PB-230 316
METHODS FOR ORGANIC PESTICIDES IN WATER
AND WASTEWATER
National Environmental Research Center
Cincinnati, Ohio
1971
DISTRIBUTED BY:
National Technical Information Service
U. S. DEPARTMENT OF COMMERCE
5285 Port Royal Road, Springfield Va. 22151
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METHODS
FOR
ORGANIC PESTICIDES
IN WATER AND WASTEWATER
1971
ENVIRONMENTAL PROTECTION AGENCY
NATIONAL ENVIRONMENTAL RESEARCH CENTER
H
CINCINNATI. OHIO
45268
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PREFACE
The use of pesticides has become a routine practice in modern agri-
culture. While these compounds have great advantages in the control
of predatory insects, they represent a possible danger to the aquatic
environment when present in even trace concentrations.
The National Technical Advisory Committee on Water Quality Criteria has
recommended "that environmental levels . . . not be permitted to rise
above 50 nanograms/liter". Many of the states have incorporated pesti-
cide criteria in their water quality standards. Therefore, the moni-
toring of surface waters for pesticides is an essential part of our
measurement of water quality.
The Analytical Quality Control Laboratory, assisted by Environmental
Protection Agency scientists experienced in the determination of pesti-
cides, has prepared the following method for organochlorine pesticides.
In the opinion of the AQC Laboratory and its advisors, this method is
the best available procedure at this time.
Because methods development and selection is a dynamic process, re-
quiring continual efforts toward improvement, comments on the appli-
cation of the method and suggestions for improvements are solicited
from the analysts in the field. These comments should be addressed to:
Director, Analytical Quality Control Laboratory
Environmental Protection Agency
National Environmental Research Center
Cincinnati, Ohio 45268
With the concurrence of the Office of Pesticides, the method has been
adopted as the EPA Method for Organochlorine Pesticides and is recommen-
ded for use by all laboratories in acquiring data on the concentration
of these materials in waters and wastewaters sampled by EPA.
Dwight G. Ballinger, Director
Analytical Quality Control Laboratory
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EPA COMMITTEE ON
ANALYTICAL METHODS FOR PESTICIDES IN WATER
Chairman: Lichtenberg, James J.
National Environmental Research Center
Analytical Quality Control Laboratory
Cincinnati, Ohio 4S268
Members: Boyle, Harvey
Division of Field Investigations
Denver Federal Center
Building 22
Denver, Colorado 80225
Dressman, Ronald C.
National Environmental Research Center
Analytical Quality Control Laboratory
Cincinnati, Ohio 45268
Eichelberger, James W,
National Environmental Research Center
Analytical Quality Control Laboratory
Cincinnati, Ohio 45268
Garza, Mike
Galveston Bay Field Station
Houston, Texas 77006
Johnson, Dewitt
Illinois Field Office
Chicago, Illinois 60605
Kahn, Lloyd
Edison Water Quality Laboratory
Edison, New Jersey 08817
Longbottom, James E.
National Environmental Research Center
Analytical Quality Control Laboratory
Cincinnati, Ohio 45268
Loy, William
Southeast Water Laboratory
Athens, Georgia 30601
Malueg, Nick
Consolidated Laboratories
Redman, Washington 98052
\l
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Muth, Gerald
Surveillance and Analysis Division
Alameda, California 94501
Streck, Larry
Robert S. Kerr Water Research Center
Ada, Oklahoma 74820
Tabri, Adib F.
National Field Investigations Center
Cincinnati, Ohio 45213
The critical review of this manual by several members of the
Office of Pesticides, the Fish and Wildlife Service, and the
U.S. Geological Survey is gratefully acknowledged.
Ml
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iv
CONTENTS
INTRODUCTION 1
PART I - RECOMMENDED PRACTICE FOR DETERMINATION OF ORGANIC
PESTICIDES IN WATER AND WASTEWATER 3
1. General Information 3
1.1 Introduction 3
1.2 Sample Collection 3
1.3 Sample Handling 4
1.4 Glassware 4
1.5 Standards, Reagents and Solvents 5
1.6 Records 7
2. Common Analytical Operations 8
2.1 Method Blank 8
2.2 Sample Transfer 8
2.3 Concentration of Extracts 8
3. Gas-Liquid Chromatography 9
3.1 Gas Chromatographic System 9
3.2 Injection into the Gas Chromatograph 16
3.3 Qualitative Analysis 16
3.4 Quantitative Analysis 17
4. Column Chromatography 19
4.1 Adsorbents 19
4.2 Packing the Column 20
4.3 Eluting the Column 20
S. Thin-Layer Chromatography 21
5.1 Equipment 21
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5.2 Layer Preparation 21
5.3 Preparation of Developing Chamber 22
5.4 Spotting the Layer 22
5.S Developing the Layer 22
5.6 Visualizing and Sectioning the Layer 22
5.7 Pesticide Removal from the TLC Plates 23
PART II - METHODS OF ANALYSIS 24
A. METHODS FOR ORGANOCHLORINE PESTICIDES 24
1. Scope and Application 24
2. Summary 24
3. Significance 25
4. Interferences 25
5, Method for Analysis Using Electron Capture Gas
Chromatography 27
5.1 Extraction of Sample 27
5.2 Clean-up and Separation Procedures 29
5.3 Gas-Liquid Chromatography 33
5.4 Confirmatory Evidence 34
5.5 Calculation of Results 34
5.6 Reporting Results 35
6. Method for Analysis Using Microcoulometric or
Electrolytic Conductivity Gas Chromatography 35
6.1 Extraction of Sample 35
6.2 Clean-up and Separation Procedures 37
6.3 Gas-Liquid Chromatography 37
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VI
6.4 Confirmatory Evidence 38
6.5 Calculation of Results 38
6.6 Reporting Results 38
REFERENCES 48
APPENDIX 51
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ORGANIC PESTICIDES IN WATER AND WASTEWATER
V
INTRODUCTION
Advances in the science of analytical chemistry in recent years are
typified by constant new developments of methods which yield greater
efficiency, selectivity, and sensitivity. As a result, the analytical
chemist now has the means to measure minute quantities of pesticides,
either singly or in combination. Laboratories capable of this degree of
measurement are now commonplace; making possible pesticide pollution data
for every major stream in the nation.
The increasing sophistication of data storage and retrieval systems
and the every expanding toxicological and ecological information on the
impact of pesticides, permits data obtained by several laboratories to be
used in the combined assessment of pollution in a given river system or
in the nation's major streams, no matter where located.
This growing use of data dictates development of effective means to
minimize procedural error within each laboratory and optimize analytical
agreement between laboratories through use of a standardized method. The
Environmental Protection Agency's "Methods for Organic Pesticides in Water
and Wastewater", presented herein, arc designed to provide the means of
obtaining such agreement and validity.
Part I of this manual, entitled "Recommended Practice for the Deter-
mination of Organic Pesticides in Water'^presents a general discussion,
helpful hints and suggestions, and precautionary measures required for
pesticide analyses. Succeeding chapters present stepwise procedures ^or
various types of pesticides and types of samples. This format was chosen
to provide short, concise, and easy-to-follow methods, to facilitate the
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inclusion of additional methods and revisions of existing methods as they
are developed and found to be acceptable, and to emphasize the analytical
quality control aspects of pesticide analysis.
The Environmental Protection Agency methods offer several analytical
alternatives, depending on the analyst's assessment of the nature and extent
of interferences and the complexity of the pesticide mixtures found. They
are recommended for use only by experienced residue analysts or under the
close supervision of such qualified persons.
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3
PART I - RECOMMENDED PRACTICE FOR DETERMINATION OF ORGANIC PESTICIDES
IN WATER AMD WASTEWATER
1. General Information
1.1 Introduction - Part I of this manual is intended to provide
general information, helpful hints and suggestions, and precautionary
measures which experience has shown to be important in producing con-
sistently reliable results. Part I is by no means complete and other ref-
erences, such as the Food and Drug Administration's "Pesticide Analytical
Manual" (1), The Canadian Department of Agriculture's "Guide to the
Chemicals Used in Crop Protection" (2), and "Official Methods of Analysis
of the Association of Official Analytical Chemists" (3), should also be
consulted by pesticide residue analysts. Other helpful references for
general practice and analytical quality control are ASTM Part 30 "Tentative
Recommended Practice for General Gas Chromatography Procedures" (4), and the
Environmental Protection Agency manual "Control of Chemical Analyses in
Water Pollution Laboratories" (S). Additional recommended references giving
fundamentals of Chromatography in general, and gas chromatography in parti-
cular, are listed in the bibliography.
1.2 Sample Collection - Wide mouth glass bottles equipped with Teflon-
lined screw caps should be used for sample collection. Plastic bottles must
not be used since they are known to introduce interference and absorb pesti-
cides. The size of the sample is dictated by the sensitivity required
for a particular purpose and the detection system to be employed. The
normal sample volume requirements are given in the individual methods of
Part II. If analysis by more than one method is to be performed, sufficient
sample must be collected to supply the need of each analysis. In addition,
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sufficient sample should be collected to permit running of .duplicate and
spiked analyses. Breakage of glass sample bottles is overcome by shipping
them in expanded polystyrene containers molded to fit the bottles. Refer
to ASTM Standards, Part 23, D510 for further sampling recommendations (6).
1.3 Sample Handling - The sample collector should provide the following
information in writing: date, time, location (coordinates or river mile,
city, etc.)i depth, suspected contaminants, type of sample (surface water,
waste discharge, etc. ), name of sample collector, as well as any other
information that may be helpful in selecting the analytical approach as
well as in interpreting results. Upon receipt in the laboratory, samples
should be logged in immediately. Due to the instability of many of the
pesticides in water (7) (8) (9), samples should be extracted and analyzed
as soon as possible after collection. If samples must be stored, they
should be placed in a cool dark place, preferably in a refrigerator.
Holding time and conditions of storage should be reported along with
results.
1.4 Glassware
1.4.1 Cleaning Procedure - It is particularly important that glass-
ware used in pesticide residue analyses be scrupulously cleaned before
initial use as well as after each analysis. The glassware should be
cleaned as soon as possible after use, first rinsing with water or the
solvent that was last used in it. This should be followed by washing with
soap water, rinsing with tap water, distilled water, redistilled acetone
and finally with pesticide quality hexane. Heavily contaminated glassware
may require muffling at 400C for 15 to 30 minutes. High boiling materials,
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such as some of the polychlorinated biphenyls (PCB's) may not be eliminated
by such heat treatment. NOTE: Volumetric ware should not be muffled. The
glassware should be stored immediately after drying to prevent accumulation
of dust or other contaminants. Store inverted or cover mouth with foil.
1.4.2 Calibration - Individual Kuderna-Danish concentrator tubes and/
or centrifuge tubes used for final concentration of extracts must be
accurately calibrated at the working volume. This is especially important
at volumes below 1 ml. Calibration should be made using a precision micro-
syringe, recording the volume required to bring the liquid level to the
individual graduation marks. Class A volumetric ware should be used for
preparing all standard solutions.
1.5 Standards, Reagents and Solvents
I.S.I Analytical Standards and Other Chemicals - Analytical reference
grade standards should be used whenever available. They should be stored
according to the manufacturer's instructions. Standards and reagents sen-
sitive to light should be stored in dark bottles and/or in a cool dark
place. Those requiring refrigeration should be allowed to come to room
temperature before opening. Storing of such standards under nitrogen is
advisable.
1.5.1.1 Stock Standards - Pesticide stock standards solutions should
be prepared in 1 ug/ul concentrations by dissolving 0.100 grams of the
standard in pesticide-quality hexane or other appropriate solvent (Acetone
should not be used since some pesticides degrade on standing in this
solvent) and diluting to volume in a 100 ml ground glass stoppered volumet-
ric flask. The stock solution is transferred to ground glass stoppered
reagent bottles. These standards should be checked frequently for signs
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of degradation and concentration, especially just prior to preparing working
standards from them.
1.5.1.2 Working Standards - Pesticide working standards are prepared
from the stock solutions using a micro syringe, preferably equipped with
a Chaney adapter. The concentration of the working standards will vary
depending on the detection system employed and the level of pesticide in
the samples to be analyzed. A typical concentration (0.1 ng/vl) may be
prepared by diluting 1 ul of the 1 ug/ul stock solution to .volume in a
10 ml ground glass stoppered volumetric flask. The standard solutions
should be transferred to ground glass stoppered reagent bottles. Prepa-
ration of a fresh working standard each day will minimize concentration
through evaporation of solvent. These standards should be stored in the
same manner as the stock solutions.
1.5.1.3 Identification of Reagents - All stock and working standards
should be labeled as follows: name of compound, concentration, date prepa-
red, solvent used, and name of person who prepared it.
1.5.1.4 Anhydrous sodium sulfate used as a drying agent for solvent
extracts should be prewashed with the solvent or solvents that it comes in
contact with in order to remove any interferences that may be present.
1.5.1.5 Cotton used at the top of the sodium sulfate column must be
pre-extracted for about 40 hours in soxhlet using the appropriate sol-
vent. A cheap grade of cotton is recommended. Red Cross cotton is not
recommended.
1.5.2 Solvents - Organic solvents must be of pesticide quality and
demonstrated to be free of interferences in a manner compatible with
whatever analytical operation is to be performed. Solvents can be checked
by analyzing a volume equivalent to that used in the analysis and concen-
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trated to the minimum final volume. Interferences are noted in terras of
gas chromatographic response - relative retention time, peak geometry,
peak intensity and width of solvent response. Interferences noted under
these conditions can be considered maximum. If necessary, a solvent must
be redistilled in glass using a high efficiency distillation system. A
60 cm column packed with 1/8 inch glass helices is effective.
1.5.2.1 Ethyl Ether - Hexane - It is particularly important that
these two solvents, used for extraction of organochlorine pesticides from
water, be checked for interferences just prior to use. Ethyl ether, in
particular, can produce troublesome interferences. [NOTE: The formation
of peroxides in ethyl ether creates a potential explosion hazard. Therefore
it must be checked for peroxides before use.] It is recommended that the
solvents be mixed just prior to use and only in the amount required for
immediate use since build-up of interferences often occurs on standing.
The great sensitivity of the electron capture detector requires that
all solvents used for the analysis be of pesticide quality. Even these
solvents sometimes require redistillation in an all glass system prior
to use. The quality of the solvents may vary from lot to lot and even
within the same lot, so that each bottle of solvent must be checked before
use.
1.6 Records
1.6.1 The progress of the sample through the analysis should be
recorded in permanently bound notebooks or on laboratory data cards.
Dates of extraction, clean-up and separation and G.C. analysis and
date of reporting results should be recorded.
1.6.2 All evidence accumulated during the analysis: gas chroma-
tograms, photographs of thin-layers, infrared spectra, etc. should be
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retained for as long as may be required to fulfill the purpose for the
analysis.
1.6.3 All results should be recorded on laboratory data cards or
bound notebook so as to provide a permanent laboratory record. Where
appropriate, the data should be entered into STORET.
2. Common Analytical Operations
2.1 Method Blank - A method blank must be determined whenever a
sample or group of samples is analyzed. This is done by following
the procedure step by step including all reagents, solvents, and
other materials in the quantity required by the method concurrently
and under conditions identical to those for the samples. Additional
blanks are required whenever a new supply of any of the reagents,
solvents, etc. is introduced.
2.2 Sample Transfer - The utmost in care and technique must be
exercised in order to assure quantitative transfer of extract solutions
from one vessel to another throughout the analysis. Careless technique
will introduce determinate errors and produce inaccurate results. The
internal wall of the vessel must be carefully rinsed several times
(usually three) with a volume of the particular solvent appropriate for
the analysis involved. Final flushing while pouring into the receiving
vessel is recommended.
2.3 Concentration of Extracts
2.3.1 Kudema-Danish (K-D) Evaporation - A Snyder column, evaporative
flask and calibrated receiver ampul are employed. The evaporative flask
should be filled to no more than 60% capacity. Set the K-D assembly over
a vigorously boiling water bath or a live steam bath. The evaporation
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must be carefully attended to avoid loss of pesticides. The water level
should be maintained just below the lower joint, and the apparatus mounted
so that the lower rounded surface of the flask i; bathed in steam. Carry
out the evaporation in a hood so that solvent vapors are exhausted. When
the solvent no longer actively distills, the K-D apparatus is removed
from the bath and allowed to cool. The condensed solvent is allowed to
drain into the ampul before dismantling.
2.3.2 h'inal Concentration - Concentration below 5 ml is usually
required when analyzing surface water samples. Final evaporation to a
minimum of 0.2 ml may be accomplished in the ampul with the aid of a
gentle stream of clean dry nitrogen or air in a warm water bath, adjusted
to the temperature prescribed by the method. Final evaporation may also
be accomplished in the ampul using a micro Snyder column to give a
final volume of 0.2-0.4 ml. In the latter case, a small sand-size
boiling chip is added to the ampul prior to evaporation. The extract
volume is reduced to 0.1 ml below the volume sought so that the internal
wall of the ampul may be rinsed. This step is carried out at least
three times. Great care must be exercised to prevent the extract from
going to dryness.
3. Gas-Liquid Chromatography
3.1 Gas Chromatographic System - The gas chromatographic system em-
ployed must be demonstrated to be suitable for the determination of pes-
ticides with a minimum of decomposition and loss of compounds of interest.
The analyst must evalup'... the individual system to demonstrate such capa-
bility. Detectors, col-uP Backings, column conditioning as well as sug-
gested operating conditions are given below.
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3.1.1 Injection Systems - Many organic compounds, pesticides in par-
ticular, decompose at elevated temperatures when they come in contact with
stainless steel. To prevent this decomposition, the inlet port of the gas
chromatograph must be capable of accepting a quartz or pyrex glass insert.
To avoid bleed off that may cause high background or discrete interferences,
the septa should be preconditioned prior to use. The septa can be treated
by heating in a vacuum oven at 250 C for two hours; changing ...he septum
at the end of each work day to allow overnight purging of the system is
also good practice.
3.1.2 Detectors - The analyst must thoroughly acquaint himself with
the descriptive and theoretical information on the detectors employed.
Review of technical papers and manufacturer's instructions on a specific
detector is necessary to become familiar with its use and limitations.
3.1.2.1 Electron Capture Detector (EC) - The electron capture
detector is extremely sensitive to electronegative functional groups,
such as halides, conjugated carbonyls, nitriles, nitrates, and organo-
metallics. It is virtually insensitive to hydrocarbons, amines, alco-
hols and ketones (10) (11). The selective sensitivity of halides makes
this detector particularly valuable for the determination or organo-
chlorine pesticides. It is capable of detecting picogram (10" gram)
quantities of many organochlorine pesticides. Organophosphorus pesti-
cides containing nitro- groups are also detected, although with much
less sensitivity.
Electron capture detectors may be of parallel plate and concentric
tube or concentral design and employ one of two ionization sources:
tritium (H ) or radioactive nickel (Ni ). The tritium detector has
temperature limit of 225 C (It should not be operated above 210 C.)
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which makes it susceptible to a buildup of high boiling contaminants which
reduce its sensitivity and require frequent clean-up. The nickel detector,
on the other h.ind, can be operated or baked out up to 400 C, to reduce
contamination and cleaning problems.
3.1.2.2 Microcoulometric Titration Detector (MC) - The microcoulo-
metric detector (12) is selective for halogen containing compounds, ex-
cept fluorides, when used with the halogen cell. Under optimum conditions,
this detector is capable of detecting 5-20 ng of organochlorine pesticides.
Although the sensitivity of this detector is not as great as that of elec-
tron capture, the high degree of specificity makes it a very valuable in-
strument for qualitative identification as well as for minimizing sample
clean-up. Under the proper oxidative-reductive conditions, the system can
be made specific for sulfur, phosphorus, and nitrogen compounds.
3.1.2.3 Electrolytic Conductivity Detector (BCD) - The electrolytic
conductivity detector has a sensitivity 2 to 3 times greater than the
microcoulometric system. Although perhaps slightly less selective than
the MC, it is, nonetheless, effective for qualitative identification, and
cleanup appears to be less of a problem (13). Use of tne electrolytic
conductivity detector in the reductive mode with a platinum catalyst is
recommended when determining halogen compounds. If the oxidative mode is
used, a scrubber must be employed to remove SO , which also responds to
the detector.
3.1.2.4 Flame Photometric Detector (FPD) - The flame photometric
detector is selective for sulphur and phosphorus (14). With the use of
a dual head, the detector is capable of simultaneously measuring both sulfur
and phosphorus as well as a normal flame ionization response. Using a
single head, either sulfur or phosphorus and normal flame ionization
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response is measured. The characteristic optical emissions of sulfur
and phosphorus are measured using filters with transmission at 394 mg
(sulfur) and 526 m\i (phosphorus). The FPD is capable of detecting sub-
nanogram quantities of both sulfur (4 x 10 gram) and phosphorus
(10'U gram).
3.1.3 Columns - A well-prepared column is essential to an acceptable
gas chromatographic analysis. The most advanced gas chromatographic in-
strumentation available is no better than the column used with it. A
well-conditioned efficient column is a must. Column packings may be pre-
pared by the analyst or purchased already prepared from a supply house
that specializes in this service. The packing materials and column di-
mensions selected by EPA are specified under the individual methods in
Part II.
3.1.3.1 Preparation of Column Packing - The analyst who prepares his
own packing must develop a highly refined technique to produce consistently
good efficient columns. Particular care should be taken to accurately
measure loadings, uniformly distribute the liquid phase, and to preserve
the structure of the fragile solid support. Improved column efficiency is
obtained when the solid support is dried at 100 C overnight prior to
coating. Several methods may be employed to coat the packing material:
slurry (15), filtration (16), and frontal analysis (17).
The slurry technique consists of dissolving a weighed amount of the
liquid phase in an appropriate solvent in a beaker and slowly pouring
the weighed solid support into it with constant stirring. The beaker
is immersed to the level of the solvent in a hot water bath and gently
stirred until the bulk of the solvent has evaporated. Extra care must
be exercised to minimize crushing of the solid support. The filtration
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technique consists of mixing the solid support with the solution of
liquid phase as above then carefully pouring into a Buchner funnel
fitted with a filter flask. The excess solution is removed by vacuum.
The frontal analysis technique consists of passing the solution through a
column of the solid support until the effluent from the column is of
the same composition as the original solution. The analyst should
select the method that provides the best results for him. Drying of
the coated support may be accomplished by spreading on a tray in a
convection or vacuum oven at 100-120 C, with a rotary evaporator (18)
or with a fluidized bed drier (19). The latter is recommended.
3.1.3.2 Column Material and Dimensions - The column should be con-
structed of borosilicate glass. The most useful length of column is
about 6 ft. with a diameter of 1/4 in. O.D. to 1/8 in. O.D., depending
on the detector employed and the volume of sample injected. Electron
capture detectors of parallel plate design and the Tracor concentral
design perform well with either 1/4 in. or 1/8 in. columns, while the
Varian-Aerograph concentric tube design operates best with a 1/8 in.
column. A 1/4 in. column for the microcoulometric detection system is
recommended.
3.1.3.3 Packing the Column - It is important that the column be
packed to a uniform density not so compact as to cause unnecessary back
pressure and not so loose as to create voids during use. Care should
be exercised so as not to crush the packing. Column tubing should be
rinsed with solvent, eg. chloroform, and dried prior to packing.
Columns are filled through a funnel connected by flexible tubing to one
end. The other end of straight or coiled tubing is plugged with about
1/2 in. of silanized glass wool and filled with the aid of gentle vibra-
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14
tion or tapping. A mild vacuum may also be applied to the plugged end.
When filled the open end is also plugged with silanized glass wool. In
a similar manner, one-half of a "U" shaped column is filled and then the
other and the ends are plugged with silanized glass wool.
3.1.3.4 Column Conditioning - Proper thermal conditioning is
essential to eliminate column bleed and to provide acceptable gas
chromatographic analyses. A number of procedures may be used for this
purpose. The procedure described below is used by the Analytical
Quality Control Laboratory with excellent results:
Install the packed column in the oven. Do not connect the column to
the detector. However, gas flow through the detector should be main-
tained. This can be done using the diluent gas line or, in dual column
ovens, by connecting an unpacked column to the detector. Heat the oven
to near the maximum recommended temperature for the liquid phase without
gas flow for 2 hours. Reduce the oven temperature to approximately 40 C
below the maximum recommended temperature and allow temperature to equili-
brate for a minimum of 30 minutes still without flow. Then adjust the
carrier gas flow to about SO ml per minute for a 1/4 inch column and
about 25 ml per minute for a 1/8 inch column. (Caution—bleed off of
liquid phase will occur if not fully temperature equilibrated.) After
one hour, increase the temperature to about 20 C above normal operating
temperature with gas flow for 24-48 hours. (Do not exceed maximum recom-
mended operating temperature.) Cool down and connect column to the detector
system, then raise to normal operating temperature. Columns prepared and
conditioned in this manner should yield good chromatograms with no further
treatment.
3.1.3.S Optimizing Operating Conditions - The analyst must determine
the optimum conditions for obtaining the best results for the compounds
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15
under study. Standard mixtures of interest should be chromatographed to
determine retention times and maximum resolution that can be achieved.
Parameters such as gas flow, temperature, column length and diameter,
as well as the electronics and detector performance, must be evaluated
and adjusted as required to achieve the desired results. Other important
requirements for attaining optimum operating conditions include a clean
injection block and a leak-free pneumatic system. The instrument must be
operated within the linear range of the detector. The recorder gain and
damping adjustments must be optimized. Regulated electric line current
may be required for the electronic system. Detailed instructions for
carrying out these operations are given in the manufacturer's instrument
manuals.
Optimum detector sensitivity for each :.iethod is defined as the
minimum acceptable response to a designated amount of a selected compound.
The detector response for other compounds that may be determined by the
method relative to the selected compound are listed in each method.
Continued optimum performance is maintained by following the routine
maintenance and check program given in the instrument manuals. Frequent
checks on the injection block, oven, and detector temperatures should be
made. To avoid the risk of system or detector contamination from impurities
in gas cylinders, replace the cylinders when the pressure reaches 200 psi.
Use of molecular sieve gas-filter driers on all gases is recommended.
Column performance is monitored by observing daily response to a
selected standard mixture and comparing it to the response obtained under
previously established conditions. Changes in elution pattern, relative
proportions of peaks, and peak geometry are signs of a deteriorating column,
when all other parts of the system are properly maintained. Columns should
be replaced when deterioration is observed.
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3.2 Injection into the Gas Chromatograph
3.2.1 Loading Syringe and Measuring Volume Injected - The analyst
must develop the ability to make accurate and reproducible injections
into the gas chromatograph. Several techniques may be used. The
analyst should select the one that provides the best results for him.
One technique, preferred by some analysts is as follows:
Wet syringe needle and barrel with solvent solution of the standard or
sample to be injected and expel all air bubbles. Draw the entire quan-
tity of solution into the calibrated barrel and note volume. Inject
into the chromatograph rapidly and withdraw syringe immediately. Then
partially withdraw plunger and note volume remaining. Determine volume
injected by subtracting this volume from the original volume. It is
important that the syringe be thoroughly cleaned after each injection.
Usually, several solvent rinses are adequate.
3.2.2 Injection of Standard Solutions - The concentration of stan-
dard solutions should be such that the injection volume of the standard
is approximately the same as that of the sample.
3.3 Qualitative Analysis - Qualitative identification of an unknown
component is made by matching the retention time (R ) of the unknown
with that of a standard obtained under identical conditions. The R
is the time lapsed from injection (time zero) to the peak maximum. The
absolute and relative retention time (RRt) are commonly recorded. The
RR is defined as the R (component) * Rt (reference compound). When
solvent response is observed, as with an electron capture, the leading
edge of the solvent peak is considered time zero. When no solvent
response is observed, as with a microcoulometric detector, time zero
is electrically or manually marked immediately after injection.
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3.3.1 Confirmatory Identification - The analyst must be aware that
a single gas chromatographic determination does not provide unequivocal
identification of an unknown component. The retention time and peak
geometry must be matched on two or more unlike columns. Co-injection
of the sample with a standard of the suspected compound will assist in
confirming the qualitative identification. Clean-up and separation
techniques such as thin-layer and column chromatography also help to
make the qualitative assignment. Identification of multicomponent
pesticides requires not only matching of all retention times and
overall peak geometry, but also the correct number and relative propor-
tion of each peak - a so called "fingerprint" of the pesticide. Further
corroboration using infrared spectroscopy and/or mass spectrometry
should be obtained whenever possible.
3.4 Quantitative Analysis - The quantity of compound present is
proportional to the area of the peak and can be used to determine the
concentration of the components in a sample. The area measurement is
usually preferred; however, peak height measurement may be more accurate
when sharp narrow peaks occur. Peak area may be measured by electronic
integrator, disc integrator, or by planimeter or may be calculated by
taking the peak height x peak width at half height. The planimeter,
although less precise than the other techniques, is recommended for
measuring the area of unsymmetrical peaks that do not originate at the
original baseline. To improve precision, measure area several times and
take the average value.
3.4.1 Unresolved Peaks - To quantitate peaks that are not completely
resolved, inject standards of the suspected compounds mixed in the same
ratio as they occur in the sample and giving response equivalent to that
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18
of the sample. An alternative method is to draw a line perpendicular
from the baseline to the low point of the valley between the peaks.
Shoulders on larger peaks may be measured, although not accurately, by
using as t.he baseline a line drawn to conform to the shape of the major
peak. Resolution may sometimes be accomplished usinvj different G.C.
columns and/or by preliminary separation using thin-layer and column
chromatography.
3.4.2 Standard Calibration
3.4.2.1 Absolute Calibration (20) - Using the absolute method,
pesticide concentrations are determined by direct comparison to a single
standard when the injection volume and response are very close to that
of the sample. The concentration of pesticide in the sample is calcu-
lated as follows:
micrograms/liter = fv'1) {\i \
A = ng std
std area
B = sample aliquot area
Vi = volume of extract injected
Vt = volume of total extract (yl)
Vs = volume of water extracted (ml)
3.4.2.2 Relative Calibration (Internal Standardization) (20) - A
relative calibration curve is prepared by simultaneously chromatograph-
ing mixtures of the previously identified sample constituent and a
reference standard in known weight ratios and plotting the weight ratios
against area ratios. An accurately known amount of the reference mater-
ial is then added to the sample and the mixture chromatographed. The
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19
area ratios are calculated and the weight ratio is read from the curve.
Since the amount of reference material added is known, the amount of
the sample constituent can be calculated as follows:
micrograms/liter = Rwy* Ws
Rw = Weight ratio of component to standard
obtained from calibration curve,
Ws » Weight of internal standard added to
sample in nanograms
Vs = Volume of sample in milliliters
Using this method, injection volumes need not be accurately measured
and detector response need not remain constant since changes in response
will not alter the ratio. This method is preferred when the internal
standard meets the following conditions:
a) well-resolved from other peaks
b) elutes close to peaks of interest
c) approximates concentration of unknown
d) structurally similar to unknown.
3.4.3 Linear range - Accurate quantitative analysis depends upon a
linear relationship between concentration and detector response. The
closer the linear relation the more accurate the analysis. The analysis
range of a detector is defined as the ratio of the largest to the smallest
concentration within which the detector is linear.
4. Column Chromatography
4.1 Adsorbents - A variety of adsorbents are used in the various
pesticide methods to remove interferences and to separate individual
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20
pesticides. These are usually purchased preactivated from the manu-
facturer. The adsorptivity of the adsorbent is checked by determining
the elution pattern of specified dyes and/or of given pesticides. Re-
coveries of the pesticides must be determined prior to using the adsor-
bent for the analysis of samples.
4.1.1 Florisil - Florisil preactivated by the manufacturer at 1200
F is used. Prior to use, the Florisil is heated for at least S hours
(overnight is convenient) at 130 C. Although the adsorbent tends to yellow
when stored in this manner for several days, it remains satisfactory for
use.
4.2 Packing the Column - To pack the column, slowly pour the adsor-
bent into the column while vigorously tapping it. This will assist in
providing a uniform packing and minimize :hinneling during elution.
4.3 Eluting the Column - Liquids should be poured slowly down the
inside wall of the column to avoid disturbing the surface of the adsor-
bent. Mixing of solvents above the adsorbent is minimized by adding
succeeding solvents just as the last of the previous solvent reaches the
adsorbent surface. However, the surface of the adsorbent must not be
allowed to run dry, since introduction of air may cause channeling and
reduce the efficiency of separation.
4.3.1 Pre-elution - Prior to addition of the sample, the column is
pre-eluted with the solvent prescribed by the procedure. This is done to
remove trapped air and trace contaminants that may interfere with the
analysis. It may be necessary to tap the column to free all of the trapped
air.
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4.3.2 Introduction and Elution of Sample - The sample is intro-
duced just as the last of the pre-eluting solvent reaches the surface
of the adsorbent. The sample container is then rinsed with a few ml
of the solvent and the rinse added to the column. Just as the last of
this solvent reaches the surface a small volume of eluting solvent is
used to rinse down the internal wall of the column. Then the remaining
eluting solvent is added. Successive eluting solvents are added in a
similar manner.
4.3.3 Eluate Composition - Since variations in elution pattern may
occur from tir. to time, it is necessary to demonstrate that the eluate
composition is proper for a given analyses. This can be done by eluting
standard pesticides from the column and/or by using the activity test
given in the "Official Methods of Analysis of the AOAC" (3).
S. Thin-Layer Chroroatography
5.1 Equipment - Special equipment required for preparing layers and
carrying out thin-layer chromatography is listed in the appendix. Layers
prepared in the laboratory or purchased precoated layers may be used. The
adsorbent is usually less tightly bound to the plate when prepared in the
laboratory and is thus somewhat easier to scrape for subsequent elution
and recovery of the pesticides.
S.2 Layer Preparation - Layers of desired thickness are prepared by
making a homogeneous slurry of adsorbent in water. The slurry is poured
into the applicator and the gate is quickly opened and the applicator is
smoothly and rapidly passed over an aligning tray holding the glass plates.
The layers are allowed to stand at room temperature for a time, then activated
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in an oven, and stored in a desiccator for future use. Layers stored
longer than one week should be reactivated before use. Prior to acti-
vation, marks should be made on the layer to define the spotting line and
the upper limit of solvent development in order to minimize exposure to
the atmosphere during the spotting operation.
5.3 Preparation of Developing Chamber - The developing solvent is
added to the chamber, and two chromatography paper wicks, one on each of
the long sides of the chamber, are placed so that the entire side is
covered and the bottom edge contacts the solvent. The chamber is closed,
shaken, and allowed to equilibrate. It is important that the chamber be
protected from drafts and large temperature changes.
5.4 Spotting the Layer - Standards are spotted in the center and at
least at one edge of the layer. The standards and samples should be dis-
solved in the same solvent and spotted in the same volume. Utmost care
must be exercised to keep the spot small (less than 10 mm diameter). A
gentle stream of clean dry air or nitrogen may be applied over the spot to
facilitate close boundary evaporation, but this gas flow exposure should
be kept to a minimum.
5.5 Developing the Layer - The spotted layer is placed in the pre-
equilibrated chamber so that the bottom edge is in contact with the
developing solvent and the lid is replaced. When the developing solvent
reaches the upper reference mark, the layer is removed from the chamber and
allowed to air-dry at room temperature.
5.6 Visualizing and Sectioning the Layer - After development, the
portion of the layer containing the standards is sprayed evenly with a
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chromogenic agent. Hie sprayed area is allowed to thoroughly dry and,
where required, is further treated by exposure to short wave UV light
or some other means. The location of each standard pesticide is marked.
5.6.1 The distance of travel for pesticides present in the unknown
samples and recovery test standards will be, respectively, the same as
those of the sprayed standards. Using this information, the vertical
zone for each sample is divided into horizontal sections depending on the
pesticides being determined.
5.7 Pesticide Removal from the TLC Plates - With the aid of a sharp
pointed object, the silica gel sections of interest are individually
ruled off. With the aid of a mild vacuum, the silica gel, first from the
periphery of the section and then from the center of the section is col-
lected. It is convenient to use a medicine dropper plugged at the tip
with filtering grade glass wool (Figure 1). The pesticides are eluted
quantitatively into a K-D ampul with a selected solvent to an appropriate
volume for gas chromatographic analysis.
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PART II - METHODS OF ANALYSIS
A. METHOD FOR ORGANOCHLORINE PESTICIDES
1. Scope and Application
1.1 This method covers the determination of various organochlorine
pesticides, including some pesticidal degradation products and related
compounds. Such compounds are composed of carbon, hydrogen, and chlor-
ine, but may also contain oxygen, sulfur, phosphorus or nitrogen.
1.2 The following compounds may be determined individually by this
method: BHC, lindane, heptachlor, aldrin, heptachlor epoxide, dieldrin,
endrin, Perthane, DDE, ODD, DDT, methr>v.ychlor, endosulfan, f-chlordane
and sulphenone. Under favorable circumstances, Strobane, toxaphene,
kelthane, chlordane (tech.) and others may also be determined.
1.3 When organochlorine pesticides exist as complex mixtures, the
individual compounds may be difficult to distinguish. High, low, or
otherwise unreliable results may be obtained through misidentification
and/or one compound obscuring another of lesser concentration. Provisions
incorporated in this method are intended to minimize the occurrence of such
interferences.
2. Summary
2.1 The method offers several analytical alternatives, dependent on
the analyst's assessment of the nature and extent of interferences and
the complexity of the pesticide mixtures found. This method is recommended
for use only by experienced residue analysts or under the close supervision
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25
of such qualified persons. Specifically, the procedure describes the
use of an effective co-solvent for efficient sample extraction; provides,
through use of thin-layer, column chromatography, and liquid-liquid par-
tition methods for the elimation of non-pesticide interferences, and
the pre-separation of pesticide mixtures. Identification is made by
selective gas chromatographic separations through the use of two or more
unlike columns. Detection and measurement is accomplished by electron
capture, microcoulometric or electrolytic conductivity gas chromatography.
Techniques for confirming qualitative identifications are suggested. Re-
sults are reported in micrograms per liter without correction for recovery
data; but, such data is to be included in the report.
3. Significance
3.1 The extensive and widespread use of persistent organochlorine
pesticides has resulted in their presence in all parts of our environment.
Their occurrence in surface waters throughout the country is common. Such
common occurrence coupled with the toxic nature of these materials is cause
for concern. The known lethal effects of these substances to fish and
wildlife and the unknown long term consequences to humans make it imperative
that we identify and quantitate the pesticides present in the environment.
Effective evaluation and control programs require such information.
3.2 Because of the concept of biological concentration, we need to
detect minute quantities (low nanogram amounts) of pesticides in water.
The method presented here is capable of detecting these small quantities.
4. Interference
4.1 Solvents, reagents, glassware, and other sample processing hardware
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26
may yield discrete artifacts and/or elevated baselines causing mis-
interpretation of gas chromatograms. All of these materials must be
demonstrated to be free from interference under the conditions of the
analysis. Specific selection of reagents and purification of solvents
by distillation in all-glass systems is required. (Refer to Part 1,
Sections 1.4 and 1.5)
4.2 Sample treatment required to remove non-pesticide materials which
cause interference may result in the loss of certain organochlorine
pesticides. Methods for eliminating or minimizing interferences are de-
scribed below in the section on Clean-up and Separation Procedures (5.2).
It is beyond the scope of this method to describe procedures for overcoming
all of the possible interferences that may be encountered, particularly,
in highly contaminated water and wastewater.
4.3 Polychlorinated Biphenyls - Special attention is called to
industrial plasticizers and hydraulic fluids such as the chlorinated bi-
phenyls (PCB's, Aroclors ) which are a potential source of interference in
pesticide analysis. Chlorinated biphenyls containing 4 to 8 chlorine atoms
per molecule have been reported in extracts of birds, fish, mussels and
water. Possible interferences from these compounds are indicated by un-
resolved peaks (shoulders and nongaussian peaks), slight discrepancies
in retention times, and peaks which elute later than p.p'-DDT. With some
dependence upon the relative concentrations, a number of chlorinated
biphenyl isomers may interfere with the determination of DDE, ODD and DDT
isomers. The authenticity of the DDT identification may be determined by
treating the extvact with alcoholic KOH which converts DDT to DDE (21).
Tradename of the Monsanto Company, St. Louis, Missouri
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27
Particularly severe PCB interference will require special separation
procedures, such as those of Reynolds (22), Armour and Burke (23), and
Mulhern, et al (24).
4.4 Phthalate Esters - These compounds, widely used as plasticizers,
respond to the electron capture detector and are a source of interference
in the determination of organochlorine pesticides using this detector.
Water leaches these materials from plastics, such as polyethylene bottles
and tygon tubing. The presence of phthalate esters is implicated in samples
that respond to electron capture but not to the microcoulometric or electro-
lytic conductivity halogen detectors or to the flame photometric detector,
since these materials are not detected by the latter three detectors.
4.5 Organophosphorus Pesticides - A number of organophosphorus
pesticides, notably those containing a nitro group, eg, parathion, also
respond to the electron capture detector and may interfere with the deter-
mination of the organochlorine pesticides. The presence of such compounds
is indicated in samples which respond to both the electron capture and
flame photometric detectors but not to the microcoulometric halogen detector.
5. Method for Analysis Using Electron Capture Gas Chromatography
S.I Extraction of Sample
5.1.1 The size of sample taken for extraction is dependent on the type
of sample and the sensitivity required for the purpose at hand. Background
information on the pesticide levels previously detected at a given sampling
site will help to determine the sample size required as well as the final
volume to which the extract needs to be concentrated. A 1-liter sample is
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28
usually taken for electron capture analysis. The extract should not be
concentrated further than required to meet the sensitivity dictated by the
purpose for the analysis. Each time a set of samples is extracted, an
aliquot of solvent equivalent to that used for extraction is carried through
the entire procedure to provide a method blank. To assist in interpretation
of results, the pH of the sample is taken prior to extraction. When the volume
of the sample permits, one set of duplicates and one do ;ed sample should also
be analyzed as a quality control check.
S.I.2 A measured volume (1-liter) of sample is drained into a 2-liter
separatory funnel equipped with a Teflon stopcock, and extracted with 60 ml
of 15% ethyl ether in hexane by shaking vigorously for two minutes. The
sample container is rinsed with each aliquot of extracting solvent prior to
extraction of the sample.
S.I.3 The mixed solvent is allowed to separate from the water; the water
is then drawn into the original sample container or into a second 2-liter
separatory funnel. The organic layer is passed through a small column of
anhydrous sodium sulfate topped with a pledget of cotton (previously rinsed
with hexane) and collected in a 500 ml Kuderna Danish flask equipped with a
10 ml ampul. The extraction is repeated and the solvent treated as above.
Approximately 35 ml of sodium sulfate saturated water is then added to the
sample and a third extraction is completed with 60 ml of hexane (not hexane-
ethyl ether). This solvent, too, is passed through the sulfate column and
collected in the flask. The column is rinsed with several small portions of
hexane and this solvent is recovered in the collection flask containing the
combined extracts. The extract is concentrated in the Kuderna-Danish evapora-
tor as described in Part I, Section 2.3.1.
5.1.4 Concentration of Extract - The final concentration volume for
samples of high pesticide content (eg. pesticide plant wastewater samples)
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29
is adjusted as necessary. Samples containing small quantities of
pesticides (low nanogram amounts, eg. most surface water samples) are
concentrated to 1 ml. The volume of the initial K-D concentrate is 5 to
6 ml. This is reduced to 1 ml in a warm water bath (70 C) with a gentle
draft of clean dry air or nitrogen. The internal wall of the ampul is
rinsed several times during this operation. Initial gas chromatographic
analysis is made on this volume. Up to 10 pi of extract are injected.
If insufficient pesticide is present for detection at this volume and
greater sensitivity is required, the extract is concentrated further to
a minimum volume of 0.2 ml in the manner described above. The extract
volume is reduced below the volume sought so that the internal wall of
the ampul may be rinsed. (See Part I, Section 2.3.2) (The volume should
never be reduced below 0.1 ml). Repeat this operation three times, exer-
cising great care to prevent the extract from going to dryness.
5.2 Clean-up and Separation Procedures
5.2.1 Interferences in the form of distinct peaks and/or high back-
ground in the initial gas chromatographic analysis, as well as, the physical
characteristics of the extract (color, cloudiness, viscosity) will indicate
whether clean-up is required. When these interfere with measurement of the
pesticides, proceed as directed below. Whether required for quantitative
analysis or not, all extracts should be subjected to these procedures,
subsequent to the initial analysis and rechromatographed for qualitative
corroboration of the results. Another clean-up technique, acetonitrile
partition, although not ordinarily required for surface water extracts, is
sometimes useful for cleaning up high organic wastewater samples. Refer to
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the FDA methods manual (1) for this procedure.
S.2.2 Florisil Column Adsorption Chromatography - The sample extract
previously concentrated to 1 ml or less is diluted to 10 ml. A 15 g charge
of activated Florisil is placed in a column over a small layer (one-half
inch) of anhydrous granular sodium sulfate. After tapping the Florisil into
the column, about a three-fourths inch layer of granular sodium sulfate is
added to the top. The column, after cooling, is pre-eluted with about 75 ml
of hexane. The pre-eluate is discarded, and just prior to exposure of the
sulfate layer to air, the sample extract is quantitatively transferred into
the column by decantation and subsequent hexane washings. The elution rate
is adjusted to about 5 ml per minute with two eluates collected separately
in 500 ml K-D apparatus equipped with 10 ml ampuls. The first elution is
performed with 200 ml of 6% ethyl ether in hexane, and the second elution with
200 ml of 15% ethyl ether in hexane. The K-D apparatus containing the
eluates are connected to three ball Snyder columns and the solvents are
evaporated as described in Part I, Section 2.3. The concentrated extract
may be analyzed directly by injecting suitable aliquots from the K-D ampuls
into the gas chromatograph. If the residues are high in total organics, they
may be further cleaned up and separated by thin-layer chromatography prior
to gas chromatographic analyses.
5.2.2.1 Eluate Composition - If the Florisil has been properly acti-
vated and stored, and if the reagents are carefully prepared, the following
eluate compositions will be obtained when the pesticides are present. The
first eluate (6% ethyl ether in hexane) will contain:
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31
lindane DDE methoxychlor
BHC ODD toxaphene
kelthane DDT Strobane
aldrin Perthane chlordane (y 6 tech)
heptachlor heptachlor epoxide endosulfan I
The second eluate (15% ethyl ether in hexane) will contain:
dieldrin endosulfan II
endrin lindane (possible trace of total)
kelthane (possible trace of total)
Pol/chlorinated biphenyls are recovered in the first eluate and phthalate
esters in the second eluate.
5.2.3 Thin-Layer Chromatography - The sample extract (5,1.4) or the
eluates from the Florisil clean-up (5.2.2) may be subjected to thin-layer
chromatography according to the procedure described below. (Refer to Part I,
Section 4.3 for general discussion and conditions for preparation of thin-
layers). Silica gel G layers, 250 u thick, are employed.
5.2.3.1 Spotting the Extract - The extract volume should be adjusted
so that no more than 100 ul must be spotted in order to retain adequate GC
sensitivity in the TLC eluates when they are reduced to the minimum volume of
0.2 ml. Up to 100 yl of the eluate is then spotted on the thin-layer.
5.2.3.2 Spotting of Standards - A mixture of reference standard
pesticides is spotted at the center and at one edge of the layer, in 10-20 ug
amounts of each pesticide, to confirm the separation of the individual
pesticides by visual observation. It is convenient to use a 100 ng/yl mixture
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32
of encrin, lindane, ODD and DDT. Standards for recovery and instrumental
measurement are spotted in the 20 to 100 nanogram range and handled just
as a sample would be treated in the subsequent steps of the procedure.
5.2.3.3 Developing the Layer - The layer is developed with carbon
tetrachloride. When the solvent front reaches the upper reference mark
(10 cm), the layer is removed from the chamber and allowed to air dry at
room temperature. The r"3veloping solvent should be checked frequently for
contamination and changed as necessary.
5.2.3.4 Visualizing and Sectioning the Layer - The portions of the
layer containing the samples are covered with a glass plate or cardboard.
The portion of the layer containing the standards is sprayed evenly with a
fairly heavy coat of Rhodamine B (0.1 mg/ml in ethanol). The sprayed area
is allowed to thoroughly dry (about 5 minutes) and then is exposed to and
viewed under short wave UV light. The pesticides show up as quenched areas
(dark) on a fluorescent background. Mark the location of each pesticide.
From this information, the vertical zone for each sample is divided into five
horizontal sections. The sections are identified with Roman numerals as
shown in Figure 2. Examples of respective Rf and Rr values for various
pesticides are listed in Table 1.
5.2.3.5 Removal of Pesticide From the TLC Layer - Using the spotting
template as a ruler, and with the aid of a sharp pointed object, the silica
gel sections of interest are individually ruled off. With the aid of a mild
vacuum, the silica gel, first from the periphery of the section and then
from the center of the section, is drawn into a medicine dropper which is
plugged at the tip with filtering grade glass wool (Figure 1). The pes-
ticides adsorbed on this silica gel are eluted quantitatively into a 10 ml
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33
K-D ampul with successive small washes of ethyl ether-petroleum ether (1+1)
to a total volume of 5 to 10 ml. The ampul is glass stoppered and the con-
tents are retained for gas chromatographic analysis. Prior to gas chromato-
graphic analysis, the extracts are concentrated as required. Refer to 5.1.4.
5.3 Gas Liquid Chromatography
5.3.1 Reasonably positive identification of a pesticide is obtained
by corroborating the results using, at least two different types of gas
chromatographic columns. To achieve this, a relatively less polar packing
[5% OV-17 on Gas Chrom Q (80-100 mesh)] and a more polar packing [5% QF-1
(FS-1265) plus 3% DC-200 on Gas Chrom Q (80-100 mesh)] are employed. Other
packings reconmended for this purpose are 3% OV-101 on Gas Chrom Q (80-
100 mesh) and 3% OV-210 on Gas Chrom Q (80-100 mesh). Packings and packed
columns can be obtained from commercial sources or may be prepared in the
laboratory.
5.3.2 Preparation of Columns - To prepare the column packings, dis-
solve 5 g of OV-17 in 225 ml of methylene chloride - chloroform (1+1) in
a 500 ml beaker and add 95 g of Gas Chrom Q. Similarly, dissolve 5 g of
QF-1 plus 3 g of DC-200 in methylene chloride - chloroform (1+1) and add
92 g of Gas Chrom Q. Dissolve 3 g of OV-101 in chloroform and add to 97 g
of Gas Chrom Q. Dissolve 3 g of OV-210 in acetone and add to 97 g of Gas
Chrom Q. Proceed as described in Part I, Section 3.1.3.1.
5.3.2.1 Columns of borosilicate glass, 6 ft. long x 1/4 in. O.D.
(5/32 in. I.D.) or 1/8 in. O.D. (1/16 in. I.D.) are packed and conditioned
according to directions in Part I, Sections 3.1.3.3 through 3.1.3.5. The
column o.ven operating temperature is approximately 210 C for the OV-17
column, 185 C for the QF-l/DC-200 column, 190 C for the OV-101 column, and
185 C for the OV-210 column. The operating conditions are optimized for
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34
the individual instrument as described in Part I. Section 3.1.3.5. Opera-
ting conditions are considered acceptable for an electron capture system
when the response to 0.3 ng of aldrin is at least 50% of full scale while
operating within the linear range of the detector,
5.3.3 Sample Measurement - The volume of sample extract (5.14), the
Florisil eluates, or the TLC eluates is noted and suitable aliquots
(5-10 jil) are analyzed by gas chromatography, employing at least two columns
of varying polarity for identification and quantitation. Standards are
injected frequently, as a check on the stability of the operating conditions.
Gas chromatograms of several standard pesticides are shown in Figures 3, 4,
5, and 6. The elution order, as well as elution ratios for various pesticides
in Table 2, are provided only as a guide. It is the responsibility of the
analyst to develop his own identification keys to fit the chosen operating
conditions of the instrument.
5.4 Confirmatory Evidence - The qualitative identification of a
pesticide should be confirmed using infrared spectroscopy or mass spec-
troscopy whenever the instrumentation is available and/or the quantity of
the compound permits. If this is not possible, gas chromatographic analysis
using additional unlike columns and other selective detectors is recommended.
Lack of response to the flame photometric detector is negative evidence
which supports the identification of organochlorine compounds. Determination
of the p-values of an unknown pesticide in several solvent systems will
assist in confirming the identification (25).
5.5 Calculation of Results - The pesticide concentrations are deter-
mined using the absolute or the relative calibration procedure described in
Part I, Section 3.4.2.
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5.6 Reporting Results - Report results in micrograms per liter with-
out correction for recovery data. The percent recoveries of known pesti-
cides added to samples or to distilled water as well as the step in the
procedure where they were added must also be reported. The recoveries of
several organochlorine pesticides from natural waters during collaborative
testing are listed in Table 3. The precision of the method within the
designated range varies with the concentration as shown in Table 4.
5.6.1 If a sample is reported negative for a given pesticide, the
minimum detectable limit for that compound should also be reported. If
favorable conditions prevail and ultimate sensitivity is required by the
purpose for the analysis, sample response of less than two times the de-
tector noise level (N) should be reported as negative. For sample response
at two times the detector noise level, list the result as presumptive.
Responses of greater than 2N should be quantified if possible. In cases
of questionable identification, the analyst should qualify the reported
resUiC to insure the subsequent misinterpretation will not occur.
6. Method for Analysis Using Microcoulometric or Electrolytic Conductivity
Gas Chromatography
6.1 Extraction of Sample
6.1.1 The size of sample taken for extraction is dependent on the
type of sample and the sensitivity required for the purpose at hand. Back-
ground information on the pesticide levels previously detected at a given
sampling site will help to determine the sample size required as well as the
final volume to which the extract needs to be concentrated. A 3-liter sample is
usually taken for microcoulometric analysis. Since the conductivity
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36
detector is 2 to 3 times more sensitive than the microcoulometric
detector, less than 3 liters may be required when using the conductivity
detector. If such isthe case, the volume of extracting solvents and other
reagents are correspondingly decreased.
6.1.2 A measured volume (3 liters) of sample is drained into a 4-liter
separatory funnel equipped with a Teflon stopcock, and extracted with 150 ml
of 15% ethyl ether in hexane by shaking vigorously for two minutes. The
sample container is rinsed with each aliquot of extracting solvent prior to
extraction of the sample.
6.1.3 The mixed solvent is allowed to separate from the water and this
water is drawn off into the original container or into a second 4-liter
separatory funnel. The organic layer is passed through a small column of
anhydrous sodium sulfate topped with a pledget of cotton (previously rinsed
with hexane) and collected in a 600 ml tall form beaker. The extraction is
repeated and the solvent treated as above. Approximately 100 ml of sodium
sulfate saturated water is then added to the sample and a third extraction
is completed with 150 ml of hexane (not hexane-ethyl ether). This solvent
too, after separation, is passed through the column of sodium sulfate. The
column is then rinsed with several small portions of hexane and this solvent
is recovered in the collection beaker containing the combined extracts. The
contents of the beaker are partially evaporated to about 300 ml in a water
bath at 70 C applying no air or vacuum and quantitatively transferred to a
500 ml K-D evaporator equipped with a 10 ml receiver ampul. The extract is
concentrated in the K-D evaporator as described in Part I, Section 2.3.1.
6.1.4 Concentration of Extract - The final concentration volume of
sample extract is adjusted as necessary according to 5.1.4. The use of
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37
a "keeper" is recommended when concentrating below 0.3 ml. Two milli-
grams of "keeper" is placed in the concentrated extract through syringe
addition of 100 pi of 20 yg/pl of Nujol in hexane. This "keeper" will not
interfere with mocrocoulometric detection and will prevent major residue
losses in this exhaustive evaporation step. Because of interference
possibilities, it is not advisable to use a "keeper" in extracts to be
analyzed by electron capture.
6.2 Clean-up and Separation Procedures
6.2.1 Interferences in the form of distinct peaks and/or high back-
ground in the initial gas chromatographic analysis, as well as the physical
characteristics of the extract (color, cloudiness, viscosity) will indicate
whether clean-up is required. When these interfere with measurement of the
pesticides, proceed as directed below. Whether required for quantitative analysis
or not, all extracts should be subjected to these procedures, subsequent to
the initial analysis and rechromatographed for qualitative corroboration of
the results. Another clean-up technique, acetonitrile partition, although
not ordinarily required for surface water extracts, is sometimes useful for
cleaning up high organic wastewater samples. Refer to the FDA methods manual
(1) for this procedure.
6.2.2 Florisil Column Adsorption Chromatography - Refer to 5.2.2.
6.2.3 Thin-Layer Chromatography - Refer to S.2.3.
6.3 Gas Liquid Chromatography
6.3.1 Reasonably positive identification of a pesticide is obtained
by corroborating the results using, at least two different types of gas
chromatographic columns. Refer to 5.3.1.
-------
38
6.3.2 Preparation of Columns - Refer to 5.3.2. Columns of boro-
silicate glass, 6 ft. long x 1/4 in. O.D. are employed. The operating
conditions are optimized for the individual instrument as described in
Part I, Section 3.1.3.5. Conditions are considered optimum when the
response to 15 ng and 30 ng of aldrin is at least 50% of full scale for
the electrolytic conductivity and microcoulometric detectors, respec-
tively, while operating within the linear range.
6.3.3 Sample Measurement - The volume of sample extract (6,1.4),
the Florisil eluates, or the TLC eluates is noted and suitable aliquots
(20-100 yl) are analyzed by gas chromatography, employing at least two
columns of varying polarity for identification and quantitation. Stan-
dards are injected frequently, as a check on the stability of the operating
conditions. Gas chromatograms of several standard pesticides are shown
in Figures 3, 4, 5, and 6. The elution order as well as the elution
ratios for various pesticides are provided in Table 2, only as a guide.
It is the responsibility of the analyst to develop his own identification
keys to fit the chosen operating conditions of the instrument.
6.4 Confirmatory Evidence - Refer to 5.4
6.5 Calculation of Results - The pesticide concentrations are
determined using the absolute or the relative calibration procedure
described in Part I, Section 3.4.2.
6.6 Reporting Results - Refer to 5.6.
-------
TABLE 1
SOME Rf AND Rr VALUES OF PESTICIDES DEVELOPED WITH CC1
4
ON SILICA GEL G THIN LAYER
Pesticide
Dieldrin
Endrin
Heptachlor Epoxide
Lindane
DDD
Y-Chlordane
Heptachlor
DDT
DDE
Aldrin
Rf Value
0.17
0.20
0.29
0.37
0.54
0.55
0.67
0.68
0.72
0.73
Rr Value
0.33
0.37
0.52
0.69
1.00
1.02
1.24
1.26
1.33
1.35
Section
II
III
IV
Rf * distance traveled by the compound divided by the distance
traveled by the solvent front.
Rr • distance traveled by the compound divided by the distance
traveled by standard p,p'-DDD.
-------
TABLE 2
RETENTION TIMES OF ORGANOCHLORINE PESTICIDES RELATIVE TO ALDRIN
Liquid Phase1
Column Temp.
Pesticide
«-BHC
Lindane
Heptachlor
Aldrin
Kelthane
Heptachlor Epoxide
Y-Chlordane
Endosulfan I
p,p'-DDE
Dieldrin
Endrin
o,p'-DDT
Endosulfan 11
p,p'-DDD
p,p'-DDT
Methoxychlor
Aldrin (Minutes
Absolute)
3% DC-200
+
5% QF-1
200 C
RRt3
0.40
0.51
0.80
1.00
1.19
1.38
1.53
1.77
1.93
2.10
2.43
2.62
2.62
2.68
3.41
5.26
3.76
$%
OV-17
200 C
RRt3
0.45
0.6'
0.79
1.00
1.52
1.58
1.82
2.00
2.67
2.54
3.21
3.97
3.97
4.13
5.19
11.17
3.84
3%
OV-101
17S C
RRt3
0.33
0.42
0.76
1.00
1.12
1.30
1.55
1.70
2.18
2.08
2.33
3.02
2.45
2.'94
3.97
6.88
.•» f •
£.V-»
1%
OV-210
160 C
RRt3
0.54
0.75
0.82
1.00
2.46
2.16
2.12
2.89
2.91
3.65
4.46
4.04
5.96
5.61
6.28
13.52
2.28
Relative
Sensitivity
to EC Detector
1.0
1.0
1.0
1.0
0.1
0.5
0,5
0.4
0.5
0.5
0.3
0.1
0.3
0.1
0.2
0.1
All columns glass, 6 ft. long x 4 mm ID, solid support Gas-Chrom Q (80/100 mesh),
nitrogen carrier flow 80 ml/rain.
Sensitivity factors relative to aldrin.
Retention times relative to aldrin.
-------
TABLE 3
Recovery of Organochlorine Pesticides from Natural Waters
Pesticide
Aldrin
Lindane
Dieldrin
DDT
Added Level
ng/liter
IS
110
10
100
20
125
40
200
Recovery
Without
Cleanup, %
69
72
97
73
108
85
101
77
Added Level
ng/liter
25
100
15
85
25
130
30
185
a
Recovery
With
Cleanup, %
68
65
94
70
70
65
118
71
isil column clean-up used.
-------
TABLE 4
Precision of Method for Organochlorine Pesticides in Natural Waters
Pesticide
Aldrin
Lindane
Dieldrin
DDT
Pretreatment
No
Cleanup
Cleanup
No
Cleanup
Cleanup
No
Cleanup
Cleanup
No
Cleanup
Cleanup
Mean Recovery
ng/ liter
10.42
79.00
17.00
64.54
9.67
72.91
14.04
59.08
21.54
105.83
17.52
84.29
40.30
154.87
35.54
132.08
Precision,
ST
4.86
32.01
9.13
27.16
5.28
26.23
8.73
27.49
18.16
30.41
10.44
34.45
15.96
38.80
22.62
49.83
a
ng/liter
so
2.59
20.19
3.48C
8.02°
3.47
11.49
S.20
7.75
17.92
21.84
5.10°
16.79°
13.42
24.02
22.50
25.31
ZS_, = Overall precision, and
S. = Single-Operator precision.
Use of Florisil column cleanup prior to analysis
S0 < ST/2
-------
FIGURE I
SILICA GEL COLLECTION ASSEMBLY
AIR FLOW /—EYE DROPPER
«-
VACUUM
HOSE ^GLASS WOOL
SILICA GEL COLLECTION ASSEMBLY
-------
FIGURE 2
DIAGRAM OF DESIGNATION OF TLC SECTIONS
IN THE CLEANUP AND SEPARATION
ON SILICA GEL PLATES
10.0 cm
8.0cm
SECTION
I
6.0cm
3.8cm
1.0 cm
SPOTTING 0-,
LINE "
ZONE FOR;
SPOT 1 SPOT 2 SPOT 3
SECTION
XE
SECTION
HI
SECTION
n
SECTION
I
I
•SOLVENT LINE
.7.8 ALDRIN
•7.5 DDE
• 7.3 DDT
• 6.5 HEPTACHLOR
--5.5 X'CHLOROANE
--5.2 ODD
--4.2 LINDANE
- 3.4 HEPTACHLOR EPOXIDE
2.4ENDRIN , DIELDRIN
I
I
I
J_
SPOT 1234
1.0 cm
8 9 10 II 12
-------
LINDANE
FIGURE 3
ELECTRON CAPTURE GAS CHROMATOGRAM
OF PESTICIDE STANDARDS
HEPTACHLOR
EPOXIDE
p.p'-DDE
"V
p.p'-ODT
I
I
4
I
I
I
12
I
6 8 10
RETENTION TIME IN MINUTES
(Chart speed one-half inch per minute)
Column Packing- 3%DC-200+5%QF-1 on Gas Chrom Q (80/100 Mesh)
Carrier Gas- Nitrogen at 80 ml/min.
Column Temperature- 200°C
14
-------
LINDANE
FIGURE 4
ELECTRON CAPTURE GAS CHROMATOGRAM
OF PESTICIDE STANDARDS
HEPTACHLOR
EPOXIDE
I
0 2 4 6 8 10
RETENTION TIME IN MINUTES
(Chart speed one-half inch per minute)
Column Picking- 3% OV-101 on Gas Chrom Q (80/100 Mesh]
Carrier Gas- Nitrogen at 80m!/min.
Column Temperature - 175°C
12
-------
LINDANE
FIGURE 5
ELECTRON CAPTURE GAS CHROMATOGRAM
OF PESTICIDE STANDARDS
HEPTACHCLOR
HEPTACHLOR
EPOXIDE
p.p'-DDT
02 46 8 10 12 14
RETENTION TIME IN MINUTES
(Chart speed one-half inch inch per minute]
Column Packing- 5%OV-17 on Gas Chrom Q (60/80 Mesh)
Carrier Gas - Nitrogen at 80ml/min.
Column Temperature- 2QO°C
16
18
-------
INDANE + HEPTACHLOR
FIGURE 6
ELECTRON CAPTURE GAS CHROMATOGRAM
OF PESTICIDE STANDARDS
HEPTACHLOR
EPOXIDE
p.p'-DDE
p,p'-DDT
I
0
T
2
T
I I I
6 8 10
RETENTION TIME IN MINUTES
(Chart speed one-half inch per minute]
Column Packing- 3% OV-210 on Gas Chrom Q (80/100 Mesh)
Carrier Gas- Nitrogen at 80 ml/min.
Column Temperature- 160°C
T
12
I
14
-------
REFERENCES CITED:
(1) "Pesticide Analytical Manual", U.S. Department of Health, Education,
and Welfare, Food and Drug Administration, Washington, D.C.,
Volumes I and II, 1968.
(2) "Guide to the Chemicals Used in Crop Protection", Canada Department
of Agriculture, Catalog Number A-43-1093, Queen's Printer, Ottawa
Canada, 5th Edition, 1968.
(3) "Official Methods of Analysis of the Association of Official
Agricultural Chemists", Association of Official Agricultural
Chemists, Washington, D.C., 20044, 10th Edition, 1965.
(4) "Tentative Recommended Practice for General Gas Chromatography
Procedures", E-260-65T, Part 30, ASTM Standards - General Test
Methods, p. 806, May, 1968.
(S) "Control of Chemical Analyses in Water Pollution Laboratories",
Environmental Protection Agency, Analytical Quality Control Laboratory,
1014 Broadway, Cincinnati, Ohio 45202. (In Press)
(6) "Standard Methods of Sampling Industrial Water", D-S10-68, Part 23,
ASTM Standards - Water; Atmospheric Analysis, p. 3, November, 1970.
(7) Gannon, H. and Bigger, J.H., Journal of Economic Entomology, 51,
No. 1, 1 (1958).
(8) Hendrick, R.D. et al, Journal of Economic Entomology, 59, No. 6
1388 (1966).
(9) Eichelberger, J.W. and Lichtenberg, J.J., "Persistence of Pesticides
in River Water", U.S. Department of the Interior, FWQA, DWQR, AQCL,
Cincinnati, Ohio, April 1970 (Accepted for publication by Environmental
Science and Technology).
(10) Lovelock, J.E. and Lipsky, S.R., Journal of the American Chemical
Society. 82_, 431 (1960).
(11) Lovelock, J.E., Analytical Chemistry, 33_, 162 (1961)
(12) Challacombe, J.A. and McNulty, J.A., "Applications of the Micro-
coulometric Titrating System as a Detector in Gas Chromatography of
Pesticide Residues", Residue Reviews, 5_, 57 (1964).
(13) Coulson, D.M., Journal of Gas Chromatography, 4_, 285 (1966)
(14) Brody, S. and Chancy, J., Journal of Gas Chromatography, £, 42 (1966)
(15) Purnell, H., "Gas Chromatography", Wiley, New York, 1962, p. 240.
-------
(16) Homing, E.G., Moscatelli, E.A., and Sweeley, C.C.',"QleW.ulIrtd,.,
751, London (1959) .
(17) Smith, E.D., Analytical Chemistry, 32, 1049 (1960).
(18) Burke, J.A., Journal of the Association of Official Analytical
Chemists. 413, 1037 (1965) ,
(19) Kruppa, R.F., Henley, R.S., and Smead, D.L., Analytical Chemistry,
39, 851 (1967).
(20) McNair, H.M. and Bonelli, E.J., "Basic Gas Chromatography",
Varian-Aerograph, Walnut Creek, California 94598, 1965.
(21) Schafer, M.L., Busch, K.A., and Campbell, J.E., Journal of Dairy
Science. XLVI, No. 10, 1025 (1963).
(22) Reynolds, L.M., Bulletin of Environmental Contamination S
Toxicology, 4_, No. 3, 128 (1969) .
(23) Armour, J.A., and Burke, J.A., Journal of the Association of
Official Analytical Chemists. 5_3_, 761 (1970).
(24) Bagley, G.E., Reichel, W.L., Cromartie, E.,
Journal of the Association of Official Analytical Chemists,
S3, 251 (1970).
(25) Beroza. M. and Bowman, M.C., Analytical Chemistry, 37, 291 (1965).
ADDITIONAL RECOMMENDED REFERENCES:
(26) Littlewood, A.B., "Gas Chromatography - Principles, Techniques and
Applications", Academic Press, New York, 1962.
(27) Noebels, H.J., Wall, R.F., and Brenner, N., "Gas Chromatography",
Second and Third International Symposium Held Under the Auspices
of the Analysis Instrumentation Division of the Instrument Society
of America, June 1959 and June 1961.
(28) Heftmann, E. ed. "Chromatography", Reinhold Publishing Corporation,
New York 1961.
(29) Stahl, E. ed., 'Thin-Layer Chromatography, A Laboratory Handbook",
(Springer-Verlag, Berlin) Academic Press, Inc., Publishers, New
York, 2nd edition, 1969.
(30) Randerath, K., "Thin-Layer Chromatography" (Verlag Chemio, CmbH,
Weinheim/Bergstr.) Academic Press, New York, 1964.
(31) Truter, E.V., "Thin Film Chromatography", Interscience Publishers,
Division of John Wiley and Sons, Inc., New York, 1963.
-------
f?'
(32) Smith., D. and Eichelherger, J.W., Journal of Hater Pollution
Control Federation. 37_, 77 (1965).
(33) Mills, P.A., Journal of the Association of Official Analytical
Chemists, 42, 734 (1959).
(34) Johnson, L., ibid., 45, 363 (1962).
(35) Goerlitz, D.F., "Methods for the Analysis of Organic Substances
in Water", Techniques of Water-Resources Investigations cf the
U.S. Geological Survey, Book 5, Chapter A2, (in review), 1970.
(36) Boyle, H.W., Burttschell, R.H., and Rosen, A.A., "Infrared
Identification of Chlorinated Insecticides in Tissues of
Poisoned Fish", published in Organic Pesticides in the
Environment, Advances in Chemistry Series-60, American
Chemical Society, Washington, D.C., 1966.
(37) Walker, K.C. and Beroza, M., Journal of the Association of
Official Analytical Chemists, 46, 250 (1963).
(38) Breidenbach, A.W., et al, "The Identification and Measurement
of Chlorinated Hydrocarbon Pesticides in Surface Waters",
Publication WP-22, U.S. Department of the Interior, Federal
Water Pollution Control Administration, Washington, D.C.
20242, 1966.
(39) Teasley, J.I. and Cox, W.S., "Determination of Pesticides in
Water by Microcoulometric Gas Chromatography after Liquid-
Liquid Extraction", Journal American Water Works Association
55_, 1093 (1963).
(40) Lamar, W.L., Goerlitz, D.F., and Law, L.M., "Identification
and Measurement of Chlorinated Organic Pesticides in Water
by Electron Capture Gas Chromatography", U.S. Geological
Survey Water Supply Paper 1817-B, 1965.
(41) "Guide to The Analysis of Pesticide Residues", U.S. Department
of Health, Education, and Welfare, Public Health Service,
Bureau of State Services, Office of Pesticides, Volumes I and
II, 1965.
-------
APPENDIX
7. SPECIAL EQUIPMENT. REAGENTS, AND SOLVENTS
7.1 Equipment
7.1.1 Gas Chromatograph - Suitable gas chromatographs are available from
many manufacturers.
7.1.2 Detectors
7.1.2.1 Electron Capture - Radioactive Source (tritium or nickel-63)
a. Concentric tube design
Varian-Aerograph
2700 Mitchell Drive
Walnut Creek, California 94598
b. Unique concentral design
Tracer, Inc.
6500 Tracer Lane
Austin, Texas 78721
c. Parallel plate design
Perkin-Elmer Corporation
Norwalk, Connecticut 06852
Also supplied by many other manufacturers.
7.1.2.2 Microcoulometric (T-300-S)
Dohrmann Instruments Company
1062 Linda Vista Avenue
Mountain View, California 94040
7.1.2.3 Electrolytic Conductivity
Tracor, Inc.
6500 Tracor Lane
Austin, Texas 78721
7.1.2.4 Flame Photometric
Also from Tracor, Inc.
-------
7.1.3 Recorder - 1 millivolt, 1 second full scale potentioraetric strip
chart. This type of recorder is supplied by many instrument
manufacturers.
7.1.4 Kuderna-Danish Glassware
Snyder Column - three ball (macro) and one ball (micro)
Evaporative Flasks - 125 ml, 250 ml, and 500 ml
Receiver Ampuls - 10 ml
Ampul Caps
Kontes Glass Company
Vineland, New Jersey 08360
Dohrmann Instruments
1062 Linda Vista Avenue
Mountain View, California 94040
7.1.5 Column Chroroatography - Pyrex column (I.D. 19 mm, length 400 mm)
with coarse fritted plate on bottom and Teflon stopcock, 250 ml
reservoir bulb at top of column with flared out funnel shape at
top of bulb--a special order.
Kontes Glass Company
Vineland, New Jersey 08360
7.1.6 Micro Syringes - (1, 5, 10, 25, 50, and 100 ul)
Hamilton Company
Post Office Box 307
Whittier, California 90608
7.1.7 Separatory Funnels - two liter and four liter funnels with Teflon
stopcock,
Pyrex or Kimble supplied through msny distributors.
-------
-TV
7.1.8 Thin-Layer Chromatography - Applicator, aligning tray, spotting
template, developing chamber, and UV light source.
Applied Science laboratories. Incorporated
Post Office Box 440
State College, Pennsylvania 16501
Brinkmann Instruments, Incorporated
Cantiague Road
Westbury, New York 11590
also - from many other suppliers
7.2 Standards, Reagents and Solven'r
7.2.1 Pesticide standards - highest available purity
City Chemical Company
132 West 22nd Street
New York, New York 10011
Applied Science Laboratories, Incorporated
Post Office Box 440
State College, Pennsylvania 16501
Environmental Protection Agency
Perrine Primate Research Branch
P.O. Box 490
Perrine, Florida 33157
also - from the manufacturer
7.2.2 Florisil (60/100 mesh) - purchased activated at 1200 F and stored
at 130 C.
Floridin Company
2 Gateway Center
Pittsburgh, Pennsylvania 15222
7.2.3 Sodium sulfate (A.C.S.) - granular, anhydrous
7.2.4 Pyrex wool - filtering grade
-------
7.2.S Solvents - hexane, diethyl ether, acetone, benzene, xylene,
carbon tetrachloride, acetonitrile, methylene chloride -
high purity, distilled in glass for pesticide analyses--
either Nanograde type or purified in lab.
Burdick and Jackson, Incorporated
1953 South Harvey Street
Muskegon, Michigan 49442
Mallinckrodt Chemical Works
2nd and Mallinckrodt Streets
St. Louis, Missouri 63160
Matheson Coleman and Bell
Post Office Box 85
East Rutherford, New Jersey 07073
7.2.6 Gas Chromatographic Column Materials
Gas-Chrom Q (80-100 mesh)
Glass Wool (silanized with dimethyldichlorosilane)
OV-17
OV-101
OV-210
UC-200 (12,500 centistokes)
QF-1 (FS-1265)
Tubing (Pyrex 1/8 in. and/or 1/4 in. O.D.)
Applied Science Laboratories, Incorporated
Post Office Box 440
State Collage, Pennsylvania 16501
Ohio Valley Specialty Chemical, Inc.
Marietta, Ohio 45750
7.2.7 Silica gel-G with gypsum binder (No. 8076)
Warner-Chilcott Laboratories
Instruments Division
200 South Garrard Boulevard
Richmond, California 94801
-------
7.3 Sample Collection Bottles and Shipping Containers
7.3.1 One-Quart Jars - Standard 32 02. 63-400 flint (C-S020),
FTK cap P/0 C63-400)
Cincinnati Container Co.
2833 Spring Grove Avenue
Cincinnati, Ohio 45225
7.3.2 Teflon Insert for Bottle Cap - 2-7/16 in. diameter, 0.020 in.
thick
Cadillac Plastics
3818 Red Bank Road
Cincinnati, Ohio 45227
7.3.3 Shipping Containers - Expanded polystyrene packer for one-quart
jars
Preferred Plastics Corp.
Route 12
North Grosvenordale, Connecticut
Polystyrene packers are also available for half-gallon and
one-gallon bottles.
Mention of products and manufacturers
is for identification only and does not imply
endorsement by the Water Quality Office,
Environmental Protection Agency.
6 «. V WnilJtttl NWIN OffKtt 197J-759-301/2113 K
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