United States
Environmental Protection
Agency
Office of Health and
Environmental Assessment
Washington DC 2O46O
EPA-600/8-84-001 F
November 1984
Final Report
Research and Development
&EPA
Health Assessment
Document for
Hexachlorocyclo-
pentadiene
Final
Report
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EPA-600/8-84-001F
November, 1984
Final Report
HEALTH ASSESSMENT DOCUMENT
FOR
HEXACHLOROCYCLOPENTADIENE
U.S. ENVIRONMENTAL PROTECTION AGENCY
Office of Research and Development
Office of Health and Environmental Assessment
Environmental Criteria and Assessment Office
Cincinnati, Ohio 45268
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DISCLAIMER
This document has been reviewed In accordance with U.S. Environmental
Protection Agency policy and approved for publication. Mention of trade names
or commercial products does not constitute endorsement or recommendation for
use.
11
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PREFACE
The Office of Health and Environmental Assessment of the Office of
Research and Development has prepared this Health Assessment, Document (HAD)
at the request of the Office of Air Quality Planning and Standards. Hexa-
chlorocyclopentadlene (HEX) 1s an Intermediate In the pesticide manufactur-
ing process and 1s currently being studied by the Environmental Protection
Agency (EPA) to determine 1f 1t should be regulated as a hazardous air
pollutant under Section 112 of the Clean Air Act.
The scientific literature has been searched and Inventoried, key studies
have been reviewed and evaluated and summaries and- conclusions have been
directed at Identifying the health effects from exposure to HEX. At several
stages 1n the HAD development process, the HEX document has been reviewed
for scientific and technical accuracy. These peer reviews have been by
scientists from Inside and outside the EPA. Observed effect levels and
dose-response relationships are discussed where appropriate in order to
Identify the critical effect and to place adverse health responses 1n
perspective with observed environmental effects.
111
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ACKNOWLEDGEMENTS
The EPA's Office of Health and Environmental Assessment (OHEA) was
responsible for the preparation of this health assessment document. The
OHEA Environmental Criteria and Assessment Office in Cincinnati (ECAO-Cin)
had overall responsibility for coordination and direction of the document
(David J. Reisman, Project Manager; Jerry F. Stara, Office Director). David
J. Reisman served as the principal author of this document. The following
people contributed substantial portions of various chapters and their
assistance has been greatly appreciated:
Finis Cavender
Dynamac Corporation
11140 Rockville Pike
Rockville, MD 20852
Shane Que Hee
Department of Environmental Health
University of Cincinnati
Cincinnati, OH
W. Bruce Peirano
Environmental Criteria and Assessment Office
U.S. Environmental Protection Agency
Cincinnati, OH 45268
Randall J.F. Bruins
Environmental Criteria and Assessment Office
Environmental Protection Agency
Cincinnati, OH 45268 :
Charles H. Nauman
OHEA - Exposure Assessment Group
U.S. Environmental Protection Agency
Washington, DC 20460
Dharm V. Singh
OHEA - Carcinogen Assessment Group
U.S. Environmental Protection Agency
Washington, DC 20460
Sheila Rosenthal
OHEA - Reproductive Effects Assessment Group
Washington, DC 20460
1v
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The following Individuals provided reviews of this publication and/or
earlier drafts of this document:
U.S. Environmental Protection Agency
Environmental Criteria and Assessment Office
Michael Dourson
Linda Erdreich
Richard Hertzberg
Franklin Mink
Jennifer Orme
William Pepelko
Office of Toxic Substances
Ralph Northrop
Carol Glasgow
Harold Day
Office of Air Quality Planning and Standards
Tim Hohin, OAQPS Project Manager
Larry J. Zaragoza
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CONSULTANTS..REVIEWERS AND CONTRIBUTORS
James R. WUhey , ,
Health and Welfare, Canada
Foods Directorate
Ross Avenue
Tunney's Pasture
Ottawa, Ontario
Canada K1A OL2
Fumlo Hatsumura
Pesticide Research-Center
Michigan State University
East Lansing, Michigan 48824
Joseph F. Borzelleca
Division of Toxicology
Department of Pharmacology
Medical College of Virginia
Richmond, Virginia 23298
Abdo
Kama! M.
NIEHS
P.O. Box 12233
Research Triangle Park, NC
27709
C. Scott Clark
Department of Environmental Health
University of Cincinnati
Cincinnati, Ohio
James 6. Col son
Occidental Chemical Corporation
Long Road
Grand Island, New York 14072
Alfred A. Levin
Velsicol Chemical Corporation
341 East Ohio Street
Chicago, Illinois 60611
Jack L. Egle
Medical College of Virginia
Richmond, Virginia 23298
DOCUMENT PRODUCTION
Technical Support Services Staff: C.A. Cooper, P.A. Daunt, E.R. Ourden,
C.L. Fessler, K.S. Mann, J.A. Olsen, B.L. Zwayer, Environmental Criteria and
Assessment Office, Cincinnati
vi
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Co-chairmen:
HmchlorocyclopintidUnt Ptir itvltw Pine! Htmben
June 29, 1983 Cincinnati, Ohio
Jerry F. Stara, ECAO-CIN
David J. Relsman, ECAO-CIN
Finis Cavender, The HHre Corporation
James Withey
Frederick Coulston
Mary Anne Zanetos
C. Ralph Buncher
Fumio Matsumura
Wyman Dorough
Joseph Borzelleca
Shane Que Hee
Charles H. Nauman
Randall J.F. Bruins
W. Bruce Pelrano
Linda S. Erdrelch
Richard C. Hertzberg
Ralph Northrop
John Komlnsky
Alfred A. Levin
Mildred S. Root
James Grutsch
Panel Members
Health and Welfare, Canada
Coulston International
Battelle Memorial Institute
University of Cincinnati
Michigan State University
University of Kentucky
Medical College of Virginia
University of Cincinnati
U.S. EPA, OHEA
U.S. EPA, ECAO-CIN
U.S. EPA. ECAO-CIN
U.S. EPA, ECAO-CIN
U.S. EPA, ECAO-CIN
U.S. EPA, OTS
U.S. OHHS, NIQSH
Velslcol Chemical Corp.
Velslcol Chemical Corp.
Velslcol Chemical Corp.
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TABLE OF CONTENTS
Page
1. INTRODUCTION I .............
2. SUMMARY, CONCLUSIONS AND RESEARCH NEEDS
2.1.
2.2.
2.3.
SUMMARY
2.1.1. Properties, Production and Uses
2.1.2. Sources, Environmental Levels, Transport and Fate .
2.1.3. Aquatic Life, Vegetation and Wildlife
2.1.4. Pharmacoklnetlcs, Toxicology, Exposure and
Health Effects
CONCLUSIONS . . .
RESEARCH NEEDS. ..........
3. PHYSICAL AND CHEMICAL PROPERTIES/ANALYTICAL METHODOLOGY
3.1.
3.2.
3.3.
3.4.
SYNONYMS, TRADE NAMES AND IDENTIFICATION
PHYSICAL AND CHEMICAL PROPERTIES
3.2.1. Physical Properties
3.2.2. Chemical Properties
ANALYTICAL METHODOLOGY
3.3.1. Air
3.3.2. Water
3.3.3. Soil
BIOLOGICAL MEDIA.
3.4.1. Sampling.
3.4.2. Analysis
4. PRODUCTION, USE, SOURCES AND AMBIENT LEVELS
4.1.
4.2.
4.3.
4.4.
4.5.
4.6.
PRODUCTION. ...
USE
SOURCES
AMBIENT LEVELS
4.4.1. Air ...
4.4.2. Water .
4.4.3. Food. .
4.4.4. Soil
RELATIVE SOURCE CONTRIBUTIONS
SUMMARY AND CONCLUSIONS
1-1
2-1
2-1
2-1
2-1
2-3
.2-4
2-6
2-6
3-1
3-1
3-1
3-1
3-4
3-6
3-6
3-8
3-11
3-12
3-12
3-13
4-1
4-1
4-1
4-2
4-4
4-4
4-4
4-6
4-6
4-8
4-8
viii
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Page
5. ENVIRONMENTAL FATE AND TRANSPORT. ...... 5-1
5.1. FATE --.... .,, ......-•. . . 5-1
5.1.1. Air • 5-1
5.1.2. Water 5-2
5.1.3. Soil 5~12
5.2. TRANSPORT 5-18
5.2.1. Air 5-18
5.2.2. Water 5-19
5.2.3. Soil •• • • 5-22
5.3. BIOCONCENTRATION/BIOACCUMULATION 5-23
5.4. SUMMARY AND CONCLUSIONS 5-30
6. ECOLOGICAL EFFECTS. 6-1
6.1. EFFECTS ON AQUATIC ORGANISMS 6-1
6.1.1. Freshwater Aquatic Life 6-1
6.1.2. Marine and Estuarine Aquatic Life 6-5
6.2. EFFECTS ON OTHER ECOSYSTEMS 6-7
6.3. EFFECTS ON TERRESTRIAL VEGETATION 6-10
6.4. EFFECTS ON WILDLIFE 6-11
6.5. SUMMARY 6-11
7. TOXICOLOGY AND HEALTH EFFECTS 7-1
7.1. PHARMACOKINETICS. . '. * 7-1
7.1.1. Absorption, Distribution, Metabolism and
Excretion , 7~1
7.1.2. Summary 7~10
7.2. MAMMALIAN TOXICOLOGY 7-11
7.2.1. Acute Toxicity. 7-11
7.2.2. Subchronic Toxicity . 7-15
7.2.3. Chronic Toxicity 7-22
7.3. MUTAGENICITY 7~24
7.3.1. Mutagenicity 7-24
7.3.2. Summary '-26
7.4. CARCINOGENICITY . 7~26
7.4.1. In vivo Carcinogenicity T,-26.
7.4.2. In vitro Carcinogenicity 7-26
7.4.3. Summary 7~27
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7.5. TERATOGENIC AND REPRODUCTIVE EFFECTS y..?-/
7.5.1. Teratogenicity 7,27
7.5.2. Reproductive Effects ! ! ! ! ! 7-28
7.5.3. Summary 7-29
7.6. HUMAN EXPOSURE AND HEALTH EFFECTS 7-29
7.6.1. Human Exposure 7-29
7.6.2. Health1 Effects 7-29
7.6.3. Summary 7...42
8. OVERVIEW 8-1
8.1. EFFECTS OF MAJOR CONCERN 8-1
8.1.1. Principal Effects and Target Organs 8-1
8.1.2. Animal! loxlclty Studies Most Useful for Hazard
Assessments 8-2
8.2. FACTORS INFLUENCING HEALTH HAZARD ASSESSMENT 8-6
8.2.1. Exposure 8-6
8.2.2. Lowest-Observed-Effect Level 8-6
8.2.3. Carcinogenic!ty . 8-7
8.3. REGULATIONS AND STANDARDS 8-8
8.3.1. Occupational Standards 8-8
8.3.2. Transportation Regulations 8-8
8.3.3. Solid Waste Regulations . '. . 8-9
8.3.4. Food Tolerances 8-10
8.3.5. Water Regulations 8-10
8.3.6. Air Regulations 8-10
8.3.7. Other Regulations 8-11
9. REFERENCES g_1
APPENDIX: Toxicity Table for Hexachlorocyclopentadiene A-l
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LIST OF TABLES
No.
3-1
3-2
3-3
3-4
4-1
4-2
4-3
5-1
5-2
5-3
5-4
6-1
6-2
fj-T
Title
cK^r-oj-farTc-firc r\f fhia Pnrpn^k T Pnllprtinn Svs ism. * *
HEX Content in the Effluent Stream of the Memphis North
Area Air Samples Collected at the Memphis North Treatment
Plant 1Q7H
Concentrations of Selected Organic Compounds in Influent
Summary of Constants Used in the Exposure Analysis
Summary of Results of Computer Simulation of the Fate and
Transport of Hexachlorocyclopentadiene in Four Typical
Microbial Degradation of HEX During 14-Day Exposure in a
Relative Distribution of HEX and Its Degradation Products .
Acute Toxicity Data for Freshwater Species Exposed to HEX .
Acute Toxicity Data on Marine Organisms Exposed to HEX. . .
Effects of 28 Days Exposure of Mysid Shrimp, Mvsidopsis
hahia. to HEX
Page
. 3-2
. 3-3
. 3-9
. 3-10
. 4-3
. 4-5
. 4-7
. 5-6
. 5-7
. 5-17
. 5-28
. 6-2
. 6-6
. 6-8
7-1
7-2
7-3
Disposition of Radioactivity Expressed as Percentage
of Administered Dose from 14C-HEX in Rats Dosed by
Various Routes
Fate of Radiocarbon Following Oral, Inhalation and
Intravenous Exposure to 14C-HEX in Rats Expressed as
Percentage of Administered Dose
Distribution of HEX Equivalents in Tissues and Excreta
of Rats 72 Hours After Oral, Inhalation and Intravenous
Exposure to 14C-HEX
7-6
7-7
7-8
xi
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Page
. No.._ Title
7-4 Acute "loxicity of HEX 7_12
7-5 Subchronic ToxicHy of HEX 7-17
7-6 lexicological Parameters for Mice and Rats Administered
Technical Grade HEX for 91 Days 7-18
7-7 Memphis HEX Monitoring Summary 7-30
7-8 Marshall HEX Monitoring Summary 7-32
7-9 Symptoms of 145 Wastewater Treatment Plant Employees
Exposed to HEX 7-35
7-10 Abnormalities for 18 of 97 Cleanup Workers at the Morris
Forman Treatment Plant 7-37
7-11 Overview of Individual Exposure - Symptomatology Corre-
lations at the Morris Forman Treatment Plant. . 7-38
7-12 Hepatic Profile Comparison of Hardeman County: Exposed
Group (November 1978) and Control Group 7-40
8-1 Oral Toxicity Data for Threshold Estimates 8-3
8-2 Inhalation Toxicity Data for Threshold Estimates 8-4
xii
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No.
1
3-1
5-1
5-2
5-3
LIST OF FIGURES
Title
Page
Structure Diagram of Hexachlorocyclopentadiene xlv
Synthesis of Chlorinated Pesticides from
Hexachlorocyclopentadiene ...... 3-5
Proposed Pathway of Aqueous HEX Phototransformation 5-5
Rate of Biodegradation of 14C-H£X to 14C02 • • 5-11
Persistence of Nonpolar 14C when 14C-HEX is Applied to
Unaltered and Altered Soils 5-15
xiii
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FIGURE 1
Structure Diagram of Hexachlorocyclopentadlene
x1v
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1. INTRODUCTION
Hexachlorocyclopentadiene (HEX) is an unsaturated, highly reactive,
chlorinated cyclic hydrocarbon of low water solubility. HEX is a chemical
intermediate in the manufacture of chlorinated pesticides and flame retar-
dants with essentially no end uses of its own. The major source of environ-
mental contamination by HEX is the aqueous discharge from production facil-
ities, with small concentrations present as contaminants in commercial prod-
ucts made from it. However, HEX is not frequently found in the environment
and, even when present, it is rapidly degraded. In view of this and 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 (an illegal act) in 1977 in Louisville, KY,
and the cleanup of a large waste disposal site in Michigan in 1983, have
brought this chemical to the forefront of environmental news.
Hexachlorocyclopentadiene is not readily absorbed via epithelial tissues
because it is highly reactive, especially with the contents of the gastro-
intestinal tract. HEX is moderately toxic when given orally, but has been
estimated to be 100 times more toxic when inhaled. The data base for
chronic toxicity of HEX is very limited. A chronic inhalation bioassay is
being conducted by the National Toxicology Program (NTP) and may provide
data regarding any carci- nogenic potential of HEX.
Several literature reviews on the health and environmental effects of
HEX are available and include the following: Equitable Environmental
Health, Inc. (1976), National Academy of Sciences (NAS, 1978), Bell et al.
(1978) and U.S. EPA (1980c). Although each of these reports is different in
scope and emphasis, a large amount of, the scientific knowledge about HEX is
1-1
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Included in these documents. To avoid unnecessary duplication, previously
reviewed material found In these documents will not be considered at great
length, except when it impinges directly upon present critical considera-
tions. The information presented in this document is current through 1984,
and contains a critical evaluation of some data which were not available at
the publication time of the previously mentioned documents.
One final note of caution for the interested reader. Some of the
reports reviewed in this document are unpublished laboratory reports. The
Agency has received copies of these documents from various sources under the
Toxic Substances Control Act (TSCA) reporting provisions. It is not the
purpose of this document to judge the quality or validity of these reports
unless there are peer review studies to compare results. The overall pur-
pose of this document is to present the research data in order to assist the
regulatory office of the Agency in developing a proposal concerning the
decision to regulate HEX under Section 112 of the Clean Air Act.
The subject matter contained in this health assessment document has been
reviewed by many Agency scientists, as well as scientists from private
corporations, other government Agencies, and the general public. A previous
draft of this publication was available for public comment. This final
publication Incorporates all of these comments and responses, as well as new
literature published since the previous draft.
1-2
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2. SUMMARY, CONCLUSIONS AND RESEARCH NEEDS
2.1. SUMMARY
2.1.1. Properties, Production and Uses. Hexachlorocyclopentadiene (HEX,
C-56) is a dense pale-yellow or greenish-yellow, nonflammable liquid with a
unique, pungent odor. HEX has a molecular weight of 272.79 and low water
solubility. It is highly reactive and undergoes addition, substitution and
Diels-Alder reactions.
Hexachlorocyclopentadiene is produced by only one company in the United
States, Velsicol Chemical Corporation. Production data are considered
proprietary; however, it has been estimated that between 8 and 15 million
pounds/year are produced. HEX has been used as an intermediate in the
production of many pesticides; however, this use has been limited by
restrictions on the production of certain organochlorine pesticides. HEX is
also used in the manufacture of flame retardants, resins and dyes.
2.1.2. Sources, Environmental Levels, Transport and Fate. HEX is
released into the environment at low levels during its manufacture and
during the manufacture of products requiring HEX. HEX can enter the
environment in low levels as an impurity and contaminant in some of the
products using HEX as an intermediate. There are only limited monitoring
data available concerning the environmental levels of HEX. The available
information suggests that HEX will be present mainly in the aquatic compart-
ment and associated with bottom sediments and organic matter, with the
exception of areas where land disposal has taken place. HEX readily adsorbs
to most soil particles.
Releases of HEX to the atmosphere can result from the production and use
of HEX, disposal of waste streams containing HEX, or from products contami-
nated with HEX. The total annual estimated release of HEX to the environ-
2-1
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ment is 11.9 Mg (12.5 tons). Because of Us physical and chemical charac-
teristics, only a small amount of this total can be expected to persist.
The fate and transport of HtX in the atmosphere, considering available
information, suggests that the compound has a tropospheric residence time
(the time required for the concentration to be reduced to 1/e) of ~5 hours.
However, atmospheric transport of HEX from an area of stored wastes and from
wet wells during treatment of industrial wastes has been demonstrated.
In water, HEX may undergo photolysis, hydrolysis and biodegradation. In
shallow water, HEX has a photolytic half-life of <1 hour. In deeper water
where photolysis is precluded, the hydrolytic half-life of HEX is several
days, while biodegradation is predicted to occur more slowly. HEX is known
to volatilize from water, but this is Influenced by turbulence and adsorp-
tion onto sediments.
HEX should be relatively immobile in soil based on its low water solu-
bility. Volatilization, which is likely to occur primarily at the soil
surface, is inversely related to the organic matter levels and water-holding
capacity of the soil. Chemical hydrolysis and microbial metabolism are
expected to reduce levels of HEX in soils.
Using model ecpsystem data, the bloconcentration/bioaccumulation/blomag-
n1f1cat1on potential of HEX would theoretically be expected to be substan-
tial based on its high lipophilicity [log octanol/water partition coeffi-
cient (log P)]. .However, experimental evidence does not support this
theory. Bioaccumulation factors derived from a short-term model ecosystem
study appear to indicate a moderate accumulation potential in algae, snails,
mosquito larvae, and mosquito fish. In addition, studies with laboratory
animals have shown that HEX is excreted rapidly within the first few hours
2-2
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after oral dosing, with little being retained in the body. The -Compound.did
not biomagnify substantially from algae to snails or from mosquito larvae'to
fish. In addition, steady-state bioconcentration factors, measured in 30 to
32-day flow-through exposures, were only 29 and <11 in fish exposed to con-
stant HEX levels of 20.9 yg/S. and 9.1 ppb, respectively. Therefore, it
would appear from these data that HEX does not persist or accumulate in any
large amounts. The degradation products of HEX have not been identified.
2.1.3. Aquatic Life, Vegetation and Wildlife. Low concentrations of HEX
have been shown to be toxic to aquatic life. Lethality in acute (48- to
96-hour) exposures has been observed in both freshwater and saltwater crus-
taceans and fish at nominal concentrations of 32-180 vg/i in static
exposure systems in which the water was not renewed during the test. In the
only studies using flowing water and measured HEX concentrations, identical
96-hour LCrn values of 7 lag/8, were obtained for freshwater fish and
50
saltwater shrimp. Chronic tests with the latter two species showed adverse
effects at levels as low as 7.3 and 0.70 vg/l. respectively.
Seven-day static tests with marine algae showed median reduction of
growth {EC,J at nominal concentrations ranging from 3.5-100 vg/l,
3 \J
depending on the species.
In aqueous media, HEX is toxic to many microorganisms at nominal concen-
trations of 0.2-10 mg/9., or levels substantially higher than those needed
to kill most aquatic animals or plants. Some microorganisms are able to
withstand HEX exposures as high as 1000 mg/fc. HEX appears to be less
toxic to microorganisms in soil than in aquatic media, probably due to
adsorption of HEX on the soil matrix.
Sufficient information is not available to determine the effects of HEX
exposure on terrestrial vegetation or wildlife, although data from
2-3
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laboratory studies summarized In the following sections could be used to
estimate effects on mammals in the wild.
2.1.4. Pharmacokinetics, Toxicology, Exposure and Health Effects. The
absorption of unchanged HEX is lessened because of its reactivity with
membranes and tissues, and especially with the contents of the gastrointes-
tinal tract. HEX is considered a primary irritant, extremely toxic by
Inhalation, and moderately toxic by oral ingestion. Radiolabeled 1/JC-HEX
is retained by the kidneys and liver of animals after oral or inhalation
dosing; after inhalation, the trachea and lungs also retain radiolabeled
material. Absorbed HEX is metabolized and rapidly excreted, predominantly
in the urine and feces with <1% of the HEX found in expired air. Following
inhalation or intravenous injection no unchanged HEX is excreted, and the
fecal and urinary metabolites have been isolated, but not identified. The
failure to identify these metabolites has been one of the mysteries concern-
ing HEX. Without this information and quantitative data, it is difficult to
assess the total effect of inhaled HEX in humans.
The acute inhalation lethal concentration {LC5 ) of 1.6 and 3.5 ppm in
male and female rats, respectively, has been demonstrated. Although there
are some interspecies 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 then dermal administration being
less toxic routes. Systemic effects of acute exposure include degenerative
changes in the lungs, liver, kidneys and adrenal glands.
Subchronic oral dosing of rats (38 mg/kg/day) and mice (75 mg/kg/day)
for 91 days produced nephrosis and inflammation and hyperplasia of the fore-
stomach. No overt signs were noted when mice or rats were exposed by
inhalation at 0.2 ppm of HEX (6 hours/day, 5 days/week) for 14 weeks.
2-t
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However, inhalation exposure of rats at 0.5 ppm for 30 weeks caused
degenerative changes in the liver, respiratory tract and kidneys. I_n yUrjo
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 effects of exposure in humans. Isolated
events have occurred which show HEX to cause severe irritation of the eyes,
nose, throat and lungs. Human exposures have included short-term irrita-
tions, with recovery after cessation of exposure. There were no statisti-
cally significant differences in liver enzymes between exposed and control
groups. The long-term health effects of continuous low-level exposure
and/or intermittent acute exposure in man are not known. Waste handlers and
sewage workers have been shown to be occupations at risk.
The data base is neither extensive nor adequate for assessing the car-
cinogenicity of HEX. The National Toxicology Program (NTP) has recently
completed a subchronic animal study and will begin a lifetime animal inhala-
tion bioassay using both rats and mice. Several epidemiologic studies were
cited in the literature; however, no increased incidences of neoplasms at
any site were reported which could be related to HEX. Accordingly, Velsicol
Chemical Corporation has on-going programs and follow-up studies in order to
study the long-term effects of HEX exposure. A final judgment of carcino-
genicity will have to be deferred until the results of the NTP bioassay are
available. Using the International Agency for Research on Cancer (IARC)
criteria, the available evidence matches the overall Group 3 category.
According to the IARC criteria, Group 3 indicates that because of major
qualitative or quantitative limitations, the studies cannot be interpreted
as showing either the presence or absence of a carcinogenic effect.
2-5
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2.2. CONCLUSIONS
This document presents the current scientific data base concerning HEX.
HEX 1s not found frequently in the environment because its emissions are low
(-12 megagrams per year) and because it rapidly degrades into other sub-
stances of unknown character. This document summarizes the known health
effects from exposure to HEX. At expected ambient concentrations, there
have been no known long-term adverse health effects. The only known effect
of HEX that might occur at current and projected exposures is odor recogni-
tion. The odor recognition threshold concentration for HEX, which is not
well-established, may be exceeded in the vicinity of the sources listed in
this chapter. At this time, available information is not sufficient from
either animal or,human data to determine the carcinogenic potential of HEX.
In addition, as listed below, there are still uncertainties in the data base
that affect the interpretation of available data. Once these voids in data
are filled, we will have a better understanding of HEX and its effect on
humans and the environment.
2.3. RESEARCH NEEDS
In the development of this document and previous drafts, there have been
many comments on the need to complete certain studies. This data would
refine the known information and give scientists a better understanding of
the effects of HEX and its properties. Because studies on the carcinogenic
potential of HEX are being done by NTP, additional research for carcinogen-
1c1ty does not appear to be warranted. In its response to the Interagency
Testing Committee regarding section 4 of TSCA, the Agency stated that, given
current manufacture, distribution in commerce, use or disposal of HEX, there
was no need to acquire more test data to make regulatory decisions under
TSCA, However, as the result of this document and its review, the following
2-6
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research areas could yield data that would provide information on the spe-
cific nature of this compound, as well as help resolve some remaining
unknowns.
An unresolved issue at the peer review workshop concerned the matter in
which external factors influence the vapor pressure of HEX. Consider-
able discussion resulted in the recommendation that a study of vapor
pressure should be included as a priority item in future research.
There is a need for a thorough metabolism study in which the metabolites
are isolated and identified.
Monitoring and study of groups exposed to continuous low levels of HEX
is warranted. Monitoring data are needed to derive estimates of expo-
sure, especially for those areas near production and formulation
facilities.
Further studies are needed to determine the fate of HEX in the
environment.
Teratogenicity studies should be conducted using various routes of
exposure, with emphasis on the inhalation route.
There is a need to measure the odor recognition threshold of HEX. One
study has been performed; however, this study was not peer reviewed.
2-7
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3. PHYSICAL AND CHEMICAL PROPERTIES/ANALYTICAL METHODOLOGY
3.1. SYNONYMS, TRADE NAMES AND IDENTIFICATION
Hexachlorocyclopentadiene (HEX) is the most commonly used name for the
compound that is designated 1,2,3,4,5,5'-hexachloro-l,3-cyclopentadiene by
the International Union of Pure and Applied Chemistry (IUPAC) system.
Table 3-1 cites the IUPAC name and synonyms, identification numbers and
molecular and structural formulas of HEX.
3.2. PHYSICAL AND CHEMICAL PROPERTIES
3.2.1. Physical Properties. Hexachlorocyclopentadiene is a nonflammable
liquid with a characteristic pungent, musty odor; the pure compound is light
lemon-yellow. Impure HEX may have a greenish tinge (Stevens, 1979). It has
a molecular weight of 272.79 and its molecular formula is C5C16. Hexa-
chlorocyclopentadiene (98%) is a dense liquid (sp. gr. 1.7019 at 25°C) with
low solubility in water (0.805-2.1 mg/st at 25°C). A detailed list of
physical properties is presented in Table 3-2. The compound is strongly
adsorbed by soil colloids. It volatilizes rapidly from water (Atallah et
al., 1980). According to the Handbook of Chemistry and Physics (Weast and
Astle, 1980), the ultraviolet visible x in heptane is 323 nm with a
II lu A
log (molar absorptivity) of 3.2. This absorption band reaches into the vis-
ible spectrum, as evidenced by the yellow color of HEX. Facile carbon-
chlorine bond scission might be expected in sunlight or under fluorescent
light. The IR spectrum has characteristic absorptions at 6.2, 8.1, 8.4,
8.8, 12.4, 14.1 and 14.7 pm. The mass spectrum of HEX shows a weak molec-
ular ion (M) at M/e 270, but a very intense (M-35) ion making this latter
ion suitable for sensitive specific ion monitoring.
3-1
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TABLE 3-1
Identity of Hexachlorocyclopentadiene
Identifying Characteristic
Name/Number/Structure
IUPAC Name:
Trade Names:
Synonyms:
CAS Number
CIS Accession Number:
Molecular Formula:
Molecular Structure:
1,2,3,4,5,5'-Hexachloro-l,3-cyclopen tadiene
C56; MRS 1655; Graphlox
Hexach1 orocyclopen tadiene
Perch1 orocyclopen tadiene
HEX
HCPD
HCCP
HCCPD
C-56
HRS 1655
Graphlox
77-47-4
7800117
C5C16
Cl
Cl
ClI fcl
cr*^
3-2
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TABLE 3-?
Physical Properties of Hexachlorocyclopentadiene
Property
Value/Description
Reference
Molecular Weight:
Physical Form (25°C)
Odor:
Electronic Absorption Max-
imum [(in 50% acetoni-
trile-water)]
Solubility (25°C)
Water (mg/9.):
Organic Solvents:
Vapor Density (air = 1}
Vapor Pressure
(mmHg, °C):
Specific Gravity:
Melting Point (°C):
Boiling Point (°C):
Octanol/Water Partition
Coefficient (log P)
(measured):
(estimated):
Latent Heat of Vaporiza-
tion
Henry's Law Constant
(atm-mVmole)
272.79 Stevens, 1979
Pale yellow liquid Hawley, 1977; Irish, 1963
Pungent Hawley, 1977; Irish, 1963
322 rirn
2.1
0.805
1.8 (28°C)
Miscible (Hexane)
9.4
0.08 (25°C)
0.975 (62°C)
1.717 (T5°C)
1.710 (20°C)
1.7019 (25°C)
-9.6
-11.34
239 @ 753 mm Hg
234
5.04+0.04
5.51
176.6 J/g
2.7xlO~2
Wolfe et al., 1982
Dal Monte and Yu, 1977
Lu et al., 1975
Wolfe et al., 1982
Bell et al., 1978
Verschueren, 1977
Irish, 1963
Stevens, 1979
Hawley, 1977
Stevens, 1979
Weast and Astle, 1980
Hawley, 1977
Stevens, 1979
Hawley, 1977; Stevens, 1979
Irish, 1963
Wolfe et al., 1982
Wolfe et al., 1982
Stevens, 1979
Atallah et al., 1980;
Wolfe et al., 1982
3-3
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3.2.2. Chemical Properties. Commercial HEX has various purities depend-
ing upon the method of synthesis. HEX made by chlorination of cyclopenta-
dlene by alkaline hypochlorite at 40°C, followed by fractional distillation,
1s only 75% pure, and contains many lower chlorinated cyclopentadienes.
Purities >90% have been (obtained by thermal dechlorination of octachloro-
cyclopentene at 470-480°C: (Stevens, 1979). The current specification for
HEX produced by Velsicol Chemical Corporation at the Memphis, IN plant,
which is used internally and sold to other users, has a 97% minimum purity
{Velsicol Chemical Corporation, 1984).
If moisture 1s excluded, HEX can be stored without harming the product
or its containers. Storage containers should not have iron in their inner
linings (Stevens, 1979).
Hexachlorocyclopentadiene is a highly reactive diene that readily under-
goes addition and substitution reactions and also participates in dels-
Alder reactions (Ungnade and McBee, 1958). The products of the Diels-Alder
reaction of HEX with a compound containing a non-conjugated double bond are
generally 1:1 adducts containing a hexachlorobicyclo(2,2,l )heptene struc-
ture; the monoene derived part of the adduct is nearly always in the endo-
position, rather than the exo-position (Stevens, 1979). Figure 3-1 illus-
trates synthetic pathways to various chlorinated pesticides for which HEX is
a precursor. Flame retardant chemicals for which HEX is a precursor include
chlorendlc acid, chlorendic anhydride and Dechlorane Plus (Stevens, 1979).
Two excellent early reviews of the chemistry of HEX were published by
Roberts (1958) and Ungnade and McBee (1958). Look (1974) reviewed the for-
mation of HEX adducts of aromatic compounds and the by-products of the Diels
Alder reaction.
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3.3. ANALYTICAL METHODOLOGY
3.3.1. A1r.
3.3.1.1. SAMPLING — The techniques used to collect samples of HEX
vapor 1n air involve the adsorption and concentration of the vapors In
liquid-filled impingers or solid sorbent-packed cartridges.
Whitmore et al. (1977) pumped airborne vapors through a miniature glass
impinger tube containing hexane or benzene and through a solid sorbent
packed (Chromosorb® 102) tube. Sampling efficiency was 97% with hexane
and ~100% with benzene, the sampling efficiency for the solid sorbent tube
was -100%. The sensitivity of the liquid impinger system was found to be <1
ppb in ambient air.
Kominsky and Wisseman (1978) collected HEX vapor on Chromosorb® 102
(20/40 mesh) sorbent previously cleaned by extraction with 1:1 acetone/
methanol. The extraction removed interfering compounds. Ihe sorbent was
packed Into a front 100-mg and a back 50-mg section separated by a 2 mm
polyurethane plug In a glass tube, 7 cm long and 4 mm i.d. The samplers
were collected using battery powered vacuum pumps operating at 0.05 or 0.20
S./m1nute. HEX was desorbed with carbon disulfide (68% efficiency) and
analyzed by gas chromatography-flame ionization detection (Neumeister and
Kurimo, 1978).
In studying the pyrolysis products of endosulfan, Chopra et al. (1978)
collected the vapors of endosulfan-treated tobacco smoke in a cold trap con-
taining pentane cooled to 0 and -80°C. The pentane extract was then pre-
pared for gas chromatographic (GC) analysis; HEX was qualitatively deter-
mined to be one of the pyrolysis products formed.
3-6
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Under contract with NIOSH, Boyd et al. (1981) and Dillon (1980) of the
Southern Research Institute developed and validated sampling and analytical
methods for air samples containing HEX. Methods were reliable below the
8-hour time-weighted-average (TWA) Threshold Limit Value (TLV) of 0.1
mg/m3 recommended by the American Conference of Governmental Industrial
Hygienists (ACGIH).
The developed NIOSH method, P & CAM 308 (NIOSH, 1979) utilized adsorp-
tion on Porapak® T (80/100 mesh), desorption with hexane (100% for 30 ng
of HEX on 50-100 mg adsorbent), and then analysis by GC-63Ni electron
capture detection. The solid sorbent was cleaned by soxhlet extraction with
4:1 (v/v) acetone/methanol (4 hours), and hexane (4 hours), and was allowed
to dry under vacuum at 50-70°C overnight before cooling in a desiccator.
The pyrex sampling tubes (7 cm long, 6 mm o.d., 4 mm i.d.) contained a front
75 mg layer of sorbent and a 25 mg backup section. Each section was held in
place with two silylated glass wool plugs. A 5 mm long airspace was neces-
sary between the front and backup sections. A battery operated sampling
pump drawing air at 0.05 and 2.0 si/minute was utilized for personal
sampling of workers. The lowest analytically quantifiable level was 25 ng
HEX/sorbent sample, assuming 1 ms. of hexane-desorbing solvent and a 1 hour
desorption time by ultrasonification. The upper limit of the method was
2500 ng/sorbent sample. The method was validated for air HEX concentra-
tions between 13 and 865 pg/m3 at 25-28°C and a relative humidity of
>90%. .
3.3.1.2. ANALYSIS— Gas chromatography is the preferred method for
analyzing HEX in air using either flame ionization collection or electron
capture detection (e.g., Chopra et al., 1978; Neumeister and Kurimo, 1978;
Whitmore et al., 1977; NIOSH, 1979). Gas chromatography/mass spectroscopy
(GC/MS) is necessary for confirmation (Eichler, 1978).
3-7
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Several sorbent materials were evaluated for collection of HEX vapor:
Amberllte® XAD-2 (20/50 mesh), Porapak® R (50/80 mesh), Ambersorb®
XE-340 (20/50 mesh), Chromosorb® 104 (60/80 mesh), lenax-GC® (35760
mesh), Porapak® T (80/100 mesh) and Porapak® 1 (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 3-3 enumerates additional factors related to the Porapak® T
collection system.
Gas chromatography with electron capture detection (ECD) was determined
to be the most sensitive analytical technique. For HEX the chromatographic
response was stated to be a linear and reproducible function of HEX concen-
tration In the range of -5-142 ng/ms. (25-710 pg injected) with a correla-
tion coefficient of 0.9993 for peak height measurement. The optimized
operating conditions for this method are shown in Table 3-4.
Validation tests were conducted according to NIOSH guidelines. The
accuracy and precision of the overall sampling and analytical procedure for
HEX were evaluated in the concentration range of -13-865 pg/m3. The
lowest analytically quantifiable level (LAQL) of HEX was determined to be
25 ng/sorbent tube. This level represents the smallest amount of HEX that
can be determined with a recovery of >80% and a relative standard .deviation
(RSD) of <10%. The desorption efficiency of 100% was determined by averag-
ing the levels ranging from near the LAQL of 25 ng to 1000 times the LAQL.
3.3.2. Water. Since HEX is sensitive to light in both organic and aque-
ous solutions, the water samples, extracts and standard HEX solutions must
be protected from light. The rate of degradation is dependent upon the
intensity and wavelength with the half-life of HEX being -7 days when the
3-8
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TABLE 3-3
Characteristics of the Porapak® T Collection System3
Characteristic
HEX Type/Value
Sorbent material
Breakthrough timec
Breakthrough volume0
Tube capacity0
Average desorption
efficiency of indicated
quantity of analyte
Sorbent tube
configuration1^
Extraction solvent
Porapak® Tb
(80/100 mesh)
>8 hour (0.2 si/minute)
>100 9.
>100 g
0.94 (27.4 ng)
75 mg sorbing layer,
25 mg backup layer
Hexane
aAdapted from Boyd et al., 1981
blhis 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 >90%. The concentration of the
analyte in the generator effluent was 1 mg/m3 of HEX.
sorbent tubes were Pyrex (7 cm long by 6 mm o.d. and 4 mm i.d.).
Silanized glass wool plugs separated the sections.
3-9
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TABLE 3-4
Optimized GC Analytical Procedure for HEXa»b
Characteristic
Type/Value
Detector
Column
E lectron capture
3% OV-1 on Gas-Chrom Q
(100/120 mesh) In glass
(4 mm i.d. by 2 m)
OPERATING CONDITIONS
Carrier gas
(20 ms./m1nute)
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 ml/minute)
Hexane
aAdapted from Boyd et al., 1981
bA Hewlett-Packard 5750A gas chromatograph was used.
cThe Injection volume was 5 yfc of sample and 1 p8. of solvent flush.
3-10
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solution was exposed to ordinary laboratory lighting conditions (BenoU and
Williams, 1981). Storing the HEX-containing solutions in amber or red (low
actinic) colored glassware is recommended for adequate protection (Benoit
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/S. vs. 0.5 ng/9. 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 qualita-
tively because of the low detection limit (Benoit and Williams, 1981).
3.3.3. Soil.
3.3.3.1. SAMPLING — In the method described by DeLeon et al.
(1980a), samples were taken from vertical borings 30 feet deep using the
split-spoon method. The samples were then placed in jars and sealed with
Teflon®-lined screw caps. During shipment, the samples were maintained at
6-10°C. Upon their arrival at the analysis site, they were maintained at
-20°C until required for analysis.
3.3.3.2. ANAYLSIS — DeLeon et al. (1980a) developed a method for
determining volatile and semivolatile organochlorlne compounds in soil and
chemical waste disposal 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 electron
capture detection; GC/MS is used for confirmation of the presence of the
chlorocarbons. The method has a lower detection limit of 10
3-11
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Spiked samples of soil were used to test the recovery and reproducibil-
Uy of the procedure. When a soil sample was spiked with a 10 yg/g con-
centration of HEX, the recovery was 59.8% (S.D. 6.1); at 100 pg/g, 95.9%
(S.D. 15.9); at 300 yg/g, 90.?% (S.D. 4.1). However, as the authors
state, some modifications may be necessary for analysis of the more volatile
one to four carbon chlorinated compounds, since some compounds may be lost
1n the concentration step before GC analysis. Of the 11 compounds tested in
three different concentration levels by the authors, the 100 ug/g HEX
sample had the highest standard deviation of all compounds. Over 100
chemical waste disposal site and soil samples have been evaluated by this
method. In this study, HEX was not detected in three typical samples each
taken from a different location within and around a chemical waste disposal
site (DeLeon et al., 1980a).
3.4. BIOLOGICAL MEDIA
3.4.1. Sampling. A method to determine levels of HEX in blood and urine
has been described by DeLeon et al. (1980b)\ This method involves isolation
of the compound from the blood or urine sample by liquid-liquid extraction,
GC analysis with electron capture detection and confirmation by GC/MS. Mean
recoveries of 28.8 and 54.5% were reported for blood samples containing 50
and 500 ng/mS., respectively; for urine, mean recoveries of 35.0 and 51.8%
were reported for samples containing 10 and 200 ng/ma, respectively. The
best recoveries were obtained in the study through the use of a toluene-
acetonitrlle extraction combination for blood assays, and petroleum ether
extraction for urine assays. The authors concluded' that this method is
useful for the detection and identification of nanogram quantities of HEX,
with low detection limits of 50 ng/ma. for blood and 10 ng/mfc for urine.
3-12
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Studies by Velsicol Chemical Corporation have shown that .up to 30% of the
HEX can be lost If the extracts are concentrated to 0.1 m. Quantitative
recovery was possible only for volumes of concentrate >0.5 ma. This
limits the sensitivity of the method. However, the method may offer a
sensitive means of monitoring occupational exposure.
3.4.2. Analysis. Velsicol Chemical Corporation (1979) has developed
three analytical methods which have been used for urine, fish fillet, beef
liver, beef skeletal muscle, beef adipose tissue, beef kidney, chicken
liver, chicken skeletal muscle and chicken adipose tissue. The respective
recoveries were: 80+10 (1-50 ppb), 81+1, 69+_4, 88+2, 86+5, 71+3, 55+9, 76+4
and 85+2%. The level of fortification for the tissue samples was 10 ppb.
For urine, up to 31% HEX could be degraded when the fortified urine sample
was stored overnight in 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, centrlfuged, and the pentane
layer transferred into a separatory funnel. The residues were then parti-
tioned into acetonitrile (3 times), water diluent added to the acetonitrile,
and then back-extracted with pentane. The pentane extract was treated with
concentrated sulfuric acid and then water, and concentrated to ~3 ml.
Upon dilution to 10 ms, with hexane, the solution was treated with a 1:1
concentrated sulfuric add/fuming sulfuric acid solution, water, and a 9:1
mixture (solid) of sodium sulfate/sodium carbonate. Packed columns (3% OV-1
on Gas Chrom Q-100/120 mesh-in 2 m x 2 mm i.d. glass column) or capillary
columns (30 m x 0.25 mm SE-30 WCOT) can be used for GC using a ^Mi-elec-
tron capture detector.
3-13
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4. PRODUCTION, USE, SOURCES AND AMBIENT LEVELS
4.1. PRODUCTION
Because there is only one producer of HEX, production statistics are
considered confidential business information (CBI) and are not available to
the general public. Production estimates for HEX, based on the manufacture
of chlorinated cyclodiene pesticides 1n the early 1970s, were -50 million
pounds per year (Lu et a!., 1975). Following restrictions in the use of
pesticides produced from HEX, production estimates were lowered to a range
of 8-15 million pounds per year (U.S. EPA, 1977). In a report prepared for
the U.S. EPA, Hunt and Brooks (1984) estimated 8300 Hg (9130 tons) of HEX
were produced in 1983. Technical grade HEX usually contains other chemicals
as contaminants of manufacture (e.g., hexachlorobenzene and octachlorocyclo-
pentene. The nature and levels of contaminants will vary with the method of
production.
4.2. USE
HEX is the key intermediate 1n the manufacture of some chlorinated
cyclodiene pesticides (see Figure 3-1). These include heptachlor, chlor-
dane, aldrin, dieldrin, endrin, mirex, PENTAC and endosulfan. Another major
use of HEX is 1n the manufacture of flame retardants such as chlorendlc
anhydride and Dechlorane Plus. HEX Is also used to a lesser extent in the
manufacture of resins and dyes (U.S. EPA, 1980c), and has been used pre-
viously as a general biocide (Cole, 1954). Currently, HEX is produced at
two locations: Memphis, TN and Marshall, IL. All of the HEX produced at
the Illinois plant is used solely for the production of chlordane, and 1s
not sold or distributed, while that produced at the Memphis plant is used to
produce heptachlor, endrin and the fire-retardant chlorendic anhydride. The
HEX produced at the Memphis plant is the same as that sold in commerce to
users of HEX (Velsicol Chemical Corporation, 1984).
4-1
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4.3. SOURCES
HEX is released into the environment in low levels during its manu-
facture and during the manufacture of products requiring HEX (U.S. EPA,
1980c). It is also found as an impurity and a degradation product in com-
pounds manufactured from HEX (Spehar et al., 1977; Chopra et al., 1978).
Limited monitoring data from production sites indicated that HEX was present
at 18 mg/S. (on February 2, 1977) in the aqueous discharge from the Memphis
pesticide plant (U.S. EPA, 1980c). In the summer of 1977, shortly after
these readings, a new wastewater treatment plant began operation. Before
construction of the plant, wastewater flowed directly into the Mississippi
River or through one of ;its tributaries (Elia et al., 1983). Voluntary
Improvements in controlling the discharge from the Memphis plant resulted in
reported levels of 0.07 ppb HEX in the Mississippi River, near the mouth of
Wolf Creek (Velsicol Chemical Corp., 1978). HEX measurements were taken
from the effluent stream of the Memphis North Sewage Treatment Plant from
February to July 1982. Monthly averages ranging from 0.15-0.61 ppb were
reported. Table 4-1 summarizes these data (Levin, 1982b). In May 1977, HEX
was also detected at 0.17 mg/8. in the aqueous discharge and at 56 ppb in
air samples collected from a waste site in Montague, MI (U.S. EPA, 1980c).
Indoor air concentrations of HEX in some Tennessee houses with contaminated
groundwater supplies ranged from 0.06 to 0.10 pg/m3 (S. Clark et al.,
T982). HEX has also been identified in the soil and river sediments down-
stream from a Virginia manufacturing plant, even after pesticide production
was discontinued (U.S. EPA, 1980c). Under contract with the U.S. EPA, the
Radian Corporation prepared a report which presented the results of a pre-
liminary source assessment on HEX (Hunt and Brooks, 1984). Some of the
results of this study are presented in the following sections.
4-2
.
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TABLE 4-1
HEX Content 1n the Effluent Stream
of the Memphis North Sewage Treatment Plant, 1982^
Month
February
March
April
May
June
July
Number of
Samples Analyzed
19
15
30
31
29
30
High
0.80
0.60
3.04
0.54
0.57
1.80
HEX Level (ppb)
Low
NDC
NOC
NDC
NOC
NOC
NOC
Average*3
0.32
0.34
0.61
0.24
0.18
0.15
aSource: Levin, 1982b
bAverage of all samples taking all NO (not detected) values as zero.
cDetection limit is <0.01 ppb
4-3
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4.4. AMBIENT LEVELS
Published reports, environmental releases and physicochemlcal properties
r
of HEX imply that it will be present mainly in the aquatic compartment and
associated with bottom sediments and organic matter. Relatively much lower
concentrations will be found in the soil and air compartments.
4.4.1. A1r. Releases of HEX to the atmosphere can result from the pro-
duction and use of HEX, disposal of waste streams containing HEX or from
products contaminated with HEX (Hunt and Brooks, 1984). Data sent to the
U.S. EPA regarding emission levels from Velsicol plants indicate that quan-
tities of HEX are emitted into the air; however, these data are not con-
sidered public information. No data were found that reported ambient atmos-
pheric levels of HEX; however, the half-life of HEX in air is <5 hours
(Cupitt, 1980), which greatly reduces the potential for measurement. The
highest reported concentration of HEX measured in Tennessee homes was 0.10
yg/m3, while air levels at the Memphis North Treatment plant ranged as
high as 39 Mg/m3 (S. Clark et al., 1982; Elia et al., 1983). A list of
values is given in Table 4-2 for these air samples. This plant handles the
wastewater from a pesticide manufacturer five miles away. The only other
air monitoring was done on an abandoned waste site in Michigan where the
average HEX emission rate was 0.26 g/hr {^.05).
4.4.2. Water. Environmental monitoring data for HEX are available from a
number of sources. The bulk of the reported levels are contained within the
STORET data base (U.S. EPA, 1980b). The available monitoring data (STORET)
do not provide specific information about the sampling site and analytical
methodology. Additionally, the STORET data has not been verified and it is
not possible, therefore, to analyze the reported data critically.
4-4
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TABLE 4-2
Area Air Samples Collected at the
Memphis North Treatment Plan, 1978a
Concentration13, v>g/m3
Date
A. WET WELL
May
June
September
October
November
8. GRIT CHAMBER
May
June
July
September
October
November
Nc
3
2
2
1
1
3
7
2
4
1
1
HEX
0.03
18
8
15
39
0.03
1.9
0.03
0.03
0.04
12
HEX-BCH
219
278
25
2
7
4.1
6.5
, 0.5
0.5
1.2
2.6
HCBCHd
87
15
200
1
85
1.9
1.7
0.7
1.1
1.0
4.3
Chlordene
45
16
44
0.1
7.8
0.9
7.5
2.3
2.7
0.8
1.0
aSource: Elia et a!., 1983
DMean values of the number of samples, N, indicated
CN designates the number of samples collected
^Heptachlorobicycloheptene
4-5
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As previously mentioned, water samples of the influent wastewater were
taken at the Memphis North Treatment plant (lable 4-3). However, in the S.
Clark et al. (1982) study, HEX was not detected in the private wells of the
Tennessee homes.
Benoit and Williams (1981) sampled both raw and drinking waters from an
Ottawa water treatment plant. Using solvent extraction analysis with a
detection sensitivity of 50 ng/8. or using the XAD-2 resin extraction
method with a detection sensitivity of -0.5 ng/8. no HEX was detected in
the raw water, but levels ranging from 57-110 ng/a. were reported in the
finished drinking water, suggesting that HEX was Introduced Into the drink-
Ing water during the treatment process. The authors did not find the source
of the HEX, and are investigating their findings further (Benoit, 1983).
4.4.3. Food. HEX was qualitatively detected in fish samples taken from
water near a pesticide manufacturing in Michigan (Spehar et al., 1977);
however, none has been detected in fish samples taken from the waters near
the pesticide manufacturing plant in Memphis (Velsicol Chemical Corp., 1978;
Bennett, 1982). No information was available regarding HEX contamination of
other foods.
4.4.4. Soil. Ambient monitoring data for the terrestrial environment are
not available. However, it appears that these concentrations should be much
lower than concentrations present in the aquatic environment. Depositing of
HEX from the atmospheric (and aquatic) compartment into the terrestrial
environment is expected to be minimal. Similarly, direct release of HEX
Into the terrestrial environment (i.e., as an impurity in chlorinated pesti-
cides) should be decreasing with the possible exceptions of disposal at
waste sites, accidental spills and other illegal disposal methods.
4-6
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TABLE 4-3
Concentrations of Selected Organic Compounds
in Influent Wastewater at Memphis North Treatment Plant, 1978a
Concentration*, yg/s.
Date
June
August6
September
October -November
Nc
1
5
2
2
HEX
3
0.8
4
0.8
HEX-BCH
334
329
292
11
HCBCHd
57
115
668
17
Chlordene
87
216
58
32
aSource: Ella et al., 1983
bflean values for the number of samples Indicated
cNumber of samples
dheptachlorobicycloheptene
elhese values are furnished by the chemist at the North plant.
4-7
-------
4.5. RELATIVE SOURCE CONTRIBUTIONS
Available data are insufficient to derive relative source contributions.
After considering the available Information, the U.S. EPA has reported that
human exposure through the environment via air or water would be extremely
low except for workers and residents near manufacturing, shipping and waste
sites, and concluded that exposure was not considered significant or sub-
stantial (U.S. EPA, 1982).'
The only other estimation of relative source contributions 1s the Radian
report previously mentioned (Hunt and Brooks, 1984). The air releases from
manufacturing processes can be from vents on reactors, process and storage
tanks, and fugitive emissions. Hunt and Brooks (1984) estimated the total
quantity of HEX released from these sources to be 8.0 Mg (8.8 tons). In
addition, HEX can be emitted to the air from the Incineration and land-
filling of wastes containing HEX, with the best estimation being 1.0 Mg (1.1
tons). The other sources Include those listed In Section 4.4.4. and other
discharges to water bodies. The total annual estimated release of HEX to
the environment 1s 11.9 Mg (12,5 tons). These are only estimates because of
the limited data and are given only to provide the relative magnitude of HEX
emissions to the environment. For the reader who wishes to examine these
data and assumptions further, the Radian report should be reviewed In Us
entirety.
4.6. SUMMARY AND CONCLUSIONS
Measured ambient concentrations of HEX are available for aquatic com-
partments (U.S. EPA, 1980b). These Include freshwater and sediments of
streams, lakes and wells. Limited data are also available for estuaries and
oceans. Additional saltwater, as well as atmospheric and terrestrial mon-
i
itoring data, are needed to determine the ambient concentrations in these
compartments.
4-8
-------
Freshwater levels of HEX are estimated to range from 0-800
based upon non-verified STORE! data. Estimates for atmospheric concentra-
tions are not available in the literature, while estimates for HEX concen-
trations in soils are limited. To achieve proper conclusions concerning the
levels of HEX in the environment, careful monitoring and analysis must be
conducted. To date, this information is very limited.
Air HEX levels in areas near previous dump sites have been shown to be
high. High concentrations of HEX have been recorded in wastewater and in
two incidents have increased the ambient 'HEX levels inside treatment facil-
ities above the ACGIH time-weighted-average.
4-9
-------
-------
5. ENVIRONMENTAL FATE AND TRANSPORT
5.1. FATE
The evidence presented In this section Indicates that HEX is not persis-
tent in the air, water or soil. Photolysis, hydrolysis and biodegradation
have been shown to be the key processes influencing the environmental fate
of HEX. i
5.1.1. Air. Little relevant information is available to predict the fate
of HEX in air. Its tropospheric residence time was estimated by CupHl
(1980) to be ~5 hours based on estimated rates of reaction with hydroxyl
radicals and ozone. The respective reaction rates were theoretically calcu-
lated to be 59xlO~12 and 8xlO"18 cm3 molecule"5 sec"1. In estima-
ting the tropospheric residence time, or time for a quantity of HEX to be
reduced to 1/e (or -37%) of Its original value, it was assumed that the rate
constants calculated at room temperature for both reactions are valid in the
ambient atmosphere and that the background concentrations of hydroxyl radi-
cal and ozone are 106 and 1012 molecules cm"3, respectively. Atmos-
pheric photolysis of HEX was also rated as "probable", since HEX has a
chromophore that absorbs light in the solar spectral region, and is known to
photolyze in aqueous media (see Section 5.1.2.1.). No attempt was made to
estimate a rate for atmospheric photolysis. Cupitt (1980) listed the theo-
retical degradation products as phosgene (Cl CO), dlacylchlorides, ketones
and free chlorine (C1-) radical, all of which would be likely to react with
other elements and compounds.
Korte (1978) demonstrated the photomineralization of HEX (1.9 g) applied
to silica gel (400 g) after 4 days Irradiaton (x > 290) in an atmosphere
of pure oxygen. The mineralization products were chloride (Cl , 44.9%),
carbon dioxide (C0?, 48.3%), chlorine gas (Cl?, 5.4%) and carbon monox-
ide (CO, 1.2%).
5-1
-------
5.1.2. Water. In the event of release into shallow or flowing bodies of
water, degradatlve processes such as photolysis, hydrolysis and biodegrada-
tlon, as well as transport processes involving volatilization and other
physical loss mechanisms, are expected to be prominent in HEX dissipation.
In deeper, nonflowing bodies of water, hydrolysis and biodegradation may
become the predominant fate processes.
5.1.2.1. PHOTOTRANSFORMATION — Zepp et al. (1979) and Wolfe et al.
(1982) reported the results of U.S. EPA studies on the rate of HEX photo-
transformation in water. Under a variety of sunlight conditions, in both
distilled and natural waters of 1--4 cm depth, phototransformation half-life
was <10 minutes. Addition of natural sediments to distilled water contain-
ing HEX had little effect on phototransformation rate. These findings indi-
cated that the dominant mechanism of HEX phototransformation was direct
absorption of light by the chemical, rather than photosensitization reac-
tions involving other dissolved or suspended materials.
The direct photoreaction of HEX in water was also studied under con-
trolled conditions in the laboratory using a monochromatic light (313 nm)
isolated by filters from a mercury lamp. Phototransformation rate con-
stants, computed for the study location (Athens, 6A, 34°N latitude), agreed
with those observed in the sunlight experiments described above. Rate con-
stants were also computed for various times of day at 40°N latitude. The
near-surface phototransformation rate constant of HEX at this latitude on
cloudless days (averaged over both light and dark periods for a year) was
3.9 hr'1, which corresponds to a half-life of 10.7 minutes (Zepp et al.,
1979; Wolfe et al., 1982).
These researchers suggested that the primary phototransformation product
was the hydrated form of tetrachlorocyclopentadienone (CgCl.O, TCPD),
5-2
-------
although U was not isolated. Several chlorinated .photoproducts with higher
molecular weights than HEX were detected by GC/MS analysis of the reaction
mixture. Photolysis of HEX in methanol gave a product identified as the
dimethyl ketal of 1CPD (Wolfe et al., 1982). According to Zepp et al.
(1979), It is likely that TCPD exists predominantly 1n its hydrated form In
the aquatic environment. Ihe compound was not isolated, supposedly because
it rapidly dimerizes or reacts to form higher molecular weight products.
To the contrary, other research indicates that formation of higher
molecular weight products is a relatively minor pathway of phototransforma-
tion. Yu and Atallah (1977b) found that at a concentration of 2.2 mg/a in
water, uniformly labeled 14C-HEX was rapidly converted to water-soluble
products upon irradiation with light from a mercury-vapor lamp (light energy
40-48% ultraviolet, 40-43% visible, remainder infrared). In exposures of
0.5-5.0 hours, 46-53% of the radiolabel was recovered as water-soluble prod-
ucts (compared with 7% at initiation), whereas the amount recovered by
organic (petroleum ether) extraction decreased with increasing exposure
duration from 25 to 6% (compared with 66% at Initiation). No HEX was
detected among the photoproducts in the organic extraction.
But^ et al. (1982) also found that 14C-HEX, when dissolved and irradi-
ated as above, was rapidly photodegraded. Failure to detect HEX after 10
minutes, with a detection limit of 0.13% of the starting amount, suggested a
photolytic half-life under these conditions of <1.03 minutes, assuming
first-order kinetics. Reaction products were extracted and radioassayed.
After both 5- and 10-minute exposures, 44% of the recovered radioactivity
was in the water-soluble fraction (total percent recovery was not speci-
fied). Photoproducts were purified by thin-layer chromatography (TLC) and
identified by GC/MS. The authors concluded that pentachlorocyclopentenone
5-3
-------
O) was the major degradation intermediate, which subsequently
degraded to water-soluble products. Dimerization of pentachlorocyclopente-
none to hexachloroindenone (CgCl60) was thought to occur by hydrolysis,
rather than phototransformation, and to represent a minor pathway {Figure
5-1). Other high molecular weight compounds identified were believed to be
artifacts of the GC/MS analysis of pentachlorocyclopentenone. The research-
ers analyzed for mirex and kepone, but did not detect either after 5 hours
Irradiation (Butz et al., 1982).
The environmental fa^e and transport of HEX was modeled in four simulat-
ed aquatic ecosystems using the Exposure Analysis Modeling System for Toxic
Organic Pollutants in Aquatic Ecosystems (EXAMS) with experimentally derived
constants (Table 5-1) (Zepp et al., 1979; Wolfe et al., 1982). The four
ecosystems considered in the model included a 35 km x 100 m river segment; a
small eutrophic pond with a 31-day retention time in the water column; and
two lakes (35 ha), one eutrophic and the other a stratified oligotrophic
lake. Results indicate that in the river, export from the system and photo-
lysis were the dominant processes (Table 5-2). In the simulated pond and
both lake environments, photolysis was predicted to be the dominant process,
accounting for more than 80% of the HEX transformation occurring at each of
these sites. Although HEX is quite reactive, the recovery times (i.e., the
times needed to reduce steady-state concentration by five half-lives) in the
pond and lakes were predicted to be on the order of 2-3 months. This was
attributed to slow release of HEX from the bottom sediments where the photo-
lytic rate is retarded (Wolfe et al., 1982).
5.1.2.2. HYDROLYSIS — Studies of the hydrolysis of HEX Indicate that
at 25-30°C and in the environmental pH range of 5-9, a hydrolytic half-life
of -3-11 days is observed (Wolfe et al., 1982; Yu and Atallah, 1977a).
5-4
-------
CI
.Cl
I; Av, zs^C
w 1^. f** •••••••^^••i^^
;>< ^ -ci
•i f+\ «OH
• i \*i
\
Cl . ^0
1 CH
li ,^^
r* i '^^^^^^'^ r* i
^^^
Cl Cl
II """
^^^^^^^^ Water-koiuBie
Pt«otoprefluct>
42'C
Hydrolylit
-JHCl
-COCI,
Cl
M«,or
Miner
FIGURE 5-1
Proposed Pathway of Aqueous HEX Phototransformatlon
Source: Butz et al., 1982
5-5
-------
• TABLE 5-1
Summary of Constants Used 1n the Exposure
Analysis Modeling System (EXAMS) at 25°C in Water3
Constants
Symbols, Units
Values Used
Water solubility
Henry's law constant
Octanol/water partition
coefficient
Photolysis
Hydrolysis
Oxidation
Biodegradation
Ks, mg/s.
KH, atm mVmole
Kp, hr x
Kox, M'1 see"1
Kg, mil org"1 hr"1
1.8
2.7xlO~2
l.lxlO5
3.9
4.0xlO~ab
aAdapted from Wolfe et al., 1982
bExtrapolated to 25°C
°Estimated value (see Wolfe et al., 1982)
dThis is a maximum value based on the observation that there was no de-
tectable difference in the hydrolysis rate in either sterile or nonsterile
studies and measured organism numbers (plate counts).
5-6
-------
TABLE 5-2
Summary of Results of Computer Simulation of the Fate and Transport
of Hexachlorocyclopentadiene in Four Typical Aquatic Environments3
River
Pond
Eutrophic Oligotrophic
Lake Lake
Accumulation factor
Distribution (percent)
Water column
Sediment
Recovery timec (days)
Load reduction (percent)
by processes:
Hydrolysis
Oxidation
Photolysis
Biodegradation
(biolysis)
Volatilization
Export0"
5.4xl04
1.22
98.78
52
8.04
0.00
18.68
0.57
0,69
72.02
2.4
14
86
81
17.85
0.00
80.39
0.23
1.33
0.20
1.7
12.97
87.03
58
8.29
0.00
89.18
0.30
1.56
0.01
54
2.91b
97.09
87
16.50
0.00
82.41
0.01
1.08
0.00
aAdapted from Wolfe et al., 1982, with correction applied.
bValue was incorrectly reported as 32.91 in original paper.
cThe time needed to reduce steady-state concentration by five half-lives.
^Physical loss from the system by any pathway other than volatilization.
5-7
-------
Hydrolysis is much slower than photolysis (see Table 5-1), but may be a sig-
nificant load-reducing process in waters where photolysis and physical
transport processes are not important (i.e., in deep, non-flowing waters).
Wolfe et al. (1982) found hydrolysis of HEX to be independent of pH over
a range of 3-10. The rate was adequately described by a neutral hydrolysis
rate constant (K^o ± standard deviation) of (1.5i0..6) x 10~6 sec"1 at 30°C,
which corresponds to a half-life of 5.35 days. The rate constant was depen-
dent on temperature at pH 7.0 with the half-life estimated to be 3.31, 1.71
and 0.64 days at 30, 40 and 50°C, respectively. The addition of various
buffers or 0.5 H NaCl did not affect the hydrolysis rate constant, suggest-
ing that the rate constant obtained would be applicable to marine environ-
ments as well. The addition of natural sediments sufficient to adsorb up to
9254 of the compound caused the rate constant to vary by less than a factor
of 2. It was therefore concluded that sorption to sediments would not
significantly affect the rate of hydrolysis (Wolfe et al., 1982).
Some variability of hydrolysis rate with changes in pH was demonstrated
by Yu and Atallah (1977a). They studied the stability of ^C-HEX in water
at pH 3, 6, 9 and 12 at 25°C and 45°C, under dark conditions. HEX was rela-
tively unstable at alkaline pH. At 25°C, the half-lives were 11.4, 11.4 and
6.0 days at pH 3, 6 and 9, respectively, and <2 hours (0.1 day) at pH 12.
At 45°C the half-lives at pH 3, 6 and 9 were 9.2, 10.6 and 4.4 days, res-
pectively. Degradation of HEX resulted in water-soluble products, and based
upon their chromatographic behavior, the hydrolysis products appear to be
polyhydroxy compounds, with CO- as a minor hydrolysis product.
In -the Wolfe et al. study (1982), a preliminary investigation was con-
ducted to determine the products from hydrolysis. The hydrolysis reaction
was conducted at 60-70°C in 40% acetonitrlie-water at 10~4 N HEX and
5-8
-------
proceeded through approximately two half-lives. After extraction and con-
centration of the lipophilic reaction products, analysis by GC/MS showed
nine major peaks in the chromatogram. Several of these were high molecular
weight compounds, but, as with the Yu and Atallah study (1977a), identifica-
tion was not positive.
The degradation of HEX by hydrolysis in the EXAMS model environments,
consisting of a simulated river, pond, eutrophic lake and oligotrophic lake
were estimated to be 8.0, 17.9, 8.3 and 16.5%, respectively, of the total
initially present (see Table 5-2). Hydrolysis in these aquatic environments
was considered to be minor relative to photolysis in the overall degradation
of HEX (Wolfe et al., 1982). The above data indicate that at neutral pH the
hydrolysis half-life is from 3-11 days, compared with a much more rapid
photolytic half-life of <10 minutes.
5.1.2.3. OXIDATION — HEX is not expected to be oxidized under ordi-
nary environmental conditions. In the laboratory, HEX has been reported to
react with molecular oxygen at 95-105°C to form a mixture of hexachlorocy-
clopentenones (Molotsky and Ballweber, 1957). However, based on an estima-
ted first order oxidation rate constant of IxlO"10 H"1 sec"1 at 25°C
in water (see Table 5-1), the EXAMS computer simulation of Wolfe et al.
(1982) predicted that HEX would not be oxidized in the simulated river,
pond, eutrophic lake or oligotrophic lake (see Table 5-2).
5.1.2.4. 8IODEGRADATION -- Tabak et al. (1981) stated that HEX is
biodegraded fairly rapidly in a static laboratory culture. Bottles contain-
ing HEX added to 5 mg yeast extract/a, as the synthetic medium were inocu-
lated with an unspecified domestic wastewater and kept in the dark at 25°C.
5-9
-------
Extractions for GC were done with 20 m». portions of methylene chloride
(neutral pH) at an efficiency of >75% HEX at 5 and 10 mg/s. (concentra-
tions exceeding the compound's aqueous solubility limits) was degraded below
the GC method minimum sensitivity limits (0.1 mg/s.) in 7 days. Volatili-
zation was stated not to occur during a 10-day period in which control
bottles having no inoculum were observed. The importance of chemical
hydrolysis was not discussed by the authors. According to studies presented
in Section 5.1.2.2., 7 days could represent as much as 1-2 hydrolytic half-
lives, accounting for loss by as much as a factor of 4. Based on nominal
concentrations, loss of HEX in these tests exceeded a factor of 50-100;
therefore, hydrolysis cannot fully account for its disappearance.
Atallah et al. (1980) reported an aqueous aerobic biodegradability study
to determine 1f, and at what rate, HEX can be degraded to C0_. The
Inoculum was a mixed acclimated culture containing secondary municipal waste
effluent and several strains of Pseudomonas putida. HEX, labeled with
X4C, was the sole source of carbon in the study, with the exception of
trace levels of vitamins. Total removal of 14C, primarily as volatile
organics, was >80% in the first day in both uninoculated (45 mg/a. HEX) and
Inoculated (4.5 and 45 mg/9. HEX) media, although removal was slightly
higher in inoculated media. 1/1C02 was released from both media, indica-
ting C02 was a product from hydrolysis as well as biodegradation. The
rate of conversion to C02 was initially higher in the uninoculated, but
after a week, became higher in the inoculated media (Figure 5-2).
These studies show clearly that HEX can be biodegraded in aquatic media
under laboratory conditions. However, Wolfe et al. (1982) stated that they
failed to detect any difference in the HEX degradation rate between hydro-
lysis experiments where sterile and nonsterile natural sediments were added
5-10
-------
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5-11
-------
to water (1.0 g/100 mfc). Thus they calculated a relatively low value
(lx!0~s mfi. org~a hr~a; see Table 5-1} as a maximum biodegradation
rate, and consequently biodegradation was estimated to be a relatively unim-
portant fate process in the EXAMS model (see Table 5-2).
5.1.2.5. ADSORPTION— On the basis of computer simulations, Wolfe et
al. (1982) predicted that HEX would adsorb strongly to sediments found in
various aquatic environments (see Table 5-2). The distribution in sediments
from a simulated river, pond, eutrophic lake and oligotrophic lake was esti-
mated to be 98.8, 86.0, 87.0 and 97.1%, respectively, of total HEX in the
system. The sorptive properties of HEX in relation to soil are discussed
below.
5.1.3. Soil. Upon release onto soil, HEX is likely to adsorb strongly to
any organic matter or humus present (Kenaga and Goring, 1980; Weber, 1979).
With time, HEX concentrations should decrease as populations of soil micro-
organisms better adapted to metabolize HEX increase (Rieck, 1977b,c; Thuma
•et al., 1978). Volatilization (See Section 5.2.3.), photolysis, and various
chemical processes may also dissipate the compound in certain soil environ-
ments.
5.1.3.1. ADSORPTION — The soil adsorption properties of compounds
such as HEX can be predicted from their soil organic carbon-water partition
coefficients (K ). Kenaga (1980) examined the adsorption properties of
UC-
100 chemicals and concluded that compounds with K values >1000 are
oc
tightly bound to soil components and are immobile in soils. Those possess-
ing values below 100 are adsorbed less strongly and are considered moderate-
ly to highly mobile. Accordingly, the theoretical K value is useful as
an Indicator of potential soil Teachability or binding of the chemical.
5-12
-------
The K values also indicate whether a chemical is likely to enter water
UC*
by leaching or by being adsorbed to eroded soil particles. Because K
values for HEX are not available in the literature, these values were
calculated using the mathematical equation developed by Kenaga and Goring
(1980) and Kenaga (1980). The equation used was:
log KQC = 3.64 - 0.55 (log WS)
where WS is water solubility (mg/8,), and the 95% confidence limits are
jfl.23 orders of magnitude (OM). The calculated range of K values for
HEX using the reported water solubility values of 2.1 mg/8, (Dal Monte and
Yu, 1977), 1.8 mg/8. (Wolfe et al., 1982) and 0.805 mg/S. (Lu et'al.,
1975) are 2903, 3159 and 4918, respectively. These calculated K values
are all >1000, suggesting that HEX is tightly bound to soil components and
immobile in the soil compartment. Similarly, Briggs (1973) concluded that
compounds with a log octanol/water partition coefficient (log P) >3.78 are
immobile in soil. The measured log P value of HEX is 5.04 (Wolfe et al.,
1982), further indicating that the compound is immobile with respect to
leaching.
The only sorption data found in the literature were for an experimental-
ly flooded soil. Weber (1979) reported that an average of 68% of applied
HEX was adsorbed to Cape Fear loam soil present in aqueous solutions. In
these experiments, aqueous solutions (50 ma) containing 0.0, 0.41 (1.5
liH), 0.82 (3.0 MM) and 1.64 (6.0 PH) mg/kg of ^C-HEX were added to
soil samples (0.50 mg) in stoppered bottles, which were shaken at room temp-
erature for 24 hours. Standards, controls and two replications were used in
all cases. The difference between the initial and the 24-hour equilibrium
concentration of radiolabel in water was considered to be the amount of HEX
adsorbed to soil. Less than 5% of the radiolabel was lost from the bottles
5-13
-------
over the 24-hour period. About 62, 66 and 75% of the applied dose was
adsorbed to the soil at 0.41, 0.82 and 1.64 ppm concentrations, respec-
tively. Weber (1979) suggested that the HEX Is very strongly adsorbed by
organic soil colloids because of Its lipophllic character.
5.1.3.2. BIODEGRADATION — The metabolism of HEX by soil micro-
organisms apparently Is an Important process In Its environmental degrada-
tion. Soil degradation Is rapid under nonsterlle aerobic and anaerobic
conditions, with Indirect evidence for mlcroblal Involvement reported by
Rleck (1977b,c). In one of his studies, Rleck (1977b) used several types of
treatments and soil pHs to determine If the blodegradatlon of HEX 1n Maury
silt loam soil was either biologically or chemically mediated, or both
(Figure 5-3). Soils were incubated in glass flasks covered with perforated
aluminum foil and kept on a laboratory shelf, presumably exposed to ambient
lighting through the flask walls. When 14C-HEX was applied to nonsterile
soil at 1 mg/kg, only 6.1% was recovered as nonpolar material (either HEX or
nonpolar degradation products) 7 days after treatment, and -71.7% was polar
and unextractable material. Adjustment of pH to 4 or 8 had little effect on
these results. By comparison, in autoclaved soil (the control), 36.1% of
the applied dose was recovered as nonpolar material and only 33.4% recovered
as polar and unextractable material (see Figure 5-3). The degradation of
HEX under anaerobic (flooded) conditions occurred at a slightly faster rate
than under aerobic conditions. However, no sterile, flooded control was
used to determine the effects of hydrolysis, which could have accounted for
the observed difference in this treatment. The mean total recovery in all
treatments decreased from 67% at 7 days to 55% at 56 days. This decrease
was attributed to volatilization of HEX and/or its degradation products.
5-14
-------
so
• UNALTERED
O pH 4
O pH 8
A AUTOCLAVE D
A SODIUM AZIDE
• FLOODED
DAYS AFTER TREATMENT
FIGURE 5-3
Persistence of Nonpolar 14C when 14C-HEX 1s Applied
to Unaltered and Altered Soils
Source: Adapted from R1eck, 19775
5-15
-------
Volatilization from soil was examined In another experiment (R1eck,
1977c). In a 14-day study, radiocarbon volatilized from nonsterile,
x4C-HEX-treated soil was trapped and assayed. Over the study duration, a
total of 20.254 of the applied 14C was trapped; 11.2% in hexane and 9.0% in
ethanolamine-water. Most of the hexane fraction (9.3% of applied 14C) was
trapped during the first day, probably representing volatilized HEX, How-
ever, the ethanolamine-water fraction, considered to represent evolved
C0?, was released gradually over the 14-day period. In the soil analysis,
nonpolar (extractable) and polar (extractable and unextractable) material
accounted for 3.4 and 40.0% of the dose, respectively, during the 14 days;
thus, total recovery was only 63.6% including volatilization. No metabolic
products were identified:1n either study by Rieck (1977b,c).
In these studies (Rieck, 1977b,c), HEX was degraded to polar material in
both sterile and nonsterile soils, indicating the occurrence of an abiotic
degradation process such as hydrolysis by soil water and possibly some
photolysis. Since degradation occurred more quickly in nonsterile soils,
biodegradation evidently was also occurring. Volatilization of HEX occurred
mainly during the first day, and apparently represented no more than 11.2%
of the total amount applied (Rieck, 1977c), although the low total recovery
in this experiment decreases the reliability of this figure.
Under contract with the U.S. EPA, Thuma et al. (1978) studied the
feasibility of using selected pure cultures (organisms not identified) to
biodegrade spills of hazardous chemicals on soils, including HEX. They
tested 23 organisms and found that from 2-76% of the HEX had been removed
from the aqueous culture medium within 14 days. Seven of the 23 organisms
degraded more than. 33% of the HEX within 14 days (Table 5-3). Losses of
HEX, other than biodegradation, were accounted for by the use of controls.
5-16
-------
TABLE 5-3
Mlcroblal Degradation of HEX During
14-day Exposure In a Test Medium*
Organism
Code Number
Control 1
Control 2
006
016
020
022
123
369
505
HEX Remaining 1n
Test Medium (ppm)
635
630
410
415
410
150
395
350
265
Percent Degraded
Relative to Control
,_
35
34
35
76
38
45
58
*Source: Adapted from Thuma et al., 1978
5-17
-------
These studies Indicate that the persistence of HEX 1n soil 1s brief,
with degradation of >90% of applied HEX to nonpolar products within ~7 days.
Factors contributing to this loss Include abiotic and biotic degradation
processes and volatilization, although the relative importance of each is
difficult to quantify given the limited information available.
5.2. TRANSPORT
5.2.1. A1r. The vapor pressure, water solubility, vapor density,
adsorption properties, rapid photolysis (Wolfe et al., 1982) and high
reactivity (Callahan et al., 1979) of HEX combine to affect its atmospheric
transport. The atmospheric transport of HEX vapor from a closed waste site
at Montague, MI was demonstrated by Peters et al. (1981). At an unspeci-
fied distance downwind of the site, HEX was detected in air at concentra-
tions of 0.032-0.053 ppb (0.36-0.59 yg/m3). Based on the concentration
ratio of HEX and a tracer gas released at a known rate, the average HEX
emission rate during the measurement period was calculated to be 0.26 g/hour
Volatilization of HEX from water may occur following either industrial
discharge [e.g., a concentration of 18 mg/s. was found in the aqueous dis-
charge at a Memphis pesticide plant (U.S. EPA, 1980c)] or accidental spill.
The tendency of HEX to adsorb to organic matter in water or soil would limit
the compound's volatility, as would suspended solids in surface water.
Transport of HEX vapor will also be limited by the estimated atmospheric
residence time, based on photolysis, hydrolysis and ozone reaction rates, of
~5 hours (Cupitt, 1980). HEX adsorbed onto aquatic or terrestrial particles
may also enter the atmosphere and be transported in the air for a time while
being transformed by photolysis or other processes.
5-18
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fVs part of an experiment with chlordane, Bevenue and Yeo (7969) found
some interesting vaporization and adsorption properties of HEX, which may
exist in an amount as large as 1% in commercially available chlordane.
Small quantities of HEX (0.5 mg) in open vials were placed in closed glass
vessels containing 20 ms. of either distilled water or isooctane, so that
only vaporized HEX could contact the solvent. Vessels were stored under
fluorescent lighting. Gas chromatographic data from the solutions of dis-
tilled water initially revealed the presence of adsorbed vapor of HEX and
Us degradation products, indicating transport from air to water. Beyond 3
days exposure, however, the chemical and its products had completely disap-
peared from the GC chrornatogram, indicating either dissipation or decomposi-
tion of the compound. The data obtained from the Isooctane solutions re-
vealed a different GC pattern. No degradation was observed after 24 hours,
while a multiple-peak chromatogram (indicating degradation products) was
obtained after the solutions were exposed 7-21 days. This chromatogram
suggests that the compound may be susceptible to atmospheric oxidation and
photodecomposition or both (NCI, 1977). The more rapid disappearance of
compound and degradation products in water than in the Isooctane solution
may further indicate the occurrence of hydrolysis.
More information on the volatility and adsorption of HEX is presented in
Sections 5.2.2. and 5.2.3., respectively.
5.2.2. Water. HEX introduced into water bodies may be transported in
either undissolved, dissolved or adsorbed forms. In its undissolved form,
HEX will tend to sink because of Its high specific gravity and may then
become concentrated in deeper waters, where photolysis and volatilization
would be precluded. Some HEX may be dissolved 1n water (up to ~2 mg/a.)
and then be dispersed with water flow (i.e., in a river). HEX tends to
5-19
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adsorb onto organic matter because of Us lipophilic nature and may then be
transported with water flow in a suspended form. Transport to the air may
occur by volatilization, which was measured in laboratory studies (Kilzer et
al., 1979; Weber, 1979) and predicted using the EXAMS model by Wolfe et al.
(1982). However, suspended solids in surface water may be a major factor 1n
reducing volatilization.
Weber (1979) measured the volatility of 14C-HEX from distilled water
following the incubation of the glass-stoppered and unstoppered test bottles
shaken at room temperature for 24 hours. Experiments were performed with
standard HEX solutions of l.SxlO"6 M (0.41 mg/a.) in distilled water,
with readings taken 24 hours later. From the full glass-stoppered bottles,
only 4-5% of the HEX was lost, while in the half-full stoppered bottles,
15-16% of the chemical was missing. This suggests that head space in the
bottle contributed to the loss of HEX. The volatility of HEX was shown by
the loss of 45-47% from the half-full, unstoppered bottles over the 24-hour
period.
Kilzer et al. (1979) determined the rate of 14C-HEX volatilization
from water as a function of the rate of water evaporation. Bottles contain-
ing aqueous HEX solutions (50 pg/fc) were kept without shaking at 25°C.
The escaping vapor condensed on a "cold finger" and was quantified by liquid
scintillation spectroscopy. Based on recovery of added label, the HEX vola-
tilization rates for the first and second hours of testing were calculated
to be 5.87 and 0.75%/mil H20, respectively. Since the water evaporation
rate was 0.8-1.5 ma/hour, rates for HEX were within the ranges of 4.7-8.8
and 0.,6-1.1%/hour, respectively. These results suggest that a fairly rapid
initial volatilization occurred at the water surface, and that by the second
hour, diffusion of HEX to the water surface may have been limiting because
of the static conditions of the test. If the rate observed during the
5-20
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.second hour had continued for the remainder of 24 hours, total loss would
have been ~18-34%, or somewhat less than that observed in the test by Weber
(1979) where unstoppered bottles were shaken.
In the aqueous biodegradation test of Atallah et al. (1980) described in
Section 5.1.2.4., a very high rate of volatilization was determined. Over
80% of the radiolabel added as 14C-HEX had disappeared after the first
day, even from uninoculated media. Most was found to have volatilized
(total recovery averaged 94%) and was primarily in organic form. The physi-
cal conditions of the test, such as covering, shaking or aeration of test
solutions, were unspecified. In addition, disappearance of label at initia-
tion was >50%. This peculiarity was not explained, but could be due to the
use of HEX concentrations of 4.5 and 45.3 mg/9,, which exceed the limit of
aqueous solubility.
Wolfe et al. (1982) also studied the evaporation rate of HEX from water
and experimentally determined the Henry's law constant (H) to be 0.027+_0.010
atm mVmole. This value corresponds with 0.0137 and 0.0357 atm mVmole
calculated from the measured vapor pressure (0.08 mm Hg at 25°C) (Irish,
1963) and the water solubilities (2.1 and 0.805 mg/a) (Dal Monte and Yu,
1977; Lu et al., 1975, respectively), according to the following equation:
Vapor pressure (atm)
H =
Water solubility (mole/m3)
The mathematical EXAMS model (see Section 5.1.2.) was used to indicate the
relative importance of volatilization and other processes in the fate and
transport of HEX (load reduction) of four aquatic systems (see Table 5-2).
The model indicated that volatilization of HEX from a river, pond, eutrophic
lake and oligotrophic lake would account for only 0.69, 1.33, 1.56 and 1.08%
of load reduction, respectively. These values are quite low compared with
the laboratory values described previously. This discrepancy is apparently
5-21
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due to the fact that the model estimates that 86-99% of the HEX present 1n
these systems will be adsorbed to sediment, and thus will not be subject to
volatilization. Experiments measuring vaporization of HEX from water-
sediment systems.have not been conducted.
Export (i.e., physical loss by methods other than volatilization) was
predicted to be a very important transport mechanism in the simulated river
environment (Wolfe et al., 1982). Using the EXAMS model, export accounted
for load reductions of 72% in the river, as compared with the three nonflow-
ing environments mentioned previously, where photolysis was the dominant
removal mechanism.
5.2.3. Soil. As indicated previously (Section 5.1.3.1.), HEX in soils is
predicted to be tightly adsorbed to organic matter and relatively resistant
to leaching by soil .water. Thus, the primary routes of transport for soil
applied HEX are by movement of particles to which it is adsorbed or by
volatilization. No. data are available pertaining to HEX transport on soil
particles; however, a few studies have determined the rate of volatilization
from soils.
Kilzer et al. (1979) determined that 14C-HEX volatilized from moist
soils (sand, loam and humus) at a faster rate in the first hour than in the
second hour of the study. HEX (50 pg/kg) was placed in bottles with each
soil type and shaken vigorously. The bottles were incubated for 2 hours at
25°C, apparently without shaking. The evaporating HEX condensed on a "cold
finger" and was quantified by liquid scintillation counting. For sand, loam
and humus, the volatilization rate was expressed as the percentages of
applied radioactivity per ma. of evaporated water and were for the first
hour 0.83, 0.33 and 0.14%, while for the second hour they were 0.23, 0.11
and 0.05%, respectively. Volatilization was much higher from the sand.
5-22
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For HEX and nine other tested chemicals, KHzer and coworkers found that the
volatilization rate from distilled water cannot be used to predict the rate
from wetted soils. Among the chemicals tested, there was no correlation
between water solubility or vapor pressure and volatilzation from soils.
The volatilization rate for HEX in soil was primarily dependent upon soil
organic matter content, mainly because of the highly adsorptive properties
of HEX.
Rieck (1977c) measured the rate of volatilization of HEX from Maury silt
loam soils (see Section 5.1.3.2.). Following the application of 100 mg of
14C-HEX to soil, the cumulative evaporation of HEX and its nonpolar meta-
bolites (penta- and tetrachlorocyclopentadlene) on days 1, 2, 3, 5, 7 and 14
were 9.3, 10.2, 10.6, 10.8, 11.0 and 11.2%, respectively. The results indi-
cate that HEX evaporation to air occurred mainly during the first day fol-
lowing application and was probably associated with the surface soil only.
When compared with data presented in the preceding section (5.2.2.),
these studies demonstrate that HEX volatilizes from soils much more slowly
than from sediment-free water. This difference is most likely due to
adsorption of HEX to the soil matrix, and possibly to slow diffusion to the
soil surface.
5.3. BIOCONCENTRATION/BIOACCUMULATION
The occurrence of toxic substances in the environment raises the issues
of whether humans may be exposed to them by air, water or food and, if so,
what are the physiological exposures? The transport and fate of HEX (see
Sections 5.1. and 5.2.) are the primary determinants of human exposure to
the environmental sources of these compounds, but the more crucial physio-
logical exposure levels are determined by the manner in which a compound
crosses biological membranes. Bioaccumulation, alternately sometimes
5-23
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expressed as biological persistence, Is the net result of the absorption and
elimination rate of a compound and, therefore, determines the level and
duration of human physiological exposure.
The terminology used in this section will follow that of Hacek et al.
(1979): bioconcentration implies that tissue residues result only from
exposure to the ambient environment (e.g., air for terrestrial or water for
aquatic species); bioaccumulatlon considers all exposures (air, water and
food) of an individual organism as the source of tissue residues; and bio-
magnification defines the increase in tissue residues observed at succes-
sively higher trophic levels of a food web.
The log octanol/water partition coefficient (log P) of HEX has been
experimentally determined to be 5.04 (Wolfe et al., 1982) and 5.51 (Veith et
al., 1979), which would Indicate a substantial potential for bioconcentra-
tion, bloaccumulation arid biomagnification. Actual determinations of bio-
concentration and bioaccumulation in several aquatic organisms, however,
indicate that HEX does not accumulate to a great extent (Podowski and Khan,
1979, 1984; Veith et al., 1979; Spehar et al., 19/9; In el. al., 1975),
mainly because 1t Is metabolized rapidly.
Podowski and Khan (1979, 1984) conducted several experiments concerning
the uptake, bloaccumulation and elimination of a/lC-HEX in goldfish
(Carasslus auratus) and concluded that the species eliminated absorbed HEX
rapidly. In one experiment, fish were transferred daily into fresh solu-
tions of 14C-HEX for 16 days. This transfer of three fish/jar resulted in
accumulative exposure of 240 pg of HEX. Nominal HEX concentrations of 10
ng/a. resulted in measured water concentrations (based on radioactivity)
in the range of 3.4-4.8 ng/S., because of rapid volatilization of the
compound. Radioactivity accumulated rapidly in fish tissue, reaching a
5-24
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maximum on day 8 corresponding to ~6 mg HEX/kg. Since an undetermined
amount of the radioactivity was present as metabolites, no bioconcentration
factor can be calculated. From day 8 to day 16, tissue levels declined in
spite of daily renewal of exposure solutions, indicating that excretion of
HEX and/or its metabolites was occurring more rapidly than uptake. In a
static exposure to an initial measured HEX concentration of 5 yg/8.,
radioactivity was taken up by the fish to a level corresponding to 1.6 mg
HEX/kg on day 2, accompanied by a slight decrease of HEX in the water. By
day 4, -50% of the absorbed activity had been excreted, and the water level
increased. Over the following 12 days, radioactivity in both water and fish
declined slowly.
Podowski and Khan (1979, 1984) also studied elimination, metabolism and
tissue distribution of HEX injected intraperitoneally into goldfish and
concluded that goldfish eliminate injected HEX both rapidly and linearly
(biological half-life -9 days). Fish (27-45 g) were Injected with 39.6 Pg
of 14C-HEX and analyzed 3 days later. Of the 97% of the radiolabeled dose
accounted for, -18.9% was eliminated by the fish, leaving a residual of
78.1%. Of the residue found in the fish, 47.2% was extractable 1n organic
solvent (little of the radiolabeled material could be identified as HEX,
which indicated that biotransformation had occurred); 10.6% was water
soluble metabolites; and 20.3% was unextractable. None of the metabolites
were identified. A biphasic elimination was observed — rapid at first,
followed by a slower phase.
In another part of these studies, residual activity in several fish
tissues was assayed 2, 4, 6 and 8 days following an injection of 38.4 jig/
fish of 14C-HEX. Results showed activity corresponding to 0.2 and 0.3 mg
HEX/kg in the spinal cord and gills, respectively. These concentrations
5-25
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were constant throughout the 8-day period of the study. Residues In the
kidneys and bile Increased within the same period from 1-3 and 0-32 mg/kg,
respectively, Indicating elimination by these routes. The authors stated
that the Increase was probably from enhanced conversion of the parent com-
pound Into polar products suitable for elimination. In the other tissues,
all residual levels dropped leaving only the liver with levels >1 mg/kg
(Podowskl and Khan, 1979, 1984). The authors did not Identify the metabo-
lites because of the complications: created by the fact that HEX and Us
metabolites are very reactive and extremely llpophlllc. When the fat was
removed to purify the HEX, over 90% of the radioactivity levels Initially
accounted for 1n the goldfish were lost.
Velth et al. (1979) determined the bioconcentratlon factor (BCF) for HEX
to be 29 in the fathead minnow (Pimephales promelas). In a 32-day flow-
through study, 30 fish were exposed to HEX at a mean concentration of 20.9
pg/2. and were sacrificed five at a time for residue analysis at 2, 4, 8,
16, 24 and 32 days. The study was conducted using Lake Superior water at
25°C (pH 7.5, dissolved oxygen >5.0 mg/8. and hardness 41.5 mg/a. as
CaC03). On the basis of its estimated octanol/water partition coefficient
alone (log P = 5.51), a BCF of -9600 would have been predicted. However,
HEX did not bioconcentrate substantially, and therefore deviated from the
log P:log BCF relationship shown for most of the other 29 chemicals tested
by these researchers.
Spehar et al. (1979) conducted a 30-day early-life-stage, flowthrough
toxicHy test at 25°C with the fathead minnow (P. promelas). HEX residues
In the fish after 30 days of continuous exposure to HEX were <0.1 mg/kg for
all concentrations tested (0.78-9.1 pg/a), and the BCF was <11 (0.1
mg/kg 1n fish divided by 9.1 vg/i in water). In addition, toxicHy
5-26
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results Indicated that a median lethal threshold (or Incipient LC^Q) was
attained within 4 days. The authors concluded that the rapid attainment of a
threshold toxicity level and the low BCF indicate that HEX is noncumulative.
Lu et al. (1975) studied the fate of HEX in a model terrestrial-aquatic
ecosystem maintained at 26.7°C with a 12-hour photoperiod. The model eco-
system consisted of 50 sorghum (Sorghum vulgare) plants (3-4 inches tall) in
the terrestrial portion; 10 snails (Physa sp.), 30 water fleas (Daphnia
magna), filamentous green algae (Oedogonium cardiacum) and a plankton cul-
ture were added to the aquatic portion. The sorghum plants were treated
topically with 5.0 mg of 14C-HEX in acetone to simulate a terrestrial
application of 1.0 Ib/acre (1.1 kg/ha). Ten early-fifth-instar caterpillar
larvae (Estigmene acrea) were placed on the plants. The Insects consumed
most of the treated plant surface within 3-4 days. The feces, leaf grass
and the larvae themselves contaminated the moist sand, permitting distri-
bution of the radiolabeled metabolites by water throughout the ecosystem.
After 26 days, 300 mosquito larvae (Culex plpiens quinquefasciatus) were
added to the ecosystem, and on day 30, three mosquito fish (Gambusia
affinis) were added. The experiment, was terminated after 33 days, and the
various parameters were analyzed. The radioactivity was then extracted from
water with diethyl ether and from organisms with acetone. The results of
TLC analysis of the extracts are presented in Table 5-4. Data were not
reported for Daphnia or the salt marsh caterpillar. Uptake in this experi-
ment occurred through food as well as water, and therefore is termed bioac-
cumulation rather than bioconcentration. Lu et al. (1975) used the term
ecological magnification (EM) to designate the bioaccumulation factor (BAF).
5-27
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TABLE 5-4
Relative Distribution of HEX and Its Degradation Products3
14C-HEX Equivalents (ppm)
Water
Mosquito
Algae Snail Larva Fish
(mg/kg) (mg/kg) (mg/kg) (mg/kg)
HEX 0.00024 0.0818 0.3922
Other extractable compound? 0.00204 0.1632 p.3824
Total extractable 14Cb 0.00228 0.2450 0.7746
Unextractable *«C 0.00750 0.0094 0.0814
Total "C13 0.00978 0.2544 0.8560
0.2230
0.2542
0.4772
0.0104
0.4876
0.1076
0.1542
0.2618
0.0982
0.3600
aSource: Lu et al., 1975
bUnderlines indicate summation
5-28
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The BAF for HEX In fish was 448 (0.1076 mg/kg in fish divided by 0.24
pg/9, 1n water) for the 3-day exposure period, indicating a moderate
potential for concentration (Kenaga, 1980). The BAF in algae (<33-day
exposure), snails {<33-day exposure) and mosquito larvae (7-day exposure)
was reported to be 341, 1634 and 929, respectively (Lu et al., 1975).
Biomagnification, measured as the ratio of HEX residues between trophic
levels (e.g., snail/algae or fish/mosquito), was far less substantial than
bioconcentration. Based on the HEX tissue residues, the snail/algae ratio
was 0.3922/0.0818 = 4.8 and the fish/mosquito ratio was 0.1076/0.2230 = 0.48.
Lu et al. (1975) also studied the metabolism of HEX by the organisms
present in the model terrestrial-aquatic ecosystem. None of the products
were identified except for HEX. The authors reported that unmetabolized HEX
represented large percentages of the total extractable 14C, being 33% In
algae, 50% in snail, 46% in mosquito and 41% in fish. Percent biodegrada-
tion was calculated for each organism [(unextractable 14C x 100)/total
14C] and reported to be: 4% for the alga (in <33 days); 10% for the snail
(in <33 days); 2% for the mosquito (in 7 days); and 27% for the fish (in 3
days). However, these values may underestimate the extent of metabolism,
since acetone extractable polar compounds were not considered in the calcu-
lations.
Velsicol Chemical Corporation (1978) conducted fish tissue residue
studies below their Memphis, TN facility and reported that HEX was not
detected in either catfish or carp, although chlorinated compounds were
detected in the fish tissue. The possible source of these other compounds
was not discussed. In a Joint Federal and state study of the Mississippi
River above, around and below Memphis, Bennett (1982) of the U.S. EPA
reported that HEX was not detected in any of the eight fish sample groups
analyzed by GC/MS.
5-29
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In contrast to the above-described findings, fish collected from the
stream in the vicinity of the Hooker chemical plant discharge in Montague,
Michigan, were reported to contain 4-18 yg/kg of HEX in the edible fil-
lets. However, there was some question as to whether the analyzed compound
was HEX or a degradation product (Swanson, 1976).
5.4. SUMMARY AND CONCLUSIONS
The fate and transport of HEX in the atmosphere is not known, but avail-
able information suggests that the compound does not persist. Cupitt (1980)
estimated its tropospheric residence time to be ~5 hours, with photolysis
and reaction with hydroxyl radicals and ozone being the key degradative pro-
cesses. However, atmospheric transport of HEX from an area of stored wastes
has been demonstrated, at least for a short distance (Peters et al,, 1981).
In water, HEX is likely to dissipate rapidly by means of photolysis,
hydrolysis and biodegradation. In shallow water (a few centimeters deep),
HEX has a photolytic half-life of -0.2 hours (Butz et al., 1982; Wolfe et
al., 1982). In deeper water where photolysis is precluded, hydrolysis and
biodegradation should become the key degradative processes when there is
little movement from the system. The hydrolytic half-life of HEX is several
days, and is not strongly affected by pH in the environmental range (5-9),
by salinity or by suspended solids (Yu and Attallah, 1977a; Wolfe et al.,
1982). Biodegradation may also be a significant process in certain waters
(Tabak et al., 1981), although the evidence is weak. HEX is known to vola-
tilize from water (KHzer et al., 1979; Weber, 1979). It is probable that
volatilization is limited by diffusion, that is, loss from deeper waters
would occur very slowly unless vertical mixing has taken place. Sorption on
sediments may also retard volatilization.
5-30
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The fate and transport of HEX in soils are affected by its strong ten-
dency to adsorb onto organic matter (Kenega and Goring, 1980; Wolfe et al.,
1982; Weber, 1979). HEX is predicted to be relatively immobile in soil
based on its high log P value (Briggs,, 1973). Volatilization, which is
likely to occur primarily at the soil surface, is inversely related to the
organic matter levels and water-holding capacity of the soil (Kilzer et al.,
1979). Leaching of HLX by groundwater should be very limited, and chemical
hydrolysis and microbial metabolism, are expected to reduce environmental
levels. HEX is metabolized by a number .of unidentified soil microorganisms
(Rieck, 1977b,c; Thuma et al., 1978). .
The bioconcentration/bioaccumulatiort/biomagnification potential of HEX
would appear to be substantial based on its high lipophilicity. BAFs de-
rived from a short-term model ecosystem study appear to indicate a moderate
accumulation potential for algae (BAF = 341), snails (1634), mosquito larvae
(9?9) and mosquito fish (448). However, the compound did not substantially
biomagnify from algae to snails -or from mosquito larvae to fish (Lu et al.,
1975). In addition, steady-state bioconcentration factors (BCFs) in fish,
measured in 30-day flow-through exposures to constant levels of HEX, were
only 29 and <11, respectively (Veith et al., 1979; Spehar et al., 1979).
Metabolism and excretion of HEX by goldfish were demonstrated by Podowski,
and Khan (1979).
5-31
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6. ECOLOGICAL EFFECTS
The effects of HEX have been reported for several aquatic organisms,
Including invertebrates and fish from freshwater and saltwater environments
and saltwater algae. The bioconcentration potential of HEX in aquatic
organisms and ecosystems has also been studied; these data have been dis-
cussed in Section 5.3. The effects on microorganisms have been examined to
some degree. However, few studies have been located which describe the
effects of HEX on terrestrial plants or vertebrates.
6.1. EFFECTS ON AQUATIC ORGANISMS
6.1.1. Freshwater Aquatic Life.
6.1.1.1. ACUTE TOXICITY — Several studies are available on the
effects resulting from exposure of freshwater aquatic life to various con-
centrations of HEX.
Two studies have reported the acute toxicity of HEX in EL magna (Bucca-
fusco and LeBlanc, 1977; Vilkas, 1977). The results are shown in Table 6-1.
The 48-hour LC5Q value ranged from 39-52 Mg/8,, and the 48-hour no-
effect level ranged from 18-32 yg/fc. In the study by Vilkas (1977),
routine water quality parameters were also analyzed. Results showed that
the pH values, determined initially and after 48 hours, increased with an
increase in HEX concentration.
Results from acute toxicity tests with HEX have been reported for a
number of freshwater fish species (Table 6-1). The 96-hour LCgQ value for
fathead minnow larvae in a flowthrough test with measured toxicant concen-
trations was 7 yg/s. (Spehar et al., 1977, 1979). Values obtained with
adult fathead minnows in static tests with unmeasured toxicant concentra-
tions ranged from 59-180 yg/8. (Henderson, 1956; Buccafusco and LeBlanc,
1977). Reported 96-hour values for goldfish, channel catfish and bluegllls
6-1
-------
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were also within this range (Podowski and Khan, 1979; Khan et al., 1981;
Buccafusco arid LeBlanc, 1977). Anomalously high values for blueglll (25,000
pg/8.) and largemouth bass (20,000 pg/S.). well above the solubility
limit of 800-2100 pg/a. (see Section 3.2.1.). were reported by Davis and
Hardcastle (1957) (see Table 6-1). These results could be high due to the
failure to properly disperse the toxicant in the test water (no carrier was
mentioned), and/or to volatilization of the compound, since the water was
aerated during the test.
Sinhaseni et al. (1982) have recently reported biological effects of HEX
in rainbow trout (Salmo qairdneri) exposed to 130 pg/a. HEX in a nonre-
circulating flowthrough chamber. Oxygen consumption, measured polarograph-
ically, increased by 193% within 80 minutes and then gradually decreased
until death in ~5 hours. Vehicle controls showed no effects after 76 hours
of exposure. HEX added to normal trout mitochondria increased basal oxygen
consumption. The authors concluded that HEX uncoupled oxidative phosphoryl-
ation.
Sinhaseni et al. (1983) continued their research on the respiratory
effects of HEX on intact rainbow trout. Acclimated rainbow trout were ex-
posed to 130 ppb HEX in a flow-through well water circuit which was designed
to permit measurements of oxygen consumption in fish. Again, HEX increased
oxygen consumption rates (186i24%), with the maximum oxygen consumption
rates being nearly the same as the previous experiment (-84 minutes). The
oxygen consumption decreased until death (-6.5 hours). Control trout
(acetone vehicle) showed no changes. The authors reported profound respira-
tory stimulation and HEX appeared to uncouple oxidative phosphorylation.
Sinhaseni et al. (1983) postulated that HEX intoxication in the intact
animal may be due to increased oxygen consumption and impaired oxidative ATP
synthesis due to the mitochondrial uncoupling action of HEX.
6-3
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6.1.1.2. SUBCHRONIC/CHRONIC TOXICITY — Spehar et al. (1977, 1979)
conducted 30-day early life stage flowthrough toxicity tests with fathead
minnows (P.. promelas). Tests were performed with measured concentrations
and were initiated with 1-day-old larvae. The 96-hour LC5Q value was
reported in the preceding section. The 96-hour mortality data indicated a
sharp toxicity threshold, such that 94% survival was observed at 3.7
pg/8,, 70% at 7.3 yg/8., and 2% at 9.1 pg/s.. At the end of the
30-day exposure period, mortality was only slightly higher, with 90% survi-
val at 3.7 jig/8., 66% at 7.3 pg/S., and 0% at 9.1 pg/fc. These
results indicated that the median lethal threshold, the lowest concentration
causing 50% mortality, was attained within 4 days. In addition, the HEX
residues found in fathead minnows during the end of the 30-day tests were
low (<0.1 pg/g) and the BCF value was reported to be <11 (Spehar et al.,
1979). The authors concluded that the toxicity data and BCF values
indicated that HEX was noncumulative in fish; i.e., did not bioconcentrate
in fish as a result of continuous low-level exposure to HEX. The growth
rate of surviving larvae, measured as both body length and weight, did not
decrease significantly at any of the concentrations tested, compared with
the controls. This was true even at 7.3 pg/fc, a level greater than the
calculated LC.-n value. Based on these toxicity and growth data, Spehar et
bu
al. (1977, 1979) concluded that 3.7 pg/a. was the highest concentration
of HEX that produced no adverse effects on fathead minnow larvae. Thus, the
maximum acceptable toxicant concentration (HATC) was In the range of 3.7-7.3
pg/S.. No other chronic toxicity data for any freshwater species were
located.
6-4
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fe."\.2. Marine and Estuarlne Aquatic Life.
6.1.2.1. ACUTE TOXICITY — Walsh (1981) reported unpublished data on
the effects of HEX on four species of marine algae, derived according to the
method described by Walsh and Alexander (1980). The 7-day EC™ was calcu-
lated as the concentration causing 50% decrease' in biomass compared with the
control, as estimated by absorbance at 525 nm. The 7-day EC™ values
reported Indicated a wide range of susceptibility between the species
tested. Isochrysis galbana and Skeletonema costatum were the most suscep-
tible species, with the average 7-day EC50 values reported were about 3.5
and 6.6 yg/8., respectively. The average value for Porphyridium cruentum
was 30 yg/a., while that for Dunaliella tertiolecta was 100 pg/a..
Other tests with S. costatum indicated that the direct, algicidal effect of
HEX was less pronounced than its effect on growth. After 48 hours of expo-
sure to HEX at 25 yg/a., mortality, as indicated by staining and cell
enumeration, was only 4% (Walsh, 1983).
Among marine invertebrates, the 96-hour LC_ values for HEX ranged
from 7-371 yg/5t (Table 6-2) (U.S. EPA, 1980a). Except where indicated,
these results were from static tests with nominal concentrations of HEX.
The organism exhibiting by far the highest LC™ was the polychaete,
Neanthes arenaceodentata. which is an infaunal organism living in the sedi-
ment. The two shrimp species tested were more sensitive to HEX by a factor
of 10 or more.
The static LC5Q value reported by U.S. EPA (1980a) for the grass
shrimp, Palaemonetes pugio. was slightly higher than that for the mysid
shrimp, Hysidopsis bahia (see Table 6-2). However, the LC,-n for the mysid
shrimp was considerably lower in a flow-through test than in the static
test. Similarly, the LC5Q value was lower when calculated from actual
6-5
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TABLE 6-2
Acute Toxlcity Data on Marine Organisms Exposed to HEX3
Species
Polychaete
Neanthes arenaceodentata
Grass shrimp
Palaemonetes pugio
Hysid shrimp
Hysidopis bahia
Hysid shrimp
Hysidopis bahia
Hysid shrimp
Hysidopis bahia
Pinfish
Laqodon rhomboides
Spot
Leiostomus xanthurus
Sheepshead minnow
Cyprinodon variegatus
Method^
S,U
S,U
S,U
FI.U
n,M
S,U
S,U
S,U
96-hour LCsoc
(pgA)
371
(297-484)
42
( ?f>-cin\
32
(27-37)
12
(10-13)
7
(6-8)
48
(41-58)
37
(30-42)
45
(34-61)
aSource: U.S. EPA, 1980a
&H = measured concentrations; S
concentrations
C95J4 confidence interval
= static; FT = flowthrough; U = unmeasured
6-6
-------
measurements of HEX concentrations in the test solutions (measured concen-
tration) than when calculated according to the concentrations based on
amounts originally added to test solutions {nominal concentrations).
The acute toxicity values for HEX were comparable for each of three
marine fish species tested (U.S. EPA, 1980a). The static 96-hour LC
values based on unmeasured concentrations for spot, sheepshead minnow and
pinfish varied only from 37-48 ng/8. (see Table 6-2).
6.1.2.2. CHRONIC TOXICITY -- In an unpublished study (U.S. EPA,
1981), groups of 40 mysid shrimp were exposed for 28 days to measured, flow-
through concentrations of HEX. From the data shown in Table 6-3, measured
concentrations were about one-half of nominal. Mortality occurred in all
concentrations except the control, but showed no consistent dose-response
relationship. Fecundity, however, was more clearly related to dose
(Table 6-3).
No other data were located on the chronic toxicity of HEX to saltwater
organisms.
6.2. EFFECTS ON OTHER ECOSYSTEMS
The effects of HEX on microorganisms in aqueous and soil systems have
been tested. Many of the aqueous concentrations tested exceeded the upper
limit of aqueous solubility of 0.8-2.1 mg/8.; these concentrations usually
were achieved by use of an organic solvent. Thus the environmental signifi-
cance of the results must be Interpreted with caution.
Cole (1953) inoculated 10 strains of common human and animal pathogens
into growth media containing various concentrations of HEX. The Inhibiting
concentration, or lowest concentration 1n which no growth was observed after
96 hours of contact, ranged from 1-10 mg/s. HEX. Addition of 5 or 10
mg/8, of HEX to sewage effluent Inoculated with Salmonella typhosa was also
6-7
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TABLE 6-3
Effects of 28 Days Exposure of Mysid Shrimp, Mysidopsis bahia. to HEX3
Concentration (yg/a.)
Nominal
Control
0.75
1.5
3.0
6.0
12.0
Measured
ND
0.30
0.70
3.0
2.9
6.2
Mortality
(%)
0
18.9
43. 6b
18. 4C
23.1
97. 5b
Total
Offspring
195
167
67
79
72
0
Offspring
per Female
15.7
11.6
5.0b
5.4b
5.5b
Ob
aSource: U.S. EPA, 1981
bS1gnificantly different from the control (p<0.05)
°No explanation was given in original text as to this value in comparison
with the next measured value of 2.9 yg/a..
NO = Not detected
6-8
-------
96 hours of contact, ranged from 1-10 mg/s. HEX. Addition of 5 or 70
mg/8. of HEX to sewage effluent inoculated with Salmonella typhosa was also
found to be more effective than similar concentrations of chlorine in reduc-
ing total bacterial count, coliforms and S. typhosa (Cole, 1954). Yowell
(1951) also reported in a patent application that HEX has antibacterial
properties; standard phenol coefficients for E.. typhus (sic) and Staphy-
lococcus aureus were 25 and 33, at 21 and 23 ppm of HEX, respectively.
These findings indicated that concentrations of HEX at or slightly above its
aqueous solubility limit were toxic to several types of pathogens.
In contrast, tests with other microorganisms have shown some ability to
withstand HEX exposure. Twenty-three strains of organisms (type unspeci-
fied), when added to aqueous medium containing HEX at 1000 mg/S., were able
to metabolize the compound to a varying degree. Analysis of the medium
after 14 days indicated a HEX removal of 2-76%, depending on the organism
used (Thuma et al., 1978).
Rieck (1977a) found no effects on natural populations of bacteria,
actinomycetes and fungi after 24 days incubation of a sandy loam soil treat-
ed with 1 or 10 pg/g (dry weight) HEX. He concluded that no significant
detrimental effects on microbial populations would result from treatment of
soils with these levels of HEX.
The effects of HEX on three ecologically important microbial processes
were recently reported by Velsicol (Butz and Atallah, 1980). Results on
cellulose degradation by the fungus Trichoderma longibrachiatum indicated
that a suspension of HEX inhibited cellulose degradation at a concentration
of 1 mg/a. and higher in a liquid medium. The calculated 7-day EC was
1.1 mg/a.. Extrapolations for the 1- and 3-day EC™ values were reported
to be 0.2 mg/S.. The decrease in toxicity in the 7-day period was attri-
buted to adaptation by T. longibrachiatum.
6-9
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HEX Inhibited anaerobic sulfate reduction by Desulfovibrio desu.lf.uricans.
when present in suspension in a liquid medium. Following a 3-hour contact
period, growth inhibition was observed at HEX concentrations of 10-100
mg/8., and no growth was evident at 500 and 1000 mg/a.. Similarly, growth
inhibition was observed at 1 and 10 mg/a. following a 24-hour contact
period, and no growth was evident at 50-1000 mg/8.. HEX was considered
slightly toxic to D_. desulfuricans (Butz and Atallah, 1980).
A third study by the same investigators {Butz and Atallah, 1980) focused
on the effects of HEX on urea ammonification by a mixed microbial culture in
moist soil. The results indicated that HEX concentrations of 1-100 pg/g
(dry weight) were not toxic to soil organisms responsible for urea ammonifi-
cation. EC5Q increased from 104 pg/g at 1 day to 1374 pg/g at 14
days. The authors suggested that the low toxicity and its decrease over
time in this experiment may have'been due to adsorption of the toxicant onto
soil particles, as well as to potential adaptation by the organism. Soil
adsorption may also account for the lack of toxicity in the test by Reick
(1977a).
6.3. EFFECTS ON TERRESTRIAL VEGETATION
In a patent application, HEX was reported to be nontoxic to plants in
concentrations at which it was an effective fungicide (Yowell, 1951). Test
solutions were prepared by adding HEX at various proportions to attaclay and
a wetting agent, and the mixture was then mixed with water. The concentra-
tions of HEX applied to plants as an aqueous spray were 0.1, 0.2, 0.5 and
1.0%. Slight injury (unspecified) to Coleus blumei was reported at 1.0%
HEX, whereas lower concentrations were not harmful. Similarly, HEX was
added to horticultural spray oil and an emulsifier at various proportions
6-10
-------
96-hour LC5Q values ranging from. 59-180
and then mixed with water. The concentations of HEX In the prepared spray
were 0.25 and 0.5%. No injury to C. blumel was observed at these concentra-
tions.
6.4. EFFECTS ON WILDLIFE
No data were available on the effects of HEX on amphibians, reptiles or
birds, or on mammals other than those typically utilized in laboratory
testing.
6.5. SUMMARY
The toxicity of HEX to several forms of aquatic life has been demon-
strated. The freshwater cladoceran Daphnia magna gave 48-hour LC™ values
of 39 and 52 yg/8. in static tests (Buccafusco and LeBlanc, 1977; Vilkas,
1977). Freshwater fish species tended to be slightly more tolerant, with
(Henderson, 1956; Bucca-
fusco and LeBlanc, 1977; Podowski and Khan, 1977). However, when fathead
minnow fry (larvae) were tested in a flowing system, a value of 7 yg/s.
was obtained (Spehar et al., 1977, 1979).
Saltwater crustaceans were of similar sensitivity as D. magna in static
tests; 96-hour LC5Q values for two shrimp species were 32 and 42 yg/Sl,
while a polychaete was more resistant with a value of 371 yg/8.. How-
ever, a flowthrough test with mysid shrimp gave a 96-hour LCfn of 7
bu
pg/s,. Three saltwater fish species all had static LC5Q values within
the range of 37-48 Pg/a. (U.S. EPA, 1980a).
The chronic HATC for the fathead minnow, based on a 30-day early
lifestage test, was between 3.7 and 7.3 vg/fc, as was the acute LC™
(Spehar et al., 1977, 1979). Thus no cumulative toxic effect was observed,
and there was also no accumulation of residues of HEX. Fish growth was
unaffected in this test. On the other hand, a 28-day chronic test with
6-11
-------
mysld shrimp gave an HATC between 0.30 and 0.70 yg/!t, well below the
acute value of 7 yg/fi. for this species. Both survival and fecundity
were reduced by toxicant exposure (U.S. EPA, 1981).
In the only tests conducted with aquatic plants, two of four saltwater
unicellular algal species tested were of comparable sensitivity as crusta-
ceans, with 7-day EC™ values of 3.5 and 6.6 pg/a, respectively. The
other species were somewhat more tolerant (Walsh, 1981).
In general, flowing toxicant concentrations produced a greater response
than static concentrations, and measured concentrations were found to be
about one-half of nominal concentrations. Thus static tests, all based on
nominal concentrations, probably underestimated HEX toxicity. Tests ini-
tiated with other than newborn animals could also have underestimated the
toxic response of natural populations exposed to HEX.
In aqueous media, HEX is toxic to many microorganisms at nominal concen-
trations of 0.2-10 mg/s., or levels substantially higher than those needed
to kill most aquatic animals or plants (Cole, 1953, 1954; Yowell, 1951).
Some microorganisms are able to withstand exposures as high as 1000 mg/8,
(Thuma et al., 1978). HEX appears to be less toxic to microorganisms in
soil than in aquatic media, probably because of adsorption on the soil
matrix (Rieck, 1977a; Butz and Atallah, 1980).
Sufficient information is not available to determine the effects of HEX
exposure on terrestrial vegetation or wildlife, although data from labora-
tory studies summarized in the following sections could be used to estimate
effects on wild mammals.
6-12
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7. TOXICOLOGY AND HEALTH EFFECTS
7.1. PHARMACOKINETICS
7.1.1. Absorption, Distribution, Metabolism and Excretion.
7.1.1.1. ORAL — Mehendale (1977) studied the absorption, metabolism,
excretion and tissue distribution of HEX In 225-250 g male Sprague-Dawley
rats. A single dose of 6 mg/Jcg 14C-HEX In corn oil was given by oral
gavage. The animals were maintained in metabolism cages for 7 days. Urine
and fecal samples were collected daily. After 7 days, the rats were sacri-
ficed and the amount of radiolabel in major organs, urine and powdered feces
was determined. Ten percent of the radiolabel was recovered in the feces
and 33% in the urine during the 7 days while only trace amounts were found
in the liver, kidney and other major organs. Since >50%'of the administered
dose was not accounted for, the author speculated that the respiratory tract
was the major route of excretion for orally administered HEX. Subsequent
studies that are reviewed later in this chapter, in which exhaled air and
lung tissues were analyzed for 14C activity, have shown that this is not
the case. Another interpretation of these results is that HEX and/or its
metabolites were volatilized and lost during sample preparation, i.e.,
powdering of the feces before analysis (Whitacre, 1978). Mehendale also
studied the subcellular distribution of radiolabel in cellular fractions of
rat liver and kidney following oral administration of i/JC-HEX. In both
organs, the majority of radiolabel was located in the cytosol. Specific
metabolites and the metabolic form of the radiolabel in various fractions
and samples were not identified in these studies.
In 1979, Dorough studied the absorption, tissue distribution and excre-
tion of HEX in male and female Sprague-Dawley rats (200-250 g) and mice
7-1
-------
(strain not specified; 25-30 g). The animals were divided into two compar-
able groups and were given a single oral dose of 2.5 or 25 mg/kg of
14C-HEX (corn oil vehicle). The animals were placed in metabolism cages
equipped with a trap to collect expired organocompounds and a trap to
collect expired carbon dioxide. Less than 1% of the radiolabel was trapped
in the expired gases over a 3-day period. The pattern of results for other
routes of elimination was similar in both sexes of each species. Therefore,
this study disproves Mehendale's (1977) speculation that the compound was
mainly excreted by exhaled air. After 3 days, animals given 2.5 mg/kg
excreted an average of 68% of the radiolabel in the feces and 15% in the
urine while animals given 25 mg/kg excreted an average of 72% of the radio -
label in the feces and 14% in the urine. Total recovery of radiolabel was
between 83 and 86%. Thus, 14-17% of the radiolabel was not accounted for in
this study. In addition, Dorough fed 1, 5 or 25 ppm HEX to rats and mice
for a maximum of 30 days. During this study, 54-70% of the radiolabel was
excreted in the feces and 6-12% in the urine. The total cumulative recovery
of radiolabel ranged between 63 and 79% with average values of 72% recovery.
This means that an average of 28% of the radiolabel was left unaccounted.
Metabolites were not identified in these studies.
In a study by Yu and Atallah (1981), male and female Sprague-Dawley rats
(240-350 g) were given a single dose of 3 or 6 mg/kg 14C-HEX in 0.5 ms.
corn oil by gavage. Radioactivity appeared in the blood (taken from the
tail) within 30 minutes, reached a maximum value at 4 hours, and then gradu-
ally decreased. Within 48 hours, 70% of the radiolabel was excreted in the
feces and 17%. in the urine while only a total of 2.8% was retained in the
liver, kidneys, fat, muscle, brain and heart. Thus, -90% of the radiolabel
I
was recovered in this study. Metabolites were not identified by various
7-2
-------
chromatographic methods although the authors stated that no unchanged HEX
(I.e., HEX that was not metabolized or bound to other molecules). ;was found
in the excreta or tissues examined after killing the animals. When HEX was
incubated iji vitro with the contents of rat gut or with fecal homogenates,
the estimated half-life of unchanged HEX was 10.1 hours and 1.6 hours,
respectively. The addition of HgCl to fecal homogenates and gut contents
resulted in decreases in the degradation; rates of HEX. On this basis, the
authors concluded that HEX was poorly absorbed in the gut and that microbial
action was responsible for the metabolis.m of HEX.
7.1.1.2. DERMAL — There were ,no pharmacokinetic studies of HEX,
involving the dermal route, found ,in our literature survey. While no
quantitative studies of HEX absorbed through the skin were found, studies
have been reported in which discoloration of the skin was observed following
the dermal application of HEX (Treon ,e,t al., 1955; IROC, 1972). Although
this does not prove absorption, toxic response leading to death was observed
in several cases. This fact would suggest that HEX is possibly absorbed
trans-dermally into the systemic circulation. These studies are discussed
in greater detail later in this chapter.
7.1.1.3. INTRAVENOUS -- Mehendal.e (1977) studied biliary excretion
following Injection of 1 yd HEX (Svifflole vehicle not identified) into
the femoral vein or artery in Sprague-Oawley rats whose common bile duct had
been cannulated. There was biexponential decay of radiolabel from the blood
with estimated half-lives of ~5 and 60 minutes. Approximately 9% of the
radiolabel was excreted in the bile in 1 hour.
Yu and Atallah (1981) administered 0.73 mg/kg ^C-HEX (10.6 mCi/mmole
in 0.3 ms, of 20% Emulphor® EL 0620 vehicle in saline solution) intraven-
ously into the lateral caudal vein, of Sprague-Dawley rats. Within 48 hours,
7-3
-------
21% of the radiolabel was excreted in the feces and 18% In the urine while a
total of -28% of the radiolabel remained 1n the liver, kidneys, fat, muscle,
brain and heart. Metabolites were not identified in this study and only 67%
of the dose was recovered.
7.1.1.4. INHALATION — In 1980, Dorough studied the absorption and
fate of inhaled HEX in female Sprague-Dawley rats (175-250 g). Animals were
exposed to vapors of 14C-HEX over a 1-hour period to achieve desired
dosing of ~24 yg/kg body weight (both measured and from theoretical calcu-
lations). Considerable difficulty was experienced in maintaining the
desired concentration of HEX throughout the exposure period. Approximately
69% of the radiolabel was recovered, with 13% in the body tissues, 23% in
the feces, and 33% in the urine. Less than 1% of the inhaled radiolabel was
recovered in the expired air following exposure.
These results were confirmed in a study by Lawrence and Oorough (1982)
in which female Sprague-Dawley rats (175-225 g) were exposed in a specially-
designed facemask system for 1 hour to concentrations ~24 yg/kg 14C-HEX.
Retained doses received by rats during inhalation exposures ranged from 1-40
yg/kg bw, but the retention of inhaled 14C-HEX was not influenced by the
quantity received within this range of doses (Lawrence and Dorough, 1982).
Following exposure, <1% of the recovered radiolabel was expired as organo-
compounds and no detectable 14C-carbon dioxide was expired. The trachea
and lungs contained the highest levels of radiolabel with 107 and 74.5 ng
equ1valent/g tissue, respectively. Radioacarbon remaining In the body after
72 hour represented 12.9 and 31.0% of the inhalation and i.v. treatments.
In their experiment studying the effects of HEX exposure on the Clara
cells of monkeys and rats, Rand et al. (1982b) hypothesized that HEX vapor
Inhalation interferes with metabolism by the peroxidation of membrane-bound
7-4
-------
unsaturated llpids. These researchers suggest that there would be a de-
crease 1n the production of pulmonary cytochrome P-450, resulting In a
modification of the microsomal enzyme system of the smooth endoplasmic
reticulum to metabolize foreign compounds. This resultant biochemical
action then changes the morphology of the secretory glands.
7.1.1.5. COMPARATIVE STUDIES — In the inhalation studies of El
Career et al. (1983), Dorough (1980) and Lawrence and Dorough (1982), and
groups of rats were given HEX by oral gavage and by intravenous (i.v.)
injection in order to compare the results for the three routes of admini-
stration. Tables 7-1, 7-2 and 7-3 summarize the results of these three
studies. The tissue distribution was different for the three routes of
administration. The results of the oral studies compare quite favorably
with the studies of Dorough (1979) and Yu and Atallah (1981).
El Dareer et al. (1983) completed a HEX disposition comparison study
using male Fischer 344 rats for the National Toxicology Program (NTP). HEX
(95-99% pure) was administered orally (4.1 and 61 mg/kg), intravenously
(0.59 mg/kg) and by inhalation (1.0 and 1.4 mg/kg). The disposition of
radioactivity from ^C-HEX in rats dosed by various routes is summarized
in Table 7-1. In this experiment after oral doses, most of the radioactiv-
ity appeared in the urine and feces within 72 hours. In comparing the oral
with the i.v. route, the percentages found in the urine and feces were
smaller with a comparitively large proportion of the radioactivity remaining
in the tissues, mostly in the liver and carcass. The rats exposed to the
vapor had a higher percentage remaining in the tissues as compared with oral
dosing, but lower in comparison with the i.v. route. Hetabolities were not
identified 1n the study.
7-5
-------
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TABLE 7-2
Fate of Radiocarbon Following Oral, Inhalation and
Intravenous Exposure to 1"C-HEX in Rats
Expressed as Percentage of Administered Dose3
Oralb
Cumulative Percent of Dose
Intravenous0
Inhalationd
Urine
Feces
Urine
Feces
Urine
Feces
Body
Total Recovery
22.2
62.2
24.0
67.7
24.4
68.2
0.2
92.8
+ 1
± 8
+ 1
± 5
+ 1
f 5
+ 0
+ 4
._
.8
.0
.9
.1
.9
.1
.2
.7
24-Hour
18
21
48-Hour
20
30
72-Hour
22
47
15
85
.3
.1
.7
.4
.1
.4
.7
.2
+ 5
+ 7
+ 5
i 1
+ 5
+ 1
+ 7
+ 4
.2
.1
.6
.7
.7
.9
.8
.8
29
17
32
21
33
23
12
69
.7
.0
.5
.0
.1
.1
.9
.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 ma. corn oil at 7 vg/kg body weight
C0oses administered in 0.2 ms, 10:4:1 saline:propylene glycolrethanol by
injection into the femoral vein at 5 yg/kg body weight
dOoses administered as vapors over a 1-hour exposure period to achieve doses
of -24 yg/kg body weight.
7-7
-------
TABLE 7-3
Distribution of HEX Equivalents3 in Tissues and Excreta of Rats
72 Hours After Oral, Inhalation and Intravenous Exposure to 14C-HEXb,c
Sample
Oral Dose
(6 mg/kg)d
Inhaled Dose
(-24 pg/kg)
Intravenous Dose
(10 yg/kg)
Trachea
Lungs
Liver
Kidneys
Fat
Remaining carcass
ng/g of Tissue
292
420
539
3272
311
63
+
7
4-
4-
4-
4-
170
250
72
84
12
40
107.
71.
3.
29.
2.
1.
0
5
6
5
8
3
4- 65
4- 55
4- 1
+ 20
4- 0
+ 0
.0
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.9
.2
.4
.6
3
14
9
22
2
0
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.9
.6
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.3
.5
+
4_^
4-
4-
4-
4-
1
1
1
0
0
0
.7
.1
.1
.6
.2
.1
Percent of Dose
Whole
Urine
Feces
Total
Body
Recovery
2.8
15.3
63.6
81.7
4- 1
+ 3
+ 8
4- 6
.1
.3
.5
.7
12
33
23
69
.9 4-
.1 +
.1 +
.1 4-
4
4
5
9
.7
.5
.7
.6
31
22
31
84
.0
.1
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.6
+ 7.8
+• 5.7
+ 1.9
+ 4.6
aOne HEX equivalent is defined as the amount of radiolabel equivalent to
one nanogram of HEX based on the specific activity of the dosing solution.
''Source: Adapted from Dorough, 1980 and Lawrence and Dorough, 1982
CA11 values are the Mean i S.D. of three replicates.
dNote that the oral dose was 250 and 600 times that of the inhaled and
i.v. doses, respectively. That was necessary since residues were not
detected in individual tissues of animals treated orally at doses of 5-25
vg/kg.
7-8
-------
This study (El Dareer et al., 1983) confirms Dorough's studies 1n that
the major routes of elimination are fecal and urinary. Little radioactivity
appeared as 14CO_ or as other volatile compounds. Since little radio-
activity was detected In the exhaled air, the respiratory tract is not a
substantial route of elimination of HEX. This substantiates the findings of
Lawrence and Oorough (1981) and negates the Mehendale (1977) conclusion.
The radioactivity found in the urine, feces and body after 72 hours were
similar to Lawrence and Dorough (1981) with the exception of a higher
percentage being found in the feces than in the urine.
Several observations have been made during the development and peer
review of this document. During inhalation and the passage of HEX through
the lung tissue to reach the blood, metabolism to water-soluble compounds
may occur and HEX would be eliminated through the kidneys. In contrast, an
i.v. dose may be bound unchanged to blood components and remain attached
until reaching the liver, whereupon it may be displaced and become associ-
ated with the liver tissue. However, Lawrence and Dorough (1982) still
conclude that regardless of the route of HEX administration, damage to the
lungs occurs and in all cases appears to be the primary cause of death in
the laboratory animals.
7.1.1.6. CONCLUSIONS REGARDING THE FATE OF HEX IN BIOLOGICAL
SYSTEMS — From the data presented in the pharmacoklnetic studies, the
following points can be made regarding the fate of HEX in biological systems;
HEX or its metabolites interact with biological tissues as
indicated by the following:
- high concentrations of HEX are found in the lung and trachea
following inhalation exposure, skin darkens in appearance
when exposed, and HEX Interacts, at a fairly rapid rate,
with gut and fecal homogenates
7-9
-------
HEX 1s not readily absorbed through the gastrointestinal tract
as Indicated by the following:
- there is a high retention of HEX in the fecal contents of
animals dosed orally and there is relatively little bilary
excretion to account for this dose
HEX equivalents are not volatilized and lost in expired air
during the first 72 hours following dosing as indicated by the
following:
- no radiolabelled carbon dioxide and only small amounts of
14C-HEX were found in animals post exposure after dosing
by the pulmonary, i.v. or oral routes
Since the recovery of radiolabel following HEX administration varies
from 43% to >90% in the pharmacokinetic studies reported, a need for a more
thorough study of the pharmacokinetics of HEX by various exposure routes is
evident. A major portion of the radiolabel may be "fixed" to tissues at the
site of administration and missed in routine recovery procedures for pharma-
cokinetics studies. No one has measured the amount of radiolabel retained
by the blood vessel walls or the gastrointestinal epithelial tissues. One
might expect binding to these tissues (as sites of uptake) after i.v. or
oral dose administration.
7.1.2. Summary. Pharmacokinetic studies designed to determine the
absorption, distribution, metabolism and elimination of HEX in rats and mice
have involved the oral, i.v. and inhalation routes of administration of
14C-HEX. The fecal excretion of radiolabel following oral dosing is 2- to
3-fold higher than for i.v. or inhalation administration which indicates
that HEX is not readily absorbed from the gastrointestinal tract. Following
inhalation, considerable radiolabel remained in the lung and trachea indi-
cating that HEX reacts with biological membranes and molecules i_n vivo. HEX
has also been shown to react with the contents of the gastrointestinal tract
7-10
-------
Ijn vitro. Since up to 57% of the radiolabel has not been accounted for even
in studies in which considerable effort has been made to recover all of the
radiolabel, HEX might possibly react with biological membranes and molecules
at all sites of administration or membrane transport. A number of studies
have been conducted to elucidate the whereabouts of HEX in body tissues
after exposure by different routes. However, since the 14C-labelled
compound used in these studies did not allow for the identification of any
of the metabolites, little, as yet, is known about the fate of HEX or Its
metabolites.
7.2. MAMMALIAN TOXICOLOGY
7.2.1. Acute Tox1c1ty. The acute toxicity of HEX is summarized in
Table 7-4. A complete toxicity table is also presented In Appendix 1.
7.2.1.1. ACUTE ORAL TOXICITY — Treon et al. (1955) conducted a
series of oral toxicity studies using female rabbits (strain unspecified)
and Carworth rats of both sexes. HEX was administered as a 5% solution in
peanut oil by oral gavage. The oral LD5Q for female rabbits was deter-
mined to be -640 mg/kg. The oral LD™ for male and female rats was -510
mg/kg and 690 mg/kg, respectively. In 1968, IROC determined the oral L0g
for albino rats to be 926 mg/kg for HEX given in corn oil by oral gavage.
In more recent studies, Dorough (1979) reported the oral LD5Q for male and
female Sprague-Dawley rats to be -651 mg/kg and for male and female mice
(strain unspecified) to be greater than 600 mg/kg. Thus, HEX is moderately
toxic when given orally. Based on FIFRA guidelines (40 CFR 162.10) HEX,
when administered orally to young adult experimental animals, would be
classified in Toxicity Category III. In addition, Southern Research Insti-
tute (SRI, 1980a) reported the oral LD5Q for male and female weanling
B C F mice to be 680 mg/kg. Also, SRI (1980a) reported the oral
Do!
7-11
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LDg0 for weanling Fischer 344 rats to be 425 mg/kg for males and 315 mg/kg
for females.
7.2.1.2. ACUTE DERMAL TOXICITY — Treon et al. (1955) reported the
dermal LD,. in female rabbits (strain unspecified) to be 780 mg/kg while
IROC (1972) reported the dermal LD™ in albino rabbits (strain unspeci-
fied) to be <200 mg/kg in males and to be 340 mg/kg in females. These data
would place HEX, when applied dermally, in Toxicity Category II.
7.2.1.3. ACUTE INHALATION TOXICITY — Treon et al. (1955) reported a
3.5-hour LC5Q of 3.1 ppm for Carworth rats of both sexes. Rand et al.
1982a reported a 4-hour LC5Q of 1.6 ppm for male Sprague-Dawley rats and
3.5 ppm for female rats. Treon et al. (1955) determined the 3.5-hour LC5Q
to be 5.2 ppm in female rabbits, 2.1 in male and female mice, and 7.1 in
male and female guinea pigs. These concentrations are in the range of
0.02-0.08 mg/S. for HEX vapor for rats and mice which would place HEX, when
inhaled, in Toxicity Category I.
7.2.1.4. EYE IRRITATION -- IRDC (1972) tested HEX for eye iritation
by instilling 0.1 mil HEX into the eyes of New Zealand white rabbits for 5
minutes or 24 hours before washing. All rabbits died on or before the 9th
day of the observation period. HEX is a strong eye irritant and would be in
Toxicity Category I based on ocular exposure.
7.2.1.5. DERMAL IRRITATION — Treon et al. (1955) reported HEX to be
a primary skin irritant in rabbits (s'train unspecified) at a dose level of
250 mg/kg. In 1972, IRDC reported HEX in New Zealand white rabbits to be a
dermal irritant based upon edema observed following application of 0.5 ma
HEX. In this study, intense discoloration of the skin was noted. These
data would place HEX in Toxicity Category II for dermal irritation. In the
7-14
-------
Treon study (1955), monkeys (strain unspecified) were also tested and dis-
coloration of the skin was noted even at low doses (0.05 ma of 10% HEX).
7.2.1.6. SUMMARY ~ The acute oral toxicity of HEX has been studied
In rats, rabbits and mice. The oral L0,-0 for adult animals is >500 mg/kg
which places HEX in Toxicity Category III. The acute dermal toxicity of HEX
has been studied in rabbits and, because <50% of the animals died at the
tested dose, the dermal LD5Q is >200 mg/kg which places HEX in Toxicity
Category II. The acute inhalation toxicity of HEX has been studied in rats,
rabbits, guinea pigs and mice. In rats and mice, the 3.5-4.0 hour LCrn
bu
for HEX is <0.2 mg/9, which places HEX in Toxicity Category I. In compari-
son, the pathological effects are observed in the lung no matter which route
of administration of HEX is used. In addition, HEX is a severe eye, skin
and pulmonary irritant.
7.2.2. Subchronlc Toxicity.
7.2.2.1. SUBCHRONIC ORAL TOXICITY --
7.2.2.1.1. Range-Finding Studies — Using small range-finding tests
Litton Bionetics (1978b) determined the oral LDg of HEX in CO-1 mice to be
76 mg/kg. However, when this expected LO was administered to mice for 5
consecutive days, all mice (24) died within the 5-day period. In a range-
finding study using groups of 5 male and 5 female Fischer 344 rats, SRI
(1980a) reported no mortality at doses of 25, 50 or 100 mg/kg when given 12
doses in 16 days. At 200 mg/kg and using the same dosing schedule, 5 of 5
males and 4 of 5 females died, and at 400 mg/kg, 5 of 5 males and 4 of 5
females died during the study. In .the same study, 86C3F, mice died when
given doses of 400 or 800 mg/kg but not at doses of 50, 100 or 200 mg/kg.
Both rats and mice exhibited pathologic changes of the stomach wall in all
but the lowest dose level.
7-15
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7.2.2.1.2. Studies 90 Days or Longer 1n Duration — The subchronic
toxlcity of HEX Is summarized 1n Table 7-5. Subchronic toxlclty studies In
B6C3F, mice and Fischer 344 rats have been conducted by SRI (1981a,b)
under contract with the National Toxicology Program (NTP). In the mouse
study (1981a), dose levels of 19, 38, 75, 150 and 300 mg/kg HEX (94.3-97.4%)
1n 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, the
liver was enlarged. Toxic nephrosis in females at doses of 75 mg/kg and
higher was characterized by lesions in the terminal portions of the con-
voluted tubules, with basophilia in the inner cortical zone and cytoplasmic
vacuolization. However, male mice at this level and higher did not show
these effects. Dose levels of 38 mg/kg HEX and above caused lesions in the
forestomach, including ulceration in both males and females. The no
observed adverse effect level (NOAEL) in mice for HEX was 19 mg/kg and the
lowest observed effect level (LOEL) was 38 mg/kg.
In the rat study (SRI, 1981b), dose levels of 10, 19, 38, 75 and 150
mg/kg HEX in corn oil were administered by gavage to groups of 10 male and
female F344 rats. At the 38 mg/kg dose and higher levels, mortality and
toxic nephrosis were noted in both males and females. The male rats treated
at the 19 mg/kg dose level showed no highly abnormal effects while female
rats exhibited lesions of the forestomach. Such lesions were observed in
males at 38 mg/kg or higher levels. There was a dose-related depression of
body weight gain relative to the controls. The NOAEL in rats for HEX was 10
mg/kg and the LOEL was 19 mg/kg.
A summary of the results of these two experiments appears in Table 7-6.
Based on these studies, a maximum tolerated dose (MTD) of 38 mg/kg for mice
7-16
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and 19 mg/kg for rats was recommended by SRI to NTP for a chronic toxicity
study.
7.2.2.2. SUBCHRONIC DERMAL TOXICITY --
7.2.2.2.1. Range-Finding Study — In a Russian study, Naishteln and
Lisovskaya (1965) studied the effects of HEX applied to the shaved area of
the skin of rabbits (strain unspecified) daily for 10 days. According to
the authors, no effects were noted in control and test animals given dally
doses of 0.5-0.6 ml of a 20 mg/9, solution of HEX.
7.2.2.3. SUBCHRONIC INHALATION TOXICITY —
7.2.2.3.1. Range-Finding Studies — Rand et al. (1982a) conducted a
range-finding study in which groups of 10 male and 10 female Sprague-Oawley
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 moribund after 5-7 exposures. These rats had
dark red eyes, labored breathing, and paleness of extremities. No mortal-
ities were noted in the other exposure groups; however, the males in the
0.11 and 0.5 ppm groups lost weight during the study and alterations In
liver weight and pathology were noted. The NOAEL for HEX exposure was 0.022
ppm and the LOEL was O.Tl ppm.
7.2.2.3.2. Studies 90 Days or Longer in Duration — Fourteen-week
inhalation studies in 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, 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 and 0.20 ppm HEX. In monkeys,
there were no mortalities, adverse clinical signs, weight gain changes,
pulmonary function changes, eye lesions, hematologic changes, clinical
7-20
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chemistry abnormalities or hlstopathologlc abnormalities at any dose level
tested. Thus, the no observed effect level (NOEL) for monkeys was 0.2 pptn
HEX and the LOEL was not determined.
Male 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 in hemoglobin concentration arid erythrocyte count in 0.01 ppm
males, 0.05 ppm females, and 0.20 ppm males and females. There were small
but not statistically significant changes in mean liver weight of all treat-
ment groups and similar changes in the kidneys of all treated males. There
were no treatment-related abnormalities in gross pathology or histopatho-
logy. On this basis, the NOEL in rats was 0.2 ppm HEX; the LOEL was not
established.
In the other study by Rand and coworkers {Rand et a!., 1982b), no
ultrastructural changes were observed that could be attributed to the inhal-
ation of HEX vapor in exposed monkeys. Exposure was identical to that of
the previous study (Rand et al., 1982a). This study took an in-depth look
at the Clara cells and the results show 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 con-
trols. According to some researchers (Evans et al., 1978), Clara cells
respond to injury by regression to a more primitive cell type. Rand et al.
(1982b) noted that some of the ultrastructural changes in the exposed
animals resemble those of the Evans study. It is not known what effect
these changes might cause. The Clara cell contributes important materials
to the extracellular lining of the peripheral airways, and if this effect
from HEX vapors causes the content of the contributed material to be
changed, then the extracellular lining may be altered and breathing may be
7-21
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subsequently Impaired (Rand et alV, '1'9'8'2'b). This observation coincides With
those of other researchers (Dorough, 1979, 1980; Lawrence and Dorough, 1981,
1982). Furthermore, in the inhalation experiments with HEX, researchers
have noted occasional statistically significant increases in hemoglobin and
red blood cells of rats, which may be manifestations of the impairment of
respiratory functions.
7.2.2.4. SUMMARY — The subchronic toxicity of HEX has been studied
in rats and mice following oral gavage and in rats and monkeys following
inhalation exposure. In oral studies, rats and mice exhibited decreased
body weight gain, lesions of the forestomach, and toxic nephrosis. Female
mice also exhibited enlarged livers. The oral LOEL was 38 mg/kg for mice
and 19 mg/kg for rats". In the inhalation studies, no abnormalities were
observed in monkeys at doses as high as 0.2 ppm HEX for 6 hours over 14
weeks. No statistically significant changes were noted in blood parameters,
and in kidney and liver weight in rats at all doses tested (range 0.01-0.2
ppm HEX). Thus, the NOEL in both rats and monkeys was 0.2 ppm; no LOEL was
established.
7.2.3. Chronic Toxicity.
7.2.3.1. CHRONIC ORAL TOXICITY -- A chronic oral toxicity study of
HEX being conducted by SRI for the National Toxicology Program was termi-
nated in April 1982 because inhalation was determined to be the more rele-
vant route of exposure. No other chronic oral toxicity data were available
for this report.
7.2.3.2. CHRONIC DERMAL TOXICITY -- There were no chronic dermal
toxicity studies found in the available literature.
7.2.3.3. CHRONIC INHALATION TOXICITY -- Treon et al. (1955) exposed
guinea pigs, rabbits, rats and mice to a concentration of 0.33 ppm HEX for 7
7-?2
-------
hours/day, 5 days/week for 25-30 exposures, Guinea pigs survived 30 expo-
sures; however, rats and mice did not.survive ,5 exposures .and 4 of 6 rabbits
did not survive 25 exposures. Using a lower concentration (0.15 ppm HEX),
guinea pigs, rabbits and rats survived 150 seven-hour exposure periods (7
months). This level was too high for a chronic study 1n mice since 4/5
animals did not survive. The rats, guinea pigs and rabbits tolerated 0.15
ppm and did not exhibit any treatment-related effects. Thus, the NOEL for
rats, guinea pigs and rabbits and the LOEL for mice was 0.15 ppm HEX. The
NOEL for mice was not established while the LOEL for rats, guinea pigs and
rabbits was 0.33 ppm HEX.
A 30-week chronic inhalation study of technical grade HEX in rats, 96%
pure with hexachlorobuta-1,3-diene and octachlorocyclopentene as impurities,
was conducted by Shell Toxicology Laboratory {0. Clark et al., 1982). Four
groups of 8 male and 8 female Wistar 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. Females in the high and medium dose groups had
lower body weights at the end of the recovery period as compared with the
controls. At 0.5 ppm, there were pulmonary degenerative changes noted in
both sexes although the males were affected more severely. At the highest
dose, there were mild degenerative changes in the liver and kidneys at 30
weeks in a few rats and kidney weights were significantly increased 1n the
females. After 30 weeks of study, there was no biologically significant
toxicity noted in animals exposed to .concentrations of 0.05 or 0.1 ppm HEX
7-23
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{D. Clark, et al., 1982). Thus, the NOEL 1n rats exposed to vapors of HEX
was 0.05 ppm; the LOEL was 0.1 ppm based on body weight, organ weight, and
hlstopathology data.
A chronic inhalation study of HEX has been scheduled by the National
Toxicology Program (Abdo, 1983).
7.2.3.4. SUMMARY — The chronic effects of HEX have been studied
primarily by inhalation exposure. No oral studies and one under-reported
dermal study were located for this review. The inhalation toxicity of HEX
has been evaluated in rats, mice, rabbits and guinea pigs. Four of five
mice did not survive exposure to 0.15 ppm HEX, while the other species did
not show effects following 150 seven-hour exposures to 0.15 ppm. In a more
recent study, chronic degenerative changes in the lung, liver and kidneys
were noted in rats exposed to 0.5 ppm HEX and the NOEL for rats was 0.05 ppm
HEX. A 2-year inhalation bioassay has been scheduled by the National
Toxicology Program to begin in 1984 (Abdo, 1983).
7.3. MUTAGENICITY
7.3.1. Mutagenicity. Goggelman et al. (1978) found that HEX was not
mutagenic before or after liver microsomal activation at 2.7xlO~3 M in an
£. coli K,2 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 cytotoxic to E_. coli. A previous report from the same laboratory
(Greim et al., 1977) indicated that HEX was also not mutagenic in S. typhi-
murium strains TA1535 (base-pair mutant) or TA1538 (frame shift mutant)
after liver microsomal activation; however, no details of the concentrations
tested were given. Although tetrachlorocyclopentadiene is mutagenic in
these systems, probably through metabolic conversion to the dienone, it
appears that the chlorine atoms at the C-l position of HEX hindered metab-
olic oxidation to the corresponding acylating dienone {Greim et al., 1977).
7-24
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A study conducted by Industrial Bio-Test Laboratories (IBT, 1977) also
suggests that HEX Is not mutagenic in S. typhimurium. Both HEX and Us
! I i "
vapors were tested with and without metabolic activation. The vapor test
was done In desiccators with only the TA-100 strain of S. typhimurium. It
is not clear from the presented data of the test with the vapors that suf-
ficient amounts of HEX or adequate times of exposure were used. Exposure
times of 30, 60 or 120 minutes were studied. Longer exposures in the
presence of the HEX vapors may be necessary for observation of a potential
mutagenic effect.
At concentrations of up to 1.25xlO~3 yg/mi in the presence of an S-9
liver activating system, HEX was not mutagenic in the mouse lymphoma muta-
tion assay. Mutagenicity could not be evaluated at higher concentrations
because of the cytotoxicity of HEX (Litton Bionetics, Inc., 1978a). This
assay uses L5178Y cells that are heterozygous for thymidine kinase (TK+/-)
and are bromodeoxyuridine (BUdR) sensitive. The mutation is scored by
cloning with BUdR in the absence of thymidine. HEX is highly toxic to these
cells, particularly in the absence of activating system (at 4xlO"s
yjl/ma.) and a positive control, dimethylnitrosamine, was mutagenic at
0.5 yaymi.
Williams (1978) found that HEX (10~6 M) was inactive in the liver
epithelial culture hypoxanthine-guanine-phosphoribosyl transferase (H6PRT)
locus/mutation assay. At 10~s M it 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 (Juodeika, 1983) also did not demon-
strate the mutagenicity of HEX. In S. typhimurium strains TA98, TA100,
7-25
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TA1535 and TA1537, levels of up to 3.3 pg/plate were not mutagenic without
activation and levels of up to 100.0 pg/plate were not mutagenic after
mlcrosomal activation. Higher levels could not be tested because of exces-
sive killing of the bacteria. In the Drosophlla sex-linked recessive lethal
test, HEX was not mutagenic. The doses used in 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 (Litton
B1onet1cs, 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 spermatogenesis (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
resorptions/pregnancy.
7.3.2. Summary. The available evidence suggests that HEX is not a
mutagen. Negative mutagenicity results were obtained in bacteria, liver
epithelial cells, Drosophila, mouse lymphoma cells and in the mouse dominant
lethal test. Furthermore, HEX did not induce unscheduled DNA synthesis in
rat hepatocytes.
7.4. CARCINOGENICITY
7.4.1. In. Vivo Carcinogenicity. Bioassays of HEX for possible carcino-
genicity have not been conducted. However, NTP has scheduled HEX for car-
cinogenlcity testing by the inhalation route in rats and mice (Abdo, 1983).
7.4.2. In. Vitro Carcinogenicity. The ability of HEX to induce morpho-
logic transformation of BALB/3T3 cells In vitro has been studied by Litton
Bionetics, Inc. (1977). The procedure employed by the investigators was
7-26
-------
similar to that of Kakunaga (1973). Evaluation of the carcinogenic activity
i
was based on the following criteria:
The endpoint of carcinogenic activity is determined by the presence
of fibroblastic-like 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 basophilic in character and variable in size.
These changes are not observed in normal cultures, which stain
uniformly.
Assays were performed at levels of 0.0, 0.01, 0.02, 0.039, 0.078 and
0.156 viS./ma.. The cultures were exposed for 48 hours followed * by an
incubation period of 3-4 weeks. The cultures were observed daily. The
selection of test doses was based on previous cytotoxicity tests using a
wide range of HEX concentrations. The doses selected allowed an 80-100%
survival of cells as compared with solvent negative controls. This high
survival rate permitted an evaluation of iji vitro malignant transformation
in cultures treated with HEX as compared with the solvent controls.
3-Methylcholanthrene at a dose level of 3 ug/ma was used as a positive
control. Results indicated that HEX was not responsible for any significant
carcinogenic activity.
7.4.3. Summary. HEX has not been demonstated to be a carcinogen In vitro
in transformation assays using BALB/3T3 cells. In. vivo bioassays have not
been conducted; however, an inhalation bioassay has been scheduled by the
National Toxicology Program.
7.5. TERAT06ENIC AND REPRODUCTIVE EFFECTS
7.5.1. Teratogenlcity. The teratogenic potential of HEX was evaluated in
pregnant Charles River CD-I rats that were administered HEX (98.25%) in corn
7-27
-------
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 ma,/kg/day. Survival was 100%, and there was
no difference in mean maternal body weight gain between dosed groups and
controls. 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 groups, and there were no statistical differences in
malformation or developmental variations compared with the controls when
external, soft tissue and skeletal examinations were performed (IRDC, 1978).
Hurray et al. (1980) evaluated the teratogenic potential of HEX (98%) in
CF-1 mice and New Zealand white rabbits. Mice were dosed at 0, 5, 25 or 75
mg/day HEX by gavage from days 6-15 of gestation while rabbits received the
same dose from days 6-18 of gestation. The fertility of both the treated
mice and rabbits was not significantly different from the control groups.
In the mice, no evidence of maternal toxicity, embryotoxicity or teratogenic
effects was observed. A total of 249-374 fetuses (22-33 litters) were
examined in each dose group.
In rabbits, maternal toxicity was noted at 75 mg/day (diarrhea, weight
loss and mortality), but there was no evidence of maternal toxicity at the
lower levels. There were no embryotoxic effects at any dose level. Al-
though there was an increase in the proportion of fetuses with 13 ribs at 75
mg/day over controls, this was considered a minor skeletal variation, and
the authors concluded that HEX was not teratogenic at the levels tested.
Studies on the teratogenic potential of inhaled HEX were not located in
the review of the scientific literature.
7.5.2. Reproductive Effects. No data were located that addressed the
reproductive effects of HEX.
7-28
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7.5.3. Summary. HEX has been tested for teratogenic potential by oral
gavage In rats, mice and rabbits. No maternal toxicity or teratogenic
effects were noted in rats or mice when HEX was administered on days 6
through 15 of gestation at doses of up to 25 and 75 mg/day, respectively.
Rabbits exhibited maternal toxicity when HEX was administered at 75 mg/day
from days 6 through 18 of gestation and an increase in fetuses with 13 ribs
was also noted at this dose level. The latter was considered to be a minor
skeletal variation by the authors. No maternal toxicity or fetal abnormal-
ities were noted in rabbits at lower doses. HEX therefore does not appear
to be teratogenic by oral gavage in the species and at the doses tested.
HEX was not tested for teratogenicity following inhalation exposures.
7.6. HUMAN EXPOSURE AND HEALTH EFFECTS
7.6.1. Human Exposure. According to a recent NIOSH estimate, 1427
workers are occupationally exposed to HEX {NIOSH, 1980). Velsicol officials
estimate that approximately 157 employees are potentially exposed to HEX in
their production facilities. A summary of monitoring results is presented
in Tables 7-7 and 7-8 for the Velsicol Memphis and Marshall plants, respec-
tively. In addition, acute human exposure has been reported in homes near
waste sites where HEX has been disposed (S. Clark et al., 1982; Elia et al.,
1983).
7.6.2. Health Effects. Very little detailed information is available
concerning the effects of HEX exposure on humans. The odor threshold has
been stated to be 0.00017 ppm, however, there has been great individual
variation. According to the data provided in a study completed by A.D.
Little for Occidental Chemical Corporation, the 100% panel recognition con-
centration was 0.0017 mg/m3 (0.00017 ppm v/v) {Levins, 1980). The study
design and methodology was not given. According to the Material Safety Data
7-29
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Sheet prepared by Hooker Chemical Corporation (1979) and based on animal
studies, 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 mists can result in the secretion of excess fluid in
the lungs; and inhalation or ingestion may cause nausea, vomiting, diarrhea,
lethargy, respiratory impairment and injury to the liver or kidneys.
7.6.2.1. EFFECTS FOLLOWING INCIDENTS OF ACUTE EXPOSURE — Treon et
al. (1955) reported that members of a group conducting toxicity tests
developed headaches when they were accidentally exposed to unknown concen-
trations of HEX, which had escaped into the room when an aerated exposure
chamber was opened.
A well-documented incident of acute human exposure to HEX occurred in
March 1977 at the Morris Forman Wastewater Treatment Plant in Louisville,
Ky. The incident has been described and reviewed in several papers
(Kominsky et al., 1980; Wilson et al., 1978; Morse et al., 1979). The com-
plete details of the original NIOSH Hazard Evaluation and Technical Assis-
tance Report Number TA-77-39 (Kominsky et al., 1978) are available from the
National Technical Information Service (NTIS).
In 1977, the Louisville treatment facility was contaminated with ~6 tons
of HEX and OCCP, a waste byproduct 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 as high as 1000 ppm, and levels in the sewer line ranged
up to 100 ppm. Air samples from the sewer line showed HEX concentrations as
high as 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
7-33
-------
the plant had closed. (The TWA for HEX was 10 ppb In 1977.) During the
cleanup of the contamination, workers using steam attempted to remove an
odoriferous and sticky substance from.the bar screens and grit collection
system. This produced a blue haze which permeated the primary treatment
area. Airborne HEX concentration of the blue haze generated by the cleanup
procedures was reported to be 19.2 ppm (Kominsky et al., 1980).
Both the Center for Disease Control (CDC) and NIOSH sent representatives
to the plant, with each group developing questionnaires seeking Information
on the type and duration of symptoms (Horse, 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
(Horse et al., 1979). A questionnaire was sent to each of these workers and
145 (75%) responded. Workers with complaints of mucous membrane irritation
were given a physical examination, and blood and urine samples were col-
lected for clinical screening by an independent laboratory. Data were also
collected on the exposure levels and symptoms in several individual cases of
acute exposure to the chemical vapors.
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 7-9), 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 27%
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 al., 1978, 1979).
7-34
-------
TABLE 7-9
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: Horse et al., 1978
7-35
-------
While there was difficulty 1n measuring the amount of exposure by the
plant workers, over half of the cleanup crew was monitored. Laboratory
tests showed no significant abnormalities, however, several minimal-to-mild
abnormalities did appear in liver function tests (Kominsky et al., 1980).
These abnormalities are listed in Table 7-10. All of these affected persons
also had physical signs of mucous membrane Irritation. In addition, more
detailed correlation of acute exposure level data to symptomatology was
reported for 9 adults (Kominsky et al., 1980). These data are reviewed in
Table 7-11. The exposure levels could not be estimated accurately because
of prior exposure or because the worker 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 occu-
pants 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. The
authors stated that no significant ambient air concentrations of HEX were
found in these areas (Kominsky et al., 1978). The same types and frequency
of symptoms reported by workers' to be associated with HEX exposure were
reported by residents in the survey which led the authors to suggest that
these symptoms were unrelated to HEX exposure (Morse et al., 1978).
Several papers have documented another similar incident in Hardeman
County, TN. (S. Clark et al., 1982; Meyer, 1983; Elia et al., 1983). While
conducting a ser1oep1demiologic study of the health risks from bacteria and
viruses associated with the treatment of municipal wastewater, potential
human exposure to organic chemicals emitted from the wastewater being
treated at one of the plants in the study was recognized (Elia et al.,
7-36
-------
TABLE 7-10
Abnormalities for 18 of 97 Cleanup Workers
at the Morris Forman Treatment Plant3
Serum
Laboratory Test Normal Range
Glutamate-
Oxalacetate Transaminase 7-40 mU/m».
Serum
Serum
Serum
Alkaline Phosphatase 30-100 mU/ma,
Total Bilirubin 0.15-10 mg/%
Lactate Dehydrogenase 100-225 mU/ma.
Abnormal
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
Results
No.b
5
1
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0
1
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lc
1
aKominsky et al., 1980
bFor individuals with more than one serial blood test, only the most
abnormal result is tabulated.
Associated with serum glutamate-oxalacetate transaminase of 66
U = Units of enzyme activity
7-37
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1983). In 1978, workers at the treatment plant began complaining of acute
symptoms similar to those found in the Louisville plant. Air' and wastewater
* ? <
monitoring was started, analysis of urine specimens, analysis of blood and
liver function tests, and an illness symptom questionnaire were used to
collect data. In the original study design, workers were compared to a
control group from another Memphis treatment plant which does 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, S. 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 differ-
ences among the values obtained for all survey groups.
About the time the wastewater treatment plant study was being performed,
residents of Hardeman County in the general area of the plant began to
complain of foul odors and bad taste in their well, water and asked for an
investigation (Meyer, 1983). In this area lies a 200 acre chemical land
dump which was operated from 1964-1972. In 1978, the U.S. Geologic Survey
(Sprinkle, 1978; Rima, 1979) confirmed the contamination of wells. However,
HEX was not detected in any samples. Urine surveys and liver function
analyses were conducted. Utilizing an unexposed group (those not exposed to
the treatment facility or the contaminated water), a comparison of various
liver enzymes was done (Table 7-12). 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 (Elia, 1983).
7-39
-------
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The Hardeman County studies have been the subject of much scrutiny and
court litigation. At the time of this publication, there has not been any
legal decision rendered. Because of questions concerning the various study
designs used in the studies, very few conclusions can be reached until
further monitoring can be completed. However, these two incidents illus-
trate the possibility of acute exposure at waste treatment facilities
receiving industrial waste.
7.6.2.2. EPIDEHIOLOGIC STUDIES — Mortality studies have been con-
ducted on the workers involved in the production of HEX or formulation of
HEX products. The Shindell report (1980) was a cohort study of workers
employed at the Velsicol Chemical Corporation plant at Marshall, Illinois
between 1946 and 1979. The purpose was to evaluate the vital status of all
former and current employees (>3 months) who were present during the manu-
facture of chlordane. In preparing the cohort, the authors noted the diffi-
culties 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 in the study. The analysis showed no significant differences in
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. popula-
tion (Shindell and Associates, 1980).
Wang and MacMahon (1979) conducted a study on a group 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 mortal-
ity ratio (SMR) of 72, not remarkable for an employed population. The 2
highest SMRs were 134 for lung cancer and 183 for cerebrovascular disease,
but only the latter was statistically significant (p<0.05). The authors
7-41
-------
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.
Shindell and Associates (1981) completed an ep1dem1olog1c study for
Velsicol. The study group consisted of over 1000 employees (93% of the
cohort) of the Memphis, Tn plant for the years 1952-1979, coinciding with
the manufacture of heptachlor. Again, the researchers found no significant
difference in mortality between the control and exposure groups and fewer
deaths in the study group. The investigators report that there was no
excess mortality by job function.
Buncher et al. (1980) studied the mortality of workers at a chemical
plant that produced HEX. The investigators reviewed personnel 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 specific for sex, age and
calendar year. The SMR was 69 which showed 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,
at the most 25 years, reduced the power of the study to detect cancers which
may have a 10-40 year latent period.
7.6.3. Summary. While there is human experience with respect to mortal-
ity, there is only limited Information on the morbidity results in those
exposed to HEX. Acute inhalation produces a high prevalence of headaches
and severe irritation of the eyes, nose, throat and lungs. Dermal contact
can cause severe burns. Epidemiologic studies have generally shown no
7-42
-------
significant differences in mortality between workers exposed to HEX fn the
workplace and the general population. Although, a significant excess of
deaths from cerebrovascular disease was reported in one study, the deaths
showed no consistent pattern with duration of employment or follow-up.
Current human exposure is 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.
7-43
-------
-------
8. OVERVIEW
8.1. EFFECTS OF MAJOR CONCERN
Although minimal quantitative information is available on the effects of
HEX on humans, transient exposure to HEX vapor has been found to cause irri-
tation to the eyes, nose and throat, as well as headaches. The levels of
exposure causing these effects are not well defined but they are at a level
close to the odor threshold, which varies individually and may be as low as
0.00017 pptn (0.0017 mg/m3). There is no information on the long-term
effects of a single exposure or of subchronic exposure. There is no Infor-
mation available on the carclnogenicity of HEX. In. vitro mutagenicity or
transformation tests were negative. The In. vivo mouse dominant lethal assay
was negative at the tested levels. HEX has not been shown to be teratogenic
in studies examining three species.
Considering all of the above facts, the major concerns of HEX exposure
are the toxic effects on the respiratory system when HEX is inhaled. Al-
though the chronic toxicity data are presently limited, the systemic toxic
effects of HEX inhalation have been demonstrated after acute and subchronic
exposure, suggesting that chronic inhalation exposure to low doses of HEX
may have adverse effects.
8.1.1. Principal Effects and Target Organs. Repeated exposure of several
animal species to levels of HEX vapor in the 0.1-0.2 ppm range has been
found to cause pulmonary degenerative changes (Treon et al., 1955; Rand et
al., 1982a,b; S. Clark et al., 1982). Treon et al. (1955) reported mild
degenerative changes in the kidneys, liver, brain, heart and adrenal
glands. Rand et al. (1982), however, did not confirm this and suggested
that the changes found by Treon et al. (1955) were caused by impurities in
the preparation of HEX. Acute exposure by oral and dermal routes also cause
8-1
-------
effects on the respiratory system (Kommineni, 1978; SRI, 1980a). Death from
acute exposure by any tested route appears to be associated with respiratory
failure (Lawrence and Dorough, 1981).
There are Insufficient data to Identify clearly the site most sensitive
to prolonged, repeated exposure to HEX. However, researchers found In com-
paring routes of admlnlstraton that regardless which route was used, damage
to the lungs occurred (Lawrence and Dorough, 1982). When HEX is adminis-
tered orally to animals, the kidneys may be the most sensitive site, since
subchronic dosing of rats and mice was found to cause nephrosis especially
in females (SRI, 1981a,b). Although the oral route may not be significant
in human exposure, the fact that the,kidneys are a possible target organ in
subchronic exposure indicates that low-level, prolonged systemic exposure
from any ambient route may affect the kidneys. The liver has also been an
affected organ as seen in many of the laboratory studies.
8.1.2. Animal ToxicUy Studies Host Useful for Hazard Assessments. The
studies most useful for prediction of hazards are those that use a variety
of dose levels, a variety of species, adequate sample sizes, and display the
full range of effect severity, from no effects through mortality. The major
quantitative goal is to estimate the threshold level for adverse effects,
I.e., the level at or above which adverse effects are observed. In this
regard, the most appropriate studies are those presenting no-observed-effect
levels (NOEL), no-observed-adverse-effect levels (NOAEL) and adverse-effect
levels (AEL), i.e., those dose rates which bracket the threshold level
(Tables 8-1 and 8-2). Dose rates labeled "EL" (for "effect level") are
associated with effects which may or may not be adverse, based upon the data
presented by the researchers. Because dosing regimens varied among studies,
a time-weighted-average (TWA) daily exposure level has been calculated to
8-2
-------
TABLE 8-1
Oral Toxicity Data for Threshold Estimates
An 1 ma 1
Rat
Rat
Mouse
Rat
House
Rat
Exposure
Duration
(days)
10
12
12
91
91
216
Exposure
Levela
10 mg/kg
30 mg/kg
100 mg/kg
25 mg/kg
50 mg/kg
50 mg/kg ,
100 mg/kg
7 mg/kg
14 mg/kg
27 mg/kg
14 mg/kg
27 mg/kg
54 mg/kg
0.2 mg/kg
2.0 mg/kg
Effect
Sever1tyb
NOEL
EL
AEL
NOAEL
AEL
EL
AEL
NOAEL
EL
AEL
NOAEL
EL
AEL
NOEL
EL
Reference
IRDC, 1978
IROC, 1978
IROC, 1978
SRI, 1980b
SRI, 1980b
SRI, 1980a
SRI, 1980a
SRI, 1981a
SRI, 1981a
SRI, 1981a
SRI, 1981b
SRI, 1981b
SRI, 1981b
Naishtein and
Lisovskaya, 1965
aTime-weighted-average daily exposure levels
bDefinitions: NOEL - No-observed-effect level
NOAEL - No-observed-adverse-effect level
EL - Effect level
AEL - Adverse effect level
8-3
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TABLE 8-2
Inhalation Toxlcity Data for Threshold Estimates
Animal
Rat
Rat,
guinea pig
Rat
Monkey
Rat
Rat, rabbit,
guinea pig
Exposure
Duration
(days)
14
42
90
90
210
216
Exposure
Level3
0.004 ppm
0.020 ppm
0.089 ppm
0.069 ppm
0.002 ppm
0.009 ppm
0.036 ppm
0.002 ppm
0.009 ppm
0.036 ppm
0.009 ppm
0.018 ppm
0.089 ppm
0.031 ppm
Effect
Severity13
NOAEL
EL
AEL
AEL
NOAEL
NOAEL
EL
NOAEL
NOAEL
NOAEL
NOEL
EL
AEL
AEL
Reference
Rand et al . ,
Rand et al.,
Rand et al . ,
Treon et al . ,
Rand et al .,
Rand et al .,
Rand et al.,
Rand et al .,
Rand et al . ,
Rand et al.,
Clark et al.,
Clark et al.,
Clark et al.,
Treon et al. ,
1982a
1982a
1982a
1955
1982a
1982a
1982a
1982a
1982a
1982a
1982
1982
1982
1955
aT1me-we1ghted-average dally exposure levels
Definitions: NOEL - No-observed-effect level
NOAEL - No-observed-adverse-effect level
EL - Effect level
AEL - Adverse effect level
8-4
-------
use as a comparison. This value assumes a continuous 24-hour ambient expo-
sure. For example, at the highest actual dose level (0.5 ppm) In the Rand
et al. (1982a,b) studies, the equation would be as follows:
X 6 hours
TWA level . 0.5 ppm x
7 days 24 hours
= 0.089 ppm.
Toxidty from Inhalation of HEX appears to be more severe than that of
oral or dermal exposure and may be the cause of so few Inhalation studies
showing minor effects. Rand et al. (1982a,b) used sufficiently low concen-
trations In a 14-day study on rats and in a 90-day study on rats and monkeys
to elicit effect levels. Clark and researchers (D. Clark et al., 1982)
found that rat groups (18 males and 18 females per group) exposed to HEX at
0.05 ppm {0.009 ppm daily TWA) for 30 weeks showed no effects. However,
Rand et al. (1982a) found their animals had demonstrated some effects at the
same level (0.009 ppm daily TWA) in only 90 days. Treon et al. (1955)
exposed their animals for 216 days and found adverse effects at 0.03 ppm
daily TWA.
Short-term oral studies by IROC (1978) and SRI (1980a,b) provide infor-
mation on toxicity to rats and mice, although the study sizes were small (5
and 10 animals per dose group, respectively). The 90-day study by SRI
(1981a,b) on rats and mice is the only short-term oral study providing no-
adverse-effect levels, and the Naishtein and Lisovskaya (1965) 6-month study
on rats is the only long-term data set giving no-effect levels. These three
studies had marginally adequate sample sizes.
The remaining studies detailed in Chapter 7, and those listed in the
toxicity table in the Appendix, provide information on more severe effects
that can be used to show consistency with the threshold estimates. By them-
selves, however, they cannot be used to estimate a threshold since none
8-5
-------
adequately describes the shape of the dose-response severity relationship.
For example,, dose, rates associated with NOFELs (no-observed-frank-effect-
levels) Indicate that no significant change in frank effects was attributed
to the exposure. Milder effects were not Investigated, so that the NOFEl.
could dramatically overestimate the threshold.
8.2. FACTORS INFLUENCING HEALTH HAZARD ASSESSMENT
8.2.1. Exposure. Data are available regarding the potential human expo-
sure to HEX. It appears that any significant exposure would be the result
of Improper disposal or accidental spill. Limited data were presented for
the air and water levels of HEX In these Incidents. Emissions data, from
which atmospheric exposure estimates could be derived, have been sent to the
U.S. EPA, but are considered confidential business Information (CBI) and are
not available 1n this report. No HEX residue was detected In fish taken
from the waters near a production,plant In Memphis in 1982. No Information
was available regarding HEX contamination of other foods. Although occupa-
tional exposure is expected to be minimal, the long-term health effects of
continuous low-level exposure and/or intermittent acute exposure in man are
not known. Waste handlers and sewage treatment workers have been shown to
be occupations at risk.
8.2.2. Lowest-Observed-Effect Level. Both single dose and short-term
range-finding inhalation studies (7 hours/day) by Rand et al. (1982a) demon-
strated "a steep dose response effect of HEX exposure with a threshold of
toxicity 1n rats between 0.11 and 0.5 ppm." This observation is based on
severe irritation of the lungs, consequent inflammation, and impaired
respiratory function in rats. The TWA daily exposure levels, from the NOAEL
to the AEL, give a range between 0.004-0.089 ppm. Subchronic exposure (-90
days) to rats and monkeys (Rand et al., 1982a) Indicate a threshold range
8-6
-------
between 0.002-0.036, ppm based on TWA daily dose rates'.--'1D. Clark et al.
(1982) .exposed rats for 30 weeks arid found adverse effects.in the p. 089 ppm
TWA range with no adverse effects at 0.009 ppm TWA. However, Treon et al.
(1955) exposed rabbits, rats and guinea pigs to a TWA level of 0.031 ppm for
216 days and caused moderate adverse effects, so the lifetime experimental
threshold is likely to be somewhat•less. No lifetime data exist for deter-
mining NOELs or NOAELs. :
As expected, the toxicity from HEX 'inhalation seems highly dependent
upon the dosing rate and regimen. In several studies, a dose change of less
than one order of magnitude separated minor effects from increased mortal-
ity. This pattern was observed for acute studies through chronic studies.
In the previous comparison of threshold levels, the difference between
effects and no-observed-adverse effects depends to a large degree on the
researchers' documentation and detailed discussion of ;.the observed effects
shown by HEX exposure. With the narrow range between these dose levels, the
determination of exact separations between effect levels and adverse effect
levels is limited by the published data. •
The short-term oral studies (IRDC, 1978; SRI, 1980a,b) indicate a lowest
effect range for daily exposure to be 25-100 mg HEX/kg bw, based on rat and
mouse data. Subchronic oral studies (SRI, 1981a,b) suggest a lowest effect
range of 7-54 mg HEX/kg bw/day based on TWA dose rates used with rats and
mice. The rats responded at lower doses than did the mice, but the meta-
bolic similarities to man are not sufficiently well understood to allow
choice of a best animal model. Chronic oral HEX exposure to 0.2-2.0 mg/kg
showed no adverse effects (Naishtein and Lisovskaya, 1965).
8.2.3. Carcinogenlcity. There are no animal bioassay data indicating
that HEX is carcinogenic to animals. An inhalation carcinogenesis bioassay
8-7
-------
1n mice and rats Is to be conducted by NIP {Abdo, 1983). No unit risk
estimate for HEX has been suggested because carcinogenic bioassay data for
HEX have not been completed.
8.3. REGULATIONS AND STANDARDS
Hexachlorocyclopentadlene has been addressed under . numerous U.S.
statutes. These have been grouped according to the type of activity or
medium being controlled.
8.3.1. Occupational Standards. There is no current QSHA standard for HEX
levels In the workplace (29 CFR 1910). However, the AC6IH has adopted a
threshold limit value (TLV), expressed as an 8-hour time-weighted average
(TWA), of 0.1 mg/m3 (0.01 ppm). A short-term exposure limit (STEL), the
maximal concentration allowable in a 15-minute period, of 0.3 mg/m3 (0.03
ppm) for HEX has also been adopted (ACGIH, 1982). The levels are based on
the Treon et al. (1955) study.
In 1978, NIOSH classified HEX as a Group II pesticide and recommended
criteria for standards for occupations in pesticide manufacturing and formu-
lating. 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 1n manufacturing and
formulating. NIOSH specifically chose not to establish scientifically valid
environmental (workplace air) limits for pesticides (except those already
promulgated), because exposure by other routes, especially dermal, had
proved to be of critical importance for many pesticides and because NIOSH
believed that "Immediate action" was needed to protect workers in pesticide
manufacturing and formulating plants (NIOSH, 1978).
8.3.2. Transportation Regulations. The Hazardous Materials Transporta-
tion Act specifies the requirements to be observed in the preparation for
8-8
-------
shipment and transport of hazardous materials (49 CFR 171-179). The trans-
port of HEX by air, land and water is regulated by the.se statutes, and the
Department of Transportation has designated HEX as a "hazardous material"
(ID Number UN 2646), a "corrosive material", and a "hazardous substance" (49
CFR 172.101). The maximum net quantity of HEX permitted in one package for
transport by passenger-carrying aircraft or rallcar has been set at 1 quart,
while the maximum net quantity for cargo aircraft has been set at 10 gallons
per package. Transport on deck or below deck by cargo vessel is also per-
mitted (49 CFR 172.101).
The Hazardous Materials Transportation Act, in conjunction with the
Comprehensive Environmental Response, Compensation and Liability Act
(CERCLA), also provides that common carriers of hazardous substances may be
held liable for releases of hazardous substances in amounts equal to or
greater than their designated reportable quantity (RQ). The RQ for HEX has
been set at 1 pound (0.454 kg) (49 CFR 172).
8.3.3. Solid Waste Regulations. Under the Resources Conservation and
Recovery Act (RCRA), EPA has designated HEX as a hazardous toxic waste,
Hazardous Waste No. U 130 (40 CFR 261.33), subject to disposal and permit
regulations of Title 40, Code of Federal Regulations, Parts 262-265 and
Parts 122-124. Hexachlorocyclopentadlene is a hazardous constituent of
wastewater treatment sludge from the production of chlordane, wastewater and
scrub water from the chlorination of cyclopentadiene in the production of
chlordane, and filter solids from the filtration of HEX in the production of
chlordane (Hazardous Waste Nos. K032, K033 and K034, respectively) which are
also designated as a hazardous waste (40 CFR 261.320) and subject to RCRA
disposal regulations.
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8.3.4. Food Tolerances. Under FIFRA, a tolerance of 0.3 ppm has been
established for technical chlordane, Us components and metabolites which
cannot contain >1% of HEX (40 CFR 180.122).
8.3.5. Water Regulations. Under section 311 of the Federal Water Pollu-
tion Control Act, HEX was designated as a hazardous substance (40 CFR 116.4)
and these regulations established a Reportable Quantity (RQ) of 1 pound
(0.454 kg) for HEX (40 CFR 117.3). Discharges equal to or greater than the
RQ into or upon U.S. waters are prohibited unless the discharge is in com-
pliance with applicable permit programs (40 CFR 117.11).
Under the Clean Water Act, EPA has designated HEX as a toxic pollutant;
i.e., priority pollutant (40 CFR 401.15). 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. Specific definitions for classes and categories are set forth
in 40 CFR Parts 402 through 699.
Under the Clean Water Act, Ambient Water Quality Criteria (AWQC) for HEX
have also been developed (U.S. EPA, 1980c). Based on available toxicity
data for the protection of public health, the level derived was 206
vig/a.. Using organoleptic data for controlling undesirable taste and
odor quality of ambient water, the estimated level was 1 yg/a.. The AWQC
for freshwater aquatic life from acute and chronic toxicity indicated con-
centrations as low as 7.0 and 5.2 yg/8., respectively. Acute toxicity to
saltwater aquatic life was indicated at concentrations as low as 7.0
yg/8. (U.S. EPA, 1980c).
8.3.6. A1r Regulations. Hexachlorocyclopentadiene is not regulated under
the Clean Air Act. The U.S. EPA will propose a decision on the need to
regulate this chemical under the Clean Air Act and publish this proposal in
the Federal Register.
8-10
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8.3.7. Other Regulations! Pursuant to rules* under sections Q(a) and 8{d)
of the Toxic Substances Control Act (44 FR 31866), all manufacturers and
processors of HEX are required to report health and safety information on
HEX to EPA's Office of Toxic Substances. The deadline for submission of
Preliminary Assessment Information Manufacturer's Report on HEX (40 CFR 712)
was November 19, 1982.
In 1979, the Interagency Testing Committee recommended that HEX be con-
sidered for health and environmental effects testing under Section 4(a) of
the TSCA (44 FR 31866). This recommendation was based on evidence of poten-
tial human exposure and a potential for environmental persistence and bio-
accumulation. In 1982, the U.S. EPA responded (U.S. EPA, 1982) in the
Federal Register. The following is an excerpt from that notice:
EPA has decided not to initiate rulemaking 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 mutagenic and
teratogenic health effects. Neither has the EPA found evidence
that there is 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 oncogenic effects unnecessary.
8-11
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9-20
-------
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