tlAU-UlN-DOO/
United States August, 1988
Environmental Protection Revised April, 1991
Agency
Research and
Development
DRINKING WATER CRITERIA DOCUMENT FOR
HEXACHLOROCYCLOPENTADIENE
Prepared for
OFFICE OF WATER
Prepared by
Environmental Criteria and Assessment Office
Office of Health and Environmental Assessment
U.S. Environmental Protection Agency
Cincinnati, OH 45268
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DISCLAIMER
This document has been reviewed 1n accordance with the U.S. Environ-
mental Protection Agency's peer and administrative review policies and
approved for publication. Mention of trade names or commercial products
does not constitute endorsement or recommendation for use.
11
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FOREWORD
Section 1412 (b)(3)(A) of the Safe Drinking Mater Act, as amended 1n
1986, requires the Administrator of the Environmental Protection Agency to
publish maximum contaminant level goals (MCLGs) and promulgate National
Primary Drinking Water Regulations for each contaminant, which, In the
judgment of the Administrator, may have an adverse effect on public health
and which 1s known or anticipated to occur 1n public water systems. The
HCLG 1s nonenforceable and 1s set at a level at which no known or antici-
pated adverse health effects 1n humans occur and which allows for an
adequate margin of safety. Factors considered 1n setting the HCLG Include
health effects data and sources of exposure other than drinking water.
This document provides the health effects basis to be considered 1n
establishing the HCLG. To achieve this objective, data on pharmacoklnetlcs,
human exposure, acute and chronic toxldty to animals and.humans, epidemi-
ology and mechanisms of toxldty are evaluated. Specific emphasis Is placed
on literature data providing dose-response Information. Thus, while the
literature search and evaluation performed 1n support of this document has
been comprehensive, only the reports considered most pertinent In the deri-
vation of the HCLG are cited 1n the document. The comprehensive literature
data base 1n support of this document Includes Information published up to
1985; however, more recent data may have been added during the review
process. Editorial changes were also made 1n 1991 when this document was
finalized.
When adequate health effects data exist, Health Advisory values for less
than lifetime exposures (1-day, 10-day and longer-term, -10% of an
Individual's lifetime) are Included 1n this document. These values are not
used 1n setting the HCLG, but serve as Informal guidance to municipalities
and other organizations when emergency spills or contamination situations
occur.
Tudor Davles, Director
Office of Science and
Technology
James Elder, Director
Office of Ground Water
and Drinking Water
111
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DOCUMENT DEVELOPMENT
Linda R. Papa, Document Manager
Environmental Criteria and Assessment Office, Cincinnati
U.S. Environmental Protection Agency
Scientific Contributors
David J. Relsman
Environmental Criteria and
Assessment Office, Cincinnati
U.S. Environmental Protection Agency
James WHhey. Author of Chapter III and Reviewer
Environmental Health Directorate
Bureau of Chemical Hazards
Tunney's Pasture
Ottawa, Ontario K1A-OL2
Scientific Reviewers
W. Bruce Pelrano
Environmental Criteria and
Assessment Office, Cincinnati
U.S. Environmental Protection Agency
Annette M. Galchett
Environmental Criteria and
Assessment Office, Cincinnati
U.S. Environmental Protection Agency
C. Ralph Buncher
Department of Environmental Health
University of Cincinnati
Cincinnati, OH
Fumlo Matsumura
Director, Pesticide Research Center
Michigan Stale University
East Lansing. MI
Charles 0. Abernathy
Yogendra Patel
Health Effects Branch
Office of Drinking Water
U.S. Environmental Protection Agency
Washington. DC
Larry ValcovU
Reproductive Effects Assessment Group
Office of Health and Environmental
Assessment
U.S. Environmental Protection Agency
Washington, DC
Paul White
Exposure Assessment Group
Office of Health and Environmental
Assessment
U.S. Environmental Protection Agency
Washington, DC
Arthur Chlu
Carcinogen Assessment Group
Office of Health and Environmental
Assessment
U.S. Environmental Protection Agency
Washington, DC
Editorial Reviewer
Judith Olsen
Environmental Criteria and
Assessment Office. Cincinnati
U.S. Environmental Protection Agency
1v
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TABLE OF CONTENTS
Page
I. SUMMARY 1-1
II. PHYSICAL AND CHEMICAL PROPERTIES 11-1
ANALYSIS H-10
SUMMARY H-16
III. TOXICOKINET1CS 111-1
INTRODUCTION 111-1
INTRAVENOUS ROUTE III-l
ORAL ROUTE III-3
INHALATION ROUTE 1II-8
PERCUTANEOUS ROUTE 111-13
COMPARATIVE STUDIES > . . . 111-13
SUMMARY III-14
IV. HUMAN EXPOSURE 1V-1
(To be provided by the Office of Drinking Mater)
V. HEALTH EFFECTS IN ANIMALS V-l
OVERVIEW V-l
ACUTE TOXICITY V-l
SUBCHRON1C AND CHRONIC TOXICITY V-7
MUTAGENICITY V-15
CARCINOGEN1CITY V-16
TERATOGEN1CITY V-17
SUMMARY V-18
VI. HEALTH EFFECTS IN HUMANS VI-1
ACUTE EXPOSURE STUDIES Vl-1
EPIDEM10LOG1C STUDIES VI-8
SUMMARY VI-10
VII. MECHANISMS OF TOXICITY Vll-1
SUMMARY VI1-?
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TABLE OF CONTENTS (cont.)
Page
VIII. QUANTIFICATION OF TOXICOLOGIC EFFECTS V1I1-1
INTRODUCTION Vlll-1
NONCARCINOGENIC EFFECTS VI11-6
QUANTIFICATION OF NONCARCINOGENIC EFFECTS VI1I-11
Derivation of T-Day HA V1I1-11
Derivation of TO-Day HA VIII-12
Derivation of Longer-Term HA VI11-13
Assessment of Lifetime Exposure and Derivation of DUEL . VI1I-14
CARCINOGENIC EFFECTS VIII-15
EXISTING GUIDELINES, RECOMMENDATIONS AND STANDARDS VII1-1S
Occupational Standards -. . . . V1I1-15
Transportation Regulations VI1I-17
Solid Waste Regulations V11I-17
Food Tolerances V1I1-17
Water Regulations V11I-17
A1r Regulations VI11-18
Other Regulations Vlll-18
SPECIAL GROUPS AT RISK VI1I-19
SUMMARY VI11-19
IX. REFERENCES IX-1
vl
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LIST OF TABLES
No. Title Page
II-l Identity of Hexachlorocyclopentadlene 11-2
11-2 Physical Properties of Hexachlorocyclopentadlene 11-3
11-3 Characteristics of the Porapak* T Collection System . . . 11-12
11-4 Optimized GC Analytical Procedure for HEX 11-13
111-1 Extractablllty of [14C] HEX and Radioactivity Derived
from Saline and Various Biological Preparations 111-12
III-2 Disposition of Radioactivity Expressed as Percentage
of Administered Dose from 14C-HEX In Rats Dosed by
Various Routes 111-15
111-3 Fate of Radiocarbon Following Oral. Inhalation and
Intravenous Exposure to a«C-HEX In Rats Expressed
as Percentage of Administered Dose 111-16
1II-4 Distribution of HEX Equivalents In Tissues and Excreta
of Rats 72 Hours After Oral, Inhalation and Intravenous
Exposure to *«C-HEX 111-17
V-l Acute Toxlclty of HEX V-2
V-2 Subchronlc Toxlclty of HEX V-B
V-3 lexicological Parameters for Mice and Rats Administered
Technical Grade HEX In Corn Oil for 91 Days V-10
Vl-1 Symptoms of 145 Wastewater Treatment Plant Employees
Exposed to HEX (Louisville, KY, March 1977) VI-4
Vl-2 Abnormalities for 18 of 97 Cleanup Workers at
the Morris Forman Treatment Plant VI-5
VI-3 Overview of Individual Exposure - Symptomatology
Correlations at the Morris Forman Treatment Plant VI-6
VI-4 Hepatic Profile Comparison of Hardeman County:
Exposed Group (November 1978) and Control Group Vl-9
V1II-1 Summary of HAs and DWEL for NoncarclnogenU Effects .... V1II-16
vll
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LIST OF ABBREVIATIONS
CDC
DMSO
DWEL
ECO
6C/MS
Gl
HA
HEX
*«C-HEX
HPLC
l.d.
1.v.
LAQL
LD50
LOAEL
LOEL
Mg
NAS
NIOSH
NOAEL
NOEL
NTP
OCCP
Radlolabeled carbon dioxide
Centers for Disease Control
Dlmethylsulfoxlde
Drinking water equivalent level
Electron capture detection
Gas chromatography/mass spectrometry
Gastrointestinal
Health advisory
Hexachlorocyclopentadlene
Radlolabeled carbon-14 hexachlorocyclopentadlene
High performance liquid chromatography
Internal diameter
Intravenous
Lowest analytically quantifiable level
Lethal dose to 50% of recipients
Lowest-observed-adverse-effect level
Lowest-observed-effect level
Megagrams equivalent to 1 metric ton
National Academy of Sciences
National Institute for Occupational Safety and Health
No-observed-adverse-effect level
No-observed-effect level
National lexicology Program
Octachlorocyclopentadlene
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LIST OF ABBREVIATIONS (cent.)
RfD Reference dose
S.D. Standard deviation
SMR Standard mortality ratio
sp. gr. Specific gravity
SRI Southern Research Institute
Ix
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I. SUMMARY
HexachlorocyclopentacMene (HEX) 1s an unsaturated, highly reactive,
chlorinated cyclic hydrocarbon of low water solubility. HEX 1s a chemical
Intermediate used 1n the manufacture of chlorinated pesticides and flame
retardants with no end uses of Its own. The major source of environmental
contamination by HEX 1s the aqueous discharge from production facilities,
with small concentrations present as contaminants In commercial products
made from It. However, HEX 1s not frequently found 1n the environment and,
even when present, H 1s rapidly degraded. The degradation products of HEX
have not been Identified. Because of recent controls on environmental
emissions, current environmental exposure to HEX Is extremely low. From
time to time. Isolated Instances, such as the sewer system disposal of HEX
wastes In 1977 In Louisville, KY, and the cleanup of a large waste disposal
site 1n Michigan In 1983, have brought this chemical to the forefront of
environmental news.
HEX Is not readily absorbed by epithelial tissues because It Is highly
reactive, especially with the contents of the GI tract. HEX Is moderately
toxic when given orally, but has been estimated to be 100 times more toxic
when Inhaled. The data base for the long-term toxlclly of HEX Is very
limited. A chronic Inhalation bloassay Is being conducted by the NIP and
may provide data regarding any carcinogenic potential of HEX.
Several literature reviews on the health and environmental effects of
HEX are available. Although each of these reports varies In scope and
03610 1-1 08/24/88
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emphasis, a large amount of the scientific knowledge about HEX 1s reviewed
In these documents. To avoid unnecessary duplication, the previously
reviewed material will not be considered at great length, except where H
Impinges directly upon present critical considerations.
.HEX Is currently produced by only one company In the United States,
Velslcol Chemical Corporation. Production data are considered proprietary;
however, estimates show that 8-15 million pounds/year are produced. HEX can
enter the environment as an Impurity/contaminant In some of the products
using HEX as an Intermediate or can be released during the-manufacture of
products requiring HEX. The total estimated environmental release of HEX Is
11.9 Mg (13.1 tons). Because of Us physical and chemical characteristics,
only a small amount of this total can be expected to persist. In water, HEX
may undergo photolysis, hydrolysis and blodegradaUon. In shallow, standing
water, HEX has a photolytlc half-life of <1 hour, while In deeper waters
where photolysis 1s precluded, HEX may persist for several days. HEX Is
known to be quite volatile; however, the rate of volatilization from natural
waters can be Influenced by turbulence and adsorption onto sediments.
In animal studies, the absorption of unchanged HEX 1s reduced because of
Its reactivity with membranes and tissues, and especially with the contents
of the GI tract. Radioactivity from *«C-HEX 1s retained by the kidneys
and livers of animals for at least 72 hours after oral or Inhalation
dosing. Absorbed HEX 1s metabolized and rapidly excreted, predominantly In
the urine and feces with <1X of the HEX found In expired air. The
metabolites of HEX have not been Identified.
03610 1-2 08/30/88
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Subchronlc oral dosing studies have been conducted on rats and mice for
91 days. At 38 and 75 mg/kg/day levels In rats and mice, respectively,
adverse effects were noted, which Included Inflammation and hyperplasla of
the forestomach and nephrosls. In. vitro test results from three species
have not shown HEX to be a mutagen. HEX was also Inactive In the mouse
dominant lethal assay.
Limited data are available on the health effects 1n humans from exposure
to HEX. Isolated Instances have occurred that show Inhalation exposure to
HEX causes severe Irritation of the eyes, nose, throat and lungs. Human
exposures from other routes have elicited effects that Include short-term
Irritations, with recovery after cessation of exposure. The long-term
health effects of continuous low-level and/or Intermittent acute exposure 1n
humans are not known.
The data base Is neither extensive nor adequate for assessing the car-
dnogenldty of HEX. The NTP has recently completed a subchronlc Inhalation
animal study and will begin a lifetime animal Inhalation bloassay using both
rats and mice. Several epldemlologlc studies were cited 1n the literature;
however, no Increased Incidences of neoplasms at any site were reported that
could be related to HEX. Accordingly. Velslcol Chemical Corporation has
on-going programs and follow-up studies In order to study the long-term
effects of HEX exposure. A Judgment of carcinogenic potential will be
deferred until the results of the NTP long-term bloassay are available.
Using the International Agency for Research on Cancer (IARC) criteria, the
available evidence would place HEX In the Group 3 category. According to
the U.S. EPA Guidelines for Carcinogen Risk Assessment, based upon present
03610 1-3 08/24/88
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data, this chemical Is classified In the Group D category. A U.S. EPA
carclnogenldty Group 0 classification was verified by the CRAVE Work Group
on 10/05/89. This classification Indicates that the available data base 1s
Inadequate to assess the carcinogenic potential of this substance.
Using a gavage study on rats, the 1-day health advisory (HA) for
children has been calculated to be 15 mg/l. The 10-day HA was derived
using a repeated-dose toxlclty study on both rats and mice, and for children
the level Is recommended to be 2 mg/i.
The longer-term HAs were developed from the SRI 13-week study also used
for the derivation of the Drinking Water Equivalent Level (DWEL). The
recommended longer-term HA for adults 1s 3 mg/8. and for children the
longer-term HA 1s 0.7 mg/i. The DWEL Is 0.3 mg/i, which 1s calculated
using an RfD of 0.007 mg/kg bw/day (verification date 10/09/85) based on a
NOAEL of 10 mg/kg for absence of adverse effects In rats.
03610 1-4 06/11/91
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II. PHYSICAL AND CHEMICAL PROPERTIES
Hexachlorocyclopentadlene (HEX, HCCPD, C-56) Is a nonflammable liquid
with a characteristically pungent, musty odor. The pure compound Is a pale
yellow but Impurities may produce a greenish tinge (Stevens. 1979). HEX Is
a dense liquid with a specific gravity of 1.7019 at 25°C and low water
solubility (0.80S-2.1 mg/i) (U.S. EPA, 1978). Studies by Nalshteln and
Llsovskaya {1965} have Indicated an odor threshold for HEX of 0.002 mg/i
and a "taste" threshold of 0.007 mg/i. HEX 1s a liquid with a high
boiling point (239°C) and a vapor pressure of 0.08 mm Hg at 25°C. Table
II-l presents Information on the Identity of the chemical while Table 11-2
lists Its physical properties.
HEX Is stable under moisture-free and Iron-free conditions (Stevens,
1979). Chemically, HEX Is a highly reactive dlene that readily undergoes
addition and substitution reactions and also participates In Dlels-Alder
reactions (Ungnade and McBee, 1958). The products of the Dlels-Alder
reaction of HEX are generally 1:1 adducts containing a hexachlorobicyclo-
(2,2,1Jheptene structure; the monoene derived part of the adduct is nearly
always 1n the endo-posltlon. rather than the exo-posltlon (Stevens, 1979).
Look (1974) also reviewed the formation of HEX adducts of aromatic compounds
and the by-products of the Dlels-Alder reaction. Two early reviews of the
chemistry of HEX were publUhPd by Roberts (1958) and Ungnade and McBee
(1958).
HEX has an absorption band In the ultraviolet range at 322 and 3?3 nm
(log e = 3.17) 1n ethanol (U.S. EPA, 1978). This absorption band reaches
03620 11-1 04/12/91
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TABLE II-l
Identity of Hexachlorocyclopentadlene*
Identifying Characteristic
Name/Number/Structure
IUPAC Name:
Trade Names:
Synonyms:
CAS Number
CIS Accession Number:
Molecular Formula:
Molecular Structure:
1.2>3,4.5t5l-Hexachloro-l,3-cyclopentad1ene
C56; MRS 1655; Graph!ox
Hexachlorocyclopentadlene
Perchlorocyclopentadlene
HEX
HCPD
HCCP
HCCPD
C-56
HRS 1655
Graphlox
77-47-4
7800117
Cl,
JC1
Cl
Cl
'Source: U.S. EPA. 1984
03620
II-2
10/16/85
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TABLE 11-2
Physical Properties of Hexachlorocyclopentadlene
Property
Value/Description
Reference
Molecular weight
Physical form (25°C)
Odor
Odor threshold: air
water
Electronic absorption
maximum [(1n 50%
acelon1tr1le-water)]
Solubility 1n water
(rag/l)
Organic solvents
Vapor density (air = 1)
Vapor pressure
(mm Hg, °C)
Specific gravity
Melting point ("CJ*
Boiling point (°C)
Conversion Factor
272.79
Pale yellow liquid
Pungent
0.03 ppm (v/v)
0.0077 ppm (w/v)
313 nm
3.4 (20°C)
2.1 (25°C)
1.8 (28°C)
Mlsclble (hexane)
9.4
0.08 (25°C)
0.975 (62°C)
1.717 (15°C)
1.710 (20°C)
1.7019 (25°C}
9.6-11.34
239
234
Stevens, 1979
Hawley, 1977; Irish. 1963
Hawley, 1977; Irish. 1963
Amoore and Hautala, 1983
Wolfe et al.. 198?
Horvath, 1982
Dal Monte and Yu, 1977
Wolfe et al.. 1982
U.S. EPA, 1978
Verschueren, 1977
Irish. 1963
Stevens, 1979
Hawley, 1977
Stevens, 1979
Weast and Astle, 1980
U.S. EPA, 1978; Hawley,
1977; Stevens. 1979
Weast and Astle. 1980
Aldrlch Chemical
Company, 1988
Hawley, 1977; Stevens, 1979
Irish, 1963
1 ppm = 11.17 mg/cu.m Irish, 1963
03620
11-3
04/12/91
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TABLE II-2 (cont.)
Property
Octane! /Hater partition
coefficient (log P)
(measured)
(estimated)
Kow
Latent heat of vapori-
zation
Henry's Law constant
(atm-mVmole)
Value/Description
5.04+0.04
5.51
1.1x10*
176.6 J/g
2.7xlO'2
Reference
Wolfe et al.,
Wolfe et al.,
Wolfe et al.,
Stevens, 1979
Atallah et al
Wolfe et all,
1982
1982
1982
., 1980;
1982
*A wide range of melting points have been reported for this chemical; this
variation may be due to chemical Impurities and/or Isomerlc structure.
03620
11-4
06/19/90
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Into the visible spectrum, as evidenced by the yellow color of HEX. Facile
carbon-chlorine bond scission might be expected In sunlight or under
fluorescent light. The 1R spectrum of the dlene has two absorption bands at
6.2 and 6.3 ym In the double bond region and three bands at 12.4, 14.1 and
14.7 ym 1n the C-C region. The mass spectrum of HEX shows a weak
molecular Ion (M) at M/e 270, but also a very Intense (H-35) Ion making this
latter Ion suitable for sensitive, specific Ion monitoring.
HEX Is produced for commercial use only by Velslcol Chemical Corporation
(SRI, 1987; USITC, 1986). Current production data are not available. U.S.
EPA (1984) estimated that between 8 and 15 million pounds/year {4000-7500
tons) are produced. In a report prepared for the U.S. EPA, Hunt and Brooks
(1984} estimated that 8300 Hg (9130 tons) of HEX were produced In the United
States In 1983.
Commercial HEX contains various Impurities depending upon the method of
synthesis. The three primary processes commonly used for the production of
HEX are chlorlnatlon of cyclopentadlene, dechlorlnatlon of octachloro-
cyclopentene and the dehydrochlorlnatlon of hexachlorocyclopentane. In the
first process, cyclopentadlene Is mixed with alkaline hypochlorlte at 40°C.
HEX 1s recovered by fractional distillation, and contains appreciable
quantities of lower chlorinated cyclopentadlenes. In the dechlorlnatlon
process, HEX Is produced by thermal dechlorlnatlon of octachlorocyclopentene
at 470-480eC (Stevens, 1979). Technical grade HEX usually . contains
contaminants such as hexachlorobenzene, octachlorocyclopentene, PCBs and
mlrex. The nature and levels of contaminants will vary with the method of
production. Commercial HEX produced by Velslcol Chemical Corporation at the
03620 11-5 08/25/88
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Memphis, TN plant, which 1s used Internally and sold to other users, has a
97% minimum purity (Velslcol Chemical Corporation, 1984).
Although HEX has essentially no end use of Us own, It has been used as
a chemical Intermediate In the production of several Insecticides/pesticides
Including aldrln, dleldrln, endrln, chlordane, heptachlor, endosulfan,
pentac, mlrex and others and 1n the manufacture of flame retardants such as
wet add chlorendlc add, and Dechlorane plus* (Stevens, 1979). With the
exception of endosulfan and pentac, the use of HEX-based pesticides has been
banned, suspended or severely restricted (U.S. EPA, 1980). Figure 11-1
Illustrates synthetic pathways for the various chlorinated pesticides. HEX
1s also used to a lesser extent In the manufacture of resins and dyes (U.S.
EPA, 1980), and has been used previously as a general bloclde (Cole, 1954).
When studying HEX 1n water, there are numerous physical properties that
Influence Us ultimate fate. In the event of release Into shallow or flow-
Ing bodies of water, degenerative processes such as photolysis, hydrolysis
and blodegradatlon, as well as transport processes Involving volatilization,
evaporation and other physical loss mechanisms, are expected to be prominent
1n HEX dissipation. In deeper, nonflowlng bodies of water, hydrolysis and
blodegradatlon may become the predominant fate processes (U.S. EPA, 1984).
Zepp et al. (1979, 1984) and Wolfe et al. (1982) reported the results of
U.S. EPA studies on the rate of HEX phototransformatlon 1n water. Under a
variety of natural sunlight conditions, 1n both distilled and natural waters
of 1-4 cm depth, phototransformatlon half-life was <10 minutes. Rate
constants were also computed for various times of the day at 40°N latitude.
03620 11-6 08/25/88
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iNtMlN
o
GO
IV)
o
o
OS
CO
CD
INOO1UITM4
«'Clj. I.Oj Oi 'Ullllt
UIIN IN CCI«OI CtIV
Ot »0,CI, . IINIOVt
Mill II
FIGURE II-l
Synthesis of Chlorinated Cyclodlene Pesticides from Hexachlorocyclopentadlene
Source: U.S. EPA, 1984
-------
The rate constant for near-surface photolysis of HEX was estimated as 3.9
hour"1. This corresponds to a half-life of -11 minutes. Addition of
natural sediments to distilled water containing HEX had little effect on the
phototransformatlon rate. These findings Indicated that the predominant
mechanism of HEX phototransformatlon (decomposition) was direct absorption
of light by the chemical, rather than photosensltlzatlon reactions Involving
other dissolved or suspended materials. Although no product was Identified,
Zepp et al. (1984) and Wolfe et al. (1982) speculated that the primary
phototransformatlon product was hydrated tetrachlorocyclopentadlenone, which
dlmerlzes or reacts to form higher molecular weight products. The dlmerlza-
tlon of hexachlorocyclopentadlene to form higher molecular weight products
represents a minor pathway according to Butz et al. (1982) and Yu and
Atallah (1977b). According to Chu et al. (1987), the photolysis half-life
of HEX 1n water was <4 minutes when exposed to sunlight. These authors
positively or tentatively Identified at least eight photoproducts:
2,3,4,4,5-pentachloro-2-cyclopentenone; hexachloro-2-cyclopentenone; hexa-
chloro-3-cyclopentenone (primary products); pentachloro-ds-2,4-pentad1eno1c
acid; 1- and E-pentachlorobutadlene, tetrachlorobutyne (secondary products)
and hexachlorolndenone (minor product).
Studies of the hydrolysis of HEX Indicate that at 25-30°C and 1n the
environmental pH range of 5-9, a hydrolytlc half-life of -3-11 days Is
observed (Wolfe et al., 1982; Yu and Atallah, 1977a). In comparison,
hydrolysis Is much slower than photolysis, but may be a significant load-
reducing process 1n waters where photolysis and physical transport processes
are not Important (I.e., In deep, nonflowlng waters). Wolfe et al. (1982)
found hydrolysis of HEX to be Independent of pH over an environmental range
03620 11-8 08/25/88
-------
of 3-10. Also, the addition of natural sediments, sufficient to adsorb <92%
of the compound, caused the rate constant to vary by less than a factor of
2. It was therefore concluded that sorptlon to sediments would not
significantly affect the rate of hydrolysis (Wolfe et al., 1982). In both
studies (Wolfe et al., 1982; Yu and Atallah, 1977a) the authors concluded
that the hydrolysis rate was higher as temperatures Increased and although
the hydrolysis products were not Identified, high molecular weight
polyhydroxyl compounds appear to be major products.
The rate constants for the oxidation of hexachloropentadlene with
singlet oxygen ^O.) and penoxy radicals 1n water were estimated at
<103 and 12 M~a hour'1, respectively (Mabey et al., 1982). If the
concentrations of 10. and ROp radicals In water are assumed to be
10'" and 10'9 M, respectively (Mill and Mabey, 1985), It 1s likely that
under ordinary environmental conditions, oxidation of HEX will not be
significant.
The Intermedia transport of HEX from water may occur through
volatilization Into air, adsorption onto suspended partkulate matters and
subsequent sedimentation and uptake by plants and animals In water. The
significance of HEX sorptlon In water was predicted by Wolfe et al. (1982)
using a computer simulated Exposure Analysis Modeling System (EXAMS). The
distribution of HEX 1n the sediments of a river, pond, eutrophlc and
ollgotrophU lake was estimated to be 98.8, 86, 87 and 97.IX, respectively,
of the total HEX In the system. Johnson and Young (1983) observed that
adsorption plays an Important role 1n reducing the concentration of HEX In
aqueous solutions. The predicted strong sorptlon of HEX In sediments is
also supported by experimental sorptlon data 1n soils.
03620 11-9 08/25/88
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However, the main removal of HEX from water bodies comes from
volatility. Weber (1979) measured the volatility of "C-HLX from
distilled water following the Incubation of the glass-stoppered and
unstoppered test bottles shaken at room temperature for 24 hours. From the
full glass-stoppered bottles containing 0.41 mg/s. HEX. only 4-5% of HEX
was lost, while 1n the half-full stoppered bottles, 15-16% of the chemical
was missing. The volatilization of HEX from the half-full, unstoppered
bottles was 45-47% over the 24-hour period. Other researchers (Kllzer el
al., 1979; Atallah et a!., 1980) have determined very high rates of
volatilization for HEX. Kllzer et al. (1979) reported that the volatility
of HEX ranged from 4.7-8.8%/hour from a static aqueous solution containing
50 yg/a HEA. Atallah et al. (1980) observed >80% volatilization In 24
hours from unlnoculated media containing 45 mg/8. HEX.
Blodegradation may also be a significant process 1n certain waters
(Tabak et al., 1981), although the evidence 1s limited. Tabak et al. (1981)
observed complete degradation of HEX at concentrations of 5 and 10 mg/a
within 7 days In a settled domestic wastewater culture system.
Blodegradation of <2.5% 14C-HEX by acclimated mixed microorganisms was
observed 1n 2-3 weeks by Atallah et al. (1980), while Wolfe et al. (1982)
observed no difference In degradation rate when sterile and nonslerile
natural sediments were added to HEX solutions.
Analysis
Gas chromatography Is the preferred method for analyzing HEX In air
using either flame lonlzatlon collection or GC-63Ni electron capture
03620 11-10 04/12/91
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detection (ECD) (Chopra et al.t 1978; Neumelster and Kurlmo, 1978; HhHmore
et al., 1977; NIOSH. 1979). GC/MS Is necessary for confirmation (Elchler,
1978).
Several sorbent materials were evaluated for collection of HEX vapor:
Amber 1 He® XAD-? (20/50 mesh), Porapak® R (50/80 mesh). Amber sorb®
XE-340 (20/50 mesh}. Chromosorb® 104 (60/80 mesh). Tenax-GC® (35/60
mesh), Porapak* T (80/100 mesh) and Porapak® T (50/80 mesh). According
to the NIOSH criterion for acceptable methods, a sorbent material must have
a demonstrated sorptlon capacity for the analyte that Is adequate for
sampling a reasonable volume of workplace air at an established rate. Table
II-3 enumerates additional factors related to the Porapak® T collection
system.
Gas chromatography with ECO was determined to be the most sensitive
analytical technique (Boyd et al., 1981). For HEX the chromatographlc
response was stated to be a linear and reproducible function of HEX
concentration 1n the range of -5-142 ng/mi (25-710 pg Injected), with a
correlation coefficient of 0.9993 for peak height measurement. The
optimized operating conditions for this method are shown In Table 11-4.
Validation tests were conducted according to NIOSH guidelines. The
accuracy and precision of the overall sampling and analytical procedure for
HEX were evaluated 1n the concentration range of -13-865 yg/m3 at
25-28°C and a relative humidity >907.. The LAQL of HEX was determined to be
25 ng/sorbent tube, assuming 1 ma. of hexane-desorblng solvent and a 1 hour
desorptlon time by ultrasonlfUatlon. The upper limit of the method was
03620 11-11 08/25/88
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TABLE II-3
Characteristics of the Porapak* 1 Collection System3
Characteristic HEX Type/Value
Sorbent material Porapak* Tb
(80/100 mesh)
Breakthrough t1mec >8 hour (0.2 l/mlnute)
Breakthrough volume0 >100 fi.
Tube capacity0 >100 g
Average desorptlon 0.94 (27.4 ng)
efficiency of Indicated
quantity of analyte
Sorbent tube 75 mg sorblng layer,
configuration0" 25 mg backup layer
Extraction solvent Hexane
aAdapted from Boyd et al., 1981
bTh1s material required cleaning by Soxhlet extraction (see text).
cFor these tests the temperature of the generator effluent was maintained
at 25-28°C and the relative humidity at >9054. The concentration of the
analyte 1n the generator effluent was 1 mg/m3 of HEX.
sorbent tubes were Pyrex (7 cm long by 6 mm o.d. and 4 mm 1.d.).
Sllanlzed glass wool plugs separated the sections.
03620 11-12 08/11/88
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TABLE II-4
Optimized GC Analytical Procedure for HEXa-b
Characteristic
Type/Value
Detector
Column
Electron capture
354 OV-1 on Gas-Chrom Q
(100/120 mesh) 1n glass
(4 mm 1.d. by 2 m)
OPERATING CONDITIONS
Carrier gas
(20 ml/minute]
Temperatures
Injection port
Column
Detector
Detector parameters
Solvent For compound0
5% CH4, 95% Ar
150°C
135°C
250°C
Detector purge, 5% CH4 with
95% Ar (80 mi/minute)
Hexane
aAdapted from Boyd et al., 1981
bA Hewlett-Packard 5750A gas chromatograph was used.
cThe Injection volume was 5 pi of sample and 1 yi of solvent flush.
03620
11-13
08/11/88
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2500 ng/sorbent sample. This 2500 ng level represents the smallest amount
of HEX that can be determined with a recovery of >80% and a relative
standard deviation (RSD) of <10X. The desorptlon efficiency of 100% was
determined by averaging the levels ranging from near the LAQL of 25 ng to
1000 times the LAQL.
For analysis of HEX 1n water, adequate precautions must be taken since
HEX 1s sensitive to light 1n both organic and aqueous solutions; therefore,
the water samples, extracts and standard HEX solutions must be protected.
The rate of degradation 1s dependent upon the Intensity and wavelength, with
the half-life of HEX being -7 days when the solution 1s exposed to ordinary
laboratory lighting conditions. Storing the HEX-conta1n1ng solutions In
amber or red (low actinic) colored glassware 1s recommended for adequate
protection (Benolt and Williams, 1981).
The XAD-2 resin extraction has been used to concentrate HEX from large
volumes of water. Solvent extraction of water has also proved successful.
The detection limit used for the organic solvent extraction technique was 50
ng/8. vs. 0.5 ng/fc for the XAD-2 method. Using the solvent extraction
method under subdued laboratory lighting conditions, the efficiency of
recovery for an artificially loaded water sample was In the range of 79-88%.
The authors concluded that the XAD-2 resin could not be used to accurately
sample HEX In water, but could be used to screen samples qualitatively
because of the poor detection limit (Benolt and Williams, 1981).
DeLeon et al. (1980) developed a method for determining volatile and
semlvolatlle organochlorlne compounds In soil and chemical waste disposal
03620 11-14 08/25/88
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site samples. This procedure Involves hexane extraction followed by
analysis of the extract by temperature-programmed gas chromatography on
high-resolution glass capillary columns using ECD; GC/HS Is used for
confirmation of the presence of the chlorocarbons. The method has a
detection limit of 10 vg/g.
When a soil sample was spiked with a 10 pg/g concentration of HEX, the
recovery was 59.8% (S.D. 6.1}; at 100 vg/g, 95.9% (S.D. 15.9); at 300
yg/g, 90.2% (S.D. 4.1). Of the 11 different compounds tested, the 100
vg/g HEX sample had the highest standard deviation Indicating that
utilizing this method for HEX may have limitations (DeLeon et al., 1980).
Velslcol Chemical Corporation (1979) has developed analytical methods
that have been used for detecting HEX 1n urine, adipose tissue, liver,
kidney and muscle tissues. The respective recoveries and standard
deviations for these tests were (1n percentages): 80+10 (1-50 ppb), 85+_2,
69*4, 71+3 and 76+4. The level of fortification for the tissue samples was
10 ppb. For urine, <31% HEX could be degraded when the fortified urine
sample was stored overnight 1n a cooler.
Urine was extracted with hexane, the hexane passed through anhydrous
sodium sulfate, and evaporated to 1 ma. The limit of detection for HEX
without concentrating the extract was 0.5 ppb. For cattle, poultry and fish
tissues, the tissues were extracted with 2:1 pentane/acetone, the homogenate
diluted with 10% sodium chloride solution, centMfuged, and the pentane/
acetone layer transferred Into a separatory funnel. The residues were then
partitioned Into acetonltrUe (3 times), water diluent added to the aceto-
nltrlle, and then back-extracted with pentane. The pentane extract was
03620 11-15 08/25/88
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treated with concentrated sulfurlc acid and then water, and concentrated to
~3 ma. Upon dilution to 10 ml with hexane, the solution was treated
with a 1:1 concentrated sulfurlc add/fuming sulfurlc acid solution, water.
and a 9:1 mixture (solid) of sodium sulfate/sodlum carbonate. Packed
columns (3% OV-1 on Gas Chrom Q-100/120 mesh-1n 2 m x 2 mm 1.d. glass
column) or capillary columns (30 m x 0.25 mm SE-30 WCOT) can be used for GC
using a 63N1-electron capture detector.
Summary
HEX 1s an unsaturated, highly reactive, chlorinated cyc-l1c hydrocarbon
of low water solubility and high volatility. HEX 1s used primarily as a
chemical Intermediate In the manufacture of chlorinated pesticides and flame
retardants with essentially no end uses of Its own. Low levels of HEX have
been detected 1n the environment; however, when present, 1t Is rapidly
degraded by a number of physical processes.
Several analytical methods have been developed for Identifying and
quantifying HEX In various media. Although HEX may be found In water,
because of Us physical and chemical characteristics, only a small amount
can be expected to persist. In water, HEX may undergo photolysis,
hydrolysis and blodegradatlon. In shallow water, HEX has a photolytlc
half-life of <1 hour. In deeper water where photolysis Is precluded, the
hydrolytlc half-life of HEX Is several days, while blodegradatlon 1s
predicted to occur more slowly. The rate of volatilization of HEX from
distilled water may be as high as 4.7-8.854/hour. The rate of volatilization
from natural waters will depend upon depth, turbulence and wind speed.
Adsorption to sediments will also greatly reduce the rate of evaporation
from natural waters.
03620 11-16 08/25/88
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111. TOX1COK1NETICS
Introduction
Limited studies on pharmacoklnetlc parameters of HEX have been reported
In the literature. Those studies that have addressed the disposition and
fate of HEX have been conducted with the 1«C-labeled compound and dealt
wUh the total radioactivity rather than HEX per se. Ho metabolites of HEX
have been unequivocally Identified; however, some characteristics, such as
their volatility and solubility, have been recorded.
The principal route of HEX entry Into the human body as a. contaminant In
water would be by the oral route, although In Industrial settings H Is
conceivable that HEX could form an aerosol mist or exist at fairly low con-
centrations as a vapor; therefore, there Is the potential for pulmonary
uptake. Dermal exposure during handling procedures or accidental spills
should also be considered as a possible route of HEX uptake.
Intravenous Route
Yu and Atallah (1981) administered a dose of 0.25 mg (-0.75 mg/kg) of
»«C-HEX (with an activity of 10.b mCl/mmole) as 0.3 mi of a solution 1n
20% Emulphor EL620/sal1ne 1rt the lateral caudal vein of male and female
Sprague-Dawley rats weighing between 240 and 350 g. Sequential blood
samples, taken over a period of 24 hours post-dosing, revealed that the
elimination of HEX followed a two-compartment .open pharmacoklnetlc model
with a half-life of -0.7 and -32 hours for the Initial and terminal phases,
respectively.
Following the administration of the l.v. dose, Yu and Atallah (1981)
found that -18% of the "C-labeled HEX was excreted In the feces and -21%
03630 II1-1 08/15/88
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1n the urine. The radlolabel was found In all examined animal tissues.
However, at 24 hours post-dosing, the bulk of the radlolabel was still 1n
the blood (-20%), with lower amounts In the liver (-5%), kidneys (-2%) and
the fat (-2%). Also, 9% of the administered radlolabel was found In the GI
tract (duodenum, large and small Intestine), which 1s consistent with the
earlier observation by Mehendale (1977) that some excretion occurred In the
bile. Of the total administered dose. 67% was recovered within 24 hours
post-dosing. Metabolites were not characterized In the part of the study
concerned with the fate of HEX after 1.v. administration.
Lawrence and Dorough (1982) examined the uptake, deposition and elimina-
tion of HEX following l.v. exposure of female albino Sprague-Dawley rats.
Intravenous doses of 0.01 mg/kg of 14C-HEX were administered In DMSO or
10:4:1 sallne-.propylene glycol :ethanol 1n 0.2 ml of solution. Recovery of
radiocarbon Immediately after treatment was 101.2+4.7%, based on calculated
doses. Elimination of the dose occurred predominantly during the first 24
hours In the feces and urine. Within 72 hours following treatment, 22% of
the radiocarbon was excreted 1n the urine, 31.4% was found In the feces and
31% was retained In the body. Of the 31% retained In the body, the kidneys
(22.3%), lungs (14.9%) and liver (9.6%) were the sites of residue
accumulation. Accumulation In adipose tissue was not significant. Biliary
excretion was <14%, Indicating that -50% of the radiocarbon excreted 1n the
feces was unabsorbed.
In a similar study, El Dareer et al. (1983) administered male Fischer
344 rats doses of 0.59 mg/kg "C-HEX In Emulphor EL-620:ethanol:water
(1:1:4 v/v) In a volume of 0.15 ma/150 g bw. A specific activity of 18
03630 III-2 08/15/88
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vCVmg HEX was used for the 1.v. dose. At 72 hours after treatment 34.0,
15.8 and 39.0% of the dose was found In the feces, urine and body tissues,
respectively. Of the 39% retained In the body, 13.9 and 18.4% were found In
the liver and carcass, respectively. About 0.1% was excreted as
"C-carbon dioxide during the observation period.
Oral Route
In the same communication In which the pharmacoklnetlcs of HEX after
l.v. administration was reported, Lawrence and Dorough (1982) also described
the results of administering a single oral dose of HEX to rats. Female
Sprague-Dawley rats (175-250 g bw) were administered 5 vg/kg 14C-HEX by
Intubation as 0.5 ml of solution 1n corn oil. Recovery of the radiocarbon
Immediately after oral treatment based on calculated dose was 98.0%. At 72
hours, 68.2% of the 5 vg/kg of 14C-HEX had been eliminated 1n the feces,
24.4% In the urine and 2.8% of the dose was retained In the body. Biliary
excretion accounted for 18% of the oral dose, Indicating the radiocarbon
eliminated In the feces (-50% of the dose) was not absorbed from the Gl
Irtct.
Mehendale (1977) administered i4C-HEX (5 pmole; 6 mg/kg) In corn oil
to four male Sprague-Dawley rats by oral Intubation. Dally urine and fecal
samples were collected. After 7 days -33% of the administered dose was
excreted 1n the urine, 87% of which was eliminated within the first 24 hours
after treatment. Fecal excretion was equivalent to 10% of the dose, -60% of
which was excreted within 24 hours. Only a small amount (-0.5%) of the
original dose was recovered In the kidneys and liver. Hehendale (1977)
speculated that. 1n view of the low total recovery of the administered dose,
03630 III-3 08/15/88
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a major part of the dose (>50?4) had been excreted through the lung.
Subsequent studies by Dorough (1979) found that <1X of an oral dose of
**C-HEX was excreted 1n the lungs. El Dareer et al. (1983) reported that,
after oral dosing, HEX and Us volatile metabolites can be readily lost If
the samples were dried and powdered In their analysis.
Dorough (1979) and Dorough and Ranlerl (1984) Investigated the accumula-
tion, distribution and excretion of radlolabeled HEX following Us adminis-
tration to male and female Sprague-Dawley albino rats and mice, either as a
single oral dose or as a component of their diet. To effectively account
for the radiocarbon administered to the test animals, two female rats were
dosed by gavage with 14C-HEX at 20 mg/kg In 0.9 ma of corn oil. The
animals were Immediately placed In separate metabolism cages through which
air was drawn at 600 ml/minute. Passing the expired air through, toluene
traps showed that <1% of the administered dose was voided as respiratory
gases.
Single dose studies (Dorough, 1979; Dorough and Ranlerl, 1984) were
conducted by administering 14C-HEX at doses of either 2.5 or 25 mg/kg bw
dissolved In 0.9 ma of corn oil for rats and 0.2-0.3 ma to mice using a
feeding needle. Animals were killed at 1. 3 and 7 days post-dosing and
samples of muscle, brain, liver, kidney, fat and either ovaries or testes
were removed and analyzed for 14C-act1vUy. Urine and feces were also
collected during the period between dosing and tissue sample collection. No
appreciable differences due to sex or species were found In the excretion
patterns. After 7 days, animals administered 2.5 and 25 mg/kg HEX had
eliminated an average of 16.4 and 15.8% of the dose In urine, and 63.3 and
03630 1II-4 08/15/88
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75.2% In the feces, respectively. The liver, kidney and fat were the most
Important sites of deposition for l4C-res1dues In both rats and mice, the
levels In the kidneys of rats and liver for mice being the highest.
In the same study, rats and mice were also placed on diets containing 0,
1, 5 or 25 ppm of 14C-HEX for 30 days. Assuming a dally Intake of 15 g
for rats and 5 g for mice, this would give dose rates of 0, 0.066, 0.330 and
1.666 mg/kg/day for rats and 0, 0.182, 0.910 and 4.55 mg/kg/day for mice.
Feed was replaced In the feeders every 12 hours to minimize the loss of
»4C-HEX (from volatilization) and killed at 1, 3, 7, 12, 1'5 and 30 days.
The surviving animals were then returned to a normal diet for <30 days and,
during this post-treatment period, animals were killed at 1, 3, 7, 15 and 30
days after the last day of treatment.
The total excretion (urine and feces} of the radlolabel ranged from
63-7954 of the consumed 14C-HEX. In all cases, the liver, kidney and fat
contained the highest amounts of 14C-label and a steady-state for these
levels appeared to be reached after 15 days of the feeding phase. A good
correlation was observed between the level of HEX 1n the diet and the
14C-levels found In all of the tissues examined. In a separate experiment
with male rats, 1n which the bile duct was cannulated and a single dose of
25 mg/kg of »*C-HEX was administered orally, only 16% of the dose was
excreted 1n the bile.
The extraction characteristics of the radiocarbon compounds In the
excreta showed that they were largely polar metabolites, some of which were
capable of being changed to organic soluble compounds after add catalyzed
hydrolysis.
03630 III-5 08/15/88
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In a comparative study of the pharmacoklnetlcs of 14C-HEX after l.v.
and oral dosing, Yu and Atallah (1981) administered single oral doses of 3
or 6 mg of J*C-HEX (specific activity. 0.267 mC1/mmole) to Sprague-Dawley
rats (240-350 g bw). The doses ranged from 8.5-25.6 mg/kg. The
l*C-act1v1ty appeared 1n the blood within 30 minutes of dosing and reached
a maximum after -4 hours.
Tissues were analyzed for "C-HEX at 8, 24, 48. 72, 96 and 120 hours
post-dosing. The kidney and liver had higher residue levels than any other
tissue after oral dosing, although these were generally much lower than
those observed after l.v. dosing. The kidneys and liver were found to
contain 0.96 and 0.7554, respectively, of the administered oral dose while
these organs retained 2.92 and 4.68%, respectively, of the administered l.v.
dose at 24 hours post-dosing. Significantly, a higher proportion (15.07%)
of the J4C-act1vHy was found 1n the digestive system (duodenum, large and
small Intestines) after oral dosing. In addition, the Increased rate and
extent of fecal excretion after oral administration (-72%), as compared with
that after l.v. dosing (-20%), would suggest that only a fraction of the
orally administered dose was absorbed. About 17% of the oral dose was
excreted In the urine.
Both urinary and fecal metabolites were characterized as polar due to
their lack of solubility 1n organic solvents. Unchanged HEX was not
detected In any of the samples examined. Only 11% of the 14C-content was
soluble In organic solvents and a further 32% was rendered organo-soluble
after acid catalyzed hydrolysis Indicating, perhaps, the formation of
metabolic ester-conjugates.
03630 111-6 08/15/88
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Yu and Atallah (1981) also examined the capability of liver, fecal and
gut homogenates to metabolize HEX ^ vitro. In an apparent first-order
process. HEX was metabolized by gut content, fecal and liver homogenates
with half-lives of 10.6, 1.6 and 14.2 hours, respectively. When mercuric
chloride (HgCl.) was added to the gut and fecal homogenates, as a bacte-
rlodde, the half-lives were Increased to 17.2 and 6.2 hours, respectively,
Indicating that there was an Important contribution of the gut and fecal
flora to the metabolism of HEX. Denaturatlon of the liver homogenate had
virtually no effect on the ^ vitro metabolic rate. Indicating that there
was limited Involvement of enzyme-dependent processes.
Lawrence and Dorough (1982), In a comparative study of the uptake,
disposition and elimination of HEX, by the 1.v.. Inhalation and oral routes,
orally dosed Sprague-Dawley rats (175 and 250 g) with either 5 vq or 6
mg/kg of radlolabeled HEX. Doses 1n the mlcrogram range were useful for
monitoring the urinary and fecal excretion of HEX; however, an oral dose of
6 mg/kg, 250 and 600 times the Inhaled and 1.v. doses, respectively, was
necessary to measure tissue residue levels. The authors attributed this to
the poor b1oava1labH1ty of HEX when administered by the oral route.
The total recovery of the radlolabel Immediately following the adminis-
tration of the oral dose was 98.0*5.3% (mean *. standard deviation). Rats
dosed orally voided 2-3 times more of the dose In the feces than those
animals dosed by the l.v. or Inhalation route. Blood levels were measured
<6 hours after dosing and reached a maximum at 2 hours. The low level of
radiocarbon (-1 ng/mi) appearing 1n the blood as compared with animals
dosed l.v. and by Inhalation Indicate that there was poor absorption of HEX
03630 111-7 08/15/88
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from the GI tract. The data were not obtained over a sufficient post-dosing
period to allow any estimate of the elimination rate.
Biliary excretion accounted for 18% of the administered dose after oral
exposure as compared with -13X after l.v. and -8% after Inhalation. This
observation agreed with the report of Yu and Atallah (1981) who adminis-
tered comparable dose levels by the oral route. Lawrence and Dorough (1982)
also reported that the fecal material contained predominantly polar or
unextractable material. This was the case even In animals where bile was
collected, Indicating that degradation of HEX to polar products was taking
place 1n the gut. The authors suggested that the poor absorption of HEX
from the GI tract may be partially related to Us high affinity for binding
with the stomach contents and gut mucosa.
In a more recent comparative study (El Dareer et al., 1983), male
Fischer 344 rats (169 g) were dosed at 4.1 and 61 mg/kg with -1 ma of a
solution of 14C-HEX dissolved 1n a 1:1:4 mixture of Emulphor EL620,
ethanol and water. Little radioactivity (-1X) appeared as exhaled
14C02, 2.4% remained 1n the tissues at 72 hours, <79.5X was excreted In
the feces, and <35.5 was eliminated In the urine.
Inhalation Route
In a report originally produced for the Velslcol Chemical Corporation,
(Dorough, 1980) and later reported by Lawrence and Dorough (1981, 1982). on
the comparative uptake of HEX when administered by various routes, rats were
exposed to 14C-HEX vapor In a specially designed single animal exposure
system. Essentially, a known amount of 14C-HEX was applied as a solution
03630 III-8 08/15/88
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In hexane to the Inside of a 3.7 l generating flask. Air was drawn
through the flask that was slowly rotated to allow the deposition of HEX on
the walls of the flask as the hexane evaporated. The animal was then
exposed to the vapors In a rodent respirator and the exhaust vapors from the
system were passed through a filter pad made from expanded polyurethane
foam. The polyurethane foam was demonstrated to entrap all of the
14C-HEX, which could then be solvent extracted for analysis.
Rats were exposed for a period of 1 hour to the HEX vapors and received
doses that ranged from 1.4-37.4 yg/kg bw (Lawrence and Dorough, 1981).
Immediately following this 1-hour exposure, the recovery of the retained
dose was found to be 91.8+8.5% (mean £ standard deviation). Tbe exposed
animals were Immediately placed In metabolism cages for 72 hours during
which time expired air, feces and urine were collected. The animals were
then killed and specific tissues analyzed for 14C activity. Less than 1%
of the retained radiocarbon was expired during a 24-hour period Immediately
following exposure and no radiocarbon was detected as 14C02- Only -69%
of the Inhaled dose was recovered, which was much lower than that recovered
after 1.v. or oral dosing (85% and 82%, respectively). Since recovery of
the dose Immediately following the administration of the Inhalation dose was
-92%, the reduced recovery during the 72-hour post-dosing period led to the
speculation that a volatile metabolite was formed during this post-exposure
period, but attempts to collect and Identify such a material were not
successful.
Blood concentration-time data during a 1-hour exposure and for >6 hours
postexposure were also presented. It would appear that the levels in the
03630 1II-9 04/12/91
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systemic circulation had barely reached steady state during the exposure
period. Elimination during the subsequent 6 hours appeared to relate to a
complex pharmacoklnetlc model with a terminal rate comparable wHh thai
reported for the l.v. route, with a half-life of -30 hours.
The elimination In the bile appeared to be low, with only -8% of the
administered dose being eliminated by this route, as compared with 13 and
18% after 1.v. or oral administration, respectively (Lawrence and Oorough,
1981, 1982).
The trachea and lungs contained the highest concentrations of 14C-HEX
when the tissues from animals killed at 72 hours after exposure were
analyzed. These concentrations were higher In these tissues than when the
dose was administered by other routes. The kidneys were also a site for
residue accumulation. Lawrence and Dorough (1981, 1982) concluded that.
Irrespective of the route of administration, the lung appeared to be the
primary target organ for toxIcHy.
The fraction of the dose recovered In the feces and urine (23 and 33%,
respectively) was about the same as that recovered after the l.v. dose,
except that more was recovered In the urine than from the feces after the
Inhalation exposure, while the reverse was observed after the 1.v. dose.
In a comparative study of the uptake, disposition and elimination of
i-C-HEX, El Oareer et al. (1983) placed Fischer 344 rats (125-190 g) In
Delmar-Roth type metabolism cages; 1.1 mg of 14C-HEX (15 pCI) In 0.05
ma of elhanol was then placed In glass U-tubes situated 1n the Incoming
03630 111-10 04/12/91
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air line of the metabolism cages. After 2 hours, >90% of the radioactivity
had volatilized and the dose received by each rat was calculated from the
total amounts of radioactivity recovered from the tissues, feces, urine and
exhaled air. The fur of the animals was not Included. The dose received by
the exposed animals was between 1.3 and 1.8 mg/kg bw.
The animals were killed at either 6 or 24 hours after removal from the
Inhalation exposure. Whole blood, plasma, liver, kidney, voluntary muscle
(gastrocnemlus). subcutaneous fat, brain, skin (ears) and the residual
carcass (except for the skin and fur) were analyzed for 14C activity. The
principal sites of deposition were the lungs, kidney and liver. Only -1J4 of
the radlolabel was Identified as 14CO?. No Intact HEX was found 1n any
of the tissues. The majority of the radlolabel extracted from the tissues
was polar (water-soluble).
In a separate experiment, El Dareer et al. (1983) Incubated 1*C-HEX
with homogenates of liver, feces and Intestinal (large and small) contents
as well as with whole blood and plasma. These _1n vitro studies were
designed to assess the binding of HEX to substrates present 1n the homogen-
ates. The mixtures were Incubated at 37°C with gentle shaking. Samples
were taken at 0, 5 and 60 minutes and extracted twice, first with chloroform
and then, after acidification with 1 ml of IN HC1, with chloroform:metha-
nol (2:1 v/v). The radiocarbon content of each of the samples was deter-
mined and selected samples were also analyzed for HEX, per se, by an HPLC
method. The results, presented 1n Table III-l, demonstrated the chemical
reactivity of HEX and Its ability to bind components of biological material.
03630 III-ll 08/15/88
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o
u>
TABLE III-l
o Extractablllty of [14C] HEX and Radioactivity Derived from Saline and Various Biological Preparations3
Percent
Total Radioactivity In Fraction
First Extraction
Preparation Time
(minutes)
Saline 0
5
60
- Liver 0
« 5
h 60
Plasma 0
5
60
Whole Blood 0
5
60
Feces 0
5
60
_ Intestinal contents 0
§ 5
£ *°
m _^ — ^^ _ _
Organlcb
99.6 (92.4)
99.1 (92.8)
98.8 (94.6)
55.0 (74.4)
42.8 (49.7)
11.1
22.2 (61.7)
19.7 (66.3)
1.4
16.2 (60.4)
2.8
0.6
90.0 (93.7)
83.4 (87.8)
40.5 (61.0)
93.7 (94.7)
82.8 (89.5)
66.3 (87.0)
Aqueous
0.4
0.9
1.2
8.0
15.2
18.8
7.2
25.0
43.4
3.8
21.6
27.4
0.6
0.8
2.8
0.6
1.6
4.6
Second Extraction
Organic
24.5
15.0
5.9
50.2
33.6
21.
27.9
13.4
12.0
8.0
9.0
31.3
4.6
8.6
15.4
Aqueous
1.0
4.7
2.4
0.8
2.0
3.9
1.2
1.6
1.4
0.2
0.6
3.0
0.2
1.0
2.4
Pellet
11.6
22.2
51.8
19.6
19.6
30.2
50.8
60.6
58.6
1.2
6.2
22.4
1.0
5.9
11.3
aSource: El Dareer et al.. 1983
mbers In parentheses represent the percent of the
Inactivity In the fraction as HEX.
-------
Percutaneous Route
There were no studies on the pharmacoklnetlcs or disposition of HEX
found An a survey of the published literature or 1n the technical research
documents provided to the U.S. EPA by Industry under the Toxic Substances
Control Act reporting requirements. While no quantitative studies or
estimates of the uptake of HEX through skin were found, studies have been
reported In which discoloration, edema and necrosis of the skin was observed
following the dermal application of HEX (Treon et a!., 1955; IRDC. 1972).
In these reports, a toxic response, leading to death, was observed In
several Instances, which would suggest that HEX was absorbed transdermally
Into the systemic circulation.
Comparative Studies
It was evident, from the available published literature, that the four
principal studies on the uptake and disposition of HEX each Involved more
than one route of uptake and that the comparison of the exposure route was
at least one objective of the study. The principal comparative observations
were as follows:
1. Irrespective of the route of application, the lung was the
target organ for toxlclty.
2. The principal routes of elimination were the urine and feces.
Considerably more of the administered dose was excreted 1n the
feces after oral administration than after dosing by the 1.v.
or Inhalation route. More of the administered dose was
excreted In the urine than the feces after Inhalation exposure
while the reverse was the case after l.v. administration.
3. Biliary excretion occurred after administration by all three
routes. For similar doses, elimination was In the following
order: oral > l.v. > Inhalation.
03630 111-13 08/15/88
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4. Results of distribution studies are presented 1n Tables III-2,
I1I-3 and III-4. The highest HEX accumulation was In the
kidneys, lung and liver following oral and l.v. exposure. Ihe
lung and trachea contained the highest concentration of HEX
after Inhalation exposure.
Summary
A number of studies are available regarding the absorption, distribution
and excretion of HEX and Us metabolites following oral. Inhalation and l.v.
exposure. Comparative pharmacokineUc studies of laC-HEX have shown
higher levels of fecal excretion following oral exposure than l.v. or
Inhalation exposure (El Career et al., 1983; Lawrence and Dorough, 1982).
Increased elimination of radiocarbon following oral exposure is consistent
with toxlcity data that Indicate that HEX Is more toxic following Inhalation
than oral exposure. Following Inhalation exposure to iaC-HEX,
considerable amounts of the radlolabel remain In the lung and trachea,
Indicating that HEX reacts with biological material In the lung (Dorough,
1980).
The low level of 1«C02 or 14C-HEX (<1%) exhaled from the lungs
following Inhalation or oral exposure Indicates that the respiratory tract
does not play a major role In the excretion of HEX (Dorough, 1979). Urinary
excretion predominates following Inhalation exposure.
Following oral exposure, the highest levels of HEX are found In the
kidney and liver. Poor absorption of HEX from the GI tract has been
attributed to the low bloavallabllHy of HEX In the gut and Its rapid
metabolism by Intestinal flora following oral exposure (El Dareer et al..
1983). Dietary exposure to HEX for 30 days showed a good correlation
03630 IH-14 04/12/91
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o
CJ
to
o
TABLE II1-2
Disposition of Radioactivity Expressed as Percentage
of Administered Dose from "C-HEX In Rats Dosed by Various foutesa
Feces
Urine
^ Tissues
en
Other volatile
TOTAL RECOVERY
Oral Dose
Lou Doseb High Doseb
(4.1 mg/kg) (61 mg/kg)
79.4 * 2.9 65.3 * 6.9
35.5 * 2.5 28.7 * 4.2
2.4 * 0.6 2.4 ± 0.1
0.8 f 0.0 0.6 f 0.0
0.2 i 0.0 0.3 * 0.0
118.3 i 3.0e 97.3 i 7.0
aSource: Adapted from El Oareer et al., 1983 (The
tlon for three rats.)
Intravenous Doseb Inhalation Dose
Group Ac Group Bb
0.59 mg/kg (1.3 mg/kg) (1.8 mg/kg)
34.Oil.Qd 28.7^4.3 47.5*6.4
15.8 i 1.4 41.0 i 4.8 40.0 » 6.6
39.0 * 1.0 28.9 * 1.6 11.5 * 0.8
0.1 * 0.0 1.4 * 0.3 1.0 * 0.5
0.1 * 0.0
89.0 * 2.0 (100) (100)
values represent the mean X of dose i standard devta-
bAt 72 hours after dosing or exposure
cAt 6 hours after
dplus Intestinal
o
OO _fc
exposure
contents
j cent recoveries for this dose are "normalized" to 100X. differences In disposition for the two doses are
minimal, an Indication that no saturable process is operative In this ilosc range.
-------
TABLE I1I-3
Fate of Radiocarbon Following Oral, Inhalation and
Intravenous Exposure to 14C-HEX 1n Rats
Expressed as Percentage of Administered Oosea
Cumulative Percent of Dose
Oralb Intravenous0 Inhalat1ond
Urine
Feces
Urine
Feces
Urine
Feces
Body
Total Recovery
22.2 + 1.8
62.2 ± 8.0
24.0 * 1.9
67.7 * 5.1
24.4 + 1.9
68.2 * 5.1
0.2 + 0.2
92.8 + 4.7
24-Hour
18.3
21.1
48-Hour
20.7
30.4
72-Hour
22.1
47.4
15.7
85.2
+ 5.2
± 7.1
+ 5.6
± I-?
* 5.7
* 1.9
* 7.8
+ 4.8
29.7
17.0
32.5
21.0
33.1
23.1
12.9
69.1
* 4.5
± 7.5
+ 5.1
± 7-5
+ 4.5
+ 5.7
+ 4.7
* 9.6
aSource: Adapted from Dorough. 1980, and Lawrence and Dorough, 1982
bDoses administered In 0.5 ml corn oil at 7 yg/kg bw
cDoses administered 1n 0.2 ml 10:4:1 sallne-.propylene glycol-.ethanol by
Injection Into the femoral vein at 5 vg/kg bw.
dDoses administered as vapors over a 1-hour exposure period to achieve
doses of -24 yg/kg bw.
03630 111-16 08/15/88
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TABLE III-4
Distribution of HEX Equivalents In Tissues and Excreta of Rats
72 Hours After Oral, Inhalation and Intravenous Exposure to 14C-HEXa'D-c
Sample
Oral Oose
(6 mg/kg)d
Inhaled Dose
(-24
Intravenous Dose
(10 ug/kg)
Trachea
Lungs
Liver
Kidneys
Fat
Remaining carcass
ng/q of Tissue
292 + 170
420 7 250
539 7 72
3272 7 84
311 7 12
63 +• 40
107.0 * 65.0
71.5 * 55.2
3.6 + 1.9
29.5 * 20.2
2.8 +• 0.4
1.3 «• 0.6
3.3 * 1.7
14.9 * 1.1
9.6 * 1.1
22.3 + 0.6
2.3 +• 0.2
0.5 «• 0.1
Percent of Dose
Whole
Urine
Feces
Total
Body
Recovery
2
15
63
81
.8
.3
.6
.7
+ 1
7 3
I 8
7 6
.1
.3
.5
.7
12
33
23
69
.9 *
.1 7
.1 *
.1 *
4.7
4.5
5.7
9.6
31
22
31
84
.0 «•
.1 7
.4 *
.6 «•
7.8
5.7
1.9
4.6
^Source: Adapted from Dorough, 1980 and Lawrence and Oorough, 1982
bOne HEX equivalent Is defined as the amount of radlolabel equivalent to
one nanogram of HEX based on the specific activity of the dosing solution.
CA11 values are -the Mean ± S.D. of three replicates.
dNote that the oral dose was 250 and 600 times that of the Inhaled and
1.v. doses, respectively. That was necessary since residues were not
detected In Individual tissues of animals treated orally at doses of 5-25
03630
111-17
08/15/88
-------
between 14C levels found 1n all tissues and levels of HEX In the diet;
steady-state levels were reached after 15 days of exposure.
There 1s very limited data available regarding the pharmacoklnetlcs of
dermal exposure to HEX. Harked discoloration of the skin has resulted
following dermal application of HEX.
Metabolites of HEX have not been characterized. At least four polar
metabolltles were separated from I issues and excreta regardless of the route-
of admlnstratlon. Fractions of the polar metabolites could be rendered
organo-soluble by treatment with an aqueous strong acid, which suggested
that these are conjugated metabolites. Very IHtle of the radlolabel In
tissues or excreta was extractable with organic solvents and no unchanged
HEX was detected 1n any of these fractions.
The available studies not are adequate for assessing the nature and
mechanisms Involved In the uptake, metabolism, distribution and excretion of
HEX by the l.v., Inhalation and oral routes. Insufficient data are
available to make any quantitative estimate of dermal uptake. The precise
characterization and chemical nature of the metabolites of HEX are not
available.
03630 111-18 06/19/90
-------
IV. HUMAN EXPOSURE
Text to be provided by the Office of Drinking Water
03640 IV-1 10/16/85
-------
V. HEALTH EFFECTS IN ANIHALS
Overview
Hexachlorocyclopentadlene toxlclty has been reported In many shorl-term
studies, several of which have been reviewed In other documents. Including
NAS (1978) and U.S. EPA (1980, 1984). Although different 1n scope and
emphasis (earlier U.S. EPA documents addressed ambient water and air
toxlclty. respectively) a large amount of the scientific knowledge
pertaining to HEX 1s Included In these documents. To avoid unnecessary
duplication, studies previously discussed 1n the above dcc-jrscr.ts will not be
reviewed In great detail In this section, except when they are used In the
presentation of critical health effects of HEX.
Acute Toxlclty
An approximate oral lethal dose (ALD) for female rabbits was determined
to be 420-620 mg/kg. The oral ALDs for male and female rats were <280 and
>280 mg/kg, respectively. An oral LD5Q of 505 mg/kg for male rats was
also calculated (Treon et al., 1955). IRDC (1968) determined the oral
L05Q for male albino rats to be 926 mg/kg for HEX given In corn oil. In a
later study, IRDC (1972) reported oral LD^s of 630, 530 and 584 mg/kg for
male, females, and male and female rats combined, respectively. In a more
recent study, Dorough (1979) reported an approximate lethal dose of 65 mg/kg
for male and female Sprague-Dawley rats and approximate lethal dose of 180
and 600 mg/kg for Sprague-Dawley male and female mice, respectively.
The acute toxlclty studies of HEX are summarized 1n Table V-l. Treon et
al. (1955) conducted a series of oral toxlclty studies using female rabbits
(strain unspecified) and Carworth rats of both sexes. HEX was administered
03650
v_-| 04/12/91
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o
OJ
o»
171
o
TABLE V-l
Acute Toxlclty of HEX*
Study
Oral LDjQ
Oral LDso
Oral LDcn
Species/Age
rat,
rat,
Rat.
young adult
adult
young adult
l»50:
LD50:
L050:
males
males
males
Results
- 505
- 926
- 630
mg/kg
mg/kg
mg/kg
Reference
Treon
I ROC.
IRDC.
et al..
1968
1972
1955
Oral 1050
Oral 1050
Inhalation 1059
Inhalation LCso
Inhalation LCjo
Primary eye Irritation
rat. young adult
mouse, young adult
rat. young adult
rat. young adult
guinea pig.
young adult
rabbit, adult
females - 530 mg/kg
males and females -
584 mg/kg
LDso: males and females -
300-600 mg/kg
LDso: males and females -
600-1200 mg/kg
3.5-hour LCso: males and
females - 3.1 ppm
4-hour LC5Q,: males - 1.6 ppm
females - 3.5 ppm
3.5-hour LCso: males and
females - 7.1 ppm
Severe eye Irritant (0.1 ml lor
5 minutes or 24 hours); all dead by
day 9 of study
SRI. 1980a
SRI. 1980a
Treon et al.. 1955
Rand et al.. 1982a
Treon et al.. 1955
IRDC. 1972
CD
CO
-------
o
u>
o»
en
TABLE V-l (cont.)
Study
Species/Age
Results
Reference
Primary dermal Irritation rabbit, adult
Primary dermal Irritation rabbit, adult
Primary dermal Irritation monkey, adult
Moderate skin Irritant (250 mg/kg)
One application
Severe skin Irritant (200 mg/kg)
All males died In study
Skin discoloration and necrosis
(0.05 mt of 10X HEX solution)
Treon et al.. 1955
IRDC, 1972
Treon et al.. 1955
'Source: U.S. EPA.
oo
CO
-------
as a 5% solution In peanut oil by gavage. In addition, Southern Research
Institute (SRI. 1980a) reported the oral LD5Q for male and female weanling
B6C3F1 mice to be 600-1200 mg/kg and the oral LD,Q for weanling Fischer
344 male and female rats to be between 300 and 600 mg/kg.
Treon et al. (1955) reported an approximate dermal lethal dose for
rabbits between 430 and 610 mg/kg while IROC (1972) reported that 1n male
New Zealand white rabbits, an approximate lethal dose for dermal exposure
was <200 mg/kg.
Treon et al. (1955) reported a 3.5-hour LC5_ of 3.1 ppm for Carworth
rats of both sexes, and 2.1 and 7.1 ppm In male and female guinea pigs.
respectively. Rand et al. (1982a) reported a 4-hour IC™ of 1.6 ppm for
male Sprague-Dawley rats and 3.5 ppm for female rats.
Treon et al. (1955) reported HEX to be a primary skin Irritant 1n
rabbits (strain unspecified) at a dose level of 250 mg/kg. Monkeys (strain
unspecified) were also tested by Treon et al. (1955). Discoloration,
necrosis and edema of the skin were noted when 0.05 mi of a 10% HEX
solution was applied for 3 consecutive days. Application of 0.01 ma of
0.1-10% solutions of HEX resulted In no skin Irritation. IRDC (1972)
reported HEX to be a dermal Irritant 1n New Zealand white rabbits based upon
edema observed following the application of 200 mg/kg HEX. In this study.
Intense discoloration of the skin was also noted.
In a Russian study. Nalshteln and Llsovskaya (1965) studied the effects
of HEX applied to the shaved area of the skin of rabbits (strain unspeci-
fied) dally for 10 days. According to the authors, no effects were noted In
03650 V-4 08/25/88
-------
control and test animals given dally doses of 0.5-0.6 ml of a 20 mg/a.
solution of HEX.
The acute oral toxlclty of HEX was tested 1n male CD-I mice (6/group)
following a single gavage dose of 0. O.OS, 0.1, 0.5, 1.0 or 5.0 g/kg HEX In
DHSO (Litton Blonetlcs, 1978b). Mortality was observed 1n 6/6 mice exposed
to both the 1.0 and 5.0 g/kg dose. At doses of 0.05, 0.1 and 0.5 g/kg. 1/6,
1/6 and 3/6 deaths were observed, respectively. In a follow-up study,
exposures of 7.6, 25.3 or 76 ing/kg HEX for 5 consecutive days resulted In
6/10, 9/10 and 10/10 deaths, respectively.
Fischer 344 rats and B6C3F1 mice (5/sex/group) were administered a
single gavage dose of 0, 75, 150. 300, 600 or 1200 mg/kg HEX and observed
for 14 days (SRI. 1980a). Treatment-related mortality was seen In 5/5 male
and 5/5 female rats at >600 mg/kg and In 2/5 females at the 300 mg/kg dose
level. No other treatment-related deaths were recorded. Animals exposed to
>300 mg/kg HEX also experienced decreased activity, ruffled fur and
diarrhea; the severity and duration was directly related to the dose level.
Animals In the 75 and 150 mg/kg dose groups experienced none of the effects
observed at the higher exposure levels with the exception of "wet fur In the
anal area", which the authors suggested may be an Indication of mild Gl
disturbances.
In the mice, all animals exposed at 1200 mg/kg died, while only one
female and one male died at the 600 mg/kg level. At the other exposure
levels (<300 mg/kg) only minor effects were observed.
03650 V-5 08/25/88
-------
In a follow-up range finding study (SRI, 1980b), rats and mice were
administered HEX In corn oil at 25. 50, 100, 200 and 400 mg/kg for rats and
50, 100, 200, 400 and 800 mg/kg for mice on a 12-day schedule (days 1-5,
8-12, 15 and 16). Five mice and five rats of each sex were dosed at each
level, with at least two consecutive dosing days before sacrifice. Body
weights were taken on days 0, 7 and 17. All animals were observed twice
dally for signs of toxlclty and clinical symptoms.
In the rat experiment (SRI, 1980b). all males and 4/5 females at the 400
mg/kg HEX dose level died, while at the 200 mg/kg dose, 1/5 males and 4/5
females died. For the remaining males, there was a dose-related decrease 1n
body weight gain of 8, 17, 43 and 135% for the 25, 50, 100 and 200 mg/kg
dose groups, respectively. Significant gross changes In the stomach Includ-
ing ulceratlon and thickening of the stomach wall were observed 1n all the
surviving animals at doses >100 mg/kg and 1n 5/5 males and 2/5 females at
the 50 mg/kg dose level.
In the mouse experiments (SRI, 1980b), all of the treated animals at the
400 and 800 mg/kg dose level died. No treatment-related mortality was seen
at <200 mg/kg and only minor effects were observed at <100 mg/kg HEX. The
weight gain depression seen 1n the rat study was not evidenced In the mouse
study.
Rand et al. (1982a) conducted a range-finding study 1n which groups of
10 male and 10 female Sprague-Dawley rats were exposed to atmospheres 0.022.
0.11 or 0.5 ppm HEX. 6 hours/day, 5 days/week for a total of 10 exposures.
Nine male rats and one female rat exposed to 0.5 ppm HEX were dead or morl-
03650 V-6 04/12/91
-------
bund after 5-7 exposures. Prior to death, these rats had dark red eyes,
labored breathing, and paleness of extremities. No deaths were noted \r\ the
other exposure groups; however, the males In the 0.022 and 0.11 ppm groups
experienced a significant (p<0.05) concentration-related reduction In mean
body weight. Mean lung weights were significantly Increased 1n males and
females exposed to 0.5 ppm. Significant reduction In kidney, adrenal and
ovary weights were also observed In the 0.5 ppm group. In males, liver
weight was reduced 8-9% of control at >0.022 ppm dose levels and 1n females
in the 0.11 and 0.5 ppm groups.
SubchronU and Chronic Toxldty
Subchronlc toxlclty studies of HEX are summarized In Table V-2. Oral
toxlclly studies In B6C3F1 mice and Fischer 344 rats have been conducted by
SRI (1981a,b; Abdo. 1984) under contract with NIP. In the mouse study (SRI,
1981a). dose levels of 0. 19. 38, 75. 150 and 300 mg/kg HEX (94.3-97.4%) In
corn oil were administered by gavage to 10 mice of each sex. 5 days/week for
13 weeks (91 days). At the highest dose level (300 mg/kg), all male mice
died by day 8 and three females died by day 14. In female mice, there was
significant dose-related enlargement of the liver. Toxic nephrosis In
females at doses of >75 mg/kg was characterized by lesions In the terminal
portions of the convoluted tubules, with basophllla In the Inner cortical
zone and cytoplasmlc vacuollzatlon. Nephrosis was not observed In male mice
at any dose level. Levels of >38 mg/kg HEX caused lesions In the fore-
stomach, Including Inflammation and hyperplasla In both males and females
and a decrease In relative body weight gain. These lesions Increased in
severity and Incidence with Increased dose. At the 19 mg/kg dose level, no
significant hlstologlc changes were seen In any of the organs examined.
03650 V-7 04/12/91
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o
CO
CO
TABLE ¥-2
Subchronlc loxtclty of HEX*
Study Species
90-Day feeding study rat
90-Day feeding study mouse
14 -Week Inhalation rat
toxlclty study
14 -Meek Inhalation monkey
toxlclty study
Dose
10. 19. 3B. TS. 150
mg/kg (by gavage)
19. 38. 75, 150 or
300 mg/kg (by gawage)
0.01. O.OS and 0.? ppn
IS days/week)
0.01. 0.05 and 0.2 ppn
(5 days/week)
Results
NOAEL
LOAEL
NOAEL
LOAEL
NOEL
LOEL
NOEL
LOEL
- 10 mg/kg
- 19 mg/kg
- 19 ng/kg
- 38 wj/kg
- 0.2 ppcn
- NE
- 0.2 ppm
- NE
Effects at LOU or Reference
Lowest Dose
Lesions of forestonuch In SRI. 1981b
female rats at 19 mg/kg
Lesions of forestonuch In SRI. 1981a
both sexes at 38 mg/kg
No statistically slgnHI- Rind et al.. 1982A
cant effects
No effects noted Alexander et al.. 1980
•Source: U.S. EPA. 1984
NE • Not established
CO
co
CO
-------
Slight Increases In organ weights were noted. A dose level of 38 mg/kg was
considered a LOAEL 1n mice based on the Increasesd Incidence and severity of
nephrosls In females and an Incidence of hyperplasla and Inflammation in
males and females. The 19 mg/kg dose level was considered a NOAEL for mice.
In the rat study (SRI. 1981b; Abdo et al., 1984), dose levels of 10, 19,
38, 75 and ISO mg/kg HEX 1n corn oil were administered by gavage to groups
of 10 male and female F344 rats, 5 days/week for 13 weeks. At doses of >38
mg/kg HEX, epithelial hyperplasla and Inflammation of the forestomach,
Increased mortality and toxic nephrosls were observed In males and females.
A significant (p<0.05) dose-related depression 1n body weight gain was
observed In males at levels of >38 mg/kg and females at >75 mg/kg.
Liver-to-brain weight ratios were significantly (p<0.05) Increased 1n
females at >38 mg/kg and k1dney-to-bra1n weight ratios at >75 mg/kg. At
doses >19 mg/kg, epithelial hyperplasla and focal Inflammation of the fore-
stomach, and toxic nephrosls were noted In females. Decreases In body
weight gain relative to the controls were observed 1n males at all dose
levels; however, these were not significant at the 10 or 19 mg/kg dose
levels. In females, dose-related decreases In body weight gain were
observed at doses >38 mg/kg/day. No significant adverse effects were
observed In rats at the 10 mg/kg dose level. A summary of these results are
presented In Table V-3.
Fourteen-week Inhalation studies 1n rats and monkeys have been performed
(Rand et al., 1982a,b; Alexander et al., 1980). Groups of 40 male and 40
female Sprague-Dawley rats, weighing 160-224 g or groups of 12 cynomolgus
monkeys (6/sex), weighing 1.5-2.5 kg, were exposed to HEX. 6 hours/day. 5
days/week, for as long as 14 weeks. Levels of exposure were 0, 0.01, 0.05
03650 V-9 04/12/91
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o
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in
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TABLE V-3
Toxlcologlcal Parameters for Mice and Rats Administered
Technical Grade HEX In Corn Oil for 91 Daysa
o
CO
Pathology
Forestomach
Species/
Strain
Hale mice/
B6C3Fi
Female mice/
B6C3F]
Male rats/
Fischer 344
Dose
(mg/kg)
0
19
38
75
150
300
0
19
3B
75
150
300
0
10
19
38
75
150
Mortality
1/10
0/10
0/10
0/10
0/10
10/10
0/10
0/10
0/10
0/10
0/10
3/10
3/10
1/10
1/10
1/10
3/10
7/10
Relative
Weight
Galnb
f36X
t 9X
- 9X
-45X
--
f!3X
-13X
-13X
-25X
-38X
„ _
- 4X
- 8X
-20X
-49X
-57X
Inflammation
0/10
0/10
2/10
7/10
7/10
7/10
0/10
0/10
2/9
6/10
10/10
7/9
0/10
0/10
0/10
4/10
9/10
8/9
Hyperplasta
0/10
0/10
2/10
8/10
9/10
8/10
0/10
0/10
2/9
9/10
10/10
9/9
0/10
0/10
0/10
5/10
9/10
8/9
Kidney
Toxic
Nephrosls
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/9
10/10
10/10
7/10
0/10
0/10
4/10
10/10
9/10
8/10
00
CO
-------
0 TABLE V-3 (cont.)
en
Pathology
Forestomach
Species/
Strain
Female rats/
Fischer 344
Dose
(mg/kg)
0
10
19
38
75
150
Mortality
1/10
2/10
1/10
1/10
3/10
5/10
Relative
Weight
Gain6
OX
«• 4X
- 5X
- 2%
-30X
-33X
Inflammation
0/10
0/10
2/10
2/10
9/10
9/10
Hyperplasla
0/10
0/10
2/10
5/10
9/10
9/10
Kidney
Toxic
Nephrosls
0/10
0/10
0/10
10/10
10/10
10/10
"Source: SRI. 1981a,b
^Relative weight gain Is calculated as:
ou
Control Group Value
Dose Group Value - Control Group Value
o
CD
oo
00
-------
and 0.20 ppm HEX. In monkeys, there were no deaths, or adverse clinical
signs, or significant changes In weight gain, pulmonary function, eye
morphology, hematologlc parameters, clinical chemistry or hlstopathology at
any dose level tested.
Hale rats had a transient appearance of dark-red eyes at 0.05 and 0.2
ppm" HEX. At 12 weeks, there were marginal but not statistically significant
Increases 1n hemoglobin concentration and erythrocyte count 1n 0.01 ppm
males, 0.05 ppm females, and 0.20 ppm males and females. There were small
but nonsignificant changes 1n mean liver weight of all treatment groups and
similar changes In the kidneys of all treated males. There were no
treatment-related abnormalities 1n gross pathology or hlstopathology. On
this basis, the NOAEL 1n rats was 0.2 ppm HEX.
In another study by Rand et al. (1982b), no treatment-related ultra-
structural changes were observed In monkeys exposed to HEX vapors. Exposure
was Identical to that described In the previous study (Rand et al., 198?a).
This study took an 1n-depth look at the Clara cells of the lung; the results
showed a statistically significant (p<0.01) Increase In the mean number of
electron-lucent Inclusions In the apex and base of the Clara cells In the
exposed animals as compared with the controls. According to some
researchers (Evans et al., 1978), Clara cells respond to Injury by regres-
sion to a more primitive cell type. Rand et al. (1982b) noted that some of
the ultrastructural changes In the exposed animals resembled those of the
Evans study. The biologic significance of these results are not known;
however, Clara cells play an Important part In producing the extracellular
lining of the peripheral airways. Alteration of this lining may result In
subsequent Impaired breathing (Rand et al.. 1982b).
03650 V-12 04/12/91
-------
Treon et al. (1955) exposed two guinea pigs, six rabbits, four rats and
five mice to a concentration of 0.34 ppm HEX for 7 hours/day, 5 days/week
for 25-30 exposures. Guinea pigs survived the 30 exposures, while only two
rabbits survived and no rats or mice survived 5 exposures. Degenerative
changes were observed In the brain, heart, liver, adrenal glands and
kidneys. Severe pulmonary edema, hyperemla and acute necrotlzlng bronchitis
was also observed. Using a lower concentration (0.15 ppm HEX), 2/2 guinea
pigs, 3/3 rabbits and 4/4 rats survived IbO seven-hour exposure periods. In
mice, only 1/5 animals survived. Slight renal and hepatic degeneration was
noted In all species and rats, mice and guinea pigs also developed lung
lesions.
In an NTP-sponsored study (Battelle Northwest Laboratories, 1984; Abdo
et al., 1986), F344 rats and B6C3F1 mice (10/group/sex) were exposed to HEX
(99.4254) at 0, 0.04, 0.15, 0.4. 1 or 2 ppm (0, 0.45, 1.67, 4.46, 11.1 or
22.3 mg/m3) 6 hours/day, 5 days/week for 13 weeks. All rats and mice
exposed to >1 ppm died; 5/10 male and 2/10 female mice at the 0.4 ppm also
died. In male rats, there were statistically significant (p<0.05)
reductions 1n body weight and Increases In relative lung weight at 0.4 ppm.
Similar but less severe effects were seen In females. Dose-related
hlstopathologlc changes were observed In the respiratory tract epithelium of
males and females at >0.4 ppm. Changes Included necrosis and acute
Inflammation. No statistically significant adverse effects were observed In
rats at <0.15 ppm.
In mice (Abdo et al., 1986), exposure to >0.4 ppm HEX resulted in a 10%
reduction 1n body weight gain In males and females. Dose-related
03650 V-13 04/12/91
-------
hlstopathologlc alterations 1n the respiratory epithelium, including
hyperplasla and metaplasia, were observed 1n all treated mice at levels
>0.15 ppm. No adverse effects were seen In mice exposed at 0.04 ppm.
A chronic oral toxlclty study of HEX being conducted by SRI for the NIP
was terminated In April 1982 because Inhalation was determined to be the
more relevant route of exposure. No other chronic oral toxldty data were
available for this report. There were no chronic dermal toxlclty studies
found In the available literature.
A 30-week Inhalation study 1n rats of technical grade HEX, 9654 pure with
hexachlorobuta-1,3-dlene and octachlorocyclopentene as Impurities, was
conducted by Shell Toxicology Laboratory (D. Clark et al., 1982). Four
groups of 8 male and 8 female Wlstar albino rats were exposed to HEX at
nominal concentrations of 0, 0.05. 0.1 and 0.5 ppm for 6 hours/day, 5 days/
week, for 30 weeks and were observed for a 14-week recovery period without
HEX exposure. At the highest dose level 4 males and 2 females died. In
males, there was a depressed body weight gain In the 0.5 ppm group relative
to controls beginning at 7 weeks of exposure and persisting throughout the
remainder of the study. In females, body weights were significantly
Increased during the first half of the exposure period but were signifi-
cantly (p<0.01) decreased at the end of the recovery period at 0.5 and 0.1
ppm. At 0.5 ppm, there were pulmonary degenerative changes noted In both
sexes although the males were affected more severely. At the 0.5 ppm dose,
there were mild degenerative changes In the liver and kidneys at 30 weeks In
a few rats and kidney weights were significantly Increased In the females.
After 30 weeks of study, there was no biologically significant toxldty
noted In animals exposed to concentrations of 0.05 or 0.1 ppm HEX.
03650 V-14 04/12/91
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HutagenUUy
Goggelman et al. (1978) found that HEX was not mutagenlc wUh or without
liver mlcrosomal activation at ?.7xlO"3 H 1n an Escherkhla con K12. back
mutation system. In this test there was 70% survival of bacteria at 72
hours. HEX was not tested at higher concentrations because It was cytotoxlc
to E.. coll. A previous report from the same laboratory (Grelm et al.. 1977)
Indicated that HEX was also not mutagenlc 1n Salmonella typhlmurlum strains
TA1535 (base-pair mutant) or TA1538 (frame shift mutant) after liver mlcro-
somal activation; however, no details of the concentrations tested «cie
given. Although letrachlorocyclopentadlene Is mutagenlc In these systems,
probably through metabolic conversion to the dlenone, It appears that the
chlorine atoms at the C-l position of HEX hindered metabolic oxidation to
the corresponding acylatlng dlenone (Grelm et al., 1977).
A study conducted by Industrial B1o-Test Laboratories (IBT, 1977) also
suggests that HEX Is not mutagenlc 1n S. typhlmurlum. Both HEX and 11s
vapors were tested with and without metabolic activation. The vapor test
was done In desiccators with only the TA100 strain of S. typhlmurlum. It Is
not clear from the data whether sufficient amounts of HEX or adequate times
of exposure were used. Exposure limes of 30, 60 or 120 minutes were studied.
At concentrations of <1.25xlO"3 ug/ml In the presence of an S-9
liver activating system, HEX was not mutagenlc In the mouse lymphoma muta-
tion assay. MutagenUHy could not be evaluated at higher concentrations
because of the cytotoxlclty of HEX (LHton Blonetlcs, Inc.. 1978a).
03650 V-15 04/12/91
-------
Williams (1978) found that HEX (10"* M) was Inactive in the liver
epithelial culture hypoxanthlne-guanlne-phosphoMbosyl transferase (HGPR1)
locus/mutation assay. At 10"5 M H also failed to stimulate DNA repair
synthesis In hepatocyte primary cultures. Negative results were also
obtained In an additional unscheduled DNA synthesis assay (Brat. 1983).
Two recent studies provided by NTP (Haworth et al., 1983) also did not
demonstrate the mutagenlclty of HEX. In S. typhlmurlum strains 1A98, TA100,
7A1535 and TA1537. levels of <3.3 yg/plate were not mutagenlc without
activation and levels of <100.0 yg/plate were not mutagenlc after micro-
somal activation. Higher levels could not be tested because of excessive
killing of the bacteria. In the DrosophUa sex-linked recessive lethal
test, HEX was not mutagenlc. The doses used 1n this study were 40 ppm by
feeding for 3 days or a single Injection of 2000 ppm.
HEX has also been assayed In the mouse dominant lethal test (LHton
Blonetlcs, Inc., 1978b). In this assay, 0.1, 0.3 or 1.0 mg/kg HEX was
administered by gavage to 10 male CD-I mice for 5 days and these mice were
then mated throughout spermatogenesls (7 weeks). This test determines
whether the compound Induces lethal genetic damage to the germ cells of
males. There was no evidence of dominant lethal activity at any dose level
by any parameter (e.g., fertility Index, Implantations/pregnancy, average
resorptlons/pregnancy).
CardnogenlcUy
An NTP bloassay of HEX for cardnogenlclty by the Inhalation route In
rats and mice Is In progress (NTP, 1991). The ability of HEX to Induce
03650 V-16 04/12/91
-------
morphologic transformation of BALB/3T3 cells \t\ vitro has been studied by
LHton BloneUcs, Inc. (1977). Evaluation of the potential for carcinogenic
activity was based on the following criteria:
The endpolnt of carcinogenic activity Is determined by the presence
of f1broblastlc-l1ke colonies, which are altered morphologically In
comparison to the cells observed In normal cultures. These (trans-
formed) cells grow In criss-cross, randomly oriented fashion with
overlapping at the periphery of the colony. The colony exhibits
dense piling up of cells. On staining the foci are deeply stained
and the cells are basophlllc In character and variable In size.
These changes are not observed 1n normal cultures, which stain
uniformly.
Assays were performed at levels of 0.0, l.OxlO'5, ?.OxlO's.
3.9xlO~5, 7.8xlO"s and 1.56xlO"4 pi/ml. The cultures were exposed
for 48 hours followed by an Incubation period of 3-4 weeks. The doses
selected allowed an 80-100% survival of cells as compared with solvent
negative controls. 3-Methylcholanthrene at a dose level of 5 pg/mi was
used as a positive control. Results Indicated that HEX was not responsible
for any significant malignant transformation.
Teratogenlclty
The teratogenlc potential of HEX was evaluated In pregnant Charles River
CD-I rats administered HEX (98.25%) In corn oil. by gastlc Intubation, at
dose levels of 3, 10 and 30 mg/kg/day from days 6 through 15 of gestation.
A control group received the vehicle (corn oil) at a dose volume of 10
mt/kg/day. Survival was 100%, and there was no difference In mean
maternal body weight gain between dosed groups and controls. A persistent
anogenHal staining was observed In dams at the highest dose. There were no
differences In the mean number of Implantations, corpora lutea. live
fetuses, mean fetal body weights or male/female sex ratios among any of the
03650 V-17 04/12/91
-------
groups. There were no statistical differences In malformation or develop-
mental variations compared with the controls when external, soft tissue and
skeletal examinations were performed (IRDC, 1978).
Murray et al. (1980) evaluated the teratogenlc potential of HEX (98%) In
CF-1 mice and New Zealand white rabbits. Mice were dosed at 0. 5, 25 or 75
mg/kg/day HEX by gavage from days 6-15 of gestation while rabbits received
the same dose from days 6-18 of gestation. In the mice, no evidence of
maternal toxlclty, embryotoxIcHy or teratogenlc effects was observed. A
total of 249-374 fetuses (22-33 litters) were examined 1n each dose group.
In rabbits, maternal toxlclty was noted at 75 mg/kg/day (diarrhea,
weight loss and mortality), but there was no evidence of maternal toxlclty
at the lower levels. There were no embryotoxlc effects at any dose level.
Although there was an Increase In the proportion of fetuses with 13 ribs at
75 mg/kg/day, this was considered a minor skeletal variation, and the
authors concluded that HEX was not teratogenlc at the levels tested.
Studies on the teratogenlc potential of Inhaled HEX were not located In
the review of the scientific literature. No data were located that
addressed the reproductive effects of HEX.
Summary
Although there are some Interspecles differences among guinea pigs,
rabbits, rats and mice, HEX vapors are toxic to all species tested. HEX
appears most toxic when administered by Inhalation, with oral and dermal
administration being less toxic routes. The acute lethal concentration
03650 V-18 06/19/90
-------
) of 1.6 and 3.5 ppm In male and female rats, respectively, has been
demonstrated. The oral LD5Q for adult animals Is >500 mg/kg; however,
approximate lethal doses of 180 and 280 mg/kg were determined for mice and
rats, respectively.
Subchronlc oral dosing of rats (38 mg/kg/day) and mice (75 mg/kg/day)
for 91 days produced nephrosls and Inflammation and hyperplasla of the
forestomach. No overt signs of toxUHy were noted when mice and rats were
exposed by Inhalation at 0.2 ppm of HEX (6 hours/day, 5 days/week) for 14
weeks. However, Inhalation exposure of rats at 0.5 ppm for 30 weeks caused
degenerative changes In the liver, respiratory tract and kidneys, in vitro
test results from three species have not shown HEX to be a mutagen. HEX was
also Inactive 1n the mouse dominant lethal assay. In rabbits, maternal
toxlclty has been noted at high oral levels of HEX (75 mg/kg/day}. but there
was no evidence of maternal toxUUy at lower levels. Other research has
not shown any maternal toxlclty to mice. Long-term studies of HEX Inhala-
tion are presently being conducted by the NTP.
03650 V-19 06/19/90
-------
VI. HEALTH EFFECTS IN HUMANS
According to a NIOSH estimate. 1427 workers are occupatlonally exposed
to HEX (NIOSH, 1980). VelsUol officials estimate that -157 employees are
potentially exposed to HEX In their production facilities. Acute human
exposure has been reported In homes near waste sites where HEX has been
disposed (C. Clark et al.. 1982; Ella et al., 1983).
Very little detailed Information Is available concerning the human
health effects of HEX exposure. The most noticeable effect Is the pungent,
Irritating odor produced by HEX. Levins (1980) reported that the odor
threshold of 0.0017 mg/m3 (0.00015 ppm) represented the recognition level
for 100% of the Individuals on a test panel. The study design and method-
ology were not given. Based upon animal studies and observations by
researchers, HEX vapors are very Irritating to all mucous membranes, causing
tearing, sneezing and salivation; skin contact can cause blisters and burns.
Inhalation of vapors or mist can result In the secretion of excess fluid In
the lung, while Inhalation or 1ngest1on may cause nausea, vomiting,
diarrhea, lethargy, respiratory Impairment and Injury to the liver or
kidneys.
Acute Exposure Studies
Treon et al. (1955) reported that members of a group conducting toxldty
tests developed headaches when they were accidentally exposed to unknown
concentrations of HEX, which had escaped Into the room when an aerated
exposure chamber was opened.
03660 VI-1 08/25/88
-------
A well-documented Incident of acute human exposure to HEX occurred In
March 1977 at the Morris Forman Mastewater Treatment Plant In Louisville,
KY. The Incident has been described and reviewed 1n several papers (Wilson
et al.t 1978; Morse et al.t 1979; Komlnsky et al.. 1980). The complete
details of the original NIOSH Hazard Evaluation and Technical Assistance
Report Number TA-77-39 are found In Komlnsky and Wlsseman (1978).
In 1977, the Louisville treatment facility was contaminated with -6 tons
of HEX and smaller amounts of OCCP. a waste by-product of HEX manufacture
(Morse et al., 1979). The contamination was traced to one large sewer line
that passed through several populated areas. Concentrations of HEX detected
In the sewage water at the plant ranged <1000 ppm, and levels In the sewer
line ranged <100 ppm. Air samples from the sewer line showed HEX
concentrations <400 ppb. Although airborne concentrations of HEX at the
time of the exposure were unknown, airborne concentrations In the primary
treatment areas (screen and grit chambers) ranged between 270 and 970 ppb 4
days after the plant had closed. (The ACGIH TWA for HEX was 10 ppb In
1977.) During the cleanup, when workers used steam to remove the
contamination, levels of 19.2 ppm HEX were reported (Komlnsky et al., 1980).
The Centers for Disease Control (CDC) and NIOSH developed questionnaires
regarding the type and duration of symptoms (Morse et al., 1979; Komlnsky et
al., 1980). A total of 193 employees were Identified as those potentially
exposed for 2 or more days during the 2 weeks before the plant was closed
(Morse et al., 1979). Of the 193 employees exposed, 145 (75%) responded.
Physical examinations and blood and urine samples were collected from
workers reporting symptoms of mucous membrane Irritation.
03660 VI-2 08/25/88
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Results of the CDC and NIOSH questionnaires showed that the odor of HEX
was detected before the onset of symptoms by 94% of the workers. The most
common symptoms reported were eye Irritation (59%), headaches (45%) and
throat Irritation (27%) (Table VI-1). Of the 41 workers physically
examined, 6 had physical signs of eye Irritation (I.e., tearing or redness)
and 5 had signs of skin Irritation. Laboratory analyses of blood and urine
specimens from these workers showed elevations of lactic dehydrogenase (LDH)
In 21% and protelnurla In 15%. However, no clinical abnormalities were
reported by the plant physician, the local hospital, or by the Independent
laboratory 3 weeks later (Horse et a!., 1978, 1979).
During clean-up procedures, clinical chemistry parameters were moni-
tored. The only abnormalities noted were several minimal-to-mild altera-
tions In liver function tests (Komlnsky et al., 1980). These abnormalities
are listed In Table VI-2. All workers showing these symptoms also had
physical signs of mucous membrane Irritation. Table Vl-3 summarizes data
for nine workers exposed at the site (Komlnsky et al.. 1980). The exposure
levels could not be estimated accurately because of possible prior exposure
and because the workers had used protective equipment.
A questionnaire was also given to a selected sample of residents of a
48-block area surrounding the contaminated sewer line. A total of 212
occupants were surveyed. Very few residents noted an unusual odor (3.8%).
The most prevalent symptoms were stomachaches (5.2%), burning or watering
eyes (4.7%) and headaches (4.7%). There was no association between symptom
rates and the distance of households from the contaminated sewer line.
03660 VI-3 08/11/88
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TABLE VI-1
Symptoms of 145 Wastewater Treatment Plant Employees
Exposed to HEX (Louisville, KY, March 1977)*
Symptom
Eye Irritation
Headache
Throat Irritation
Nausea
Skin Irritation
Cough
Chest pain
Difficult breathing
Nervousness
Abdominal cramps
Decreased appetite
Decreased memory
Increased saliva
No. of Employees
with Symptom
86
65
39
31
29
28
28
23
21
17
13
6
6
Percent of Employees
with Symptom
59
45
27
21
20
19
19
16
14
12
9
4
4
*Source: Morse et al., 1978
03660
VI-4
09/19/85
-------
TABLE VI-2
Abnormalities for 18 of 97 Cleanup Workers
at the Morris Forman Treatment Plant3
Abnormal Results.
b
Laboratory Test Normal Range
Serum Glutamate-
Oxalacetate Transamlnase (SG01) 7-40 mil/mi
Serum Alkaline Phosphatase 30-100 mU/ms.
Serum Total B1l1rub1n 0.15-10 mg/%
Serum Lactate Dehydrogenase 100-225 mU/mi
Range
40-49
50-59
60-69
70-79
80-89
90-99
100-109
110-119
120-129
1.0-1.9
230-239
No.
5
1
4
0
1
1
3
1
1
lc
1
aSource: Komlnsky et al., 1980
bFor Individuals with more than one serial blood test, only the most
abnormal result Is tabulated.
Associated with serum glutamate-oxalacetate transamlnase of 66
U = Units of enzyme activity
03660
VI-5
08/11/88
-------
o
CO
TABLE VI-3
Overview of Individual Exposure - Symptomatology Correlations at the Morris Forman Treatment Plant9
Case
No.
Estimated Airborne Exposure
Immediate Symptoms
Persistence of Symptoms
Laboratory Abnormalities
19.200 ppb HEX and 650 ppb
OCCP for several seconds (No
protective equipment)
2.3.4 7083 ppb HEX and 446 ppb
OCCP for several seconds.
(Half-face respirator)
5.6 40-52 ppb HEX and 9-21 ppb
OCCP (Half-face respirator)
7,8 Exact exposure unknown
(Half-face respirator)
980 ppb HEX for 15
minutes; OCCP not measured
(No protective equipment)
Lacrlmatlon; skin Irritation on face and
neck; dyspnea and chest discomfort;
nausea (several minutes later)
Lacrlmatlon; Irritation of exposed
skin
Slight eye Irritation
Slight skin Irritation
Irritated eyes
Nasal Irritation and sinus congestion
after 2 weeks of Intermittent exposures
1.5 hrs. post-exposure: Fatigue;
erythema of exposed skin; eye
Irritation subsided In 1 day; chest
discomfort persisted several days.
Asymptomatic at 2 hours, except
for soreness around eyes
No residual after cessation of
exposure
Faces felt 'puffy* and •wlndburned"
for 1-2 days after exposure. This
was noted also by friends and
family. No residual skin lesions.
Eyes felt "dry and Irritated1 for
2-3 days after exposure. Nasal
Irritation ceased within 1-2 days
of cessation of exposure.
Lab work 4 days post expo-
sure was normal1'
Lab work 7 days post expo-
sure was normal on one
workerb
Normal 7 days later on one
None available
None available
aSource: Adapted from Komlnsky et al.. 1980
(•Laboratory work was same as done on cleanup crew
CO
CO
CO
-------
The authors slated that no significant ambient air concentrations of HEX
were found 1n these areas (Komlnsky and Hlsseman, 1978). Residents reported
the same types and frequency of symptoms reported by workers associated with
HEX exposure, which led the authors to suggest that these symptoms were
unrelated to HEX exposure or may be due to other confounding environmental
factors (Horse et al., 1978}.
Several papers have documented a similar Incident of HEX exposure 1n
Hardeman County, TN. 1C. Clark et al., 1982; Meyer, 1983; Ella et al.,
1983}. In 1978, workers at the treatment plant began complaining of acute
symptoms similar to those found In the Louisville plant. Air and wastewater
levels were monitored, and urine, blood and liver function tests were
analyzed, as well as data obtained from an Illness symptom questionnaire.
In the original study design, workers were compared with a control group
from another Memphis treatment plant, which did not receive wastes from the
pesticide manufacturing plant. In a later survey, workers at two other
municipal facilities were used for comparison. In the analysis of the
various monitoring tests, C. Clark et al. (1982) found no statistical
difference In urine samples from both of the Memphis treatment facilities.
In the liver function tests, there were no statistically significant
differences among the values obtained for all survey groups.
At the same time the wastewater treatment plant study was being
performed, an Investigation of residential wells In Hardeman County was
conducted In response to complaints of foul odors and bad taste (Meyer,
1983). In this area also lies a 200 acre chemical land dump, which was
operated from 1964-197?. In 1978, the U.S. Geologic Survey {Sprinkle, 1978;
Rlma, 1979) confirmed the contamination of wells. However, HEX was not
03660 Vl-7 04/12/91
-------
detected In any samples. Urine surveys and liver function analyses were
conducted on residents. A summary of this data 1s presented 1n Table VI-4.
The situation at the Memphis treatment facility Is the only known existing
case of essentially continuous low-level chronic exposures with Intermittent
higher acute exposures, especially during an accidental discharge from the
nearby pesticide manufacturing facility (Ella et al., 1983).
Ep1dem1olog1c Studies
Mortality studies have been conducted on workers Involved In the produc-
tion of HEX or formulation of HEX products. The SMndell and Associates
(1980) report was a cohort study of workers employed at the Velslcol
Chemical Corporation plant at Marshall. Illinois between 1946 and 1979. The
purpose was to evaluate the vital statistics of all former and current
employees (>3 months) who were present during the manufacture of chlordane.
In preparing the cohort, the authors noted the difficulties In tracing some
of the employees. In the final cohort of 783 Individuals, 97.4% of the
employees were located and their vital status Included 1n the study. The
analysis showed no significant differences 1n mortality rates between these
employees and the U.S. population. The observed deaths for all causes,
Including heart disease and cancer, were fewer than the calculated expected
deaths among members of the U.S. population (SMndell and Associates, 1980).
Wang and MacMahon (1979) conducted a study of 1403 males employed at the
Marshall and Memphis plants for >3 months. There were 113 observed deaths
compared with 157 expected, yielding a standardized mortality ratio (SMR) of
72. The two highest SMRs were 134 for lung cancer and 183 for cerebrovascu-
lar disease, but only the latter was statistically significant (p<0.05).
03660 VI-8 08/11/88
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TAl )J-4
Hepatic Profile Comparison of Hardoman County: Exposed Group (November 1978) and Control Group3
o>
o
Parameter1*
Results
November 1978
Exposed Group
Control
Group
Significance of
Difference (t lest)
Alkaline phosphatase (32-72 mil/ml
age 21. 25-150 mil/ml age 21}
Serum gamma glutamlc transamlnase
(SGG1) (5-29 mU/ml)
i> Albumin (3.5-5.0 g/dft)
Total blllrubln (0.1-1.1 mg/dft)
Serum glutamlc pyruvlc transamlnase
(SGOT) (8-22 mil/ml)
Meant 88.1 61.5
Range 34-360 31-220
No. above normal/ 17/36 8/56
total tested
Meanc 9.47 11.56
Range 2-54 4-56
No. above normal/ 3/36 3/56
total tested
Heanc 4.35 4.93
Range 3.9-4.8 4.2-6.2
No. above normal/ 0/36 0/57
total tested
Meant 0.240 0.51
Range 0.1-0.8 0.2-1.7
No. above normal/ 0/31 4/52
total tested
Meanc 19.5 16.08
Range 12-36 9-140
No. above normal/ 11/36 7/56
total tested
0.016
0.430
0.0001
0.0001
0.001
§ aSource: Meyer. 1983
^ Normal range Indicated In parentheses
S cGeometrlc mean
U = Units of enzyme activity
-------
The authors suggested that these effects were unrelated to exposure because
the deaths showed no consistent pattern with duration of employment or with
duration of follow-up.
A study of >1000 employees (93% of the cohort) at the Memphis, TN plant
from 1952-1979 was conducted by Shlndell and Associates (1981). The
researchers found no significant difference 1n mortality between the control
and exposure groups and fewer deaths In the study group than predicted from
vital statistics rates. The Investigators also reported there was no excess
mortality by job function.
Buncher et al. (1980) studied the mortality rates of workers at a
chemical plant that produced HEX. The Investigators reviewed personnel
records for those who worked for at least 90 days between October 1, 1953
and December 31, 1974. There were 341 workers (287 male and 54 female) who
fit the criteria. Health status was ascertained through 1978 and expected
numbers of deaths were calculated based upon the U.S. population and
specifics for sex, age and calendar year. The SMR was 69 showing the
workers to be healthier than the general population. Deaths caused by
specific cancers, all cancers, disease of the circulatory and digestive
systems were fewer than the expected numbers. The authors noted that the
time since Initial exposure, <25 years, reduced the power of the study to
detect cancers, which may have a 10-40 year latent period.
Summary
There 1s only limited Information on the effects of HEX In humans.
Acute Inhalation exposure has resulted In headaches and severe Irritation of
the eyes, nose, throat and lungs. Dermal contact may cause severe burns and
03660 VI-10 08/25/88
-------
skin Irritation. Epldemlologlc studies have generally shown no significant
differences In mortality rates of workers exposed to HEX In the workplace
and the general population. Although a significant excess of deaths from
cerebrovascular disease was reported 1n one study, no consistent pattern was
observed with duration of employment or follow-up.
Current human exposure 1s limited to Improper handling and disposal and
proximity to either manufacturing sites utilizing HEX or disposal sites. No
other chronic human health effects data from HEX exposure have been located
In the literature.
03660 Vl-11 08/25/88
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VII. MECHANISMS OF TOXICITY
The mechanisms underlying HEX toxlclty are not well understood, mainly
because of Its high chemical reactivity and ability to polymerize even at
low temperature. HEX has been shown to react with oleflns and other organic
molecules, such as aromatic compounds. In vitro studies by El Dareer et al.
(1983) have Indicated that HEX readily binds to organic material and can be
easily extracted.
While the available literature does not provide any single mechanism to
account for HEX toxlclty, Lawrence and Borough (1982) showed that all routes
of exposure caused necrotlc lung tissue. Exposure to HEX vapors results In
Irritation of the respiratory tract and may lead to bronchopneumonla and
respiratory failure (D. Clark et al., 1982). In comparison, the degenera-
tive changes 1n the liver and kidneys are mild and unlikely to contribute to
the chemical's toxlclty (NAS, 1978; SRI, 1980a,b).
Absorption from the Gl tract was reported to be relatively Inefficient.
This may be due to the poor bloavallabllHy of HEX when administered by the
oral route or to Us rapid hepatic metabolism and biliary excretion (El
Dareer et al., 1983; Lawrence and Dorough, 1962). Studies by Yu and Atallah
(1981) Indicate that the gut and fecal flora may have a major role 1n the
metabolism of HEX and that enzyme dependent processes may be limited.
Absorption following dermal exposure resulted In marked distinct skin
discoloration, and In some cases, mortality. This might suggest a "site of
uptake" Interaction, which may be similar to that observed In the lung after
pulmonary uptake.
03670 VII-1 08/25/88
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Following Inhalation and the diffusion of HEX through the lung tissue to
the blood, metabolism to water-soluble compounds may occur, but attempts to
collect and Identify metabolites have been unsuccessful. The relatively
slow elimination of the radlolabel from the systemic circulation after l.v.
dosing with 14C-HEX (approximate terminal half-life of 30 hours) suggests
potential for bloaccumulatlon or repeated dosing (Lawrence and Dorough,
1981, 1982).
Rand et al. (1982b) showed that significant changes at the cellular
level In lung tissue occurred after HEX Inhalation. HEX vapors affect the
extracellular lining, and In some cases, cause significant Increases In
hemoglobin and red blood cells. Although more severe effects seem to occur
by Inhalation, Impaired breathing was seen 1n most experiments regardless of
the route of exposure.
Summary
Little Is known about the mechanism of HEX toxldty. Because of Us
high reactivity and volatility, attempts to Identify the reactive metabo-
lites of HEX and to characterize Us mechanism of toxldty have been largely
unsuccessful. Data gaps remain 1n several areas: 1) the question of
whether similar metabolites are formed following administration by different
routes; 2) comparison of toxldty of the parent compound and Its metabolites
1n eliciting the toxic effects seen following HEX exposure; and 3) Inter-
action of HEX with organic compounds such as hemoglobin. Finally, HEX Is
often contaminated by products used 1n Us manufacture or In the manufactur-
ing of other products; therefore, the possibility for co-exposure may exist.
03670 VI1-2 08/30/88
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VIII. QUANTIFICATION OF TOXICOLOGIC EFFECTS
Introduction
The quantification of toxlcologlc effects of a chemical consists of
separate assessments of noncardnogenlc and carcinogenic health effects.
Chemicals that do not produce carcinogenic effects are believed to have a
threshold dose below which no adverse, noncarclnogenic health effects occur.
while carcinogens are assumed to act without a threshold.
In the quantification of noncarclnogenic effects, a Reference Dose
(RfD), [formerly termed the Acceptable Dally Intake (ADI)] 1s calculated.
The RfD Is an estimate (with uncertainty spanning perhaps an order magni-
tude) of a dally exposure to the human population (Including sensitive
subgroups) that Is likely to be without an appreciable risk of deleterious
health effects during a lifetime. The RfD Is derived from a no-observed-
adverse-effect level (NOAEL), or lowest-observed-adverse-effect level
(LOAEL), Identified from a subchronlc or chronic study, and divided by an
uncertainty factor(s) times a modifying factor. The RfD is calculated as
follows:
RfD .
-------
U.S. EPA (1991) employs a modification to the guidelines proposed by the
National Academy of Sciences (MAS, 1977, 1980) as follows:
Standard Uncertainty Factors (UFs)
Use a 10-fold factor when extrapolating from valid experimental
results from studies using prolonged exposure to average healthy
humans. This factor 1s Intended to account for the variation
In sensitivity among the members of the human population. [10H]
Use an additional 10-fold factor when extrapolating from valid
results of long-term studies on experimental animals when
results of studies of human exposure are not available or are
Inadequate. This factor 1s Intended to account for the uncer-
tainty In extrapolating animal data to the case of humans.
[10A]
Use an additional 10-fold factor when extrapolating from less
than chronic results on experimental animals when there Is no
useful long-term human data. This factor 1s Intended to
account for the uncertainty In extrapolating from less than
chronic NOAELs to chronic NOAELs. [10S]
Use an additional 10-fold factor when deriving an RfD from a
LOAEL Instead of a NOAEL. This factor Is Intended to account
for the uncertainty In extrapolating from LOAELs to NOAELs.
[10L]
Modifying Factor (MF)
Use professional judgment to determine another uncertainty
factor (MF) that Is greater than zero and less than or equal to
10. The magnitude of the MF depends upon the professional
assessment of scientific uncertainties of the study and data
base not explicitly treated above, e.g., the completeness of
the overall data base and the number of species tested. The
default value for the MF 1s 1.
The uncertainty factor used for a specific risk assessment Is based
principally upon scientific Judgment rather than scientific fact and
accounts for possible Intra- and Interspecles differences. Additional
considerations not Incorporated In the NAS/ODW guidelines for selection of
an uncertainty factor Include the use of a less than lifetime study for
deriving an RfD, the significance of the adverse health effects and the
counterbalancing of beneficial effects.
03680 VII1-2 04/12/91
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From the RfD, a Drinking Water Equivalent Level {DUEL} can be calcu-
lated. The DVIEL represents a medium specific (I.e., drinking water)
lifetime exposure at which adverse, noncardnogenlc health effects are not
anticipated to occur. The DUEL assumes 100% exposure from drinking water.
The DUEL provides the noncardnogenlc health effects basis for establishing
a drinking water standard. For Ingestlon data, the DUEL 1s derived as
follows:
IRfDl x (Body weight In kg)
' — = mq/8.
Drinking Water Volume In I/day
where:
Body weight = assumed to be 70 kg for an adult
Drinking water volume = assumed to be 2 ft/day for an adult
In addition to the RfD and the DUEL, Health Advisories (HAs) for expo-
sures of shorter duration (1-day, 10-day and longer-term) are determined.
The HA values are used as Informal guidance to municipalities and other
organizations when emergency spills or contamination situations occur. The
HAs are calculated using an equation similar to the RfD and DWEL; however,
the NOAELs or LOAELs are Identified from acute or subchronlc studies. The
HAs are derived as follows:
HA . (NOAEL or LDAEL) x (bw) =
(UF) x ( a/day) y
Using the above equation, the following drinking water HAs are developed
for noncardnogenlc effects:
1. 1-day HA for a 10 kg child Ingesting 1 I water per day.
2. 10-day HA for a 10 kg child Ingesting 1 ft water per day.
3. Longer-term HA for a 10 kg child Ingesting 1 I water per day.
4. Longer-term HA for a 10 kg adult Ingesting 2 ft water per day.
03680 VII1-3 04/10/88
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The 1-day HA calculated for a 10 kg child assumes a single acute
exposure to the chemical and Is generally derived from a study of <7 days
duration. The 10-day HA assumes a limited exposure period of 1-2 weeks and
Is generally derived from a study of <30 days duration. The longer-term HA
1s derived for both the 10 kg child and a 70 kg adult and assumes an
exposure period of -1 years (or 10% of an Individual's lifetime). The
longer-term HA Is generally derived from a study of subchronlc duration
(exposure for 10% of animal's lifetime).
The U.S. EPA categorizes the carcinogenic potential of a chemical, based
on the overall welght-of-evldence, according to the following scheme:
Group A: Human Carcinogen. Sufficient evidence exists from
epidemiology studies to support a causal association between
exposure to the chemical and human cancer.
Group B: Probable Human Carcinogen. Sufficient evidence of
carclnogenlclty In animals with limited (Group Bl) or Inade-
quate (Group B2) evidence 1n humans.
Group C: Possible Human Carcinogen. Limited evidence of
carclnogenlcUy 1n animals In the absence of human data.
Group D: Not Classified as to Human CarclnogenlcUy. Inade-
quate human and animal evidence of carclnogenlcUy or for which
no data are available.
Group E: Evidence of Noncarclnogenklty for Humans. No
evidence of carclnogenlcUy 1n at least two adequate animal
tests 1n different species or In both adequate epldemlologlc
and animal studies.
If toxlcologlc evidence leads to the classification of the contaminant
as a known, probable or possible human carcinogen, mathematical models are
used to calculate the estimated excess cancer risk associated with the
Ingestlon of the contaminant In drinking water. The data used In these
03680 VI11-4 04/12/91
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estimates usually come from lifetime exposure studies using animals. In
order to predict the risk for humans from animal data, animal doses must be
converted to equivalent human doses. This conversion Includes correction
for noncontlnuous exposure, less than lifetime studies and for differences
In size. The factor that compensates for the size difference Is the cube
root of the ratio of the animal and human body weights. It Is assumed that
the average adult human body weight 1s 70 kg and that the average water
consumption of an adult human Is 2 a of water per day.
For contaminants with a carcinogenic potential, chemical levels are
correlated with a carcinogenic risk estimate by employing a cancer potency
(unit risk) value together with the assumption for lifetime exposure from
Ingestlon of water. The cancer unit risk 1s usually derived from a linear-
ized multistage model with a 95% upper confidence limit providing a low dose
estimate; that 1s, the true risk to humans, while not Identifiable, Is not
likely to exceed the upper limit estimate and. In fact, may be lower.
Excess cancer risk estimates may also be calculated using other models such
as the one-hit, Welbull, loglt and problt. There 1s little basis In the
current understanding of the biologic mechanisms Involved In cancer to
suggest that any one of these models Is able to predict risk more accurately
than any other. Because each model 1s based upon differing assumptions, the
estimates derived for each model can differ by several orders of magnitude.
The scientific data base used to calculate and support the .setting of
cancer risk rate levels has an Inherent uncertainty that Is due to the
systematic and random errors 1n scientific measurement. In most cases, only
studies using experimental animals have been performed. Thus, there Is
03680 VI11-5 04/12/91
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uncertainty when the data are extrapolated to humans. When developing
cancer risk rate levels, several other areas of uncertainty exist, such as
the Incomplete knowledge concerning the health effects of contaminants 1n
drinking water, the Impact of the experimental animal's age, sex and
species, the nature of the target organ system(s) examined and the actual
rate of exposure of the Internal targets In experimental animals or humans.
Dose-response data usually are available only for high levels of exposure
and not for the lower levels of exposure closer to where a standard may be
set. When there 1s exposure to more than one contaminant, additional
uncertainty results from a lack of Information about possible synerglstlc or
antagonistic effects.
NoncarclnogenU Effects
Limited quantitative Information 1s available regarding the human health
effects resulting from HEX exposure; however, transient or acute exposure to
HEX vapor has been associated with Irritation to the eyes, nose and throat,
as well as headaches and nausea. In animals, comparative studies have
Indicated that HEX exposure Is more toxic following Inhalation than oral
exposure.
Short-term studies by IROC (1978) and SRI (1980a,b, 1981a,b) provide
substantial data on oral toxldty to rats and mice. The Southern Research
Institute (SRI. 1981a,b) studies are of longer duration and provide NOAELs
and LOAELs. A Russian study by Nalshteln and Llsovskaya (1965) was
conducted for 6 months and also yielded no-effect levels; however, an HA
cannot be recommended since the complete study design was not reported.
03680 VIII-6 06/19/90
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More recent studies have been published (SRI, 1980a.b, 1981a,b), which
provide greater detail, use more appropriate toxlcologlc endpoints and
define NOAELs and LOAELs than the earlier study by Nalshteen and Llsovskaya
(1965) which served as the basis for the 1980 Ambient Hater Quality Criteria
Document (U.S. EPA, 1980). These studies are reviewed below and have been
used In the calculations of the 1-day, 10-day and longer-term HAs and the
lifetime DUEL.
SRI (1980a) conducted a gavage study using HEX with male and female F344
rats and B6C3F mice. Dose levels of 0, 75, 150, 300, 600 and 1200 mg/kg
HEX In corn oil were administered to both rats and mice by gavage. One dose
at each of the five levels was given to five animals of each sex.. Clinical
observations related to the administration of HEX Included a decrease In
activity, ruffled fur, wet fur In the anal area and diarrhea. These
alterations occurred within 6-24 hours of dosing; severity and duration were
related to the dose level. All rats In the 600 and 1200 mg/kg dose groups
died, while In the 300 mg/kg dose group, two female rats died. In the 150
and 75 mg/kg rat dose groups, ruffled fur occurred, but no deaths. In the
mouse study, all 1200 mg/kg dose group animals died. At the 600 mg/kg
level, only one female and one male mouse died. At the other levels, only
minor effects were observed. The 150 mg/kg rat level was observed to be the
NOAEL In rats, while the 300 mg/kg was chosen as the NOAEL In mice.
The SRI (1980b) conducted a repeated-dose study In which F344 rats and
B6C3F] mice were exposed to 0, 25, 50, 100. 200 and 400 mg/kg HEX and 0,
50, 100, 200, 400 and 800 mg/kg HEX 1n rats and mice, respectively. All
doses were administered by gavage with corn oil used as the vehicle. Five
03680 VIII-7 04/12/91
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mice and five rats of each sex were used for each dose level on a schedule
of 12 dosing days (days 1-5, 8-12, 15 and 16), with at least 2 consecutive
dosing days before the terminal sacrifice began.
In the rat study (SRI. 1980b), all males and 4/5 females at the 400
mg/Jcg HEX dose level died, while at the POO mg/kg dose, 1/5 males and 4/5
females died. Clinical observations were performed twice dally. At the 100
mg/kg dose level, ruffled fur occurred 1n all rats. The Investigators noted
that there was significant weight depression In both the males and the
females at this dose rate, but there were no deaths or life-threatening
clinical and gross observations. At the 50 mg/kg HEX dose level, there were
no significant clinical signs, but there were gross changes to the stomach
wall and a depression In weight gain. The gross observations at the 50
mg/kg dose level Included discolored raised areas on the mucosal surface of
the stomach In 3/5 females and 3/5 males. The decrease In weight gain 1n
males was -17% and females -22%. At the 25 mg/kg level, the clinical and
gross observations did not find any Irregularities. The weight gain
depressions were 8% for males and 11% for females. The 25 mg/kg dose level
was selected as the NOAEL for rats.
In the mouse study (SRI. 1980b), all of the animals treated at the 800
and 400 mg/kg dose level died. However, one of these deaths was attributed
to Improper gavage technique. At the 200 mg/kg dose level, two animals died
because of Inappropriate experimental techniques, as did one at the 50 mg/kg
level and two of the controls. One animal's death at the 200 mg/kg level
was attributed to chemical toxldty. The 100 mg/kg HEX dose was determined
to be the NOAEL, since there were effects but none considered adverse.
03680 VI1I-8 04/12/91
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In a 13-week oral toxUHy study (SRI, 1981a,b; Abdo et al.. 1984), HEX
was administered In corn oil by gavage to groups of 10 male and 10 female
F344 rats at doses of 0, 10, 19, 38, 75 and 150 mg/kg and to 10 mice of each
sex at doses of 0, 19, 38, 75, 150 and 300 mg/kg. The doses were glve'n
dally, 5 days a week for 13 weeks. Compared with the previously-reviewed
shorter-term studies, HEX was found to be more toxic at the lower dose
levels. All 10 male and 3 female mice died at the 300 mg/kg HEX dose
level. At the 150 mg/kg level, six male rats died, while one rat In the 75
mg/kg group died as a result of chemical toxlclty. Other deaths occurred,
but these were attributed to Improper gavage technique by the Investi-
gators. Stomach lesions (hyperplasla and focal Inflammation) were observed
1n male and female rats at the 38. 75 and 150 mg/kg dose groups and only
female rats at the 19 mg/kg dose level. At higher doses (>150 mg/kg) toxic
nephrosls In females was observed. According to the Investigators, these
lesions decreased In severity with decreasing dose, with hyperplasla being
the primary lesion. One female rat at the 19 mg/kg dose level had a
squamous cell papllloma of the epithelial surface. The only effect reported
for the 10 mg/kg dose was a slight depression In body weight In both males
and females.
In mice (SRI, 1981a), the stomach lesions occurred at the 38 mg/kg and
higher dose levels, Including deaths at the two highest levels. In the
published results (Abdo et al., 1984), the "no observed toxic effect level"
of HEX was stated to be at the 19 mg/kg level for rats. In addition, the
authors stated that rats of both sexes and female mice are approximately
equally susceptible to the toxic effects of HEX administered by gavage over
a 90-day period. Therefore, the NOAEL was 10 mg/kg HEX for rats and 19
03680 VIII-9 04/12/91
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mg/kg for mice. The lower level In rats was selected because the lesions
were still considered toxic effects, but In the long-term may or may not be
adverse. Therefore, by using this level, an added margin of safety 1s
Inherent In the calculation of the DUEL. Because HEX toxlclty Increase's
with duration, this NOAEL would accommodate the possibility of these effects
becoming adverse over the longer-term.
Quantification of Noncarclnogenlc Effects
Although there Is some question as to the route of HEX transport In the
body, the kidneys, liver and lungs appear to be the major target organs.
Investigators have observed sex and species differences 1n toxic response to
HEX. In addition, some observed effects In these studies may have resulted
from a combined exposure to HEX and other compounds. In the Abdo et al.
(1984) study, hexachloro-l,3-butad1ene (HCBD) was present as a contaminant
of HEX. Since HCBD Is a known nephrotoxln In rodents, some of the adverse
effects seen In the study may be attributed to either or both compounds.
Derivation of 1-Day HA. In the range-finding study by SRI (1980a)
rats exposed to a single oral dose of 150 mg/kg HEX experienced "wet fur In
anal area" and ruffled fur when observed for 14 days. No other toxic
effects were seen at this dose level. At higher doses, mortality and
diarrhea were observed. A NOAEL of 150 mg/kg can be derived based on the
absence of adverse effects seen at this level.
The 1-day HA for a child Is calculated as follows:
HA m 150 mq/kq x 10 kq =
100 x 1 I/day
03680 VII1-10 04/12/91
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where:
ISO mg/kg = determined to be the NOAEL of the SRI (1980a) rat
study
10 kg = assumed body weight of a child
1 l/day = assumed water consumption by a child
100 = uncertainty factor chosen In accordance with
NAS/ODU and Agency guidelines 1n using a NOAEL from
an animal study
This HA Is equivalent to 15 mg/day or 1.5 mg/kg/day. It should be noted
that this level (15 mg/i) 1s above the reported water solubility of HEX
(1.2-3.4 mg/l) under normal ambient conditions; however, under certain
conditions (e.g., Increased temperature) the water solubility of HEX can
Increase.
Derivation of 10-Dav HA. The SRI (1980b) repeated-dose toxlclty study
was used. In this study a NOAEL of 25 mg/kg, based on no significant
decreases In weight gain In male and female rats was defined following
-exposure to 25, 50, 100, 200 and 400 mg/kg HEX for 12 days (days 1-5, 8-12,
and 15 and 16).
The 10-day HA for a child 1s calculated as follows:
10-day HA = 2S m(l/kq x 10 kq x °'714 = 1.8 mg/l (rounded to 2 mg/a)
100 x 1 l/day
where:
25 mg/kg = determined to be the NOAEL of the SRI (1980b) female
rat study
10 kg = assumed body weight of a child
0.714 = the correction factor (5/7) to adjust for continuous
exposure from a 5- to 7-day exposure
03680 VIII-11 06/11/91
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100 = uncertainty factor chosen in accordance wHh NAS/QDW
and Agency guidelines for use with a NOAEL from an
animal study
1 l/day = assumed water consumption by a child
This HA 1s equivalent to l.B mg/day or 0.18 mg/kg/day.
Derivation of Longer-Term HA. The 13-week oral toxicity study by SRI
(19Blb) is the only longer-term oral study of sufficient duration and
experimental design that can be used for calculating longer-term HAs. In
this study, HEX was administered in corn oil by gavage to groups of 10 male
and female f344 rats at doses of 0, 10, 19, 38, 75 and 150 mg/kg.
Treatment-related dose-dependent adverse effects Including Inflammation of
the stomach were seen with increasing severity at >19 mg/kg. The only
effect reported at the 10 mg/kg dose was a slight depression in body weight
in males and females.
The longer-term HA for a child Is calculated as follows:
i • u* 10 mg/kq x 10 kg x 0.714 ., _ m..
Longer-term HA = a , * •—a = 0.7 mg/J.
100 x 1 i/day
This HA is equivalent to 0.07 mg/kg/day.
where:
10 mg/kg = NOAEL from the SRI (1981b; Abdo et al.t 1984) study
based on absence of stomach lesions
10 kg = assumed body weight of a child
0.714 = the correction factor (5/7) to adjust for
continuous exposure from a 5- to 7-day exposure
100 = uncertainty factor chosen In accordance with
NAS/ODW and Agency guidelines for use with a NOAEL
from an animal study
03680 VJII-12 04/12/91
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1 a/day = assumed water consumption by a child
For an adult:
10 mg/kg x 70 kg x 0.714 9 c .
Longer-term HA . 2 i/day x 100 (rSundSfto 3 mg/i)
where:
10 mg/kg = NOAEL from the SRI (1981D; Abdo et al.. 1984) study
based on absence of stomach lesions
70 kg = assumed body weight of an adult
0.714 = the correction factor (5/7) to adjust for
continuous exposure from a 5- to 7-day exposure
2 ft/day = assumed water consumption by an adult
100 = uncertainty factor chosen In accordance with
NAS/ODW and Agency guidelines for use with a NOAEL
from an animal study
Assessment of Lifetime Exposure and Derivation of DWEL. In a 13-week
oral toxlclty study by SRI (1981b), rats (10/sex/group) were exposed by
gavage with 0. 10, 19, 38, 75 or 150 mg HEX/kg bw Tor 5 days/week.
Epithelial hyperplasla, focal Inflammation of the forestomach and stomach
lesions, and nephrosls were observed with Increased Incidence and severity
1n both sexes at >38 mg/kg bw. At the 19 mg/kg/day dose similar effects
were observed In males only. A slight but nonsignificant depression In body
weight gain was seen at 10 mg/kg 1n males and 1n females at 19 mg/kg.
Therefore the 10 mg/kg dose was considered a NOAEL and the 19 mg/kg dose a
LOAEL.
Step 1: Determination of RfD
RfD = 10 mq/kq x 0.714 = Q Q0714 mg/kg/day (rounded to 0.007 mg/kg/day)
100 x 10
03680 VIII-13 06/11/91
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where:
10 mg/kg = NOAEL from the SRI (1981b; Abdo et al.. 1984) study
based on a lack of adverse effects
0.714 = the correction factor (5/7) to adjust for continuous
exposure from a 5- to 7-day exposure
100 -= uncertainty factor chosen In accordance with NAS/OOW
and Agency guidelines for use with a NOAEL from an
animal study
10 = uncertainty factor appropriate for use with study data
that are significantly less-than-Hfetlme 1n duration
Step 2: Determination of the Drinking Water Equivalent Level (DWEL)
DWEL __ 0.007 mq/kq/dayx 70 kq = Q ^ M t() Q 3
2 a/day
where:
0.007 mg/kg/day = RfD
70 kg = assumed weight of an adult
2 l/day = assumed water consumption by an adult
A summary of noncarclnogenU effects Is listed In Table VIII-1.
Carcinogenic Effects
The data base Is neither extensive nor adequate for assessing the
cardnogenldty of HEX. The National Toxicology Program (NTP) has completed
a subchronlc animal Inhalation study and Is presently conducting a lifetime
animal Inhalation bloassay using both rats and mice. A judgment of
carcinogenic potential will be deferred until the results of the long-term
NTP bloassay are available. Using the IARC criteria, the available evidence
matches the overall Group 3 category. According to the U.S. EPA Guidelines
for Carcinogen Risk Assessment, this chemical Is classified In the Group D
03680 VIII-14 06/11/91
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TABLE VIII-1
Summary of HAs and DUEL for Noncarclnogenlc Effects
Drinking Water
Concentration
(mg/i)
Reference
1-Day HA (10 kg child)
10-Day HA (10 kg child)
Longer-term HA (10 kg child)
Longer-term HA (70 kg adult)
DUEL
15
2
0.7
3
0.3
SRI, 1980a
SRI, 1980b
SRI. 1981b
SRI, 1981b
SRI. 1981b
03680
VIII-15
06/12/91
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category, which Indicates that the available data base Is Inadequate to
assess the carcinogenic potential of this substance (U.S. EPA, 1986).
ExIsUIng Guidelines. Recommendations and Standards
EPA Guidelines. An RfO of 0.007 mg/kg/day was verified by the RfD
Work Group orr 10/09/85 (U.S. EPA, 1991). The CRAVE Work Group verified a
classification of Group D (not classifiable as to cardnogenldty for
humans) for HEX on 10/05/89 (U.S. EPA. 1991).
Occupational Standards. There 1s no current Occupational Safety and
Health Administration (OSHA) standard for HEX levels In the workplace.
However, the American Conference of Governmental Industrial Hyg1en1sts
(ACGIH, 1986) has adopted a TLV. expressed as an 8-hour TWA of*0.1 mg/m3
(0.01 ppm). The levels are based on the data from the Inhalation study by
Treon et al. (1955).
The National Institute for Occupational Safety and Health (NIOSH, 1978)
classlfed HEX In the Group II pesticide category and recommended criteria
for standards for occupations 1n pesticide manufacturing and formulating.
These standards rely on engineering controls, work practices and medical
surveillance programs, rather than workplace air limits, to protect workers
from the adverse effects of pesticide exposure In manufacturing and formu-
lating (NIOSH, 1978).
Transportation Regulations. The Hazardous Materials Transportation
Act specifies the requirements to be observed 1n the preparation for ship-
ment and transport of hazardous materials. The transport of HEX by air,
03680 VIII-16 04/1?/91
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land and water Is regulated by these statutes, and the Department of Trans-
portation has designated HEX as "hazardous material," a "corrosive material"
and a "hazardous substance." The maximum net quantity for transport by
passenger-carrying aircraft or rallcar has been set at 10 gallons per
package. Transport on deck or below deck by cargo vessel 1s also permitted.
Solid Haste Regulations. Under the Resource Conservation and Recovery
Act (RCRA), the U.S. EPA has designated HEX as a hazardous toxic waste,
Hazardous Waste No. U 130, subject to disposal and permit regulations (40
CFR 262-265 and 122-124).
Food Tolerances. Under the Federal Insecticide Fumlgant and Rodentl-
clde Act (F1FRA), a tolerance of 0.3 ppm has been established for chlordane
residues, which are not to contain >1X of HEX {40 CFR 180.122).
Water Regulations. Under Section 311 of the Federal Hater Pollution
Control Act, the U.S. EPA designated HEX as a hazardous substance and estab-
lished a reportable quantity (RQ) of 1 pound (0.454 kg) for HEX. Discharges
equal to or greater than the RQ Into or upon U.S. waters are prohibited
unless the discharge 1s In compliance with applicable permit programs.
Under the Clean Water Act, the U.S. EPA has designated HEX as a toxic
pollutant (I.e., priority pollutant). Effluent limitations guidelines, new
source performance standards, and pretreatment standards have been developed
or will be developed for the priority pollutants for 21 major Industries.
Under the Clean Water Act, an ambient water quality criteria level for
HEX was also developed (U.S. EPA, 1980). Based on available toxlclty data
03680 V11I-17 04/12/91
-------
for the protection of public health the level derived was 206
Using organoleptlc data for controlling undesirable taste and odor of
ambient water, the estimated level was 1 pg/a. (U.S. EPA, 1980).
AU Regulations. HEX 1s not regulated under the Clean A1r Act. The
U.S. EPA, after evaluating the current exposure data and ambient air levels
of HEX, Issued a decision not to list HEX as a hazardous air pollutant.
Other Regulations. Pursuant to rules under sections 8(a) and 8(d) of
the Toxic Substances Control Act (TSCA), all manufacturers of HEX are
required to report health and safety Information on HEX to the U.S. EPA's
Office of Toxic Substances. The deadline for submission of Preliminary
Assessment Information Manufacturer's Report on HEX was November 19, 198?.
In 1979, the Interagency Testing Committee (ITC) recommended that HEX be
considered for health and environmental effects testing under Section 4(a)
of the TSCA (44 FR 31866). This recommendation was based on evidence of
potential human exposure and a potential for environmental persistence and
bloaccumulatlon. The U.S. EPA (1982) responded In the Federal Register.
The following Is the statement from that notice:
EPA has decided not to Initiate rulemaklng to require testing of
HEX under section 4 of TSCA because EPA does not believe that there
Is a sufficient basis to find that current manufacture, distribu-
tion In commerce, processing, use or disposal of HEX may present an
unreasonable risk of Injury to the environment or of mutagenlc and
teratogenlc health effects. Neither has the EPA found evidence
that there 1s substantial or significant environmental release of
HEX. In addition, certain new studies have become available since
the ITC's report or are underway, making additional testing for
chronic and oncogenlc effects unnecessary.
03680 V1II-18 04/1P/91
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Special Groups at Risk
While very little data are available concerning the effects of HEX
exposure on humans, It Is apparent that those Individuals with respiratory
problems or diffidences will be most sensitive to HEX exposure. Workers
repeatedly exposed to HEX vapors are also at high risk.
03680 VIII-19 04/12/91
-------
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