1. Test/QA Plan for Verification of Rapid Toxicity Technologies, Battelle, Columbus, Ohio,
June 2003.
2. United States Environmental Protection Agency, National Secondary Drinking Water
Regulations: Guidance for Nuisance Chemicals, EPA/810/K-92/001, July 1992.
3. U.S. EPA Method 531.1, "Measurement of n-Methylcarbamoyloximes and
n-Methylcarbamates in Water by Direct Aqueous Injection HPLC with Post Column
Derivatization," in Methods for the Determination of Organic Compounds in Drinking
Water—Supplement 111, EPA/600/R-95/131, 1995.
4. U.S. EPA Method 335.1, "Cyanides, Amenable to Chlorination," in Methods for the
Chemical Analysis of Water and Wastes, EPA/600/4-79/020, March 1983.
5. SW846 Method 8141 A, "Organophosphorous Compounds by Gas Chromatography:
Capillary Column Technique," Revision 1, September 1994.
6. U.S. EPA Method 200.8, "Determination of Trace Elements in Waters and Wastes by
Inductively-Coupled Plasma Mass Spectrometry," in Methods for the Determination of
Organic Compounds in Drinking Water, Supplement I, EPA/600/R-94/111, 1994.
7. U.S. EPA Method 180.1, "Turbidity (Nephelometric)," Methods for the Determination of
Inorganic Substances in Environmental Samples, EPA/600/R-93/100, 1993.
8. American Public Health Association, et al. Standard Methods for the Examination of Water
and Wastewater. 19th Edition, 1997. Washington, DC.
9. U.S. EPA, Methods for Chemical Analysis of Water and Wastes, EPA/600/4-79/020.
10. U.S. EPA Method 524.2, "Purgeable Organic Compounds by Capillary Column GC/Mass
Spectrometry," Methods for the Determination of Organic Compounds in Drinking
Water—Supplement 111, EPA/600/R-95/131.
11. U.S. EPA Method 552.2, "Haloacetic Acids and Dalapon by Liquid-Liquid Extraction,
Derivatization and GC with Electron Capture Detector," Methods for the Determination of
Organic Compounds in Drinking Water—Supplement III, EPA/600/R-95/131.
29
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12. Quality Management Plan (QMP)for the ETV Advanced Monitoring Systems Center,
Version 4.0, U.S. EPA Environmental Technology Verification Program, Battelle,
Columbus, Ohio, December 2002.
30
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