EPA 600/1-81-024
March 1981
METHOD DEVELOPMENT FOR THE ASSESSMENT OF POSSIBLE
HUMAN EXPOSURE TO PESTICIDES AND INDUSTRIAL CHEMICALS
by
Thomas R. Edgerton
Robert F. Moseman
Lynn H. Wright
Analytical Chemistry Branch
Environmental Toxicology Division
Health Effects Research Laboratory
Research Triangle Park, North Carolina 27711
U. S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF RESEARCH AND DEVELOPMENT
HEALTH EFFECTS RESEARCH LABORATORY
RESEARCH TRIANGLE PARK, NORTH CAROLINA 27711
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DISCLAIMER
This report has been reviewed by the Health Effects Research
Laboratory, U. S. Environmental Protection Agency, and approved for
publication. Approval does not signify that the contents necessarily
reflect the views and policies of the U. S. Environmental Protection
Agency, nor does mention of trade names or commercial products constitute
endorsement or recommendation for use.
11
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FOREWORD
The many benefits of our modern, developing, industrial society are
accompanied by certain hazards. Careful assessment of the relative risk
of existing and new man-made environmental hazards is necessary for the
establishment of sound regulatory policy. These regulations serve to
enhance the quality of our environment in order to promote the public
health and welfare and the productive capacity of our Nation's
population.
The Health Effects Research Laboratory, Research Triangle Park,
conducts a coordinated environmental health research program in toxicology,
epidemiology, and clinical studies using human volunteer subjects.
These studies address problems in air pollution, non-ionizing radiation,
environmental carcinogenesis and the toxicology of pesticides as well as
other chemical pollutants. The Laboratory participates in the development
and revision of air quality criteria documents on pollutants for which
national ambient air quality standards exist or are proposed, provides
the data for registration of new pesticides or proposed suspension of
those already in use, conducts research on hazardous and toxic materials,
and is primarily responsible for providing the health basis for non-ionizing
radiation standards. Direct support to the regulatory function of the
Agency is provided in the form of expert testimony and preparation of
affidavits as well as expert advice to the Administrator to assure the
adequacy of health care and surveillance of persons having suffered
imminent and substantial endangerment of their health.
This report represents a research effort in the development of
methods which will allow for assessment of humans to possible exposure
to pesticides and industrial chemicals.
The emphasis of this project was to develop methodology for deter-
mining the metabolites of the chemicals from rat feeding studies and
apply these developed methods to humans. Such data is required for the
continued development of criteria for assessing human exposure to this
group of chemicals.
F. G. Hueter, Ph.D.
Director
Health Effects Research Laboratory
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ABSTRACT
The determination of chlorinated phenols in urine can be used as a
means for assessing exposure to pesticides and industrial chemicals in
the human population.
A method was developed for the analysis of chlorinated phenols
which involves the derivatization of metabolites from the urine of rats
fed hexachlorobenzene (HCB) and pentachlorophenol (PCP). This method
was then applied to urine samples taken from the general human population
to gain a background level. Pentachlorophenol was detected in greater
than 90% of the human samples analyzed. The only other metabolites
detected were tetrachloropyrocatechol and tetrachlorohydroquinone. The
urine of a worker occupationally exposed to PCP exhibited quantifiable
amounts of tetrachloropyrocatechol and tetrachlorohydroquinone along
with large amounts (greater than 3 ppm) of PCP. Pentachlorothiophenol,
a major metabolite of HCB fed to rats, was not detected in human urine.
The analysis of human urine for underivatized chlorinated phenols
using a direct gas chromatographic method not requiring derivatization
detected quantifiable levels of 2,5-dichloro-, 2,4,5-trichloro-, 2,3,4,6-
tetrachloro- and pentachlorophenol in greater than 90% of the samples
examined. Approximately 50% of the samples contained detectable levels
of 2,6 and 3,5-dichlorophenol and 2,4,6-trichlorophenol.
IV
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CONTENTS
Foreword , iii
Abstract iv
List of Figures vi
List of Tables vii
Acknowledgment viii
1. Introduction 1
2. Conclusions 4
3. Materials and Methods 5
Hexachlorobenzene and Pentachlorophenol ... 5
Underivatized Chlorinated Phenols 10
Confirmation Techniques 12
4. Results 14
Hexachlorobenzene and Pentachlorophenol
Studies 14
Confirmation of Metabolites by GC/MS 24
Analysis for Underivatized Chlorinated
Phenols 29
References 37
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FIGURES
Number Page
1 Gas Chromatogram of Urine Extract From
a Rat Fed PCP 18
2 Gas Chromatogram of Urine Extract From
a Rat Fed HCB 19
3 Gas Chromatogram of Fraction 2, Acid Alumina
Column. 2 ml Urine Extract. PCP Exposed Worker. . 26
4 Metabolites Isolated and Confirmed From
PCP Feeding Study 27
5 Metabolites Isolated and Confirmed From
HCB Feeding Study 28
6 Chromatograms of (I) XAD-4 Reagent Blank and (II)
Human Urine 35
7 Chromatogram of Human Urine 36
VI
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TABLES
Number Page
1 Possible Origins of Various Chlorinated Phenols . . 2
2 Recoveries of Metabolites from Fortified Urine ... 15
3 Recoveries of Methylated Metabolites of HCB and
PCP from an Acid Alumina Column 16
4 Relative Retention Data for Methylated Metabolites of
HCB and PCP 17
5 Hexachlorobenzene Metabolites in Rat Urine...
100 ppm in Diet, Results in ppm 20
6 Pentachlorophenol Metabolites in Rat Urine...
100 ppm in Diet, Results in ppm 21
7 Control Urine - plain Chow Results in ppm .... 22
8 Human Urine - Results in ppm 25
9 Retention Data for Chlorinated Phenols on a Double
Support Bonded DECS Column 29
10 Retention Data for Chlorinated Phenols on a Support
Bonded BDS Column 30
11 Recoveries of Chlorinated Phenols from Fortified
Urine 31
12 Human Urine - Chlorinated Phenols, Results in ppb . 33
VI 1
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ACKNOWLEDGMENTS
The valuable technical assistance of S. Quinn, E. Yarbro, and S. Todd
is greatly appreciated. A sincere thanks is extended to R. E. Linder for
assistance in the metabolism and feeding studies.
The authors wish to thank D. W. Bristol for his helpful suggestions and
the professionals of the Analytical Chemistry Branch, Environmental Toxicolocy
Division for their support.
The constant encouragement of E. 0. Oswald and R. G. Lewis is deeply
appreciated.
vm
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SECTION 1
INTRODUCTION
Chlorinated phenols have been of concern to the enivronmental
scientist for many years. Their toxicity to fish and other aquatic life
1 2
is well documented. ' The presence of chlorophenols in industrial
3
effluents has also been demonstrated. Methodology was developed to
analyze for chlorinated phenols in urine to assess possible human exposure
to chemicals known or suspected to give these compounds as metabolites. ' '
The most widely used and studied chlorinated phenol is pentachlorophenol
(PCP). It has been used extensively in the wood products industry as a
preservative and in agriculture as a fungicide. A discussion of its
uses, toxicity, and fate in the environment is found in the literature.
The occurrence of PCP in humans from occupational exposed workers and
Q Q
the general human population is also well documented. '
The metabolism of PCP and other chemicals which give rise to PCP
and other chlorinated phenols as metabolites can be found in the litera-
~| n —~i c
ture. A listing of possible sources of chlorinated phenols is
found in Table 1. This list is by no means complete, but does give some
insight into possible origins of various chlorinated phenols that might
be encountered in exposure assessment work.
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Table 1. Possible Origins of Various Chlorinated Phenols
Metabolite
2,6-dichlorophenol
2,4-dichlorophenol
2,3-dichlorophenol
2,5-dichlorophenol
3,4-dichlorophenol
3,5-dichlorophenol
2,3,4-trichlorophenol
2,3,5-trichlorophenol
2,3,6-trichlorophenol
2,4,5-trichlorophenol
2,4,6-trichlorophenol
3,4,5-trichlorophenol
2,3,5,6-tetrachlorophenol
2,3,4,6-tetrachlorophenol
2,3,4,5-tetrachlorophenol
pentachlorophenol
Origin
Lindane
Lindane
VC-13
m-dichlorobenzene
2,4-D
Lindane
o-dichlorobenzene
Lindane
2,4,5-T
p-dichlorobenzene
Lindane
o-dichlorobenzene
Diuron
Lindane
PCP
Lindane
Lindane
PCP
Lindane
Lindane
Ronnel
Erbon
2,4,5-T
HCB
Tetrachlorvinphos
Lindane
Lindane
HCB
PCP (impurity)
Lindane
PCP
Lindane
HCB
PCP
Lindane
HCB
PCNB
Type Pesticide
Insecticide
Insecticide
Insecticide
Fumigant
Herbicide
Insecticide
Fumigant
Insecticide
Herbicide
Insecticide
Fumigant
Herbicide
Insecticide
Insecticide
Insecticide
Insecticide
Insecticide
Insecticide
Herbicide
Herbicide
Fungicide
Insecticide
Insecticide
Fungicide
Fungicide
Insecticide
Fungicide
Insecticide
Fungicide
Fungicide
Insecticide
Fungicide
Fungicide
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Metabolism studies and analytical methods development were conducted
using rats fed two chemicals, pentachlorophenol (PCP) and hexachlorobenzene
(HCB), which were known or suspected to metabolize to chlorinated phenols.
The method was then applied to urine samples from the general human
population and the urine of a worker occupationally exposed to penta-
chlorophenol .
The literature contains no apparent methodology for the determination
of lower chlorinated phenols in urine at the low ng/g level. Because of
this, a method was developed in an attempt to detect mono-, di-, and
trichlorophenols in urine. No metabolism studies were performed, but
the method was applied to general human population urine samples.
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SECTION 2
CONCLUSIONS
Pentachlorophenol, 2,5-dichlorophenol, 2,4,5-trichlorophenol and
2,3,4,6-tetrachlorophenol were found in greater than 90% of the human
urine samples analyzed. Since all but one of the samples was from the
general population, it is difficult to determine a common exposure
route. The urine of workers occupationally exposed to other chemicals
which can give chlorinated phenol metabolites and further animal feeding
studies need to be conducted before definitive exposure assessments can
be made.
The determination of chlorinated phenols in urine can possibly be
used as an index of exposure to numerous chemicals.
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SECTION 3
MATERIALS AND METHODS
HEXACHLOROBENZENE AND PENTACHLOROPHENOL17
Animals
Adult female Sherman rats, 3-4 months of age and weighing 215-275 g,
were distributed into the following treatment groups: group 1, 4
rats on 575 ppm cornstarch in chow; group 2, 6 rats on 100 ppm HCB;
group 3, 6 rats on 100 ppm PCP. The HCB and PCP diets were prepared by
mixing a quantity of HCB or PCP with a small amount of cornstarch (resulting
in a starch concentration of about 575 ppm in the final mix) in a mortar,
then making the appropriate dilution with ground chow and mixing in an
electric mixer.
Rats were housed 2 per cage, identified with ear tags, and provided
their respective diets and water ad 1ibitum. Animals were weighed
weekly and food consumption measured at least 2 days each week. Indi-
vidual urine samples were collected overnight, after 30 days and after
107 days. Rats were fed plain chow during urine collection to avoid
non-ingested parent compound in the collected urine.
No clinical signs of toxicity were observed. Weight gain and food
consumption were comparable in all rats. Average HCB or PCP ingestion
over the first 30 days was 6.5 mg/Kg/day for both compounds.
Apparatus
Tracer MT-220, gas chromatograph equipped with a nickel-63 electron-
capture detector, was operated in the pulsed linearized mode. Borosilicate
glass columns (1.8 m x 4 mm i.d.) were packed with 80/100 mesh Gas Chrom Q
coated with 5% OV-210, 3% OV-1, 3% Silar 10-C, 4% SE-30/6% OV-210 or
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1.5% 0V-17/1.95% QF-1. A 5% DECS coated on 80/100 mesh Gas Chrom P was
also used. The OV-1, Silar 10-C, SE-30/OV-210, and OV-17/QF-1 columns
were operated at 170°C with a 5% methane in argon carrier gas flow rate
of 60 mL/min. The OV-210 column was operated at 160°C with a 5% methane
in argon flow rate of 40 mL/min. The DEGS column was operated at 170°
with a 5% methane in argon flow rate of 90 mL/min. Detector, inlet, and
transfer line temperatures were 300°, 235°, and 220°, respectively.
Analytical results were confirmed on a Finnigan Model 3200 quadru-
pole mass spectrometer equipped with a Model 9500 Gas Chromatograph and
Model 6100 Data System. Methane was used as the reagent gas for operation
in the chemical ionization mode with a source temperature of 120°C,
pressure 117 Pa, 110 eV electron energy and 1.0 ma emission current.
For gas chromatography-mass spectrometry (GC/MS), a borosilicate
glass column (1.2 m x 2 mm i.d.) was packed with 80/100 mesh Gas Chrom
Q coated with 5% OV-210. The column was operated at 90°C isothermal for
1 minute, then programmed at 4°C per minute to a final temperature of
160°C. Methane carrier gas flow rate was 20 mL/min. Inlet, transfer
line, and ion source temperatures were 200, 250, and 120°C, respectively.
Reagents and Materials
Anhydrous, granular sodium sulfate and sodium bisulfite were
Soxhlet extracted for 4 h with hexane and oven dried at 130°.
Acid alumina, Brockman Activity I (Fisher Scientific), was dried
for 24 h at 130°C and stored in a desiccator.
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Potassium hydroxide and hydrochloric acid were reagent grade.
N-Methyl-N'-nitro-N-nitrosoguanidine (Aldrich Chemical Co., Milwaukee,
Wis.), should be handled carefully since it is a known carcinogen.
All solvents were pesticide quality or equivalent.
2,3,4,6-tetrachlorophenol, 2,3,5,6-tetrachlorophenol, pentachloro-
phenol, pentachlorothiophenol, 2,3,4,5-tetrachlorophenol, were obtained
from Aldrich Chemical Co. ; tetrachloropyrocatechol from Pfaltz and
Bauer, Inc., Flushing, NY; and tetrachlorohydroquinone from K&K Labora-
tories, Inc., Plainview, NY. Pentachlorothiophenol, tetrachlorohydro-
quinone and tetrachloropyrocatechol were recrystallized prior to use.
The purity of HCB (based on EC-GC) used for dietary feeding was
greater than 99.5%. No apparent impurities were found by EC-GC analysis.
PCP contained 0.8% 2,3,4,6-tetrachlorophenol as the only detectable
impurity.
Methylating Reagent
Potassium hydroxide (2.3 g) was dissolved in 2.3 ml of distilled
water in a 125 ml Erlenmeyer flask and cooled to room temperature.
Twenty-five milliliters of ethyl ether was then added and the flask was
cooled in a refrigerator. The following step was carried out in a glove
box or high-draft hood. N-Methyl-N'-nitro-N-nitrosoguanidine (1.5 g)
was added in small portions to the flask with vigorous shaking. The
ether layer, which contained diazomethane, was decanted into a scintil-
lation vial and stored in a freezer.
Preparation of Standard Solutions
A standard of each phenol was prepared in hexane and stored at
-15°C in brown glass bottles. Urine fortifications were made from
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acetone dilutions of the seven mixed phenol standards. A volume con-
taining 10 |jg of each phenol was pipeted into separate 15 mL graduated
centrifuge tubes. Methylation was accomplished by adding 5 mL of dia-
zomethane reagent in a high draft hood to each tube (CAUTION: Diazome-
thane is toxic and may be explosive under certain conditions). The
phenol standards were allowed to stand for 30 minutes before EC-GC
determination. Nitrogen was bubbled through the individual standard
solutions to remove any excess diazomethane prior to EC-GC analysis.
For determination of elution patterns on an acid alumina column, a known
amount of each phenol was methylated as a mixture, and allowed to stand
for 30 minutes. The methylated phenol mixture was concentrated to 0.2
to 0.3 mL under a gentle stream of nitrogen prior to being placed on the
acid alumina column.
Preparation and Elution of Acid Alumina Column
A size 22-9 chromaflex column (Kontes 420530) was loosely plugged
with a small amount of glass wool. Acid alumina (4 g) was added in
small increments with tapping. Anhydrous, granular Na^SCL (1.6 g) was
added on top of the alumina. Thirty milliliters of 40% benzene in
hexane was used to wash the column free of interferences. After thor-
oughly air drying, the column was placed in an oven at 130°C overnight
prior to use.
A prepared column was removed from the oven just prior to use and
allowed to cool to room temperature. The column was wetted with 7 mL of
hexane. When the solvent layer reached the top of the NapSCL, an aliquot
of methylated sample or methylated standard phenol mixture in 0.2 to 0.3
-------
ml was placed on top of the column. Quantitative transfer of the sample
or standard was accomplished with three 0.5 ml rinsings with hexane. An
additional 3.5 ml of hexane was added and the hexane fraction (5.0 ml)
was collected and discarded. 2,3,4,6-Tetrachlorophenol, 2,3,5,6-tetra-
chlorophenol, pentachlorophenol, and pentachlorothiophenol were eluted
with 20 ml of 10% benzene in hexane (Fraction I). The remaining phenols,
2,3,4,5-tetrachlorophenol, tetrachloropyrocatechol, and tetrachlorohydro-
quinone, were eluted with 20 ml of 40% benzene in hexane (Fraction II).
Fractions I and II were adjusted to an appropriate volume for injection
into the gas chromatograph.
Analysis of Urine
Two milliliters of urine were transferred into a 20 mm x 125 mm
(R)
Teflon lined screw cap culture tube and 100 mg of sodium bisulfite was
added. The urine was acidified by the addition of 0.5 ml of concentrated
HC1. The tube was sealed and placed in a boiling water bath for 1 h
with periodic shaking. After hydrolysis, an additional 100 mg of sodium
bisulfite was added to the cooled urine sample and extracted twice for
one hour each on a mechanical rotator at 30 to 50 RPM using two 5 ml
portions of benzene. The samples were centrifuged after each extraction
and the extracts combined in an aluminum foil wrapped 15 ml centrifuge
tube. The benzene extract was concentrated to a volume of 0.3 to 0.5 ml
in a water bath at 30°C under a gentle stream of nitrogen and methylated
with 5 ml of diazomethane reagent. The methylated extract was allowed
to stand for 1 h. Prior to column cleanup, the methylated urine
extract was concentrated to approximately 0.3 ml under a gentle stream
of nitrogen. Two milliliters of hexane were added, and the solution was
again reconcentrated to a volume of 0.3 ml.
9
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UNDERIVATIZED CHLORINATED PHENOLS
Apparatus
A Tracer MT-222 gas chromatograph, equipped with a nickel-63 electron
capture detector, was operated in the pulsed linearized mode. Borosili-
cate glass columns (1.8 m x 4 mm I.D. or 0.6 m x 4 mm I.D.) were packed
with double support-bonded diethylene glycol succinate (DSB-DEGS) or
support-bonded butane 1,4-diol succinate (SB-BDS) on 80/100 mesh acid
18 19
washed Chromosorb W. ' The columns were operated at 180°-210° with a
5% methane in argon carrier gas flow-rate of 60-80 mL/min. Other tempera-
tures were: detector, 300°; inlet 225°; transfer line, 220°.
The following glassware was used: Chromaflex column, plain, teflon
stopcock, 250 x 10.5 mm I.D. , (Kontes Cat. No. K-420280); Kuderna-Danish
concentrator assembly (K-570000); 25-ml graduated tubes, size 2525
(K-570050); 15 x 125 mm screwcap culture tube.
Reagents and Materials
Chlorinated phenol reference standards were obtained from Aldrich
Chemical Co. (Milwaukee, Wis., U.S.A). For fortification purposes, the
phenols were converted to their respective sodium salts prior to addi-
tion to urine.
All solvents were pesticide quality or equivalent.
Reagent materials (3 N HC1, 0.1 N NaOH, NaHS03, deionized water)
were extracted with hexane and toluene prior to use.
The macroreticular resin, XAD-4, was obtained from Rohm and Haas
(Philadelphia, Pa., U.S.A.). The fines were removed by slurrying in
20
methanol and decanting. The remaining beads were purified by Soxhlet
extraction with 3 N HC1 for 18 h followed by neutralization with
10
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water in a Buchner funnel. The neutralized resin was washed with six 50
ml volumes of 0.1 N NaOH. The resin was again neutralized with water
and allowed to dry. The dried resin was then sequentially extracted
with methanol, acetonitrile, acetone and hexane in a Soxhlet extractor
20
for 8 h per solvent. The purified resin was stored under methanol in
a glass stoppered bottle.
Preparation of XAD-4 Column
A small plug of hexane-extracted glass wool was placed in a 250 mm
x 10.5 mm I.D. chromaflex column. Approximately 6 cm (1.5-2 gm dry
weight) XAD-4 resin in methanol was added to the column. The flow of
methanol was halted when the level of solvent reached the top of the
resin bed. A second plug of glass wool was placed on top of the resin
bed.
Preparation of Urine
Four milliliters of urine were placed in a 15 x 125 mm screw-cap
culture tube. To this was added 100 mg of sodium bisulfite and 1 ml of
®
concentrated HC1. The tube was sealed with a Teflon lined screw-cap
and placed in a boiling water bath for 1 h with periodic shaking. The
sample was removed and cooled to room temperature.
Column Elution and Regeneration
The XAD-4 column was rinsed with 10 ml of deionized water. When
the water level reached the top of the resin bed, 15 ml of 3 N HC1 was
added. After approximately 5 ml of acid eluted through the column, the
flow was halted and the column allowed to equilibrate for 5 min. After
equilibration the flow of acid was continued and a phenol standard or
hydrolyzed urine sample was added to the column when the level of acid
11
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reached the top of the resin bed. Quantitative transfer of the sample
was accomplished with two 1 ml rinsings with 3 N HC1. The column was
eluted with an additional 23 ml of 3 N HC1 followed by 25 ml of deionized
water. Both acid and water eluates were discarded. When the water
level reached the top of the resin bed, a Kuderna-Danish (K-D) concen-
trator assembly was placed under the column and the phenols were eluted
from the column with 100 ml of 10% 2-propanol in hexane (V/V). After
the first 5 ml was eluted and collected, the flow was halted and the
column allowed to equilibrate for 5 min. After equilibration, the flow
was continued and the eluate collected. The K-D assembly was removed
and the column was regenerated by washing with approximately 25 ml of
methanol which was discarded. A volume of methanol was kept in the
column until further use.
Concentration
The K-D assembly was placed on steam bath and the sample was concen-
trated to a final volume of 10-15 ml (two phases). The sample was
cooled and the hexane layer (upper) transferred to a 15 ml centrifuge
tube. The extract was then further concentrated to a volume of 1-2 ml
for analysis by EC-GC.
CONFIRMATION TECHNIQUES
Results from the hexachlorobenzene and pentachlorophenol studies
were confirmed by the GC/MS technique as was previously described.
12
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Samples from the genera] chlorinated phenol method were confirmed
by a separate gas chromatograph column (Butane diol succinate).
13
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SECTION 4
RESULTS
HEXACHLOROBENZENE (HCB), PENTACHLOROPHENOL (PCP) STUDIES
Results of recovery studies for the suspected chlorinated phenol
metabolites from HCB and PCP from fortified urine and through acid
alumina column cleanup are given in Tables 2 and 3. Recovery of penta-
chlorothiophenol was low, possibly due to an oxidative reaction or to
binding by components in urine. Difficulty was also encountered in
analyzing for di- and trichlorophoenols using the derivatization technique
because of the method's lack of reproducibility in derivatizing these
compounds.
Retention times for the methyl ethers of the suspected chlorinated
phenol metabolites found in HCB and PCP feeding study are listed in
Table 4. A 5% OV-210 column was found to give adequate separation after
column cleanup for these particular chlorinated phenol methyl ethers. A
5% DECS column was used only for separation of 2,3,5,6 from 2,3,4,6-
tetrachlorophenol.
Representative chromatograms for these studies are illustrated in
Figures 1 and 2. Tables 5, 6 and 7 list actual analytical results from
the PCP and HCB feeding studies. Note the difference in rat metabolism
of PCP from that of HCB. Pentachlorothiophenol and 2,3,5,6-tetrachloro-
phenol appear to be metabolites of HCB but not PCP.
The developed methodology was then applied to human urine samples.
Table 8 is a listing of these results. As can be seen from the table,
the levels of the two dihydroxy compounds in the urine of the occupationally
14
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Table 2. Recoveries of Metabolites from Fortified Urine
Metabolite
2,3,5,6-tetra-
chlorophenol
2,3,4,6-tetra
chlorophenol
2,3,4,5-tetra-
chlorophenol
Pentachloro-
phenol
Tetrachloro-
pyrocatechol
Tetrachloro-
hydroquinone
Pentachloro-
thiophenol
ppm Added
1.0
0.3
0.1
0.03
0.01
1.0
0.3
0.1
0.03
0.01
1.0
0.3
0.1
0.03
0.01
1.0
0.3
0.1
0.03
0.01
1.0
0.3
0.1
0.03
0.01
1.0
0.3
0.1
0.03
0.01
1.0
0.3
0.1
0.03
0.01
% Range
89.3-92.3
85.7-92.3
82.0-87.9
78.0-85.6
79.1-87.5
88.9-92.4
86.1-91.8
83.1-88.3
80.8-84.2
79.6-86.8
89.3-95.6
89.0-94.3
86.0-91.0
85.8-90.3
82.8-90.5
95.2-97.8
91.5-95.2
86.0-95.0
93.8-100.4
90.6-96.3
78.6-81.4
78.7-85.7
76.3-83.0
59.1-71.4
60.1-69.7
80.2-82.7
77.0-84.5
77.0-84.0
75.6-86.0
71.3-77.3
69.9-73.6
69.3-73.3
61.0-73.0
49.8-57.2
41.5-51.3
Avg. % Rec.
91.1
88.8
85.3
82.3
82.8
90.9
88.9
86.0
82.5
82.6
93.1
91.8
88.2
87.4
85.6
96.5
93.4
92.0
97.2
93.2
80.1
81.6
79.8
65.6
63.7
81.5
81.4
80.9
80.4
74.6
71.9
71.1
66.4
53.3
47.3
% Rel. Std. Dev.
±1.3
±2.7
±2.5
±3.3
±3.6
±1.5
±2.4
±2.5
±1.7
±3.2
±2.7
±2.3
±2.1
±2.0
±3.4
±1.1
±1.8
±4.1
±2.8
±2.5
±1.2
±2.9
±3.2
±5.4
±4.3
±1.0
±3.2
±2.9
±4.6
±2.5
±1.5
±1.7
±6.0
±3.3
±4.2
Four determinations for each.
15
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Table 3. Recoveries of Methylated Metabolites
of HCB and PCP from an Acid Alumina Column
Metabolite
Amount added, |jg Amount recovered,
Recovery
2,3,4,6-tetra-
chlorophenol
2,3,5,6-tetra-
chlorophenol3
5.00
1.00
0.10
5.00
1.00
0.10
4.650
0.970
0.092
4.723
0.970
0.093
93.0
97.0
92.0
94.5
97.0
93.0
Pentachloro-
phenola
Pentachloro-
thiophenol
2,3,4,5-tetra-
chlorophenol
5.00
1.00
0.10
5.00
1.00
0.10
5.00
1.00
0.10
4.800
0.930
0.094
4.850
0.880
0.091
4.720
0.893
0.091
96.0
93.0
94.0
97.0
88.0
91.0
94.4
89.3
91.0
Tetrachloro-,
pyrocatechol
Tetrachloro-,
hydroquinone
5.00
1.00
0.10
5.00
1.00
0.10
4.700
0.950
0.095
4.850
0.960
0.091
94.0
95.0
95.0
97.0
96.0
91.0
Fraction 1 -- 20 ml 10% benzene in hexane
^Fraction 2 -- 20 ml 40% benzene in hexane
16
-------
Table 4. Relative Retention Data for Methylated Metabolites of HCB and PCP
Metabolite
2 ,3 ,4, 6-tetrachl orophenol
2,3,5,6-tetrachlorophenol
2,3,4, 5- tetrachl orophenol
Pentac hi orophenol
Tetrachl oropyrocatechol
Tetrachlorohydroquinone
Pentachlorothiophenol
4% Se-30/6% OV-210
0.23
0.23
0.38
0.46
0.45
0.48
0.95
1.5% OV-17/1.95% QF1
0.33
0.33
0.51
0.55
0.55
0.56
1.06
5% OV-210
0.24
0.24
0.46
0.49
0.52
0.59
1.00
3% OV-1
0.22
0.22
0.34
0.44
0.42
0.42
0.91
3% Silar 10-C
0.36
0.37
0.46
0.68
0.88
0.83
1.59
5% DEGSb
1.21
1.13
1.66
2.58
--
--
--
Retention time relative to Aldrin
Underivitized phenols
-------
10
12
TIME, minutes
Figure 1. Gas chromatograms of urine extract from a rat fed PCP. (I) Fraction 1
acid alumina column, (a) PCP, 12.3 ppm; (II) Fraction II acid alumnia column.
(b) 2, 3, 4, 5-tetrachlorophenol, 1.02 ppm; (c) tetrachlorophyrocatechol, 0.18
ppm; (d) tetrachlorohydroquinone, 24 ppm. Column: 5% OV-210 on 80/100
mesh Gas Chrom Q. Oven temperature '160°C 5% methane in argon; flow rate
40 mL/min.
-------
10
12
TIME, minutes
Figure 2. Gas chromatograms of urine extract from a rat fed HCB. (I) Fraction 1 acid
alumina column, (a) 2, 3, 5, 6-tetrachlorophenol, 74 ppb; (b) PCP, 405 ppb; (c)
pentachlorothiophenol, 500 ppb. (II) Fraction II acid alumina column; (d) 2, 3, 4,
5-tetrachlorophenol, 20 ppb; (e) tetrachloropyrocatechol, 8 ppb; (f) tetrachloro-
hydroquinone, 435 ppb. Column: 5% OV-210 on 80/100 mesh Gas Chrom Q. Oven
temperature 16QOC. 5% methane in argon; flow rate 40 mL/min.
-------
Table 5. Hexachlorobenzene Metabolites in Rat Urine--100 ppm in Diet
Results in ppm
Days
on
Chow
30
107
30
107
30
107
Jl 30
A 107
30
107
30
107
Sample
No.
2240
2240
2241
2241
2242
2242
2243
2243
2244
2244
2245
2245
Pentachloro-
phenol
0.081
0.405
0.167
0.420
0.085
0.355
0.066
0.189
0.079
0.412
0.168
0.471
Tetrachloro-
hydroquinone
0.079
0.182
0.238
0.413
0.031
0.339
0.027
0.090
0.114
0.170
0.139
0.349
Pentachloro-
thiophenol
0.424
0.500
0.390
0.303
0.560
0.136
0.485
0.200
0.332
0.197
0.747
0.363
Tetrachloro-
pyrocatechol
0.001
0.003
0.002
0.007
0.001
0.004
0.001
0.002
0.002
0.002
0.002
0.007
2,3,5,6-tetra-
chlorophenol
0.029
0.074
0.033
0.074
0.027
0.048
0.025
0.055
0.023
0.044
0.042
0.085
2,3,4,5-tetra-
chlorophenol
0.005
0.008
0.013
0.015
0.001
0.010
0.002
0.004
0.005
0.008
0.005
0.012
-------
Table 6. Pentachlorophenol Metabolites in Rat Urine—100 ppm in Diet
Results in ppm
Days
on
Chow
30
107
30
107
30
107
30
107
30
107
30
107
Sample
No.
2246
2246
2247
2247
2248
2248
2249
2249
2250
2250
2251
2251
Pentachloro-
phenol
16.8
12.3
9.2
10.8
12.2
6.6
21.0
10.1
19.4
15.3
7.4
8.1
Tetrachloro-
hydroquinone
48.8
24.0
25.9
28.4
40.4
10.8
42.8
20.4
38.4
29.7
27.0
14.3
Pentachloro-
thiophenol
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
Tetrachloro-
pyrocatechol
0.16
0.18
0.15
0.17
0.15
0.08
0.11
0.16
0.40
0.42
0.31
0.28
2,3,5,6-tetra-
chlorophenol
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
2,3,4,5- tetra-
chlorophenol
2.17
1.02
1.05
0.93
1.33
0.36
1.18
1.00
1.48
1.31
1.19
0.87
-------
Table 7. Control Urine—Plain Chow
Results in ppm
Days
on
Chow
30
107
30
107
30
107
30
107
Sample
No.
2236
2236
2237
2237
2238
2238
2239
2239
Pentachloro-
phenol
0.002
0.004
0.003
0.002
0.033
0.021
0.041
0.036
Tetrachloro-
hydroquinone
0.008
0.006
0.004
0.012
0.006
0.011
0.037
0.043
Pentachloro-
thiophenol
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
Tetrachloro-
pyrocatechol
0.004
0.002
0.002
0.002
<0.001
0.003
0.007
0.009
2,3,5,6-tetra-
chlorophenol
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
2,3,4,5-tetra-
chlorophenol
<0.001
<0.001
0.001
0.004
0.015
0.021
0.011
0.008
-------
exposed worker are in nearly equal proportion. This may indicate a
possible path for human metabolism of PCP and also be used as a further
indicator of exposure to this compound.
Pentachlorophenol was identified in ten of the eleven urine samples
from the general human population using the described analytical method-
ology, and ranged from 1 to 80 ppb. The presence of 2,3,4,6-tetra-
chlorophenol in the urine can possibly be attributed to its presence as
an impurity in preparations of PCP. The only measurable metabolites
from the general human population samples were tetrachlorohydroquinone
and tetrachloropyrocatechol. A representative chromatogram from the
urine of an occupationally exposed worker is shown in Figure 4. The
occupationally exposed worker contained a high level of PCP and measurable
levels of tetrachlorohydroquinone and tetrachloropyrocatechol.
As can be seen from these results, pentachlorothiophenol in urine
can be used as an indicator of possible exposure to HCB. PCP exposure
would be indicated by a high level of PCP and the presence of tetra-
chlorohydroquinone and tetrachloropyrocatechol in the urine.
CONFIRMATION OF METABOLITES BY GC/MS
The phenolic metabolites in urine extracts from the feeding study
and human population were confirmed as their methyl ethers by combined
GC-MS. Chemical ionization, using methane reagent gas, produced fairly
strong M + 1 protonated-molecular ion cluster beginning at m/z 245 for
the three isomers of tetrachlorophenol, m/z 279 for pentachlorophenol,
24
-------
m/z 295 for pentachlorothiophenol and m/z 275 for tetrachloropyrocatechol
and tetrachlorohydroquinone. In addition, a fairly strong M + 1 protonated
molecular ion isotope cluster beginning at m/z 241 was tentatively
identified as an isomer of the methyl ether of trichlorodihydroxybenzene
from the PCP feeding study samples. The phenolic metabolites in the
urine from the occupationally exposed worker were confirmed by GC/MS as
the methyl ethers of tetrachloropyrocatechol and tetrachlorohydroquinone.
The structures of the metabolites isolated and confirmed from the HCB/PCP
feeding studies are shown in Figures 4 and 5.
25
-------
Table 8. General Human Population Urine Samples - Results in ppm
Sample
No.
1
2
3
4
5
6
7
8
9
LO
11
12*
Pentachloro-
phenol
0.006
0.012
0.004
<0.001
0.080
0.004
0.015
0.012
0.009
0.038
0.018
3.60
Tetrachloro-
hydroquinone
<0.001
<0.001
<0.001
<0.001
0.002
<0.001
0.003
0.006
<0.001
0.008
<0.001
0.024
Pentachloro-
thiophenol
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
Tetrachloro-
pyrocatechol
<0.001
<0.001
0.002
<0.001
0.001
<0.001
0.004
0.005
<0.001
0.007
<0.001
0.024
2,3,4,6-tetra-
chlorophenol
0.004
0.002
0.003
<0.001
0.013
0.002
0.004
0.002
0.003
0.009
0.003
0.123
2,3,5,6-tetra-
chlorophenol
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
2,3,4,5-tetra
chlorophenol
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
<0.001
0.005
*0ccupationally exposed to PCP
-------
LU
CO
z
o
CL.
CO
DC
LU
Q
DC
O
O
LU
DC
4 6
TIME, minutes
10
Figure 3. Gas chromatogram of Fraction 2, Acid Alumina Column.
2 ml urine extract, PCP exposed worker (a) tetrachloropyrocatechol,
equiv. 24 ppb; (b) tetrachlorohydroquinone, equiv. 24 ppb. 5%
OV-210, 1600C, 40 mL/min.
-------
OH
Cl
tetrachlorohydroquinone
Cl
(OH)2
Cl
Cl
trichlorodihydroxybenzene
OH
OH
OH
Cl
tetrachloropyrocatechol
OH
2,3,4,5-tetrachlorophenol
Cl
Cl
Cl
Cl
Figure 4. Metabolites isolated and confirmed from PCP feeding study.
-------
2,3,,5,6-tetrachlorophenol
2,3,4,5-tetrachlorophenol
pentachlorophenol
tetrachlorohydroquinone
Cl
pentachlorothiophenol
tetrachloropyrocatechol
Cl Cl^v> ^Cl
Figure 5. T Metabolitesjsolated and confirmed from HCB feeding study.
-------
ANALYSIS FOR UNDERIVATIZED CHLORINATED PHENOLS
Relative retention times of the chlorinated phenols analyzed as the
free phenol are presented in Tables 9 and 10. The difference in elution
patterns of SB-BDS from that of DSB-DEGS allows confirmation of chlorinated
phenols in urine. A double support-bonded DECS column packing was
chosen over conventional and single support-bonded packings because of
its efficiency, low bleed and longevity. The column was prepared by
double heat treating stabilized diethylene glycol succinate on acid
18 19
washed Chromosorb W. ' A 0.6 m column was used for the rapid elution
of pentachorophenol. Work is continuing in our laboratory on the use of
HPLC for the separation and confirmation of chlorinated phenols. These
results will be presented in a later publication.
As shown in Table 11, recoveries of chlorinated phenols from forti-
fied urine averaged better than 80%. Method sensitivity is dependent
upon chlorine substitution and elution time of the phenol through the
gas chromatographic column. Method sensitivity for di- and trichlorophenols
(except 3,4- and 3,5-dichlorophenol and 3,4,5-trichlorophenol) is 1 ppb.
The method sensitivity for the tetrachlorophenols and pentachlorophenol
is 2 ppb. Because of the possible effect of meta-meta substitution,
method sensitivity for 3,5-dichlorophenol is 15 ppb and 150 ppb for
3,4-dichlorophenol and 3,4,5-trichlorophenol.
Table 12 lists results from the application of this analytical
methodology to general population human urine samples. (Different
samples from those of Table 8.) Only those chlorinated phenols that had
identical retention times with standards on DSB-DEGS and SB-BDS are
29
-------
Table 9. Retention Data for Chlorinated Phenols on a
Double Support-Bonded DECS Column*
DDT
Compound minutes 2,4,5-TCP
2,6-dichlorophenol 2.95 0.35
2,4-dichlorophenol 3.19 0.37
2,5-dichlorophenol 3.19 0.37
2,3-dichlorophenol 3.39 0.40
2,4,6-trichlorophenol 5.47 0.64
2,3,5-trichlorophenol 6.81 0.80
2,3,6-trichlorophenol 6.85 0.80
2,4,5-trichlorophenol 8.54 1.00
2,3,4-trichlorophenol 8.94 1.05
2,3,5,6-tetrachlorophenol 13.15 1.54
2,3,4,6-tetrachlorophenol 14.84 1.74
3,5-dichlorophenol 19.29 2.26
3,4,5-trichlorophenol 20.47 2.40
2,3,4,5-tetrachlorophenol 21.18 2.48
3,4-dichlorophenol 25.83 3.02
pentachlorophenol 35.20 4.12
*Double Support-Bonded DECS - 1.8 m x 4 mm; 170°C, 60 ml/min.
30
-------
Table 10. Retention Data for Chlorinated Phenols
on a Support-Bonded BDS Column*
DDT
Compound minutes 2,4,5-TCP
2,6-dichlorophenol 0.98 0.30
2,4-dichlorophenol 0.98 0.30
2,5-dichlorophenol 1.02 0.31
2,3-dichlorophenol 1.02 0.31
2,4,6-trichlorophenol 2.83 0.86
2,3,5-trichlorophenol 2.83 0.86
2,3,4-trichlorophenol 3.15 0.95
2,4,5-trichlorophenol 3.31 1.00
2,3,6-trichlorophenol 4.25 1.29
3,5-dichlorophenol 6.69 2.02
3,4-dichlorophenol 8.19 2.48
2,3,4,5-tetrachlorophenol 11.10 3.36
2,3,4,6-tetrachlorophenol 17.56 5.31
3,4,5-trichlorophenol 24.62 7.26
2,3,5,6-tetrachlorophenol 27.76 8.39
pentachlorophenol
^Support-Bonded BDS - 0.6 m x 4 mm; 190°C, 60 mL/um
31
-------
Table 11. Recoveries of Chlorinated Phenols from Fortified Urine
ppm added*
2,6-dichlorophenol
2,4-dichlorophenol
2,3-dichlorophenol
2,5-dichlorophenol
3,4-dichlorophenol
3,5-dichlorophenol
2,3,4-trichlorophenol
2,3,5-trichlorophenol
2,3,6-trlchlorophenol
2,4,5-trichlorophenol
2,4,6-trichlorophenol
3,4,5-trichlorophenol
2,3,5,6-tetrachlorophenol
2,3,4,6-tetrachlorophenol
2,3,4,5-tetrachlorophenol
pentachlorophenol
*Av. 5 determinations each fortification level.
**Range for all fortification levels.
0.01
Av % Rec 1,
83 ±
86 ±
86 ±
80 ±
85 ±
80 ±
88 ±
87 ±
85 ±
87 ±
87 ±
84 ±
79 ±
; SD
7.7
12.3
3.4
1.3
4.6
2.8
7.7
5.0
15.0
5.3
3.6
6.8
5.6
0.05
Av % Rec
96
89
83
83
86
82
87
81
92
89
98
82
91
94
92
86
% SD
± 5.6
± 5.3
±4.2
± 3.0
± 6.3
± 3.2
± 6.3
± 5.4
± 7.5
± 6.6
± 8.9
± 4.3
±9.2
± 7.7
± 10.3
± 4.9
0.10
Av % Rec
90
89
87
89
87
81
88
85
90
81
93
86
88
92
82
85
% SD
± 9.0
± 7.4
± 3.2
± 4.0
± 4.0
± 1.9
± 4.4
± 4.2
± 7.3
± 3.8
± 5.7
± 4.6
±7.2
± 5.3
± 7.1
± 4.9
0.50
Av % Rec
83
84
89
84
87
81
89
87
90
86
84
84
87
88
91
88
%
± 2
± 2
± 3
± 2
± 4
± 2
± 7
± 7
± 5
± 3
± 2
± 2
± 6
± 5
± 6
± 5
SD
.5
.8
.8
.4
.5
.3
.6
.5
.6
.1
.9
.8
.7
.1
.7
.2
1.00
Av % Rec
84
86
88
86
86
82
88
86
83
86
88
86
92
94
97
93
% SD
± 5.3
±4.1
± 3.8
± 4.1
±4.9
± 1.6
± 4.0
± 3.8
± 6.7
± 4.9
± 7.5
± 5.6
±7.6
± 6.0
± 4.7
±5.1
Range**
75-104
70-103
78-92
79-92
80-96
77-88
80-101
74-100
79-102
76-94
70-110
78-92
75-106
81-102
74-102
70-101
-------
listed. In addition, the presence and levels of chlorinated phenols in
pooled urine samples were confirmed by HPLC/MS. These samples were
taken from individuals who have no known exposure which can give chlorinated
phenols as metabolites in urine. These results allow for a background
or control level of expected chlorophenols in urine.
Figure 6 and 7 are representative chromatograms for a general human
urine sample free phenol analysis. As can be seen from the chromatographs,
a wide variety of GC operating parameters can be employed to analyze for
both early and late eluting chlorophenols.
33
-------
Table 12. General Human Population Urine Sample - Chlorinated Phenols
Residues in ppb
Sample Number 2,6-dcp 2,5-dcp 3,5-dcp 2.4,6-tcp 2.4,5-tcp 2,3,4,6-tcp PCP
1 <1 21 <15 2 7 27
2 <1 13 <15 6 7 49
3 38 20 <15 <1 7 24
4 <1 11 <15 1 2 26
5 <1 14 <15 1 8 3 11
6 <1 21 <15 1 3 2 14
7 <1 6 <15 1 3 28
8 <1 161 <15 4 9 15 74
9 <1 100 16 3 6 3 23
10 <1 10 <15 <1 <1 39
11 31 2 <15 2 2 <2 5
12 112 30 44 3 5 38
13 14 81 20 1 8 4 15
14 31 117 <15 3 3 6 11
15 76 159 29 <1 4 27 29
16 11 71 53 <1 <1 7 19
17 31 6 16 <1 17 5 11
18 <1 45 42 <1 2 8 16
19 21 117 21 <1 4 6 11
20 31 15 <15 <1 3 69
21 <1 13 <15 <1 2 43
22 40 132 <15 <1 5 76
23 <1 13 <15 <1 <1 35
24 <1 156 18 <1 37 99
25 <1 9 36 <1 4 7 14
26 <1 179 <15 <1 <1 6 13
27 <1 25 30 <1 <1 36
28 <1 23 11 2 4 78
29 <1 33 19 4 8 14 5
30 <1 56 16 5 16 4 26
31 <1 30 11 3 5 15 18
32 18 104 6 2 2 5 25
33 <1 208 10 4 18 6 25
34 12 35 9 5 5 9 28
-------
2 4 6 8 10 12 14 16 18 20 22 24 26
TIME, minutes
Figure 6. Chromatograms of (I) XAD-4 reagent blank and (II) human urine:
(a) 2, 6-dichlorophenol, equiv. 112 ppb; (b) 2, 4 - and/or 2, 5-dichlorophenol,
equiv. 30 ppb; (c) 2,4, 6-trichlorophenol, equiv. 3 ppb; (d) 2,4, 5-trichloro-
phenol, equiv. 5 ppb; (e) 2, 3, 4, 6-tetrachlorophenol, equiv. 3 ppb; (f) 3, 5-
dichlorophenol, equiv. 44 ppb. Column: DSB - DECS (1.8 x 4 mm 1.0) on
80/100 chrom W (AW); oven temperature, 170<>C; 5% methane in argon; flow
rate 60 mL/min.
-------
CO
z
O
O.
co
LU
DC
DC
LU
Q
DC
O
O
LJJ
DC
0
6 8
TIME, minutes
10
12
14
Figure 7. Chromatogram of human urine, (e) 2, 3, 4, 5-tetrachlorophenol, equiv. 3 ppb;
(f) 3, 5-dichlorophenol, equiv. 40 ppb; (g) pentachlorophenol, equiv. 8 ppb. Column:
DSB - DECS (0.6 m x 4 mm I. D.) on 80/100 Chrom W (AW); oven temperature 195OC;
5% methane in argon; flow rate 60 mL/min.
-------
REFERENCES
1. Burttschell, R. H., A. A. Rosen, F. M. Middleton, and M. B. Ettinger.
J. Am. Water Works Assoc. 51, 205, 1959.
2. Farrington, D. S. , and J. W. Munday, Analyst, 101, 639, 1976.
3. Lamparski, L. L., and T. V. Nestrick, J. Chromatogr., 156, 143,
1978.
4. Menzie, C. M. , Metabolism of Pesticides, U. S. Dept. of Interior,
Fish and Wildlife Service Special Scientific Report. Wildlife No.
127 (1969).
5. Menzie, C. M., Metabolism of Pesticides, An Update, U. S. Depart.
of Interior, Fish and Wildlife Service Special Scientific Report.
Wildlife No. 184 (1974).
6. Menzie, C. M., Metabolism of Pesticides, Update II, U. S. Dept. of
Interior, Fish and Wildlife Special Scientific Report. Wildlife
No. 212 (1978).
7. Bevenue, A., and H. Beckman, Residue Rev., 19, 83-134, 1967.
8. Bevenue, A., J. R. Wilson, L. J. Casarett, and H. W. Klemmer, Bull.
Environ. Contam. Toxicol. 2, 319-332, 1967.
9. Wyllie, J. A., J. Gabica, W. W. Benson, and J. Yoder, Pestic. Monit. J.
9, 150-153, 1975.
10. Lui, H., and G. D. Sweeney, Fed. Environ. Biochem. Soc. Lett., 51,
225, 1975. ~
11. Mehendale, H. M., M. Fields and H. B. Mathews, J. Agr. Food Chem.,
23, 261, 1975.
12. Koss, G. , W. Koransky and K. Steinbach, Arch. Toxicol., 35, 107,
1976.
13. Engst, R., R. M. Macholz and M. Kujawa, Bull. Environ. Cont. Toxicol.,
16, 250, 1976.
14. Renner, G., and K. P. Schuster, Toxicol. Appl. Pharm., 39, 355,
1977. ~~
15. Jakobson, I., and S. Yllner, Acta Pharmacol. Toxicol., 29, 513,
1971. ~
16. Ahlborg, U. G., J. E. Lindgren and M. Mercier, Arch. Toxicol., 32,
271, 1974. '
37
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17. Edgerton, T. R. , R. F. Moseman, R. E. Under, and L. H. Wright,
J. Chromatogr., 170, 331-342, 1979.
18. Edgerton, T. R., and R. F. Moseman, J. Chromatogr. Sci. , 18, 25-29,
1980.
19. Moseman, R. F., J. Chromatogr., 166, 397, 1978.
20. Junk, G. A., J. J. Richard, M. D. Geiger, D. Witiak, J. L. Witiak,
M. D. Argnello, R. Vick, H. J. Svec, J. S. Fritz, and G. V. Calder,
J. Chromatogr., 99, 745, 1974.
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TECHNICAL REPORT DATA
(Please read [nuntcrions on [he reverse before completing)
i. REPORT NO.
EPA-600/1-81-024
2.
3. RECIPIENT'S ACCESSION>NO.
4. TITLs AND SU3TITLS
METHOD DEVELOPMENT FOR THE ASSESSMENT OF POSSIBLE
HUMAN EXPOSURE TO PESTICIDES AND INDUSTRIAL CHEMICALS
5. REPORT OATS
March 1981
6. PERFORMING ORGANIZATION COOe
7. AUTHCR(S)
Thomas R, Edgerton, R.F, Moseman and L.H, Wright
3. PERFORMING ORGANIZATION REPORT NO
9. PERFORMING ORGANIZATION NAME AND ADDRESS
Analytical Chemistry Branch
Environmental Toxicology Division
Health Effects Research Laboratory
Research Triangle Park, NC 27711
10. PROGRAM ELEMENT NO.
ACAE1A
11. CONTRACT/GRANT NO.
12. SPONSORING AGENCY NAME AND ADDRESS
Health Effects Research Laboratory
Office of Research and Development
U.S. Environmental Protection Agency
Research Triangle Park, NC 27711
13. TYPE OF REPORT AND PERIOD COVERED
RTP, NC
14. SPONSORING AGENCY CODE
EPA^SOO/11
15. SUPPLEMENTARY NOTES
1S. ABSTRACT
The determination of chlorinated phenols in urine can be used as a means for
assessing exposure to pesticides and industrial chemicals in the human population,
A method was developed for the analysis of chlorinated phenols which involves
the derivatization of metabolites from the urine of rats fed hexachlorobenzene
(HCB) and pentachlorophenol (PCP). This method was then applied to urine samples
taken from the general human population to gain a background level. Pentachloro-*
phenol was detected in greater than 90% of the human samples analyzed, The only
other metabolites detected were tetrachloropyrocatectiol and tetrachlorohydroquinone
along with large amounts (greater than 3 ppm) of PCP. Pentachlorothiophenol (/ a
major metabolite of HCB fed to rats i' was not detected in human urine.
The analysis of human urine for underivatized chlorinated phenols using a
direct gas chromatographic method not requiring derivatization detected quantifiable
levels of 2,5-dichloro-,1 2i'4i;5>trichlorqT, 2,i3,4,6wtetrachloro> and penta;cjilorqV
phenol in greater than 90% of the samples examined. Approximately 50% of the
samples contained detectable levels of 2,'6 and 3l/5^ichlorophenol and
2,4,6^trichlorophenol,
17.
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cPA rorm 2220-1 (9-73)
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