United States
Environmental Protection
Agency
Office of Health and
Environmental Assessment
Washington DC 2O46O
EPA/600/8-84/015F
January 1985
Research and Development
SERA
Health Assessment
Document for
Chlorinated
Benzenes
Final
Report
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EPA/600/8-84/015F
January 1985
Final Report
Health Assessment Document
for
Chlorinated Benzenes
Final Report
Environmental Criteria and Assessment Office
Office of Health and Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268
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DISCLAIMER
This document has been reviewed \n 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)
on chlorinated benzenes at the request of the Office of A1r Qu'allty, Plan-
ning and Standards. The chlorinated benzenes are a group of 12 chlorinated
cyclic aromatic compounds which are currently being studied by the Environ-
mental Protection Agency (EPA) to determine If they should be regulated as
hazardous air pollutants under the Clean A1r Act.
In the development of this assessment document, the scientific litera-
ture 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 the various chlorinated benzenes. At
several stages In the HAD development process, the chlorinated benzenes
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 1n order to Identify the critical effect and to place adverse
health responses 1n perspective with observed environmental levels.
111
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Authors, Contributors and Reviewers
The EPA Office of Health and Environmental Assessment (OHEA) was respon-
sible for the preparation of this health assessment document. The OHEA
Environmental Criteria and Assessment Office (ECAO-C1nc1nnat1) had overall
responsibility for coordination and direction of the document and production
effort {W. Bruce Pelrano, Project Manager, Jerry F. Stara, Director, ECAQ-
C1nc1nnat1). W. Bruce Pelrano served both as the Project Manager and as the
principal author of this document. The following Individuals contributed
substantial portions 'of various chapters and their assistance has been
greatly appreciated:
D1pak Basu
Hike Neal
Shane Que Hee
David J. Relsman
Linda S. Erdrelch
Robert E. McGaughy
Chao W. Chen
William E. Pepelko
Muriel M. Llppman
Sheila L. Rosenthal
Syracuse Research Corporation
Syracuse Research Corporation
University of Cincinnati
College of Medicine
U.S. EPA, ECAO-C1nc1nnat1
U.S. EPA, ECAO-C1nc1nnat1
U.S. EPA, CAG, Washington, OC
U.S. EPA, CAG, Washington, DC
U.S. EPA, CAG, Washington, DC
ERNACO, Inc.
U.S. EPA, REAG, Washington, DC
The following people also contributed
document:
to the development of this
David Dellarco
Charles H. Nauman
Phil Wlrdzek
Larry J. Zaragoza
U.S. EPA, OTS, Washington, DC
U.S. EPA, EA6, Washington, DC
U.S. EPA, OTS, Washington, DC
U.S. EPA, OAQPS, RTP
The following Individuals were asked to review the final draft of this
document:
Albert E', Munson
James R. WHhey
Medical College of Virginia
Health and Welfare, Canada
1v
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The following Individuals were asked to review earlier drafts of this
document:
George T. Bryan University of Wisconsin
Derek J. CMpps University of Wisconsin
Erma Durden U.S. EPA, ECAO-C1nc1nnat1
Erdogan Erturk University of Wisconsin
Richard W. Lambrecht University of Wisconsin
Carl R. Morris U.S. EPA, OTS, Washington, OC
Henry A. Peters University of Wisconsin
James Wlthey Health and Welfare, Canada
Jennifer Orme U.S. EPA, ECAO-C1nc1nnat1
The following members of the ECAO-C1nc1nnat1 Technical Services Staff
were responsible for document production:
Cynthia Cooper Karen Mann
Patricia Daunt Judith Olsen
Cindy Fessler Bette Zwayer
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CHLORINATED BENZENES PEER REVIEW PANEL MEMBERS
July 25-26, 1983 Cincinnati, Ohio
Chairman:
W. Bruce Pelrano, ECAO-CIN
Julian B. Andelman
01pak Basu
Gary P. Carlson
Herbert H. Cornish
Fred Coulston
Diane Courtney
David Dellarco
Christopher DeRosa
Chris Dlppel
Linda S. Erdrelch
Charallnggayya Hlremath
Muriel M. Llppman
Oebdas Mukerjee
Albert Munson
Charles H. Nauman
Hike Neal
William E. Pepelko
Shane Que Hee
Martha J. Radlke
David J. Relsman
John F. Rlsher
Sheila L. Rosenthal
Jerry F. Stara
Norman M. Trleff
Phil Wlrdzek
Members
University of Pittsburgh
Syracuse Research Corporation
Purdue University
University of Michigan
Coulston International Corporation
EPA, HERL-RTP
EPA, OTS, Washington, DC
University of Maine
Dynamac Corporation
EPA, ECAO-CIN
EPA, CAG, Washington, DC
ERNACO, Inc.
EPA, ECAO-CIN
Medical College of Virginia
EPA, EAG, Washington, DC
Syracuse Research Corporation
EPA, ECAO-CIN
University of Cincinnati College of Medicine
University of Cincinnati College of Medicine
EPA, ECAO-CIN
EPA, ECAO-CIN
EPA, REAG, Washington, DC
EPA, ECAO-CIN
University of Texas Medical Branch
EPA, OTS, Washington, DC
v1
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TABLE OF CONTENTS
Page
1. INTRODUCTION 1-1
2. SUMMARY AND CONCLUSIONS 2-1
2.1. SUMMARY 2-1
2.1.1. Properties, Production and Use 2-1
2.1.2. Environmental Levels, Transport and Fate 2-2
2.1.3. Ecological Effects 2-4
2.1.4. Pharmacok1net1cs 2-5
2.1.5. Effects on Humans 2-10
2.1.6. Mammalian Toxicology 2-12
2.2. CONCLUSIONS 2-20
2.3. NEEDS FOR FUTURE RESEARCH 2-21
3. PHYSICAL AND CHEMICAL PROPERTIES/ANALYTICAL METHODOLOGY 3-1
3.1. SYNONYMS, TRADE NAMES AND IDENTIFICATION NUMBERS 3-1
3.2. PHYSICAL AND CHEMICAL PROPERTIES 3-1
3.3. ANALYTICAL METHODOLOGY 3-15
3.3.1. Chemical Analysis 1n A1r 3-16
3.3.2. Chemical Analysis 1n Water 3-18
3.3.3. Chemical Analysis 1n Soil, Sediment and Chemical
Waste Disposal Site Samples 3-20
3.3.4. Chemical Analysis 1n F1sh and Other Foods 3-20
3.4. SUMMARY 3-23
4. PRODUCTION, USE AND ENVIRONMENTAL LEVELS 4-1
4.1. PRODUCTION 4-1
4.2. USE 4-5
4.3. SOURCE AND ENVIRONMENTAL LEVELS ! 4-5
4.3.1. Levels 1n A1r 4-12
4.3.2. Water 4-18
4.3.3. Food 4-26
4.3.4. Soil and Sediments 4-28
4.3.5. Human Tissue Residues 4-29
4.4. RELATIVE SOURCE CONTRIBUTIONS TO TOTAL EXPOSURE 4-34
4.4.1. A1r 4-36
4.4.2. Water 4-36
4.4.3. Food 4-38
4.5. SUMMARY 4-38
V11
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Page
5. ENVIRONMENTAL TRANSPORT AND FATE 5-1
5.1. TRANSPORT 5-1
5.1.1. Air 5-1
5.1.2. Water 5-2
5.1.3. Soil 5-4
5.2. FATE 5-7
5.2.1. A1r 5-7
5.2.2. Water . 5-8
5.2.3. Soil 5-10
5.3. BIOCONCENTRATIQN, BIOACCUHULATION AND BIOMAGNIFICATION. . . 5-13
5.4. SUMMARY 5-20
6. ECOLOGICAL EFFECTS. 6-1
6.1, EFFECTS ON THE AQUATIC ENVIRONMENT 6-1
6.1.1. Effect on Freshwater and Marine F1sh 6-1
6.1.2. Effect on Aquatic Crustaceans 6-14
6.1.3. Embryotoxlc and Reproductive Effects. . 6-16
6.1.4. Effect on Aquatic Plants 6-24
6.1.5. Residues. 6-30
6.2. EFFECTS ON NONAQUATIC ENVIRONMENTS 6-34
6.2.1. Plants 6-34
6.2.2. Insects ...... ... 6-35
6.2.3. Birds 6-37
6.2.4. Residues 6-38
6.3. SUMMARY 6-39
7. MONOCHLOROBENZENE 7-1
7.1. PHARMACQKINETICS. . 7-1
7.1.1. Absorption 7-1
7.1.2. Distribution 7-1
7.1.3. Metabolism 7-2
7.1.4. Excretion . . 7-5
7.1.5. Summary 7-10
7.2. EFFECTS ON HUMANS 7-10
¥111
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Page
7.3. MAMMALIAN TOXICITY 7-12
7.3.1. Acute Toxldty 7-12
7.3.2. Subchronlc Toxldty 7-15
7.3.3. Chronic Toxldty 7-23
7.3.4. Mutagen1c1ty 7-24
7.3.5. Carclnogenldty 7-24
7.3.6. Reproductive and Teratogenlc Toxlclty 7-30
7.4. INTERACTIONS 7-30
7.5. SUMMARY 7-31
8. OICHLOROBENZENES 8-1
8.1. PHARMACOKINETICS 8-1
8.1.1. Absorption 8-1
8.1.2. Distribution 8-3
8.1.3. Metabolism 8-6
8.1.4. Excretion . . .......... 8-8
8.1.5. Summary 8-9
8.2. EFFECTS ON HUMANS 8-10
8.2.1. Occupational Studies 8-10
8.2.2. Case Studies 8-11
8.2.3. Summary 8-17
8.3. MAMMALIAN TOXICOLOGY 8-17
8.3.1. Acute Toxldty 8-17
8.3.2. Subchronlc Toxldty 8-22
8.3.3. Chronic Toxldty . 8-32
8.3.4. Mutagenldty 8-34
8.3.5. Carc1nogen1c1ty ......... . . 8-36
8.3.6. Reproductive and Teratogenlc Toxldty 8-40
8.4. INTERACTIONS 8-41
8.5. SUMMARY 8-41
9. TRICHLOROBENZENES 9-1
9.1. PHARMACOKINETICS 9-1
9.1.1. Absorption 9-1
9.1.2. Distribution 9-2
9.1.3. Metabolism. ....... ...... 9-2
9.1.4. Excretion 9-6
9.1.5. Summary ....... 9-8
9.2. EFFECTS IN HUMANS 9-9
1x
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Page
9.3. MAMMALIAN TOXICOLOGY 9-9
9.3.1. Acute ToxIcHy 9-9
9.3.2. Subchronlc Tox1c1ty 9-14
9.3.3. Chronic Tox1c1ty 9-23
9.3.4. MutagenlcUy 9-24
9.3.5. Cardnogenldty 9-25
9.3.6. Reproductive and Teratogenlc ToxIcHy 9-26
9.4. INTERACTIONS 9-28
9.5. SUMMARY 9-29
10. TETRACHLOROBENZENES 10-1
10.1. PHARMACOKINETICS 10-1
10.1.1. Absorption 10-1
10.1.2. Distribution 10-2
10.1.3. Metabolism 10-7
10.1.4. Excretion 10-10
10.1.5. Summary 10-13
10.2. EFFECTS ON HUMANS 10-14
10.3. MAMMALIAN TOXICOLOGY 10-15
10.3.1. Acute ToxIcHy 10-15
10.3.2. Subchronlc ToxIcHy 10-21
10.3.3. Chronic ToxIcHy 10-23
10.3.4. Mutagenldty 10-23
10.3.5. Cardnogenldty 10-24
10.3.6. Reproductive and Teratogenlc Effects 10-24
10.4. INTERACTIONS 10-27
10.5. SUMMARY 10-28
11. PENTACHLOROBENZENE 11-1
11.1. PHARMACOKINETICS 11-1
11.1.1. Absorption 11-1
11.1.2. Distribution 11-2
11.1.3. Metabolism 11-7
11.1.4. Excretion 11-11
11.1.5. Summary 11-13
11.2. EFFECTS ON HUMANS 11-13
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Page
11.3. MAMMALIAN TOXICOLOGY. 11-13
11.3.1. Acute Tox1c1ty . 11-13
11.3.2. Subchronlc ToxIcHy 11-16
11.3.3. Chronic ToxIcHy 11-18
11.3.4. MutagenlcHy 11-19
11.3.5. CarclnogenlcHy 11-19
11.3.6. Reproductive and Teratogenlc ToxIcHy 11-19
11.4. INTERACTIONS 11-26
11.5. SUMMARY 11-26
12. HEXACHLOROBENZENE 12-1
12.1. PHARMACOKINETICS 12-1
12.1.1. Absorption 12-1
12.1.2. Distribution 12-3
12.1.3. Metabolism 12-16
12.1.4. Excretion 12-20
12.1.5. Summary 12-27
12.2. EFFECTS ON HUMANS 12-29
12.2.1. Ep1dem1olog1c Studies . 12-29
12.2.2. Accidental Ingestlon 1n Turkey . 12-32
12.2.3. Summary 12-37
12,3 MAMMALIAN TOXICOLOGY 12-40
12.3.1. Acute ToxIcHy 12-40
12.3.2. Subchronlc ToxIcHy 12-42
12.3.3. Chronic ToxIcHy . 12-56
12.3.4. MutagenlcHy . 12-59
12.3.5. CardnogenlcHy . 12-60
12.3.6. Reproductive and Teratogenlc Effects 12-122
12.4. INTERACTIONS 12-127
12.5. SUMMARY 12-131
13. OVERVIEW OF EFFECTS OF MAJOR CONCERN 13-1
13.1. PRINCIPAL EFFECTS AND TARGET ORGANS ... 13-1
13.2. ANIMAL TOXICITY STUDIES USEFUL FOR HEALTH ASSESSMENT
AND ESTIMATED TOXICITY THRESHOLDS 13-5
13.2.1. Animal ToxIcHy Studies 13-5
13.2.2. Estimated ToxIcHy Thresholds 13-32
13.3. CARCINOGENICITY STUDIES 13-32
13.4. HUMAN STUDIES . 13-40
xl
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Page
13.5. FACTORS INFLUENCING HEALTH HAZARD ASSESSMENT . . 13-41
13.5.1. Exposure 13-41
13.6. REGULATIONS AND STANDARDS 13-46
13.6.1. Occupational Standards 13-46
13.6.2. Transportation Regulations 13-52
13.6.3. Solid Haste Regulations 13-53
13.6.4. Food Tolerances 13-55
13.6.5. Water Regulations . 13-55
13.6.6. A1r Regulations ......... 13-56
14. REFERENCES 14-1
APPENDIX A: Comparison Among Different Extrapolation Models A-l
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LIST OF TABLES
No. Title Page
3-1 Synonyms, Trade Names and Identification Numbers of the
Chlorinated Benzenes 3-3
3-2 Physical Properties of the Chlorinated Benzenes 3-7
3-3 Vapor Pressures and Vapor Densities of the Chlorinated
Benzenes 3-8
3-4 Reported Composition of Commercial l,2-D1chlorobenzene. . . . 3-11
4-1 United States Production of Chlorinated Benzenes for
Selected Years 4-2
4-2 U.S. Producers and Estimated Annual Production Capacities
(1983) of Chlorobenzenes 4-6
4-3 A Summary of the Uses of the Chlorinated Benzenes 4-8
4-4 Estimated Quantities of Chlorobenzenes Lost During
Manufacture, and to the Environment Compared with
Total Production In 1983 • 4-9
4-5 Estimated Quantities of Hexachlorobenzene (HCB) 1n
Industrial Wastes and Byproducts In 1972 4-11
4-6 Chlorinated Benzene Levels 1n Ambient A1r from
Different Locations 1n the U.S 4-13
4-7 Concentrations of Chlorinated Benzenes at Three Sites .... 4-16
4-8 Overall and Site-Specific Mean Atmospheric Levels of
Chlorobenzenes throughout the United States 4-17
4-9 Atmospheric Levels of Hexachlorobenzene Around Selected
Industrial Plants 4-19
4-10 Chlorinated Benzenes 1n Surface Water 4-20
4-11 Chlorobenzene Concentrations 1n Drinking Water from
Ontario, Canada 4-24
4-12 Frequency and Range of Concentrations of Chlorinated
Benzenes Pollutants 1n Industrial Wastewaters 4-27
4-13 Chlorinated Benzene Residues 1n Human Adipose Tissue 4-30
4-14 Chlorinated Benzenes 1n the Blood of Nine Residents of
Love Canal 1n Niagara Falls, New York 4-33
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Ngu. Title Page
4-15 Chlorinated Benzenes 1n the Breath and Urine of Nine
Residents of Love Canal 1n Niagara Falls, New York 4-35
4-16 Estimated Yearly Exposure to Several Chlorinated Benzenes
Via Inhalation 4-37
5-1 Predicted Transport and Fate of Chlorinated Benzenes
Released from Landfills and Lagoons 5-5
5-2 Transport of Chlorinated Benzenes 1n Sandy Soil 5-6
5-3 Estimated Atmospheric Residence Time and Dally Loss Rates
for Several Chlorinated Benzenes 5-9
5-4 Aqueous B1odegradab1l1ty Studies of Chlorinated Benzenes. . . 5-11
5-5 Octanol/Water Partition Coefficients, B1oconcentrat1on
Factors and Biological Half-lives for Chlorinated Benzenes
1n F1sh 5-15
5-6 B1oconcentrat1on Factor and Slope of the Elimination Curve
for Gupples (PoeclUa retlculata) Exposed to Six Chlorinated
Benzenes. . 5-19
6-1 Acute Toxldty Data for F1sh Species Exposed to Chlorinated
Benzenes 6-2
6-2 Chronic Toxldty Values of Chlorinated Benzenes 1n F1sh . . . 6-12
6-3 B1oconcentrat1on Factors of Some Chlorinated Benzenes
1n Two F1sh Species 6-15
6-4 Acute Toxldty Data for Crustaceans Exposed to Chlorinated
Benzenes 6-17
6-5 Embryo-Larval Toxldty of Honochlorobenzene to Goldfish,
Largemouth Bass and Rainbow Trout In Soft and Hard Water. . . 6-21
6-6 Results of 1,2,4,5-Tetrachlorobenzene Tests with Embryo
to Juvenile Sheepshead Minnows 1n Continuous-Flow Natural
Seawater. 6-23
6-7 Adult Llfespan and Reproductive Performance of Brine
Shrimp Exposed to 1,3,5-Trlchlorobenzene 6-25
6-8 Acute Toxldty Data for Aquatic Algae Exposed to
Chlorinated Benzenes 6-26
6-9 Chlorinated Benzene Concentrations (yg/8,) 1n Water and
Sediment 6-31
xlv
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No.
6-10
6-11
6-12
7-1
7-2
7-3
7-4
7-5
7-6
7-7
8-1
8-2
8-3
8-4
8-5
8-6
8-7
8-8
9-1
9-2
Title
Chlorinated Benzene Concentrations 1n a Variety of Marine
Species . ,
Emergence of Adult HousefHes 8 Days Following Exposure of
Pupae to "Saturation Concentration" of Dlchlorobenzene
Vapors
Chlorinated Benzene Residues 1n Bird Eggs .... .
Percentage of Isomers of Chlorophenol from Metabolism
of Monochlorobenzene . ..
Species Variation 1n Urinary Metabolites of 14C~Mono-
chlorobenzene ....
Acute Toxldty of Monochlorobenzene
Summary of Subchronlc Toxldty Studies on Monochlorobenzene .
Mutagen1c1ty Testing of Monochlorobenzene
Nonneoplastlc Lesions 1n F344 Rats Given Chlorobenzene by
Savage for 2 Years ,
Statistical Comparisons of Liver Tumors in Male Rats
Treated with Chlorobenzene and Vehicle Controls
Tissue Concentrations of 1 ,4-D1chlorobenzene 1n Adult
Female CFY Rats
Chromosomal Alterations 1n Persons Accidentally Exposed
to 1 ,2-D1chlorobenzene
Case Reports Involving Dlchlorobenzenes (DCB)
Acute Toxldty of 1 52-01chlorobenzene
Acute Toxldty of 1 ,4-D1chlorobenzene
Subchronlc Toxldty of 1 ,2-01chlorobenzene.
Subchronlc and Chronic Toxldty of 1 ,4-D1chlorobenzene. . . .
NTP Bloassay of 1 ,2-D1chlorobenzene Analysis of
Primary Tumors 1n Male Rats: Adrenal Pheochromocytomas. . . .
Distribution of 14C-Labeled 1 ,2,4-Tdchlorobenzene In Rat
Tissues after Oral Dosing with 181.5 mg/kg/day for 7 Days . .
Summary of Subchronlc and Chronic Toxldty Studies
on Trlchlorobenzenes
Page
6-32
6-36
6-40
7-4
7-9
7-14
7-16
7-25
7-27
7-28
8-5
8-12
8-13
8-19
8-20
8-23
8-25
8-37
9-3
9-15
XV
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No. TUIe Page
10-1 Percentage of 1,2,4,5-Tetrachlorobenzene Steady-State
Reached at Specific Times In Fat and Plasma of Dogs 10-4
10-2 Time Required to Reach Various Percentages of 1,2,4,5-
Tetrachlorobenzene Steady-State 1n Fat and Plasma of Dogs . . 10-5
10-3 Unchanged Tetrachlorobenzene In Rabbit Tissues 6 Days
After Oral Dosing 10-6
10-4 Urinary Metabolites of Tetrachlorobenzene Isomers 1n
Rabbits 6 Days After Oral Dosing 10-9
10-5 Summary of Excretion of the IsomeMc Tetrachlorobenzenes
as Metabolites or as Unchanged Compound 1n Rabbits Dosed
Orally 10-11
10-6 Excretion of Unchanged Tetrachlorobenzenes In the Expired
A1r of Rabbits After Oral Dosing 10-12
10-7 Frequency of Chromat1d-type Chromosome Aberrations In
Peripheral Lymphocytes 10-16
10-8 Frequency of Labile Chromosome-type Aberrations 10-17
10-9 Frequency of Stable Chromosome-type Aberrations 10-18
10-10 Summary of Toxlclty Studies on Tetrachlorobenzenes 10-19
11-1 Distribution of Pentachlorobenzene Residues 1n the
Tissues of Maternal Rats after Oral Administration 11-3
11-2 Distribution of Pentachlorobenzene Residues 1n the
Tissues of Fetal Rats after Oral Administration to Dams . . . 11-4
11-3 Distribution of Pentachlorobenzene and/or Metabolites on
the 40th Day In the Rhesus Monkey Following a Single Oral
Dose of 0.5 mg/kg Body Weight 11-6
11-4 Distribution of Pentachlorobenzene 1n Chinchilla Doe
Rabbits Expressed as a Percentage of Administered Dose. . . . 11-8
11-5 Percentage of Pentachlorobenzene and Its Metabolites
Identified 1n Urine, Feces and Various Organs of Rhesus
Monkeys Dosed 0.5 mg/kg Body Height Pentachlorobenzene. . . . 11-9
11-6 Cumulative Urinary and Fecal Excretion of Pentachlorobenzene
and Metabolites During 40 Days Following a Single Oral Dose
of 0.5 mg/kg 1n Male and Female Rhesus Monkeys 11-12
11-7 Acute Oral Toxlclty of Pentachlorobenzene 11-15
xv 1
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No. Title Page
11-8 Summary of Subchronlc, Reproductive and Teratogenlc
Tox1c1ty Studies on Pentachlorobenzene 11-17
11-9 Reproductive Effects 1n Litters of female Rats Fed Diets
Containing Pentachlorobenzene ........ 11-21
11-10 Toxic Effects of Pentachlorobenzene on Reproduction 1n
Rats Dosed on Each of Gestation Days 6-15 .......... 11-23
11-11 Skeletal and Soft-Tissue Abnormalities Observed 1n Rat
Utters of Dams Treated with Pentachlorobenzene on Each
of Gestation Days 6-15 11-24
11-12 Fetal Wlstar Rat Residues of Pentachlorobenzene 11-25
12-1 Storage and Excretion of 14C-HCB Administered Orally
1n Arachls 011 1n Rats 12-4
12-2 Tissue Concentration (ppm) of l"C-Hexachlorobenzene
and Its Metabolites 1n Sprague-Dawley Rats 12-6
12-3 Tissue Levels of HCB (ppm) 1n Adult Female Rhesus Monkeys . . 12-7
12-4 HCB Concentrations 1n Tissues of Male Beagles Receiving
Single Intravenous Doses of 1 mg/kg bw 1n Olive Oil 12-9
12-5 Mean (+SE) Hexachlorobenzene Radioactivity (dpm/g) of
Selected European Ferret Tissues. . 12-14
12-6 Mean (tSE) HCB Radioactivity (dpm x 103) of European
Ferret Kits 12-15
12-7 Concentrations of HCB and Us Metabolites (mg/kg) In the
Liver and Kidneys of Male and Female Rats 12-19
12-8 Hexachlorobenzene and Its Major Metabolites In the Excreta
of Different Animal Species 12-21
12-9 Results of Blood and Urine Analysis 1n Men Employed 1n a
Chlorinated Solvents Plant, 1974-1977 . . 12-31
12-10 HCB Plasma Levels 1n Exposed Individuals and Controls .... 12-33
12-11 Clinical Signs and Symptoms 1n Humans 25 Years After
Exposure to Low Levels 1n HCB 1n Turkey, 1955-1959 12-36
12-12 Porphyrln Levels 1n Patients and Controls ... 12-38
12-13 Laboratory Test Results of Turkish Patients . 12-39
12-14 Summary of Toxldty Studies on Hexachlorobenzene 12-43
xv11
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No. Title Page
12-15 Porphyrln Content and Uroporphyrlnogen Decarboxylase
Activity 1n the Liver Cytosol of Female Rats Pretreated
with TOO mg/kg HCB Every Other Day for 6 Weeks 12-53
12-16 Tumor Incidence In Hamsters Given HCB In the Diet 12-62
12-17 Effect of HCB on Hamsters: Liver Tumors and Other Liver
Lesions 12-66
12-18 Liver Tumor Incidence 1n Mice Fed HCB 12-68
12-19 Tumor Data on Mice Fed HCB 12-69
12-20 Body Weights of Female Agus Rats Fed Hexachlorobenzene
for 90 Weeks 12-73
12-21 Growth Rates for Female Agus Rats on a Diet Containing
100 ppm HCB 12-74
12-22 Dosage Levels 1n the Chronic Feeding Study of
Hexachlorobenzene In Sprague-Dawley Rats. . 12-78
12-23 Liver and Kidney Tumors In Sprague-Dawley Rats Given
Hexachlorobenzene 1n the Diet for up to 2 Years 12-79
12-24 Adrenal Tumors 1n Sprague-Dawley Rats Given
Hexachlorobenzene 1n the Diet for up to 2 Years 12-81
12-25 Intake of Hexachlorobenzene (mg/kg/day) 1n the Chronic
Feeding, 2-Generat1on Study of Hexachlorobenzene 1n
Sprague-Dawley Rats 12-83
12-26 Tumors 1n Organs that Showed Statistical Differences
from Control 1n F-j Sprague-Dawley Rats Treated with
Hexachlorobenzene . 12-84
12-27 Parathyroid and Adrenal Pheochromocytomas 1n Sprague-
Dawley Rats Maintained on Synthetic Diets of Varying
Vitamin A Content and With or Without Hexachlorobenzene . . . 12-86
12-28 Qualitative Comparison of Tumor Development 1n Rats
Following Hexachlorobenzene Administration 1n Different
Studies 12-89
12-29 Tumor Incidences 1n Male and Female Hamsters Given
Hexachlorobenzene 1n Diet 12-104
12-30 Incidence of Liver Cell Tumors 1n Male and Female
Swiss Mice Given Hexachlorobenzene Diet 12-105
XV111
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No. Title Page
12-31 Liver and Kidney Tumor Incidence Rates 1n Hale and
Female Sprague-Oawley Rats Given Hexachlorobenzene 1n Diet. . 12-106
12-32 Incidence Rate of Adrenal Pheochromocytoma 1n Female
Sprague-Dawley Rats (F-| generation) 1n a 2-6enerat1on
Feeding Study 12-107
12-33 The Carcinogenic Potency of Hexachlorobenzene, Calculated
on the Basis of 14 Data Sets, Using the Linearized
Multistage Model 12-110
12-34 Upper-Bound (Point) Estimation of Risk, Based on
Hepatocellular Carcinoma 1n Female Rats 12-112
12-35 Relative Carcinogenic Potencies Among 54 Chemicals
Evaluated by the Carcinogen Assessment Group as
Suspect Human Carcinogens 12-115
12-36 Significantly Increased Incidence of Tumors 1n
Animals Given Hexachlorobenzene 1n Diet 12-121
12-37 Analysis of the Excreta from Rats Administered Hexa-
chlorobenzene After an Initial Treatment with Dlethyl-
stllboestrol 12-129
13-1 Summary of Subchronlc Toxldty Studies on Monochlorobenzene . 13-6
13-2 Subchronlc Toxldty of 1,2-01chlorobenzene 13-9
13-3 Subchronic and Chronic Toxldty of 1,4-D1chlorobenzene. . . . 13-11
13-4 Summary of Subchronlc and Chronic Toxldty Studies on
TMchlorobenzenes 13-13
13-5 Summary of Toxldty Studies on Tetrachlorobenzenes 13-15
13-6 Summary of Subchronlc, Reproductive and Teratogenlc
Toxldty Studies on Pentachlorobenzene 13-17
13-7 Summary of Toxldty Studies on Hexachlorobenzene 13-18
13-8 Comparison of Toxic Effects of Chlorinated Benzenes
1n Rats 13-23
13-9 Comparison of Toxic Effects of Chlorinated Benzenes
1n Mice 13-25
13-10 Comparison of Toxic Effects of Chlorinated Benzenes
1n Rabbits 13-27
x1x
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No. Title Page
13-11 Comparison of Toxic Effects of Chlorinated Benzenes
In Dogs 13-29
13-12 Comparison of Toxic Effects of Chlorinated Benzenes
In Monkeys 13-31
13-13 Tox1c1ty Data for Threshold Estimates 13-33
13-14 Summary of Tumors Induced 1n Rodents by HCB , 13-38
13-15 Comparison of Chemical and Physical Properties of
Chlorinated Benzenes 13-42
13-16 Comparison of Chlorinated Benzenes BCF and Water
Concentrations 13-43
13-17 Estimated Yearly Exposure to Several Chlorinated
Benzenes Via Inhalation 13-45
13-18 Occupational Standards for Monochlorobenzene 13-47
13-19 Occupational Standards for 1,2-D1chlorobenzene ... 13-49
13-20 Occupational Standards for 1,4-D1chlorobenzene ... 13-50
13-21 The Chlorinated Benzenes as Constituents of
Hazardous Wastes from Specific Sources 13-54
13-22 Ambient Water Quality Criteria for Chlorinated
Benzenes—Aquatic Life 13-57
13-23 Ambient Water Quality Criteria for the Chlorinated
Benzenes for the Protection of Human Health 13-58
13-24 Maximum Imm1ss1on Concentration Standards for
Monochlorobenzene 13-60
xx
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LIST OF FIGURES
No. TUIe Page
3-1 Chemical Structure of the Chlorinated Benzenes 3-2
7-1 Metabolism of Monochlorobenzene . . 7-7
9-1 Metabolic Pathways for Trlchlorobenzene (TCB) Isomers
Through Arene Oxide Intermediates 1n Rabbits 9-7
12-1 Histogram Representing the Frequency Distribution of
the Potency Indices of 54 Suspect Carcinogens Evaluated
by the Carcinogen Assessment Group 12-114
xx1
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1. INTRODUCTION
The purpose of this document 1s to summarize the current knowledge of
the effects of exposure to the chlorinated benzenes on human health.
The chlorinated benzenes are a group of 12 compounds 1n which 1 to 6
chlorine atoms have been substituted for the hydrogens on a benzene ring.
They are used as chemical Intermediates 1n the synthesis of pesticides and
other chlorinated compounds, and as solvents, pesticides, dye carriers,
space deodorants and other products. Environmental contamination results
from emissions to air and water during the manufacture and use of the
chlorinated benzenes and from the disposal of wastes from a number of pro-
cesses. These compounds are resistant to chemical and biological degrada-
tion and tend to accumulate 1n I1p1d-conta1n1ng tissues of animals and
humans. The ubiquitous environmental distribution of the chlorinated ben-
zenes and their bloconcentratlon 1n humans are a basis for concern over the
consequences of chronic exposure to human health.
The rationale for structuring the document Is based primarily on two
major Issues, exposure and response. The first portion of the document 1s
devoted to the chlorinated benzenes 1n the environment: physical and chemi-
cal properties, the monitoring of the chlorinated benzenes 1n various media,
natural and human-made sources, the transport and distribution of the
chlorinated benzenes within environmental media, and the levels of expo-
sure. The second part 1s devoted to biological responses 1n laboratory
animals and humans Including metabolism, pharmacok1net1cs, as well as the
toxlcologlcal effects of the chlorinated benzenes.
1-1
-------
This assessment 1s based on original publications, although the overall
knowledge covered by a number of reviews and reports was also considered.
The references cited were selected to reflect the current state of knowledge
on those Issues which are most relevant for a health assessment of the
chlorinated benzenes 1n the environment.
1-2
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2. SUMMARY AND CONCLUSIONS
2.1. SUMMARY
2,1.1. Properties, Production and Use. The chlorinated benzenes are a
group of cyclic aromatic compounds 1n which 1-6 hydrogen atoms of a benzene
ring have been replaced by up to six chlorine substltutents. This substitu-
tion yields 12 compounds: monochlorobenzene, three 1somer1c forms of
dlchlorobenzene, three Isomers of trlchlorobenzene, three Isomers of tetra-
chlorobenzene, pentachlorobenzene and hexachlorobenzene. The physical prop-
erties of these compounds vary with the degree of substitution of each and
are, 1n general, low water solubility (solubility decreasing with Increasing
chlorlnatlon), low flammabllHy, moderate to high octanol/water partition
coefficients (coefficients Increasing with Increasing chlorlnatlon) and low
to moderate vapor pressures (vapor pressures decreasing with Increasing
chlorlnatlon). They are chemically unreactlve and exist as liquids or
solids at environmental conditions. Analyses of airborne chlorobenzenes are
usually accomplished by adsorption onto sorbent cartridges, followed by
thermal desorptlon and analysis by gas chromatography (GC). For water
samples, the purge-trap method 1s used to concentrate the volatile halo-
genated benzenes before analysis by GC. For less volatile chlorinated ben-
zenes, solvent extraction followed by column chromatographlc cleanup of the
extract and electron capture/gas chromatography (EC/GC), 1s the most com-
monly used method for the Isolation, detection and quantification. Methods
similar to those used for wastewater samples are commonly used for the
analysis of chlorinated benzenes 1n biological matrices.
Annual production of these 12 chlorinated benzenes In 1983 was on the
order of 450 million pounds, the majority of which 1s accounted for by mono-
chlorobenzene and dlchlorobenzenes. These compounds are used 1n a number of
2-1
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organic chemical syntheses, Including the synthesis of other chlorobenzenes,
and have applications as solvents, electrical equipment Insulators, pesti-
cides, herbicides and fungicides. Emissions of chlorobenzenes are most
Hkely to occur during their manufacture or 1n their use as Intermediates
and from the disposal of waste products from manufacturing operations.
Hexachlorobenzene 1s Imported but not produced commercially 1n the United
States, and occurs as a by-product In the synthesis of nine other chlori-
nated hydrocarbons; 2-5 million pounds may be generated each year.
2.1.2. Environmental Levels, Transport and Fate. Chlorinated benzenes
have been Identified 1n air, food and soil, and 1n surface, ground and
drinking water. The highest concentrations have been found 1n or near manu-
facturing and waste disposal sites, although no study has attempted to char-
acterize the contribution of any one source to the total environmental con-
tamination by chlorobenzenes. Ambient air and water levels are 1n the
m1crogram/cub1c meter and m1crogram/Hter range, respectively, although
monitoring studies for finished water have been limited. The most fre-
quently detected chlorinated benzenes 1n air and water were monochloroben-
zene and the d1- and trlchlorobenzenes. Penta- and hexachlorobenzene have
been found more frequently 1n food and soil, although their detection may
reflect more the concern over their use as pesticides and fungicides, or
their presence as contaminants 1n pesticides or fungicides, rather than the
absence of the other chlorobenzenes.
The transport and fate of the chlorinated benzenes In the environment
have not been well characterized although, from laboratory and field studies
and from the known chemical and physical properties, several generalizations
can be made. After emission Into air, the chlorobenzenes are likely to be
widely dispersed by the prevailing wind and degrade slowly through chemical
2-2
-------
and photolytlc reactions. One study estimated residence times for three of
the chlorobenzenes to be 1n the range of from 13-116 days. When released
Into water, these compounds, because of their low water solubility, will
evaporate from the surface rapidly. Small amounts are likely to remain 1n
solution or be removed through sedimentation. Some of the chlorobenzenes
can undergo mlcroblal degradation, and all show a propensity for bloaccumu-
latlon. After release of chlorobenzenes Into soil, very Uttle will be
removed by leaching with water because of low water solubility and high soil
adsorption; the latter Increases with the number of substHuent chlorines.
Evaporation 1s likely to occur from the upper soil layers. Overall, the
less chlorinated chlorobenzenes will tend to partition from soil and water
Into air, there to be dispersed and degraded. The chlorobenzenes will also
tend to enter the atmosphere, either as participates or vapors, and
disperse, degrade or precipitate out.
The chlorinated benzenes are llpophlllc compounds that bloaccumulate In
animal and human tissues from ambient air, water and food. The bloconcen-
tratlon factor (BCF) (tissue concentration/media concentration) 1s an Indi-
cator of bloaccumulatlon and 1s sometimes expressed 1n terms of physico-
chemical parameters such as the water solubility or the octanol/water parti-
tion coefficient, when biological data are not available, which reflect the
number of substHuent chlorine atoms. The BCF In various fish species range
from 12-46 for monochlorobenzene to >44,000 for hexachlorobenzene. Physio-
logical exposure levels {the levels of exposure, concentration, at the site
of the compounds Interaction, sequestration or observed effects) are deter-
mined by absorption, metabolism, elimination and storage In adipose tissue;
thus, biologically persistent compounds, like the chlorinated benzenes, may
result 1n prolonged physiological exposures.
2-3
-------
No comprehensive study of human exposure to the chlorinated benzenes has
been conducted, although their ubiquity 1n the environment and the detection
of measurable residues 1n human tissue Indicate that human exposure and
absorption do occur. The contribution of the chlorinated benzenes from all
three media (air, water and food), to estimate a person's total exposure,
cannot be made from the limited environmental monitoring data. The avail-
able data, however, Indicate that human Inhalation exposure to chlorinated
benzenes may be higher than 1ngest1on exposure either through drinking water
or through foods.
2.1.3. Ecological Effects. As has been demonstrated 1n acute toxldty
bloassays, the LC5_ 1n fish generally decreases as the number of substHu-
ent chlorine atoms on the molecule Increases (Isomers vary). Chlorinated
benzenes cause adverse reproductive effects 1n Invertebrates and fish.
Honochlorobenzene tested 1n goldfish and largemouth bass, 1,3,5-tr1chloro-
benzene tested 1n brine shrimp, and the exposure of sheepshead minnows to
1,2,4,5-tetrachlorobenzene resulted 1n decreased hatching of eggs or embryo
lethality and decreased survival of juvenile fish.
Adverse effects of chlorinated benzenes were also apparent 1n terres-
trial organisms. Mitosis 1n seeds and seedlings was disrupted by l,4-d1-
chlorobenzene; 1,2,4,5-tetrachlorobenzene affected seed germination and
seedling growth depending on soil type. Soil application rates of 224 kg/ha
or higher of 1,2,4,5-tetrachlorobenzene were found to be toxic to mature
cotton plants. Dlchlorobenzene vapors at "saturation concentrations"
Inhibited the emergence of housefly pupae, while l,2-d1chlorobenzene and
trlchlorobenzene each in dlesel oil were toxic to Douglas fir beetles.
2-4
-------
Contact with residues of 1,3,5-tMchlorobenzene shortened the Hfespan of
female wasps, and their eggs suffered high mortality within 7 days of
exposure.
Although effects of chlorinated benzenes (mortality, decreased reproduc-
tion) on natural populations have not been adequately studied, tissue
concentrations of several Isomers were measured 1n a number of different
species. Aquatic organisms (fish and Invertebrates) and terrestrial species
have been found to contain chlorinated benzenes levels. Tissue concentra-
tions of the measured chlorinated benzenes were highest for hexachloro-
benzene. The detection 1n North America and Europe of hexachlorobenzene 1n
the eggs of birds and subcutaneous fat of wild animals suggests Us wide-
spread distribution 1n the environment.
2.1.4. Pharmacok1net1cs. Monochlorobenzene 1s readily absorbed through
the respiratory system and the gastrointestinal tract, but the quantitative
extent 1s not known. It 1s deposited 1n body I1p1ds and metabolized by
mlcrosomal oxidation. Ox1dat1ve reactions, via the mixed function oxldase
enzymes, are believed to lead to the formation of metastable arene oxide
Intermediates; these epoxldes are metabolized further to the ortho-, meta-
or para-chlorophenols. The chlorophenols can conjugate with glutathlone and
be detoxified by conversion to the corresponding mercapturlc adds and
excreted via the urine or they can bind to cellular proteins. Binding to
cellular protein appears to be correlated with necrotlc pathological changes
1n the kidneys and livers of rodents. In addition to conjugation with
glutathlone, metabolites of monochlorobenzene (monophenols and dlphenols)
can conjugate with glucuronlc add or with sulfate and be excreted 1n the
urine. Monophenols are the major metabolites; the dlphenols are formed to a
lesser degree. The arene oxides, 3-chlorobenzene oxide or 4-chlorobenzene
2-5
-------
oxide, can also be converted to the dlhydrodlol by epoxlde hydrase and
dehydrogenated to form chlorocatechols. There appear to be species differ-
ences 1n the profile of urinary metabolic conjugates, and end metabolites
may vary depending on the availability of tissue glutathlone. Detoxifica-
tion by conjugation with glutathlone 1s Important 1n the modulation of toxic
effects especially at high exposure levels. Saturation of these metabolic
pathways has been demonstrated at relatively low exposure levels.
The available data for rats, rabbits and humans Indicate that the
dlchlorobenzenes are absorbed through the lungs, gastrointestinal tract and
Intact skin, though actual determinations of absorption rates were not
located 1n the available literature. Once absorbed through either Inhala-
tion or 1ngest1on, the dlchlorobenzenes are rapidly distributed to many
tissues by the systemic circulation, Including adipose, kidney, liver, lung,
heart, brain and muscle tissues. Distribution 1s primarily to adipose
tissue, which has Initial levels 10-32 times the blood concentrations and to
lung and kidney tissues to a greater extent than liver, muscle and plasma.
Single-dose and repeated exposures by both Inhalation and 1ngest1on show
similar patterns of distribution. Elimination of the dlchlorobenzenes and
their metabolites occurs within 5-6 days after exposure, although elimina-
tion from adipose tissue 1s slowest and 1,2-d1chlorobenzene and metabolites
are eliminated slightly more rapidly than 1,4-d1chlorobenzene. The
dlchlorobenzenes are primarily metabolized by hydroxylatlon to their respec-
tive dlchlorophenols, which are excreted 1n the urine 1n the form of glucu-
ronlc and sulfate conjugates. Some metabolites are excreted 1n the bile,
although the majority are then reabsorbed by the enterohepatlc pathway and
reexcreted 1n the urine. Intermediates of the metabolism of 1,2-d1chloro-
benzene, possibly arene oxides, bind to liver protein and may be Involved 1n
the Induction of hepatotoxldty.
2-6
-------
The limited comparative pharmacoklnetlc data available on the trlchloro-
benzenes prevent specification of the absorption, distribution, metabolism
and excretion of the Individual Isomers. From the available data, 1t
appears that metabolism 1n at least three animal species has a common first
step, the production of an arene oxide Intermediate. Subsequent metabolic
steps, however, vary among the species examined, at least for the most
studied Isomer, 1,2,4-tr1chlorobenzene.
In general, the pharmacoklnetlcs of the trlchlorobenzenes are similar to
those described for the other halogenated aromatlcs. These compounds are
UpophlUc and their metabolism and excretion depend on conversion to polar
metabolites. In addition, their UpophlUc character provides for ready
absorption from the gastrointestinal tract and Initial distribution to the
more highly perfused tissues, particularly the liver, after which they are
either metabolized and excreted or redistributed to adipose tissue or skin.
Additional experiments are needed to clarify the relationship of these
studies to the metabolism of trlchlorobenzenes In humans.
No studies describing the absorption, distribution, metabolism or excre-
tion of 1,2,3,4-, 1,2,3,5- or 1,2,4,5-tetrachlorobenzene following Inhala-
tion exposure were located 1n the available literature. The pharmaco-
klnetlcs of the tetrachlorobenzene Isomers following oral administration 1s
well characterized In rabbits, but not 1n other animal species. The Upo-
phlUc characteristics of the tetrachlorobenzene Isomers would allow effi-
cient transepithellal absorption at the gastrointestinal and respiratory
surfaces. Once absorbed, the tetrachlorobenzene Isomers administered orally
to rabbits were rapidly accumulated 1n fat, metabolized primarily to tetra-
chlorophenols and conjugated partly as glucuronldes and ethereal sulfates or
eliminated unchanged 1n the expired air or feces.
2-7
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No pharmacoklnetlc data were available for humans, except for a report
of 1,2,4,5-tetrachlorobenzene 1n adipose tissue (range of 0.006-0.039 mg/kg
bw; mean of 0.019 mg/kg bw) of 15 Tokyo residents. The tetrachlorobenzene
Isomers are both 1_n_ vivo and ln_ vitro metabolites of the pesticides, Undane
and hexachlorobenzene; therefore, human exposure via air, food and drinking
water may occur from the environmental degradation of these pesticides.
Although studies of the absorption of pentachlorobenzene Indicated that
absorption does occur through the gastrointestinal tract, the rate or extent
of absorption has not been determined. A study 1n rabbits Indicated that up
to 50% of a dose was absorbed within 3-4 days. Oral administration to
monkeys Indicated 95% absorption within 4 days. Absorption resulting from
Inhalation has not been studied, and absorption from dermal exposure was
found to be rather poor 1n rats. Once absorbed, pentachlorobenzene 1s dis-
tributed to many tissues, with the highest levels appearing 1n fat and bone
marrow. A study 1n rats demonstrated that transport across placental
membranes occurred readily and that accumulation of pentachlorobenzene 1n
the fetus 1s highest 1n the liver. No studies were encountered that
described the distribution of pentachlorobenzene after Inhalation or dermal
exposure.
The metabolism of pentachlorobenzene 1s not fully understood, but some
studies suggested that metabolic activity other than the hepatic cytochrome
P-450, xenoblotlc metabolizing system may be Involved. Metabolism appeared
to be primarily via oxidation to two major metabolites, pentachlorophenol
and 2,3,4,5-tetrachlorophenol, which were excreted 1n the urine. Metabolism
and excretion occurred at a slow rate; an estimated elimination half-life
for a single dose 1n primates was 2-3 months.
2-8
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The pharmacoklnetlcs of hexachlorobenzene 1n a number of mammalian
species have been studied 1n detail following oral administration and, to a
lesser extent, following Intravenous or 1ntraper1toneal Injection. No
Information was present 1n the available literature on hexachlorobenzene
metabolism following Inhalation or topical application. Absorption of hexa-
chlorobenzene from the Intestinal tract appeared to depend on the vehicle
used during test material administration. Thus, when hexachlorobenzene was
administered 1n olive oil, -80% of the dose was absorbed; when 1t was admin-
istered 1n an aqueous solution, 1n 1% methyl cellulose or 1n a crystalline
form, relatively little (<20%) was absorbed. Intestinal absorption of
hexachlorobenzene occurred primarily through lymphatic channels, with only a
minor portion being absorbed Into the portal circulation.
Following absorption, hexachlorobenzene was distributed to tissues that
have a high llpld content. The adipose tissue accumulated the greatest
concentrations of hexachlorobenzene 1n all species studied, although bone
marrow and skin, which contain large amounts of Uplds, also accumulated
hexachlorobenzene. The adrenal cortex accumulates hexachlorobenzene at
concentrations approaching those of fat. Other body compartments (e.g.,
liver, kidneys, lungs, heart, spleen and blood) generally contain much lower
amounts of hexachlorobenzene. Intravenous Injection of hexachlorobenzene
results 1n a tissue distribution similar to that seen following oral admin-
istration. Hexachlorobenzene 1s transported via the placenta and 1s dis-
tributed 1n fetal tissue as indicated by studies 1n rabbits, rats, mice,
mink and ferrets.
Hexachlorobenzene 1s metabolized slowly Into other chlorinated benzenes,
chlorinated phenols and other minor metabolites and forms glucuronlde and
glutathlone conjugates. For this reason tissues were found to contain
2-9
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mainly unchanged hexachlorobenzene together with small amounts of metabo-
lites. Similarly, only small amounts of hexachlorobenzene metabolites were
detected 1n feces, whereas most of the metabolites were excreted 1n the
urine together with small amounts of unchanged hexachlorobenzene. There are
Indications that females produce and excrete more hexachlorobenzene metabo-
lites than do males.
The excretion of hexachlorobenzene from treated animals 1s slow and
occurs mainly through the feces with relatively little being excreted 1n the
urine. It 1s characterized by an Initial rapid phase followed by one or
more slow phases. This slow phase of excretion can be enhanced by the
administration of mineral oil, paraffin or n-hexadecane. Both biliary and
Intestinal excretion contribute to fecal excretion. A three-compartment
mammlllary model has been reported for the behavior of hexachlorobenzene 1n
beagles and rhesus monkeys following 1.v. Injection of a single dose.
Radioactivity was not detected 1n exhaled air following l.p. Injection of
14C-hexachlorobenzene. Hexachlorobenzene has been detected 1n the milk of
nursing mammals.
2.1.5. Effects on Humans. No ep1dem1olog1c studies regarding the effects
of exposure to monochlorobenzene are available. Human exposure to mono-
chlorobenzene by Inhalation or by accidental 1ngest1on can cause neurotoxlc
effects. It 1s not known 1f the effects are reversible after long-term
exposure or 1f there are other sites of toxldty.
Ep1dem1olog1c data on dlchlorobenzenes are Insufficient to evaluate
dose-response association. Possible chronic effects of exposure to the
dlchlorobenzenes are Indicated by case reports of the chronic exposure of
Individuals, I.e., repeated exposures over a period of more than a year,
suggesting a common set of toxic effects, those of the ret1culoendothel1al
and hematopoletlc systems and those of the liver. Of the 23 exposure cases
2-10
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found In the literature, 17 Involved pathological changes In the blood or
liver, Including chronic lymphold leukemia, acute hemolytlc anemia, aplastlc
anemia and bone marrow hyperplasla. Although the exposures 1n these cases
are not well defined 1n time and often Involve other toxic substances,
together they suggest a common pathologic action of the dlchlorobenzenes on
bone marrow and other organs of the blood-forming system. The one available
ep1dem1olog1c study supports this generalization 1n that the reported
short-term exposure to 1,2-d1chlorobenzene (8 hours/day for 4 days) produced
alterations 1n the chromosomes of leukocytes. This study did not establish
an association between chromosomal alterations and the pathologic changes
that characterize the case studies.
Human exposure to 1,2,4-tr1chlorobenzene at 3-5 ppm causes eye and
respiratory Irritation. The only other data on human exposure are Individ-
ual case reports of aplastlc anemia of persons exposed occupatlonally or
domestically.
Only one ep1dem1olog1c study was available regarding the effects of the
tetrachlorobenzenes on humans and this study examined peripheral lymphocytes
for chromosomal abnormalities 1n blood. The blood was collected from
Hungarian workers engaged 1n the production of 1,2,4,5-tetrachlorobenzene.
There were observed chromosome aberrations 1n the lymphocytes; however, no
airborne concentrations or exposures were determined.
No ep1dem1olog1c or case studies of effects on humans resulting from
exposure to pentachlorobenzene were available for review.
A few ep1dem1olog1c studies with occupatlonally-exposed workers have
been reported, together with studies conducted 1n Turkey and 1n the United
States (I.e., Louisiana) on the general population following accidental
exposure to hexachlorobenzene. These studies qualitatively support the
toxlclty of hexachlorobenzene, but give Uttle dose-response Information.
2-11
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Biological monitoring of plasma levels clearly show more hexachlorobenzene
1n the plasma of exposed compared to nonexposed Individuals, although no
biologically significant adverse health effects were seen during the obser-
vation periods. The exposure of humans to hexachlorobenzene 1n Turkey
during 1955-1959 caused an epidemic of hexachlorobenzene-lnduced porphyrla
cutanea tarda (PCT), also known as porphyrla turdca, which 1s manifested by
disturbed porphyrln metabolism, cutaneous lesions and hyperp1gmentat1on.
The authors estimated that from 0.05-0.2 g/day were Ingested. In exposed
children under 1 year of age, pink sore was observed as well as 95% mortal-
ity 1n these Infants.
Follow-up studies conducted with patients 20-25 years after the onset of
porphyrla showed that a few subjects still had active porphyrla, whereas
>SO% exhibited hyperplgmentatlon scarring, as well as other dermatologlc,
neurologic and skeletal features of hexachlorobenzene toxlclty, Hexachloro-
benzene residues were also found In the blood, fat and breast milk of some
patients.
A correlation was found between hexachlorobenzene levels In blood and
the number of years worked In a chlorinated solvents plant. The concentra-
tion of urinary uroporphyrlns and coproporphyrlns ranged from 21-37 and
67-101 vig/H, respectively, for the period between 1974 and 1977. An
ep1dem1olog1c survey conducted with 86 residents 1n the vicinity of this
chlorinated solvents plant showed elevated hexachlorobenzene residues In
plasma. Higher levels of hexachlorobenzene residues were found 1n males
than 1n females, but these were not associated with race or food consumption.
2.1.6. Mammalian Toxicology. Acute exposure to monochlorobenzene by
Inhalation causes sensory Irritation of the respiratory system after a few
minutes; exposure for several minutes to several hours causes narcosis and
2-12
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central nevous system depression, which can result 1n death. Monochloroben-
zene Is also toxic by the oral or parenteral routes. Systemic effects of
acute toxic doses Include kidney damage. Subchr'onlc Inhalation exposure at
1.0 mg/m3 {contlnously for 60 days) causes neurotoxlc effects In rats, an
Increase 1n blood chollnesterase and abnormal chronaxla of the muscles.
Repeated exposure of rats to monochlorobenzene at 250 ppm {1157 mg/m3)
causes slight changes 1n the liver, kidneys and adrenal cortex. Repeated
oral dosing of rats or dogs {100-200 mg/kg/day) causes some toxic manifesta-
tion 1n the liver and kidneys. Gavage administration of monochlorobenzene
to mice and rats 5 times/week for 13 weeks resulted 1n Increased mortality
1n the higher dose groups (>250 mg/kg), urinary porphyrla and dose-dependent
Injury to the liver, kidney, bone marrow, spleen and thymus. A set of sim-
ilar studies were conducted 1n mice and rats for 2 years and resulted 1n
some Increased mortality 1n the male monochlorobenzene exposed groups when
compared with controls. Only equivocal evidence for mild monochlorobenzene-
Induced hepatocellular necrosis was found 1n rats.
Although one study 1n Streptomyces antlblotlcus found monochlorobenzene
to Induce reversion to vitamin B, prototrophy and one study 1n Saccharo-
myces cerevlslae showed Induction of DMA damage, several other studies using
bacterial, fungal and mammalian tissue culture systems were negative. The
carcinogenic activity of monochlorobenzene was tested by the NTP bloassay
program 1n two rodent species at doses of 60 and 120 mg/kg bw/day 1n male
and female rats and female mice, and at 30 and 60 mg/kg bw/day 1n male mice,
Carc1nogen1c1ty was not definitively demonstrated for monochlorobenzene 1n
this study, but high dose male rats had a significant Increase 1n neoplastlc
nodules of the Hver.
Repeated exposures to monochlorobenzene at 2.0 mg/a. {vapors) or 272.5
mg/kg/day (oral) were found to cause atrophy of the epithelial tissue 1n the
2-13
-------
seminiferous tubules and decreased spermatogenesls 1n dogs and rats and
Increased gonad weight/body weight ratios 1n female rats. These effects 1n
dogs, however, were seen only at levels sufficiently toxic that the dogs
died or were moribund.
Studies of the acute and subchronlc toxldty of the dlchlorobenzene
"Isomers Indicate that generally these compounds have similar target organs
and effects. At oral doses ranging from 125-1000 mg/kg over periods of up
to 6 months, the dlchlorobenzenes cause central nervous system depression,
Injury to liver, kidney, heart, thymus and spleen, and hepatic porphyrla;
however, one study reported that a dose of 0.01 mg/kg over a 5-month period
Inhibited erythropolesls and bone marrow activity. The subchronlc oral
toxlclty studies 1n rats provide two estimates of no-observed-effect level
(NOEL) values: 0.001 mg/kg for l,4-d1chlorobenzene and 18.8 mg/kg for 1,2-
and for l,4-d1chlorobenzene. The National Toxicology Program (NTP, 1982)
subchronlc oral study on l,2-d1chlorobenzene 1n mice provided higher esti-
mated NOEL values of 125 and 250 mg/kg for males and females, respectively.
A 2-year NTP chronic oral gavage study on l,2-d1chlorobenzene In rats and
mice, conducted primarily as a carclnogenesls bloassay at the 60 and 120
mg/kg dose levels, resulted only 1n Increased mortality 1n the male rats
given 120 mg/kg. Acute and subchronlc Inhalation studies of dlchloroben-
zenes Indicate similar toxic effects and target sites as seen 1n the oral
studies. The effects occurred at doses >900 rag/m3; Inhalation NOELs were
reported as 580 mg/m3 and -450 mg/ma for l,4-d1chlorobenzene, and 290
mg/ra3 for l,2-d1chlorobenzene.
Studies of the mutagenlc activity of dlchlorobenzenes show little or no
activity 1n a range of bacterial systems, Including Salmonella, with and
without metabolic activation. However, these studies were lacking 1n exper-
imental detail. Several studies with mold and plant cultures treated with
2-14
-------
dichlorobenzenes have reported mutations and chromosomal alterations. The
carcinogenic activity of l,2-d1ehlorobenzene, was tested 1n the NTP bloassay
program 1n two rodent species at doses of 60 and 120 mg/kg. No evidence of
carcinogenic activity was found under the test conditions. The carcinogen-
Id ty of l,4-d1chlorobenzene was tested 1n two rodent species using long-
term Inhalation exposure. Again, no evidence for carc1nogen1c1ty was noted.
Since neither study may have used the maximum tolerated dose, the evidence
must be considered Inadequate for developing conclusions concerning the
carc1nogen1c1ty of 1,2- or 1,4-d1chlorobenzene 1f the IARC criteria for
classifying carcinogens are used.
The effects 1n mammals of acute exposure by various routes to trlchloro-
benzenes Include local Irritation, convulsions and death. Livers, kidneys,
adrenals, mucous membranes and brain ganglion cells appear to be target
sites with effects Including edema, necrosis, fatty Infiltration of livers,
Increased organ weights, porphyrln Induction and mlcrosomal enzyme Induction.
Quantitative data on the toxic effects of trlchlorobenzene following
subchronlc exposure by various routes were obtained 1n a variety of species.
In general, these studies Indicate that the liver and kidney are target
organs. Inhalation of 1,2,4-trlchlorobenzene at >74.2 mg/m3 (10 ppm) for
6 hours/day, 5 days/week for up to 26 weeks Induced hepatocytomegaly and
hyaline degeneration 1n several species, although these effects may be to
some extent reversible. One study Identified 22.3 mg/m3 {3 ppm) as a no-
observed-adverse-effect level (NOAEL) 1n rats, while another study reported
that some rats exposed by Inhalation to 1,3,5-trlchlorobenzene at 7423
mg/m3 (1000 ppm) for 13 weeks showed squamous metaplasia and focal hyper-
plasla of the respiratory epithelium, which appeared to be reversible.
2-15
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Subchronlc oral studies have also found that the trlchlorobenzenes Induce
hepatic xenoblotlc metabolism and porphyrla. Subchronlc dermal exposure
resulted 1n mild to moderate Irritation.
One chronic study on the effects of trlchlorobenzene (0.03 ml) painted
on the skin of mice for 2 years reported Increased mortality 1n females at
the low dose (30% solution 1n acetone) and 1n both sexes at the high dose
(60K solution).
Results of two reports on mutagenlclty tests with Salmonella tvphlmurlum
test strains were negative. However, this test system 1s generally Insensi-
tive to chlorinated compounds. One carc1nogen1c1ty study, a 2-year skin
painting study In mice, failed to demonstrate a conclusive tumor1gen1c
effect. A multlgeneratlon study of the reproductive effects of oral expo-
sure of rats to trlchlorobenzene failed to show effects on reproduction.
Oral teratogenldty studies 1n rats showed mild osteogenlc changes 1n pups
and significantly retarded embryonic development as measured by fetal growth
parameters.
The only mammalian toxicology data available for tetrachlorobenzenes are
the result of oral exposures. The oral LDgQ for 1,2,4,5-tetrachloroben-
zene was reported as 1035 mg/kg In mice and 1500 mg/kg 1n rats and rabbits
when administered In sunflower oil and 2650 mg/kg 1n mice when administered
1n 1.5% starch solution. Subchronlc oral exposure of rats and rabbits to
1,2,4,5-tetrachlorobenzene resulted In statistically significant effects on
biochemical parameters, Including retlculocytosls, Increased blood chollnes-
terase activity, erythremla and an Indication that glycogen formation was
Impeded; at higher doses of 1,2,4,5-tetrachlorobenzene, rats also had
Increased kidney and liver weights, and renal and hepatic hlstologlc changes.
2-16
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Reversible effects on serum alkaline phosphatase and total blUrubln
were reported In dogs given 5 mg/kg bw/day 1,2,4,5-tetrachlorobenzene 1n the
diet for 2 years.
1,2,4,5-Tetrachlorobenzene was not mutagenlc 1n the sex-linked recessive
lethal assay with Drosophlla melanogas ter. However, because only an
abstract of the study was available, experimental details were too sparse to
permit an evaluation of this negative result. Both 1,2,3,5- and 1,2,4,5-
tetrachlorobenzenes were negative 1n the reverse mutation assay with Salmo-
nella typh1mur1um strains TA98, TA100, TA1535, TA1537 and TA1538. These
results were reported 1n an abstract with Insufficient experimental detail.
Also, a negative result for chlorinated compounds 1n the Salmonella rever-
sion assay 1s not unexpected.
No Information was available regarding the carclnogenlclty of any of the
three tetrachlorobenzene Isomers In either animals or humans.
The tetrachlorobenzene Isomers have been found to Induce appreciable
maternal toxldty, mild fetotoxlclty and negligible teratogenlcl ty 1n rats
following oral administration.
Oral ID™ values were determined for pentachlorobenzene In adult rats
(1080-1125 mg/kg) and mice (1175-1370 mg/kg), and for weanling rats (940
mg/kg). No clinical signs of toxldty were observed 1n adult rats following
dermal application of 2500 mg/kg pentachlorobenzene. Also, 1t was demon-
strated that pentachlorobenzene caused an Increase 1n the liver content of
cytochrome P-450, mlcrosomal drug metabolizing enzymes and mlcrosomal
proteins.
A subchronlc feeding study Indicated that the primary toxic effects are
on the liver and kidneys, although slight changes 1n some hematologlc param-
eters (e.g., decreased erythrocyte count, hemoglobin and hematocrlt; and
2-17
-------
Increased leukocyte count) occurred 1n the high dose groups, H1stolog1c
examination Identified pathologic changes 1n the livers of the female rats
fed 500 and 1000 ppm In the diet for 180 days and 1n the 1000 ppm male rats
treated for 100 days. These data were sufficient to Identify a subchronlc
lowest-observed-adverse-effect level (LOAEL) of 500 ppm (-27-63 mg/kg/day)
and a NOEL of 250 ppm (-16-31 mg/kg/day).
No mutagenlc activity was detected 1n five strains of Salmonella, typhl-
murlum when tested at five unspecified concentrations of pentachlorobenzene
1n the presence and absence of rat Hver mlcrosomes Induced by Aroclor 1254.
These results were reported 1n an abstract with Insufficient experimental
details presented. A negative result 1s not unexpected, because the Salmo-
nella test system has been found to be generally Insensitive to chlorinated
compounds.
Studies also have shown that pentachlorobenzene 1s capable of causing
reproductive and developmental effects. Female rats fed diets containing
pentachlorobenzene during mating and gestation produced Utters with reduced
pup survival and body weights at weaning, and Increased Hver-to-body weight
ratios. No adverse effects were observed 1n the offspring of the dams
exposed to 125 ppm (6-16 mg/kg/day).
Single oral doses of pentachlorobenzene given dally to pregnant rats
durlng-gestatlon Increased the Incidence of fetal death at all tested doses,
Identifying a LOAEL of 50 mg/kg/day. Sternal defects and an Increase In the
Incidence of extra ribs also were observed at doses of 200 mg/kg/day and 50,
100 and 200 mg/kg/day, respectively.
In a study of possible reproductive and teratogenlc effects, doses of 50
and 100 mg/kg/day of pentachlorobenzene administered by gavage to pregnant
mice had no adverse effect on fetal development or survival of the pups.
2-18
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The acute oral toxlclty of hexachlorobenzene has been found to be low
with LD-g values ranging from 1700-10,000 mg/kg. Subchronlc oral toxlclty
studies with a number of mammalian species Indicated a significant Increase
In liver and kidney weights 1n hexachlorobenzene-treated animals. Some
studies have shown Increases 1n other organ weights as well. The livers
from hexachlorobenzene-exposed animals have shown hlstologlc changes such as
Irregular shaped and moderately enlarged liver mitochondria and Increases In
the size of the centrllobular hepatocytes. Chronic toxlclty studies
revealed similar effects to those seen 1n the subchronlc studies, plus
hexachlorobenzene associated life-shortening and various hepatic and renal
pathologies. These subchronlc and chronic effects were usually dose-
related. Other effects Included multiple alopecia and scabbing, together
with neurologic effects 1n rats, mice and dogs. A dose-related hlstopatho-
loglc change 1n the ovaries of monkeys has also been reported.
Increased porphyrln levels 1n the liver and 1n urine have been reported
for all species studied except the dog. Hexachlorobenzene was found to
cause the accumulation of B-H-stero1ds which Induce porphyrln biosynthesis
and to Inhibit uroporphyrlnogen decarboxylases. The Inhibition of uropor-
phyrlnogen decarboxylase appears to be from pentachlorophenol, a hexachloro-
benzene metabolite. Indications are that females are more susceptible to
hexachlorobenzene-lnduced porphyrla than are males, which may be related to
the females estrogen levels and greater hexachlorobenzene metabolism.
Hexachlorobenzene was reported to produce a mixed-type Induction of cyto-
chromes resembling that produced by a combination of phenobarbltal (P-450)
and 3,4-benzpyrene (P-448), In addition, the activities of several hepatic
mlcrosomal enzymes were found to be Induced by hexachlorobenzene.
2-19
-------
Hexachlorobenzene did not Induce dominant lethal mutations In two stud-
ies but was reported to be mutagenlc In a yeast, S.. cerevlslae. assay at a
concentration of 100 ppm. Hexachlorobenzene possessed no detectable levels
of mutagenlc activity 1n the Salmonella h1st1d1ne reversion assay. The
chronic toxldty studies provide sufficient evidence of the carc1nogen1c1ty
of hexachlorobenzene 1n animals since there was an Increased Incidence of
malignant tumors of the liver 1n two species (haemang1oendothel1oma In ham-
sters and hepatocellular carcinoma 1n rats) as well as reports of hepatoma
1n mice, rats and hamsters. Hexachlorobenzene given to pregnant mice was
found to produce cleft palates and renal agenlsls 1n exposed pups. Cer-
tain chemicals were found to alter the toxldty of hexachlorobenzene 1n mam-
mals, whereas hexachlorobenzene pretreatment was reported to Increase CC1,
toxldty and alter the Immune responses of treated animals.
2.2. CONCLUSIONS
The chlorinated benzenes are a group of 12 cyclic aromatic compounds 1n
which 1-6 hydrogen atoms of a benzene ring have been replaced by up to six
chlorine substltuents. As the benzene ring Is Increasingly chlorinated
there are physlochemlcal trends towards Increased melting points, boiling
points, densities and log partition coefficients, and decreased volatility
and water solubility of the compounds.
A wide range and severity of chlorinated benzenes-Induced health effects
have been reported In rodents and other laboratory animals. Some of these
same effects have also been observed 1n chlorinated benzenes-exposed humans
as well, but the human reports are not as extensive or complete as the
animal studies. A review of the animal chlorinated benzenes health effects
literature also Indicates that there are some large data gaps existing for
several of the chlorinated benzene Isomers, especially for 1,3-dlchloro-
benzene, the trlchlorobenzenes and the tetrachlorobenzenes. The animal
2-20
-------
studies Indicate a trend of Increasing toxldty with Increased chlorlnatlon
of the benzene ring, e.g., hexachlorobenzene 1s more porphyrlnogenlc than
monochlorobenzene. Adequate evidence of the carc1nogen1c1ty of the
different chlorinated benzenes has only been shown for hexachlorobenzene.
Hexachlorobenzene has been classified as a probable carcinogen 1n humans.
2.3. NEEDS FOR FUTURE RESEARCH
In the development of this document and previous drafts, there have been
many comments on the need to complete certain studies and to Initiate other
research. These new data would refine the known Information and give scien-
tists a better understanding of the effects of the chlorinated benzenes and
their properties. Some of the health-related data might become available as
Indicated In 48 FR 54836. However, as the result of this document and Its
review, the following research needs were Identified which would yield data
that would provide further Information on the specific nature and health
effects of the chlorinated benzenes, as well as help to resolve many remain-
Ing unknowns.
Further studies should be conducted to determine detailed pharmaco-
klnetlcs of each of the chlorinated benzene Isomers (I.e., absorp-
tion, distribution, metabolism and excretion).
Further studies should be conducted to determine more thoroughly
the long-term toxldty and, 1n some cases, the carc1nogen1c1ty of
many of the chlorinated benzene Isomers, except for hexachloro-
benzene where sufficient data already exists.
Further mutagen1c1ty studies should be conducted on those chlori-
nated benzene Isomers which do not have sufficient mutagenlclty
data available.
Studies should be conducted to assess the potential of the chlori-
nated benzenes to cause ONA damage.
Teratogen1c1ty, fetotoxlclty and reproductive studies should be
conducted using various routes of exposure, with emphasis on the
Inhalation route, on all the chlorinated benzene Isomers.
2-21
-------
Studies on the neurotoxlc effects of the chlorinated benzene
Isomers should be conducted using various routes of exposure, with
emphasis on the Inhalation route.
Studies should be conducted to assess for possible chlorinated
benzenes effects on alterations to the endocrine, hematopoletlc and
Immunologlc systems 1n humans and animals.
Further studies need to be conducted on the porphyrla-produdng
properties of the chlorinated benzenes [I.e., the properties of the
chlorinated benzene molecules or their metabollte(s) which are
responsible for this adverse health effect 1n humans and animals].
Investigations need to be conducted Into the quantitative
structure-activity relationships of the chlorinated benzenes with
an effort to relate biological and health effects to physlochemlcal
properties.
Studies are needed to Identify the extent of human exposure from
each of the chlorinated benzene Isomers and the relative contri-
bution of the various environmental medlas to the total human
exposure.
Exposure and health assessments of Indoor air pollution by chlori-
nated benzenes need to be made. This Is Important especially for
the dlchlorobenzenes which are present In household space deodor-
ants and moth repellents,
Epidemiclogic studies need to be conducted on Individuals who are
occupationally exposed to the chlorinated benzenes, with particular
emphasis on those adverse health effects already observed In the
human and animal studies.
Further follow-up studies are needed concerning the health of the
Turkish Individuals who were exposed to hexachlorobenzene In the
1950's, with particular emphasis on their cancer Incidences.
2-22
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3. PHYSICAL AND CHEMICAL PROPERTIES/ANALYTICAL METHODOLOGY
The chlorinated benzenes are the group of substituted benzene compounds
1n which 1-6 hydrogen atoms of benzene are replaced by chlorine atoms with
no substUuents present other than chlorine and hydrogen. The chlorlnatlon
of benzene can yield 12 different compounds: monochlorobenzene
(CcHrCl); 1,2-, 1,3- and 1,4-d1chlorobenzene (M,C1 ); 1,2,3-,
b b 642
1,2,4- and 1,3,5-tr1chlorobenzene (C6H3C13>J 1,2,3,4-, 1,2,3,5- and
1,2,4,5-tetrachlorobenzene (C6H2C1«); pentachlorobenzene (C6HC15);
and hexachlorobenzene (CtCl,). The chemical structures of these com-
0 0
pounds are shown 1n Figure 3-1.
3.1. SYNONYMS, TRADE NAMES AND IDENTIFICATION NUMBERS
Synonyms, trade names and Identification numbers for the 12 chlorinated
benzenes are listed 1n Table 3-1.
3.2. PHYSICAL AND CHEMICAL PROPERTIES
Some physical properties of the chlorobenzenes are shown 1n Tables 3-2
and 3-3. In general, the chlorinated benzenes have low water solubility,
moderate to high octanol/water partition coefficients and low to moderate
vapor pressures at 25°C, and low flammablllty. Apart from hexachloroben-
zene, they are considered to be volatile compounds because their Henry's Law
constants are greater than 10~4 atm m3 q • mol"1 (MacKay et al.,
1979).
The chlorobenzenes are chemically very unreactlve compounds and are
generally stable under ambient conditions 1n the laboratory. Because of the
electron-withdrawing character of the chlorine atom relative to carbon, the
chlorobenzenes are highly resistant to electrophHlc attack (e.g., chlorlna-
tlon), and each additional chlorine substltuent further lowers the reactiv-
ity of these compounds. Hydroxylatlons occur only at high temperatures 1n
3-1
-------
o
NONOCHLOftO*tN2ENE
o
1. 2-D4CHLOAMENZENE 1.1-OICHI.OftMENZENE 1,4-O
-------
TABLE 3-1
Synonyms, Trade Names and Identification Numbers of the Chlorinated Benzenes3
Chemical
Identification Number
Synonyms and Trade Names
Honochlorobenzene
CAS No. 108-90-7
TSL No. CZ017500
NCI No. C54886
EPA Haz Waste No. U037
EPA Haz Waste No. F002
DOT Haz Mat No. UNI 134
Dlchlorobenzene
1,2-
CAS No. 95-50-1
TSL No. CZ4500000
NCI No. C54944
EPA Haz Waste No.
EPA Haz Waste No.
U070
F002
DOT Haz Mat No. UNI591
Chlorobenzene
Benzene chloride
Phenyl chloride
Chlorobenzol
MCB
Chlorbenzene
Monochlorbenzene
Benzene, chloro-
Chlorobenzeen (Dutch)
Chlorobenzene (Polish)
Clorobenzene (Italian)
Monochlorobenzene (Dutch)
Monochlorobenzol (German)
Monochlorobenzene (Italian)
£-01Chlorobenzene
o-D1chlor benzol
DCB
Dowtherm Eb
ODB
o-DCB
p_-D1chlorobenzol
OrthodlChlorobenzene
OrthodlChlorobenzol
Chlorobenb
D1zeneb
Dlchlorobenzene, ortho, liquid
Special Termite Fluid
Term1tk1l
Cloroben
Benzene, 1,2-d1chloro-
Benzene, £-d1chloro-
ODCB
D1lant1n DB
3-3
-------
TABLE 3-1 (cont.)
Chemical
Identification Number
Synonyms and Trade Names
Dlchlorobenzene
1,3-
CAS No. 541-73-1
EPA Haz Waste No. U071
1,4-
CAS No. 106-46-7
TSL No. C24550000
NCI No. C54955
EPA Haz Waste No. U072
DOT Haz Hat No. UNI592
Benzene, m-d1chloro-
Benzene, I,3~d1chloro-
m-Phenylene dlchlorlde
m-D1chlorobenzol
m-D1chlorobenzene
ineta-D1chlorobenzene
D1-chlor1c1de
Paramoth
p_-D1chlorobenzene
PDB
Paraclde
Paradlchlorobenzene
Paradl
Paradow
Santochlor
P.-DC8
p_-D1chlorobenzeen (Dutch)
l,4-01chloorbenzeen (Dutch)
p_-D1chlorbenzol (German)
l,4-D1chlor-benzol (German)
p_-D1chlorobenzol
Dlchlorobenzene, para, solid
l,4-D1chlorobenzene (Italian)
p_-D1clorobenzene (Italian)
para Crystals
Paradl chlorobenzo'l
Paranuggets
Parazene
Benzene, p_-d1chloro-
Benzene, 1,4-d1chloro-
p_-Chlorophenyl chloride
Evola
Persla-Perazol
3-4
-------
TABLE 3-1 (cont.)
Chemical
Identification Number
Synonyms and Trade Names
TMchlorobenzene
1,2,3-
Trlchlorobenzene
1,2,4-
1,3,5-
Tetrachlorobenzene
1,2,3,4-
1,2,3,5-
1,2,4,5-
Pentachlorobenzene
CAS No. 87-61-6
CAS No. 120-82-1
TSL No. DC2100000
CAS No. 108-70-3
CAS No. 634-66-2
CAS No. 634-90-2
CAS No. 95-94-3
TSL No. DB9450000
EPA Haz Waste No. U207
CAS No. 608-93-5
EPA Haz Waste No. U183
TSL No. OA6640000
v1c-Tr1ch1orobenzene
1,2,6-Tr1chlorobenzene
v-Tr1chlorobenzene
Benzene, 1,2,4-tr1chloro-
asym-Trlchlorobenzene
TCB
Trojchlorobenzen (Polish)
1,2,4-Tr1chlorobenzol
Hostetex L-Pec
s-Tr1chlorobenzene
sym-Tr1chlorobenzene
TCB
TCBA
Benzene, 1,3,5-tr1chloro-
Benzene, 1,2,3,4-tetrachloro-
Benzene, 1,2,3,5-tetrachloro-
Benzene tetrachlorlde
Benzene, 1,2,4,5-tetrachloro-
s-Tetrachlorobenzene
1,2,3,4,5-Pentachlorobenzene
QCB
Benzene, pentachloro-
Qulntochlorobenzene
3-5
-------
TABLE 3-1 (cont.)
Chemical
Identification Number
Synonyms and Trade Names
Hexachlorobenzene
CAS No. 118-74-1
TSL No. DA2975000
EPA Haz Haste No. U127
Esaclorobenzene (Italian)
Amatln
An11carle
Bunt-Cure
Bunt-No-More
Co-op Hexa
Granox NM
HCB
HEXA C.B.
Hexachlorobenzol {German)
Hexachlorobenzene
Julln's Carbon Chloride
No Bunt
No Bunt 40
No Bunt 80
No Bunt Liquid
Pentachlorophenyl Chloride
Perchlorobenzene
Phenyl Perchloryl
Sanoclde
Smut-Go
Sn1ec1otox
aSource: National Library of Medicine (NLH), Toxicology Data Bank (TDB)
^Formulations which contain 1,2-d1chlorobenzene
3-6
-------
TABLE 3-2
Physical Properties of the Chlorinated Benzenes3
Chemical
Monochlorobenzene
Dlchlorobenzene
1,2-
1,3-
1,4-
Irlchlorobenzene
1,2,3-
1,2,4-
1,3,5-
Tetrachlorobenzene
1,2,3,4-
1,2,3,5-
1,2,4,5-
Pentachlorobenzene
Hexachlorobenzene
Molecular
Height
112.56
147.01
147.01
147.01
181.46
181.46
181.46
215.90
215.90
215.90
250.34
284.79
Melting
Point
CO
-45.6
-17.0
-24.7
53.1
52.6
16.95
63.4
47.5
54.5
139.5
86
230
Boiling
Po1ntb
CC)
132
180.5
173
174
221
213.5
208.4
254
246
246
277
322.9
Henry's Law
Oens1tyc Constantd x 10
(g/ral) (atm m3 moT1}
1.1 2.6
1.30 1.3
1.28(25)
1.25 2.4
1.69 1.0
1.45 4.3
1.39(64)m
NA
NA
1.86(22)
1.83(16.5)
1,57(23) 0.12
-a Log pod
2.84f
3.38f
3.38f
3.39f
4.121
NA
NA
NA
5.63"
5.8'
Water
Solubility
(mg/i)e
500(20)9
1459
1239
799
31. 5k
34. 6k
6.6k
4.3k
3 5k
0.60k
'0.56k
0.005k
Flash
Point
CC or °F)
85 F/ccn :
151 F/cc
NA
150 F/cc
113 C
110 C
107 C
NA
311 F
311 F
NA
468 F
Index of
Refraction
at CC)
1.5241(20)
1.5515(20)
1.5459(20)
1.5285(60)
1.5776(19)
1.5717(20)
1.5662(19)
NA
NA
NA
NA
NA
(lala are from the National Library of Medicine (NLM), Toxicology Data Bank (TOB), except as noted.
"At 760 mm
cAt 20°C, except as noted
dMacKay et al., 1979
eAt 25"C, except as noted
fLeo et al., 1971
^Verschueren, 1977
These are data from closed cup (cc) experiments .
Monsanto, 1978
•'isomer unspecified
kYalkowsky and Valvanl, 1980
Hansch and Leo, 1981
"Wvath, 1982
"U.S. EPA, 1980t>
P° = Partition coefficient at 25°C
NA = Not available
-------
TABLE 3-3
Vapor Pressures and Vapor Densities of the Chlorinated Benzenes
Chemical Vapor Pressure Specific Vapor Density
(mm Hg) (air = 1)
Honochlorobenzene 8.8 at 20°Ca 3.88a'b'c
10 at 22.2°Cb 3,9d
11.8 at 25°Cb
15 at 30°Ca
Dlchlorobenzene
1,2- 1 at 20°Ca 5,05b
1.28 at 25°ce 5.0?a.c
1.5 at 25°Ca
1.9 at 30°ca
1,3- 1 at 12.1°Cb 5.08b
1.89 at 25°Cd
1,4- 0.6 at 20°Ca 5.07C
1.0 at 25°Cf 5.08b
1.8 at 30°Ca
Trlchlorobenzene
1,2,3- 0.07 at 25°Cd 6.26b
1 at 40°Cb
1,2,4- 0.29 at 25°C<1 6.26b
1 at 38.4°Cb
1,3,5- 0.15 at 25°Cd 6.26b
10 mm at 78°Cb
Tetrachlorobenzene
1,2,3,4- 1 at 68.5°C9 NA
0.04 at 25°Ch
1,2,3,5- 1 at 58.2°C9 NA
0.07 at 25°Ch
1,2,4,5- 0.05 at 25°C1 7.4b
0.05 at 25°Ch
3-8
-------
TABLE 3-3 (cont.)
Chemical Vapor Pressure Specific Vapor Density
(mm Hg) (air = 1)
Pentachlorobenzene 1 at 98.6°C9 NA
Hexachlorobenzene 1 at 114°C9 9.84a
1.68xl(T5 at 25°C3
1.089xlO~s at 20°Ck
aVerschueren, 1977
bSax, 1979
cLowenhe1m and Moran, 1975
dNLM, 1982a
eR1chardson, 1968
^Martin and Worthing, 1977
9Weast, 1980
hMacKay et al., 1982
1Ware and West, 1977
^Leonl and Darca, 1976
kFarmer et al., 1980
NA = Not available
3-9
-------
very alkaline conditions. A description of each of the chlorinated benzenes
follows.
Honochlorobenzene, which 1s the most polar of the chlorinated benzenes,
1s a colorless, volatile liquid with a pleasant almond-Uke odor that 1s
classified as a flammable liquid by the U.S. Department of Transportation
(NLH, 1982a). Honochlorobenzene 1s soluble 1n water to the extent of 499+8
mg/a, between 20 and 30°C {Verschueren, 1977). It 1s mlsclble 1n all
proportions 1n ethyl alcohol and dlethyl ether, and 1s very soluble 1n car-
bon dlsulflde and benzene (NLH, 1982a). No established trade specifications
exist for monochlorobenzene. Kao and Pottenberger (1979) reported two
Impurities for a typical analysis of monochlorobenzene: dlchlorobenzenes at
<0.1 wt percent and benzene at <0.05 wt percent. This Implied a purity of
99.8% or higher for the sample. A product data sheet (Dow Chemical Company,
1977) listed a 99.9% purity for monochlorobenzene, while Allied Chemical
Corporation (1973) stated a purity of 99.0% for Us product (U.S. EPA, 1980a)
1,2-D1chlorobenzene 1s a clear, volatile liquid with a pleasant odor
(NLH, 1980) and 1s combustible. It has a solubility of 145 mg/a 1n water
at 25°C {Verschueren, 1977). 1,2-D1chlorobenzene 1s mlsclble with alcohol,
ether, benzene, carbon tetrachlorlde, and acetone {NLH, 1980). The lack of
Industry-wide standards of purity for this chlorinated benzene 1s Illus-
trated by the compositions reported for 1,2-d1chlorobenzene by different
sources shown 1n Table 3-4.
1,3-D1chlorobenzene 1s a colorless liquid that 1s combustible. It can
react violently with aluminum (NLH, 1981a). It has a solubility of 123
mg/2. 1n water at 25°C (Verschueren, 1977). l,3-D1chlorobenzene 1s soluble
1n alcohol, ether and benzene, and 1s mlsclble with acetone, carbon tetra-
chlorlde and petroleum ether (NLH, 1981a).
3-10
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TABLE 3-4
Reported Composition of Commercial 1,2-D1chlorobenzene
Composition (%}
Constituent
C6H5C1
1,2-C6H4C12
lt3-C6H4Cl2
1,4-C6H4C12
C6H3C13 (a11 Comers)
1,2,4-C6H3C13
Standard
Grade3
NA
80
2
17
NA
NA
Standard
6radeb
0.07
82.7
0.5
15.4
1.6
NA
Mechanical
6radec
NA
75-85
0.5
15-25
NA
NA
High Purity
6radec
NA
99.0
"balance"
NA
NA
NA
Technical
6raded
<0.05
80.0
<19.0
NA
<1.0
NA
Purified
Graded
<0.05
98.0
NA
NA
NA
<0.2
aDow Chemical Company, 1977
bAH1ed Chemical Company, 1973
CMCA, 1974
dKao and Poffenberger, 1979
NA = Not available
-------
1,4-D1chlorobenzene 1s a combustible crystalline solid that tends to
sublime at ordinary temperatures. It possesses a distinctive odor that 1s
noticeable at concentrations between 30 and 60 ppm (NLM, 1981a). It has a
solubility of 79 mg/a. 1n water at 25°C (Verschueren, 1977). It 1s soluble
at 25°C 1n ether, chloroform, carbon dlsulflde and benzene, and 1s mlsdble
with alcohol and acetone (NLM, 1981b). The commercially available technical
grade 1,4-d1chlorobenzene may contain <0.5 wt percent of the other two
Isomers and also may contain <0.1 wt percent of monochlorobenzene and
trlchlorobenzene (Kao and Poffenberger, 1979). A product data sheet (Dow
Chemical Company, 1977) stated a purity of 99.95% for that company's 1,4-
dlchlorobenzene. Product Information from Montrose Chemical (1972)
described a mixture of 35% 1,2-d1chlorobenzene and 65% 1,4-d1chlorobenzene
(U.S. EPA, 1980a).
1,2,3-Tr1chlorobenzene 1s a white crystalline solid (platelets from
alcohol) that 1s volatile with steam. It 1s slightly soluble (31.5 mg/a)
at 25°C 1n water, slightly soluble 1n alcohol, soluble 1n benzene and carbon
dlsulflde, and very soluble 1n ether (NLM, 1981e; Yalkowsky and Valvanl,
1980).
1,2,4-Tr1chlorobenzene 1s a colorless liquid at 25°C but may also take
the form of rhombic crystals because of Its low melting point of 16.95°C.
It possesses a distinctive odor, similar to that of 1,4-d1chlorobenzene, and
1s considered volatile with steam (NLM, 1981f). It 1s slightly soluble 1n
water, 34.6 mg/fc at 25°C (Yalkowsky and Valvani, 1980); mlsdble with
benzene, petroleum ether and carbon dlsulflde; slightly soluble 1n ethanol;
and very soluble 1n dlethyl ether (NLM, 1981f). An Information sheet (Dow
Chemical Company, 1977) listed a purity of 100% for Us product. Kao and
Poffenberger (1979) reported that commercial 1,2,4-tr1chlorobenzene may
3-12
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contain monochlorobenzene (<0,1 wt percent) and d1~ and tetrachlorobenzenes
(<0.5 wt percent and <0,5 wt percent) with the 1,2»4-tr1chlorobenzene
content being around 97%.
1,3,5-Tr1ch1orobenzene takes the physical form of white crystals or
needles. It 1s very slightly soluble (6.6 mg/a at 25°C) 1n water; spar-
ingly soluble 1n alcohol; and soluble In ether, benzene, petroleum ether,
carbon dlsulflde and glacial acetic add (NLM, 1982c; Yalkowsky and Valvanl,
1980).
1,2,3,4-Tetrachlorobenzene 1s a white crystalline solid that appears as
needles from alcohol (NLM, 1981c). It 1s very slightly soluble In water
(4.3 mg/a, at 25°C); slightly soluble 1n alcohol; soluble 1n hot alcohol;
and very soluble 1n ether, carbon dlsulflde, acetic add and petroleum ether
(NLM, 1981c; Yalkowsky and Valvani, 1980).
1,2,3,5-Tetrachlorobenzene Is a solid that appears 1n the form of
needles or white flakes. It 1s very slightly soluble 1n water (3.5 mg/a
at 25°C), slightly soluble 1n alcohol, and very soluble In carbon dlsulflde
and petroleum ether (NLM, 1981d; Yalkowsky and Valvanl, 1980).
1,2,4,5-Tetrachlorobenzene appears as white flakes or needles. It takes
the form of monocllnlc prisms from ether, alcohol or benzene. It 1s
practically Insoluble 1n water (0.6 mg/a at 25°C), slightly soluble 1n hot
alcohol, and soluble 1n ether, chloroform and carbon dlsulflde (NLM, 1982b;
Yalkowsky and Valvanl, 1980). A commercial 1,2,4,5-tetrachlorobenzene was
analyzed as 97.0% pure; Impurities were not Identified (Kao and Potten-
berger, 1979; Dow Chemical Company, 1977).
Pentachlorobenzene Is a needle-like solid (NLM, 1979b). It 1s slightly
soluble 1n water (0.56 mg/a at 25°C); slightly soluble 1n ether, benzene
and chloroform; and soluble 1n hot alcohol and carbon dlsulflde (NLM, 1979b;
Yalkowsky and Valvanl, 1980).
3-13
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Hexachlorobenzene 1s a colorless crystalline (monocl1n1c prisms) solid.
Its water solubility was reported as 0.005 mg/9, at 25°C (Yalkowsky and
Valvanl, 1980). Hexachlorobenzene 1s sparingly soluble 1n cold alcohol and
soluble 1n benzene, chloroform and ether (NLM, 1979a). Impure commercial
preparations may contain pentachlorobenzene (10-81,000 ppm), octachlorodl-
benzo-p-d1ox1n (0.05-212 ppm) and octachlorodlbenzofuran (0.35-58.3 ppm)
(Vllleneuve et al., 1974).
According to the CRC Atlas of Spectral Data and Physical Constants for
Organic Compounds (GrasselH, 1973) the following group trends 1n spectro-
scoplc properties can be seen:
There 1s a red shift 1n ultraviolet x^x for the aromatic
•a to ir* transition with Increasing chlorlnatlon (245 to 272 nm
for monochlorobenzene; 291 to 301 nm for hexachlorobenzene). This
Implies that the more chlorinated the chlorinated benzene, the more
likely Is photodegradatlon at sea level by sunlight. Diagnostic
Infrared absorptions for all the chlorinated benzenes occur around
3.2-3.3 and 6.2-6.4 vm. All the proton NMR aromatic signals for
chlorinated benzenes 1n carbon tetrachlorlde or deuterated chloro-
form solvents occur between 6.9 to 7.5 ppm with respect to tetra-
methylsllane. The mass spectra for all compounds are characterized
by very Intense molecular Ions (H) (100% for all compounds except
for pentachlorobenzene), and Intense M-35 peaks. Thus, specific
1on monitoring using the molecular Ions and M-35 peaks 1s possible,
Increasing the sensitivity of analysis.
The atmospheric chemistry of chlorobenzenes has been studied under
laboratory conditions. D1ll1ng et al. (1976) studied the photocatalzyed
degradation of monochlorobenzene 1n an atmosphere containing 5 ppm nitric
oxide and reported Its half-life to be 8.7 hours under strong light at room
temperature. Kanno and Nojima (1979) Irradiated monochlorobenzene with
light from a xenon lamp 1n the presence of nitric oxide and air and found
the products to be chlorinated nltrobenzenes and nltrophenols. Uyeta et al.
(1976) found that Irradiation of several chlorobenzenes with natural sun-
light for periods up to 56 days yielded polychlorlnated blphenyls. Whether
PCBs are formed under atmospheric conditions 1s unknown, but 1t 1s con-
3-14
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sldered unlikely because of the low concentration, Yanaglharl et al, (1977)
studied the degradation of monochlorobenzene In a smog chamber (2 ppm chlo-
robenzene, 1 ppm NO ) and found 7.5% degradation 1n 5 hours. Using higher
A
concentrations (5000 ppm chlorobenzene and 1000 ppm NO), Kanno and Nojlma
(1979) found similar rates of degradation and Identified one chloronltroben-
zene and three chloronltrophenols as products. Rates of reaction of chlo-
robenzene with hydroxyl radical (Anbar and Neta, 1967) and singlet oxygen
(Graedel, 1978) are also available which allows half-life estimations of 0.5
and 2.6 years, respectively. Yanohlhara et al. (1977) also studied the
atmospheric photochemistry of o~d1chlorobenzene and found 21.5% degradation
1n 5 hours and Nojlma and Kanno (1980) found 7.6% of p_-d1chlorobenzene (high
concentrations of test chemical and NO) In 5 hours of Irradiation.
One study has examined the possibility of photocatalyzed degradation of
the chlorinated benzenes. Oliver et al. (1979) exposed 1.4 dlchlorobenzene
1n saturated aqueous solutions with various suspended sediments (titanium
oxide, clays and samples of river sediments) to ultraviolet light. Degrada-
tion of the dlchlorobenzene occurred only 1n the titanium oxide solution,
possibly because of a shielding of the chemical from the light by the other
sediments or as a consequence of a catalytic effect of titanium oxide, and
did not occur 1n natural sediment systems. Korte et al. (1978) and Hustert
et al. (1981) demonstrated that hexachlorobenzene was photochemlcally
stable. The Hustert et al. (1981) study consisted of sunlight Irradiation
of an aqueous solution.
3.3. ANALYTICAL METHODOLOGY
The usual sampling and analytical methods for airborne chlorobenzenes
Involve the adsorption and concentration of airborne vapors on sorbentpacked
cartridges followed by thermal desorptlon and gas chromatographlc (GC) anal-
ysis using either flame 1on1zat1on detection, electron capture detection, or
3-15
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photo1on1zat1on detection. The purge and trap method 1s the most common
method used for the sampling of volatile chlorobenzenes 1n water. Headspace
analysis using GC with flame 1on1zat1on detection or electrolytic conductiv-
ity detection are also used for analysis of aqueous samples. Methods that
are slightly modified from the analytical procedures for aquatic samples are
used for the analysis of chlorobenzenes 1n soil, food and human tissues.
The following sections provide examples of these analytical methods.
3.3.1. Chemical Analysis 1n A1r. Krost et al. (1982) described a method
whereby ambient air was drawn through a cartridge containing a 1.5x6.0 cm
bed of Tenax-SC (35/60 mesh) so that vapors were collected completely on the
resin. The sample was then thermally desorbed and the vapors passed through
a cryogenlcally cooled trap and subsequently Introduced Into a gas chromato-
graph-mass spectrometer (GC-MS). Estimated detection limits for three
chlorobenzenes were as follows: monochlorobenzene, 2.1 ng/m3; 1,2-dl-
chlorobenzene, 1.0 ng/m3; and 1,3-d1chlorobenzene, 0.7 ng/m3. However,
the precision of this method for field sampling and analysis has been deter-
mined to range from £10 to ±40% relative standard deviation. A similar
method has been used for the monitoring of mono- and dlchlorobenzenes by
Barkley et al. (1980), Pell1zzar1 (1982) and Bozzelli (1981).
Lewis and MacLeod (1982) have developed and evaluated a portable low-
volume air sampling system for Indoor air monitoring of semlvolatlle organic
chemicals. Two types of sampling cartridges were used to sample for chloro-
benzenes. Using polyurethane foam (PUF), a mean collection efficiency of
94.0% with a relative standard deviation of 12% was determined for five
1 vg samples of pentachlorobenzene. For five 0.5 and 1.0 pg samples of
hexachlorobenzene, the reported mean collection efficiency was 94.5% with a
relative standard deviation of 8%. The tr1- and tetrachlorobenzenes were
3-16
-------
poorly trapped using this PUF plug, with collection efficiencies of 6.6 and
62.3%, respectively. However, using a dual-sorbent trap consisting of a
0.6 g layer of Tenax-GC (35-60 mesh) sandwiched between two 3.8 cm PUF
plugs, a collection efficiency of 98% was obtained for both compounds.
Theoretical detection limits, using GC-electron capture detection, are
expected to be at least one order of magnitude lower (1n the range of
0.06-0.1 yg/m3). Storage stability of the PUF cartridges was tested
under adverse storage conditions. The amount of chlorobenzenes recovered
from the cartridges after 15 days of storage at 32°C ranged from 57% for the
trlchlorobenzenes to 98% for hexachlorobenzene. Billings and Bldleman
(1980) reported that hexachlorobenzene was very poorly retained by porous
PUF, but efficiently collected by Tenax-GC. Oehme and Stray (1982), how-
ever, reported high recovery (76-115%) of tr1-, tetra-, penta- and hexa-
chlorobenzene with PUF plugs.
Langhorst and NestMck (1979) used an air sampling tube packed with two
sections of Amberllte XAO-2 resin separated by a sllanlzed glass wool plug
to collect the chlorobenzenes. The adsorbent was desorbed with carbon
tetrachlorlde and analyzed by GC using a photo1on1zat1on detector. Using
the method described, the minimum detection limits for mono-, d1-, tr1-,
tetra-, penta- and hexachlorobenzene were 15, 20, 30, 35, 45 and 70 ppb
(v/v), respectively. Collection and desorptlon efficiencies for all chloro-
benzenes (air concentrations between 5 ppb and 15 ppm) were -95% with a
precision of ^12%.
Thompson et al. (1980) described a sampling technique using the Trace
Atmospheric Gas Analyzer coupled with negative atmospheric pressure chemical
1on1zat1on for analysis of hexachlorobenzene after gas chromatography. This
system 1s portable with close to real-time capability and a detection limit
of -30 ppt (v/v) 1n air.
3-17
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Van Tassel et al. (1980) pointed out that there are disadvantages 1n
using some sorbent materials because of Interferences, such as relative
humidity and high concentrations of carbon dioxide. They described a method
for determining m1crogram-per-cub1c meter levels of monochlorobenzene 1n air
using sampling cartridges containing Porapak N, followed by elutlon with
methanol and analysis by GC. Both electron capture detection and photo-
1on1zat1on detection can be used with detection limits of 1 pg/m3 and 5
yg/m3, respectively. This technique reportedly allows for greater
flexibility. Results are reproducible at various relative humidity levels
and varying concentrations of carbon dioxide. Advantages over thermal
desorptlon techniques Include ease of operation and the ability to use
various columns to achieve analytical precision.
NIOSH (1977) has developed sampling and analytical methods for occupa-
tional air monitoring for monochlorobenzene, 1,2-d1chlorobenzene and 1,4-
dlchlorobenzene. All three methods Involve trapping the compound 1n a char-
coal tube, desorblng the analyte with carbon dlsulflde, and analyzing the
sample 1n a gas chromatograph using flame 1on1zat1on detection.
3.3.2. Chemical Analysis 1n Water. The purge-trap method 1s the most
commonly used method for analyzing volatile organlcs 1n water. Otson and
Williams (1982) evaluated the use of the dynamic headspace or the purge-trap
method 1n combination with GC technique for a number of organlcs Including
monochlorobenzene and dlchlorobenzene. For monochlorobenzene, a recovery
rate of 91% was measured using flame 1on1zat1on detection and 96X was mea-
sured using electrolytic conductivity detection. The corresponding recovery
rates for 1,4-d1chlorobenzene were 65 and 58%. Detection limits ranged from
<0.1 yg/l for monochlorobenzene to 0.2 pg/8, for 1,2-d1chlorobenzene.
3-18
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The relative standard deviation ranged from 6.3-9.6% using flame 1on1zat1on
detection. Storage of samples for several weeks did not affect results by
more than +15%, The dynamic headspace or the purge-trap method has been
used by other researchers to determine the levels of mono- and dlchloroben-
zenes 1n water samples (Perelra and Hughes, 1980; Jungclaus et al., 1978).
The purge-trap technique 1s also recommended by U.S. EPA Method 602 (1982)
for the determination of mono- and dlchlorobenzenes 1n wastewater. Cowen
and Baynes (1980) concluded that headspace analysis was applicable for
monochlorobenzene and dlchlorobenzenes, using flame 1on1zat1on detection at
the 5 vg/l concentration 1n water. Minimum detectable quantities using
electrolytic conductivity detection were 0.15 and 0.20 ng for monochloro-
benzene and dlchlorobenzenes, respectively. The static headspace technique
was employed by Otson et al. (1982) for the quantification of mono- and
dlchlorobenzene 1n Canadian potable waters.
The purge-trap technique does not provide quantitative recoveries for
compounds with low volatilities, such as trlchlorobenzenes and higher
chlorinated benzenes. Therefore, a solvent extraction and cleanup method 1s
normally used to produce organic extracts suitable for GC/MS analysis. The
U.S. EPA (1982) (Method 612) has recommended the use of Flor1s1l column
chromatography as a cleanup step before the quantification of the samples by
GC with electron capture detector. This recommended method 1s applicable
for the determination of d1-, trl-, tetra-, penta- and hexa-chlorobenzene 1n
drinking water and wastewater. The recovery of dlchlorobenzenes and hexa-
chlorobenzene by this method was found to be 82-89% and 95%, respectively.
The percent standard deviation of the method for dlchlorobenzenes and
hexachlorobenzene ranged from 10-20% (U.S. EPA, 1982).
3-19
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3.3.3, Chemical Analysis 1n Soil, Sediment and Chemical Waste Disposal
Site Samples. A method for the determination of hexachlorobenzene 1n son
and chemical waste disposal site samples has been developed by DeLeon et al.
(1980). The procedure Involves methane extraction followed by temperature-
programmed GC analysis using electron capture detection. Recoveries of
samples spiked at the 10, 100 and 300 pg levels were 96.5% (±3,6), 93.1%
(±8.1) and 78.OX (±2.6), respectively. The lower detection limit for this
method 1s around 10 pg/g. The solvent extraction method was used by
Lopez-Avlla (1983) to determine chlorobenzenes 1n sediment samples. In this
method, the solvent extract was subjected to acid-base fractlonatlon. The
base/neutral fraction containing the chlorobenzenes was fractionated by
silica gel chromatography. The final separation and quantification was
accomplished by GC-MS. The recovery of 1,3-d1chlorobenzene, 1,2-dlchloro-
benzene, 1,2,4-tMchlorobenzene and hexachlorobenzene by this method was 63,
66, 67 and 46%, respectively, at a spike level of 400 ng/g of dry sediment.
3.3.4. Chemical Analysis 1n F1sh and Other Foods.
3.3.4.1. FISH — The levels of pentachlorobenzene and hexachloroben-
zene have been determined In fish samples using solvent extraction, solvent
and sulfurlc acid partitioning and GC with electron capture detection (Lunde
and Ofstad, 1976).
H1att (1981) compared three analytical methods used to quantify mono-
chlorobenzene levels 1n fish tissue. His data Indicate that recoveries of
64±15%, 32±8X, and 68% were reported for the vacuum extraction method,
direct purge and trap method, and a modified purge and trap method, respec-
tively. In the modified purge and trap procedure, the homogenized fish
tissue was purged, with the concentration trap Immersed 1n an Ice water bath
for 5 minutes, followed by Immersion In a 55°C water bath for an additional
3-20
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7 minutes. This modification provided reproducible purging efficiencies. A
similar modified purge-trap method has been used by Easley et al. (1981) for
the determination of monochlorobenzene 1n fish samples.
The determination of trlchlorobenzenes and other higher chlorinated ben-
zenes in fish samples can also be accomplished by a solvent extraction
method. In one method, Kuehl et al. (1980) subjected the solvent extract to
Florlsll and gel permeation on chromatographlc separation, followed by GC-MS
Identification and quantification of tMchlorobenzene and other higher
chlorinated benzenes 1n fish samples. Murray et al. (1980), however, used
solvent partitioning, silica gel chromatography, followed by SC-electron
capture detection for the quantification of hexachlorobenzene 1n fish
samples.
3.3.4.2. HUMAN MILK — A method to detect ppb concentrations of hexa-
chlorobenzene 1n human milk has been used by Brevlck (1978). This method
involves solvent extraction and GC analysis using electron capture detec-
tion. A mean recovery of 98.6*10.8% was reported for 10 samples containing
5 ppb hexachlorobenzene. The solvent extraction method was also used by
Tessari and Savage (1980) for the determination of hexachlorobenzene In
human milk. In this method, the extract was subjected to Flor1s1l and
silica gel column chromatographlc cleanup, followed by GC-electron capture
detection. The method gave 68% recovery at a fortification level of
5.7 ng/g.
The quantification of more volatile halogenated benzenes, such as mono
and dlchlorobenzenes 1n milk samples, was performed by a purge and trap
technique at an elevated temperature of 50°C. The trapped gas was thermally
desorbed and quantified by the GC-MS method (Michael et al., 1980). The
average recovery of monochlorobenzene by this method was determined to be
88%.
3-21
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3.3.4.3. OTHER FOODS -- Rice, vegetables, meat, milk, eggs and fish
have been analyzed for hexachlorobenzene residues using GC with electron
capture detection (Seklta et al., 1980); 6C-HS was used to confirm the anal-
ysis. A similar solvent extraction method, followed by solvent partitioning
and Flor1s1l column cleanup, and GC-electron capture detection was used for
the quantification of hexachlorobenzene In different crops from 37 states
(Carey et al., 1979).
3.3.4.4. OTHER BIOL06ICAL MATRICES — Gas chromatography using elec-
tron capture detection has been employed to determine levels of pentachloro-
benzene and hexachlorobenzene 1n blood samples (Lunde and Bjorseth, 1977)
and to determine levels of 1,4-d1chlorobenzene and Us major metabolites 1n
urine and serum samples (HcKlnney et al., 1970). Blood and urine samples
have also been analyzed for the chlorobenzenes by GC using photo1on1zat1on
detection (Langhorst and Nestrlck, 1979). Using carbon tetrachlorlde
extraction, silica gel column chromatography and concentration with a
Kuderna-Oanlsh concentrator, chlorobenzene recoveries from blood and urine
samples averaged 83£12% for concentrations between 1 and 500 ppb.
A method of hexachlorobenzene determination and confirmation 1n adipose
tissue has been described by Watts et al. (1980). In this method, the
solvent extract 1s subjected to a Florlsll cleanup and one-fraction elutlon.
Hexachlorobenzene 1s determined by direct GC with electron capture detec-
tion. Confirmation Is made by analysis of the b1s-1sopropoxytetrachloroben-
zene derivative, which 1s formed by reaction with Isopropanol. Average
recoveries ranged between 87.4+6.854 and 92.6ilO.054. This method 1s particu-
larly useful for the determination of hexachlorobenzene 1n the presence of
M1rex.
3-22
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The determination of the less volatile chlorinated benzenes, such as
trl-. tetra-. penta- and hexachlorobenzene 1n biological tissue samples, has
been done by solvent extraction of the tissue, followed by column chromato-
graphlc cleanup of the sample and final separation and quantification by GC
with electron capture detection (Lamparskl et al., 1980; Mes et al., 1982).
For more volatile chlorinated benzenes, such as mono- and dlchlorobenzenes,
the modified purge-trap method 1n combination with capillary GC and flame
1on1zat1on detection or preferably more specific detection method can be
used (Michael et al., 1980).
3.4. SUMMARY
The chlorinated benzenes are a group of compounds 1n which 1-6 chlorine
substltuents have been added to a benzene ring yielding a total of 12 1so-
meMc forms. In general, these compounds have low water solubility (solu-
bility decreasing with Increasing chlorlnatlon), low flammabHlty, moderate
to high octanol/water partition coefficients (coefficients Increasing with
Increasing chlorlnatlon) and low to moderate vapor pressures (vapor pres-
sures decreasing with Increasing chlorlnatlon). They are chemically unreac-
tlve and exist as liquids or solids at environmental conditions. Analysis
of airborne chlorobenzenes 1s usually accomplished by adsorption onto
sorbent cartridges, followed by thermal desorptlon and analysis by GC. For
water samples, the purge-trap method Is used to concentrate the volatile
halogenated benzenes before analysis by GC. For less volatile chlorinated
benzenes, solvent extraction followed by column chromatographlc cleanup of
the extract and GC with electron capture detection, 1s the most commonly
used method for the Isolation, detection and quantification. Methods
similar to those used for wastewater samples are commonly used for the
analysis of chlorinated benzenes In biological matrices.
3-23
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4. PRODUCTION, USE AND ENVIRONMENTAL LEVELS
4.1. PRODUCTION
Industrial synthesis of chlorinated benzenes Is achieved through the
controlled catalytic chlorlnatlon of benzene and 1s described In the K1rk-
Othmer Encyclopedia of Chemical Technology (Hardle, 1964), In general,
monochlorobenzene and the dlchlorobenzenes are synthesized at 30-50°C 1n the
presence of a ferric chloride catalyst. The output product 1s then purified
by distillation, and the Isomers are Isolated by fractional distillation or
crystallization. Trlchlorobenzene 1s produced by the chlorlnatlon of
dlchlorobenzene using ferric chloride and temperatures of 25-30°C, An
aluminum catalyst, trlchlorobenzene and chlorine are used to produce tetra-
chlorobenzene, which 1n turn can serve as a precursor for pentachloroben-
zene. Hexachlorobenzene can be obtained by the chlorlnation of benzene at
150-200°C using a ferric chloride catalyst or from the distillation of resi-
dues from the production of tetrachloroethylene. Because these reactions
are not completely controlled and purification processes are not 100% effec-
tive, 1t 1s Hkely that any commercially available chlorinated benzene will
also contain unwanted 1somer1c chlorobenzenes as Impurities, and this 1s
particularly true for I,2~d1chlorobenzene.
The TSCA Inventory (U.S. EPA, 1981) provides production data on the
chlorinated benzenes for Individual facilities. The data for the largest
producers (>lx!0* pounds/year) are expressed as ranges of estimated pro-
duction for 1977 1n Table 4-1. Total yearly production data are published
for high-volume, synthetic chemical Intermediates by the U.S. International
Trade Commission (USITC); data for 1980 are available only for monochloro-
benzene, 1,2-d1chlorobenzene and 1,4-d1chlorobenzene (USITC, 1981), and are
Incorporated Into Table 4-1. A 1983 11st of producers and the estimates of
4-1
-------
TABLE 4-1
United States Production of Chlorinated Benzenes for Selected Years
Chemical/
Manufacturers
Production
Location Estimates
for 1977a
(Ib x 10«)
Production
for 1980°
(Ib x 10«)
Honochlorobenzene:
Dow Chemical Co.
PPG Industries, Inc.
Hontrose Chemical
Corp. of CA
Allied Chemical Corp.
Monsanto Co.
NAd
NAd
1»2-D1chlorobenzene:
Dow Chemical Co.
PPG Industries, Inc.
Monsanto Co.
Montrose Chemical
Corp. of CA
Allied Chemical Corp.
1,3-D1chlorobenzene:
PPG Industries, Inc.
NAd
Midland, MI 50-100
New Mart1nsv1lle, WV 10-50
Henderson, NV 10-50
Solvay, NY 1-10
Sauget, IL 50-100C
NAd 10-50
NAd 1-10
Total: 132-370
Midland, MI 1-10
New Mart1nsv1lle, WV 10-50
Sauget, IL 1-10
Henderson, NV 1-10
Solvay, NY 1-10
Total: 14-90
New Mart1nsv1lle, WV 0.1-1
NAd 0.1-1
Total: 0.2-2
284
49
NA
4-2
-------
TABLE 4-1 (cent.)
Chemical/
Manufacturers
1 ,4-D1chlorobenzene:
Dow Chemical Co.
PPG Industries, Inc.
Monsanto Co.
Montrose Chemical
Corp. of CA
Allied Chemical Corp.
Dover Chemical Corp.
NAd
1 ,2»3-Tr1chlorobenzene;
Dow Chemical Co.
1 ,2,4-Tr1chlorobenzene:
Dow Chemical Co.
NAd
L^S-Trlchlorobenzene:
Chemical Systems Division
1 ,2,3,4-Tetrachlorobenzene:
Dow Chemical Co.
NAd
Production
Location Estimates Production
for 1977a for 1980b
(Ib x ID6) (Ib x ID6)
Midland, MI
New Mart1nsv1lle, WV
Sauget, IL
Henderson, NW
Solvay, NY
Dover, OH
NAd
Total:
Midland, MI
Total:
Midland, MI
NAd
Total:
San Jose, CA
Total:
Midland, MI
NAd
Total:
1-10
10-50
1-10
1-10
1-10
1-10
1-10
16-110
1-10
1-10
1-10
10-50
11-60
0.01-0.1
0.01-0.1
1-10
0.1-10
1.1-20
75
NA
NA
NA
NA
4-3
-------
TABLE 4-1 (cent.)
Chemical/
Manufacturers
1 ,2.3.5-Tetrachlorobenzene:
(No manufacturers listed)
1 ,2,4,5-Tetrachlorobenzene:
Dow Chemical Co.
Chem South Corp.
NAd
Pentachlorobenzene:
OUn Corp.
Hexachlorobenzene:
{No manufacturers listed)
Production
Location Estimates Production
for 1977a for 1980b
{lb x 10«) {lb x 106)
Midland, MI 10-50
Chlldersburg, AL 0.1-lc
NAd 1-1 Oc
Total: 11.1-61 NA
Mclntosh, AL 1-10
Total: 1-10 NA
NA
aSource: U.S. EPA, 1981
bSource: U.S. ITC, 1981
CA11 production at this site was processed within the facility and was not
distributed outside the facility as a chemical or in a mixture.
^Producer and location not listed in the TSCA inventory.
NA = Not available
4-4
-------
their production capacities for chlorobenzenes are available from SRI
(1983), who 11st -the producers of chlorobenzenes and their estimated produc-
tion capacities as of January 1983 (Table 4-2). The names of the chloroben-
zene manufacturers given 1n Table 4-2 are slightly different from those
given in Table 4-1, because Table 4-2 11st only the manufacturers as of
January, 1983. More recent Information Indicates that the Dow Chemical
Corporation no longer produces any chlorinated benzenes, that Standard
Chlorine Chemical now produces trlchlorobenzenes (mixed Isomers), and that
pentachlorobenzene is no longer produced In the U.S. by any manufactures
(Chlorobenzenes Producers Association, 1984).
As mentioned already, hexachlorobenzene 1s not manufactured commercially
in the United States but does occur 1n waste streams during the production
of some organic chemicals (e.g., perchloroethylene, trlchloroethylene,
carbon tetrachloride and chlorine) and pesticides.
4.2. USE
Chlorinated benzenes are used 1n the manufacture of Intermediates in the
production of organic chemicals, Including other chlorinated benzenes,
herbicides, pesticides, dyes and rubber chemicals and as dye carriers,
process solvents, pesticides, fungicides and deodorizing agents (U.S. EPA,
1980). A summary of these uses 1s presented 1n Table 4-3.
4,3. SOURCE AND ENVIRONMENTAL LEVELS
No comprehensive studies have been conducted on the sources of chloro-
benzenes released into the environment. In general, these releases would
occur during the manufacture and transport of chlorobenzenes, through their
use as pesticides, solvents and other Industrial and consumer products, and
through the disposal of wastes from the manufacturing process. Estimates of
releases (Table 4-4) have been made for monochlorobenzene, dlchlorobenzenes
and trlchlorobenzenes. Dow (1978) estimated that 30-50% of the monochloro-
4-5
-------
TABLE 4-2
U.S. Producers and Estimated Annual Production
Capacities (1983) of Chlorobenzenes*
Chemical/
Manufacturer
Location
Annual Capacity
(Ib. x 10*)
Honochlorobenzene:
Dow Chemical Co.
Honsanto Co.
PPG Industries, Inc.
Standard Chlorine Chem.
1.2-D1chlorobenzene:
Dow Chemical Co.
Honsanto Co.
PPG Industries, Inc.
Standard Chlorine Chem.
1,3-Dlchlorobenzene:
NA
1,4-Dlchlorobenzene:
Dow Chemical Co.
Honsanto Co.
PPG Industries, Inc.
Standard Chlorine Chem.
1.2.3-Tr1chi or obenzene:
Standard Chlorine Chem.
Hldland, MI
Sauget, IL
Natrium, WV
Delaware City, DE
Total:
Midland, MI
Sauget, IL
Natrium, WV
Delaware City, DE
Total:
NA
Midland, MI
Sauget, IL
Natrium, WV
Delaware City, DE
Total:
Delaware City, DE
170
150
45
150
515
30
6
20
50
106
NA
30
12
30
75
147
NA
4-6
-------
TABLE 4-2 (cont.)
Chemical/
Manufacturer
Location
Annual Capacity
(lb. x 10«)
1,2,4-Tr1ch1orobenzene:
Dow Chemical Co,
Standard Chlorine Chem.
1,3.S-Tr1chlorobenzene:
Southland Corp.
Trlchlorobenzene. MixedIsomers;
PPG Industries, Inc.
1,2,3.4-Tetrachlorobenzene:
NA
1,2,3,S-Tetrachlorobenzene;
NA
1f2,4,5-Tetrachlor obenzene:
Dow Chemical Co.
Standard Chlorine Chem.
Pentachlorobenzene:
NA
Midland, MI
Delaware City, DE
Great Meadows, NJ
Natrium, WV
NA
NA
Midland, MI
Delaware CHy, DE
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
*Source: SRI, 1983
NA = Not available
4-7
-------
TABLE 4-3
A Sucmary of the Uses of the Chlorinated Benzenes
Chemical
Major Uses
Reference
4*
I
CO
Honochlorobenztnt
1,2-Dlchlorobenzene
l»3-D1chlorobenzene
1,4-D1ehlorobenzene
1,2,3-Trlchlorobenzene
1,2,4-TMchlorobenzene
Intermediate 1n the manufacture of chloronltrobenzenes, dlphenyl oxide, U.S. EPA, 1980
DOT and slllcones; as a process solvent for methylene dllsocyanate,
adheslves, polishes, waxes, Pharmaceuticals and natural rubber; as
a degrading solvent.
In the manufacture of 3,4-d1chloroan1Hne; as a solvent for a wide range Hawley, 1977
of organic materials and for oxides of non-ferrous metals; as a solvent
carrier In production of toluene dllsocyanate; In the manufacture of dyes;
as a fumlgant and Insecticide; In degreaslng hides and wool; 1n metal
polishes; In Industrial odor control; In cleaners for drains.
As a fumlgant and Insecticide Hawley, 1977
As a moth repellent, general Insecticide, germicide, space deodorant; Hawley, 1977
In the manufacture of 2,S-d1chloroan1l1ne and dyes; as an Inter-
mediate; 1n Pharmaceuticals; 1n agricultural fumlgants.
Other than chemical Intermediate usage, the uses of this compound U.S. EPA, 1980
are the same as 1,2,4-tMchlorobenzene.
As an Intermediate In the manufacture of herbicides; as a dye carrier, U.S. EPA, 1980
as a dielectric fluid; as a solvent; as a heat-transfer medium.
1,3,5-Trlchlorobenzene
1,2,3,4-Tetrachlorobenzene
1,2,3,5-Tetrachlorobenzene
1,2,4,5-Tetrachlorobenzene
Pentachlorobenzene
Hexachlorobenzene
Solvent for high-temperature melting products; as a coolant In
electrical Insulators; as a heat-transfer medium, lubricant and
synthetic transformer oil; as a termite preparation and Insecticide;
1n dyes.
As a component In dielectric fluids; 1n the synthesis of fungicide.
NA
As an Intermediate for herbicides and defoliants; as an Insecticide;
moisture-resistant Impregnant; 1n electric Insulation; In packing protection.
In a pesticide used to combat oyster drills; as a chemical Intermediate.
As a fungicide; Industrial waste product In the manufacture of perchloro-
ethylene, chlorinated solvents, pesticides and nltroso rubber.
SUmak et al., I960
Hawley, 1977
U.S. EPA, 1980
Hawley, 1977
Clement Associates, 1979
Ware and West, 1977
Courtney, 1979
NA = Not available
-------
TABLE 4-4
Estimated Quantities of Chlorobenzenes Lost During Manufacture,
and to the Environment Compared with Total Production In 1983*
(All figures are 1n megagram)
Chlorobenzene
Quantity Lost During
Manufacture
Quantity Lost to
Environment
Total Industrial
Production
Hono-
01-
1,2-
1,3-
1,4-
Trl-
1,2,3-
1,2,4-
1,3,5-
Tetra-
Penta-
Hexa-
191-303
118-206
0.185-0.608
178-284
0.598-2.111
3.39-10.2
Import
NA
not manufactured
not manufactured
153-259
29.95
NA
166-269
<0.100
0.364-0.924
Import
NA
NA
NA
88,769-128,755
18,301-21,479
147-460
28,310
23.3-74.1
1,253-3,668
111-210
NA
NA
NA
*Source: 47 FR 26992
NA = Not available
Note: The Quantity Lost During Manufacture Includes the estimated Quantity
Lost to Environment.
4-9
-------
benzene produced annually was released Into the air. Virtually all the
monochlorobenzene used as a solvent In herbicide formulations Is probably
released Into the atmosphere. Approximately 0.1% of monochlorobenzene
produced annually was estimated to enter water (Dow, 1978). The U.S. EPA
(Gruber, 1975) estimated that, assuming all of the monochlorobenzene was
produced using a batch process, 95% of this
total. These four products and the estimated quantities of hexachloroben-
zene produced are listed 1n Table 4-5. Hexachlorobenzene Is also a consti-
tuent of a seed treatment called Grannox NM that 1s Imported and used In the
United States. In 1975, -200,000 pounds of Grannox NM, a formulation of
hexachlorobenzene also containing the pesticide Haneb, entered the United
States (IARC, 1979). Table 4-4 shows the most recent official figures for
quantities lost from Industrial sources, and quantities released Into the
environment compared with total Industrial production (47 FR 26992).
4-10
-------
TABLE 4-5
Estimated Quantities of Hexachlorobenzene {HCB)
1n Industrial Wastes and Byproducts 1n 1972*
Product
Perchloroethylene
Trlchloroethylene
Carbon tetrachlorlde
Chlorine
Total HCB (ID3 Ibs)
1750-3500
230-450
200-400
160-390
HCB (Ibs/ton of
4.8-9.5
1.1-2.1
0.4-0.8
0.02-0.04
product)
*Source: Mumma and Lawless, 1975
4-11
-------
4.3.1. Levels 1n A1r. Investigations of the occurrence of chlorobenzenes
In air have been conducted 1n Japan and the United States utilizing both
grab and sorbent cartridge techniques. These studies, which have Included
the sampling of polluted air and urban and rural air, have reported the
detection of monochlorobenzene, and various Isomers of d1-, tr1-, tetra- and
hexa-chlorobenzenes. Analysis of Indoor air has Indicated the presence of
monochlorobenzene and the dlchlorobenzenes; other studies have measured
monochloro- and 1,4-d1chlorobenzene 1n occupational settings.
Horlta and Oh1 (1975) sampled ambient air for the determination of
l,4-d1chlorobenzene levels at six central and suburban Tokyo sites 1n Japan
and found concentrations ranging from 1.5-4.2 pg/m3. Determinations
were also performed on "Indoor" samples from a closet, a bedroom and a
wardrobe; these concentrations were -25-400 times greater than the highest
reported ambient levels.
PelUzzaM et al. (1979) presented the results of the analysis of air
samples collected from a number of outdoor locations 1n the United States.
Samples from each location were obtained from several sites at a given loca-
tion and at numerous times. The samples were analyzed for monochlorobenzene
and the d1- and trlchlorobenzenes. Table 4-6 1s a compilation of these data.
Monochlorobenzene and the dlchlorobenzenes were also measured by
WoJ1nsk1 et al. (1979) 1n samples from an Industrially produced cloud that
periodically appears over Henderson, Nevada, an Industrialized town 10 miles
southeast of Las Vegas. Monochlorobenzene levels were nearly 50 times
greater 1n the Henderson samples (mean: 24325 ng/m3) as compared with the
Las Vegas samples (mean: 458 ng/m3). The mean 1,2-d1chlorobenzene concen-
trations were ~5 times greater over Henderson than over Las Vegas (10291
ng/m3 compared with 3087 ng/m3). However since the methodology was
different, the significance of this finding 1s uncertain.
4-12
-------
TABLE 4-6
Chlorinated Benzene Levels 1n Ambient A1r from Different Locations 1n the U.S.3
Concentration range, ng/m3
SHe
K1n-Buc Disposal
Site, Edison, NJ
Baton Rouge, LA
Houston, TX
Niagara Falls, NY
Different NO S1tesc
Date
Sampled
1976/
1978
1977
1977
NR
1978
HCB
ND-12,791
ND-900
ND-132
ND-119
ND-6072
1,2-DCB
ND-12,433
ND-87
ND-86
ND-444b
ND-5513
1,3-DCB
ND-33,783
ND-102
ND-86
ND-444&
ND-3392
1,4-DCB
ND-7000
ND
ND
ND-444b
ND
KB
ND-1327
ND
ND
ND-4346
ND
TeCB
NR
NR
NR
ND-451
NR
PeCB
NR
NR
NR
ND-17
NR
aSource: Taken from PelUzzarl et al., 1979
^These are the values for the imseparated Isomers
cThe sites Include: Edison, Ground Brook, Paterson, Hoboken, Clifton, Fords, Passalc and Sayrevllle
ND = Not detected; NR = not reported
MCB = Monochlorobenzene; DCB = dlchlorobenzene; TCB = trlchlorobenzene; TeCB = tetrachlorobenzene; PeCB =
pentachlorobenzene
-------
In a study of contamination from a hazardous waste site, Barkley et al.
(1980) provided data on chlorobenzene levels from the homes of nine resi-
dents of the "Old Love Canal" area of Niagara Falls, New York. Monochloro-
benzene was detected Inside three of the homes at concentrations ranging
from 60-600 ng/m3. The dlchlorobenzenes (Isomers unseparated) were found
Inside the homes at concentrations ranging from trace levels to 31,000
ng/m3. Samples taken outside the homes over 6-16 hour periods contained
monochlorobenzene, dlchlorobenzenes (Isomers unseparated), trlchlorobenzenes
(Isomers unseparated) and tetrachloro-benzenes (Isomers unseparated) from
nondetectable amounts up to 440 ng/m3, the highest level being found at
one location for the dlchlorobenzene Isomers,
In 1978, the Department of Environmental Protection of New Jersey
Initiated a study of selected volatile organic chemicals 1n ambient air.
Over a 5-month period, a total of 330 samples were obtained at five sites
that Included a mixture of Industrial, residential and semlrural locations.
The results, reported by BozzelH and Kebbekus (1979), Indicated the pres-
ence of 1,2- and l,4-d1chlorobenzene at all sites. The average concentra-
tions (trace amounts were averaged as the lower detection limit of 0.01 ppb)
were 2096 ng/ma and 1703 ng/m3 for 1,2- and 1,4-d1chlorobenzene, respec-
tively. Peak concentrations were 46,780 ng/m3 and 93,560 ng/m3, respec-
tively. In a follow-up study, Harkov et al. (1981) sampled air at six sites
1n New Jersey over 24-hour periods every 6 days for a year. Monochloro-
benzene was detected 1n 86% of the samples at an average level of 2.53
yg/m3 and a maximum level of 1.36 pg/m3.
Field studies were conducted by Singh et al. (1981) 1n California and
Arizona to characterize the atmospheric levels and fate of several organic
chemicals. The samples, collected over 24 hours during a 2-week period at
-------
each site, were analyzed for four chlorobenzenes: monochlorobenzene, l,2-d1-
chlorobenzene, 1,3-d1chlorobenzene and 1,2,4-tr1chlorobenzene. Table 4-7
presents the results of the analysis.
The atmospheric concentrations of the chlorinated benzenes around
different locations In the United States have been measured by a number of
other Investigators. The monitoring sites can be broadly divided into three
different categories, namely rural/remote sites, urban/suburban sites and
sites near source areas. In a recent report, Brodzlnsky and Singh (1982)
Integrated most of the available U.S. air monitoring data relating to the
levels of chlorobenzenes along with a number of other organlcs Into a
coherent data base. The overall and site-specific mean atmospheric levels
of monochlorobenzene, dlchlorobenzenes, trlchlorobenzenes and tetrachloro-
benzenes compiled 1n this report {Brodzlnsky and Singh, 1982) are shown In
Table 4-8.
The levels of pentachlorobenzene and hexachlorobenzene 1n ambient air
samples rarely have been reported. Considering their volatility, the
abundance of these compounds 1n the atmosphere can be speculated to be lower
than the other chlorobenzenes. However, tetrachlorobenzenes, pentachloro-
benzene and hexachlorobenzene have been detected, but not quantified, 1n fly
ashes from municipal Incinerators (Elceman et al., 1979, 1981). High volume
air samples collected from Boston, Massachussetts, and Columbia, South
Carolina, using a glass fiber filter and polyurethane foam trap were
subjected to Interlaboratory analysis of hexachlorobenzene along with other
parameters (Bldleman, 1981). The average concentrations of hexachloroben-
zene In Boston (Massachussetts) and Columbia (South Carolina) air were found
to be >0.057 ng/m3 and 0.012 ng/m3, respectively. The percent relative
standard deviations for the two results were determined to be 35 and 43%,
respectively.
4-15
-------
TABLE 4-7
Concentrations of Chlorinated Benzenes at Three Sites3
Mean Concentration 1n na/m3 — 1 Standard Deviation^
Chemical Los Angeles, CA
Monochlorobenzene -936
1
1
1
,2-D1chlorobenzene 75.1 + 59.5
,3-01chlorobenzene 46.3 + 33.7
,2,4-Tr1chlorobenzene 52,0 + 36.9
Phoenix, AZ Oakland, CA
-936 -468
135.8 + 209.1 24.0 ± 30.1
52.3 i 35.5 39.1 ± 17.4
23.4 + 15.8 22.6 + 18.1
aSource: Singh et a!., 1981
&The conversion of ppt unit to ng/m3 1s based on a temperature of 20°C
and a pressure of 1 atmosphere.
4-16
-------
TABLE 4-8
Overall and Site-Specific Mean Atmospheric Levels
of Chlorobenzenes throughout the United States*
Chemical
Monochlorobenzene
1 ,2-Q1ch1orobenzene
1 ,3-D1chlorobenzene
1 ,4-D1chlorobenzene
Trlchlorobenzenes
Tetrachlorobenzenes
Total Number
of
Localities
56
51
38
24
35
3
Mean
Overall
3087
1142
571
1563
136
3502
Concentration,
Rural -Remote
Area
ND
11
40
NO
ND
198
ng/m3
Urban-
Suburban
Areas
3742
1142
499
1743
128
6196
Areas of
Production
936
1202
902
16
181
853
*Source: Brodzlnsky and Singh, 1982
ND - Not detected
4-17
-------
In a survey of contamination by hexachlorobenzene around eight Individ-
ual plants, L1 et al. (1976) reported the detection* of up to 24 v9/m3
(1.9 ppb) hexachlorobenzene at a distance of 90 feet from one plant.
«>
Table 4-9 summarizes the data from this Investigation. Concentrations of
hexachlorobenzene at distances 400-3000 feet downwind from the plants ranged
from 0.02-2.7 v9/m3. The authors noted that the highest levels of hexa-
chlorobenzene contamination were associated with the production of lower
chlorinated hydrocarbons as opposed to the production of chlorine and herbi-
cides, and that plants with onslte landfill and open pit waste disposal
sites had the highest levels of airborne concentrations of hexachlorobenzene.
Chlorinated benzenes are also present 1n the air of occupational
settings. Ware and West (1977) reported that the air of facilities manufac-
turing 1,4-d1chlorobenzene contained an average of 204 mg/m3 dlchloroben-
zene (from 42-288 mg/m3) for certain processes and that no levels <48
mg/m3 were detected throughout the plant. More recently, a survey of
monochlorobenzene exposure was conducted at the chemical companies by the
National Institute for Occupational Safety and Health (NIOSH) (Cohen et al.,
1981). Personal sampling data Indicated that monochlorobenzene levels
ranged from below the detection level to 18.7 mg/m3.
4.3.2. Water. Chlorinated benzenes have been detected 1n ground, surface
and drinking water and 1n Industrial and municipal wastewater. The most
prevalent compound 1s monochlorobenzene. The dlchlorobenzenes, trlchloro-
benzenes, tetrachlorobenzenes, pentachlorobenzene and hexachlorobenzene are
detected Infrequently.
Numerous investigations have Identified chlorinated benzenes In samples
of surface water (Table 4-10). The U.S. EPA STORET system also Includes
monitoring data on the chemicals.
4-18
-------
TABLE 4-9
Atmospheric Levels of Hexachlorobenzene Around
Selected Industrial Plants*
Company/Location
Products
Hexachlorobenzene
Concentrations,
High
Low
Vulcan Materials
Wichita, KS
Stauffer Chemical
Louisville, KY
Dow Chemical
Pittsburgh, CA
Du Pont
Corpus Chr1st1, TX
Diamond Shamrock
Deer Park, TX
Ol1n
Mclntosh, AL
C1ba-Ge1gy
St. Gabriel, LA
PPG
Lake Charles, LA
Perchloroethylene, carbon
tetrachlorlde, chlorine
Perchloroethylene, carbon
tetrachlorlde, methylene
chloride, chlorine
Carbon tetrachlorlde, perchlo-
roethylene, chlorine
Carbon tetrachloride
Trlchloroethylene, perchloro-
ethylene, chlorine
Pentachloronltrobenzene,
chlorine
Atrazlne, propazlne,
slmazlne
Trlchloroethylene, perchloro-
ethylene, vinyl chloride, vinyl
1dene chloride, chlorine, etc.
24 0.53
7 0.24
0.08 <0.02
ND ND
ND ND
2.2 0.03
0.02 NO
1.7 NO
*Source: L1 et a!., 1976
ND = Not detected {<0.02
4-19
-------
TABLE 4-10
Chlorinated Benzenes 1n Surface Water
Chemicals
Levels3
Location
Reference
•*»
i
O
Tr1chlorobenzeneb
Monochlorobenzene
1,4-D1chlorobenzene
Honochlorobenzene
Dlchlorobenzene
Trlchlorobenzene
Tetrachlorobenzene
Pentachlorobenzene
Hexachlorobenzene
Hexachlorobenzene
Honochlorobenzene
Dlchlorobenzeneb
Tr1chlorobenzeneb
Honochlorobenzene
1»4-D1chlorobenzene
Dlchlorobenzeneb
Honochlorobenzene
Dlchlorobenzene
Trlchlorobenzene
Tetrachlorobenzene^
Pentachlorobenzene
Hexachlorobenzene
100-500
4-
30-900
ND-17.4 (2.7)
ND-7000
ND-400
ND-1000
NO->10,000
ND->10,000
+ 1n 7% of all surface
water and 1n 3% of all
groundwater samples
100-8000
100-200,000
NO-100,000
8000-30,000
MerMmack River, MA H1tes, 1973
Glatt River, Germany G1ger et al., 1976
River Waters, U.S.
Ohio River
600 sites 1n NJ
Drainage streams
Niagara Falls, NY
Shackelford and Keith, 1976
Tiber River, Italy Leonl and D'Arca, 1976
Delaware River Sheldon and HHes, 1978
ORVHSC, 1982
Page, 1981
Elder et al., 1981
-------
TABLE 4-10 (cont.)
Chemicals
Levels3
Location
Reference
I
IV)
D1chlorobenzenec
Tr1chlorobenzenec
Tetrachlorobenzenec
Pentachlorobenzenec
Hexachlorobenzene
01chlorobenzenec
Tr1chlorobenzenec
Tetrachlorobenzenec
Pentachlorobenzene
Hexachlorobenzene
3-71 (27)
0.1-1.6 (0.5)
ND-0.8 (0.12)
NO-0.6 (0.12)
0.02-0.1 (0.05)
NO-77 (11)
NO-8.7 (2.1)
NO-0.2 (0.05)
NO-0.1 (0.05)
0.02-0.1 (0.06)
Great Lakes
Oliver and Nlchol, 1982
Grand River, Canada
Oliver and Nlchol, 1982
aRange (mean) 1n ng/s, unless Indicated
^Unidentified Isomers
CA11 Isoraers
NO = Not detected; f = detected
-------
Drinking water supplies also have been sampled for chlorobenzene contam-
ination. In a survey of three water treatment plants of the city of New
Orleans, Louisiana, U.S. EPA (1975a) reported finding only one chloroben-
zene, 1,3-d1chlorobenzene, at two of the plants. The reported concentration
was <3 v9/8" The U.S. EPA also found monochlorobenzene and all three
Isomers of dlchlorobenzene at levels <1.0 yg/8. 1n three of five raw
water supplies that were sampled as part of the National Organlcs Reconnais-
sance Survey (U.S. EPA, 1975b). A follow-up study (U.S. EPA, 1975c) report-
ed concentrations of monochlorobenzene 1n samples of finished drinking water
from the four Initial locations and at five additional sites.
Con1gl1o et al. (1980) reported data on concentrations of volatile
organic chemicals from drinking water treatment plants 1n the United States.
The frequency of occurrence of 1,2-d1chlorobenzene and 1,2,4-tr1chloroben-
zene 1n finished water originating from surface water was 12.5 and 11.5%,
respectively. Of the facilities using groundwater, 12.9 and 7.1% had drink-
Ing water samples containing 1,4-d1chlorobenzene and monochlorobenzene,
respectively. The other chlorobenzenes were detected 1n <4.5% of the
samples. The authors also reported data from a groundwater survey conducted
1n New Jersey. Of the chlorobenzenes, only the dlchloro- and trlchloroben-
zenes (Isomers unspecified) were detected at concentrations ranging from
<1-100 vg/8. 1n 1 and 3% of the samples, respectively.
Fielding et al. (1981) sampled untreated and finished groundwater, raw
and finished river water and rainwater from 13 unspecified sites throughout
Great Britain. 1,4-D1chlorobenzene was detected at a concentration of 0.08
jjg/fi. In one finished groundwater sample. Page (1981) also surveyed
groundwater and surface water at over 1000 sites throughout New Jersey, 1n a
study designed to compare the relative degree of chemical contamination of
4-22
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ground and surface water. All Isomers of chchlorobenzene were found 1n -3%
of all the groundwater samples and 4% of the surface water samples.
Analysis of these data and data on 52 other toxic chemicals Indicated that
New Jersey groundwater has a similar pattern and degree of contamination as
surface water. Oliver and Nlchol (1982) sampled drinking water 1n three
cities on Lake Ontario, both before and after chlorlnatlon. Individual
Isomers of dlehloro- through hexaehlorobenzene were found In mean concentra-
tions ranging from non-detectable to 13 ng/St. No Increase 1n the level of
concentration was noted 1n these compounds after chlorlnatlon. The levels
of chlorobenzenes In the drinking water of three cities bordering Lake
Ontario are given 1n Table 4-11.
As part of a study of the contamination of drinking water by leachate
from a pesticide waste dump site in Hardeman County, TN, Clark et al. (1980)
reported data from a U.S. EPA survey of chemicals 1n private wells.
Monochlorobenzene was found 1n 23 of 25 wells at levels from trace amounts
to 41 vg/8. with a median value of 5.0 vg/fc. In another study of
possible contamination of drinking water by the disposal of toxic chemicals,
Barkley et al. (1981) found monochlorobenzene and dlchlorobenzenes (Isomers
unspecified) 1n tapwater from all nine of the houses 1n Old Love Canal,
Niagara Falls area. Concentration levels for these two compounds ranged
from 10-60 ng/a and from 10-800 ng/a, respectively. The tetrachloro-
benzenes and pentachlorobenzene also were found at concentrations up to 2000
and 240 ng/J., respectively.
The chlorobenzenes have been Identified In wastewaters from Industrial
processes and In Influents and effluents at municipal sewage treatment
plants. Gaffney (1976) sampled water at four municipal facilities 1n
Georgia that, 1n addition to handling sewage, also treated wastewater from
4-23
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TABLE 4-11
Chlorobenzene Concentrations 1n Drinking Water from Ontario, Canada3
Chemical
1 ,2-D1chlorobenzene
1 ,3-D1chlorobenzene
1 ,4-D1chlorobenzene
1 ,2,3-TMchlorobenzene
1 , 2, 4-Tr1 chlorobenzene
1 ,3,5-TMchlorobenzene
1 ,2,3,4-Tetrachlorobenzene
1 ,2,3,5-Tetrachlorobenzene
1 ,2,4, 5-Tetrachlorobenzene
Pentachlorobenzene
Hexachlorobenzene
Concentration
Range
ND-7
ND-2
8-20
0.1-0.1
1-4
NDb
0.1-0.4
NDb
NDb-0.3
0.03-0.05
0.06-0.2
ng/8.
Mean
3
1
13
0.1
2
N0b
0.3
NDb
0.2
0.04
0.1
aSource: Oliver and Nlchol, 1982
bL1m1ts of detection were -0.1 ng/SL for the trlchlorobenzenes and -0.05
ng/2. for the tetrachlorobenzenes.
ND = Not detected
4-24
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local synthetic carpet rams. Average concentrations reported for dlchloro-
benzenes In the Incoming and outgoing water ranged from 3-146 yg/8, and
0-268 vig/l, respectively. For the trlchlorobenzenes, the levels ranged
from 1-60 pg/a, and 0-13 yg/St for Influent and effluent, respec-
tively. The author concluded that the Increase 1n the dlchlorobenzene
levels was a result of chloMnatlon performed during the secondary phase of
wastewater treatment.
A U.S. EPA survey of wastewater throughout the United States found that
dlchlorobenzenes and trlchlorobenzenes occurred 1n discharges from Indus-
trial and municipal plants (Ware and West, 1977). The reported concentra-
tions ranged from 15-690 vig/S. for 1,2-d1chlorobenzene and from 0.25-500
yg/8, for 1,2,4-tr1ch1orobenzene. Young and Hessan (1978) also reported
the presence of several chlorobenzenes 1n the wastewater of major municipal
facilities 1n southern California. The highest concentrations they found
were for water discharged by the Los Angeles Hyperion Treatment Facility
during December: 1,2-dlchlorobenzene, 435 iig/2.; 1,4-d1chlorobenzene, 230
yg/a,; 1,2,4-tr1chlorobenzene, 130 yg/a. and 1,3,5-trlchlorobenzene
<0.2 yg/8.. For the other sites, the levels of dlchlorobenzene Isomers
ranged from 0.2-6 yg/a,. None of the facilities used chlorlnatlon to
treat the water.
Neptune (1980) compiled data for organic priority pollutants analyzed In
samples taken during several Industrial wastewater surveys conducted from
September 1978 through October 1979. The resulting data were grouped by
their occurrence 1n each of 35 standard Industrial categories. The most
prevalent of the chlorobenzenes was monochlorobenzene, which was present In
waste streams from 14 Industrial categories; 1,2-, 1,3- and 1,4~d1chloro-
benzene were detected In 13, 10 and 14 categories, respectively, with
4-25
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1,2,4-tr1chlorobenzene present 1n 7 categories. The frequency and range of
concentrations found are summarized 1n Table 4-12.
4.3.3, Food. Investigation of the occurrence of chlorobenzenes 1n food
has been limited primarily to the measurement of hexachlorobenzene. This
concern derived from Its use as a fungicide on the seeds of several food
crops and from Its ability to bloaccumulate 1n the food chain. The bio-
accumulation potential of hexachlorobenzene and the other chlorobenzenes 1s
discussed 1n Section 5.3.
Johnson and Hanske (1976) reported the detection of hexachlorobenzene at
levels of 0.0006-0.041 mg/kg 1n food samples from 30 U.S. cities obtained
from the Total Diet Program of the U.S. Food and Drug Administration (FDA).
Based on these and other data, FDA estimated the average dally Intake of
hexachlorobenzene from foods 1n fiscal years 1973 and 1974 to be 0.3978 and
0.0725 yg/day, respectively (IARC, 1979). Leonl and D'Arca (1976), using
analysis data on cooked foods served 1n the Italian Navy, estimated an aver-
age dally Intake of 4.11 yg of hexachlorobenzene. In addition, these
authors surveyed uncooked foods available to the public and found mean hexa-
chlorobenzene levels ranging from none detected to 133.0 ppb (0.133 mg/kg),
with the highest levels occurring In butter, lard and pork meat. The aver-
age dally Intake of hexachlorobenzene from uncooked diets was calculated to
be 4.32 yg, a value similar to Intake from cooked diets. Hexachloroben-
zene has also been detected 1n Navy foodstuffs available 1n Japan (Morlta et
a!., 1975a,b; Seklta et al., 1980) and was measured 1n beef (12 yg/kg),
salmon (9 yg/kg), pork (7yg/kg) and other animal sources of protein
(Morlta et al., 1975a,b), In a survey of over 300 suppliers of cow's milk
1n southern Ontario 1n 1977, hexachlorobenzene residues were Identified 1n
6854 of the samples at a level of 0.002 mg/kg 1n fat. The hexachlorobenzene
4-26
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TABLE 4-12
Frequency and Range of Concentrations of Chlorinated
Benzenes Pollutants 1n Industrial Wastewaters*
Chemical
Monochlorobenzene
1 ,2-D1chlorobenzene
1 ,3-D1chlorobenzene
1 ,4-01chlorobenzene
1 ,2,4-Tr Ichlorobenzene
Total
Samples
31,194
3,268
3,268
3,268
3,266
Samples
Number
of Samples
147
80
44
88
30
Containing >
Range
11-6,400
12-860
10-39
10-410
12-607
10 ug/8.
Mean
Concentration
667
141
21
79
161
*Source: Neptune, 1980
4-27
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levels found 1n the 1977 survey were significantly below the levels detected
1n a similar survey conducted 1n 1973 (Frank et al., 1979).
Some Information on the presence of other chlorobenzenes 1n food was
also available. Morlta et al. (1975c) reported detecting 1,4-d1chloroben-
zene 1n fish Including mackerel caught 1n Japanese coastal waters. Oliver
and Nlchol (1982) reported detecting all Isomers of dlchlorobenzene, tr1-
chlorobenzene, tetrachlorobenzene, pentachlorobenzene and hexachlorobenzene
1n trout from the Great Lakes. The highest levels were detected for penta-
and hexachlorobenzene and the mean concentrations were 5.5 and 46.8 vg/kg,
respectively. Residues of pentachlorobenzene have also been found 1n oils,
fats and shortening (0.001-0.11 mg/kg) and sugar (-0.002 mg/kg) (U.S. EPA,
1980).
4.3.4. Soil and Sediments. The study of soil contamination by chloro-
benzenes has focussed on hexachlorobenzene, although more recent surveys
have Included all the chlorobenzenes.
Hexachlorobenzene has been detected 1n sediment samples taken from lakes
throughout Germany (Buchert et al., 1981) and was measured at 0.04 ppm 1n
soil from a farming region 1n Italy where high mortality among birds had
taken place (Leonl and D'Arca, 1976). In 1975, the U.S. EPA examined soil
and aquatic sediments at 26 locations along a 150-mile transect of Louisiana
and found that 46% of the soil samples were contaminated with hexachloroben-
zene at levels of 20-440 ppb. The aquatic sediments contained hexachloro-
benzene at levels of 40-850 ppb (Blackwood and Spies, 1979). In a survey of
nine Industrial plants (see Table 4-9 for site locations) producing chloro-
carbon compounds, L1 et al. (1976) detected hexachlorobenzene 1n soil
samples taken within the plant area at levels >1000 vg/g (1000 ppm) at
three plants. Soil taken from the cornfield adjacent to one plant contained
4-28
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(1100 ppb) and >3000 pg/g were detected along a boundary road
of another plant.
Elder et al. (1981) sampled sediments 1n streams draining the Love Canal
area of Niagara Falls, NY, and detected all of the chlorobenzenes (Isomers
unspecified) at levels 1n the ppm range. Oliver and Nlchol (1982) studied
the fate and distribution of chlorobenzenes 1n the Great Lakes and reported
detecting all Isomers 1n the sediments of Lakes Superior, Huron, Erie and
Ontario. The most contaminated lake was Lake Ontario, for which the mean
levels 1n the sediments of the Individual Isomers ranged from 11-94 ng/g for
the dlchlorobenzenes, from 7-94 ng/g for the trlchlorobenzenes, and from
6-52 ng/g for the tetrachlorobenzenes. The levels of penta- and hexachloro-
benzene were measured as 32 and 97 ng/g, respectively.
4.3.5. Human Tissue Residues. Studies of the transport, fate and bio-
accumulation of the chlorinated benzenes reviewed above Indicate that human
exposure 1s likely from air, food and drinking water (Sections 5.1., 5.2.
and 5.3.). In this section, human ambient exposure 1s confirmed by the
reported levels of chlorobenzene 1n human adipose tissue, blood, breath and
urine; unfortunately, the environmental concentrations were not available
for comparison with the observed tissue concentrations.
Due to the HpophlUc character of the chlorinated benzenes, as Indicat-
ed by their octanol/water partition coefficient discussed 1n Section 5.3.,
adipose and other fatty tissues are the major tissue depots for chlorinated
benzenes. The measured levels of several chlorobenzene Isomers In human
adipose tissue are shown 1n Table 4-13.
Since human milk has a high fat content, chlorinated benzenes Ingested
by pregnant and nursing mothers would be likely to distribute to this depot
and, on repeated exposure, to bloaccumulate. Thus the suckling offspring
would be susceptible to a high exposure via this Intake. Stacy and Thomas
4-29
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TABLE 4-13
Chlorinated Benzene Residues 1n Human Adipose Tissue
Compound
Country
Tissue
Concentration
Cmg/kg)*
Reference
1 ,4-D1chlorobenzene
1,2,4,5-
Tetrachlorobenzene
Hexachlorobenzene
Japan
Japan
Japan
Japan
Japan
United States
Italy
Great Brltlan
Germany
New Zealand
Canada
Canada
Sweden
2.3
1.88
1.7
0.019
0.21
0.03-0.47
0.491
0.05
6.3
0.31
0.001-0.52
0.01-0.67
0.029-0.071
Horlta and Oh1, 1975
MorHa and Oh1, 1975
Morlta et a!., 1975?
Horlta et al . , 1975?
Horlta et al., 1975?
Barquet et al., 1981
Leonl and O'Arca, 1976
Abbott et al., 1972
Acker and Shulte, 1970
Solly and Shanks, 1974
Mes et al., 1979
Mes et al., 1982
Noren, 1983
*Values are for adipose tissue
4-30
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(1975) analyzed breast milk samples from 20 urban and 20 rural Australian
mothers and found the concentration of hexachlorobenzene 1n rural milk
(0.079 mg/kg milk) significantly greater than that In urban milk (0.028
mg/kg). A study of another group of Australian mothers showed the opposite
results: rural milk contained 0.042 mg/kg while urban milk contained 0.063
mg/kg (Newton and Greene, 1972). In France, 18 of 49 breast milk samples
contained hexachlorobenzene at concentrations of 0.001-0.17 mg/kg whole milk
(0.50-3.50 mg/kg on fat basis) (Goursaud et a!,, 1972). Relatively low con-
centrations of pentachlorobenzene and hexachlorobenzene (0.002 mg/kg and
0.006 mg/kg, respectively) were found 1n milk samples from Yugoslavian women
(Kodr1c-Sm1t et a!., 1980). In another study, 50 milk samples from Helsinki
women In 1982 (Wlckstrom et al., 1983) contained 0.7-6 ^g hexaehloroben-
zene/kg whole milk (14-240 jig hexachlorobenzene/kg milk fat). No detect-
able hexachlorobenzene was found, however, 1n 57 samples of breast milk from
women of rural Arkansas and Mississippi (Strassman and Kutz, 1977). Levels
1n two Swedish women varied from 0.029+0.002 mg hexachlorobenzene/kg milk
fat In one to 0.07H0.005 mg hexachlorobenzene/kg milk fat In the other
(Noren, 1983). Courtney (1979) reviewed some of these and other studies
that substantiate the ubiquity of hexachlorobenzene by the fact that people
with no known exposure to the chlorobenzene had measurable tissue con-
centrations.
In a study Involving 28,000 people across the United States (Murphy et
al., 1983), hexachlorobenzene was found 1n 4% of 4200 blood serum samples
using a method with a detection limit between 1 and 2 yg/8,. In addi-
tion, hexachlorobenzene was found In 93% of 785 adipose tissue samples,
using a method with detection limits around 10-20 ng/$t. These findings
were Interpreted as signifying non-occupational exposures. No actual levels
were provided In this study.
4-31
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From the data on chlorobenzene concentrations found In human blood and
plasma, U 1s apparent that newborns to adults, even those from Industrially
remote areas or those with no known chlorobenzene exposure, experience
exposures to these compounds.
Astolfl et al. (1974) reported hexachlorobenzene at 19 ng/l 1n the
umbilical cord blood of Infants born 1n Argentina. Ninety-seven rural and
97 urban children from Upper Bavaria all had detectable levels of hexa-
chlorobenzene 1n their blood ranging from 2.8-77.9 ppb (ng/g); the average
concentration was 16.5 ppb (Rlchter and Schmld, 1976). An average concen-
tration of 22 ppb hexachlorobenzene was measured 1n the blood of nonexposed
Australians, whereas occupatlonally exposed people had an average concentra-
tion of 55.5 ppb (range 21-100 ppb) 1n their blood (SlyaU, 1972). MorHa
and Oh1 (1975) analyzed the blood of four male and two female residents of
Tokyo for 1,4-d1chlorobenzene and reported an average of 9.5 yg/Sl.
Wastes containing hexachlorobenzene were spread on a landfill 1n western
Louisiana as a fly control measure (Burns and Miller, 1975). Blood levels
of hexachlorobenzene 1n 22 husband-wife pairs living near the landfill were
analyzed. The average blood level for the men was 5.10 ppb, which was
significantly higher than that for the women, which was 1.70 ppb. Forty-six
Louisiana residents not living 1n the Immediate vicinity of the landfill had
average blood levels of 0.5 ppb hexachlorobenzene, while chemical plant
workers In the area had a blood concentration range of 14-233 ppb. The
levels of chlorinated benzenes 1n the blood of nine residents of the Love
Canal area In Niagara Falls, New York, were measured and are shown 1n Table
4-14 (Barkley et al., 1980).
Although the chlorobenzenes bloaccumulate 1n human adipose tissue and
are detected In human blood, the levels are tempered by the elimination pro-
cesses. The expired breath and urine of nine residents of the Love Canal
4-32
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TABLE 4-14
Chlorinated Benzenes 1n the Blood of Nine Residents
of Love Canal 1n Niagara Falls, New York*
Compound
Honochlorobenzene
D1-1somers
Tetra-lsomers
No. of Positive
Results
8
9
1
Blood
Concentration
(ng/ms.)
0.05-17.0
0.15-68
2.6
*Source: Barkley et a!., 1980
4-33
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area contained measurable levels of the chlorobenzenes as shown 1n Table
4-15 (Barkley et al., 1980).
The concentrations of chlorinated benzenes reported for human tissue
(see Table 4-13) and blood, breath and urine (see Tables 4-14 and 4-15)
Indicate that humans absorb and store chlorinated benzenes. The bloaccumu-
latlon of the chlorinated benzenes 1s offset by metabolism and elimination
from the body.
4.4. RELATIVE SOURCE CONTRIBUTIONS TO TOTAL EXPOSURE
The monitoring studies discussed 1n the preceding section Indicate that
chlorinated benzenes are present 1n the environment and that human exposure
to one or more of these substances 1s likely to result from the Inhalation
of air or the 1ngest1on of water or food. The Intent of this section 1s to
estimate the relative degree that these three media — air, water and food
— contribute to a person's overall exposure. There are several limitations
to this approach. First, no comprehensive study of human exposure to the
chlorobenzenes has been conducted; the available monitoring data Indicate
the presence of the substances under the conditions of a given study and do
not establish universal levels of exposure. Consequently, the studies that
are cited and discussed 1n this section were selected on the basis of being
the most likely to represent general population exposure. Data on Instances
of gross contamination; I.e., local pollution from landfill or 1n an occupa-
tional setting, were not used. Second, no single study has analyzed any one
medium for all of the chlorobenzenes. Hence, only data from a single study
were used 1n the calculations for one type of exposure; aggregate or com-
bined data were not used. Third, all monitoring studies are limited either
1n terms of sampling duration or the number of locations sampled; studies
with the widest geographical sampling locations and longest duration of
4-34
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TABLE 4-15
Chlorinated Benzenes 1n the Breath and Urine of Nine
Residents of Love Canal 1n Niagara Falls, New York*
Compound
Monochlorobenzene
D1-1somers
Tr1-1somers
Tetra-lsomers
Pentachlorobenzene
No. of Posl
Breath
1
7
2
2
1
tlve Results
Urine
6
7
0
0
0
Concentration
Breath
(ng/m3)
T
T-5000
T-90
30-180
70
Range
Urine
(ng/Jt)
20-120
40-39,000
NO
ND
NO
*Source: Barkley et a!., 1980
T = Trace; NO = not detected
4-35
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sampling were favored for use 1n the calculations. Finally, quantitative
data on the absorption of the various chlorobenzenes by humans through the
lungs, skin or gastrointestinal tract 1s not available. For this reason,
the data 1n this section are estimates of yearly average ambient exposure
levels (I.e., the amount potentially Inhaled or Ingested) and are not
physiological exposure levels.
4.4.1. A1r. The monitoring data used for the estimation of Inhalation
exposure (Table 4-16) are taken from the overall mean concentration values
given 1n Table 4-7. In addition, this table presents estimates of the total
yearly exposure of an adult man, adult woman, child and Infant using stan-
dard respiratory volumes of 8.4xlQ«, 7.7x10*. 5.5x10* and 1.4xl06
a/year, respectively (ICRP, 1975). The Inhalation exposure estimate will
be different for rural/remote, urban/suburban and source areas.
4.4.2. Water. The estimation of exposure of chlorobenzenes from drinking
water requires that the mean or median concentrations of these compounds 1n
finished water originating from a large number of both U.S. surface and
groundwater be known. As discussed 1n Section 4.3.2., only a limited number
of monitoring data for the levels of chlorobenzenes 1n finished water
samples are available. Therefore, a realistic assessment of the exposure of
chlorobenzenes through the 1ngest1on of drinking water cannot be made at the
present time. However, the maximum concentrations of the chlorobenzenes
found 1n U.S. drinking water are the following: monochlorobenzene, 5.6
pg/st; 1,3-d1chlorobenzene, <3 yg/st; and trlchlorobenzene (Isomer
unspecified), 1.0 yg/2. (NAS, 1977). If the maximum fluid Intake by an
Individual 1s assumed to be 711.8 8,/year (ICRP, 1975), the maximum expo-
sure of an Individual chlorobenzene Isomer through 1ngest1on of finished
water can be estimated to be <4 mg/year.
4-36
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TABLE 4-16
Estimated Yearly Exposure to Several
Chlorinated Benzenes Via Inhalation
Exposure (mg/yr}
Chemical
Monochlorobcnzenes
1 ,2-D1chlorobenzene
1 ,3-D1chlorobenzene
1 ,4-D1chlorobenzene
Trlchlorobenzenes
Tetrachlorobenzenes
Mean Ambient Con-
centration (ng/ma)*
3087
1142
571
1563
136
3502
Adult
Han
25.9
9,6
4.8
13.1
1.1
29.4
Adult
Woman
23.8
8.8
4.4
12.0
1.0
27.0
Child
(10 yr)
17.0
6.3
3.1
8.6
0.7
19.3
Infant
(1 yr)
4,3
1.6
0.8
2.2
0.2
4.9
*Hean levels obtained from Table 4-8
4-37
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4.4.3. Food. Hexachlorobenzene 1s the only chlorinated benzene whose
presence in food has been systematically Investigated. Based on data from
the Total Diet Program, the FDA estimated the average dally Intake of
hexachlorobenzene for fiscal year 1974 to be 0.0725 vg/day (IARC, 1979).
This would result 1n a yearly exposure of 0.03 mg hexachlorobenzene from
food sources.
4.5. SUMMARY
Annual production of chlorinated benzenes 1n 1983 1s on the order of 450
million pounds, the majority of which 1s accounted for by monochlorobenzene
and dlchlorobenzenes. Production of the tr1- and tetrachlorobenzenes and
pentachlorobenzene 1s on the order of millions of pounds/year. Hexachloro-
benzene 1s not currently produced as a commercial product 1n the United
States (IARC, 1979), although 1t 1s a constituent of several Imported
products and Is a byproduct or waste material 1n the production of many
chemicals (Mumnta and Lawless, 1975). These compounds are used 1n a number
of organic chemical syntheses, Including the synthesis of other chloroben-
zenes, and have applications as solvents, electrical equipment Insulators,
pesticides, herbicides and fungicides. Emissions of chlorobenzenes are most
Hkely to occur during their manufacture or use as Intermediates and from
the disposal of waste products from manufacturing operations. Hexachloro-
benzene, for example, which 1s Imported but not produced commercially 1n the
United States, occurs as a byproduct In the synthesis of nine other chloro-
carbons; 2-5 million pounds may be generated each year.
Chlorinated benzenes have been Identified 1n air, food and soil, and 1n
surface, ground and drinking water. The highest concentrations have been
found near manufacturing and waste disposal sites, although no study has
attempted to characterize the contribution of any one source to the total
4-38
-------
environmental contamination by ehlorobenzenes. Ambient air and maximum
water levels are 1n the »ig/m3 and mg/ft. range, respectively, although
monitoring studies for finished water have been limited. The most frequent-
ly detected compounds 1n air and water were monochlorobenzene and the d1-
and trlchlorobenzenes. Penta- and hexachlorobenzene are more frequently
found 1n food and soil, although their detection may reflect more of the
concern over their use as pesticides and fungicides, or their presence as
contaminants 1n pesticides or fungicides, rather than the absence of the
other ehlorobenzenes.
No comprehensive study of human exposure to the chlorobenzenes has been
conducted, although their ubiquity 1n the environment and the detection of
measurable residues In human tissue (see Section 5.3., B1oaccumulat1on/B1o-
concentratlon) Indicate that human exposure and absorption occur. The
contribution of the chlorobenzenes from all the three media (air, water and
food) to a person's total exposure cannot be made with the limited environ-
mental monitoring data. The available data, however, Indicate that human
Inhalation exposure to chlorobenzenes may be higher than Ingestlon exposure
either through drinking water or through foods.
4-39
-------
5. ENVIRONMENTAL TRANSPORT AND FATE
The following sections consider the transport and fate of the chlori-
nated benzenes through the three environmental media (air, water and soil)
and their potential to accumulate or concentrate 1n plant, animal and,
ultimately, human tissues. Transport between the various environmental
media 1s governed by the physical and chemical characteristics of the com-
pounds and their Interaction with components of the environment. Evapora-
tion rates and solubilities Influence transport from water and soil Into
air. Leaching rates, adsorption, rainfall, soil type and desorptlon affect
the movement of chlorobenzenes from soil and sediment Into water and ground-
water, as well as from water Into sediment and soil. The fate of chloroben-
zenes 1n the environment depends on degradatlve processes, either abiotic
degradation by chemical reactions or photolysis, or blotlc degradation by
microbes, and on the rate at which these compounds are stored or accumulated
by plants, animals and humans.
5.1. TRANSPORT
5.1.1. A1r. The transport and distribution of the chlorobenzenes 1n the
atmosphere has not been Investigated. One study has suggested that distri-
bution of one of the chlorobenzenes 1n air may be global. Atlas and Slam
(1981) reported detecting hexachlorobenzene at a mean level of 0.10 ng/m3
in air samples taken at a remote North Pacific Ocean atoll where the only
source could be air transport. These data led the authors to suggest that
hexachlorobenzene 1s well mixed 1n the atmosphere and has wide distribution
In the Northern Hemisphere. A study of environmental contamination by hexa-
chlorobenzene from Industrial plants (L1 et al., 1976) provided some data
that Indicated such emissions can be spread by wind from point sources. The
5-1
-------
authors reported that the emissions, which were 1n both vapor and partlcu-
late form, were detected at levels from 0.1-24.0 yg/m3 near the produc-
tion facilities and decreased to 0.10-0.50 as much 200 feet or more down-
wind. The tendency of the hexachlorobenzene to remain 1n the atmosphere was
not studied.
Entry Into the atmosphere from other media 1s determined mainly by the
substance's molecular weight, water solubility and vapor pressure. Chloro-
benzenes have vapor pressures ranging from 0.05-11.8 mmHg at 20°C (see
Section 3.3.). In general, these vapor pressures decrease with the Increase
1n the number of chlorine substltuents. Chlorobenzenes are likely to enter
the atmosphere as a result of evaporation from soil and water and these
types of studies are discussed 1n the following sections.
5.1.2. Water. Chlorobenzenes have low solubility 1n water, with the solu-
bility decreasing as the number of chlorine substltuents Increases, although
some variation 1s evident among the Isomers (Hawley, 1977; Sax, 1979; Weast,
1979) (see Section 3.3.). Once dissolved 1n water, despite their relatively
low vapor pressures and high molecular weights, the chlorinated benzenes
tend to evaporate quickly (Mackay and Wolkoff, 1973). Two laboratory
studies Indicate that evaporation of some of the Chlorobenzenes from an
aqueous solution could be as rapid as a few minutes to a few days.
Garrison and H111 (1972) found that >99% of mono-, 1,2- and l,4-d1- and
1,2,4-trlchlorobenzene had evaporated within 4 hours from aerated distilled
water solutions and within 72 hours from nonaerated solutions. Mono-, 1,2-
d1- and 1,4-d1chlorobenzene volatilized completely 1n <1 day from aerated
solutions containing mixed cultures of aerobic microorganisms. l,2,4-Tr1-
ch robenzene also evaporated, but less rapidly, with 2% of the Initial
cc :entrat1on remaining after 80 hours. Lu and Hetcalf (1975) provided
5-2
-------
evidence of monochlorobenzene's volatility from water through their study of
this chemical's blodegradablHty 1n a model ecosystem. They noted that
after 48 hours, 96% of the radloactlvely labeled compound added to the model
system was found 1n the traps that sampled the system's atmosphere.
A 1-year field study of the transport of 1,4-dlchlorobenzene 1n Lake
Zurich, Switzerland, also Indicated an Important role for evaporation 1n the
removal of chlorobenzenes from water (Schwarzenbach et a!., 1979). The
authors found that the main Input of 1,4-d1chlorobenzene Into the lake was
from wastewater treatment plants and that the half-life of the chemical was
-100 days. From a comparison of the seasonal variation 1n evaporation
rates, they concluded that transport Into the atmosphere 1s the predominant
Influence on the loss of 1,4-d1chlorobenzene from the lake. Their data
Indicated that of the 90 kg/year entering the lake, 60 kg was lost to the
air, 2 kg entered lake sediments and 28 kg was 1n the lake's outflow.
In addition to laboratory and field Investigations, theoretical studies
of the transport of chlorobenzenes In aquatic systems may be useful 1n
predicting the distribution of these compounds and their removal from water
by evaporation and sedimentation. Using Henry's Law Constant and various
assumptions of water depth, air speed, etc., the half-life of evaporation
from water can be calculated. For chlorobenzene, 1,2-dlchlorobenzene and
1,2,4-tr1chlorobenzene, these values are 4.6 minutes, 8.1 minutes and 0.75
hours, respectively. Falco et al. (1982) developed a mathematical model for
assessing the transport and degradation of materials released from land-
fills and waste storage lagoons. The parameters Incorporated Into the model
Included coefficients for the following: (1) octanol/water partition, (2)
hydrolysis rate, (3) photolysis rate, (4) bacterial degradation rate, (5)
oxidation rate, (6) overall degradation rate and (7) volatilization rate.
The predictions made using the model are therefore limited by the available
5-3
-------
data base and rate coefficients. In their modeling, Falco et al. (1982)
used the model to predict the transport and persistence of chlorobenzenes
for the following types of surface waters: (1) a river capable of trans-
porting a chemical 50-100 miles 1n 5 days, (2) a pond with an average
retention time of 100 days and (3) a lake or reservoir with a retention time
of 1 year. For comparative purposes, a summary of the authors' results for
the lake or reservoir 1s presented 1n Table 5-1.
5.1.3. Soil. Chlorobenzenes have an Intermediate to high potential for
adsorption onto soils, which tends to Increase with Increasing number of
chlorine substltuents. Once adsorbed, their movement within the soil 1s
dependent on the soil type and the nature of the solvent or leachate. In
the absence of a solvent, transport Into adjacent soil and the atmosphere 1s
likely to result from vapor phase diffusion.
Wilson et al. (1981) studied the transport, over a 21-day period, of a
mixture of monochlorobenzene, 1,4-d1chlorobenzene, 1,2,4-tr1chlorobenzene
and 10 other organic chemicals through a column of sandy soil having a low
organic matter content. Using water as a solvent, these Investigators noted
that for the chlorobenzenes the retardation factors (velocity of the sol-
vent/velocity of a compound) Increased with the chlorine number regardless
of the Initial concentration of the compounds. These authors also reported
that up to 50% of the applied monochlorobenzene evaporated and -50% of the
amount of all three chlorobenzenes was degraded or unaccounted for (Table
5-2). These results Indicated that chlorinated benzenes are likely to leach
Into groundwater and this mobility 1n groundwater was confirmed 1n a field
study by Roberts et al. (1980).
Studies on the transport of hexachlorobenzene Indicate a high potential
for soil adsorption and for volatilization from porous soils. Ausmus et al.
5-4
-------
TABLE 5-1
Predicted Transport and Fate of Chlorinated Benzenes
Released from Landfills and Lagoonsa
Percentage of
Total Amount of Chlorobenzene Entering Lake
Property
1,2- and
Mono- 1,4-01- Tr1-b Tetra-b Penta-
Movement from point of
entry to outlet
Potential for degradation
or elimination
Amount absorbed onto
suspended sediments
Amount taken up by fish
Estimated volatilization
to atmosphere
5-9
83-94
-------
TABLE 5-2
Transport of Chlorinated Benzenes In Sandy Soil*
Chemical
Monochlorobenzene
1
1
»4-D1chlorobenzene
»2»4-Tr1chlorobenzene
Percentage
Volatilized
27-54
ND
ND
of Total Chlorobenzene
Degraded or
Not Accounted For
20-40
51-63
54-61
Applied
Column
Effluent
26-33
37-49
39-46
*Source: Wilson et al., 1981
NO = Not determined
5-6
-------
(1979) applied C^-labeled hexachlorobenzene to soil cores taken from a
pine forest and monitored Its evaporation and leaching by water over 21
days. Of the amount applied, <1X was lost by volatilization or 1n the
leachate, and none was degraded as Indicated by the absence of labeled
CO-. Farmer et al. (1980a) examined the vapor phase diffusion of hexa-
chlorobenzene through a high clay, low organic material soil (39 and IX,
respectively) and reported diffusion to be Increased by the soil porosity
and decreased by the soil's water content. The same authors (Farmer et al.,
1980b) also found that highly compacted wet soil covers were most effective
1n reducing hexachlorobenzene volatilization after dumping Into a land-fill.
A water cover 1n a temporary storage lagoon was also effective. Each 10°C
rise 1n soil temperature Increased volatilization fluxes 3.5-fold. Griffin
and Chou (1981) Investigated the adsorption and mobility of polychlorlnated
and polybromlnated blphenyls and hexachlorobenzene 1n seven different soil
types with Increasing amounts of organic carbon. The adsorption of hexa-
chlorobenzene Increased with Increasing amounts of organic carbon. Further,
they noted that hexachlorobenzene was Immobile and was not leached from the
three soils that were tested with water and a leachate from a landfill.
Soil sorptlon coefficient (K ) values for chlorinated benzenes are:
chlorobenzene (537), 1,2-d1chlorobenzene (977), 1,4-d1chlorobenzene (1259),
1,2,3-trlchlorobenzene (2630), 1,2,4-tr1chlorobenzene (2042) and hexachloro-
benzene (38,000) (Calamarl et al., 1983).
5.2. FATE
5.2.1. A1r. The degradation of chlorobenzenes 1n air has been studied 1n
a fair amount of detail. In theory, chlorobenzenes dispersed 1n air may be
degraded by chemical- or sunlight-catalyzed reactions or may be adsorbed
onto particles that settle or are removed from the atmosphere by rain. A
measure of the effectiveness of these factors 1s the atmospheric residence
5-7
-------
time. One study has made estimates of residence times for various chloro-
benzenes. Singh et al. (1981) conducted field studies 1n California and
Arizona and analyzed air samples over 2-week periods for 33 organic chemi-
cals Including monochlorobenzene, the dlchlorobenzenes and an unspecified
Isomer of trlchlorobenzene. The estimated-, residence times of these chemi-
cals and dally percentage of each lost from the atmosphere are presented 1n
Table 5-3.
5.2.2. Water. The fate of chlorobenzenes 1n aquatic systems has not been
completely characterized, although Initial studies Indicate that degradation
of chlorobenzenes 1s possible by microblal communities 1n wastewater treat-
ment plants and 1n natural bodies of water. Other Investigations have Indi-
cated that chlorobenzenes have a high potential for bloaccumulatlon and bio-
concentration by aquatic species (Section 5.3.). Removal of chlorobenzenes
by adsorption onto suspended material, which 1n turn settles and Is Incorpo-
rated Into sediments, has not been demonstrated.
Lee and Ryan (1979) examined the degradation of various chlorinated com-
pounds by microbes 1n samples of water and sediments taken from a river 1n
Georgia. They observed that the degradation rates fit first-order expres-
sions, although the degradation of the chlorinated compounds 1n water was
slow. In the sediment samples, monochlorobenzene was found to have a
half-life of 75 days, which was longer than the chlorinated phenols, but
more rapid than the degradation of hexachlorophene and DDE. In contrast,
hexachlorobenzene showed no degradation by water or sediment microbes.
Davis et al. (1981) conducted a similar experiment using samples of mic-
roblal populations from Industrial and municipal wastewater treatment plants
and 1,2-d1chlorobenzene along with other compounds. The dlchlorobenzene at
a concentration of 50 ng/s, was degraded by both systems within 7 days.
5-8
-------
TABLE 5-3
Estimated Atmospheric Residence Time and Dally Loss
Rates for Several Chlorinated Benzenes3
Chemical
Monochlorobenzene
1 ,2-D1chlorobenzene
1 ,3-D1chlorobenzene
1 , 4-D1chlorobenzene
Tr Ichlorobenzene^
Residence Times
{days)b
13
38.6
38,6
38.6
116.0
Dally Loss Rate
(percent)0
7.4
2.6
2.6
2.6
0.9
aSource; Singh et a!., 1981
^Calculated assuming an average dally (24-hour) abundance of OH radicals
of 106 molecules/cm3
cFor 12 hours of sunlight
^Isomer unspecified
5-9
-------
This rate, which was more rapid than the rate for phenol but slower than
that measured for benzene, was described as "comparatively rapid."
Using a model aquatic ecosystem, Lu and Hetcalf (1975) Investigated the
blodegradatlon and bloaccumulatlon of various chemicals Including monochlo-
robenzene and hexachlorobenzene. Both compounds were found to have high
"ecological magnification" Indices and accumulated 1n both aquatic plants
and animals. Both chemicals had low b1odegradab1!1ty Indices and were, 1n
general, metabolized to monochloro- and pentachlorophenol, respectively.
A number of other Investigators have studied the b1odegradabH1ty of
chlorinated benzenes and these results are summarized 1n Table 5-4, In
general, these results suggest that the b1odegradabH1ty decreases as the
number of chlorine substltuents Increases, In addition to these laboratory
studies, Zoeteman et al. (1980) Indicated that chlorobenzene, o-d1chloroben-
zene, j)-d1chlorobenzene, 1,2,4-tr1chlorobenzene and hexachlorobenzene
degrade 1n river water with half-lives of 0.30, 3-2, 1.1-25, 1.8-28 and 0.5
days, respectively, as Indicated by monitoring at various stations along the
Rhine River. These half-lives are likely to be very Inaccurate since only a
limited number of samples were taken.
Roberts et al. {1980} studied the transport and degradation of monochlo-
robenzene, £-, m-, g-chlorobenzenes and 1,2,4-trlchlorobenzene 1n ground-
water after Injection by analyzing monitoring wells at different distances
from the Injection well. No degradation was noted.
5.2.3. Soil. Studies on the fate of dlchlorobenzenes, trlchlorobenzenes,
pentachlorobenzene and hexachlorobenzene 1n soil have Indicated that the
chlorobenzenes are usually resistant to microblal degradation (however,
compare Ballschmlter and Scholz, 1980} and that chlorophenols are likely
degradation products. Beck and Hansen (1974) studied the blodegradatlon of
5-10
-------
TABLE 5-4
Aqueous BlodegradabllHy Studies of Chlorinated Benzenes
Method
Warburg
Mineral salts
shake flask
tn HITI BOD Test
i
Warburg (phenol
acclimated
cultures)
BOD 5-day
Natural water
Warburg (sewage)
MCB
3.9 BOOT
100*
Resistant to
degradation
16.1 80DT
1.5 BOOT
Degradation fast
1n fresh water,
slower tn estuar-
1ne and marine
water
Results (X Degradation)
o-DCB B-DCB 1.2,4-TCB HCB
Trace of 3.4 BOOT No degradation
degradation
18-66* 0-61* 0-70* 0-56*
Resistant to Resistant to
degradation degradation
2.4 BOOT
Halaney
Tabak et
Kawasaki
Chambers
Reference
and McKlnney. 1966
al., 1981
, 1980
et al., 1963
Heukeleklah and Rand. 1965
slow degradation
0-54
Pfaender
1982
Gaffney,
and Bartholomew,
1976
*Percent degradation after acclimation (subculture every 7 days)
-------
Qulntozene, a fungicide, and two of Its Impurities, penta- and hexaehloro-
benzene. Soil samples treated under laboratory conditions with penta- and
hexachlorobenzene at rates equal to 10 mg/ha were monitored over a period of
600 days. From the slopes of the degradation curves, the authors estimated
the half-lives of penta- and hexachlorobenzene to be 194-345 and 969-2089
days, respectively. Beall (1976) applied hexachlorobenzene at an amount
equivalent to 750 g/ha to sections of turf 1n a greenhouse. Within 2 weeks,
55% of the hexachlorobenzene had disappeared from the top 2 cm of soil, most
Hkely a result of evaporation. Very little of the chemical disappeared
from the 2-4 cm-deep soil layer over the next 19 months. Isensee et al.
(1976) also found hexachlorobenzene to be highly persistent 1n soil. Hexa-
chlorobenzene was applied to samples of sterile and nonsterlle soil to
create levels of 0.1, 1, 10 and 100 ppm. After storage of the samples under
aerobic (sterile and nonsterlle) and anaerobic (nonsterlle) conditions for 1
year, analysis Indicated that none of the hexachlorobenzene had degraded 1n
any sample.
Studies with the d1- and trlchlorobenzenes have Indicated that these
compounds are also persistent, but not to the degree reported for hexachlo-
robenzene. Ballschmlter and Scholz (1980) Investigated the metabolism of
1,2-, 1,3- and 1,4-d1chlorobenzene by a soil microbe of the Pseudomonas
genera. In culture, the soil microbe was capable of degrading the compounds
to dlchlorophenols and dlchloropyrocatechols. Similar cultures of Pseudo-
monas also metabolized the tr1- and tetrachlorobenzenes to their respective
chlorophenols. In an experiment that more closely duplicated conditions 1n
nature, Marlnucci and Bartha (1979) treated fresh field soil with radio-
labeled 1,2,3- and 1,2,4-tr1chlorobenzene. They found very slow rates of
degradation for these compounds, 0.35 and 1.00 nmol/day/20 g of soil,
respectively. These authors also noted that the amount of organic material
5-12
-------
1n the soil had no effect on the rate, but It did appear to reduce evapora-
tion of the chlorobenzenes. The primary degradation products were chloro-
phenols. Haider et al. (1974), using 14C-labeled compounds 1n soil, found
18.3, 1.1 and 1.1% C0? after 1 week of Incubation of monochlorobenzene,
o-d1chlorobenzene and p_-chlorobenzene, respectively.
5.3. BIOCONCENTRATION, BIOACCUMULATION, AND BIOMASNIFICATION
The occurrence of toxic substances 1n the environment raises the Issues
of whether humans may be exposed to them via air, water or food and, 1f so,
what are the physiological exposures. The transport and fate of the chloro-
benzenes (see Sections 5.1. and 5.2.) are primary determinants of human
exposure to the environmental sources of these compounds, but the more
crucial physiological exposure levels are determined by the ease with which
a compound crosses biological membranes. B1oaecumulat1on 1s a process 1n
which blood and tissue levels of a xenoblotlc, to which there 1s continuous
or repeated exposure, continue to Increase. Usually this phenomenon 1s a
consequence of a slow elimination rate (which Includes excretion and
metabolism) and relatively rapid absorption rate. Ultimate steady state
levels, usually reached when the rate of elimination equals the rate of
uptake or when tissue levels become saturated, will be proportional to the
exposure concentration (as 1n the case of an Inhalation exposure) or, as 1n
the case of Intermittent oral exposure, to the dosing Interval as well.
When such pharmacoklnetlc conditions exist, as they appear to do for the
chlorinated benzenes, the potential for the physiological Insult to be
prolonged beyond the exposure time 1s very great.
The terminology used 1n this section will follow the suggestion of Hacek
et al. (1979): bloconcentratlon Implies that tissue residues result only
from exposure to the ambient environment (I.e., air for terrestrial or water
5-13
-------
for aquatic species); bloaccumulatlon considers all exposures (air, water
and food) of an Individual organism as the source of tissue residues; and
b1omagn1f1cat1on defines the Increase 1n tissue residues observed at suc-
cessively higher trophic levels of a food web.
Tissue concentrations of the various chlorinated benzenes 1n laboratory
and field populations are discussed 1n Chapter 6, Ecological Effects. It 1s
sufficient to state here that the chlorinated benzene Isomers do reach mea-
surable tissue levels 1n exposed organisms. The factors limiting their
accumulation, however, are pertinent for discussion 1n this section.
Studies of the accumulation of xenoblotlcs from environmental sources
Into living cells and tissues have been conducted mainly with aquatic
species and food chains. Under the general experimental design, the organ-
Isms are exposed to sublethal concentrations of the test material under
static or flowing water conditions. After exposure, the concentration of
the test material 1n the organism 1s quantified and a bloconcentratlon
factor (BCF) 1s calculated as the ratio of the concentration 1n tissue (or
the whole organism) to the concentration 1n the water or food; air concen-
tration 1s substituted Into the denominator for calculating the BCF for
terrestrial organisms (Macek et al., 1979; Velth et al., 1980).
From such studies, 1t appears that concentration 1s determined by water
solubility, the octanol/water partition coefficient (Lu and Metcalf, 1975)
or the number of chlorine atoms on the molecule (Barrows et al., 1980). All
three parameters correlate well with the BCF (Kenaga and Goring, 1980;
Metcalf, 1977; Lu and Metcalf, 1975). Table 5-5 shows the direct relation-
ships between Increasing chlorlnatlon, Increasing I1p1d solubility as
Indicated by the octanol/water partition coefficient and the Increase 1n the
5-14
-------
TABLE 5-5
Octanol/Hater Partition Coefficients, Bloconcentratlon Factors
and Biological Half-lives for Chlorinated Benzenes 1n Fish
I
tn
Compound
Honochlorobenzene
1
1
1
1
1
1
1
,2-D1chlorobenzene
,3-01chlorobenrene
,4-01chlorobenzene
,2,4-Trlchlorobenzene
,3,5-Tr1chlorobenzene
,2,3-Trlchlorobenzene
,2,3,5-Tetrachlorobenzene
Octanol/Water
Partition
Coefficient3
690
NR
NR
2,511
2,510
2,754
2,750
2,400
2,344
2,400
2.340
NR
NR
2,400
3,388
MR
17,000
17,000
NR
17,000
10,500
15,000
14,100
15,850
12,900
15,850
28,800
87,100
Species
NS
fathead minnow
rainbow trout
blueglll
rainbow trout
blueglll
rainbow trout
blueglll
blueglll
rainbow trout
rainbow trout
rainbow trout
rainbow trout
trout
guppy
fathead minnow
fathead minnow
green sunflsh
blueglll
rainbow trout
rainbow trout
NS
rainbow trout
guppy
rainbow trout
guppy
blueglll
guppy
BCFb
12
450
46
89
270-560
66
420-740
15
60
214
370-720
32-107
80
231
100
1700
2100
2300
182
890
1300-3200
491
1800-4100
760
1200-2600
700
1800
3900
Biological
Half-11fec
{days}
NR
NR
NR
<1
NR
<1
NR
<7
<1
m
m
<}
NR
NR
0,7
<7
NR
NR
>1<3
NR
NR
NR
NR
1.7
NR
1,5
>2<4
2.5
Reference
Kenaga and Goring, 1980<*
Velth et al., 1979
Branson, 1978
Velth et al., 1980
Oliver and N11m1. 19B3
Velth et al., 1980
Oliver and Nllml, 1983
U.S. EPA, 1980
Velth et al,, 1980
Neely et al., 1974
Oliver and Nllml, 1983
Galassl et al., 1982
CalamaM et al., 1982
U.S. EPA, 1980
Konemann and Van Leeuwen,
1980
Koslan et al., 1981
Velth et al., 1979
Velth et al., 1979
Barrows et al., 1980
Velth et al., 1979
Oliver and Nllml, 1983
Kenaga and Goring, 1980
Oliver and N11m1, 1983
Konemann and Van Leeuwen,
1980
Oliver and N11ro1, 1983
Konemann and Van Leeuwen,
1980
Velth et al., 1980
Konemann and Van Leeuwen,
1980
-------
TABLE 5-5 (cont.)
tn
Compound
1 ,2,4,5-Tetrachlorobenzene
1 ,2,3,4-Tetrachlorobenzene
Pentachlorobenzene
Hexachlorobenzene
Octanol /Water
Partition
Coefficient3
33,100
47,000e
NR
28,800e
NR
87,096
87.1 DO?
154,000
490,000
NR
170,000
170,000
'170,000
169,824
316,0009
168,000
NR
NR
Species
rainbow trout
NS
carp
rainbow trout
carp
blueglll
rainbow trout
NS
guppy
fathead minnow
fathead minnow
green sunflsh
rainbow trout
rainbow trout
rainbow trout
NS
largemouth bass
sheepshead minnow
BCFb
5300-13,000
4500
4000-4900
5200-12,000
3800-4500
3400
13.000-20,000
-5000
14,000
35,000
16.200-18,500
21,900
5500
7762
12,000-20,000
8600
18,214-44,437
20,000
Biological
Half-HfeC
{days}
m
NR
NR
NR
NR
>7
3.8
>7<21
NR
NR
NR
NR
NR
NR
>4<9
NR
Reference
Oliver and N11ra1, 1983
Kenaga and Goring, 1980
Kltano, 1978
Oliver and N11m1, 1983
Kltano, 1978
Velth et a!., 1980
Oliver and N11m1, 1983
Kenaga and Goring, 1980
Koneroann and Van Leeuwen,
1980
Koslan et al , 1981
VeUh et al. 1979
Velth et al. 1979
Velth et al. 1979
Neely et al. 1974
Oliver and Nllml, 1983
Kenaga and Goring, 1980
Laseter et al., 1976
Parrlsh et al., 1978
aDeterm1ned experimentally or by calculation from relative chromatographlc retention time
^Tissue concentration/water concentration (1n flowing water)
C0epurat1on time for tissue concentration to decrease by one-half
dKenaga and Goring (1980) reported these data from various authors; therefore, each entry In a row may be from a different study.
eKonemann et al. (1979)
fBanerjee et al. (1980)
9Ch1ou et al. (1982)
NR = Not reported; NS «. Not specified
-------
BCF for chlorinated benzenes 1n fish. For example for salmon (Oliver and
N1mm1, 1983) all the chlorobenzenes except hexachlorobenzene obeyed:
log BCF = -0.632 + (1.022 ± 0.057) log K at the high exposure end,
and
log BCF = -0.869 + (0.997 +_ 0.056) log K at the low exposure end.
Accordingly, the octanol/water partition coefficient 1s a good first
approximation of the BCF 1n aquatic organisms (r = 0.948, n = 8 1n flowing
water) (Kenaga and Goring, 1980).
B1oaccumulat1on 1n aquatic species 1s a function of the total environ-
mental exposure of the organism Including both water and the food consumed.
Macek et al. (1979) showed, however, that uptake of 1,2,4-tMchlorobenzene
from the ambient water (bloconcentratlon) accounted for 93% of the total
body burden, while diet accounted for 6-7% of the 14C-1,2,4-tr1chloroben-
zene measured 1n bluegllls, Lepomls macrochlrus. after 28 days of exposure.
Similar conclusions were reached by Laseter et al. (1976) using bass and
bluegllls exposed to hexachlorobenzene.
Although the chlorinated benzenes do bloaccumulate and tissue concentra-
tions are established 1n equilibrium with the environment (Kenaga and
Goring, 1980; VeHh et al., 1980; Lu and Metcalf, 1975), the biological
(referring to Individual organisms) and ecological (referring to blomagnlfl-
catlon) persistence of the substance may be the more Important parameter.
The longer biological half-life of persistent compounds 1s most likely a
result of their relative tissue-binding kinetics and the rate of their
blotransformatlon. Ware and West (1977) concluded that halogenatlon of a
compound Increased Its resistance to blotransforraatlon. This, together with
the high affinity for adipose tissue, suggests that the chlorobenzenes are
persistent compounds; this 1s shown for fish 1n Table 5-5.
5-17
-------
The extent of halogenatton also affects the rate of depuration. Gup-
pies, PoeclHa retlculata. were exposed to a standardized mixture of six
chlorobenzenes for 19 days and then allowed to depurate for 9 weeks
(Konemann and Van Leeuwan, 1980). The ambient water concentration of each
chlorobenzene, the BCF and the slope of the elimination curve are shown 1n
Table 5-6. While the chlorobenzenes as a group are persistent, halogenatlon
Influences their rate of elimination.
The significance of biological persistence lies 1n the Increased time of
physiological exposure for an Individual organism and the greater probabil-
ity for human exposure via environmental media.
No studies were available on the bloaccumulatlon of the chlorobenzenes
1n terrestrial food webs. There are, however, no Immediately apparent
reasons why the relationships between bloaccumulatlon and the phys1cochem1~
cal parameters demonstrated for aquatic systems are not applicable to the
terrestrial environment. Generally, for air-breathing terrestrial species
such as humans, the atmospheric concentration of a compound 1s the primary
determinant of bloaccumulatlon because the frequency of air Intake 1s much
greater compared to the 1ngest1on of food or water. This was apparent from
the analysis of ambient air and household tapwater samples taken from nine
homes 1n the Love Canal area of Niagara Falls, New York (Barkley et al,,
1980). From these data, the expected total dally Intake of dlchloroben-
zenes by a 70 kg adult male for example, was nearly 300-fold greater from
air (0.119 mg/day) than from tapwater (3.36xlO~4 mg/day). This topic Is
discussed more extensively 1n Sections 4.3. and 4.4.
Although the chlorobenzenes are volatile compounds and Inhalation 1s the
expected primary route of human exposure, potentially high Intake by other
routes cannot be Ignored. Therefore, bloaccumulatlon and Internal exposure
5-18
-------
TABLE 5-6
B1oconcentrat1on Factor and Slope of the Elimination Curve for
Guppies (Pgec111 a retlculata) Exposed to Six Chlorinated Benzenes3
Compound
1 ,4-01chlorobenzene
1 ,2,3-Tr1ehlorobenzene
1 ,3,5-Tr1chlorobenzene
1 ,2,3,5-Tetrachlorobenzene
Pentachlorobenzene
Hexachlorobenzene
Average Concentra-
tion Measured 1n
Water (ng/ms.)
116
48
43
12
1.2
0.3
BCfb
1.8x10*
1.3xl04
1.4x10*
7.2x10*
2.6x10=
2.9X105
Slope of
Elimination
Curve (day"1)0
1.00
0.45
0.40
0.28
0.18
0.062
aSource: Konemann and Van Leeuwan, 1980
^Calculated on the basis of fat weight (average fat weight = 5.4%)
C0nly 1,4-dl- and hexachlorobenzene had single-phase elimination curves;
the second-phase slopes for the other compounds are excluded for simplicity.
5-19
-------
are multlfactorial parameters dependent upon the chlorinated benzene concen-
tration 1n each of three environmental compartments and upon the relative
rate of absorption and elimination for each compound.
5.4. SUMMARY
The chlorinated benzenes are a group of volatile compounds the lower
chlorinated members of which readily evaporate to the atmosphere from soil
and water. Point source releases of the chlorinated benzenes are readily
carried by prevailing winds and may be the primary source of measurable
hexachlorobenzene in Industrially remote areas, although there also may be
concentration gradients around these point sources. The high vapor pressure
and low water solubility of these compounds promotes their release to the
atmosphere from open water systems or their association with organic
material that may either be Incorporated Into sediments or flow out of the
system. Soils, depending on their type, readily allow the evaporation of
chlorobenzenes from pore spaces to the atmosphere, or, depending on the
relative affinity of the compound, release 1t as leachate.
Little Information 1s available on the fate of the chlorinated benzenes
1n air, but one study concluded that the atmospheric residence time In-
creased with an Increase 1n chlorine substltuents. Laboratory studies with
smog chambers suggests photocatalysls may produce nitrobenzene, and nltro-
phenol or polychlorlnated blphenyls (DHUng et al., 1976; Kanno and No^lma,
1979; Uyeta et al., 1976). The fate of the chlorobenzenes 1n water and soil
are similar, but the rates differ for each process (I.e., blodegradatlon,
loss to the atmosphere, accumulation 1n the biota, physical removal by
outflow or leaching, or sequestration of the unaltered compound).
The chlorobenzenes are UpophlUc compounds that bloaccumulate 1n animal
and human tissues after uptake from ambient air, water and food. The BCF
5-20
-------
(tissue concentration/media concentration) Is an Indicator of bloaccumula-
tlon and 1s determined by physlochemlcal parameters such as the water solu-
bility, the octanol/water partition coefficient and the number of substl-
tuent chlorine atoms (Kenaga and Goring, 1980). Physiological exposure
levels are determined by absorption, distribution, metabolism, elimination,
and storage In adipose tissue; thus, biologically persistent compounds, such
as the chlorobenzenes, produce prolonged physiological exposures.
5-21
-------
6. ECOLOGICAL EFFECTS
As mentioned briefly 1n the previous chapters, chlorinated benzenes
occur 1n both the aquatic and terrestrial environments. The concentrations
of these compounds 1n some areas suggest that wildlife may be exposed to
higher levels of chlorinated benzenes than those encountered by humans.
Although aquatic and terrestrial organisms are exposed, no data are avail-
able on the toxic effects of chlorobenzenes at environmental concentrations
1n natural populations. Laboratory testing has shown that chlorobenzenes
have toxic effects on aquatic and terrestrial species and bloaccumulate 1n
exposed organisms.
6.1. EFFECTS ON THE AQUATIC ENVIRONMENT
Data on the effects of chlorinated benzenes on freshwater or marine
organisms 1n their natural environment were not available. Chlorobenzenes
have been shown to be acutely toxic to aquatic species 1n laboratory bio-
assays. The results of such acute toxldty bloassays can be used to deter-
mine relative tox1c1t1es of the various chlorobenzenes to various species.
6.1.1. Effect on Freshwater and Marine Fish, The acute toxldty of mono-
chlorobenzene has been reported for several species of freshwater and marine
fish (Table 6-1). The most sensitive species appears to be the rainbow
trout, Salmo galrdnerl. with 96-hour median lethal concentration (LC,__)
, ,jy
values ranging between 3-5 mg monochlorobenzene/st (Brosler, 1972; Dow
Chemical Company, 1978b; Calaraarl et a!., 1983; Dallch et a!,, 1982). Blue-
gill sunflsh (Lepprols macrochlrus), fathead minnows (Plmephales promelas)
and gupples (Leblstes retlculatus) were moderately tolerant with mean
96-hour LC values ranging from 15.9-24.0, 31.5-33.9 and 45.5 mg/8.,
respectively (U.S. EPA, 1978; Pickering and Henderson, 1966). The goldfish,
Carasslus aurajus, was the species most tolerant of monochlorobenzene with a
96-hour LCcr. value of 51.62 mg/8, (Pickering and Henderson, 1966).
3U
6-1
-------
TABLE 6-1
Acute Toxlclty Data for Fish Species Exposed to Chlorinated Benzenes
I
r\>
Compound Species Duration
(hour)
Honochlorobenzene rainbow trout 96
(Salmo galrdnerl)
24
8
96
96
48
blueglll sunflsh 24
(Lepomls nacrochlrus) 48
72
96
96
24
48
96
24
96
fathead minnows 24
(Plmephales promelasj
48
96
24
48
96
Mean
Concentration
(mg/l)
3.58
1.8
5-10
3-5
4.7
4.ia
16.9
15.9
15.9
15.9
7.80
24.00
24.00
24.00
17.0
16.0
31.53
31.53
31.53
39.19
34.98
33.93
Method
constant-flow
NR
static
static
constant-flow
IRSA
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
Effect
"50
"50
LClOO
"50
"50
L050
"50
"50
"50
"50
None
"50>>
LC50b
LC50b
"50
"50
LC50b
LC50b
LC50b
LC50c
LC50c
LC50c
Reference
Dow Chemical Co.,
1978b
Glngerlch and
Dallch, 1978
Brosler, 1972
Brosler, 1972
Dallch et al.,
1982
Calamarl et al.,
1983
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Buccafusco et al.,
1981
Buccafusco et al.,
1981
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
-------
TABLE 6-1 {cont.)
er>
i
to
Compound Species
Honochlorobenzene (cont,) goldfish
{Carasslus ayratys)
gupples
(leblstes retlculatus)
sheepshead minnow
(Cyprlnodon varlegatus)
Brachydanlo rerlo
l,2-D1chlorobenzenc rainbow trout
(Sal mo galrdneM )
blueglll sunflsh
(Lepopls macrochlrus)
fathead minnow
(Plmephales promelas)
Duration
(hour)
24
48
96
24
48
96
24
48
96
96
48
96
48
24
48
72
96
96
24
96
96d
96
48
96
Mean
Concentration
(mg/l)
73.03
56.00
51.62
45.53
45.53
45.53
>19.9
8.94
10.50
6.20
10. 5a
1.67
2.33
6.26
6.06
5.59
5.59
<3.20
6.3
5.6
27.0
57.0
76.3
57.0
Method
static
static
static
static
static
static
static
static
static
static
IRSA
constant-flow
IRSA
static
static
static
static
static
static
static
static
static
static
static
Effect
LC50b
LC50b
LC50b
LCSO
LCSO
LC50
USD
LC50
LC50
None
LCso
LCSO
"50
LC50
LC5Q
LC50
"SO
None
LC50
LC50
LC50
LC50
"SO
LC50
Reference
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
Pickering and
Henderson, 1966
U.S. EPA, 1978;
Heltmuller et al,,
1981
CalamaM et al.,
1983
Dow Chemical Co.,
1978b
CalamaH et al.,
1983
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Buccafusco et al..
1981
Buccafusco et al. ,
1981
Dawson et al. , 1977
Curtis and Ward,
1981
Curtis et al.,
1979
Curtis et al.,
1979
-------
TABU 6-1 Jcont.)
ar»
i
Compound Species
1,2-Dlchlorobenzene (cent.) tidewater silverslde
(Henldla berylllna)
sheepshead minnow
(Cyprlnodon varlegatus)
Brachydanlo rerlo
1,3-Dlchlorobenzene blueglll sunflsh
(Lepomis macrochlrus)
Duration
(hour)
96
-------
TABLE 6-1 (cent.)
IT*
I
Compound Species
l,4-D1ch1orobenzene (cont.) fathead minnow
(Plmephales promelas)
sheepshead minnow
(Cyprlnodon varlegatus)
rainbow trout
(SaTmo galrdnerl)
Br achy dan 1o rerlo.
1,2,3-Trlchlorobenzene rainbow trout
(Salmo galrdnerD
Brachydanlo rerlo
1,2,4-Trlchlorobenzene rainbow trout
(Salmo galrdnerl)
Brachydanlo rerlo
blueglll sunflsh
(Lepomls macrochlrus)
sheepshead minnow
(Cyprlnodon varlegatus)
Duration
(hour)
96
24
48
96
24
48
12
96
96
48
48
48
48
48
48
24
48
72
96
96
24
96
24
48
72
96
96
Mean
Concentration
(mg/lj
30.0
35.4
35.4
33.7
7.5-10.0
7.17
7.40
7.40
5.6
1.183
4.25a
0.713
3.1«
1.95a
6.3a
109.0
13.0
3.59
3.36
<1.70
109.0
3.4
>46.8
>46.8
>46.8
21.4
14.6
Method
static
static
static
static
static
static
statk
static
static
IRSA
IRS A
IRSA
IRSA
IRSA
IRSA
static
static
static
static
static
static
static
static
static
static
static
static
Effect
LC50
LC50
LC50
LCSO
LC50
LC-50
LC50
LC50
None
LC50
LC50
LC50
LC50
LC50
LC50
LC50
LC50
LC50
LC50
None
LC50
"50
LC50
LC50
LC50
LC50
None
Reference
Curtis and yard.
1981
Curtis et al., 1979
Curtis et al., 1979
Curtis et al., 1979
U.S. EPA, 1978;
Heltmuller et al. ,
1981
Calaroarl et al..
1983
Calamarl et al.,
1983
Calamarl et al..
1983
Calamarl et al,,
1983
Calamarl et al. ,
1983
Calamarl et al. ,
1983
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Buccafusco et al.,
1981
Buccafusco et al. ,
1981
U.S. EPA, 1978;
Heltrauller et al.,
1981
-------
TABLE 6-1 (cont.)
Compound
1 ,2,3,5-Tetrachlorobenzene
1 ,2,4,5-Tetrachlorobenzene
Pentachlorobenzene
Species
bluegUl sunflsh
(Lepoiiils macrochlrus)
sheepshead minnow
(Cyprlnodon varleqatus)
blueflll sunflsh
(Leponils macrochlrus)
sheepshead minnow
(Cyprlnodon varleqatus)
blueglll sunflsh
(Lepomls macrochlrus)
Duration
(hour)
24
48
72
96
96
24
96
24
48
72
96
96
24
48
72
96
96
24
96
24
48
72
96
96
96
24
48
72
96
96
24
Kean
Concentration
57.8
11.5
8.34
6,42
<1.70
59.0
6.4
>7.5
5.59
4.68
3.67
1.0
5.69
4.35
1.55
1.55
0.6B
5.7
1.6
>1.80
0.90
0.84
0.84
0.32
0.33
2.27
0.55
0.30
0.25
<0.088
2.30
Method
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
flowthrough
static
static
static
static
static
static
Effect
"50
"50
"50
"50
None
"so
"50
"50
"SO
"50
"SO
None
"50
"so
"50
"50
None
"50
"50
"50
"50
"50
"JO
None
"50
"50
"so
"50
None
"50
Reference
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Buccafusco et al.,
1981
Buccafusco et al.,
1981
U.S. EPA, 1978;
Heltmuller et al..
1981
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Buccafusco et al.,
1981
Buccafusco et al.,
1981
U.S. EPA, 1978;
Heltrauller et al..
1981
Hard et al., 1981
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Buccafusco et al.,
96
0.25
static
1981
Buccafusco et al.,
1981
-------
TABLE 6-1 (cont.)
i
•••j
Compound
Pentachlorobenzene (cont.)
Hexachlorobenzene
Species
sheepshead minnow
(Cyprlnodon varlegatus)
largemouth bass
(Hlcropterus salmqldes)
sheepshead minnow
(Cyprlnodon varleqatus)
plnflsh
(Lagodon rhomboldes)
rainbow trout
(Sajmo galrdnerl)
Brachydanlo rerlo
Duration
( hour )
24
48
72
96
96
240
360
96
96
48
48
Mean
Concentration
(mg/l)
>32.0
9.55
3.2-10.0
0.83
0.32
0.009-0.01
0.022-0.026
0.13
l.fle
<0.03a
<0.03a
Method
static
static
static
static
static
static
static
constant-flow
constant-flow
IRS A
IRSA
Effect
LC50
LCso
LC50
None
None
None
None
None
LC50
LC50
Reference
U.S. EPA, 1978
Heltmuller et
1981
Laska et al. ,
Laska et al. ,
Parrlsh et al.
1974
Parrlsh et al.
1974
Calamarl et al
1983
Calamarl et al
1983
.
al.,
1978
1978
»
t
* I
* 1
aSoft water conditions: pH « 7.4; hardness = 320 mg CaC03/l; oxygen = >7QX; temperature = 15°C for trout and 23°C for Brachydanlo
&Soft water conditions: pH = 7.5; alkalinity = 18 mg/l; hardness * 20 rng/1
cHard water conditions: pH = 8.2; alkalinity = 300 mg/1; hardness = 350 mg/i
^Estimated based on 24, 48, 72 and 96-hour toxlclty tests
eNom1nal concentration; because of solubility, actual concentration would be less
NR = Not reported
-------
The marine sheepshead minnow, Cyprlnodon varlegatus, was relatively sensi-
tive with a 96-hour LC value of 10.5 mg/si (U.S. EPA, 1978; Heltmuller
et al., 1981).
The acute toxlclty of 1,2-d1chlorobenzene was studied 1n several fresh-
water and marine fish (see Table 6-1). Rainbow trout, S_. galrdnerl. was the
most sensitive species reported with an LC value of 1.67 mg/fi, follow-
ing a 96-hour exposure (Dow Chemical Company, 1978b). The U.S. EPA (1978)
and Buccafusco et al. (1981) reported 96-hour LC values near 5.6 mg/8.
for the blueglll sunflsh, L_. macrochlrus. while Dawson et al. (1977)
reported a value of 27.0 mg/8. for this species. The fathead minnow, P_.
promelas, was the most resistant species tested, having a 96-hour LC,._
value of 57.0 mg/8. (Curtis et al., 1979; Curtis and Ward, 1981). Two
marine species, the tidewater sllverslde (Hen1d1a perylUna) and the sheeps-
head minnow (C_. varlegatus), were moderately sensitive with 96-hour LC
values of 7.3 and 9.7 mg/a., respectively (Dawson et al., 1977; U.S. EPA,
1978; Heltmuller et al., 1981).
The 1,3- Isomer of dlchlorobenzene was tested for acute toxldty 1n two
species of freshwater fish and a single marine species. The 24, 48, 72 and
96-hour LC_. values for blueglll sunflsh, L. macrochlrus. were 21.8, 10.7,
bU
5.02 and 5.02 mg 1,3-d1chlorobenzene/B., respectively (U.S. EPA, 1978;
Buccafusco et al., 1981). The no-observed-effect level (NOEL) was 1.7
mg/a 1n the blueglll (U.S. EPA, 1978). The fathead minnow, P. promelas.
had a static 96-hour LC value of 12.7 mg 1,3-d1chlorobenzene/fi. (Curtis
and Ward, 1981). In the marine species, sheepshead minnow (C. varlegatus).
24, 48, 72 and 96-hour LC values were 8.46, 8.04, 8.04 and 7.77 mg/a,
respectively. The NOEL was 4.18 mg/a (U.S. EPA, 1978; HeUmuller et al.,
1981).
6-8
-------
Rainbow trout, blueglll sunflsh, fathead minnows and sheepshead minnows
were the species tested to study the static acute toxlclty of 1,4-d1chloro-
benzene. Rainbow trout, S. galrdnerl. was the most sensitive species
tested, with 48-hour LC values of 1.18 mg/8, (Calamarl et al., 1983).
The bluegUl sunflsh (L. macrochlrus) showed 24, 48, 72 and 96-hour LC5Q
values of 4.54, 4.37, 4.37 and 4.28 mg/a, (U.S. EPA, 1978; Buccafusco et
al., 1981). The NOEL for this species was reported to be <2.8 mg 1,4-d1-
chlorobenzene/8, (U.S. EPA, 1978). The 24, 48, and 96-hour static LCgo
values for fathead minnows (P_. promelas) were 35.4, 35.4 and 33.7 mg/8,,
respectively (Curtis et al., 1979). The marine sheepshead minnow, C_.
varlegatus. was Intermediate 1n sensitivity to 1,4-d1chlorobenzene, having a
96-hour LC of 7.4 mg/a, and a NOEL of 5.6 mg/a (U.S. EPA, 1978;
HeHmuller et al., 1981).
1,2,4-Tr1ch1orobenzene has been tested for acute toxlclty to fish
species. The 48-hour LC value for rainbow trout, £. galrdnerl, was 1.95
mg/a (CalamaM et al., 1983). In the blueglll sunflsh (L_. macrochlrus)
estimated LC5Qs, based on nominal concentrations, were reported for 24,
48, 72 and 96-hour exposures at 109.0, 13.0, 3.59 and 3.36 mg 1,2,4-tM-
chlorobenzene/a (U.S. EPA, 1978; Buccafusco et al., 1981). The NOEL was
<1.7 mg/8, for the sunflsh. The sheepshead minnow, C_. varlegatus. was more
tolerant with 24, 48 and 72-hour LC values >46.8 mg/a, and the 96-hour
LC50 value of 21.4 mg/a. The NOEL for this marine species was 14.6
mg/8, (U.S. EPA, 1978; HeHmuller et al., 1981). For 1,2,3-tMchloroben-
zene, rainbow trout, S. galrdnerl. showed a 48-hour LCrn value of 0.71
~ bU
mg/a, (Calamarl et al., 1983), and 1s thus more aquatlcally toxic than the
1,2,4- Isomer. The correspor
mg/a (Calamarl et al., 1983).
1,2,4- Isomer. The corresponding LC value for Brachydanlo rerlo was 3.1
6-9
-------
The toxlclty of only 1,2,3,5- and 1,2,4,5-tetrachlorobenzene has been
tested In fish. These two Isomers differ dramatically 1n their lethality to
blueglll sunflsh and sheepshead minnows. The 24, 48, 72 and 96-hour LC,.
values for the 1,2,3,5- Isomer 1n bluegllls (L_. macrochlrus) and sheepshead
minnows (C_. yarlegatus) were 57.8, 11.5, 8.34 and 6.42 mg/H and >7.5,
5.59, 4.68 and 3.67 mg/£, respectively (U.S. EPA, 1978; Buccafusco et al.,
1981; Heltmuller et al.f 1981; Ward et al., 1981). The NQELs for the
blueglll and sheepshead minnow were <1.70 and 1.0 mg 1,2,3,5-tetrachloro-
benzene/a, respectively. The 1,2,4,5- Isomer was, 1n some cases, 10-11
times more lethal to the fish species tested. For example, the 24, 48, 72
and 96-hour LC^ values 1n the blueglll sunflsh were 5.69, 4.35, 1.55 and
1.55 rag/8,. In the sheepshead minnow, the LCgo values ranged from
>1.80-0.33 for 24 through 96-hour exposures (see Table 6-1 for comparison)
{U.S. EPA, 1978; Heltmuller et al., 1981; Ward et al., 1981). NOELs for the
1,2,4,5- Isomer were reported to be 0.68 and 0.32 mg/SL for bluegUl
sunflsh and sheepshead minnows, respectively.
The acute toxldty of pentachlorobenzene was studied 1n the freshwater
blueglll sunflsh and the marine sheepshead minnow (U.S. EPA, 1978; Bucca-
fusco et al., 1981; Heltmuller et al., 1981). The static LCgo values for
24, 48, 72 and 96-hour exposures were 2.27, 0.55, 0.30 and 0.25 mg/8, for
the blueglll sunflsh (L. macrochlrus) and >32.0, 9.55, 3.2-10.0 and 0.83
mg/fi. for the sheepshead minnows (£. varlegatus). NOELs for blueglll sun-
fish and sheepshead minnows were <0.088 and 0.32 mg pentachlorobenzene/8,,
respectively.
Because of the low water solubility of hexachlorobenzene, acute toxlclty
testing of this compound has been conducted at low concentration levels
6-10
-------
only, Largemouth black bass, Mlcropterus salmoldes. exposed for 10 days at
9-10 yg/8. or exposed for 15 days at 22-26 yg/a, showed no toxic
effects (Laska et a!., 1978). Sheepshead minnows, £. yarlegatus, exposed at
0.13 mg/H and plnflsh, Lagodon rhomboldes, exposed to a nominal concentra-
tion of 1.0 mg/8. (actual concentration would be less because of low
aqueous solubility) for a 96-hour period showed no toxic effects (Parrlsh et
a!., 1974). But rainbow trout, £. galrdnerl, and Brachydanlo rerlo showed
48-hour LC50 values of <0.03 mg/a (Calamarl et a!., 1983).
Subchronlc toxlclty testing has been conducted on monochlorobenzene 1n
rainbow trout, S. galrdnerl (Dallch et al., 1982). Groups of fish were
exposed to 2.1 or 2.9 mg monochlorobenzene/a for 15 or 30 days. Treated
fish did not accept food during at least the first 15 days of treatment.
Neither concentration of monochlorobenzene resulted 1n any deaths during the
exposure periods, but loss of equilibrium was reported 1n most treated fish.
Liver toxlcity, determined by enzyme levels, and hlstologlcal hepatic
changes were reported 1n trout treated at both exposure levels (Dallch et
al., 1982).
Studies conducted by the U.S. EPA (1978, 1980a) resulted 1n chronic
toxlclty values (NOELs) for many of the chlorinated benzenes 1n fathead
minnows and/or sheepshead minnows (Table 6-2).
During bloaccumulatlon testing with the blueglll sunflsh, L_. macro-
chlrus, fish were exposed to 1,2-d1chlorobenzene (7.89 yg/a), l,3-d1~
chlorobenzene (107.0 wg/a) and 1,4-d1chlorobenzene (10.1 yg/fc) for
14 days. Similarly, 1,2,4-trlchlorobenzene (2.87 yg/a), 1,2,3,5-tetra-
chlorobenzene (7.72 yg/a) and pentachlorobenzene (5.15 yg/St) were
tested for 28 days 1n the blueglll. No deaths or toxic effects were
reported for any of the chlorinated benzenes at the exposure levels tested
(Barrows et al., 1980).
6-11
-------
TABLE 6-2
Chronic Toxldty Values of Chlorinated Benzenes 1n F1sh
Chemical
l,2-D1chlorobenzene
1 ,3-D1chlorobenzene
T 1 ,4-D1chlorobenzene
t>o
1 ,2,4-Tr1chlorobenzene
1 ,2,3,4-Tetrachlorobenzene
1 ,2,4,5-Tetrachlorobenzene
Species
fathead minnow
(Plmephales promelas)
fathead minnow
(Plmephales promelas)
fathead minnow
(Plmephales promelas)
fathead minnow
(Plmephales promelas)
sheepshead minnow
(Cyprlnodon vaMegatus)
fathead minnow
(Plmephales promelas)
sheepshead minnow
(Cyprlnodon varlegatus)
Chronic Value*
2000
1510
763
286
705
222
318
129
Range
1600-2500
1000-2270
560-1040
200 410
499-995
150-330
245-412
92-180
Reference
U
U
U
U
U
U
U
U
.S.
.S.
.S.
.S.
.S.
.S.
.S.
.S.
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
1978
1980a
1980a
1978
1980a
1978
1980a
1978
*NOELs
-------
Limited data are available on the pharmacoklnetlcs of chlorinated
benzenes 1n fish. Uptake of 1,2,4-trlchlorobenzene from the water {0.012
mg/a) was rapid 1n the rainbow trout, $>. galrdnerl. with bile and liver
concentrations exceeding 100 times the water levels within hours (Melancon
and Lech, 1980). N11m1 and Cho (1980) reported that rainbow trout absorbed
and accumulated hexachlorobenzene from their diet and body levels could
Increase 15 pg/kg body weight/day In a dose-dependent manner. Later,
Oliver and N11m1 (1983) reported evidence Indicating that all chlorinated
benzenes studied (!,2-d1, l,3-d1, l,4-d1, l,3,5-tr1, l,2,4-tr1, !,2,3-tr1,
1,2,4,5-tetra, 1,2,3,4-tetra, penta- and hexachlorobenzene) could be
absorbed from the aqueous environment. Z1tko and Hutzlnger (1976) reported
the uptake and accumulation of hexachlorobenzene from food or water 1n
juvenile Atlantic salmon, Salmo salar.
Monochlorobenzene seems to be metabolized by the liver since liver
toxldty, Including degeneration of hepatocytes and necrosis, was reported
In treated rainbow trout (GlngeMch and Dallch, 1978). A modeling study by
Lu and Metcalf (1975) suggested that chlorobenzene 1s metabolized to o- and
p-chlorophenol 1n the mosquito fish, Gambusla affinis.
Studies on the metabolism and b1otransformat1on of 1,2,4-trlchloro-
benzene 1n rainbow trout (S. galrdnerl) and carp (Cyprlnus carplo) suggested
that conjugated metabolites occur In the liver and bile (Melancon and Lech,
1980). A hepatic mixed-function oxldase Inducer (p-naphthoflavone)
elevated the hepatic and biliary levels of blotransformatlon products of
1,2,4-tr1chlorobenzene. In the mosquito fish, G. aff1n1s. absorbed hexa-
chlorobenzene 1s predominantly unchanged, but two unidentified metabolites
were reported (Lu and Metcalf, 1975).
6-13
-------
Accumulated chlorinated benzenes and/or their metabolites seem to be
distributed throughout the body 1n fish. The highest concentrations have
been detected 1n the bile, liver and muscle (Melancon and Lech, 1980). The
bloconcentratlon of chlorinated benzenes Increased as the degree of chlorl-
natlon of the test compound Increased (Oliver and N11m1, 1983). Bloconcen-
tratlon factors {BCFs) for many of the chlorinated benzenes 1n gupples
(Poedlla retlculata) and rainbow trout (S. galrdnerl) are shown 1n Table
6-3 (Konemann and van Leeuwen, 1980; Oliver and N11m1, 1983). More complete
data on BCFs 1n fish are reported 1n Section 5.3 of this document.
The excretion rate of chlorinated benzenes 1n fish Is related to the
extent of chlorlnatlon of the compound. Konemann and van Leeuwen (1980)
reported that 1,4-d1chlorobenzene 1s excreted within several days, while
trlchlorobenzenes required nearly 25 days, tetrachlorobenzene nearly 50
days, and penta- and hexachlorobenzene required >50 days for elimination.
After termination of exposure, 1,2,4-tr1chlorobenzene and metabolites are
eliminated In two stages. The first had a half-life of elimination of 0.4
days, while the second was eliminated more slowly (t,/2 = 50 days). In
comparison, Branson et al. (1975) reported half-lives for elimination of
dlchlorobenzene and hexachlorobenzene to be 1.1 and 12.1 days, respectively.
Sanborn et al. (1977) estimated the half-life for elimination of hexachloro-
benzene 1n the green sunflsh, L_. cyanellus, to be 8.0-19.6 days. The
longest time was for elimination from the liver. The biological half-life
of hexachlorobenzene was estimated to be from 7 months to several years 1n
rainbow trout (N11m1 and Cho, 1981).
6.1.2. Effect on Aquatic Crustaceans. In addition to fish, freshwater
and marine crustaceans, which are an Important element 1n aquatic food
chains, are exposed to chlorobenzenes 1n the environment (Grzenda et al.,
6-14
-------
TABLE 6-3
Bloconcentratlon Factors of Some Chlorinated Benzenes In Two F1sh Species
Species Compound
Rainbow trout l,2-d1-
Salmo galrdnerl
1,3-dl-
1,4-dl-
l,3,5-tr1~
l,2,4-tr1-
l,2,3-tr1~
1,2,4,5-tetra-
1,2,3,4-tetra-
penta-
hexa-
Guppy l,4-d1-
PoeclHa retlculata l.2,3-tr1-
l,3,5-tr1-
1,2,3,5-tetra-
penta-
hexa-
Exposure
Level
(pg/a)
47.0
940.0
28.0
690.0
28.0
670.0
2.3
45.0
3.2
52.0
4.3
72.0
1.0
21.0
1.4
26.0
0.34
9.3
0.32
8.0
116.0
48.0
43.0
12.0
1.2
0.3
BCF Reference
270 Oliver and N11m1,
560 1983
420
740
370
720
1,800
4,100
1,300
3,200
1,200
2,600
5,300
13,000
5,200
12,000
13,000
20,000
12,000
20,000
1,800 Konemann and
13,000 van Leeuwen, 1980
14,000
72,000
260,000
290,000
6-15
-------
1964), Laboratory testing of the chlorinated benzenes has provided acute
toxldty data for several species of crustaceans (Table 6-4).
The U.S. EPA (1978) reported most of the available data 1n which mono-,
l,2-d1-, l,3-d1-, l,4-d1-, 1,2,4-trl-, 1,2,3,5-tetra-, 1,2,4,5-tetra- and
pentachlorobenzene tox1cH1es were tested 1n the water flea (Daphnia magna)
and the mysld shrimp (Hys1dops1s bahla). Other available studies on
specific chlorinated benzenes tested 1n specific species were noted 1n Table
6-2. Generally, the more chlorinated benzenes appear to be more toxic. For
example, the 96-hour LC,-0 values 1n mysld shrimp were 16.4, 1.97, 0.34 and
0.16 mg/a, for mono-, I,2~d1-, 1,2,3,5-tetra- and pentachlorobenzene,
respectively. Data on the toxldty of the 1,2,3,5- and 1,2,4,5-tetrachloro-
benzenes Indicate that crustaceans, unlike that 1n fish, the 1,2,3,5- Isomer
1s more toxic. Because of the very low solubility of hexachlorobenzene 1n
aqueous solutions, data on the toxldty of this compound are limited. One
study (Laska et al., 1978) reported no toxic effects 1n crayfish, Procam-
barus c1ark11. exposed (unspecified Interval) to a saturated aqueous solu-
tion of hexachlorobenzene (estimated to be -0.02 mg/a). The 24-hour
Immobilization concentrations of several chlorobenzenes for water fleas,
Daphnia magna, using the AFNOR test were: monochlorobenzene (4.3 mg/si);
1,2-d1chlorobenzene (0.78 mg/a.}; 1,4-d1chlorobenzene (<0.03 mg/t)
(Calamarl et al., 1983).
6.1.3. Embryotoxlc and Reproductive Effects. Wild Atlantic salmon (Salmo
salar) eggs, collected from different sites, contained different levels of
hexachlorobenzene (0.086, 0.132, 0.142 and 0.159 jig/g I1p1d 1n eggs). No
correlation between hexachlorobenzene levels and egg-hatchabH1ty was demon-
strated (Z1tko and Saunders, 1979). Eggs also contained other environmental
contaminants such as PCBs and organochlorlde pesticides.
6-16
-------
TABLE 6-4
Acute Toxldty Data for Crustaceans Exposed to Chlorinated Benzenes
Compound
I
_J
~J
Species
Mean
Duration Concentration
(hour) (mq/t)
Method
Effect
Reference
nochlorobenzene water flea
(Oaphnla roagna)
rnysld shrimp
(Hysldopsls bah la)
?-D1ch1orobenzene water flea
(Daphnla magna)
mysld shrimp
(Hysldopsls pallia)
grass shrimp
(Palaeroonetes puglo)
i-D1chlorobenzene water flea
(Daphnla magna)
mysld shrimp
(Hysldopsls bahja.)
24
48
48
24
24
48
72
96
96
24
48
48
24
24
48
72
96
96
24
48
96
96
24
48
48
24
48
72
96
96
140.0
86.0
10.0
4.3
24.7
24.7
24.7
16.4
<11.1
2.44
2.44
0.36
0.78
4.75
4.52
3.88
1.97
<1.29
14.3
10.3
9.4
10.4
47.8
28.1
6.0
7.31-13.06
5.14
4.06
2.85
<1.30
static
static
static
AFNOR
static
static
static
static
static
static
static
static
AFNOR
^
&K
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
LC50
LC50
None
1^50
LC50
LC50
LC50
LC50
None
LC50
LC50
None
"50
"-C50
LC50
LC50
None
LC5Q
LC50
LC50
LC50
LC50
I-C50
None
LCso
LC50
LC50
LC50
None
U.S. EPA, 1978;
LeBlanc, 1980
Calatnarl et al..
1983
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978;
LeBlanc, 1980
Calamarl et al..
1983
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA. 1978
Curtis et al.,
1979
Curtis et al.,
1979
Curtis et al..
1979
Curtis and Mard,
1981
U.S. EPA, 1978;
LeBlanc, 1980
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S: EPA, 1978
-------
TABLE 6-4 {cont.J
cr>
co
Compound Species
1,4-Dlchlorobenzene water flea
(Oaphnla magna )
mysld shrimp
(Hysldopsls bahla)
grass shrimp
(Palaemonetes puqlo)
1,2,3-Trlchlorobenzene water flea
(Oaphnla magna)
1,2,4-Trlchlorobenzene water flea
(Oaphnla magna)
mysld shrimp
(Hysldopsls bahla)
1,3,5-Trlchlorobenzene brine shrimp
(Artemla sauna)
1 ,2,3,5-Tttrachlorobenzene water flea
(Oaphnla magna)
mysld shrimp
(Hysldopsls bahla)
Duration
(hour}
24
48
48
24
24
48
72
96
96
48
96
96
24
24
48
48
24
24
48
72
96
96
168
24
48
48
24
48
72
96
96
Hean
Concentration
(mg/l)
41.5
• 11.0
0.68
1.6
5.6-10.0
5.35
4.31
1.99
<1.0
129.2
69.0
60.0
0.35
114.0
50.2
<2.4
1.2
>1.46
>1.46
0.76
0.45
0.09
10.0
18.1
9.71
<1.1
0.96
0.36
0.34
0.34
0.10
Method
statk
statk
static
AFNOR
static
static
statk
statk
statk
statk
statk
statk
AFNOR
statk
statk
statk
AFNOR
static
statk
statk
statk
statk
statk
statk
static
statk
statk
statk
static
statk
statk
Effect
LC50
'-'•50
None
ICso
"50
"50
None
LCSO
'-''50
"50
ic50
"50
None
50
"50
"50
50
50
None
"100
LC50
"50
None
"50
"SO
"50
LC50
None
Reference
U.S. EPA, 1978;
LeBlanc, 1980
Calawarl et al.,
1983
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Curtis et al.,
1979
Curtis et al.,
1979
Curtis and Ward,
1981
Calamarl et al.,
1983
U.S. EPA, 1978;
LeBlanc, 1980
Calamarl et al.,
1983
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Srosch, 1973
U.S. EPA, 1978;
LeBlanc. 1980
U.S. EPA, 1978
U.S. EPA, 1978
U.Sf EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
-------
TABLE 6-4 (cont.)
Compound Species
1,2,4,5-Tetrachlorobenzene water flea
(Daphnla magna)
mysld shrimp
(Hys1dops1s bahja.)
Pentachlorobenzene Mater flea
(Daphnla magna)
*r>
i
»*» rays Id shrimp
(Hys1dops1s bahla)
Hexachlorobenzene water flea
(Daphnla magna)
swamp crayfish
(Procambarus dark 11.)
! shrimp
i (Crangon sept ems plnosa)
Duration
(hour)
24
48
48
24
48
72
96
96
24
48
48
24
48
72
96
96
24
NR*
96
Mean
Concentration
(mg/l)
>530.0
>530.0
320.0
3.2-5.6
1,99
1,48
1.48
0.6
17.2
5.28
1.3
0.75
0.72
0.24
0.16
<0.06
<0.03
saturated*
0.0072
Method
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
AFNOR
static or
f lowthrough*
static
Effect
LC50
LC50
None
LC50
LC50
"SO
LC50
None
LC50
LC50
None
LC50
LC50
LC50
LC50
None
icso
No toxic
effects
No mortality
Reference
U.S. EPA, 1978;
LeBlanc, 1980
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978;
LeBlanc. 1980
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
CalamaH et al.,
1983
Laska et al.,
1978
HcLeese and
Metcalfe, 1980
*ToKldty testing was conducted for an unspecified period with a saturated aqueous solution of hexachlorobenzene In both static and f lowthrough
systems.
NR •» Not reported
= Lethal concentration for SOX of animals;
= Immobilization concentration for SOS of animals
-------
The toxic effects of monochlorobenzene on egg and embryo development
were studied 1n the laboratory with largemouth bass (Hlcropterus salmoldes).
goldfish (Carasslus auratus) and rainbow trout (Sajmo galrdnerl) using a
flowthrough system with both hard (200 mg/8, CaCQ ) and soft (50 mg/S.
CaCO_) water (B1rge et a!., 1979). WHh trout, exposure to 0.09, 0.31,
1.60, 4.27 and 32.0 mg monochlorobenzene/st was Initiated 20 minutes after
fertilization and continued for 16 days (hatching time for trout 1s 23
days). Complete lethality of the trout embryos occurred at all monochloro-
benzene concentrations within the exposure period 1n hard and soft water
conditions (Table 6-5). The LC for trout embryos was therefore reported
to be <0.09 mg/4 (B1rge et a!., 1979). Largemouth bass embryos/larvae
were exposed 1-2 hours postfertH1zat1on through hatching until 4 days
posthatchlng. (Average hatching time for bass 1s 3.5 days.) Chlorobenzene
concentrations ranged from 0.013-27.3 mg/a, for soft water and 0.009-23.2
mg/8, for hard water conditions. Percent hatchabmty was reduced to 72,
25 and 4% of controls at 0.15, 3.10 and 23.2 mg/a,, respectively, 1n hard
water. Percent survival of bass larvae at 4 days posthatchlng was 80, 60
and 24% after exposure to 0.013, 0.038 and 0.16 mg monochlorobenzene/a,
respectively, 1n soft water conditions. The LC,.-. value at 4 days post-
3U
hatching for bass larvae was reported to be 0.05-0.06 mg/a, while the
LCpn value for embryos exposed until hatching was 0.34-0.39 mg/8, (see
Table 6-5). Goldfish, C_. auratus. were more tolerant to monochlorobenzene
exposure during development. (Average hatching time for goldfish 1s 4
days.) The LC,-0 values for embryos exposed until hatching and embryos/
larvae exposed until 4 days post-hatching ranged from 2.37-3.48 mg/a, and
0.88-1.04 mg/a, respectively (see Table 6-5). Abnormal bass larvae
6-20
-------
TABLE 6-5
Embryo-Larval Toxldty of Monochlorobenzene to Goldfish, Largemouth Bass
and Rainbow Trout 1n Soft and Hard Water*
Soft Water
(50 mg/a. as CaC03)
Species
Goldf1shb
Largemouth bassc
Rainbow troutd
Exposure In
Days Beyond
Egg Hatching
0
4
0
4
£/
LC50
(mg/a)
3.48
0.88
0.34
0.05
<0.09
95%
Confidence
Limits
3.08-3.87
0.67-1.12
0.22-0.51
0.04-0.07
NA
Hard Water
(200 mg/l as CaC03)
LC50
(rag/I)
2.37
1.04
0.39
0.06
<0.09
95%
Confidence
Limits
1.96-2.86
0.86-1.25
0.25-0.58
0.04-0.08
NA
aSource: B1rge et a!., 1979
^Require ~4 days from spawning to hatching; thus, exposure of the
hatched larvae for 4 additional days resulted 1n a total of 8 days of
continuous exposure.
cRequ1re ~3.5 days from spawning to hatching; thus, exposure of the
hatched larvae for 4 additional days resulted In a total of 7.5 days of
continuous exposure.
^Require ~23 days from spawning to hatching; all exposed embryos were
dead by 16 days after fertilization.
NA = Not applicable
6-21
-------
occurred 1n 2, 13, 42 and 100% of those hatching after exposure to 0.04,
0.15, 3.1 and 23.2 mg/8., respectively, during embryonic development.
Abnormal goldfish larvae were less prevalent (B1rge et al., 1979).
The embryo and larval toxldty of 1,2,4,5-tetrachlorobenzene was tested
1n sheepshead minnows, £. varlegatus. Within 4 hours after assurance of
fertilization, embryos were exposed to 0.06, 0.09, 0.18, 0.30 and 0.52 mg
1,2,4,5-tetrachlorobenzene/8, until hatching; thereafter, exposure of
larval and juvenile fish was continued for an additional 28 days. Hatching
success of embryos was not significantly decreased at any exposure level.
Juvenile mortality was significantly (p<0.05) Increased 1n fish exposed to
>0.18 mg 1,2,4,5-tetrachlorobenzene/8. (Table 6-6). The maximum acceptable
toxicant concentration (HATC) for embryos and juvenile sheepshead minnows
exposed to 1,2,4,5-tetrachlorobenzene was estimated to range between
0.09-0.18 mg/a..
The embryo and larval toxldty of trlchlorobenzene (Isomer not speci-
fied) was studied In American oysters (Crassostrea vlrglnlca) and the hard
clam (Hercenarla mercenarla) (Davis and Hindu, 1969). Exposure which
commenced soon after fertilization and embryo development was determined 48
hours later. To determine larval survival, 2-day-old larvae (hatched under
normal conditions) were exposed for 10 days (for clams) or 12 days (for
oysters) before quantitative sampling. At 1.0 and 10.0 mg trlchloroben-
zene/8,, egg survival and normal embryo development In oysters was 59 and
21%, respectively, of control cultures. In clams treated with 1.0 and 10.0
mg trlchlorobenzene/8., embryo development was reduced to 72 and 58% of
controls. Survival of clam larvae exposed to 1.0 and 10.0 mg trlchloroben-
zene/8, was 108 and 69% of controls, respectively, with no change 1n larval
length. Based on toxldty data, Davis and Hindu (1969) reported a 48-hour
6-22
-------
TABLE 6-6
Results of 1,2,4,5-Tetrachlorobenzene Tests with Embryo to Juvenile
Sheepshead Minnows In Continuous-Flow Natural Seawater3
Nominal
Concentration
(mg/ft)
Control
Solvent control
0.12
0.25
0.5
1.0
2.0
Measured
Concentration*3
(mg/l)
ND
ND
0.06+0.04
0.09+0.04
0 . 1 8+0 . 07
0.30+0. 16
Q.52i0.33
Hatching
Success
(X)
84
85
76
81
91
83
67
Juvenile
MortaHtyc
(X)
21
25
16
41
54d
79d
98d
Standard Length
of Juveniles
(mm)
11+2
12+3
10+3
12±2
10+3
12+1
12+Qe
aSource: Ward et a!., 1981
^Values expressed as mean +_ standard deviation
cAt 28 days after hatching
dS1gn1f Icantly greater than control at p<0.05
eOnly one fish survived: the 96-hour LCsg for Juveniles was 0.33 mg/a
with 95% confidence limits of 0.12-0.94 mg/8,.
NO = Not detectable (<0.007 mg/8,}
6-23
-------
LCg0 of 3.13 mg/8, for oyster embryos and 48-hour and 12-day LC5Q
values of >10.0 mg/8, for clam embryos and larvae.
The effects of 1,3,5-tMchlorobenzene on the reproductive performance 1n
brine shrimp, Artemla sallna. were reported by Grosch (1973). Ten pairs of
adult shrimp were exposed to 10 ppm 1,3,5-tr1chlorobenzene for 24 hours and
studied for their lifetime for reproductive performance. The Hfespan of
treated adult females was significantly (p<0.05) reduced. The number of
broods, number of zygotes, and larval survival rate were all significantly
reduced 1n exposed cultures (Table 6-7). The author discussed the possi-
bility that brood number and zygote number were related to the decreased
Hfespan of adult females, but discounted this as the sole cause after
computations showed a decrease 1n the brood size (Grosch, 1973). Cultures
of Artemla sallna that were continuously exposed to 10 mg 1,3,5-tMchloro-
benzene/a, survived <1 week and produced no viable embryos.
6.1.4. Effect on Aquatic Plants. The 96-hour EC (effective concen-
tration for 50% of the algae to show the effect) for reduced chlorophyll a
content 1n the freshwater algae, Selenastrum capMcornutum. treated with
monochlorobenzene was 232 mg/8, (Table 6-8). The 96-hour ECC_ for 1nh1-
bu
bltlon of growth and the reported NOEL were 224 and <111 mg/8., respec-
tively (U.S. EPA, 1978). (For more complete toxldty data for algae refer
to Table 6-8.). Toxldty of 1,2-, 1,3- and 1,4-d1chlorobenzene was somewhat
varied when comparing 96-hour EC values for reduced chlorophyll content
of 91.6, 179 and 98.1, respectively. The general trend of Increasing
toxldty with Increased chlorine substitution 1s seen with 1,2,4-tr1chloro-
benzene, tetrachlorobenzenes and pentachlorobenzene (see Table 6-8). The
1,2,3,5- Isomer of tetrachlorobenzene appears to be 2- to 3-fold more toxic
than the 1,2,4,5- Isomer 1n this freshwater algae, S. capMcornutum. The
6-24
-------
TABLE 6-7
Adult Llfespan and Reproductive Performance of Brine Shrimp
Exposed to 1,3,5-Tr1chlorobenzenea»k
Brine
Solution
Controls
Acetone
Controls
Exposed to
l,3,5-Tr1-
chlorobenzene
Adults:
Survival (1n days)
Males0
d
Females
Number of broods (per pair)
49.6±4.0 47.6j4.0 44.2*3.8
50.0i5.0 50.1*5.5 37.6*4.2
11.3*1.6 11.8-1-1.& 5.3*0.8
Offspring;
Total number of zygotes produced
Cysts produced (%)
Cysts hatched (%)f
Survival of larvae (%)
Sex ratio (no. males/no, females)
Adaptive values (ratio of average
no. of matured offspring per pair
exposed to 1,3»5-tr1chlorobenzene/
average no. of matured offspring
per pair 1n acetone controls)^
1828
29.0
46*5
7&.3i5.0
0.91
1884
30.6
48*7
75.6±4.7
0.94
1.00
456
11.4
18*10
30.3*11.5
0.82
0,11
aSource: Srosch, 1973
bTests performed with TO mating pairs exposed at 10 mg/a. for 24 hours;
each pair then returned to separate fresh brine solutions.
cControl and treatment means not statistically different.
^Statistically significant difference between control and treatment means
at 0.05 level.
6Stat1st1cally significant difference between control and treatment means
at 0.005 level.
^Statistical analyses not reported for difference between control and
treatment means.
6-25
-------
TABLE 6-8
Acute Toxldty Data for Aquatic Algae Exposed to Chlorinated Benzenes
i
1X9
*T»
Compound Species
Monochlorobenzent freshwater alpe
(Selenastrum capricornutum)
marine algae
(Skeletonema costatuml
Green algae
(Scendesmus quadMcauda)
1,2,-Dlchlorobenzene freshwater algae
(Selenastrum caprlcornutum)
marine algae
(Skeletonema costatuin)
Green algae
(Scendesmus quadrkauda)
1 ,3-D1chlorobenzene freshwater algae
(Seletrastrum capricornutum)
Duration
(hours)
243
48a
96a
96b
%c
96
24a
483
96a
96*>
96=
168
24a
483
?2a
96a
96»
96C
96
243
483
723
96a
96&
96=
168
243
483
72a
96a
96b
96C
Mean
Concentration
(Big/D
330.0
264,0
232.0
224.0
390,0
84.8
138.0
119.0
91.6
98.0
<12.9
2.2
66.7
45.1
45.6
44.2
44.1
<12.8
>100.0
180.0
170.0
162.0
179.0
149.0
41,8
Method
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
Effect
"SO
"50
"50
"50
None
"SO
"50
"50
"50
"50
None
"3d
"50
"50
"50
"SO
"50
None
"50
"50
"50
"50
"50
"50
None
"3d
"50
"50
"50
"50
"50
None
Reference
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Calamarl et al.,
1983
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA. 1978
U.S. EPA, 1978
U.S. EPA, 1978
BMngmann and
Kuhn, 1980
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA. 1978
U.S. EPA, 1978
U.S. EPA, 1978
Calamarl et al. ,
1983
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
Brlngmann and
Kuhn, 1980
U.S. EPA, 1978
U.S. EPA. 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
U.S. EPA, 1978
-------
TABLE 6-8 (cent.)
Compound Species
1 ,3-01chlorobenzene (cont.) marine a Ig3e
(Skeletonema costatum)
1 ,4-D1chlorobenzene freshwater algae
(Selenastrum capjrlcornutum)
marine algae
(Skeletonema costatum)
1 ,2,3-Tr1chlorobenzene freshwater algae
(Se1ena.stru» caprlcornutum)
1 ,2,4-Trlchlorobenzene freshwater algae
(Selena strum caprlcornutum)
marine algae
(Skeletonema costatum)
Duration
(hours)
24*
48*
723
96*
96b
96C
243
48*
72a
96*
96b
96C
96
243
48*
72a
96*
96b
9&c
96
24*
48*
72*
96a
96b
96C
96
24*
48*
72*
96a
96b
9&c
Mean
Concentration
55.8
41.9
62.3
52.8
49.6
7.3
76.9
61.6
77.5
98.1
96.7
5.6
1.6
61.9
56.6
50.6
54.8
59.1
10.0
0.9
55.0
32.8
31.8
35.3
36.7
<8.2
1.4
13.5
1.46-2.63
1.46-2.63
8.75
8.93
<1.46
Method
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
Effect
EC50
EC50
"50
"50
None
EC50
"50
EC50
EC50
"50
None
EC50
EC50
EC50
EC50
EC50
EC50
None
ECSO
EC50
EC50
EC50
"50
None
ECSO
"50
ECSO
ECSO
EC50
EC50
None
Reference
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
Calamarl
1983
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
Calamar!
1983
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
Calamar 5
1983
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
U.S. EPA
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
et al.,
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
et al.,
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
et al.,
, 1978
, 1978
, 1978
, 1978
, 1978
, 1978
-------
TABLE 6-8 (cont.)
on
trs
Compound Species
1,2,3,5-Tetrachlorobenzene freshwater algae
(Selenastrum caprlcornutum}
marine algae
(Skeletonema costaturc)
1,2,4,5-Tetrachlorobenzene freshwater algae
(Selenastrum caprlcornutum)
marine algae
(Skeletonema costatum)
Pentachlorobenzene freshwater algae
(Selenastrum caprlcornuturaj
marine algae
(Skeletonema costatum)
Duration
(hours)
243
483
723
963
96b
96C
24a
48a
723
963
96b
96C
243
48a
723
96a
96b
96C
24a
48a
723
96a
96b
96c
243
48a
723
96*
96b
9&c
243
483
723
963
96b
96C
Mean
Concentration
(mg/l)
27
28
14
17
17
<3
2
2
1
0
0
<0
50
54
47
52
46
<3
>18
9
8
7
7
<1
>32
8
13
6
6
0
5
1
1
2
1
<0
.4
.0
.7
.2
.7
.2
.83
.53
.39
.83
.70
.1
,4
.9
.3
.9
.8
.2
.0
.39
.56
.10
.32
.0
.0
.25
.0
.78
,63
.10
.53
.57
.94
.23
.98
.1
Method
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
static
Effect
"50
EC50
"50
"50
"50
None
"SO
"50
"50
"50
"50
None
"50
"50
"50
"50
"50
None
"50
EC50
"50
"50
"50
None
"50
"50
"50
"50
"50
None
"50
"50
"50
"50
"50
None
Reference
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
U
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
,s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
.s.
EPA,
EPA,
EPA,
EPA.
EPA,
EPA.
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
EPA,
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
1978
-------
TABLE 6-8 (cont.J
*r>
i
tf>
Compound
Hexachlorobenzene
Species
Freshwater algae
( Sell enast rum caprlcornutumi
Tetrahymena pyr If orals
Nixed culture; diatom/green
algae (Thalassloslra
pseudonana/Duna 11 e 1 la
tertlolecta)
Green algae
(Chlorella pyrenoldosa)
Duration
(hours)
96
240
12
16
Mean
Concentration
<0.03
0.001
0.1
10.0
Method Effect
static ECso
static Growth
reduction6
static No growth
Inhibition
static Growth
reduction^
Reference
CalamaM et al.,
1983
Gelke and
Parasher, 1976
Biggs et al.,
1979
Parasher et a1.,
1978
Effective on chlorophyll a content
^Effective on cell growth
CMOEL
^A 3% change In growth measured by turbidity
eGrowth reduced to 66X of control cultures; measured by dry mass
fGrowth reduced to 87.5X of control cultures; measured by dry mass
Concentration Inhibiting the growth of SOX of the population
-------
U.S. EPA (1978) also conducted similar toxlclty tests on the chlorinated
benzenes with the marine algae, Skeletonema costatum. The 24, 48, 72 and
96-hour EC,.- values and the 96-hour NOELs for the chlorinated benzenes
DU
studied are shown In Table 6-8. Effective toxldty concentrations of each
chlorinated benzene are within the same range for both the freshwater and
marine algae. Data from other studies (Brlngmann and Kuhn, 1980; Gelke and
Parasher, 1976; Biggs et a!., 1979; Parasher et a!., 1978) using various
algal species are also reported In Table 6-8.
6.1.5. Residues. Residue concentrations of the chlorinated benzenes {In
sediment and water) were determined 1n the Great Lakes (Superior, Huron,
Erie and Ontario), drinking water of surrounding cities, wastewater
effluents from area Industries and from the Grand and Niagara Rivers (Oliver
and N1eol, 1982). These data, reviewed 1n Table 6-9, Indicate that almost
all chlorinated benzenes exist In measurable quantities 1n the Great Lakes
and can occasionally be traced to point sources. Oliver and Nlcol (1982)
Indicate that these substances are persistent 1n the sediment and are
bloconcentrated by fish.
Bjerk and Brevlk (1980) collected sediment core samples (0-5 cm deep)
and reported concentrations of 0.87 mg/kg pentachlorobenzene and 0.528 mg/kg
hexachlorobenzene (dry weight basis) 1n the Oslo fjord at Asstranda, Norway.
Deeper samples contained less contaminants (0.064 and 0.317 mg/kg, respec-
tively). At Ora, Norway, sediment samples contained lower levels of penta-
chlorobenzene (0.003 mg/kg dry weight). In a wide variety of species tested
(algae, crustaceans, mollusks and fish), penta- and hexachlorobenzene appear
to bloaccumulate, usually about 20-fold over environmental levels
(Table 6-10).
6-30
-------
TABLE 6-9
Chlorinated Benzene Concentrations (ng/a) 1n Water and Sediment3
Chemical
1,3-Dlchlorobenzene
1 ,4-Dlchlorobenzene
1,2-Dlchlorobenzene
. , 1,3,5-Trlchlorobenzene
1 ,2,4-Trlchlorobenzene
i
OJ
"" 1,2,3-Trlchlorobenzene
1,2,3,5-Tetrachlorobenzene
1 ,2,4,5-Tetrachlorobenzene
1 ,2,3,4-Tetrachlorobenzene
Pentachlorobenzene
Hexachlorobenzene
Lake
Superior
NA
2
NA
5
NA
1
NA
0.2
NA
1
NA
0.2
NA
0.1
NA
0.3
NA
0.3
NA
0.1
NA
0.2
Lake
Huron
ND
2
4
16
ND
8
ND
0.7
0.2
6
ND
0.3
ND
0.4
ND
1
0,05
1
0.04
1
0.04
2
Lake
Erie
NA
4
NA
9
NA
2
NA
1
NA
3
NA
0.4
NA
0.3
NA
1
NA
0.7
NA
1
NA
3
Lake
Ontario
ND
74
45
94
5
11
0.1
60
0.6
94
0.1
7
ND
6
0.1
52
0.1
33
0.2
32
0.06
97
City
Drinking
Water
1
NA
13
NA
3
NA
ND
NA
2
NA
0.1
NA
ND
NA
0.2
NA
0.3
NA
0.04
NA
0.1
NA
Hastewater
Effluents
14
NA
660
NA
13
NA
0.3
NA
11
NA
2
NA
0.4
NA
1.2
NA
1.6
NA
0.9
NA
1.5
NA
Niagara1*
River
IB
94
56
fl
107
38
3
31
126
22
17
Grand
River
1
NA
10
NA
6
NA
ND
NA
2
NA
0.1
NA
ND
NA
ND
NA
0.05
NA
0.05
NA
0.06
NA
Sample
Type
W
S
U
S
W
S
M
S
H
S
M
S
W
S
W
S
W
S
H
S
W
S
aSource: Oliver and N1col» 1982
^Highest value of four sampling sites reported
NA = Not available; ND = Not dectected; S = Hean concentration In surfldal sediment sample; H = Mean concentration In water samples
-------
TABLE 6-10
Chlorinated Benzene Concentrations 1n a Variety of Marine Species
i
CO
Mean Concentration (mq/kq) of Chlorinated Benzene
Species/Tissue
Cod (Gadus mgrhua)
Cod, homogenate
Cod liver
Cod liver
Cod fillet
Whiting
Sprat
Sprat o1la
Plaice
Eel
Rainbow trout
(Salmo galrdnerl )
Brown trout
(Salmo trutta)
Arctic char
(Salvellnas alplnus)
Atlantic salmon
(Salmo salar)
Number
Analyzed
7
6
6
3
3
2
4.
3
3
10
6
5
6
TM-
0.4
NA
NA
2,7
0.4
1.1
0.5
<0.01-0.5
0.2
0.3
0.6
NA
NA
NA
Tetra-
0.3
NA
NA
0.8
0.14
0.3
0.3
<0.01-0.4
0.4
0.3
1.5
NA
NA
NA
Penta-
3.8
0.79
NA
12.7
1.1
4.3
4.7
0.01-3.7
0.7
0.7
3.5
NA
NA
NA
Hexa-
55.6
19.9
30.9
170
31
56
29
0.04-16
13
13
32.7
31.7
30.0
46.0
Reference
Ofstad et a!,, 1978
Bjerk and Brevlk, 1980
Bjerk and Brevlk, 1980
Ofstad et a!,, 1978
Ofstad et al., 1978
Ofstad et al., 1978
Ofstad et al., 1978
Lunde and Ofstad, 1976
Ofstad et al., 1978
Ofstad et al., 1978
Oliver and N11m1, 1983
Skaftason and
Johannesson, 1982
Skaftason and
Johannesson, 1982
Skaftason and
Johannesson, 1982
-------
TABLE 6-10 (cont.)
Mean Concentration (ma/kg) of Chlorinated Benzene
Species/Tissue
Coho salmon
(Oncorhynchus kisuteh)
Liver
Muscle
Brittle star
(Ophlura ajblda)
er>
is Hermit Crab
" (Pagurus sp. )
Snail
(Littorina IHtorea)
Sea star
(Asteroldea)
Saithe, homogenate
(Pollachius vlrens)
Number
Analyzed
28
28
15
3
3
12
13
Tri-
NA
NA
NA
NA
NA
NA
NA
Tetra-
NA
NA
NA
NA
NA
NA
NA
Penta-
NA
NA
1,10
0.88
NA
0.78
1.11
Hexa-
0.
0.
21.
4.
13.
1.
21.
065b
097b
2
3
9
03
8
Reference
Norstrom
Norstrom
Bjerk
Bjerk
Bjerk
Bjerk
Bjerk
and
and
and
and
and
et a!.,
et al. ,
Brevlk
Brevik
Brevik
Brevik
Brevik
1978
1978
, 1980
, 1980
, 1980
, 1980
, 1980
aValues are the concentration ranges for five sampling sites around Norway.
^Concentrations expressed as wet weight of fish.
-------
Fish and Invertebrates collected from contaminated waters have been
shown to contain various levels of chlorinated benzenes. Only Ofstad et al.
(1978) collected water and sediment samples for chlorinated benzene analysis
and quantitatively confirmed the presence of tr1- through hexachlorobenzenes
In the area where contaminated fish were collected. Concentrations of
chlorinated benzenes 1n the fish were Inversely related to the distance of
the collection site from a chlorinated benzene discharge point. Data from
several reports of tissue levels of tr1-, tetra-, penta- and hexachloroben-
zene 1n fish from the United States, Canada and Norway are presented 1n
Table 6-10. Brunn and Hanz (1982) collected 72 samples of various fish
species from several ponds, streams and rivers of Germany. Residues of
hexachlorobenzene were present 1n 66 samples (92%) at average concentrations
ranging from 0.265 mg/kg fat In fish from ponds without a flowing surface-
water connection to 0.463 mg/kg fat In fish from rivers.
Additional data on tissue levels of chlorinated benzenes In fish and
BCFs were discussed 1n Section 5.3.
6.2. EFFECTS ON NONAQUATIC ENVIRONMENTS
6.2.1. Plants. Plant seedlings and germinating seeds are commonly
exposed to 1,4-d1chlorobenzene to disrupt or arrest mitosis and facilitate
chromosome study (Meyer, 1948). Sharma and Bhattacharyya (1956) exposed
healthy root tips of 10 monocotyledons and 6 dicotyledons to a saturated
solution of 1,4-dlchlorobenzene. Chromosome fragmentation was observed 1n
all species after 1.5-4.5 hours of exposure. Barley, oat and wheat seed-
lings were raised 1n greenhouse pots of sand, sandy loam, clay loam or clay
treated with 1,2,4,5-tetrachlorobenzene at application rates equivalent to
0, 1.9, 5.6, 16.9, 50.6 or 151.9 kg/ha (Ameen et al., 1960). Eighteen days
after planting, a decrease was observed In seedling germination and 1n
6-34
-------
heights and root lengths of seedlings of all three varieties and 1n all four
soil types. A gradient of severity was reported, however, decreasing from
sand to sandy loam, clay loam and clay. No effects were noted 1n any
variety grown 1n any son type treated at the highest application rate
(151.9 kg/ha) 1f planting was delayed 125 days. Mature cotton plants grown
1n Norfolk sandy loam soil were observed 30 days after soil treatment with
1,2,4,5-tetrachlorobenzene at application rates of 0-4483 kg/ha to control
nematode parasites (Adams and Rodrlquez-Kabana, 1976). There was 100%
mortality 1n plots treated at >224 kg/ha. No effects on the cotton plants
were observed at application rates of 0-112 kg/ha.
6.2.2. Insects. Pupae of the housefly, Husca v1c1na. were exposed to
"saturation concentration" vapors of each of the three dlchlorobenzene
Isomers for 3, 6 or 10 hours (Levlnson, 1955). The emergence of adult flies
8 days after exposure 1s shown 1n Table 6-11. The actual concentrations of
the various exposure atmospheres, however, were not reported.
Solutions of 1,2-d1chlorobenzene 1n dlesel oil (1:3 or 1:5 ratio
d1chlorobenzene:o1l) and of an unspecified trlchlorobenzene Isomer 1n dlesel
oil (1:5 ratio) effectively eliminated all broods of the Douglas-fir beetle,
Dendroctonus pseudotsugae. when sprayed on both fallen logs and standing
trees (Gibson, 1957). The actual volumes of spray or total weight of the
chlorobenzene applied were not specified.
Fifteen virgin female wasps, Bracon hebetor, were each placed overnight
Inside glass vials, the sides of which had been uniformly coated with a 10
ppm solution of 1,3,5-tr1chlorobenzene 1n 0.25 ms, of acetone (Grosch and
Hoffman, 1973). The mean Hfespan of the females was shortened (15.7^1.1
days) compared with controls (22.0+0.8 days). Embryo mortality, measured by
the number of unhatched eggs, 1n the control and treated groups was similar
6-35
-------
TABLE 6-11
Emergence of Adult Houseflles 8 Days Following Exposure of Pupae to
"Saturation Concentration" of Dlchlorobenzene Vapors3
Emergence of Houseflles (%) Resulting from
Exposure Period of:
Chem1calb
1 ,2-D1chlorobenzene
1 ,3-D1chlorobenzene
1 ,4-D1ch1orobenzene
3 hours
46±10C
15±5
2+2
& hours
15±6
0
0
10 hours
0
0
0
aSource: Levlnson, 1955
^Percent emergence of controls = 94^2%
cln an average of 10% of the cases recorded as unhatched pupae, the flies
had died after having pushed through the pupal skin with their ptlUnum,
but with their thorax and abdomen Inside the pupal skin.
6-36
-------
for the first 5 days after treatment, but by the seventh day, 71% of the
eggs from treated females were unhatched.
6.2.3. Birds. The toxldty of hexachlorobenzene was tested 1n Japanese
quail (Coturnlx coturnlx japonlca) by dietary administration at 0, 1, 5, 20
or 80 mg/kg 1n the diet for 90 days (Vos et a!., 1971). A NOEL was reported
at 1 mg/kg. At higher concentrations liver damage and porphyrln excretion
Increased 1n a dose-related fashion. At 80 mg/kg, 5 of 15 quail died during
the exposure period. There was a dose-related decrease 1n the hatchabUHy
of eggs, especially 1n groups treated at 20 and 80 mg/kg (Vos et a!., 1971).
Carpenter et al. (1983) also reported hepatic toxldty and porphyrla in
quail treated orally with 500 mg/kg/day hexachlorobenzene for 1, 2, 5 or 10
days. Most treatment-related changes occurred after the first dose of
hexachlorobenzene.
Studies on the effects of chlorinated benzenes, predominantly hexa-
chlorobenzene, on wild birds have primarily focused on the accumulation of
contaminants In eggs and their effects on embryo survival and reproductive
parameters. GUbertson and Fox (1977) determined hexachlorobenzene levels
1n eggs of Herring Gulls, Larus argentatus. from Lake Erie, Lake Ontario and
1n northern Alberta (used as an "uncontamlnated" control). Hexachloro-
benzene residue levels 1n eggs were 1.37, 4.30 and 0.21 mg/kg (dry matter
basis), respectively. There was a relationship between the number of
embryos that developed to pipping stages and the final percent hatching, and
the area from which they were collected. Of the eggs collected 1n
"uncontamlnated" areas (n=14), 85% developed to pipping and 69% hatched. Of
those collected at Lake Erie (n=25), 83 and 53%, respectively, pipped and
hatched. Lake Ontario-collected eggs (n=47) showed a significant (p<0.05)
decrease 1n survival to pipping (39%) and hatchabUHy (26%). Liver weights
6-37
-------
and porphyrln levels In embryos from Lake Ontario and Lake Erie were greater
than those of the control group, Oilman et al, (1977) reported that hatch-
Ing success for Herring Gull eggs from Lakes Superior, Huron, Erie and
Ontario were 80, 72, 63 and 19%, respectively, which supports the data of
GUbertson and Fox (1977). Additional data on residue levels of chlorinated
benzenes 1n eggs and wild birds will be discussed 1n Section 6.2.4.
6.2.4. Residues. Harp seals, PhagophHus groenlandlcus. having a high
percentage of body fat, were found to contain hexachlorobenzene residues
(Rosewell et al., 1979). Forty of 42 seal pups contained hexachlorobenzene
(concentrations unspecified), which was concluded to be transferred from
adult to fetus and also through maternal nursing of the pups.
Subcutaneous adipose tissue from wild foxes, boars and deer (1n Germany)
was analyzed for hexachlorobenzene content (Koss and Hanz, 1976). The
average tissue levels (ranges) were 0.29 (0.02-0.77) mg/kg 1n 21 foxes, 0.71
(0.05-3.11) mg/kg 1n 7 wild boars and 0.03 (0.00-0.05) mg/kg In 6 female
deer. The detection of l,4-d1~, l,2,4-tr1-, 1,2,3,4-tetra-, 1,2,4,5-tetra-,
penta- and hexachlorobenzene (concentrations not reported) 1n samples of
pooled body Upld from Lake Ontario Herring Gulls (L_. argentatus) was
reported by Hallett et al. (1982). Similarly, Szaro et al. (1979) reported
that 8 of 28 Great Black-Backed Gulls, collected 1n Maine, had average
tissue levels of 0.03 mg hexachlorobenzene/kg (wet weight). Ohlendorf et
al. (1981) reported hexachlorobenzene residues at an average concentration
of 0.23 mg/kg (wet weight) among 12 of 105 herons, Including great blue
herons, Ardea herodlas. During the period 1971-1974, Barbehenn and Relchel
(1981) examined 101 bald eagles, Hallaetus leu.cgcep.ha1 us, and found 19
carcasses to contain an average concentration of 8.0 mg hexachlorobenzene/kg
(I1p1d basis; 2.2% body weight as Upld). Kaiser et al. (1980) reported
6-38
-------
that 23 of 168 bald eagles collected during 1975-1977 had mean carcass
levels of 0.08 mg hexachlorobenzene/kg wet weight. An Osprey, Pandlon
hallaetus (0,2 mg/kg), a great horned owl, Bubo v1rg1n1anus (0.7 mg/kg),
Swalnson's hawk (up to 5.2 mg/kg) and starlings, Sturnus vulgar Is {0.21
mg/kg) also were found to contain hexachlorobenzene residues (Wlemeyer et
al., 1980; Blus et al., 1983; Bechard, 1981; White, 1979).
Reports on the residue levels of some chlorinated benzenes In bird eggs
are summarized In Table 6-12. Hexachlorobenzene has been the most prevalent
and persistent chlorinated benzene Identified.
6.3. SUMMARY
As demonstrated In acute toxldty bloassays, the LC In fish gener-
ally decreases as the number of substltuent chlorine atoms on the molecule
Increases (Isomers vary). Chlorinated benzenes have adverse effects on the
reproduction of Invertebrates and fish. Monochlorobenzene tested 1n gold-
fish and largemouth bass, 1,3,5-tr1chlorobenzene tested In brine shrimp and
the exposure of sheepshead minnows to 1,2,4,5-tetrachlorobenzene resulted 1n
decreased hatching of eggs or embryo lethality and decreased survival of
juvenile fish.
Adverse effects of chlorinated benzenes were also apparent 1n terrestri-
al organisms. Mitosis 1n seeds and seedlings was disrupted by 1,4-d1chloro-
benzene; 1,2,4,5-tetrachlorobenzene affected seed germination and seedling
growth depending on soil type. Soil application rates of 224 kg/ha or
higher of 1,2,4,5-tetrachlorobenzene were found to be phytotoxlc to mature
cotton plants, Olchlorobenzene vapors at "saturation concentrations"
Inhibited the emergence of housefly pupae, while 1,2-d1chlorobenzene and
trlchlorobenzene each 1n dlesel oil were toxic to Douglas-fir beetles.
6-39
-------
TABLE 6-12
Chlorinated Benzene Residues 1n Bird Eggs
Compound Species
Tetrachlorobenzenes Herring Gull
(Larus argentatusl
Pentachlorobenzene Herring Gull
(Larjjs_ argentatus)
cr>
o
Hexacnlorobenzene Herring Gull
(Larus argentatusl
Great Black-Backed Gull
Common tern
(Sterna hlrundol
Double-Crested Cormorant
(Phalacrocorax aurUusl
Number
Analyzed
65
10
13
65
20
20
13
20
20
65
20
20
20
20
20
28
13
9-10
Mean
Concentration
(rug/kg)
0.026
0.015
0.024
0.039
0.024
0.025
0.025
0.022
0.021
0.451
0.315
0.09
0.115
0.115
0.12
0.03
?.67C
0.016
Location
Lake Ontario3
Lake Ontar1ob
Lake Er1eb
Lake Ontario3
Lake Ontar1ob
Lake Er1eb
Lake Huronb
Lake Super 1orb
Lake M1ch1ganb
Lake Ontar1oa
Lake Ontario11
Lake Er1eb
Lake Huronb
Lake Super1orb
LakeiM1cMgan'J
Maine
Lake Ontario
Bay of Fundy
Reference
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Hallett et al.
Szaro et al.,
Gilbert son and
1972
ZHko. 1976
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
, 1982
1979
Reynolds,
-------
TABLE 6-12 (cont.)
J
•e*
Compound
Hexachlorobenzene
(cont.)
Species
Canvasback Duck
(Ay thy a vallslneria)
Red-Breasted Merganser
(Mergus serrator)
Common Merganser
(Hergus merganser)
Brown Pelican
(Pelecanus occtdentalls)
Great Horned Owl
(Bubo v1rg1n1anus)
Number
Analyzed
11
51
114
92
2
115
4
Mean
Concentration
(mg/kg)
0.02
0.01
0.06
0,05
0.05
0.03
0.2
Location
Nevada
Manitoba
Lake Michigan
Lake Michigan
Lake Michigan
South Carolina
Ohio
Reference
Stendell et al., 1977
Stendell et al., 1977
Haseltlne et al., 1981
Haseltlne et al., 1981
Haseltlne et al., 1981
Blus et al., 1979
Springer, 1980
aOata collected 1n 1977
bData collected In 1978
cBased on dry weight of egg
-------
Contact with residues of 1,3,5-tr1chlorobenzene shortened the Hfespan of
female wasps, and their eggs suffered high mortality within 7 days of
exposure.
Although effects (mortality, decreased reproduction) of chlorinated
benzenes on natural populations have not been adequately studied, tissue
concentrations of several Isomers were measured 1n a number of different
species. Aquatic organisms (fish and Invertebrates) and terrestrial
species, alike, have been found to contain chlorinated benzenes. Tissue
concentrations of the quantltated chlorinated benzenes were highest for
hexachlorobenzene. The detection In North America and Europe of hexachloro-
benzene 1n the eggs of birds and subcutaneous fat of wild animals suggests
Its widespread distribution 1n the environment.
6-42
-------
7. HONOCHLOROBENZENE
Between 88.7 and 128.7 million kilograms of monochlorobenzene 1s
estimated to be produced 1n the United States in 1983 (U.S. EPA, 1983),
Monochlorobenzene 1s used primarily as an Intermediate 1n the synthesis of
organic chemicals and as a solvent 1n herbicides and paints (Hawley, 1977).
It has been detected 1n samples of urban, ambient and Indoor air, as well as
1n surface, drinking and Industrial wastewater, and 1n water and sediments
In a stream draining an abandoned waste site (see Section 4.3.). Residues
of monochlorobenzene have been found 1n human adipose tissue, and studies
Indicate that 1t bloaccumulates 1n fish and other aquatic organisms (see
Sections 5,3. and 5.4.). In addition to the exposure of workers Involved 1n
organic chemical synthesis, humans may be exposed to monochlorobenzene via
Inhalation of air and 1ngest1on of water.
7.1. PHARNACOKINETICS
7.1.1. Absorption. Quantitative studies on the absorption of monochloro-
benzene are lacking. Toxic effects reported 1n humans after 1ngest1on or
inhalation Indicate that monochlorobenzene 1s absorbed via these routes
(Reich, 1934; Rosenbaum et a!., 1947; Tarkhova, 1965). Studies of the
metabolism of monochlorobenzene 1n a number of mammalian species Indicate
that absorption from the gastrointestinal tract does occur (Williams, 1959),
Given the UpophlUc character of monochlorobenzene and the dermal absorp-
tion of other chlorobenzenes, some degree of absorption through the skin
would be expected but definitive studies are lacking.
7.1.2. Distribution. The only available study regarding the distribution
of monochlorobenzene 1s the Inhalation pharmacokinetic experiments of
Sullivan et al. (1983). Male Sprague-Dawley rats were exposed to 100, 400
or 700 ppm of 14C-monochlorobenzenene, 8 hours/day for 1 or 5 consecutive
7-1
-------
days. Following a single exposure period, radioactivity was found 1n all
tissues examined both Immediately and 48 hours post-exposure, with the
highest concentrations located 1n the fat. With the exception of the
kidney, five repeated exposures did not result 1n significantly higher
tissue concentrations than did a single exposure, Indicating that a steady-
state concentration 1s reached during the first 8 hours of exposure. The
tissue concentration was proportional to the exposure concentration, with
the exception of the fat, 1n which the tissue levels Increased 8- to 10-fold
when the exposure concentration was Increased from 100-400 ppm and 3- to
5-fold when the exposure concentration was Increased from 400-700 ppm.
7.1.3. Metabolism. Shlmada (1981) administered monochlorobenzene to rats
(strain and dose unspecified) by subcutaneous Injection and analyzed the
urine by high performance liquid chromatography. They detected £-chloro-
phenylmercapturlc add and monoglucuronlde and ethereal sulfate conjugates
of 4-chlorocatechol. Based on this Information, they proposed that the
metabolism of monochlorobenzene Involves an Initial oxidation to form
4-chlorobenzene-l,2-epox1de. This Intermediate may then 1) form a gluta-
thlone conjugate, resulting 1n the excretion of £-chlorophenylmercaptur1c
add, 2) be converted to 4-chlorophenol, conjugated, and excreted, or 3) be
converted to 4-chlorocatechol, conjugated, and excreted.
NakaJIma and Sato (1979) found that fasting enhanced the activity of
liver enzymes for monochlorobenzene 1n both male and female Wlstar rats.
They found that fed male rats metabolized most of the hydrocarbons tested
more rapidly than fed female rats; however, there were no significant
differences 1n the Initial metabolic rate for monchlorobenzene between sexes.
Selander et al. (1975) Investigated the metabolism of monochlorobenzene
1n perfused rat livers and 1n a variety of cell-free hepatic preparations
7-2
-------
(Table 7-1). They found that monochlorobenzene 1s converted to chloro-
phenols by three different enzymes. Two of these enzymes form arene oxide
Intermediates (3- and 4-chlorobenzene oxides) resulting 1n the formation of
o- and g-chlorophenol. m-Chlorophenol appeared to occur via a direct oxlda-
tlve pathway. Under the conditions of these assays, conjugation of the
arene oxide with glutathlone or hydration did not occur to a significant
extent.
Smith et al. (1972) administered 75 MC1 (Q.59 gm) of 14C-monochloro-
benzene emulsified 1n Cremophor E.L. and physiological saline to two -1.5 kg
female Dutch rabbits by gavage, twice a day for 4 days. The major urinary
metabolites recovered were g-chlorophenylmercapturlc add and the conjugates
of 4-chlorocatechol. Other minor metabolites detected were qulnol, 3-ehlo-
rocatechol, o-chlorophenylmercaptur1c add and m-chlorophenylmercaptur1c
add. The Identified metabolites accounted for over 98% of the urinary
radioactivity and consisted of 3,4-d1hydro-3,4-d1hydroxychlorobenzene
(0.57%), monophenols (2.84%), dlphenols (4.17%), mercapturlc acids (23.80%),
ethereal sulfates (33.88%) and glucuronldes (33.57%).
7.1.3.1. TISSUE BINDING -- Reid (1973) and Reid et al. (1973) have
studied tissue distribution and tissue binding of monochlorobenzene and the
related halobenzene, bromobenzene. Treatment of C57B6J mice with a single
Intraperltoneal dose of 4.58 mmol/kg bromobenzene or 6.75 mmol/kg monochlo-
robenzene produced necrosis of the proximal convoluted tubules of the
kidneys within 48 hours. This was associated with covalent binding of an
unidentified 14C labeled metabolite to the site of necrosis prior to
manifestation of the hlstologlc effect. Pretreatment of animals with
pyrazole butoxlde blocked the binding as well as the toxic effect. Six
hours after administration of 1 mmol/kg (112 mg/kg), -0.332 nmol equiva-
lents of 14C-monochlorobenzene/mg protein were covalently bound. Although
7-3
-------
TABLE 7-1
Percentage of Isomers of Chlorophenol from
Metabolism of Honochlorobenzene*
System
Perfused liver
Phenobarbltal treated
Hethylcholanthrene treated
Hlcrosomes
Phenobarbltal treated
Methylcholanthrene treated
ortho-
40
46
69
18
32
59
Isomer (%)
meta-
20
10
2
7
6
6
para-
40
44
9
75
62
35
*Source: Selander et al.» 1975
7-4
-------
tissue distribution was not studied with monochlorobenzene, a metabolite of
bromobenzene which was also used In this study was strongly bound by tissues
from the liver, lungs and kidneys but not by tissue from the heart, spleen
or testes, and this binding correlated with necrotlc changes. Hlcrosomes
from the lungs and liver (1n vitro) oxidized bromobenzene, whereas micro-
somes from the kidneys, heart, spleen and testes did not, which Indicated
that metabolic activation took place 1n lungs and liver and that an active
metabolite was transported to the kidneys before binding. Pretreatment with
3-methylcholanthrene enhances the overall metabolism of monochlorobenzene;
however, this pretreatment reduces the extent of covalent binding to
cellular macromolecules and prevents centrolobular hepatic necrosis (Reid et
al,, 1971). Similar results of preventing chlorobenzene-elldted liver
necrosis have been obtained by Inhibiting epoxlde hydrase with cyclohexene
oxide (Oesch et al., 1973). Jergll et al. (1982) found that when monochlo-
robenzene was Incubated with liver mlcrosomes 1t was bound to mlcrosomal
proteins of molecular weights of 72,000 and 50,000-60,000 daltons. The
metabolite probably was bound to the sulfhydryl groups of proteins, since
the addition of glutathlone blocked the binding.
7.1.4. Excretion. Sullivan et al. (1983) exposed male Sprague-Dawley
rats to atmospheres containing 14C-monochlorobenzene (100, 400 or 700 ppm)
8 hours/day for 1 or 5 days. Following treatment, the label was detected In
the expired air and urine of the rats. The urine contained metabolites of
monochlorbenzene, Including mercapturlc acids, glucuronlde conjugates and
sulfate conjugate; the respiratory elimination consisted of unmetabollzed
compound. The percentage of the dose excreted by respiration Increased with
Increasing exposure, Implying that the metabolic elimination of monochloro-
benzene can be saturated.
7-5
-------
Smith et al. (1972) orally dosed two female Dutch rabbits with 0.5 g (75
vC1) of inC-monochlorobenzene emulsified 1n Cremophor E.L. and physio-
logical saline twice a day for 4 days, and collected urine and feces
throughout the 7 days of the study. The urine contained 19.6% of the admin-
istered label, the feces (methanol extracted) contained 1.05% and the
tissues contained 0.05%. Radlolabeled 14C 1n expired air was not mea-
sured. Williams (1959) has reported that 27% of a 0.5 g/kg dose orally
administered to rabbits was excreted 1n expired air over a 1-2 day period.
Lindsay-Smith et al. (1972) found that the conjugated metabolites were
both mono- and d1phenol1c 1n the rabbit, but the monophenollcs were predom-
inant. p_-Monochlorophenol was the predominant Isomer 1n the urine. The
distribution of Isomers for free and conjugated monochlorophenol combined
was: ortho-, 4.9%; meta-, 22.9%; and para-, 72.2%. For free monochloro-
phenols, the distribution of Isomers was 5.9, 33.6 and 60.4% for ortho-,
meta- and para-1somers, respectively. The major diphenollc metabolite was
reported to be 4-chlorocatechol; small amounts of chloroqulnol, 3-chloro-
catechol and quinol also were found. Although there was not adequate proof
1n these studies, 1t was proposed that metabolism proceed through the forma-
tion of an arene oxide (3,4-chlorobenzene oxide). Conjugation of this arene
oxide with glutathlone followed by further metabolic reactions would account
for the meta- and para-chlorophenyl mercapturlc adds but not the ortho-
Isomer. Hydratlon of the arene oxide, followed by dehydrogenatlon, would
lead to chlorocatechol. Pathways for metabolism have been proposed based on
the Ijn vivo and in vitro studies (Figure 7-1).
The profile of urinary metabolites varies from species to species. For
example, Williams et al. (1975) reported on 13 species and Indicated that
19-65% of l4C-ur1nary metabolites of monochlorobenzene was p_-chloro-
7-6
-------
Cl
Cl
4-chloTOphenol
conjugation
[ glucuronidej
J
B
OH s-glutathione
dehydrogenise
1
Cl
A-chlorocatechol
SCH -CHCOOH
2|
NHCOCH.
Cl
•OH
3-chlorophenol
conjugation
"mercapturic acid
glucuTonide
sulfste
4-chlorophenylnercapturic acid
FIGURE 7-1
Metabolism of Monochlorobenzene
Adapted from: Williams, 1959; Lindsay-Smith et al.. 1972;
Selander et al., 1975; Shlmada, 1981; Sullivan, 1981
7-7
-------
phenyl mercapturlc add. The principal metabolites 1n humans were the same
as those 1n animals, but the proportions of metabolites were different
(Table 7-2). In humans, 19% appeared as the mercapturlc add and 33 and
31%, respectively, were excreted as 4-chlorophenol and 4-chlorocatechol
sulfate, and glucuronlde conjugates. The ultimate urinary metabolites would
also be expected to vary depending on saturation of metabolism or on the
nutritional state of the animals. If glutathlone levels are depleted, the
metabolic fate can vary. Sullivan (1981) found that monochlorobenzene
metabolism was saturable 1n rats. Male Sprague-Dawley rats were exposed via
Inhalation to 100, 400 or 700 ppm monochlorobenzene vapor for an 8-hour
period. Urinary metabolite profiles and tissue glutathlone concentrations
were measured at 16 and 48 hours after exposure. The capacity of metabolic
oxldases and the conjugation of metabolites to glutathlone were saturated at
the two higher levels of exposure. Saturation of detoxification mechanisms
can Increase the Incidence and severity of toxldty. Recent studies have
examined the profile of urinary metabolites after Inhalation exposure
(Sullivan, 1981) or after subcutaneous administration (Shlmada, 1981).
Essentially, the results of these studies are consistent with results 1n
other species that were administered monochlorobenzene orally.
Ogata and Shlmada (1982) compared the metabolism of monochlorobenzene 1n
rats and humans. The compound was diluted with polyethylene glycol and
Injected 1ntraper1toneally Into rats or administered to rats and human
volunteers orally. Urine specimens were also collected from two workers at
a factory Involved 1n distilling monochlorobenzene. In rats, the major
metabolite detected was p_-chlorophenylmercaptur1c add, accounting for 6-10
times the amount of material excreted as conjugates of 4-chlorocatechol.
In a human volunteer, only trace amounts of p_-chlorophenylmercaptur1c
7-8
-------
TABLE 7-2
Species Variation 1n Urinary Metabolites of ^C-Monochlorobenzene*
Species
Han
Rhesus monkey
Squirrel monkey
Capuchin monkey
Dog
Ferret
Hedgehog
Rabbit
Rat
Mouse
Gerbll
Hamster
Guinea pig
Percentage
4-Chlorophenol
33
19
14
19
14
33
20
29
23
20
13
15
27
of 24-Hour Excretion
4-Chlorocatechol
31
37
37
36
45
31
12
38
22
31
26
23
35
of !«C
4-Chlorophenyl-
Mercapturlc Add
19
40
50
41
42
24
65
26
49
42
51
43
21
*Source: Data cited by Williams et al., 1975
7-9
-------
add were detected; however, conjugates of 4-chlorocatechol were the major
metabolites observed. These findings suggest that urinary 4-chlorocatechol
conjugates may be useful to monitor human exposure to monochlorobenzene.
7.1.5. Summary. Monochlorobenzene 1s readily absorbed by Inhalation and
by the gastrointestinal tract but the quantitative extent 1s not known. It
1s deposited 1n body Uplds and metabolized by mlcrosomal oxidation. Oxlda-
tlve reactions are believed to lead to the preliminary formation of metast-
able arene oxides; these epoxldes are metabolized further to the ortho-,
meta- or para-chlorophenols or they may Interact with tissue. The chloro-
phenols may conjugate with glutathlone and be detoxified by conversion to
the corresponding mercapturlc adds and excreted 1n the urine or they may
bind to cellular proteins. Binding to cellular protein appears to be
correlated with necrotic pathologic changes 1n the kidneys and livers of
rodents. In addition to conjugation with glutathlone, metabolites of mono-
chlorobenzene (monophenols and dlphenols) can conjugate with glucuronlc add
or with sulfate and be excreted 1n the urine. Monophenols are the predomi-
nant metabolites; the dlphenols are minor. The arene oxides, 3-chloroben-
zene oxide or 4-chlorobenzene oxide, also can be converted to the dlhydro-
dlol by epoxlde hydrase and dehydrogenated to form chlorocatechols. There
appear to be species differences 1n the profile of urinary conjugation of
metabolites, and end metabolites may vary depending on the availability of
tissue glutathlone. Detoxification by conjugation with glutathlone 1s
Important 1n the modulation of toxic effects especially at high exposure
levels. Saturation of these metabolic pathways has been demonstrated at
relatively low exposure levels.
7.2. EFFECTS ON HUMANS
No ep1dem1olog1c studies regarding the effects of exposure to monochlo-
robenzene are available. Several case studies and one clinical study,
7-10
-------
however, provide some Information regarding the toxic effects of the chem-
ical 1n humans. Maximum allowable air concentrations range from 75 ppm In
the United States and Switzerland to 11 ppm 1n Sweden (MeMan, 1980).
Reich (1934) reported the case of a 2-year-old boy who swallowed -5-10
mfi, of monochlorobenzene. Within 2 hours, the child was cyanotlc, and he
had no detectable reflexes. He became unconscious and cyanotlc and
displayed head and neck twitching. He regained consciousness after ~3 hours
and all signs returned to normal within 8 hours. There was no followup on
the patient.
Glrard et al. (1969) reported the case of a 70-year-old woman who had
worked for 6 years with a glue containing 70% monochlorobenzene. From the
time when she began using the glue, her symptoms Included headaches and
Irritation of the upper respiratory tract and the eye mucosa. After 6 years
of exposure hematologlc examination resulted 1n a diagnosis of medullar
aplasla.
Rosenbaum et al. (1947) examined 28 factory workers who had been exposed
to monochlorobenzene for 1-2 years. Many of the workers complained of head-
aches and showed signs of somnolescence and dyspepsia. Eight of the 28 had
tingling, numbness and stiffness of the extremities, eight had hyperesthesla
of the hands, nine had spastic contractions of the finger muscles, and two
had spastic contractions of the gastronemlus muscle. Twenty-six workers who
had either short-term exposure (<1 year) to monochlorobenzene or exposure to
combinations of benzene and monochlorobenzene fumes displayed no neurotoxlc
signs,
Tarkhova (1965) exposed 4 humans to 0.02, 0.04 or 0.06 ppm (0.1, 0.2 or
0.3 mg/m3) of monochlorobenzene and monitored electroencephalographlc
patterns. At the lowest concentration there were no effects, but within
7-11
-------
minutes at the higher exposures, there were changes 1n response patterns to
10-nanosecond light flashes of 8-10 Hz.
Human exposure to monochlorobenzene by Inhalation or by accidental
1ngest1on can cause neurotoxlc signs (Reich, 1934; Rosenbaum, 1947). It 1s
not known 1f the effects are reversible after long-term exposure or If there
are other sites of toxldty.
7.3. MAHHALIAN TOXICITY
7.3.1. Acute Toxldty. Treatment with monochlorobenzene has been demon-
strated to produce a variety of changes 1n enzymatic and physiological func-
tion, Including the slight depression of mltochondrlal oxldatlve
phosphorylatlon 1n male Oonryu rats (Ogata et a!., 1981), Increased flow of
bile duct-pancreatic fluid 1n male Holtzman rats (Yang et a!., 1979),
stimulation of the activity of 6-am1nolevul1n1c add synthetase and
hemeoxldase 1n male Wlstar rats (Ar1yosh1 et al., 1981) and decreased
hepatic cytochrome P-450 1n female Wlstar rats (Ar1yosh1 et al., 1975).
Varshavskaya (1967) Investigated the toxlcologlcal, olfactory and gusta-
tory properties of monochlorobenzene and ortho- and para-dlchlorobenzene.
The olfactory and gustatory thresholds were found to be 0.01-0.02 mg/8, for
monochlorobenzene. In the oral toxldty tests with albino rats, the highest
concentration of monochlorobenzene that produced no observed toxic effect
was 0.001 mg/kg.
R1m1ngton and Zlegler (1963) administered monochlorobenzene to male
albino rats by dally gastric Intubation, using an escalating dosage regimen.
Monochlorobenzene was less effective 1n producing porphyrla than were
1,4-d1chlorobenzene, 1,2,4-tr1chlorobenzene or 1,2,3,4-tetrachlorobenzene
(hexachlorobenzene was not studied). Monochlorobenzene also has been
observed to produce bronchlolar necrosis (Reid et al., 1973) and centro-
lobular hepatic necrosis (Reid and Krishna, 1973).
7-12
-------
A summary of the acute lethal doses of monochlorobenzene 1s presented 1n
Table 7-3. In a majority of the studies reviewed, fatalities were the
result of central nervous system depression. Irish (1963) reported that
cats tolerated monochlorobenzene at concentrations of 220-660 ppm for 1
hour. Narcotic signs were noted at levels of 1200 ppm, and death occurred
after 7 hours of exposure at 3700 ppm. Cats exposed at 8000 ppm for 30
minutes died 2 hours after exposure. By the oral route, LD™ values 1n
rats and rabbits were reported to be 2.91 and 2.83 g/kg bw, respectively.
Bonnet et al. (1982) reported that 6-hour Inhalation exposures to rats and
mice resulted 1n LC_-s of 2965 and 1886 ppm, respectively.
Administration of sublethal doses of monochlorobenzene causes toxic
signs that are manifest within 24 hours. When mice are given a single
1ntraper1toneal dose of 6.75 mmol/kg (760 mg/kg), they develop coagulation
necrosis of the proximal tubules of the kidneys. Rats are slightly less
sensitive. Doses of 9.3 mmol/kg (1047 mg/kg) have been reported to cause
swollen, vacuolated, convoluted tubules (Reid et al., 1971).
Monochlorobenzene causes sensory Irritation of the respiratory system
after inhalation exposure. A comparison of the Index of sensory irritation
for 22 chemicals was made based on a short Inhalation experiment 1n mice (De
Ceaurrlz et al., 1981). Mice were exposed usually for 5 minutes at varying
concentrations, and respiratory rates were measured with a plethysmograph.
An RD value for mice, which 1s the concentration that causes a 50%
decrease 1n respiratory rate, was calculated. An uncomfortable human dose
was predicted to be 0.1 RDcn» and a no-effect dose was predicted to be
0.01 RDcf]- For monochlorobenzene, the RD™ was 1054 ppm, and the
predicted no-effect human dose was 11 ppm. For comparison, the RD™ for
formaldehyde and toluene d11socyanate were 5.3 and 0.24 ppm, respectively.
7-13
-------
TABLE 7-3
Acute Toxldty of Honoehlorobenzene
Species
Rat
Cat
Rat,
Sprague-Dawley
House
Rat
Rat
Rabbit
Rat
Guinea pig
Rabbit
Route
Inhalation
Inhalation
Inhalation
Inhalation
oral
oral
oral
1.p,
1.p.
dermal
Dose
22,000 ppm
9,000 ppm
3,700 ppm
8,000 ppm
2,965 (2787-3169)* mg/kg
1,886 (1781-1980)* mg/kg
2,144 mg/kg
400-1600 mg/kg
2,830 mg/kg
7,400 mg/kg
4,100 mg/kg
>10 g/kg
Exposure
Duration
(hour)
2.5
3.0
7.0
0.5
6.0
6.0
NR
NR
NR
NR
NR
NR
Lethal
Effect
Level
LC50
LC67
LClOO
LClQO
LC50
LC50
"-D50
LD50
LDSO
LD50
LD50
LD50
Reference
Eastman Kodak,
Irish, 1963
Bonnet et al.,
Bonnet et al,,
Monsanto, 1965
Eastman Kodak,
Eastman Kodak,
NIOSH, 1982
NIOSH, 1982
Konsanto, 1965
1978
1982
1982
1978
1978
*95% confidence limits 1n parentheses
NR = Not reported
-------
B\oche«\\cal manAfestatAons of the acute toxic effects of monochloroben-
zene may be associated with the binding of liver and kidney protein by
metabolites of the compound (the arene oxide or monochlorophenol) as dis-
cussed 1n Section 7.1.2. Ogata et al. (1981) found that 0.24 mM monochloro-
benzene, 1n an In vitro assay, caused a slight depression of rat liver mlto-
chondrlal oxldatlve phosphorylatlon. This effect was much less than the
effect caused by the more highly chlorinated congeners. A slight decrease
1n hepatic cytochrome P-450 was observed 1n female rats administered 200
mg/kg monochlorobenzene 1ntraper1toneally 24 hours before analysis (Ar1yosh1
et al., 1975).
7.3.2. Subchronlc Toxldty. The subchronlc toxlclty data are summarized
1n Table 7-4. Several Investigators have studied the subchronlc Inhalation
toxlclty of monochlorobenzene. 011 ley (1977) exposed groups of 32 male
Sprague-Dawley rats (125 g) or male rabbits (2.0-2.5 kg) to monochloroben-
zene (99+X) at 0, 75 and 250 ppm for 7 hours/day, 5 days/week, for 24 weeks.
After exposure for 11 weeks (55 exposures), the rats showed Increased Hver-
to-body weight ratios. After 120 exposures at 250 ppm, the rats showed an
Increase 1n liver-to-body and kidney-to-body weight ratios as well as
decreased food consumption. Slight changes were also observed 1n three
hematologlc parameters (retlculocyte, white blood cell, and platelet
counts). H1stopatholog1c changes were seen 1n the kidneys, liver and
adrenals of rats at 11 and 24 weeks; the kidneys had regenerating cortical
tubules with basophlUc Inclusions 1n the cytoplasm of cells, the livers
were congested and the adrenals had vacuolatlon of cells 1n the zona fascl-
culata. It was suggested that 75 ppm may be the marginal toxic concentra-
tion for dally Inhalation. Effects were less marked 1n rabbits than 1n
rats; no hlstologlc or hematologlc changes were found relating to monochlo-
robenzene exposures at 24 weeks.
7-15
-------
1A8LE 7-4
Summary of Subchrontc Toxlclty Studies on Honochlorobeniene3
Species
Dog
(beagle)
Rat
Rat
Rat
Rat
Rat
Rat
Rabbit
Route
1nhalat1onb
Inhalation
Inhalation
Inhalation
Inhalation
Inhalation
Inhalation
Inhalation
Dose
0.75 mg/l, 6 hrs/day,
5 days/week (162 ppm)
1.50 mg/l, 6 hrs/day,
5 days/week {424 ppm)
2.00 mg/l, 6 hrs/day,
5 days/week
0.75, 1.50 or 2 mg/l
6 hrs/day, 5 days/week
0.1 or 1.0 mg/m3
(continuous)
0.1 mg/m^ (continuous)
1.0 mg/m3 (continuous)
0.1, 1.25 or 1.5 mg/l
0.1 mg/l, 3 hr/day
(alternate days)
75 and 250 ppm, 7 hrs/day
5 days/week
75 and 250 ppm, 7 hrs/day,
5 days/week
Duration
(days)
62
exposures
over 90 days
62
exposures
over 90 days
62
exposures
over 90 days
62
exposures
over 90 days
72-80
60
60
49-98
37 weeks
120
exposures
120
exposures
Effects Reference
None Honsanto, 1978
Weight loss; conjunctivitis; moribund at
31 days
Height loss; hypoactlvUy and conjunctivitis;
vacuolated hepatocytes; cytoplasmlc vacuolatlon
of renal collecting tubules; bilateral atrophy
of seminiferous tubules; lower total leukocyte
counts, elevated SAP, SGOT, SGPT; aplastlc bone
marrow; mortality 1n 5/8 dogs after 25-29 days
None Honsanto, 1978
Liver necrosis and regeneration; kidney Khanln, 1977
hyperplasla; encephalopathy; pneumonia
None Tarkhova, 1965
Inhibited chronaxla of antagonistic muscles
at 39 days; Increased blood chollnesterase
ChronaxImetMc Inhibition Plslaru, 1960
Inhibition of extensor tlblalls 7-14 weeks; Gabor and Raucher,
normal by 20 weeks 1960
Focal lesions of adrenal cortex; lesions 1n Dllley, 1977
tubules of kidneys; congestion of liver and
kidneys; decreased SGOT
Decreased SGOT after 24 weeks of exposure Dllley, 1977
-------
TABLE 7-4 (cont.;
Species
Route
Dose
Duration
(days)
Effects
Reference
Mouse oral (gavage)
I
->J
Rat
Oral (gavage)
60 mg/kg/day, 5 days/week 13 weeks
125 mg/kg/day, 5 days/week 13 weeks
250 mg/kg/day, 5 days/week 13 weeks
500 mg/kg day, 5 days/week 13 weeks
750 mg/kg/day, 5 days/week 10 weeks
60 mg/kg/day. 5 days/week 13 weeks
125 mg/kg/day, 5 days/week 13 weeks
250 mg/kg/day, 5 days/week 13 weeks
500 mg/kg/day, 5 days/week 13 weeks
750 mg/kg day, 5 days/week 13 weeks
one male with hepatic necrosis NTP, 1983
Increased liver weights 1n males one male
with hepatic necrosis
>50X reduction 1n weight gain, Increased
excretion of coproporphyrlns In females,
Increased liver weights, lesions of the
liver, kidney, bone marrow, spleen and
thymus
100X lethal to males within 1 week,
reduced body weight gains, polyuHa
In females, Increased liver weights,
lesions of the liver, kidney, bone
marrow, spleen and thymus.
100X lethal to male mice within 1 week
and to Female mice within 10 weeks,
lesions of the liver, kidney, bone marrow,
spleen and thymus at death
None NTP, 1983
None
Minimal centrolobular hepatocellular
necrosis
Decreased body weights gain, Increased
GGTP and alkaline phosphatase In females.
Increased excretion of porphyrlns, con-
trolobular hepatocellular necrosis,
nephropathy 1n males, myelold depletion
of bone marrow.
Decreased body weight gain and survival
of animals, hematologlc effects, Increased
GGTP and alkaline phosphatase 1n females,
polyurla In males, Increased excretion of
porphyrlns, centrolobular hepatocellular
necrosis, nephropathy, lymphold depletion
of thymus and spleen, myelold depletion of
bone marrow.
-------
TABLE 7-4 (cont.)
Species Route Dose
Dog oral (capsule) 27.3 mg/kg/day
54.6 mg/kg/day
272.5 mg/kg/day
i
GO Rat oral (diet) 12.5 or 50 mg/kg/day
100 mg/kg/day
250 mg/kg/day
Rat oral (diet) 14.4 mg/kg/day
144 and 288 mg/kg/day
Duration
(days)
90
90
90
93-99
93-99
93-99
192
192
Effects
None
Diarrhea and vomiting; conjunctivitis
4/8 died 1n 3-5 weeks; Increased Immature
leukocytes; elevated SCOT and SAP, bH1rub1n
and cholesterol; low blood sugar; hlstopatho-
loglc changes 1n liver, kidneys, spleen, and
seminiferous tubules
None
Increased liver and kidney weights
Increased liver and kidney weights;
retarded growth 1n males
None
Increased liver and kidney weights;
Increased salivation and hair loss
Reference
Monsanto, 1967a
Honsanto, 1967b
Irish, 1963
aSource: Updated from U.S. EPA, 1980a
bl ppm -4.60 mg/m1, 1 mg/l -219 ppm (Irish, 1963)
-------
Wonsanto (1978b) exposed by Inhalation Charles River albino rats to
monochlorobenzene at 0, 0.76, 1.47 and 2,00 mg/fc (0, 165, 319 and 434
ppm), 6 hours/day, 5 days/week for 62 exposures. Fifteen rats of each sex
were exposed at each dose level. Erythema and hair loss were noted 1n 2 of
30 animals at the lowest dose. Hematology, clinical chemistry values and
urlnalysls parameters were found to be similar between the treated and
control groups, and no hlstopathologlc changes attributable to monochloro-
benzene were found.
Beagle dogs exposed under the same regimen as the rats had toxic mani-
festations. Although no effects were noted at 0.75 mg/st, at 1.5 mg/a,, 2
of 8 dogs were moribund and sacrificed at 30 days; they were hypoactlve, had
decreased weight gain, and conjunctivitis. No clinical or hlstopathologlc
examination was made on this group. At the 2.0 trig/a, level, all the dogs
displayed weight loss, hypoactlvlty, and conjunctivitis. The mean leukocyte
counts of these dogs were lower than In controls at 45 and 90 days, and SAP
and SSOT were elevated at 38 days. Five dogs were moribund and therefore
sacrificed between days 25 and 38. Hlstopathologlc examination revealed
vacuollzatlon of hepatocytes 1n 5 of 8 dogs, aplastlc bone marrow in 5 of 8
dogs, abnormalities of collecting tubules of the kidneys In 4 of 8 dogs, and
bilateral atrophy of seminiferous tubules 1n 2 of 4 dogs.
Tarkhova (1965) exposed by Inhalation adult male rats to monochloroben-
zene at 0.1 or 1.0 mg/m3 (0.02 or 0.2 ppm) for 60 days of continual
exposure. No effects were seen at the lower level, but neurotoxlc effects
were noted at the higher level. In the high dose group, the chronaxy ratio
of antagonistic muscles was reversed at day 39 (I.e., the conduction speeds
of nerve Impulses to sets of flexor and extensor muscles had changed). Blood
7-19
-------
chollnesterase was Increased before the chronaxlmetrlc changes developed.
Similar neurotoxlc effects 1n rats were reported by Plslaru (1960) and Gabor
and Raucher (1960).
Subchronlc toxlclty studies regarding the effects of monochlorobenzene
administered to rats and dogs via gavage (oral administration) have been
reported by Monsanto (1967a,b). Male and female rats (18 of each sex 1n
each group) were dosed with 0, 12.5, 50, 100 or 250 mg/kg monochlorobenzene
1n corn oil for 5 days/week for 13 weeks. There were no effects on mortal-
ity, no clinical signs of abnormality and no hlstopathologlc lesions. A
slight decrease 1n growth rate over controls 1n males receiving the highest
dose level and a dose-related Increase 1n salivation 1n the animals were
noted.
Groups of 4 male and 4 female beagle dogs were given repeated doses of
27.3, 54.6 and 272.5 mg/kg of monochlorobenzene by capsule for 5 days/week
for 13 weeks. At the highest dose, two animals died and two were moribund
and sacrificed 1n the Interval between 14 and 21 doses. All animals given
doses of 272.5 mg/kg had weight loss and hlstologlc changes 1n the liver,
kidneys, gastrointestinal mucosa and hematopoletlc tissues. Minimal hlsto-
loglc changes were seen at 54.6 mg/kg, and no effects were noted at the
lowest dose. Animals that survived the higher dose had Increases In SGPT,
SAP, b1!1rub1n and cholesterol.
Subchronlc toxldty studies on monochlorobenzene were conducted under
the auspices of the National Toxicology Program (NTP, 1983b). The Investi-
gations were completed using 10 male and 10 female B6C3F mice and using
10 male and 10 female F344/N rats. The monochlorobenzene was administered
by gavage using a corn oil vehicle, 1n a volume of 5 ma/kg bw, 5 days/week
for 13 weeks. The monochlorobenzene doses used were 0, 60, 125, 250, 500
and 750 mg/kg bw.
7-20
-------
The mouse study resulted 1n 13-week survival rate of 100% (10/10), 100%
(10/10), 100% (10/10), 44% (4/9), 0% (0/10) and 0% (0/10) 1n male mice and
90% (9/10), 100% (10/10), 100% (10/10), 60% (6/10), 30% (3/10) and 0% (0/10)
1n female mice for the 0, 60, 125, 250, 500 and 750 mg/kg dose groups,
respectively (NTP, 1983b). Body weight gains during the 13 weeks were
decreased when compared with control animals 1n the surviving male mice, 27%
for the 60 and 125 mg/kg groups, and 82% for the 250 mg/kg group. A
decrease 1n body weight gains 1n surviving female mice was seen only 1n the
250 and 500 mg/kg dose groups (50% decrease 1n both groups). No clear
compound-related effects were found 1n the surviving monochlorobenzene-
treated mice from the hematologlc and clinical analyses performed. PolyuMa
was noted 1n the 750 mg/kg male group and the 500 mg/kg female group.
Significantly Increased excretion of coproporphyrlns were observed 1n sur-
viving female mice receiving 250 and 500 mg/kg. No changes 1n liver
porphyrln concentrations were observed 1n any of the male or female mice.
At sacrifice Increased liver weights were observed 1n surviving male mice at
125 and 250 mg/kg and surviving female mice at 250 and 500 mg/kg. Dose
dependent monochlorobenzene-lnduced Injury was revealed after hlstologlc
examination of liver, kidney, bone marrow, spleen and thymus. Except for
two male mice each with hepatic necrosis 1n the 60 and 125 mg/kg dose
groups, the observed tissue Injuries, which were graded as severe, only
occurred 1n the 250, 500 and 750 mg/kg dose groups. The liver lesions
consisted of focal hepatocytlc necrosis and centrllobular hepatocyte degen-
eration at 250 mg/kg and centrllobular hepatocellular necrosis at 500 and
750 mg/kg dose levels. Nephropathy was observed 1n female mice at 250 mg/kg
dose, and 1n male mice at 250, 500 and 750 mg/kg doses. Both sexes of mice
had myelold depletion of the bone marrow at doses >250 mg/kg. Doses of >250
7-21
-------
mg/kg caused necrosis of the thymus and doses of >500 mg/kg caused lymphold
depletion 1n the thymus. Based on these results, 60 mg/kg 1n corn oil
should be considered a lowest-observed-adverse-effect level (LOAEL). But
this was the lowest dose used.
The rat study resulted 1n a 13-week survival rate of 90% (9/10), 100%
(10/10), 100% (10/10), 100% (10/10), 60% (6/10) and 10% (1/10) 1n male rats
and 100% (10/10), 100% (10/10), 100% (10/10), 100% (10/10), 70% (7/10) and
20% (2/10) 1n female rats for the 0, 60, 125, 250, 500 and 750 mg/kg dose
groups, respectively (NTP, 1983b). Body weight gains over the 13-week
period were depressed by 10% or more 1n the male rats receiving doses >250
mg/kg and 1n female rats receiving 500 and 750 mg/kg doses. The only
hematologlc effects noted were at the 750 mg/kg dose level 1n surviving
males (Increased retlculocyte percentage) and females (decreased white blood
cell count). The only consistent effects observed 1n the serum chemistries
were slightly Increased activities of y-glutamyl transpeptldase and
alkaline phosphatase 1n female rats receiving 500 and 750 mg/kg. The 750
mg/kg male rats were observed to have a doubling of their 24-hour urine
output. Increased urinary excretion of uroporphyrlns was observed 1n male
rats at 750 mg/kg dose and of coproporphyrins 1n male rats at 500 and 750
mg/kg doses and 1n female rats at 500 mg/kg dose. No changes were observed
1n hepatic porphyMn levels. At sacrifice, monochlorobenzene-related
hlstologlcal changes were found 1n the liver, kidney, bone marrow, spleen
and thymus. Liver lesions were classified as centrllobular hepatocellular
necrosis (minimal at 250 mg/kg, minimal to moderate at 500 mg/kg, and moder-
ate at 750 mg/kg for both sexes of rats). M1ld to moderate nephropathy was
observed 1n male and female rats at 750 mg/kg and 1n male rats at 500 mg/kg.
Both male and female rats exhibited lymphold depletions of the thymus and
7-22
-------
spleen at the 750 mg/kg dose and myelold depletion of the bone marrow at the
500 and 750 mg/kg doses. From these rat data the lowest-observed-adverse-
effect level (LOAEL) Is 250 mg/kg and the NOEL 1s 125 mg/kg when given 1n
corn oil.
7.3.3. Chronic Toxldty. Two-year chronic bloassay studies using
monochlorobenzene were conducted under the auspices of the National Toxico-
logy Program (NTP, 1983b). The Investigations were conducted using 50 male
and 50 female B6C3F, mice and 50 male and 50 female F344/N rats. Mono-
chlorobenzene was administered by gavage In a corn oil vehicle, at a volume
of 5 mil/kg, 5 days/week for 103 weeks. The dosage groups used were
untreated, 0, 60 and 120 mg/kg for male and female rats and female mice, and
untreated, 0, 30 and 60 mg/kg for male mice. The test compounds were 99%
pure.
The mouse study revealed no monochlorobenzene-related clinical signs of
toxlclty or differences In mean body weights among test groups during the
105-week test period (exposure duration 103 weeks). Survival rates over the
test period In the male mice were 70% (35/50), 78% (39/50), 56% (28/50) and
58% (29/50) for the untreated control, vehicle control (0), 30 and 60 mg/kg
dose groups, respectively. Survival rates for the female mice were 74%
(37/50), 80% (40/50), 82% (41/50) and 76% (38/50) for the untreated
controls, vehicle controls (0), 60 and 120 mg/kg dose groups, respectively.
The only monochlorobenzene dosed group found to be significantly different
from controls 1n survival rates was the 30 mg/kg male group (p=0.031).
Hlstological findings of neoplasms will be discussed 1n Section 7.3.5,
Carcinogen1c1ty. No statistically significant Increased or decreased
Incidences 1n site-specific tumors or non-neoplast1c pathology were found 1n
either the male or female mice (NTP, 1983b).
7-23
-------
The rat study revealed no monochlorobenzene-related clinical signs of
toxldty during the 104-week study period (exposure duration 103 weeks).
The only differences noted 1n body weights during this study were Increased
body weights 1n the monochlorobenzene-treated females during the second
study year. The only significant differences 1n survival rates were
observed 1n the male 120 mg/kg dose group which had significantly reduced
survival rates (p=0.014 as compared with vehicle control). The survival
rates during this study were 68% (34/50), 78% (39/50), 64% (32/50) and 52%
(26/50) 1n male rats and 74% (37/50), 58% (29/50), 60% (30/50) and 62%
(31/50) 1n female rats for the untreated controls, vehicle controls (0), 60
and 120 mg/kg dose groups, respectively. H1stolog1cal findings of neoplasms
will be discussed 1n Section 7.3.5. Carc1nogen1c1ty. H1stolog1cal evalua-
tion of liver tissue provided equivocal evidence for mild monochlorobenzene-
Induced hepatocellular necrosis. The control rat livers were observed to
have more basophlUc cytoplasmlc changes than the monochlorobenzene-treated
rats (NTP, 1983b).
7.3.4. Hutagen1c1ty. Studies of the mutagenlcHy of monochlorobenzene
have yielded mixed results, with the greater proportion of the studies being
negative. These are summarized 1n Table 7-5.
7.3.5. Carc1nogen1c1ty. The only study available on the assay of mono-
chlorobenzene for carcinogenic potential 1s one conducted by the National
Toxicology Program (NTP), 1983. The conditions of this experiment were
described 1n Section 7.3.3.
7.3.5.1. RAT STUDY — In the case of the F344/N rats, dose selection
was made as a result of observations 1n the 13 week subchronlc study as
described 1n Section 7.3.2.
7-24
-------
TABLE 7-5
Mutagenldty Testing of Monochlorobenzene
Test System
AspeMqlllus nldulans
Salmonella strains
TA1S35, TA1537, TA1538,
TA92, TA98, TA100
Salmonella typhlmurlum
strains
Salmonella typhlmurlum
strains
Saccharomyces cerevlslae
Saccharomyces cerevlslae
Mouse lymphoma L5178Y
(forward mutation of TK)
DNA repair:
EscheMchla coll
(polAVpolA')
Bacillus subtnis
(rec~/rec*)
Streptomvces antlblotlcus
Metabolic Concentration
Activation
200 ng/ms.
+ 0.1-0.5 vsi/plate
-i- 100 jjg/plate
+ 150-3000 vg/plate
* 0.05-6%
4- 0.01-5 Mst/plate
0.001-0.1 yi/ma.
+ 0.0001-0.01 yi/mft
10-20 vft/plate
10-20 yil/plate
NR
Result
negative
negative
negative
negative
positive
negative
negative
negative
negative
positive
Reference
Prasad, 1970
Simmon et al.,
1979
Merck, 1978
DuPont, 1977
Simmon et al.,
1977
Monsanto, 1976
Monsanto, 1976
Simmon et al.,
1979
Simmon et al.,
1979
Kesklnova, 1968
NR = Not reported
-------
In the 2-year study survival of males at the 120 mg/kg groups was
significantly reduced when compared with vehicle but not with untreated
controls. Additionally, four accidental deaths occurred among the high dose
males, two at the low dose and one 1n a vehicle control male. Among females
there were seven accidental vehicle control deaths, four at the low dose and
two at the high dose.
The hlstopathology review 1n the 2-year study resulted 1n conflicting
Interpretation by different pathologlsts with respect to hepatocellular
necrosis, hepatocellular basophlUc cytoplasmlc changes and granulomatous
Inflammation. The findings of the 2 different reviewers are given, as they
appear 1n the NTP report, 1n Table 7-6. It 1s not clear whether these
differing Interpretations of non-neoplast1c lesions have any bearing on the
single set of results reported for neoplastlc nodules and carcinomas (Table
7-7). In males no carcinomas were observed 1n the treated groups, but there
was a statistically significant Increase 1n neoplastlc nodules 1n the high
dose group and a marginally significant dose-response trend. Neither neo-
plastlc nodules nor hepatocellular carcinoma were Increased 1n female rats.
In this study Interstitial cell tumors of the testls showed a signifi-
cant positive trend and the Incidence In the high dose group was signifi-
cantly different from the vehicle control 1n the life-table test. These
statistics are, however, without biological significance since the untreated
controls had Incidence of 100%. The vehicle control had 93.7% Incidence and
the low dose 97.7% while the high dose had 100%.
Both pituitary tumors (adenomas 1n female rats and combined adenomas and
carcinomas 1n male rats) and endometrlal stromal polyps of the uterus showed
significant negative trends.
7-26
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TABLE 7-6
Nonneoplastlc Lesions 1n F344 Rats Given Chlorobenzene
by Gavage for 2 Years*
Males
UC VC
Low
Dose
High
Dose
Females
UC VC
Low
Dose
High
Dose
Number of livers
examined
50 50
49
49
49 50
50
50
Hepatocellular
necrosis
Cytoplasmlc
basophlUa change
Inflammation
First Reading
214 5
25 27 6 3
993 0
38 27 18 10
23 21 11 11
Hepatocellular
necrosis
Cytoplasmlc
basophlUa change
Second Reading
325 1
28 40
12
12
1 1
43 34 26
18
*Source: NTP draft, 1983b
UC = Untreated controls; VC = vehicle controls
7-27
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TABLE 7-7
Statistical Comparisons of Liver Tumors In Male Rats Treated with
Chlorobenzene and Vehicle Controls3
Neoplastlc nodule
Overall
Adjusted
Terminal
Life Table
Incidental Tumor Test
Untreated
Control
4/50(8%)
10.4%
2/34(6%)
Vehicle
Control
2/50(4%)
4,5%
0.39(0%)
P=O.OOS
P=0.011
60 mg/kgb
4/49(8%)
12.5%
4/32(13%)
P=0.255
P=0.290
120 mg/kgb
8/49(16%)
29.3%
7/26(27%)
P=0.010
P=0.021
Coehran-Armltage Trend,
Fisher Exact Tests
Carcinoma
P=0.027
P=0.329
aSource: NTP draft, 1983b
^Results are compared with those of vehicle control.
N = Negative trend
P=0.043
Overall
Adjusted
Terminal
Life Table
Incidental Tumor Test
Coehran-Armltage Trend,
Fisher Exact Tests
Neoplastlc Nodule or Carcinoma
Overall
Adjusted
Terminal
Life Table
Incidental Tumor Test
Coehran-Armltage Trend,
Fisher Exact Tests
0.50(0%)
0.0%
0/34(0%)
4/50(8%)
10.4%
2/34(6%)
2/50(4%)
5.1%
2/39(5%)
P=0.139N
P=0.139N
P=0.098N
4/50(8%)
9.4%
2/39(5%)
P=0.033
P=0.054
P^O.121
0/49(0%)
0.0%
0/32(0%)
P=0.283N
P=0.283N
P=0.253N
4/49(8%)
12.5%
4/32(13%)
P=0.532
P=0.570
P=0.631
0/49(0%)
0.0%
0/26(0%)
P=0.331N
P=0.331N
P=0.253N
8/49(16%)
29 . 3%
7/26(27%)
P=0.048
P=0.083
P=0.168
7-28
-------
In the F344/N rats, therefore, the significant Increase 1n neoplastlc
nodules 1n the liver of male animals at the 120 mg/kg/day dose group
provides some evidence for tumor1gen1dty of monochlorobenzene.
7.3.5.2. HOUSE STUDY — The choice of dose for the chronic study 1n
mice was based on the results of a 13-week subchronlc test as described 1n
Section 7.3.2. On the basis of these data 1t could be concluded that doses
up to 120 mg/kg probably could have been tolerated 1n the chronic study of
male mice, whereas only 60 and 30 mg/kg were actually used. However, the
NTP draft document (dated Feb. 28, 1983} stated that "doses of 30 and 60
mg/kg were selected for male mice because of a perceived greater suscepti-
bility of this sex to the toxic effects of chlorobenzene".
The survival and body weight data 1n males during the chronic study also
suggest that larger doses could have been tolerated. Body weights 1n both
dose groups and survival 1n the high dose group were comparable to controls.
Although survival was reported to be significantly reduced 1n the low dose
group (30 mg/kg), two animals that died had foreign material In the lungs,
suggesting that gavage errors rather than toxldty was responsible for the
reduced survival 1n that group. These two animals were Included as deaths
from natural causes.
After hlstopathologlcal analysis the NTP found that both tumor Incidence
and non-neoplast1c pathology were comparable to controls at all sites 1n
both male and female treated groups. The test In mice therefore provided no
evidence of carclnogenlcity at doses as high as 60 mg/kg. Note, however
that the F344/N rats did not develop neoplastlc nodules until the dose was
as high as 120 mg/kg.
In summary, the evidence for the carclnogenlcity of monochlorobenzene
from the NTP study on F344/N rats and B6C3F-J mice consists of the finding
7-29
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of a significant Increase 1n neoplastlc nodules 1n the Hver 1n male rats
that received 120 mg/kg, for 5 days/week for 2 years. If the IARC criteria
for classifying carcinogens were used, this evidence would be characterized
as limited to Inadequate In animals. Since there 1s no human evidence
relating to carc1nogen1c1ty, the overall IARC classification 1s category 3,
and no conclusions can be made concerning the carclnogenlclty of monochloro-
benzene 1n humans.
7.3.6. Reproductive and Teratogenlc Toxldty. Honsanto Company (1978)
reported effects on the gonads of dogs exposed to monochlorobenzene vapor at
0, 0.76, 1.47 and 2.0 mg/s, for 6 hours/day, 5 days/week for a total of 62
exposures. Two of four male dogs 1n the high dose group developed bilateral
atrophy of epithelial tissue 1n the seminiferous tubules. These effects are
consistent with an earlier Monsanto (1967a) study where four male and four
female dogs were orally given monochlorobenzene at 27.3, 54.6 and 272.5
mg/kg/day doses for 13 weeks. Three of the four male dogs 1n the high dose
group had decreased spermatogenesls and this group also had tubular atrophy
and epithelial degeneration. These effects, however, were seen only at
levels sufficiently toxic that the dogs died or were moribund.
Rats exposed to monochlorobenzene vapor at 0, 0.76, 1.47 and 2.0 mg/ft,
for 6 hours/day, 5 days/week for a total of 62 exposures showed less
definite gonadal responses (Honsanto, 1978). The 2.0 mg/st exposed female
rats exhibited significantly higher gonad-to-body-we1ght ratio when compared
to control females.
No studies regarding the teratogenldty of monochlorobenzenes were
available for review.
7.4. INTERACTIONS
Monochlorobenzene produces a variety of alterations 1n enzyme function
and would, therefore, be expected to Influence the metabolism and toxlclty
7-30
-------
of a variety of compounds or vice-versa. Shelton and Weber (1981) Investi-
gated the hepatotoxldty of a mixture of CC1. and monochlorobenzene (1:38
molar ratio) to male CF-1 mice. The mixture was given 1ntraper1toneally In
corn oil (0.01 mfc/g bw). For the mixture of CC1. and monochlorobenzene,
the plasma anallne amlnotransferase dose-response curve did not deviate from
that predicted on the basis of dose addition.
7.5. SUMMARY
Acute exposure to monochlorobenzene by Inhalation causes sensory Irrita-
tion of the respiratory system after a few minutes; exposure for several
minutes to several hours causes narcosis and central nevous system depres-
sion, which can result 1n death. It 1s also toxic by the oral or parenteral
routes. Systemic effects of acute toxic doses Include kidney damage.
Subchronlc Inhalation exposure at 1.0 mg/m3 (contlnously for 60 days)
causes neurotoxlc effects In rats, an Increase 1n blood chollnesterase and
abnormal chronaxla of the muscles. Repeated exposure of rats to monochloro-
benzene at 250 ppm (1157 mg/m3) causes slight changes In the liver,
kidneys and adrenal cortex. Repeated oral dosing of rats or dogs (100-200
mg/kg/day) causes some toxic manifestation 1n the liver and kidneys.
Gavage administration of monochlorobenzene to mice and rats 5 times/week
for 13 weeks resulted 1n Increased mortality 1n the higher dose groups (>250
mg/kg), urinary porphyria and dose-dependent Injury to the liver, kidney,
bone marrow, spleen and thymus. A set of similar studies were conducted 1n
mice and rats for 2 years and resulted 1n some Increased mortality In the
male monochlorobenzene exposed groups when compared with controls. Only
equivocal evidence for mild monochlorobenzene-lnduced hepatocellular
necrosis was found 1n rats.
7-31
-------
Although one study 1n Streptomyces found monochlorobenzene to Induce
reversion to vitamin B, prototrophy and one study 1n Saccharomyces cere-
vlslae showed Increased mUotlc crossing over (Indication of ONA damage),
several other studies with bacterial, fungal and mammalian tissue culture
K
systems were negative. The carcinogenic activity of monochlorobenzene was
tested 1n the NTP bloassay program 1n two rodent species at doses of 60 and
120 mg/kg bw/day 1n male and female rats and female mice, and at 30 and 60
mg/kg bw/day 1n male mice. The significantly Increased Incidence of
neoplastlc nodules 1n the livers of high dose males provided some, but not
definitive, evidence for carcinogenic activity of monochlorobenzene.
Repeated exposures to monochlorobenzene at 2.0 mg/a, (vapors) or 272.5
mg/kg/day (oral) were found to cause atrophy of the epithelial tissue of the
seminiferous tubules and decreased spermatogenesls 1n male dogs and rats and
Increased gonad weight/body weight ratios 1n female rats. These effects 1n
dogs, however, were seen only at levels sufficiently toxic that the dogs
died or were moribund.
7-32
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8. DICHLOROBENZENES
The 1983 annual production of dlchlorobenzenes 1n the United States 1s
estimated to be between 46.7 and 50.2 million kilograms (U.S. EPA, 1983).
These materials are used primarily as fumlgants, Insecticides, solvents, dye
carriers and space deodorants (Hawley, 1977). Measurable levels of
dlchlorobenzenes have been reported 1n ambient urban and rural air and 1n
samples of Indoor air, 1n ground, surface and wastewater and 1n runoff from
hazardous waste sites (see Section 4.3.). Residues have been found 1n fish
and other aquatic organisms and 1n samples of human fat, blood, breath and
urine (see Section 4.3.3.). Human exposure 1s most likely through the
Inhalation of air and 1ngest1on of contaminated food and drinking water.
8.1. PHARMACOKINETICS
8.1.1. Absorption. The dlchlorobenzenes have low water solubility and
high Upld solubility and therefore are likely to diffuse through most bio-
logical membranes, Including the surfaces of the lungs and gastrointestinal
tract and the skin. The absorption of dlchlorobenzenes by humans 1s
Indicated by poisonings that have resulted from exposures by Inhalation or
1ngest1on. Quantitative studies of the absorption of dlchlorobenzenes 1n
humans and animals are lacking. The available data Indicate that absorption
does occur fairly rapidly through the lungs and gastrointestinal tract.
Skin absorption has not been tested adequately.
Twenty-three cases of poisoning by dlchlorobenzenes have been reported
1n the available literature and provide evidence of human absorption
(Downing, 1939; PerMn, 1941; Petit and Champeix, 1948; Sumers et a!., 1952;
Weller and CrelUn, 1953; Cotter, 1953; Hallowell, 1959; Frank and Cohen,
1961; Gadrat et al., 1962; Nalbandlan and Pearce, 1965; Glrard et al., 1969;
Campbell and Davidson, 1970; Ware and West, 1977; Harden and Baetjer, 1978).
8-1
-------
Of these cases, 5 Involved 1,2-d1chlorobenzene as the principal or signifi-
cant source of exposure and 11 Involved l,4-d1ehlorobenzene. Inhalation was
the primary route of exposure for 17 of the cases and 3 Involved 1ngest1on.
Three of the cases also mentioned previous dermal exposures that may have
contributed to the reported Intoxication.
Hawkins et al. (1980) exposed ten female CFY rats to a nominal air con-
centration of 1000 ppm of 1»4-d1chloro-[l4C]benzene, 3 hours/day for up to
10 days. In parallel experiments, groups of 20 female CFY rats were given
dally oral or subcutaneous doses (250 mg/kg/day) of 1,4-d1chloro-[l4C]~
benzene dissolved 1n sunflower oil. Twenty-four hour tissue concentrations
of 14C were similar for each treatment route, with the highest concentra-
tions occurring 1n the fat, kidneys, liver and lungs. l,4-D1chlorobenzene
appears to be well absorbed through both the lungs and gastrointestinal
tract; however, no quantitative measures of absorption were attempted.
Klmura et al. (1979) administered 200 or 800 mg/kg of 1,4-d1chloroben-
zene 1n corn oil orally to male Wlstar rats and monitored the appearance of
the chemical 1n blood, adipose, kidney, Hver, lung, heart and brain tissue.
At the first time point, 30 minutes after dosing, all these tissues con-
tained measurable amounts of dlchlorobenzene, with liver and adipose tissue
having 2 and 10 times the concentrations seen In the blood, respectively.
Three other studies have suggested that dlchlorobenzenes can be almost
completely absorbed from the gastrointestinal tract. Azouz et al. (1955)
dosed chinchilla rabbits 1ntragastr1cally with 1.5g 1,4-d1chlorobenzene/
rabbit 1n olive oil and did not detect any of the compound 1n the feces,
Implying that under the conditions of this study, a total absorption had
occurred. Hawkins et al. (1980) administered a single dose of labeled
1,4-d1chlorobenzene (250 mg/kg) to rats with cannulated bile ducts, which
8-2
-------
prevented fecal excretion of the absorbed metabolized compound. During the
following 24 hours, 9% of the label was present 1n the feces, representing
the unabsorbed portion of the dose. 1,2-D1chlorobenzene and other organic
contaminants of water were administered to rats 1n the diet at levels of
0.4-2 mg/kg/day (Jacobs et al., 1974a,b). The accumulation of the compound
1n several tissues Indicated that absorption occurs after the 1ngest1on of
low levels of 1,2~d1chlorobenzene.
Riedel (1941) has investigated the dermal absorption of I,2~d1ehloro~
benzene In rats. No quantitative measurements were made; however, five
applications were lethal when the compound was applied directly to a 10
cm2 area of abdominal skin.
In summary, we may say that absorption of dlchlorobenzenes can occur
through the lungs, skin and gastrointestinal tract. Quantitative studies of
absorption through the lungs and gastrointestinal tract are lacking, as well
as good quantitative studies on dermal absorption.
8.1.2. Distribution. The low water and high I1p1d solubility of the
dlchlorobenzenes facilitates the diffusion of the dlchlorobenzenes through
membranes and therefore enhance their tissue distribution. Several studies
in animals have quantified the degree and time course of the distribution of
dichlorobenzenes after Inhalation or oral 1ngest1on and Indicate rapid
distribution to blood, adipose, kidney, liver, lung, heart, brain and muscle
tissue.
Hawkins et al. (1980) investigated the distribution and excretion of
1,4-d1chlorobenzene 1n adult female CFY rats (derived from Sprague-Dawley
rats) after repeated Inhalation, oral and subcutaneous doses. Radioactive
labeled 1,4-d1chlorobenzene was administered by exposing groups of 10 rats
to the compound at an atmosphere of 1000 ppm for 3 hours/day for 10 consecu-
8-3
-------
tlve days or by oral or subcutaneous doses of 250 mg/kg/day for 10 days.
The Investigators reported tissue concentrations after 2, 4, 6, 8 and 10
doses. Radlolabel was widely distributed following each route of adminis-
tration, with the highest concentrations occurring 1n fat, kidney, liver and
lungs (Table 8-1).
Klmura et al. (1979) also provided Information on the distribution of
1,4-d1chlorobenzene. Tissue levels of the compound were monitored at
Intervals from 30 minutes to 120 hours in male Wlstar rats given a single
200 mg/kg oral dose 1n corn oil after a 16-hour fast. At the first
Interval, fat and liver levels were 10 and 2 times the blood levels (~9
vg/mi), respectively, with lower concentrations of dlchlorobenzene
appearing 1n all of the other tissues examined (kidney, lung, heart and
brain). Levels 1n fat, kidney and liver tissue peaked between 6 and 12
hours (at -50, 2 and 0.5 times blood levels, respectively) and thereafter,
decreased along with the levels 1n the other tissues. After 48 hours, con-
centrations of 1,4-d1chlorobenzene were below the detection limit 1n all
tissues examined, except for fat tissue, which had detectable levels equal
to approximately one-fiftieth of peak concentrations at 120 hours post-
administration.
Tissue distribution after subchronlc feeding of 1,2-d1chlorobenzene was
Investigated by Jacobs et al. (1974a,b), who administered the compound 1n a
mixture of other organic chemicals at doses of 0.4, 0.8 and 2 mg/kg/day for
4-12 weeks to rats. A dose-related accumulation of 1,2-d1chlorobenzene was
reported 1n abdominal and renal adipose tissue to an extent greater than
that seen 1n liver, heart and blood tissues.
Studies of tissue distribution of dlchlorobenzenes after repeated Inha-
lation exposure and single and subchronlc oral exposure Indicated that the
chemicals appeared 1n all of the major tissues soon after dosing with the
8-4
-------
TABLE 8-1
Tissue Concentrations of l,4-D1chlorobenzene 1n Adult Female CFY Ratsa«b'c
(ppm)
Number
of Doses
CD
i
en
2
4
6
8
10
Liver
Inhalation0 Oral
14
22
28
16
18
11
18
14
15
9
Subcutaneous
21
22
24
21
20
Inhalation
24
40
43
28
27
Kidneys
Oral Subcutaneous
2?
29
23
18
16
30
32
47
41
32
Inhalation
9
12
11
10
10
Lunqs
Fat
Oral Subcutaneous Inhalation Oral
7
13
10
11
9
18
12
14
21
17
418
579
597
433
337
218
369
170
131
257
Subcutaneous
372
302
269
554
383
aSource; Adapted from Hawkins et al., 1980
••Feaale rats were exposed dally to 1,4-dlchlorobenzene via: Inhalation, 1000 ppm for 3 hours/day; oral, 250 mg/kg In sunflower oil;
subcutaneously, 250 mg/kg 1n sunflower oil and killed 24 hours after last dosing.
C¥a1ues represent the average from two animals
-------
highest levels, In descending order, 1n adipose, kidney, liver and lung
tissue. Peak concentrations are reached 1n all tissues within 4-12 hours,
followed by almost total elimination (Section 8.1,4.). The pattern of dis-
tribution after Inhalation, subcutaneous and oral administration 1s similar.
8.1.3. Metabolism. The metabolism of the dlchlorobenzenes has been
Investigated primarily In rabbits and rats; few data were available on
metabolism In humans. Several studies have shown the primary metabolites to
be dlchlorophenols that are conjugated with glucuronlc and sulfurlc adds
and excreted. Formation of the dlchlorophenols from 1,2- and l,3-d1chloro-
benzene appears to Involve epoxldase enzymes and the formation of arene
oxide Intermediates.
Azouz et al. (1955) studied the metabolism of 1,2- and 1,4-d1chloroben-
zene In rabbits given oral doses of 500 mg/kg suspended 1n water for
l,2-d1chlorobenzene and 1n olive oil for 1,4-d1chlorobenzene. The compounds
were metabolized primarily through oxidation to 3,4-d1chlorophenol (from
1,2-d1chlorobenzene) and 2,5-d1chlorophenol (from 1,4-d1chlorobenzene) and
excreted 1n the urine 1n the form of glucuronlc and sulfurlc add conjug-
ates. Minor metabolites of 1,2-d1chlorobenzene Included the 4,5- and 3,4-
dlchlorocatechols and 3,4-d1chlorophenylmercaptur1c add; a minor metabolite
of 1,4-d1chlorobenzene 1s 2,5-d1chloroqu1nol. Metabolism and complete
elimination required 5-6 days for l,2-d1chlorobenzene and >6 days for
1,4-dlchlorobenzene. Klmura et al. (1979) found similar results 1n male
Wlstar rats; oral administration of 200 or 800 mg/kg of 1,4-d1chlorobenzene
resulted 1n the formation of one major metabolite, 2»5-d1chlorophenol, and
two minor sulfur-containing metabolites (<0.03% of the total dose). These
two compounds, Identified as 2,5-d1chlorophenol methoxy sulfoxlde and
2,5-d1chlorophenol methyl sulfone, were excreted over a 5-day period and
were detected 1n blood, fat, kidney and liver tissues.
8-6
-------
Hawkins et al. (1980) Investigated the metabolism of radlolabeled
1,4-d1ehlorobenzene 1n female CFY rats after repeated Inhalation (1000 ppm
for 3 hours/day), oral or subcutaneous (250 mg/kg/day) exposures. After
dosing for 10 consecutive days, all of the label was metabolized and elimi-
nated within 192 hours (8 days). Both routes of exposure resulted 1n simi-
lar urinary and biliary metabolites, primarily 2»5-d1chlorophenol sulfate
(46-54% of the total excreted) and 2,5-d1chlorophenol glucuronlde 1n the
urine (31-34%) and bile (30-42%). Two minor metabolites were Identified as
dlhydroxydlchlorobenzene and a mercapturlc add of l»4-d1chlorobenzene.
Parke and Williams (1955) studied the metabolism of 1,3-d1chlorobenzene
1n rabbits and found dlchlorophenol to be the major metabolite, accounting
for 40% of the total amount of excreted metabolites. 2,4-D1chlorophenyl-
mercapturlc add and 3,5-d1chlorocatechol were also detected. No analogous
studies have been conducted 1n humans although Pagnotto and Walkley (1966)
reported that 2,5-d1chlorophenol was present 1n the urine of men occupation-
ally exposed to 1,4-d1chlorobenzene by Inhalation. Several studies have
Indicated that the dlchlorobenzenes are also capable of Inducing hepatic
mlcrosomal enzymes and enhancing the synthesis of porphyrlns. R1m1ngton and
Zlegler (1963), Poland et al. (1971) and Ar1yosh1 et al. (1981) have report-
ed the Induction of 5-amlnolevul1n1c add synthetase 1n rats by dally
doses of 250-1000 mg/kg of dlchlorobenzenes. This enzyme Is Involved 1n the
rate-limiting step of the synthesis of porphyrlns and Us Induction 1s
necessary for an Increase 1n the activity of cytochrome P-450 and other
xenoblotlc metabolizing enzymes.
8.1.3.1. COVALENT BINDING — Metabolism of dlchlorobenzenes results
In the formation of reactive species, which may bind covalently to cellular
macromolecules. This binding may lead to some toxic effects of the
8-7
-------
dlchlorobenzenes. Reid and Krishna (1973) studied the relationship between
the binding of metabolites of halogenated aromatic hydrocarbons and the
Induction of hepatic necrosis. Labeled bromobenzene, 1,2- and 1,4-dlchloro-
benzene, as well as other aromatic compounds, were Injected 1ntraper1toneal~
ly Into Sprague-Dawley rats 1n 0.5 mmol/kg doses. A correlation between
covalent binding of bromobenzene to protein and the time course and degree
of hepatic centrolobular necrosis was established. 1,2-D1chlorobenzene also
was found to bind to Hver protein and the binding was enhanced by pretreat-
ment with phenobarbltal. 1,4-D1chlorobenzene showed little binding. The
authors Interpreted these results to mean that the hepatic Injury Induced by
1»2-d1chlorobenzene was a result of the binding to protein of reactive
Intermediates whose synthesis was Increased by the Induction of hepatic
xenob1ot1c-metabo!1z1ng enzymes. l,4-D1chlorobenzene Is less hepatotoxlc
than 1,2-d1chlorobenzene and does not bind to the same degree. Similar
results were obtained for the bronchlolar necrosis occurring 1n lung tissue
(Reid et a!., 1973).
8.1.4. Excretion. Hawkins et al. (1980) measured the excretion of 14C
1n female CFY rats following whole body exposure by Inhalation (1000 ppm, 3
hours/day, 2-10 days), by oral (250 mg/kg/day) or subcutaneous (250 mg/kg/
day) routes of 1,4-d1chloro[l4C]benzene. Excretion occurred primarily via
the urine (91-97% of the total excreted) over a 5-day period after repeated
doslngs had stopped, with only minor amounts occurring In the feces and
expired air. Following a single oral dose to blle-duct-cannulated rats,
46-63H of the 14C excreted during the first 24 hours was found 1n the
bile. This Implies that enterohepatlc redrculatlon occurs to a major
extent with this compound. Excretion seemed to Involve a rapid Initial
phase followed by a slower extended excretion phase.
8-8
-------
Klmura et al. (1979) observed similar excretion patterns 1n male Wlstar
rats. They suggested that the prolonged excretion of 1,4-d1chlorobenzene
resulted from the release of unmetabollzed material from fatty depots and
the slow excretion rates for 2,5-d1chlorophenyl methyl sulfone and
2,5-d1ehlorophenyl methyl sulfoxlde, two metabolites of 1,4-d1chlorobenzene.
Azouz et al, (1955) compared the excretion of 1,2~d1chlorobenzene and
1,4-d1chlorobenzene given to chinchilla rabbits by stomach tube In an olive
oil solution. Excretion rates were not determined; however, the excretion
of the l,2-1somer appeared to be complete within 5-6 days after dosing.
With the para Isomer, appreciable excretion of metabolites still occurred
after § days. Excretion of the meta Isomer 1n chinchilla rabbits was found
to be virtually complete within 5 days after dosing by stomach tube using an
olive oil solution (Parke and Williams, 1955). One study has suggested that
similar metabolic products occur 1n humans. Pagnotto and Walkley (1966)
reported that the appearance of dlchlorophenol 1n the urine of workers that
were exposed to 1,4-dlchlorobenzene began soon after exposure, peaked at the
end of the shift and continued for several days.
8.1.5. Summary. The available data for rats, rabbits and humans Indicate
that the dlchlorobenzenes are absorbed through the lungs, gastrointestinal
tract and Intact skin, though actual determinations of absorption rates were
not located. Once absorbed, through either Inhalation or 1ngest1on, the
dlchlorobenzenes are rapidly distributed to many tissues, Including blood,
adipose, kidney, liver, lung, heart, brain and muscle. Distribution Is
primarily to adipose tissue, which has Initial levels 10-32 times the blood
concentrations and to lung and kidney tissues to a greater extent than
liver, muscle and plasma. Single-dose and repeated exposures by both
Inhalation and 1ngest1on show similar patterns of distribution. Elimination
8-9
-------
of the dlchlorobenzenes and their metabolites occurs within 5-6 days after
exposure, although elimination from adipose tissue 1s slowest and 1,2-dl-
chlorobenzene and metabolites are eliminated slightly more rapidly than
l,4-d1chlorobenzene. The dlchlorobenzenes are primarily metabolized by
hydroxylatlon to their respective dlchlorophenols, which are excreted 1n the
urine 1n the form of glucuronlc and sulfurlc add conjugates. Some metabo-
lites are excreted 1n the bile, although the majority are then reabsorbed by
the enterohepatic pathway and reexcreted 1n the urine. Intermediates of the
metabolism of l,2-d1ehlorobenzene, possibly arene oxides and the metabolite
conjugates, bind to liver protein and may be Involved 1n the Induction of
hepatotoxldty.
8.2. EFFECTS ON HUMANS
8.2.1. Occupational Studies. One occupational study was available for
review. Zapata-Gayon et al. (1982) performed chromosomal studies on 8 males
and 18 females who were accidentally exposed to vapors of 1,2~d1chloroben-
zene for four 8-hour workdays. Karyotypes of cells from samples of periph-
eral blood from the exposed subjects were compared with those obtained from
16 controls (8 male, 8 female). Exposed subjects and controls had similar
occupational histories: all worked 1n a biological laboratory performing
electron microscope and tissue culture work. Recent history of prolonged
X-ray exposure, Infection or exposure to other toxic chemicals was not found
among the subjects. The exposure to l,2-d1chlorobenzene, which resulted
from Its use as a pest control 1n the basement of a one-story building,
caused dizziness, headache, fatigue, nausea and eye and nose Irritation 1n
all but four of the subjects. Karyotype analysis, performed Independently
by two cytogenetldsts, found that the total number of altered cells, Iden-
tified as having clastogenlc chromosomal alterations, was greater 1n the
17posed versus control groups (8.9 vs. 2.0%, p<0.001, multiple ch1-square
8-10
-------
tests). In addition, the total number of single chromosomal breaks (6,2 vs.
0.9%, p<0.001) and double breaks (6.4 vs. 1.6%, p<0.001) were different. A
follow-up study was conducted on 15 of the original exposed cases 6 months
after the Initial exposure. The Investigators reported that the number of
altered cells and single breaks was not significantly different (p<0.05)
from the original control frequencies, but that the number of double breaks
was Increased (3.7 vs. 1.6%, p<0.01). No analysis of the number of altered
cells/person was performed, although these data, reported 1n the form of a
histogram, showed distinct differences (Table 8-2). The Investigators also
noted the presence of other aberrations (polyploldy and ring formation) that
were not statistically significant.
8.2.2. Case Studies. Numerous case studies have been reported 1n the
literature Involving both long-term occupational exposure and accidental or
deliberate acute exposure. Of these cases (a total of 23), 17 have Involved
exposure primarily through Inhalation, 3 through 1ngest1on and 3 most likely
through dermal absorption. The principal agent 1n 16 of these exposures was
1,4-d1chlorobenzene; the remainder Involved 1,2-d1chlorobenzene or mixtures
of all three dlchlorobenzene Isomers. In all of these cases, toxic effects
have been reported 1n one or more of the following: liver; blood, Including
ret1culoendothel1al system; central nervous system; and respiratory tract.
A summary of these reports, which were compiled 1n U.S. EPA (1980c) with the
exception of Hardln and Baetjer (1978), Is given 1n Table 8-3.
Two surveys of the health of workers occupatlonally exposed to
1,4-d1chlorobenzene during Its manufacture have been reported. HolUngs-
worth et al. (1956) reported that periodic medical examinations showed no
evidence of Injury or adverse changes 1n hematology or eye lenses 1n workers
8-11
-------
TABLE 8-2
Chromosomal Alterations 1n Persons Accidentally
Exposed to l,2-D1chlorobenzene*
Number of Altered Cells per Person
0-1
2-3
4-5
>6
Percentaqe
Control
(n=16)
83
19
0
0
Exposed
(n=22)
5
35
29
31
*Source: Adapted from Zapata-Gayon et al.» 1982
8-12
-------
TABLE 8-3
Case Reports Involving Dlchlorobenzenes (DCS)*
Chemical/Mixture
Subject and Exposure
Effects
Reference
CO
i
1,2-OCB (vapor)
1,2-DCB solvent mixtures:
80* 1,2-OCB;
m 1,4-OCB;
2X 1,3-DCB
1,2-OCB solvent ralxure:
95X 1,2-OCB;
5X 1,4-DCB
1,2-OCB and other chlorobenzenes
1,2-DCB 1n a mixture
1,2-DCB {37X 1n solution)
1,2-DCB solvent mixture:
80% 1,2-OCB;
15X 1,4-OCB;
2X 1,3-DCB
1,4-DCB primarily
Sewage workers; occupational; Inhalation; efflu-
ent from dry cleaning establishment
Hale, 40 years; occupational; use of solvent to
clean equipment; chronic dally exposure probably
via Inhalation of vapors, and dermal absorption
from clothing
Female, IB years; occupational; chronic dally
Inhalation exposure to vapors as pressing-Ironing
worker
Hale, 60 years; occupational; filling barrels
with 1,2-OCB and other chlorobenzenes; chronic
Inhalation of vapors (last 3 years); perhaps
also skin contact
Male, 47 years; occupational; handling window
sashes dipped In mixture; chronic skin contact
(also Inhalation)
Female, 15 years; nonoccupatlonal; chronic
repeated dermal contact from compulsive use of
cleaning solution on clothing
Female, 55 years; nonoccupatlonal; chronic
repeated Inhalation of vapors from use of
solution to clean clothes; 1-2 l/year
Female, 30 years; occupational; chronic Inhala-
tion and dermal contact from 2 years of selling
mothballs and Insecticide products containing
1,4-DCB
Eye and upper respiratory
tract Irritation; vomiting
Heakness, fatigue; periph-
eral lymphadenopathy;
chronic lymphold leukemia
Severe acute hemolytk
anemia; leukocytosls, poly-
nucleosls; fatigue, nausea,
headache; bone marrow hyper-
plasla; possible Inherent
predisposing factor
Anemia
Contact eczematold derma-
titis on hands, arms, face,
erythema, edema; bullae In
response to skin test
Acute myeloblastlc leukemia
progressing to 100X leuko-
blastosls, hemorrhage and
death
Oupont, 1938
Glrard et al., 1969
Gadrat et al., 1962
Glrard et al., 1969
Downing, 1939
Glrard et al., 1969
Acute myeloblastlc leukemia Glrard et al., 1969
Heakness, nausea, spleno-
megaly, "severe hepato-
cellular derangement, and
ensuing portal hyper-
tension" with esophageal
varlces
Sumers et al., 1952
-------
TABLE 8-3 (cont.)
Chemical/Mixture
Subject and Exposure
Effects
Reference
1,4-DCB primarily
CD
1,4-DCB
1,4-DCB primarily
1,4-DCB
1,4-DCB
1,4-DCB
1,4-OCB
1.4-DCB
Female, 34 years; occupational; chronic Inhala-
tion from demonstrating 1,4-DC8 products 1n booth
1n department store
Hale, 52 years; occupational; chronic Inhala-
tion of high vapor levels 1n a fur warehouse
Female, 19 years; occupational; crushing, pouring,
sieving, filling containers; poor ventilation;
chronic Inhalation of vapors
Female, occupational; casting 1,4-DCB 1n molds;
chronic Inhalation
Hale, 20 years and workmates; occupational;
1,4-DCB manufacturing activities, 1-7 months
of exposure; Inhalation
Hale, 62 years; nonoccupatlonal; used "moth
killer" product In bathroom at home, chronic
Inhalation of vapors, and wearing of Impreg-
nated clothing (possible skin exposure)
Female, 36 years; nonoccupatlonal; use of
commercial moth killer In home (presumably
Inhalation of vapors)
Hale, 60 years; nonoccupatlonal; 3-4 months
exposure to "moth gas vapor" 1n home
Halalse, then acute nausea,
vomiting, headache, jaun-
dice, hepatomegaly, spleno-
megaly, esophageal vaHces,
and hemorrhoids; subacute
yellow atrophy and cirrhosis
of liver
Weakness, nausea, hemateme-
s1s, jaundice, emaciation,
petechla, hemorrhages;
hepatomegaly, splenomegaly,
hemorrhoids; protelnuHa,
b1!1rub1nur1a; hematuMa;
anemia, leukopenla; subacute
yellow atrophy of liver
Harked asthenia, dizziness,
weight loss; anemia and
reactlonal leukocytosls
Severe anemia
Weight loss, exhaustion,
and decreased appetite;
methemoglob1nem1a and
other blood pathologies
Asthenia, dizziness;
anemia, hypogranulocytosls
(similar to cases of In-
toxication by benzene)
Acute Illness with Intense
headache, profuse rhinitis,
peMorbltal swelling
Headache; weight loss;
diarrhea; numbness; clumsi-
ness; jaundice; enlarged
liver; anemia; neutropenla;
ascltes; death; acute
yellow atrophy of liver
noted at autopsy
Cotter, 1953
Cotter, 1953
Petit and Champelx, 194B
PerMn, 1941
Ware and West, 1977
PerMn. 1941
Cotter, 1953
Cotter, 1953
-------
TABLE 8-3 (cont.)
Chemical/Mixture
Subject and Exposure
Effects
Reference
1,4-OCB
Female, wife of above, nonoccupatlonal; prolonged
severe exposure to "moth gas vapor"
1,4-OCB
CD
i
1,4-OCB
1,4-OCB
1,4-DCB
Female, 53 years; nonoccupatlonal; used moth
eradlcator product heavily In home for 12-15
years, odor always apparent; chronic Inhalation
of vapor
Hale, 3 years; nonoccupatlonal; played with
canister of demothlng crystals, spreading on
floor, handling; Ingestlon, likely acute
Female, 21 years; nonoccupatlonal; Ingestlon
during pregnancy (pica) of toilet air freshener
blocks at rate of 1-2/week
Female, 19 years; nonoccupatlonal; Ingestlon
(pica), 4-5 moth pellets dally for 2.5 years
Gradual loss of strength
and weight, then abdominal
swelling and jaundice
before acute Illness; ele-
vated temperature and pulse,
dilated vessels, swollen
liver, toxic granulocytosls;
died 1 year later; acute
yellow atrophy of liver,
Laennec's cirrhosis and
splenomegaly noted at autopsy
Chronic progressive cough
and dyspnea with mucold
sputum, wheezing, fatigue,
diminished breath sounds
and rales; abnormal lung
field on X-ray; flbrotlc,
rubbery lung with hlsto-
loglc changes; diagnosis:
pulmonary granulomatosls
Llstlessness, Jaundice,
ollgurla, methemoglob1nur1a
and other urine abnormali-
ties, anemia, hypothermia;
diagnosis: acute hemolytlc
anemia
Fatigue, anorexia, dizzi-
ness, edema of ankles;
hypochromlc mlcrocytlc
anemia; bone marrow normo-
blastlc hyperplasla; diag-
nosis: toxic hetnolytlc
anemia; complete recovery
Increased skin pigmentation
1n areas 3-7 cm In diameter
on limbs; mental sluggish-
ness; tremor; upon with-
drawal, unsteady gait along
with decrease 1n pigmentation
Cotter, 1953
Heller and Crellln, 1953
Hallowell, 1959
Campbell and Davidson,
1970
Frank and Cohen, 1961
-------
TABLE 8-3 (cont.)
Chemical/Mixture
Subject and Exposure
Effects
Reference
1,4-DCB
Hale, 69 years; nonoccupatlonal; dermal exposure,
presumably Interrupted; episode precipitated by
use of chair treated with 1,4-DCB
CO
i
1,4-DCB (and naphthalene)
Female, 68 years; occupational; Inhalation and
dermal exposure to mothproofing agents for 1
month/year for 39 years
Dyspnea followed by stiff
neck; "tightness" 1n chest,
"gas pains" In abdomen;
symmetrical petechla and
purpura on extremities,
swelling discomfort; stool
occult blood positive,
blood cells 1n urine; and
Increased BUN; basophll
degranul. test positive
for 1,4-DCB; diagnosis:
allergic (anaphylactold)
purpura and acute glomer-
ulonephrltls
Aplastlc anemia
Nalbandlan and Pierce,
1965
Harden and Baetjer, 1978
•Source: U.S. EPA, 1980c
-------
exposed to airborne concentrations averaging 270-630 mg/m3. Workers com-
plained of eye and nose Irritation at levels >800 mg/m3. Another survey,
conducted at a 1,2-d1chlorobenzene manufacturing facility, reported ambient
levels of 6-264 mg/ma {90 mg/m3 average) {HolUngsworth et al., 1958).
Occasional medical examinations, Including hematology and urlnalysls,
revealed no evidence of Injury or adverse hematologlc effects attributable
to the exposure.
8.2.3. Summary. Ep1dem1olog1c data are Insufficient to evaluate
dose-response associations. Possible chronic effects of exposure to the
dlchlorobenzenes are Indicated by case reports of the chronic exposure of
Individuals, I.e., repeated exposures over a period of more than a year,
suggesting a common set of toxic effects, those of the ret1culoendothel1al
and hematopoletlc systems and those of the liver. Of the 23 cases 1n the
literature, 17 Involved pathological changes 1n the blood or Hver, Includ-
ing chronic lymphold leukemia, acute hemolytlc anemia, aplastlc anemia and
bone marrow hyperplasia. Although the exposures 1n these cases are not well
defined 1n terms of concentrations or duration of exposure and often
Involved other toxic substances, together they suggest a common pathologic
action of the dlchlorobenzenes on bone marrow and other organs of the blood-
forming system. The one available ep1dem1olog1c study (Zapata-Gayon et al.»
1982) supports this generalization 1n that the reported short-term exposure
to 1,2-d1chlorobenzene (8 hours/day for 4 days) produced alterations 1n the
chromosomes of leukocytes. This ep1dem1olog1c study did not establish an
association between chromosomal alterations and the pathologic changes that
characterize the case studies.
8.3. MAMMALIAN TOXICOLOGY
8.3.1. Acute Toxldty. Many studies have Investigated the acute toxldty
of 1,2- and 1,4-dlchlorobenzene, but no studies were available on
8-17
-------
1,3-d1chlorobenzene. In general, the acute toxic effects of 1,2- and
1,4-d1chlorobenzene have shown a similar profile of effects 1n all of the
species tested and depend to a certain degree on the route of administra-
tion. For oral administration, these effects Include, Initially, Increased
lacrlmatlon, salivation and excitation followed by ataxla, dyspnea and death
from respiratory paralysis, usually within 3 days. On autopsy, the animals
were found to have enlarged Hvers with necrotlc areas, submucosal hemor-
rhages of the stomach, necrotlc changes of the kidneys and brain edema.
After acute Inhalation, the toxic effects observed were eye and nose Irrita-
tion, liver and kidney necrosis and central nervous system depression.
Lethal doses for both oral and Inhalation routes for 1,2-d1chlorobenzene
tend to be one-half to two-thirds of the values for 1,4-d1chlorobenzene.
Acute dermal application of 1,2-d1chlorobenzene results 1n local Irritation
and absorption of amounts which can be lethal. Acute toxldty data for
1,2- and 1,4-d1chlorobenzene, as compiled by U.S. EPA (1980d), are given 1n
Tables 8-4 and 8-5, respectively.
HolUngsworth et al. (1956, 1958) determined the acute oral toxldty of
1,2-d1chlorobenzene (50% 1n olive oil) 1n 10 guinea pigs of mixed sex and
l,4-d1chlorobenzene (20 or 50% 1n olive oil) 1n rats and (50% 1n olive oil)
1n guinea pigs. The Intubation of guinea pigs with 1,2-dlchlorobenzene 1n
single oral doses of 800 mg/kg resulted 1n loss of body weight, but was
survived by all subjects, whereas 2000 mg/kg doses were fatal to all
subjects. Intubation of rats and guinea pigs with 1,4-d1chlorobenzene 1n
single oral doses of 1000 mg/kg and 1600 mg/kg bw, respectively, were
survived by all the test animals, while doses of 4000 mg/kg and 2800 mg/kg
bw were found to be lethal to rats and guinea pigs, respectively.
8-18
-------
TABLE 8-4
Acute Toxldty of 1,2-Dlchlorobenzene*
Species
Rat
Rat
Rat
Guinea pig
Guinea pig
CD
J_, Guinea pig
Rabbit
Rat
House
Guinea pig
Rat
Mouse
Rabbit
Route of
Administration
Inhalation
Inhalation
Inhalation
Inhalation
oral
oral
oral
oral
oral
oral
dermal
Intravenous
Intravenous
Concentration
or Dose
5872 mg/rn*
4243 mg/m'
3239 mg/ra«
4808 mg/m*
2000 rug/kg
800 mg/kg
1875 mg/kg
2138 nig/kg
2000 mg/kg
3375 mg/kg
unspecified
dally for 5
applications
520 mg/kg
330 mg/kg
Regimen
7 hours
7 hours
7 hours
24 hours
single
single
single
single
single
single
twice
single
single
Effects
lethal 1n 4/5
lowest lethal concentration
eye Irritation, CMS depression,
liver and kidney damage
lowest lethal concentration
100X mortality
weight loss
L050
LD50
L°50
LOfiO
lethality
lowest lethal concentration
lowest lethal concentration
Reference
HolUngsworth et al., 1958
Chrlstenson and Falrchlld,
1976
HolUngsworth et al., 1958
Chrlstenson and Falrchlld,
1976
HolUngsworth et al., 1958
HolUngsworth et al., 1958
Varshavskaya, 1967a
Varshavskaya, 1967a
Varshavskaya, 1967a
Varshavskaya, 1967a
Rledel, 1941
Chrlstenson and Falrchlld,
1976
Chrlstenson and Falrchlld,
1976
•Source: U.S. EPA, 1980d
-------
CD
ro
o
TABLE 8-5
Acute Toxldty of 1,4-Dlchlorobenzene*
Species
Rabbit
Rat
Guinea pig
Guinea pig
Guinea pig
Rabbit
Rat
Rat
Mice
Route
Inhalation
Inhalation
Inhalation
oral
oral
oral
oral
oral
s.c.
Concentration
or Dose
10s mg/m3
10s mg/m3
10s mg/m3
2800 mg/kg
1600 mg/kg
2812 mg/kg
500 mg/kg
4000 mg/kg
5145 mg/kg
Regimen
30 minutes
dally
30 minutes
dally
30 minutes
dally
single
single
single
single
single
Effects
CNS depression, eye
and nose Irritation
CNS depression, eye
and nose Irritation
Irritation, CNS de-
pression, and death
100% lethal
100% survival
"50
LD50
100% lethal
L050
Reference
Domenjoz, 1946
Oomenjoz, 1946
Domenjoz, 1946
HolUngsworth
et a!., 1958
HolUngsworth
et a!., 1958
Varshavskaya,
1967a
ChMstenson and
FalrchUd, 1976
HolUngsworth
et a!., 1958
Ir1e et a!.,
1973
*Source: Modified from U.S. EPA, 1980d
CNS = Central nervous system; s.c. = subcutaneous
-------
IMe et al. (1973) reported the toxldty of 1,4-dlchlorobenzene adminis-
tered subcutaneously to mice. They reported an L0,-0 of 5.145 g/kg.
Inhalation of 1,4~d1chlorobenzene (dose not specified) resulted 1n meta-
chromasla of the nuclei and cytoplasm of liver cells.
The Induction of hepatic porphyrla by oral administration of dlchloro-
benzene has been reported 1n several studies. R1m1ngton and Zlegler (1963)
gave rats 1,2- and 1,4-d1chlorobenzene at levels that Increased over several
days to 455 and 770 mg/kg, respectively. Clinical observations of toxldty
Included anorexia, weakness, clonlc contractions, hepatomegaly and liver
degeneration and focal necrosis. The metabolic alterations seen were
Increased urinary excretion of uroporphyrln, porphoblUnogen and amlnolevu-
I1n1c acid (1,4~d1chlorobenzene only). The authors noted that 1,2-d1chloro-
benzene appeared more acutely toxic and damaging to the liver, while !,4-d1-
chlorobenzene was more porphyrogenic. Poland et al. (1971) also Induced
hepatic porphyrla 1n rats by the dally gastric administration of 800 mg/kg
1,3-dlchlorobenzene 1n peanut oil over a 9-day period. Urinary copropor-
phyrln excretion Increased 1 day after the first dose, peaked at day 3 and
then decreased to a level 3 times the pre-dos1ng concentration. The Invest-
igators also found that the administration of 1, 3 or 5 doses of l,3-d1chlo-
robenzene enhanced the metabolism of hexabarbltal and b1shydroxycoumar1n»
and Interpreted these results to Indicate that 1,3-d1chlorobenzene Induced
drug-metabolizing enzymes and enhanced Us own degradation.
Enhancement of xenoblotlc metabolism of the liver by the dlchloroben-
zenes and other halogenated benzenes has been confirmed by other studies.
Ar1yosh1 et al. (1975a) treated female Wlstar rats orally for 3 days with
250 mg/kg/day of each of the dlchlorobenzene Isomers. The activities of
8-21
-------
several hepatic drug-metabolizing enzymes were Increased by these treat-
ments, although none of the Isomers Increased the liver content of cyto-
chrome P-450. Carlson and Tardlff (1976) also studied the effect of
1,4-d1chlorobenzene and other halogenated benzenes on hepatic metabolism.
Rats orally administered 10-40 mg/kg of the compound for 14 days showed
Increased activity of several metabolic enzymes, glucuronyltransferase and
the detoxification of hexobarbltal and 0-ethyl-0-p-n1trophenyl~phenyl-phos-
phonothloate (EPN).
8.3.2. Subchronlc Toxldty. Many subchronlc toxlclty studies of 1,2- and
l,4-d1chlorobenzene have been conducted by the oral and Inhalation routes of
administration. Although the majority of these studies have been with
1,4-d1chlorobenzene, the available data Indicate that effects similar to
those for 1,4-d1chlorobenzene result from exposure to 1,2- and 1,3-d1chloro-
benzene. In the subchronlc Inhalation studies (I.e., those using repeated
doses over a period of weeks or months), the toxic effects noted at low
doses (<1000 mg/m3 but >600 mg/m3) were growth depression, Increased
Hver and kidney weight and Hver necrosis. At higher Inhalation doses
(>1000 mg/m3), the toxic effects were Hver, lung and kidney pathology,
central nervous system depression, granulocytopenla and death. The lowest
level at which no adverse effects were found was 580 mg/m3 (96 ppm) of
1,4-d1chlorobenzene administered via Inhalation to several species for 7
hours/day, 5 days/week, over a 6- to 7-month period (HolUngsworth et al.,
1956). Subchronlc and chronic toxlclty data for 1,2- and 1,4-d1chloroben-
zene are presented 1n Tables 8-6 and 8-7, respectively.
HolUngsworth et al. (1958) exposed via Inhalation groups of 20 rats, 8
guinea pigs and 2 rabbits of each sex plus 2 female monkeys to repeated
exposures of 560 mg/m3 (93 ppm) 1,2-d1chlorobenzene for 7 hours/day,
8-22
-------
TABLE 8-6
Subchronlc ToxIcHy of 1,2-D1chlorobenzene*
TO
to
Route Concentration
or Dose
Inhalation 560 mg/m3
290 mg/m3
455 mg/m»
Oral 376 mg/kg (tube)
188 mg/kg (tube)
18.8 mg/kg (tube)
0.01-0.1 mg/kg/day
500 mg/kg
250 mg/kg
125 mg/kg
60 mg/kg
30 mg/kg
Regimen
7 hours/day, 5 days/week,
6-7 months
7 hours/day, 5 days/week
6.5 months
dally up to 15 days
5 days/week, 138 doses
5 days/week, 138 doses
5 days/week, 138 doses
5 months
5 days/week, 13 weeks
5 days/week, 13 weeks
5 days/week, 13 weeks
5 days/week, 13 weeks
5 days/week, 13 weeks
Subject
rat, guinea
pig, rabbit,
monkey
rat, guinea
pig
rat
rat
rat
rat
rat
rat
rat
rat
rat
rat
Effect
No effect on several parameters
except decreased spleen weights
In male guinea pigs
No effect on several parameters
Hepatic porphyHa
Liver, kidney weight Increase;
cloudy swelling 1n liver.
Increase In liver and kidney
weight
No effects noted
Hematopoletlc system; altered
conditioned reflexes; Increased
prothromb time and altered
enzyme activities
Increased liver weights; polyuMa
1n males; Increased urinary por-
phyHns; hepatic necrosis and
degeneration; renal tubular
degeneration; thymlc lymphold
depletion; and hematologlc and
clinical changes
Increased liver weights; hema-
tologlc and clinical changes;
hepatic necrosis
Increased liver weights; hema-
tologlc and clinical changes;
some hepatic necrosis
Hematologlc and clinical changes
Hematologlc and clinical changes
Reference
Holllngsworth et al. ,
1958
Holllngsworth et al. .
1958
R1m1ngton and
Zlegler. 1963
Holllngsworth et al.,
1958
Holllngsworth et al. ,
1958
Holllngsworth et al. ,
1958
Varshavskaya, 1967a
NTP, 1982
NTP, 1982
NTP, 1982
NTP, 1982
NTP, 1982
-------
TABLE 8-6 (cont.|
Route
Concentration
or Dose
Regimen
Subject
Effect
Reference
Oral (cont.) 500 rag/kg
CO
i
250 mg/kg
5 days/week, 13 weeks
5 days/week, 13 weeks
30, 60, 125 mg/kg 5 days/week, 13 weeks
Subcutaneous unspecified repeated
mouse
mouse
mouse
rabbit
Increased mortality; Increased
liver weights; Increased urinary
and liver porphyrlns; hepatic
necrosis and degeneration; heart
and skeletal muscle lesions;
lymphold depletion of thyntus and
spleen
Hepatic necrosis and degeneration
1n males; no effects 1n females
No effects
Blood dyscraslas, (agranulo-
cytosU)
NTP, 1982
NTP, 1982
NTP, 1982
Hare and West, 1977
•Source: Hodlfed from U.S. EPA. 1980d
-------
TABLE 8-7
Subchronlc and Chronic ToxIcHy of l,4-D1chlorobenzene*
Route
Concentration
or Dose
Regimen
Subject
Effect
Reference
CO
I
Inhalation 10s mg/m*
4800 mg/ma
4600-4800 mg/ffl»
2050 mg/m*
1040 mg/m*
950 mg/ms
900 mg/m'
580 mg/ms
500 ppm
(-3000 mg/m3)
75 ppm
(~450 mg/ms}
0.5 hours/day, 5-9 days
8 hours/day, 5 days/week,
up to 69 exposures
8 hours/day, 5 days/week,
7 hours/day, 5 days/week,
6 months
7 hours/day, 5 days/week,
16 days
7 hours/day, 5 days/week,
157-219 days
8 hours/day, 2 weeks
7 hours/day, 5 days/weeks
6-7 months
5 hours/day, 5 days/week,
for 76 weeks followed by
36 weeks with no exposure
5 hours/day, 5 days/week,
for 76 weeks followed by
36 weeks with no exposure
rabbit
rat, guinea pig,
rabbit
rabbit
rat, guinea pig
rat, guinea pig
rat, guinea pig,
rabbit, mouse,
monkey
mouse
rat, guinea pig,
mice, rabbit,
monkey
rat
rat
Branulocytopenla; Irritation; CNS
and lung toxldty; death (12/18)
Severe Irritation; CNS depression
and collapse; liver, kidney, lung
pathology; deaths
Tremors, weakness, nystagmus;
some deaths
Growth depression, Increased liver,
kidney weight; liver pathology
(necrosis, fatty degeneration,
swelling, flbrosls)
Increased liver, kidney weight
(rat); lung, liver pathology
Growth depression (guinea pig);
Increased liver, kidney weight;
histologlcal liver changes
(cloudy swelling, granular
degeneration) 1n rats, no adverse
effects reported 1n rabbit, mouse
or monkey
Respiratory excitation; liver
pathology, deaths; at serum
concentration of 39 mg/i
No adverse effects on several
parameters
Slightly elevated protein and
coproporphyrln outputs, Increased
liver and kidney weights.
Some Increases In liver weights
Zupko and Edwards,
1949
Holllngsworth et al.,
1956
P1ke, 1944
Holllngsworth et al.,
1956
Holllngsworth et al.,
1956
Holllngsworth et al.,
1956
Irle et al., 1973
Holllngsworth et al.,
1956
Loeser and LHchfleld,
1983
Loeser and LUchfleld,
1983
-------
TABLE 8-7 (cont.)
CO
I
Route
Inhalation
(cont.)
Oral
Concentration
or Dose
500 ppra
(-30DO ppffl)
200 ppra
(-1200 mg/m»)
75 ppm
(-450 mg/m»)
1000 mg/kg per
dose (tube)
770 mg/kg/day
500 mg/kg/day
(tube)
5000 mg/kg diet
500 mg/kg/day
(tube)
376 mg/kg/day
250 mg/kg/day
188 mg/kg/day
20-40 mg/kg/day
18.8 mg/kg/day
Regimen Subject
6 hours/day from days rat
6-15 of pregnancy
6 hours/day from days rat
6-15 of pregnancy
6 hours/day from days rat
6-15 of pregnancy
92 doses 1n 219 days rabbit
up to 5 days rat
5 days/week, 20 doses rat
up to 35 days Peking duck
5 days/week, 263 doses 1n rabbit
367 days
5 days/week, 138 doses 1n rat
192 days
3 days rat
5 days/week, 138 doses 1n rat
192 days
2 weeks rat
5 days/week, 138 doses 1n rat
192 days
Effect
5 dams out of 20 delivered Utter
1 day early, one fetus with
agnatMa and cleft palate
1 dam out of 20 delivered Utter
1 day early, one fetus with
gastroschlsls and malrotatlon
of hlndUmb
1 dam out of 20 delivered Utter
1 day early, one fetus with
gastroschlsls and malrotatlon
of hlndllmb
CMS depression; weight loss;
liver degeneration and necrosis;
deaths
Hepatic. porphyMa
Hepatic centrolobular necrosis;
cloudy swelling, renal tubular
epithelium, and casts
Death 1n 3/10. Retarded growth
CMS depression; weight loss; liver
pathology
Increased liver and kidney weight;
liver cirrhosis and focal necrosis
Induced liver metabolism enzyme
system
Increased liver and kidney weight
Induced liver metabolism enzyme
system
No adverse effects detected
Reference
Loeser and LHchfleld,
1983
Loeser and LHchfleld,
1983
Loeser and LHchfleld,
1983
HolUngsworth et al.,
1956
R1m1ngton and Zlegler,
1963
HolUngsworth et al.,
1956
HolUngsworth et al.,
1956
HolUngsworth et al.,
1956
HolUngsworth et al.,
1956
Ar1yosh1 et al.,
1975a,b
HolUngsworth et al.,
1956
Carlson and Tardlff,
1976
HolUngsworth et al.,
1956
•Source: U.S. EPA, 1980d
-------
5 days/week for periods ranging up to 7 months. They reported that this
exposure regimen did not result 1n any adverse effects on any of the animal
species tested. Inhalation-exposed groups of 20 rats and 8 guinea pigs of
each sex plus 10 female mice to repeated exposures of 290 mg/ma (49 ppm)
1,2-d1chlorobenzene for 7 hours/day, 5 days/week, for 6.5 months again
resulted 1n no adverse effects on any of the tested animals.
Several species of laboratory animals were exposed to 1,4-d1chloroben-
zene vapor at each of five concentrations for 7 hours/day (8 for the highest
dose group), 5 days/week (HolUngsworth et al., 1956). Effects 1n animals
(rats, guinea pigs, rabbits) exposed to 4800 mg/m3 (798 ppm) for up to 69
exposures Included: some deaths (up to 25%); marked tremors, weakness,
collapse, eye Irritation, and reversible eyeground changes 1n rabbits, but
no lens changes; weight loss, liver degeneration and necrosis, cloudy
swelling of renal tubular epithelium (rats); and lung congestion and emphy-
sema (rabbits). Effects 1n rats and guinea pigs exposed at 2050 mg/m3
(341 ppm) for 6 months Included: growth depression (male guinea pigs);
Increased liver and kidney weights (male rats); and liver pathology (cloudy
welling, fatty degeneration, focal necrosis, cirrhosis) 1n some of the
animals. Effects In animals exposed for as high as 12 exposure over 16 days
at 1040 mg/m3 (173 ppm) were: Increased liver, spleen and kidney weights
(guinea pigs); pulmonary edema, congestion, hemorrhage; hepatic centro-
lobular congestion and granular degeneration (rats). Effects 1n animals
exposed to 950 mg/m3 (158 ppm) for 157-219 days Included: growth depres-
sion (guinea pigs); Increased liver weights (rats, guinea pigs) and In-
creased kidney weights (rats); and centrolobular hepatocellular cloudy
swelling or granular degeneration (rats). No adverse effects were observed
1n rats, guinea pigs, rabbits, mice or a monkey exposed at 580 mg/m3 for
6-7 months.
8-27
-------
In the corresponding subchronlc oral studies, female rats (10/group,
strain not specified) were dosed via stomach tube with 18.8, 188 or 376 mg
1,2-d1chlorobenzene/kg/day, 5 days/week, for a total of 138 doses over 192
days (50% 1,2-dlchlorobenzene 1n olive oil) (Holllngsworth et al., 1958).
No adverse effects on growth or mortality were observed at any dose level.
A dose of 376 mg/kg/day resulted In slightly Increased liver and kidney wet
weights, a slight decrease 1n spleen wet weight and slight to moderate
cloudy swelling 1n the liver. Slight Increases 1n liver and kidney wet
weights were observed at the Intermediate dose and no effects were noted at
the lowest dose (18.8 mg/kg/day). Application of 1,2-d1chlorobenzene to the
eyes of two rabbits resulted 1n slight to moderate pain and slight conjunc-
tiva! Irritation which cleared completely within 7 days.
1,4-D1chlorobenzene was dissolved In olive oil and given by stomach tube
to male adult rats (2/group) at 10, 100 or 500 mg/kg 5 days/week for 4
weeks. Centrolobular hepatic necrosis and marked cloudy swelling of renal
tubular epithelium with cast formation occurred 1n animals given 500 mg/kg.
No adverse effects were observed at the lower dose levels (Holllngsworth et
al., 1956).
White female rats (10/group) were fed either 18.8, 188 or 376 mg/kg of
l,4-d1chlorobenzene 1n olive oil (emulsified with acacia) by stomach tube 5
days/week for a total of 138 doses 1n 192 days (Holllngsworth et al,, 1956).
At the highest dosage level of 376 mg/kg/dose, Increased liver and kidney
weights, and slight hepatic cirrhosis and focal necrosis were observed. No
adverse effects were noted at the lowest dose level (18.8 mg/kg). No cata-
racts were observed 1n these exposures. The same Investigators fed rabbits
(5/group) with 1,4-d1chlorobenzene 1n olive oil by Intubation for up to 92
doses In 219 days at a level of 1000 mg/kg/dose. Another group received a
8-28
-------
dose level of 500 mg/kg/dose 5 days/week for a total of 263 doses 1n 367
days. Effects at the higher dose level (1000 mg/kg) Included: weight loss,
tremors, weakness, hepatic cloudy swelling and a few areas of focal
necrosis, and deaths. Similar changes, but no deaths, were noted in rabbits
on the lower dose regimen. No cataracts were observed. Peking ducks
(10/group) fed 1,4-d1chlorobenzene 1n their diet at 0.5% (5000 mg/kg diet)
for 35 days experienced retarded growth and 30% mortality 1n 28 days, but no
cataracts were observed (HolHngsworth et al., 1956).
Varashavskaya (1967) administered 1,2-d1chlorobenzene 1n sunflower oil
orally to rats at doses of 0.001, 0.01 and 0.1 mg/kg/day over a 5-month
period. At the highest dose level, Inhibition of erythropolesls and bone
marrow activity was' observed. In addition, at this level, adrenal weight
and ascorbic add content decreased, serum alkaline phosphatase and trans-
amlnase activity Increased, and serum glutathlone decreased. Similar
effects were noted 1n the Intermediate dose level animals, but not at the
lowest dose. These results are 1n distinct contrast to those of HolUngs-
worth et al. (1956) who found no effects at a dose level of 18.8 mg/kg after
a 6-month administration period,
Subchronlc toxldty studies on 1,2-d1chlorobenzene were conducted under
the auspices of the National Toxicology Program (NTP, 1982). The Investiga-
tions were conducted using 10 male and 10 female B6C3F, mice and 10 male
and 10 female F344/N rats/group. The 1,2-d1chlorobenzene was administered
by gavage using a corn oil vehicle, 5 ml/kg, 5 days/week for 13 weeks.
The 1,2-d1chlorobenzene doses used were 0, 30, 60, 125, 250 or 500 mg/kg.
The 1,2-d1chlorobenzene mouse study resulted 1n a decreased survival
rate 1n the male 250 mg/kg dose group with a mortality rate of 10% (1/10)
and 1n the male and female 500 mg/kg dose groups with mortality rates of 40%
8-29
-------
(4/10) and 30% (3/10), respectively (NTP, 1982). Body weight gains were
depressed 47% 1n males and 67% 1n females at the 500 mg/kg/day dose. For
all other groups body weight gains were within 95% of that of controls.
Liver weight/body weight ratios were significantly Increased 1n both males
and females at 500 mg/kg. Spleen weight/body weight ratios were decreased
1n all l,2-d1chlorobenzene dosed female groups and uterus weight/body weight
ratios were decreased 1n the 500 mg/kg female group. No biologically
significant changes were found during the hematologlcal evaluations. Female
mice receiving 500 mg/kg 1,2-dlchlorobenzene were found to excrete 6 times
more coproporphyrlns 1n their urine and had a 2-fold Increase 1n liver
porphyrln concentrations when compared with control mice. No hlstologlcal
effects were observed 1n the 0, 30, 60 or 125 mg/kg dose groups. The 250
mg/kg dose male mice group was found to have hepatocellular necrosis and
degeneration while the females receiving this dose were found to be
unaffected. Ninety percent of both the male and female 500 mg/kg dose
groups were observed with centrllobular necrosis, necrosis of Individual
hepatoeytes or hepatocellular degeneration. The hearts of the 500 mg/kg
dosed animals had mineralization of the myocardlal fibers (multiple foci)
and the skeletal muscles were observed with some necrosis, myosltls and
mineralization. Both the male and female 500 mg/kg groups were observed
with lymphold depletion of the thymus and spleen and a yellow-green pigmen-
tation (considered to be hemoslderln) 1n some of their Hvers. Based on
these results NOELs were determined to be 125 mg/kg for male mice and 250
mg/kg for female mice.
The 1,2-d1chlorobenzene rat study resulted 1n a dose-dependent depres-
sion 1n mean body weight gains over the 13-week period (NTP, 1982). This
depression In weight gain averaged 9.1%, 11.5% and 32.8% 1n males and 8.9%,
8-30
-------
11.IX and 15.5% 1n females dosed with 125, 250 and 500 mg/kg/day l,2-d1-
chlorobenzene, respectively. A dose-related Increase 1n liver weights was
also observed 1n both sexes with significant Increases 1n liver weight/body
weight ratios 1n the 125, 250 and 500 mg/kg male and female dose groups.
Decreases 1n spleen and thymus weights and organ weight/body weight ratios
were observed 1n the male 500 mg/kg group. Minimal changes 1n hematologlc
parameters were observed 1n the 500 mg/kg dose groups. An Increased number
of platelets were found 1n female rats receiving 60, 125 and 500 mg/kg doses
of 1,2-d1chlorobenzene. A dose-related Increase 1n serum cholesterol levels
were found 1n males receiving 30, 125, 250 and 500 mg/kg and 1n females
receiving 125-500 mg/kg. A decrease 1n serum trlglycerldes was observed at
500 mg/kg (males) and 250 mg/kg (females), and a dose-related Increase 1n
total serum protein was observed at 250-500 mg/kg (males) and at 30-500
mg/kg (females). Female rats were observed with minimal Increases 1n serum
glucose levels at 1,2-d1chlorobenzene doses of 30, 125, 250 and 500 mg/kg.
Polyurla was observed 1n males receiving the 500 mg/kg dose. A 3- to 5-fold
Increase 1n urinary uroporphyrlns and coproporphyrlns were seen 1n males and
females at 500 mg/kg. The liver porphyrln levels were not altered by
1,2-d1chlorobenzene at any dose level. Hepatocellular necrosis and focal
hepatic necrosis were observed 1n some of the rats at the 125 mg/kg dose.
More hepatocellular necrosis was seen 1n both males and females at 250
mg/kg. Host of the rats 1n the 500 mg/kg dose groups had Hver lesions,
either centrllobular degeneration or hepatic necrosis. The 500 mg/kg male
group also had renal tubular degeneration and thymlc lymphold depletion. A
yellow-green to gold pigment (believed to be hemos1der1n) was also observed
1n the livers of rats at 250 and 500 mg/kg. Based on these results, a LOAEL
for 1,2-d1chlorobenzene 1n rats was determined to be 30 mg/kg.
8-31
-------
The effect of subchronlc treatment with 1,4-d1chlorobenzene has been
Investigated 1n guinea pigs (Salamone and Coppola, 1960; Totaro, 1961;
Coppola et a!., 1963; Totaro and L1car1, 1964). Intramuscular Injections of
12i rag 1,4-d1chlorobenzene (50% 1n almond oil) dally for 20 days were found
to produce weight loss (5-10%), Increased blood serum transamlnase and
Increased clotting times.
8.3.3. Chronic Toxldty. Two-year chronic bloassay studies using l,2-d1-
chlorobenzene were conducted under the auspices of the National Toxicology
Program (NTP, 1982). The Investigations were conducted using 50 male and 50
female B6C3F1 mice and 50 male and 50 female F344/N rats. 1,2-D1chloroben-
zene was administered by gavage 1n a corn oil vehicle, at a volume of 5
mst/kg, 5 days/week for 103 weeks. The dosage groups used were 0 (vehicle
control), 60 and 120 mg/kg. The 1,2-d1chlorobenzene was >99% pure with the
major Impurity found to be 0.84% v/v of 1,4-d1chlorobenzene. The stability
of the l,2-d1chlorobenzene preparation was monitored.
The l,2-d1chlorobenzene mouse study resulted 1n a 105-week (exposure
duration 103 weeks) survival rate of 52% (26/50), 64% (32/50) and 70%
(35/50) 1n male mice and 66% (33/50), 80% (40/50) and 76% (38/50) 1n female
mice for the 0, 60 and 120 mg/kg dose groups, respectively (NTP, 1982).
Mean body weights were comparable between 1,2-d1chlorobenzene dosed and
control male mice but were slightly greater In the female dosed mice than
controls. H1stolog1cal findings of neoplasms In dosed and control groups
will be discussed In Section 8.3.5. Carc1nogen1c1ty. No apparent Increase
In non-neoplast1c lesions In the liver, kidney, bone marrow, spleen or other
organs of male and female mice were observed as a result of administration
of 1,2-dlchlorobenzene over the 105-week study period.
8-32
-------
The 1,2-chchlorobenzene rat study resulted 1n 104-105 week (exposure
duration 103 weeks) survival rates of 84% (42/50), 72% (36/50) and 38%
(19/50) [significantly different from the 60 mg/kg group (p=0.014) and 0
mg/kg group (p<0.001)] In male rats and 62% (31/50), 66% (33/50) and 64%
(32/50) 1n female rats for the 0, 60 and 120 mg/kg dose groups, respectively
(NTP, 1982), Among the high dose males 17 were accidentally killed as a
result of the gavage procedure. If these 17 animals had not died, survival
would have been comparable to that of the low dose and control groups.
Slightly lower mean body weights were observed 1n the male 120 mg/kg group
when compared with the 0 and 60 mg/kg male groups. This was contrasted by
higher mean body weights In the female animals In the 120 mg/kg group when
compared with the female controls. H1stolog1cal findings of neoplasms 1n
dosed and control groups will be discussed 1n Section 8.3.5. Cardnogenlc-
Ity. No apparent Increase In non neoplastlc lesions 1n the liver, kidney,
bone marrow, spleen, thymus or other organs of male and female rats were
observed as a result of administration of 1,2-d1chlorobenzene over the
105-week study period.
Loeser and LHchfleld (1983) reported on a long-term Inhalation study on
1,4-dlchlorobenzene 1n rats and mice. Groups of 76-79 male and female rats
(SPF, Alderly Park W1star-der1ved strain) and 75 male and female mice (SPF,
Alderly Park Swiss strain) were exposed 5 hours/day, 5 days/week to 0, 75 or
500 ppm 1,4-dlchlorobenzene. The rats were exposed for 76 weeks and the
survivors were held unexposed for an additional 36 weeks. The mice were
exposed for 57 weeks and the surviving females were held unexposed for an
additional 19 weeks. The male mice were terminated at 57 weeks of exposure,
when mortality reached 80%, due to early fighting among the males and a
probable occurrence of respiratory Infection, which resulted 1n little data
8-33
-------
being collected from the male mice. No treatment related changes 1n body
weight, food and water Intake or mortality rates were seen between exposed
and control groups. In the rats dose-related changes 1n blood biochemistry
and hematology were not noted along with no Increases 1n hepatic amlnopyrene
demethylase activity. The 500 ppm rat groups showed a slightly elevated
urinary protein and coproporphyMn output along with Increased liver and
kidney weights and small Increases 1n heart and lung weights. There were
some Increased liver weights seen 1n the 75 ppm rat groups. The cumulative
mortality (32-40% at week 72) observed 1n the female mice did not appear to
be related to 1,4-d1chlorobenzene exposure. The female mice of all groups
had a high background Incidence of respiratory disease, which made Interpre-
tation of respiratory tract changes difficult to assess. No evidence of any
treatment-related non-neoplast1c effects 1n any examined female mice were
reported. Findings of neoplasms 1n rats and mice dosed and control groups
will be discussed 1n Section 8.3.5.
8.3.4. Mutagenldty. The capability of the dlchlorobenzenes to Induce
mutations or other alterations of genetic structure has not been extensively
Investigated, although a recent study (Zapata-Gayon et al., 1982) Indicates
such research 1s warranted. As cited 1n Section 8.2., a higher Incidence of
chromosomal breaks was observed 1n the leukocytes of humans accidentally
exposed for a short period of time to 1,2-d1chlorobenzene vapors (Zapata-
Gayon et al., 1982).
Anderson et al. (1972) reported that 1,2-d1chlorobenzene did not Induce
point mutations when tested 1n Salmonella typhlmurlum (8 unspecified
strains) without activation. No conclusions can be drawn from this study
because of the lack of details provided and because metabolic activation was
not used. In an abstract, Lawlor et al. (1979) evaluated the ability of
8-34
-------
chlorinated phenols, benzenes and hexanes to Induce mutations or DNA damage
1n bacteria. Tests of 1,2- and 1,4-dichlorobenzene (doses not specified)
were negative 1n five strains of Salmonella (TA98, TA100, TA1535, TA1537 and
TA1538) with and without rat liver mlcrosomal activation. DNA repair tests
with two Salmonella and two £. coli strains with and without activation
Indicated the ability of two unspecified chlorobenzenes to cause preferen-
tial killing of the DNA repair deficient strains. Because these results
were reported 1n an abstract with Insufficient experimental detail, the
results cannot be critically evaluated. These negative findings 1n bacteria
were supported by studies conducted for the National Toxicology Program
(Appendix M of NTP, 1982). In these studies, 1,2-d1chlorobenzene was
negative 1n four Salmonella strains (TA98, TA100, TA1535 and TA1537) when
tested with and without metabolic activation at doses as high as 1300
yg/plate.
Prasad and Pramer (1968) reported testing all three Isomers of dlchloro-
benzene 1n an auxotrophlc strain of Aspergl11 us nldulans. a soil mold. All
three compounds Increased the frequency of back mutations 1n the following
descending order: 1,4-, 1,3- and 1,2-d1chlorobenzene. Abnormal numbers of
chromosomes and abnormally shaped nuclei were observed 1n the root cells of
A111 urn exposed for 4 hours to l,4-d1chlorobenzene vapors (Carey and
HcDonough, 1943). Sharma and Bhattacharyya (1956) reported chromosomal
breakage and nondlsjunctlon 1n the root tips and flower buds of Nothoscordum
fragans, which were treated with saturated aqueous solutions of l,4-d1chlo-
robenzene. Various mltotlc abnormalities were also found 1n the somatic
cells and chromosomes of the root tips of several plant species treated with
1,4-dichlorobenzene (Srivastava, 1966). The aberrations Included shortening
and thickening of chromosomes, early separation of chromatlds, tetraplold
8-35
-------
cells, blnucleale cells, chromosome bridges and chromosome breaks 1n the
heterochromatlc regions. Sarbhoy (1980) exposed germinating root tips of
Lens esculenta to 1,4-d1chlorobenzene vapors and also observed chromosome
fragmentation, condensation and bridges and polyplold cells.
8.3.5. Carc1nogen1c1ty. The National Toxicology Program (NTP) conducted
a 2-year study on !,2-d1chlorobenzene with F344/N rats and with B6C3F,
mice {NTP, 1983). Exposure conditions and noncarclnogenlc effects have been
described 1n Section 8.3.3.
8.3.S.I. RAT STUDY -- In the case of the F344 N rats, dose selection
was made as a result of observations 1n the 13-week subchronlc study as
described 1n Section 8.3.2. The findings on survival, weight decrement and
pathology formed the basis for selection of the 60 and 120 mg/kg dosages for
the 2-year study 1n the rats.
As stated previously 1n Section 8.3.3., body weights were either
unaffected or Increased during chronic exposure, while the only Instance of
Increased mortality (high dose males) could be explained by Increased
Incidence of accidental deaths.
The results of the hlstopathologlcal analysis showed that non-neoplast1c
lesions were not significantly Increased 1n the treated rats nor were neo-
plasms other than adrenal pheochromocytoma. The latter (Table 8-8) was
significantly Increased 1n the low dose group when compared to controls by
the life table test but not 1n the other statistical tests. The terminal
Incidence of adrenal pheochromocytoma 1n males was 36% (13/36) 1n the low
dose group compared with 19% (8/42) 1n controls. The historical Incidence
of adrenal tumors 1n male F344/N rats receiving corn on by gavage, based on
data from seven different laboratories 1s 153/986 (15.5%).
8-36
-------
TABLE 8-8
NIP Bloassay of 1,2-D1ch1orobenzene
Analysis of Primary Tumors 1n Male Rats: Adrenal Pheochrocnocytomas*
Vehicle 60 mg/kg 120 mg/kg
Control
Tumor Rates
Overall 9/50 (18%) 16/50 (32%) 6/49 (12%)
Adjusted 20.9% 40.5% 21.7%
Terminal 8/42 (19%) 13/36 (36%) 2/18 (11%)
Statistical Tests
Life Table p = 0.201 p = 0.039 p = 0.380
Incidental Tumor Test p = 0.499 N p =0.070 p = 0.420 N
Cochran-Armltage Trend, p = 0.285 N p = 0.083 p = 0.303 N
Fisher Exact Tests
*Source: NTP, 1982
8-37
-------
In rats, therefore, under conditions of this test, carclnogenldty was
not demonstrated. However, based on the following observations from the
2-year study, a larger dose possibly could have been tolerated:
1) there was probably no Increase 1n mortality 1n treated groups
when compared with controls,
2) there was no loss of weight In the treated groups compared to
the controls,
3) there was no evidence of life-threatening pathology 1n the
treated groups compared to the controls.
On the other hand, decreased weight gains 1n the 13-week range finding study
at 125, 250 and especially at 500 mg/kg/day doses Indicate that much larger
doses probably would not have been tolerated.
In summary, the assay of 1,2-d1chlorobenzene In F344 rats did not give
evidence of carclnogenldty. However, slightly higher doses possibly could
have been tolerated and the assay may not have been as sensitive as 1t could
have been.
8.3.5.2. MICE — In a range finding study described in Section
8.3.2., mice were exposed to doses of up to 500 mg/kg/day 1,2-d1chloroben~
zene. As a result of Increased mortality and decreased weight gain at the
500 mg/kg/day dose, along with limited evidence for liver pathology at the
250 mg/kg/day dose, 60 and 120 mg/kg/day were the dose levels selected for
the chronic study. Details of the experiment and noncardnogenlc toxic
effects are described 1n Section 8.3.3.
The combined Incidence of all types of lymphomas was not significantly
affected by exposure. A dose-related decrease 1n the Incidence of hepato-
cellular adenomas 1n males was significant, Alveolar/bronchlolar carcinomas
occurred 1n male mice with a statistically significant positive trend
(p=0.037; 4/50, 2/50 and 10/50, In the Cochran-Armltage test), but 1f the
8-38
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Incidence of alveolar/bronchlolar adenomas 1s combined with the carcinomas
(8/50, 8/50 and 13/50) no significant changes could be detected. Under the
conditions of these studies, therefore, there 1s no convincing evidence for
the cardnogenldty of 1,2-d1chlorobenzene 1n B6C3F, mice.
Loeser and Utchfleld (1983) conducted a long-term Inhalation study In
rats and mice using 1,4-d1ehlorobenzene. Groups of 76-79 rats/sex/group
(SPF, Alderly Park Wlstar derived strain) and 75 swiss mice/sex/group
(Alderly Park strain) were exposed 5 hours/day, 5 days/week to 75 or 500 ppm
1,4-dlchlorobenzene. The rats were exposed for 76 weeks, then survivors
were held for an additional 36 weeks. The mice were exposed for 57 weeks
and the females then held an additional 19-20 weeks before a terminal sacri-
fice. The male mice were terminated at the end of the 57 week exposure when
fighting resulting 1n a mortality of 80% occurred. No treatment-related
changes were seen 1n body weight, food and water Intake or mortality rates
In either species. No treatment-related effects 1n the Incidence of tumors,
their multiplicity or malignancy were seen.
Based upon an estimated minute volume of 0.22 mVday for a 350 g rat,
the dally Inhaled dose would equal -400 mg/kg bw or ~285 mg/kg averaged over
7 days/week. The lack of body weight changes or mortality Increases Indi-
cated that the maximum tolerated dose had not been reached. If however,
toxldty was similar to that of the l,2-1somer and absorption via Inhalation
was equal to that of gavage the high dose used should be closer to the HTD
than the one used 1n the NTP (1983) study.
In conclusion, neither the rat nor the mouse gavage study nor the Inha-
lation experiments gave evidence of cardnogenldty under the test condi-
tions, but the doses selected were probably below the MTD 1n both species,
reducing the sensitivity of the assays. The marginal Increase 1n adrenal
8-39
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pheochromocytoma In rats dosed via gavage should be noted as this lesion
appears with hexachlorobenzene, also at a relatively low dose.
8.3.6. Reproductive and Teratogenlc Toxldty. No data on the reproduc-
tive and teratogenlc toxldty of 1,2-d1chlorobenzenes was available for
review; however, dlchlorobenzenes have been demonstrated to cross the
placenta (Dowty et al., 1976).
Loeser and L1tchf1eld (1983) reviewed an unpublished 1977 report from
the Imperial Chemical Industries Ltd., Central Toxicology Laboratory on a
rat embryotoxlclty and teratogenldty Inhalation study. This study Involved
the Inhalation exposure of 20 pregnant SPF rats/group to 1,4-d1chlorobenzene
for 6 hours/day from day 6-15 (Inclusive) of pregnancy to atmospheres of 0
(control), 75, 200 or 500 ppm 1,4-d1chlorobenzene. During the study the
dams were observed for clinical signs, body weights, and on day 21 of preg-
nancy caesarean sections were performed and the Intact uterus was Inspected
for the number of viable fetuses (their sex and weight), resorptlons and
copora lutea. Maternal results showed only one dam at 75 ppm, one at the
200 ppm level and five dams at the 500 ppm level delivering Utters 1 day
earlier than expected, otherwise no differences among the dams were noted.
No l,4-d1chlorobenzene-1nduced effects on the numbers of Implantations,
resorptlons, viable fetuses, runts, skeletal variants, corpora lutea, mean
fetal weight, Utter size or sex ratios were noted. The only fetal effects
noted 1n the 1,4-d1chlorobenzene-exposed groups were one fetus with gas-
troschlsls and malrotatlon of left hlndUmb 1n the 75 ppm group, one fetus
with gastroschlsls and malrotatlon of right hlndUmb 1n the 200 ppm group,
and one fetus with agnathla and cleft palate 1n. the 500 ppm group. The
report concluded that no evidence of embryotoxlclty or teratogenldty 1n the
study were found, which seems surprising since effects were noted 1n the 75,
200 and 500 ppm groups and none were noted 1n the control group.
8-40
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8.4. INTERACTIONS
As Indicated 1n Section 8.2., halogenated benzenes. Including the
dlchlorobenzenes, have the ability to Induce hepatic xenoblotlc metabolizing
enzymes (Arlyoshl et a!., 1975a,b; Carlson and Tardlff, 1976; Carlson,
1977). This type of Induction will alter the metabolism of other compounds
which are blotransformed by the same metabolic pathway; thus, the toxldty
resulting from the concurrent exposure to the dlchlorobenzenes and other
compounds will probably be different from the exposure to the Individual
chemicals. One study was available that Investigated the effect of dlchlo-
robenzene on the toxldty of other compounds (Townsend and Carlson, 1981).
Mice were orally administered 0.1 mmol/kg (18 mg/kg bw) of 1,4-d1chloroben-
zene and other chlorinated and bromlnated benzenes dally for 7 days, after
which the mice were used In the determination of LDcri values for four
3U
organophosphorus Insecticides. The treatment with 1,4-dlchlorobenzene was
found to decrease the lethality of parathlon and paraoxon by ~5Q%, although
other compounds were much more effective. In addition, Carlson and Tardlff
(1976) observed that administration of 1,4-d1chlorobenzene (10-40 mg/kg for
14 days to rats) enhanced the detoxification of hexobarbltal and EPN.
Harden and Baetjer (1978) reported a human case of aplastlc anemia
following exposure to 1,4-d1chlorobenzene and naphthalene. While a single
case report cannot be considered convincing evidence for an Interactive
effect, the possibility of Interactions cannot be dismissed.
8.5. SUMMARY
The available data on the pharmacoklnetlcs of the dlchlorobenzenes
Indicate that these compounds are absorbed through the lungs, skin and
gastrointestinal tract and rapidly distributed to many tissues, especially
those with a high I1p1d content. Metabolism 1s accomplished by oxidation to
8-41
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cUchlorophenols which maybe excreted per se or conjugated as glucuronldes
and sulfates. Elimination, primarily through the urine, appears to be
rapid, although the data are Insufficient to make quantitative estimates of
the rate. Biliary excretion does occur but Uttle of the biliary excreted
dlchlorobenzene has been found 1n the feces, probably due to enterohepatlc
redrculatlon. The dlchlorobenzenes, as well as the other chlorinated ben-
zenes, are capable of bloaccumulatlon (see Section 5.3.),
Data on effects 1n humans were available 1n a number of case reports and
1n a single ep1dem1olog1c study. The case studies demonstrate the ability
of the dlchlorobenzenes to be absorbed through the lungs and gut and their
acute and subchronlc toxlclty. Many of these reports, 1n which exposure may
have occurred over several years, noted toxic effects 1n the blood, such as
chronic lymphold leukemia and anemia, as well as effects on the liver. The
one available occupational study reported chromosomal alterations 1n leuko-
cytes resulting from a short-term exposure to 1,4-d1chlorobenzene. Taken
together, these studies suggest a possible toxic action of dlchlorobenzenes
on bone marrow and other organs of the blood-forming system.
Studies of the acute and subchronlc toxlclty of the dlchlorobenzene
Isomers Indicate that, 1n general, these compounds have similar target
organs and effects. At oral doses ranging from 125-1000 mg/kg over periods
of up to 6 months, the dlchlorobenzenes cause central nervous system depres-
sion, pathological Identified Injury to liver, kidney, heart, thymus and
spleen, and hepatic and urinary porphyrla; however, one study reported that
a low dose of 0.01 mg/kg over a 5-month period Inhibited erythropolesls and
bone marrow activity. The subchronlc oral toxlclty studies 1n rats provide
two estimates of NOEL values: 0.001 mg/kg (Varashavskaya, 1967) for
l,4-d1chlorobenzene and 18.8 mg/kg for 1,2- and for 1,4-d1chlorobenzene
8-42
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(HoTMngsworth et al., 1956, 1958). The NTP (1982) subchronlc oral study on
1,2-d1chlorobenzene 1n mice provided higher estimated NOEL values of 125 and
250 mg/kg for males and females, respectively. A 2-year NTP chronic oral
gavage study on 1,2-d1chlorobenzene 1n rats and mice, conducted primarily as
a cardnogenesls bloassay at the 60 and 120 mg/kg dose levels, resulted 1n
only Increased mortality 1n the male rats given 120 mg/kg. Acute and sub-
chronic Inhalation studies of dlchlorobenzenes Indicate similar toxic
effects and target organs as seen 1n the oral studies. The effects occurred
at doses >900 mg/m3; Inhalation NOELs were reported as 580 mg/ma and
-450 mg/m3 for 1,4-d1chlorobenzene (HolUngsworth et al., 1956; Loeser and
LHchfleld, 1983) and 290 mg/m3 for 1,2-d1chlorobenzene (HolUngsworth et
al., 1958).
The mutagenldty studies with bacteria were lacking 1n experimental
detail, but suggested that the dlchlorobenzenes are probably not mutagenlc
1n bacteria. However, several studies with mold and plant cultures treated
with dlchlorobenzenes have reported mutations and chromosomal aberrations.
Because chromosomal aberrations were also observed 1n human workers exposed
to 1,2-d1chlorobenzene, the weight of available evidence suggests that the
dlchlorobenzenes are clastogens. The carcinogenic activity of 1,2-dlchloro-
benzene, was tested 1n the NTP bloassay program 1n two rodent species at
doses of 60 and 120 mg/kg. No evidence of carcinogenic activity was found
under the test conditions. The carc1nogen1c1ty of 1,4-d1chlorobenzene was
tested 1n two rodent species using long-term Inhalation exposure. Again, no
evidence for carc1nogen1c1ty was noted. Since 1t 1s possible that the maxi-
mum tolerated dose was not used 1n either study, then the evidence 1s not
considered definitive for developing conclusions concerning the carclnogen-
1c1ty of 1,2- or 1,4-d1chlorobenzene 1f the IARC criteria for classifying
carcinogens are used.
8-43
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9. TRICHLOROBENZENES
The tMchlorobenzenes are produced 1n relatively small amounts (1.3-7
million kg/year Is the estimated 1983 production) (U.S. EPA, 1983; Chloro-
benzene Producers Association, 1984) and are used primarily as chemical
Intermediates, solvents, Insecticides, and coolants and Insulators 1n
electrical equipment (Hawley, 1977; SUmak et a!., 1980), Trlchlorobenzenes
have been detected 1n all environmental media Including drinking water (see
Section 4.3.), and have been found to bloaccumulate 1n fish (see Section
5.3.). In addition to the exposure of humans during the manufacture and use
of trlchlorobenzenes, exposure could result from Inhalation of contaminated
air and 1ngest1on of contaminated food and water.
9.1. PHARHACOKINETICS
9.1.1. Absorption. No quantitative studies on the absorption of the trl-
chlorobenzenes from the gastrointestinal tract, skin or lungs were found.
Information on absorption may be obtained from data describing elimination.
Hale Charles River rats (16 1n the group) excreted a mean of 84%, and two
female rhesus monkeys excreted a mean of 40% of the orally (by gavage)
administered dose of 10 mg 14C-1,2,4-tr1chlorobenzene/kg In the 24-hour
urine, while fecal elimination accounted for only 11 and 1%, respectively
(L1ngg et al., 1982). The results Indicate that In these species, this
Isomer Is well absorbed from the gastrointestinal tract. Two Chinchilla
female rabbits given doses of 500 mg 1,3,5-tr1chlorobenzene/kg 1n araehls
oil by gavage expired -10% of the administered dose via the lungs over a
period of 9 days (Parke and Williams, 1960). These Investigators also
observed elimination of urinary and fecal metabolites, but quantities or
percentages were not reported.
That the trlchlorobenzenes are absorbed by the respiratory tract and by
the skin can be Inferred from systemic effects observed 1n toxldty studies
9-1
-------
using the Inhalation (Kodba et al., 1981) and dermal (Brown et al., 1969)
routes of exposure. These studies, however, were not designed to give
Information on rates of absorption,
9.1.2. Distribution. Smith and Carlson (1980) examined the distribution
of "C-l,2,4-tr1chlorobenzene 1n groups of four male Sprague-Dawley rats
on days 1, 6, 11 and 16 after oral dally dosing with 181.5 mg/kg (1 mmol/kg)
1n corn oil for 7 days. Their data Indicate that the adrenals Initially had
the highest concentration of radlolabel. This level declined rapidly;
however, by day 11 1t was less than twice the background of the other
tissues. Abdominal fat had the highest concentration at the end of 1 day
(Table 9-1) and maintained detectable concentrations (20% of the day 1
level) for the duration of the observation period (16 days). The liver also
maintained detectable levels throughout the recovery period, retaining ~30%
of the day 1 level by day 16. These authors also found that starvation for
4 days had no observed effect on the distribution of X4C-tr1chlorobenzene
1n fat or liver.
Parke and Williams (1960) reported the distribution of 1,3,5-tr1chloro-
benzene 1n one rabbit on day 8 following oral administration of a single
dose of 500 mg/kg as follows: 13% of the administered dose was detected 1n
the feces, 23% (4% as raonochlorobenzene) 1n the gut, 5% 1n the pelt, 5% 1n
depot fat (exclusive of pelt) and 22% 1n the carcass.
9.1.3. Metabolism. No metabolic studies following the Inhalation of trl-
chlorobenzenes were available for review, but the metabolic fate following
oral and/or Intravenous (1.v.) or 1ntraper1toneal (1.p.) administration has
been characterized 1n rabbits (Jondorf et al., 1955; Parke and Williams,
1960; KohH et al., 1976) and 1n rats and monkeys (L1ngg et al., 1982).
Jondorf et al. (1955), using spectrophotometrlc analysis, studied the
metabolism of all three Isomers of trlchlorobenzene 1n groups of 3 or 4
9-2
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TABLE 9-1
Distribution of 14C-Labeled 1,2,4-Tr1chlorobenzene 1n Rat Tissues
after Oral Dosing with 181.5 mg/kg/day for 7 Days3
Tissue
Abdominal fat
Liver
Adrenals0
Muscle
Kidney
Heart
Spleen
Day 1
2033i439
1075+87
754+132
400+30
147H167
438+14
404+14
Activity (dpm/a t1ssue)b
Day 6 Day 11
642+54 342+10
442+.22 308+_21
246+22 d/
d/
404+43 d/
d/
d/
Day 16
408+39
317+18
aSource: Smith and Carlson, 1980
*>Each value Is the mean +_ SE for 4 rats, except for abdominal fat on day 1,
which was for three rats.
cTotal for both adrenals; they were not weighed,
dyalue less than twice background; further analyses were not performed.
9-3
-------
Chinchilla rabbits given a single oral dose of 500 mg/kg In arachls oil.
The results Indicated that the 1,2,3- Isomer was metabolized to 2,3,4-tr1-
chlorophenol (TCP), to 3,4,5-TCP to a lesser degree, and to small amounts of
3,4,5-trlchlorocatechol. During the 5 days after administration, 50% of the
dose was excreted 1n the urine as glucuronlc add conjugates, 12% as
sulfurlc add (sulfate) conjugates and 0.3% as 2,3,4-trlchlorophenylmercap-
turlc add. The 5-day urinary metabolites of 1,2,4-tr1chlorobenzene were
represented by glucuronlde conjugates (27%), sulfurlc add conjugates (11%)
and 2,3,5- and 2,4,5- tr1chlorophenylmercaptur1c acid (0.3%). The major
phenols formed were 2,4,5- and 2,3,5-TCP. For the 1,3,5- Isomer, 20% was
excreted as glucuronlde and 3% as sulfurlc add conjugates. No mercaptuMc
add was found, 2,4,6-tMchlorophenol was the only phenol detected 1n the
urine, and some unchanged 1,3,5-tMchlorobenzene was present 1n the feces.
To further characterize and clarify the metabolic fate of the 1,3,5- Isomer,
Parke and Williams (1960) followed the 9-day urinary excretion 1n 2 or 3
female Chinchilla rabbits treated orally with a single dose of 500 mg of the
1somer/kg. For the first 3 days, the rabbits eliminated 2,4,6-TCP along
with some minor monochlorophenols, while from day 4 to 9, 4-chlorophenol was
detected more prominently along with 2,4,6-TCP and -1% of the dose as
4-chlorocatechol.
Using GC-HS analysis, KohH et al. (1976) examined the metabolism of the
three trlchlorobenzene Isomers following a single 1.p. Injection of 60-75
mg/kg doses 1n vegetable oil to male rabbits (number and strain not re-
ported). In agreement with the results of Jondorf et al. (1955), the major
urinary metabolites of 1,2,4-tMchlorobenzene were 2,4,5- and 2,3,5-TCP.
The major metabolite of 1,2,3-tr1chlorobenzene was 2,3,4-TCP, with 2,3,6-
and 3,4,5- TCP as minor urinary metabolites. The 1,3,5- Isomer was
9-4
-------
metabolized to 2,3P5- and 2,4,6-TCP and a third, more polar metabolite, was
tentatively Identified as a dlchlorobenzene with 2 hydroxyl and 1 methoxyl
substHuents.
L1ngg et al. (1982) Investigated the metabolism of 1,2,4-trlchloroben-
zene 1n groups of 16 male Charles River rats and groups of 2 female rhesus
monkeys following a single oral or 1.v. administration of 10 rug/kg doses and
found similar phenolic metabolites to those observed In the rabbit. These
researchers were also able to characterize some species specific conjugates.
An Homeric pair of 3,4,6-tr1chloro-3,5-cyclohexad1ene-l,2-d1ol glucuronldes
accounted for 48-61% of the 24-hour urinary metabolites 1n the monkeys.
Also found were glucuronldes of 2,4,5- and 2,3,5-TCP and unconjugated TCP,
which accounted for 14-37 and 1-37% of the urinary metabolites, respec-
tively. In the rat, the 2,4,5- and 2,3,5- Isomers of N-acetyl-S-{tr1chloro-
phenyl)-L-cyste1ne accounted for 60-62% of the urinary metabolites. Minor
urinary metabolites Included 2,4,5- and 2,3,5-tr1chloroth1ophenol and free
2,3,5- and 2,3,4-TCP, which accounted for 28-33 and 1-10% of the material
excreted, respectively.
On the basis of the studies of L1ngg et al. (1982) and Kohll et al.
(1976), 1t Is apparent that there may be differences among species 1n the
metabolism of 1,254-tr1chlorobenzene. It seems likely that these differ-
ences will extend to the other Isomers of trlchlorobenzene as well. Both
reports postulated the same first step 1n metabolism (I.e., Initial forma-
tion of arene oxide Intermediates), but Indicated differences 1n the subse-
quent metabolic reactions. In the rat, conjugation of the Intermediate with
glutathlone was postulated to account for the sulfur-containing urinary
metabolites. In the monkey, hydrolysis of the arene oxide to the dlhydro-
dlol and the absence of sulfur-containing metabolites seemed to preclude the
Involvement of glutathlone (L1ngg et al., 1982). As proposed by Kohll et
9-5
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al. (1976) and Illustrated In Figure 9-1, formation of the 1somer1c tr1-
chlorophenols from the arene oxide Intermediates can proceed either by
direct opening of the C-0 bond or by the NIH shift of chlorine.
Differences 1n the rate of metabolism of the different Isomers within a
species have been attributed to the positions of the chlorine atoms on the
benzene ring, with the presence of two adjacent unsubstltuted carbon atoms
facilitating the formation of the arene oxide Intermediate. Halogenated
benzenes without adjacent unsubstltuted carbons may still be metabolized via
an arene oxide Intermediate but at a reduced rate, and should show evidence
of a NIH shift (Matthews and Kato, 1979).
9.1.4. Excretion. L1ngg et al. (1982) measured the 24-hour excretion of
radioactivity 1n the urine and feces of 16 male Charles River rats and 2
rhesus monkeys given a single 10 mg/kg 1.v. or oral dose of 14C-1,2,4-tr1-
chlorobenzene. In the rat, 84% of the oral dose and 78% of the 1.v. dose
were excreted 1n the urine by 24 hours; 11 and 7%, respectively, were the
amounts Identified 1n the feces 1n the same period. In the monkeys, 40% of
the oral dose and 22% of the Injected dose appeared 1n the urine and <1% 1n
the feces. Smith and Carlson (1980) orally administered 181.5 mg/kg/day (1
mmol/kg/day) of 14C-1,2,4-tr1chlorobenzene 1n corn oil to 4 Sprague-Dawley
rats for 7 days and followed the excretion of radioactivity In the feces and
1n the urine during administration and up to 21 days after the first dose.
Fecal elimination rose slightly during the first 3 days of dosing, after
which 1t declined rapidly and was essentially complete at 15 days of collec-
tion, accounting for -4% of the total dose. Urinary excretion followed a
similar pattern; however, at 21 days after the first dose, radioactivity was
still detectable. Total urinary excretion to this time accounted for ~72%
of the total administered radioactivity. As noted by L1ngg et al. (1982),
9-6
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>
i
1. 1. 1-TCf
\
\
ON
ON
\
NO'
t, i.
J. 4. i-I
1.1,1-fCS
I
a
a
1.1.«-res
\
/ \ /
ON
t. *. •-U»
t. s. •-«»
a
i.tt-19
TO » TWCMKMOMMMt
ttP • fMCtHONOMMMM.
FIGURE 9-1
Metabolic Pathways for TMchlorobenzene (TCB) Isomers Through Arene Oxide Intermediates
1n Rabbits
Source: Adapted from KohH et al., 1976
-------
the differences in the excretion rate between the rat and monkey may be
attributable to their different pathways of metabolism, since the monkey
required two steps beyond the arene oxide to produce Its urinary metabolite,
while the rat required only one.
Differences 1n the rates of excretion between the Isomers of trlchloro-
benzene have also been reported. Jondorf et al. (1955) found that rabbits
given oral doses of 500 mg/kg of 1,2,3-, 1,2,4- or 1,3,5-tr1chlorobenzene
excreted 78, 42 or 9%, respectively, of the administered dose as monophenols
1n the 5-day urine collection.
U.S. EPA (1980b), using data from Williams (1959) and Parke and Williams
(1960), estimated the following half-lives of excretion 1n the rabbit: 2,
5.5 and 8.5 days for 1,2,3-, 1,2,4- and 1,3,5-tr1chlorobenzene, respec-
tively. The rate of metabolism and subsequent excretion 1s most likely
related to the position of the chlorine atoms on the benzene ring. Matthews
and Kato (1979) hypothesized that two adjacent unsubstltuted carbon atoms
facilitate the formation of the arene oxide Intermediate and Increase the
rate of metabolism and excretion.
9.1.5. Summary. The limited comparative pharmacoklnetlc data available
on the trlchlorobenzenes prevent specification of the absorption, distribu-
tion, metabolism and excretion of the Individual Isomers. From the avail-
able data, 1t seems relatively clear that metabolism 1n at least three
species has a common first step, the production of an arene oxide Intermedi-
ate. Subsequent metabolic steps, however, vary among the species examined,
at least for the most studied Isomer, 1,2,4-tr1chlorobenzene.
In general, the pharmacoklnetlcs of the trlchlorobenzenes are similar to
those described for the halogenated aromatlcs by Matthews and Kato (1979).
The authors observed that these compounds are Upophllic and that their
9-8
-------
metabolism and excretion depends on their conversion to polar Intermedi-
ates. In addition, their llpophlUc character provides for ready absorption
from the gastrointestinal tract and Initial distribution to the more highly
perfused tissues, particularly the liver, after which they are either metab-
olized and excreted or redistributed to adipose tissue or skin. Additional
experiments are needed to clarify the relationship of these studies to the
metabolism of trlchlorobenzenes 1n humans.
9,2. EFFECTS IN HUMANS
Information on the health effects of trlchlorobenzenes 1n humans 1s
limited to case reports. Rowe (1975) found that an Individual exposed to
1,2,4-tr1chlorobenzene at 3-5 ppm had eye and respiratory Irritation.
Glrard et al. (1969) reported two cases, one 1n which a 68-year-old woman,
who often soaked her husband's work clothes 1n trlchlorobenzene, developed
aplastlc anemia, and the other 1n which a 60-year-old man, who had been
occupatlonally exposed to DOT as well as to mono-, d1- and trlchlorobenzenes
for over 30 years, developed anemia.
9.3. MAMMALIAN TOXICOLOGY
9.3.1. Acute Tox1c1ty. Studies of the acute toxldty of the trlchloro-
benzenes have been performed 1n several species using various routes of
administration.
Information on the effects of acute Inhalation exposure to trlchloroben-
zenes Is limited. In an abstract of a study from the Russian literature
(Gurfeln and Pavlova, 1960), a single high Inhalation exposure {exposures of
0.005-0.01 mg/2, 1n air or 5-10 mg/m3 were used) of an unspecified Isomer
of trlchlorobenzene to rats resulted 1n Immediate nervousness and plnkness
of mouth, ears and paws. These effects were followed by convulsions and
death within 30 minutes, with edema of livers and kidneys observed upon
9-9
-------
necropsy. Unpublished results of a study performed by Treon (1950) were
reported by Coate et al. (1977) and Indicated that the target organs of non-
lethal acute Inhalation exposure to trlchlorobenzenes (a weight-to-weight
mixture of 8% 1,2,3- and 92% 1,2,4-tr1chlorobenzene) 1n cats, dogs, rats,
rabbits and guinea pigs Included the liver, ganglion cells at all levels of
the brain and mucous membranes. Lethal doses resulted 1n local Irritation
of the lungs and functional changes 1n respiration 1n animals dying after
exposure. Levels and duration of exposure were not given.
Brown et al. (1969) reported the single-dose oral LD_n for l,2,4-tr1-
50
chlorobenzene 1n CFE rats to be 756 mg/kg (95% confidence limits 556-939
mg/kg). In CF mice, the single-dose oral LD was 766 mg/kg (95% confi-
dence limits 601-979 mg/kg). Death occurred within 5 days 1n rats and 3
days 1n mice.
R1m1ngton and Zlegler (1963) studied the porphyr1a-1nduc1ng ability of
1,2,4- and 1,2,3-tr1chlorobenzenes administered by gavage to male albino
rats for various time periods (5-15 days). Doses of the Isomers were gradu-
ally Increased until porphyrln excretion was high but fatalities were few.
Porphyrla was Induced by 1,2,4-tr1chlorobenzene when the Isomer was given
for 15 days at 730 mg/kg (3 rats) as evidenced by peak rises 1n urinary
coproporphyMn, uroporphyrln, porphoblUnogen and 6-am1nolevul1n1c add.
At a dose of 500 mg/kg for 10 days (1n 5 rats), peak liver levels of copro-
porphyrln, protoporphyMn, uroporphyrln and catalase were reached. For the
1,2,3- Isomer, urinary excretion of these Indicators peaked at 785 mg/kg for
7 days (3 rats), but to a lesser extent than for the 1,2,4- Isomer. Only
the liver uroporphyrln levels were Increased by administration of l,2,3-tr1-
chlorobenzene at this dose and duration. Glutathlone was found to have a
protective effect on tr1chlorobenzene-1nduced porphyrla.
9-10
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Brown et al. (1969} determined the single-dose percutaneous L05Q 1n
CFE rats (4 of each sex) to be 6139 mg/kg (95% confidence limits 4299-9056
mg/kg) for 1,2,4-tr1chlorobenzene administered topically on the shorn dorso-
lumbar skin and covered with an Impermeable dressing. All deaths occurred
within 5 days. In skin Irritation studies, 1,2,4-trlchlorobenzene was
applied to the skin of rabbits and guinea pigs. In the first experiment.
two 2x2 cm patches of Unt, each containing 1 ml, of the compound, were
applied to the shorn backs of rabbits (4 of each sex) for 6 hours/day for 3
consecutive days and covered with an Impermeable dressing. For another
experiment, rabbits (1 of each sex) and guinea pigs (5 of each sex) received
single uncovered applications of 1,2,4-tr1chlorobenzene on the shorn mid-
dorsal skin (1 mS. for rabbits, 0.5 mi for guinea pigs) 5 days/week for 3
weeks. The results Indicated that trlchlorobenzene was not very Irritating,
although flssurlng was noted during the 3-week exposure. Some guinea pigs
that died during the 3-week regimen had focal necrosis of the liver.
Hepatotoxic effects (fatty Infiltration and necrosis) were reported by
Cameron et al. (1937) following s.c. and/or 1.v. Injection of 500 mg (range
of doses was 1-500 mg) trlchlorobenzene 1n liquid paraffin to rats; the
toxlclty was less than that of mono- and o-d1chlorobenzene. Further details
of strain, number of animals or Isomers were not reported.
Robinson et al. (1981), 1n an acute toxldty study to assess the
Increased adrenal weight which was noted in a multlgeneration study, gave
groups composed of 9-10 preweanlng female Charles River rats 1.p. Injections
of 0, 250 or 500 mg of 1,2,4-tr1chlorobenzene/kg in corn oil at 22, 23 and
24 days of age. Significant changes (p<0.05) from control values were
9-11
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observed upon necropsy at 25 days of age as follows: decreased body weight
and Increased adrenal weight at the high dose; decreased uterus and
Increased liver weights at both doses.
Hale Holtzman rats (number not specified) were given single Intraperl-
toneal Injections of 1,2,4- or 1,3,5-tr1chlorobenzene at a dose of 37 mg/kg
(5 mmol/kg) as a 50% solution 1n sesame oil 1n a volume of 1 ml/kg (Yang
et a!., 1979). Controls received an equal volume of sesame oil. After 24
hours, the femoral veins and the common bile duct were cannulated. Both
Isomers produced significant Increases (p<0.05) In bile duct-pancreatic
fluid (BDPF) flow with the 1,2,4- Isomer being 4 times more effective than
the 1,3,5- Isomer. S6PT activity was elevated by treatment with l,3,5-tr1-
chlorobenzene and bile flow was elevated by the 1,2,4- Isomer. Both Isomers
caused a decrease 1n BDPF protein concentration.
Several studies have demonstrated the ability of the trlchlorobenzenes
to enhance xenoblotlc metabolism. Carlson, 1n a series of reports (Carlson
and Tardlff, 1976; Carlson, 1977a, 1978, 1981; Smith and Carlson, 1980),
examined the ability of 1,2,4-trlchlorobenzene to Induce a variety of mlcro-
somal functions and enzymes Including cytochrome c reductase, 0-ethyl
0-p-n1trophenyl phenylphosphothlonate (EPN) detoxification, cytochrome
P-450, glucuronyltransferase, benzpyrene hydroxylase and azoreductase. In a
14-day study by Carlson and Tardlff (1976), dally doses of 1,2,4-trlchloro-
benzene 1n corn oil were administered orally to groups of 6 male albino rats
at 10, 20 and 40 mg/kg. All the above functions and enzymes Increased sig-
nificantly (p<0.05) except benzopyrene hydroxylase. In a 90-day study by
the same Investigators, all the functions and enzyme activities, Including
benzopyrene hydroxylase, Increased significantly (p<0.05) at 10-40 mg/kg/day
and remained significantly elevated after a 30-day recovery period. In a
9-12
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similar study, Smith and Carlson (1980) administered 1,2,4-tr1chlorobenzene
at 181.5 mg/kg/day (1 mmol/kg/day) to rats for 7 days, and measured recovery
at 1, 6, 11 and 16 days. EPN detoxification was still significantly
(p<0.05) elevated at 11 days; p-n1troan1sole demethylatlon at 16 days; cyto-
chrome c reductase at 6 days; and cytochrome P-450 at 11 days. In a similar
study by Carlson (1977b), 7-day administration of 1,3,5-tr1chlorobenzene at
100-200 mg/kg/day significantly (p<0.05) Increased EPN detoxification, UOP
glucuronyltransferase, and cytochrome c reductase, and significantly
decreased hepatic 6-6-P; benzpyrene hydroxylase, azoreductase and serum
Isodtrate dehydrogenase were not significantly affected at 200 mg/kg/day.
In the same study, In vivo hepatotoxldty of carbon tetrachlorlde (one dose
of 0.5 ml/kg) was significantly (p<0.05) enhanced by 14-day pretreatment
of rats with 1,2,4-trlchlorobenzene. Glucose-6-phosphatase activity was
significantly (p<0.05) decreased by pretreatment with 1,2,4-trlchlorobenzene
at 5 mg/kg/day, and Isodtrate dehydrogenase was decreased by pretreatment
at 20 mg/kg/day.
The 1,2,4- Isomer, and to a lesser extent the 1,3,5- Isomer, were also
shown to Induce hepatic esterases (Carlson et al., 1979; Carlson, 1980). In
studies similar to those previously described, rats receiving dally oral
doses of 18.2 mg 1somer/kg (0.1 mmol/kg) for 14 days were killed 24 hours
later and hepatic mlcrosomes were prepared. The l,2,4-1somer was an
effective Inducer of both acetanlUde esterase and acetanlUde hydroxylase,
while the l,3,5-1somer Induced only the esterase and to a lesser degree than
did 1,2,4-trlchlorobenzene (Carlson et al., 1979). The l,2,4-1somer also
Induced hepatic arylesterase, while 1,3,5-tr1chlorobenzene did not (Carlson,
1980). Pretreatment of rats with 181.5 mg/kg/day (1 mmol/kg/day) of either
Isomer resulted 1n Induction of procalne esterase (Carlson et al., 1979).
9-13
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In a series of experiments, Ar1yosh1 et al. (1975a,b,c) studied the
effects of the tMchlorobenzenes on Induction of hepatic mlcrosomal pro-
teins, phosphollplds and enzymes, especially 1n relation to the activity of
J~am1nolevul1n1c add synthetase, the rate limiting enzyme 1n the bio-
synthesis of heme. The three tMchlorobenzene Isomers were administered
orally to groups of 2-6 female Wlstar rats at a dose of 250 mg/kg/day for 3
days, after which the rats were killed and mlcrosomes were prepared. The
results Indicated that trlchlorobenzenes Increased the levels of mlcrosomal
proteins, phosphollplds and cytochrome P-450, and enhanced the activities of
aniline hydroxylase, amlnopyrlne demethylase and A-am1nolevul1n1c add
synthetase, with the l,2,4-1somer being the most effective (Ar1yosh1 et al.,
1975a,b). The dose response of these effects to 1,2,4-tr1chlorobenzene were
determined (Ar1yosh1 et al., 1975c) for groups of 2-6 female Wlstar rats
treated orally with single doses of 0, 125, 250, 500, 750, 1000 and 1500
mg/kg. The results Indicated that 24 hours after the administration of the
Isomer, mlcrosomal protein was elevated at >750 mg/kg and glycogen content
was decreased at >500 mg/kg. The activities of amlnopyrlne demethylase and
aniline hydroxylase and the content of cytochrome P-450 were Increased at
>250 mg/kg, as was 6-am1nolevul1n1c add synthetase activity.
9.3.2. Subchronlc Toxldty. The effects of trlchlorobenzene following
subchronlc Inhalation, as well as oral and dermal exposure, have been
Investigated 1n a variety of species. Toxldty data for the trlchloroben-
zenes can be found 1n Table 9-2.
Kodba et al. (1981) exposed 20 male Sprague-Dawley rats, 4 male New
Zealand rabbits and 2 male beagle dogs by Inhalation to 1,2,4-tMchloroben-
zene (99.4X pure) at levels of 0, 223 mg/m3 (30 ppm) or 742 mg/m3 (100
ppm) for 7 hours/day, 5 days/week for a total of 30 exposures 1n 44 days.
9-14
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TABLE 9-2
Suranary of Subchronk and Chronic Toxlclty Studies on Trlchlorobenzenes
Species Route
Rat Inhalation
Rats, rabbits, Inhalation
two dogs
Rat Inhalation
Rat Inhalation
VD
i
_j
tn
Rabbits, Inhalation
monkeys
Honkey oral
Rat oral
Rat oral
House oral
Dose
74,2, 742 or
7423 ng/in»
of 1,3,5-TGi
223 or 742 mg/m»
of 1,2,4-TCB
22.3 or
74.2 iflg/ffl»
of 1,2,4-TCB
186, 371 or
742 mg/m»
of 1,2,4-TCB
186, 371 or
742 rag/m*
of 1,2,4-TCB
1, 5, 25, 90,
125 or 173.5
mg/kg/day
of 1,2.4-TCB
50, 100 or
200 rug/kg/day
of 1,2,4-TCB
10, 20 or
40 mg/kg/day
of 1,2.4-TCB
600 ppm diet
(0.078 mg/kg/
day) of
1,2,4-TCB
Duration
6 hr/day, 5 day/wk
for up to 13 wk
7 hr/day, 5 day/wk;
total of 30 expo-
sures In 44 days
6 hr/day, 5 day/wk,
3 no
7 hr/day, 5 day/wk,
26 wk
7 hr/day, 5 day/wk,
26 wk
30 days
30, 60, 90 or
120 days
90 days
6 mo
Effects Reference
No hepatotoxlclty; three high-dose rats had Sasmora and Palmer,
squamous metaplasia and focal hyperplasla 1981
of respiratory epithelium, believed to be
reversible
Increase 1n urinary excretion of porphyrla Koclba et al., 1981
1n exposed rats; Increase In liver weights
In high-dose rats and dogs; Increased kid-
ney weights In high-dose rats
Increase In urinary porphyrln excretion In Uatanabe et al., 1978
high-dose rats; no effects In 22.3 mg/u"
group
Enlarged hepatocytes and nondose-dependent Coate et al., 1977
hepatocytes vacuollzatlon, liver granulance,
biliary hyperplasla and kidney hyaline de-
generation at 4 and 13 wk; no hlstopathology
evident at 26 wk
No treatment related changes at 26 wk . Coate et al., 1977
<25 mg/kg/day - no effects observed; Smith et al., 1978
>90 mg/kg/day - observed toxic Hy and death
Increases In liver weights, liver porphyrlns Carlson, 1977b
and urine porphyrlns, dose and time related
Increase In llver-to-body weight ratio In Carlson and Tardlff,
high-dose group; changes In enzyme actlva- 1976
tlon at all doses
Mo effects Goto et al., 1972
-------
TABLE 9-2 (cent.)
Species
Guinea pig
Mouse
Rats
Rats
Rabbits
Route Dose
dermal 0.5 ml/day
of 1,2,4-TCB
dermal 0.003 ml/paint-
ing of 30 and
60X solution 1n
acetone of
1,2,4-TCB
oral 25, 100 or
(drinking 400 mg/i
water) of 1,2,4-TCB
oral 36, 120, 360 or
1200 mg/kg/day
of 1,2,4-TCB
dermal 30, 150 or
450 mg/kg/day
of 1,2,3-TCB
Duration
5 day/wk, 3 wk
2 t1mes/wk, 2 yr
FQ to F2
generations
days 9-13 of
gestation
5 day/wk, 4 wk
Effects
Death following extensor convulsion; livers
showed necrotlc fod
Painting Induced excitability, panting and
epidermal thickening, Inflammation and
kerat1n1zat1on; Increased organ weights and
mortality
Enlarged adrenals 1n FQ and F] generations
1200 mg/kg dose all dead by the 3rd day,
360 mg/kg dose caused 22% mortality In
dams and moderate hepatocellular hyper-
trophy and non-significant Increases In
embryonic lethality and significantly
retarded embryonic development, 36 and
120 mg/kg groups not observed for embryonic
effects, but slight hepatocellular hyper-
trophy was reported 1n one 120 mg/kg dam
Dose-related skin Irritation; Increase 1n
urinary coproporphyrln In high-dose males
and slight pallor of liver 1n males and
females
Reference
Brown et al., 1969
Yamamoto et al., 1957
Robinson et al., 1981
Kltchln and Ebron,
1983a
Rao et al.. 1982
1,2,3-TCB = 1,2,3-tMchlorobenzene; 1,2,4-TCB = 1,2,4-tHchlorobenzene; 1,3,5-TCB - 1,3,5-tMchlorobenzene
-------
There were no significant effects on body weight, hematologlc Indices or
serum biochemistry tests. Upon necropsy, gross and comprehensive hlstologl-
cal examination revealed no significant treatment-related effects In any of
the species. At the 742 mg/m3 level, Increased liver weights were
detected 1n dogs and rats and Increased kidney weights In rats. Urinary
excretion of porphyrln was Increased In rats exposed to 1,2,4-tr1chloroben-
zene at 223 or 742 mg/m3, which the Investigators Interpreted as a com-
pound-specific physiologic effect rather than a toxic effect. A follow-up
study supported this Interpretation. The same Investigators exposed male
and female Sprague-Oawley rats to 1,2,4-trlchlorobenzene at 0, 22.3 mg/m3
(3 ppm) or 74.2 mg/m3 (10 ppm) for 6 hours/day, 5 days/week for 3 months.
The results, which were reported 1n an abstract (Watanabe et al., 1978),
Indicated that urinary excretion of porphyrlns was slightly Increased 1n the
74.2 mg/m3 group during exposure, but returned to control range 2-4 months
post-exposure. Since this appeared to be the most sensitive Indicator 1n
rats, and exposure to trlchlorobenzene at 22.3 mg/m3 did not cause
Increased porphyrln excretion, 22.3 mg/m3 was considered a no-observed-
adverse-effect level (NOAEL) for rats by the authors.
Sasmore and Palmer (1981) exposed male and female outbred albino CO rats
(20/group) to 1,3,5-trlchlorobenzene vapor at 0, 74.2 mg/m3 (10 ppm), 742
mg/ms (100 ppm) or 7423 mg/m3 (1000 ppm) for 6 hours/day, 5 days/week
for up to 13 weeks. No significant effects were observed on body weights,
food consumption, standard hematologlc and clinical chemistry parameters or
on methemoglobln and porphyrln levels. In a subgroup of animals killed
after 4 weeks of exposure, there was a significant Increase In the I1ver-to-
body weight and I1ver-to-bra1n weight ratios 1n the male rats of the high
exposure level group, but these effects were not observed at 13 weeks.
9-17
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Since gross and microscopic pathologic examinations of the liver revealed no
treatment-related abnormalities, the authors concluded that the exposure did
not cause hepatotoxldty. Microscopic examinations, however, revealed that
three high dose rats had squamous metaplasia and focal hyperplasla of the
respiratory epithelium, which the authors believed to be reversible.
Coate et al. (1977J exposed groups of 30 male Sprague-Oawley rats, 16
male New Zealand rabbits and 9 male monkeys (Hacaca fasc1cu1ar1s) to 99.07%
pure 1,2,4-trlchlorobenzene vapor at levels of 0, 186 mg/ma (25 ppm), 371
mg/m3 (50 ppm) or 742 mg/ma (100 ppm) for 7 hours/day, 5 days/week for
26 weeks. Pulmonary function and operant behavior tests In the monkeys,
ophthalmic examinations 1n the rabbits and monkeys, and measurements of body
weight, hematologlc Indices and serum biochemistry parameters 1n all species
were conducted before and during the exposure period. Subgroups of 5 rats
each were killed after 4 and 13 weeks of exposure; all remaining rats were
killed after 26 weeks for hlstologlcal examination of selected tissues. No
treatment-related effects at any observation time were seen with respect to
body weight, survival, hematology or serum chemistry for any of the
species. No ophthalmic changes were observed 1n rabbits or monkeys. Pul-
monary function and operant behavior were unaffected In monkeys. Hlstologl-
cal examination of rat tissues revealed that treated animals had enlarged
hepatocytes that were more prominent at 4 weeks than at 13 weeks after expo-
sure, and at 371 and 742 mg/m3 than at 186 mg/m3. Other changes In
treated rats that did not appear to be dose-dependent were vacuollzatlon of
hepatocytes at 4 and 13 weeks, slightly more severe granuloma of the Hver
at 4 weeks and biliary hyperplasla at 4 and 13 weeks. A nondose-related In-
crease 1n the severity of kidney hyaline degeneration was observed 1n test
rats at 4 weeks. This lesion was slightly more severe 1n the high dose
9-18
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group at 13 weeks. These effects appeared to be transient; rats necropsled
after 26 weeks of exposure had none of these changes. Likewise, hlstologl-
cal examination of selected tissues from rabbits and monkeys revealed no
treatment-related changes after 26 weeks of exposure.
Carlson and Tardlff (1976) assessed the effects of 14- or 90-day oral
administration of 1,2,4-tr1chlorobenzene 1n corn oil compared to corn oil
controls In male CD rats. In the 14-day studies, the effects examined were
lethality, hepatotoxldty and the Influence on hexabarbltal sleeping time
and other parameters of xenoblotlc metabolism. A dose of 600 mg/kg/day, the
highest dose administered, caused no deaths during the 14-day administration
period. Hepatotoxldty was evaluated by dosing at 0, 150, 300 or 600
mg/kg/day and determining serum 1soc1trate dehydrogenase and liver glucose-
6-phosphatase activities. Although no dose-related changes 1n serum iso-
cltrate dehydrogenase activity was observed, Hver glucose-6-phosphatase
activity was significantly decreased at >300 mg/kg (p<0.05). Hexabarbltal
sleeping time was significantly decreased at 600 mg/kg/day (the only dose
examined); this effect persisted through a 14-day recovery period. In rats
receiving 14 dally doses at 0, 10, 20 or 40 mg/kg, there was a significant
dose-related Increase 1n Hver-to-body weight ratio at >10 mg/kg/day
(p<0.05). Significant dose-related Increases were also observed 1n activi-
ties or contents of cytochrome c reductase (at >10 mg/kg), cytochrome P-450
(at >2Q mg/kg), glucuronyltransferase (at >2Q mg/kg), azoreductase (at >10
mg/kg) and the rate of detoxlcatlon of EPN (at >10 mg/kg). These results
Indicated that the doses, while causing a slight degree of hepatic Injury,
significantly enhanced xenoblotlc metabolism.
In the 90-day studies by Carlson and Tardlff (1976), the effects of oral
dosing of male CD rats (6 animals/group) at 0, 10, 20 or 40 mg/kg/day with
9-19
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1,2,4-tMchlorobenzene 1n corn oil on weight gain, liver weight, hemoglobin
content, packed cell volume and the Indicators of xenoblotlc metabolism were
evaluated. No effects on weight gain and no consistent alteration 1n hemo-
globin content or packed cell volume were observed. At 40 mg/kg, there was
a statistically significant Increase (p<0.05) 1n I1ver-to-body weight ratios
that persisted throughout a 30-day recovery period. Following 90-day admin-
istration, cytochrome c reductase activity was Increased at >10 mg/kg, with
recovery after 30 days; cytochrome P-450 levels Increased at >20 mg/kg, fol-
lowed by recovery; glucuronyltransferase activity decreased at >10 mg/kg;
EPN detoxlcatlon Increased at >20 mg/kg; benzopyrene hydroxylase activity
Increased 2-fold at 40 mg/kg; and azoreductase activity Increased at >10
mg/kg.
Groups of 5 female rats (strain not reported) received dally oral doses
of 0, 50, 100 or 200 mg 1,2,4-tr1chlorobenzene/kg/day 1n corn oil for 30,
60, 90 or 120 days (Carlson, 1977b). Significant Increases were observed 1n
liver porphyrlns at >100 mg/kg after 30 days exposure and 1n urinary
porphyrlns at 200 mg/kg after 30 days. For the 30-day study, slight but
significant Increases were also observed 1n liver weights at 200 mg/kg.
When the compound was administered for 60 days, only the liver weights were
Increased. The administration of 1,2,4-tr1chlorobenzene for 90 days
resulted 1n slight but significant Increases 1n liver weights at >50 mg/kg,
1n liver porphyrlns at >100 mg/kg and 1n urine porphyrlns at 200 mg/kg. A
significant Increase was observed for liver porphyrlns when the compound was
given at >50 mg/kg for 120 days. The excretion of 4-am1nolevul1n1c add
and porphoblUnogen 1n the urine was not Increased at any dose given for any
duration. When the author compared the 1,2,4-tMchlorobenzene results with
the results for hexachlorobenzene, he concluded that tMchlorobenzene
9-20
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Induced porphyrla was very small compared to the hexachlorobenzene Induced
porphyrla (Carlson, 1977b).
A 90-day oral study by Smith et al. (1978), reported 1n an abstract, was
reviewed by U.S. EPA (1980b), who gave further details of the study after
communication with the authors. Rhesus monkeys (4/group) were given 1,2,4-
trlchlorobenzene 1n dally oral doses of 1, 5, 25, 90, 125 or 173.6 mg/kg.
No toxic effects were observed at <25 mg/kg, while doses of >90 mg/kg were
observed to be toxic, and the 173.6 mg/kg dose was lethal within 20-30 days.
There were no deaths observed 1n the 1, 5 and 25 mg/kg groups; one death
occurred in each of the 90 mg/kg and 125 mg/kg groups and two deaths
occurred in the 173.6 mg/kg group. Animals on the highest dose exhibited
severe weight loss and predeath fine tremors. All of the animals 1n the
highest dose group had elevated BUN, Na*. K*. CPK, S60T, SGPT, LDH and
alkaline phosphatase as well as hypercalcemla and hyperphosphatemla from 30
days on. Smith et al. (1978) have been using the urinary pattern of chlor-
guanide metabolites as an Indication of cytochrome P-450 dependent drug
metabolism. At the high doses, monkeys showed evidence of the hepatic
Induction as well as Increased clearance of 1.v. doses of labeled l,2,4-tr1-
chlorobenzene. Further Information on the study (Smith, 1979) gave evidence
of liver enzyme Induction 1n the 90, 125 and 174 mg/kg animals. There were
some pathological changes noted 1n the livers of the high dose groups,
primarily a fatty Infiltration. The point at which there was no effect
related to the compound was at the 5 mg/kg level. Since only an abstract of
this study was available and since the Interpretation of this study was
complicated by the use of other drugs and weight losses 1n the control
animals, a valid no-observed-effect level (NOEL) cannot be derived from
these data.
9-21
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Two subchronlc studies have assessed the dermal toxldty of the trlchlo-
robenzenes. Powers et al. (1975) applied technical grade 1,2»4-tr1ehloro-
benzene at concentrations of 5 or 25% 1n petroleum ether, or 10054 l,2,4-tr1~
chlorobenzene topically 1n 0.2 mfi, volumes to the ventral surface of the
ears of New Zealand rabbits (groups of 12 each), 3 times weekly for 13
weeks; a control group received petroleum ether only. Rabbits exposed to 5%
trlchlorobenzene and controls had slight redness and scaling. Dermal
responses at 25 and 100% of the compound Included slight to severe erythema,
severe scaling, desquamatlon, encrustation, and some hair loss and scarring.
The responses were characterized by acanthosls and keratosls, typical of
moderate to severe Irritation and probably attributable to degreaslng
action. No overt signs of systemic toxldty were noted, body weight gain
was comparable 1n all groups, and none of the animals showed meaningful
changes 1n gross pathology. The Investigators noted that this contrasted
with the findings of Brown et al. (1969), who reported that some guinea
pigs, exposed topically to 1,2,4-tr1chlorobenzene at 0.5 mil/day, 5 days/
week for 3 weeks, died following extensor convulsions and their livers
showed necrotlc foci. This difference 1n results may be attributed to the
site of application (Brown et al., 1969, used the dorsal rnldHne for appli-
cation, a more extensive exposure site), the volume applied (0.5 ma vs.
0.2 mil), the species used, and more frequent (5 times/week vs. 3 times/
week) application, although the total number of exposures was less (5x3
weeks vs. 3x13 weeks).
Rao et al. (1982) applied technical grade trlchlorobenzene [1,2,4- (70%)
and 1,2,3-tr1ehlorobenzene (30%)] 5 days/week for 4 weeks, at doses of 0,
30, 150 or 450 mg/kg/day, to the dorsal skin (4x4 Inch area) of groups (5 of
each sex) of New Zealand rabbits weighing ~3 kg. One rabbit died after 18
applications, but the Investigators were unable to determine the cause of
9-22
-------
death by either gross or hlstologlc examination. Gross and histologic
examination of the skin showed evidence of moderate Irritation at the high-
est dose and less Irritation at the lower doses. This Irritation evidence
consisted of epidermal scaling, thickening, fissures, ulcers and erythema.
No treatment-related change was observed 1n clinical chemistry (BUN, glu-
cose, SGPT, SAP) or hematology. A slight but significant Increase 1n
urinary coproporphyrln was observed 1n high-dose males (450 mg/kg/day) at
day 24; none was seen 1n females. This slight porphyrla and a slight gen-
eralized pallor of the liver (3/5 males, 4/4 females) were the only signs of
systemic toxldty. Extensive hlstologlc examination of numerous tissues
failed to show any treatment-related abnormalities. The volume of tr1-
chlorobenzene applied at the dose levels 1n this study can be calculated as
=0.06 mS, (30 mg/kg), 0.31 ml (150 mg/kg) and 0.93 mi(450 ing/kg) by
multiplying the dose 1n g/kg by the weight of the rabbits (3 kg) and divid-
ing by the density of trlchlorobenzene (1.45).
9.3.3. Chronic Tox1c1ty. No studies on the effects of the trIchloroben-
zenes following chronic Inhalation exposure were available for review; how-
ever, a chronic skin painting study was encountered. Goto et al. (1972)
conducted a 6-month feeding study In mice using hexachlorocyclohexane
Isomers and their metabolites, Including 1,2,4-tr1chlorobenzene. Male mice
(20/group) of the ICR-JCL strain (age at Initiation 5 weeks, average weight
26.5 g) received a diet containing 600 ppm of trlchlorobenzene (78 yg of
compound/kg body weight, assuming mice consume 13% of their body weight 1n
food per day). The weight gain of treated mice did not differ from controls
during the 6-month exposure. At 26 weeks, 10 mice were killed and liver,
heart and kidneys were weighed; no abnormal weight changes were observed.
Macroscopic and hlstologlc examination of the Hver revealed no hepatic
tumors or any other lesions,
1832A 9-23
-------
Yamamoto et al. (1957) studied the toxlclty of 1,2,4-trlchlorobenzene
when painted on the skin of Slc:ddy mice 2 tiroes/week for 2 years. Groups
consisted of 75 mice/sex receiving 0.03 mi applications of the compound as
30 or 60% solutions 1n acetone. Controls consisted of 50 mice/sex and
received only acetone. The skin painting produced general symptoms of
excitability and panting, local skin thickening, kerat1n1zat1on and Inflam-
mation of the epidermis. These effects were not observed 1n controls. For
the 30% trlchlorobenzene groups, mortality was Increased 1n females (5/75
survived for 83 weeks compared with 11/50 controls). The mean survival days
were 357+125.4 for treated females compared with 423.8+145.0 for controls
(p<0.01). The survival of males at this exposure level was not signifi-
cantly different from that of controls. Spleen weights were significantly
Increased (p<0.05) and left adrenal weights were significantly decreased
(p<0.01) for treated males when compared with controls. Hematologlc and
blood chemistry Indices were essentially unchanged with the exception of
Increased red blood cell counts 1n treated males (p<0.05) and decreased
Cl" concentration (p<0.01). For the 60% solution, 6/75 treated females
survived for 83 weeks. Mean survival days were 320.2+147.7 for treated
females compared with 423.8+145.0 for controls (p<0.001). Eight of 75
treated males survived for 83 weeks compared with 9/50 control males. Mean
survival days were 288.0+.173.7 for treated males and 363.9+173.9 for con-
trols (p<0.05). Significant differences 1n organ weights from control
values were seen 1n the spleens of males (p<0.01) and the adrenals of
females (p<0.05). Hematologlc and blood biochemistry changes were seen 1n
Increased lymphocyte counts 1n treated females (p<0.05), and 1n Increased
SCOT (p<0.05), SGPT (p<0.001) and BUN (p<0.01) for treated males.
9,3.4. Mutagenldty. Schoeny et al. (1979) and Lawlor et al. (1979)
examined the mutagenlc potential of 1,2,4-trlchlorobenzene 1n Salmonella
9-24
-------
typhlnmrlum tester strains TA98, TA100, TA1535 and TA1537, using the plate
Incorporation technique. Schoeny et al. (1979) used 8 concentrations of
trlchlorobenzene ranging from 102 pg/plate to 1.4xlOs pg/plate. The
toxic dose was determined as 1599 pg/plate (killing of one or more strain
on mutagenesls plates). Trlchlorobenzene was negative for mutagenldty 1n
the absence and presence of S-9 mlcrosomal fractions from unlnduced rats,
from rats Induced by the polychlorlnated blphenyl, Aroclor 1254, and from
rats homologously Induced with trlchlorobenzene.
The study of Lawlor et al. (1979), reported 1n an abstract, used the
TA1538 strain of S. typhlmurlum 1n addition to the strains previously men-
tioned. Negative results were obtained for five unspecified concentrations
tested 1n the presence and absence of rat liver mlcrosomes Induced by
Aroclor 1254. Because these results were reported 1n an abstract without
the details of the experimental procedures used, the results cannot be crit-
ically evaluated.
The negative results 1n the Salmonella hlstldlne reversion assay are not
unexpected because this test system Is generally Insensitive to highly
chlorinated compounds (Rinkus and Legator, 1980).
9.3.5. Car dnogenl city. Yamamoto et al. (1957) applied 1,2,4-trlchloro-
benzene In acetone to the skin of Slc.ddy mice twice weekly for 2 years.
The solution of 1,2,4-trlchlorobenzene was 60% for the high dose and 30% for
the low dose and the volume applied was 0.03 ml/application. Each treated
group contained 75 animals and there were 50 control animals for each sex.
Growth rates 1n treated and control mice were comparable through 83 weeks.
Mean survival days were significantly reduced in the 60% 1,2,4-trlchloroben-
zene groups of males and females and also 1n the 30% treatment group of
females.
9-25
-------
Hlstopathology showed some organs sites had Increased non-neoplast1c
lesions. Assuming that all 75 animals 1n the treated groups were examined
and all 50 1n the control groups were examined, there would be Increases 1n
lesions 1n the males 1n lung, liver, kidney, adrenal, spleen and lymph node
at the high dose, and 1n all of these organs except lymph node 1n the
females at the high dose. Unfortunately, the English translation of
Japanese text 1s not very specific 1n describing the nature of the lesion
making 1t difficult to use this Information 1n the Interpretation of the
tumor findings.
No single tumor type was Increased significantly over the control Inci-
dence but among males nine different tumors were found 1n the high dose
group as compared with two 1n the low dose and two 1n the control group. In
females there were 11 different tumors In the high dose group as compared
with three In the low dose and eight 1n the control group. The authors do
not state whether these tumors were all found 1n different Individual ani-
mals or whether these were multiple tumors 1n the same animal. Therefore,
the actual Incidence 1n terms of the number of tumor bearing animals 1s not
known.
Further Information from this study 1s necessary for full Interpreta-
tion. This single study 1s clearly Inadequate for making any conclusions
about carc1nogen1c1ty 1n humans.
9.3.6. Reproductive and Teratogenlc ToxIcHy. Studies on the reproduc-
tive or teratogenlc effects of trlchlorobenzenes following Inhalation expo-
sure were not found 1n the available literature. Robinson et al. (1981)
reported a multlgeneratlon study of the reproductive effects of l,2,4-tr1-
chlorobenzene following oral administration. Charles River rats were con-
tinuously exposed to the compound at 0, 25, 100 or 400 ppm 1n drinking
9-26
-------
water. The authors calculated the dosages for the Fp generation based on
water consumption data to bet for females at 29 days of age, 8,3+0.8,
28.(HI,2, 133.2+13.4 mg/kg/day, respectively; for males at 29 days of age,
8.5+0.6, 27.6+J.6, 133.6+15.6 mg/kg/day, respectively; for females at 83
days of age, 3.7+0.1, 14.8+J.O, 53.6^3.9 mg/kg/day, respectively; for males
at 83 days of age, 2.5+0.1, 8.9+0.3, 33.0+J.4 mg/kg/day, respectively. The
exposure period began with the birth of the FQ generation and continued
through 32 days of age of the F? generation. Each treatment group consis-
ted of 17-23 Utters. No treatment-related effects were noted with respect
to fertility, neonatal weights, maternal weights, Utter sizes, preweanlng
viability or postweanlng growth 1n any generation. Treatment-related dif-
ferences were seen with respect to food Intake and water consumption 1n F
males and females, but they were Inconsistent and did not occur 1n other
generations. Blood chemistry analyses and locomotor activity measurements
revealed no overt hematologlc or neurologic effects, and hlstologlcal exami-
nation of the livers and kidneys of the F. generation rats revealed no
damage. At the 400 ppm dose level, significantly enlarged adrenals 1n both
sexes of the F and F rats were observed at 95 days of age (p<0.006).
A follow-up acute toxldty study showed that this effect could result from
three dally 1.p. Injections of 500 mg 1,2,4-tr1chlorobenzene/kg.
Black et al. (1983) reported 1n an abstract a teratogenlclty study In
pregnant Wlstar rats using 1,2,4-, 1,2,3- or 1,3,5-tr1chlorobenzene adminis-
tered by gavage 1n doses of 75-600 mg/kg on days 6-15 of gestation (gesta-
tlonal day 0 or 1 not defined). Upon necropsy (gestatlonal day not speci-
fied), thyroid and liver lesions and reduced hemoglobin and hematocrlt
values were observed 1n treated dams (doses not specified). No teratogenlc
effects were observed In the pups; however, pups exposed to the 1,2,4- and
1,3,5- Isomers {doses not specified) had mild osteogenlc changes.
9-27
-------
K1tch1n and Ebron (1983a) conducted a maternal hepatic toxlclty and
embryotoxlclty study where they administered 1,2,4-tr1chlorobenzene (>99%
pure) dissolved 1n corn oil (2 ml/kg) orally to pregnant Sprague-Dawley
(CD strain) rats (6 or more/group) on days 9-13 of gestation and the dams
were then sacrificed on day 14 of gestation. The dosing groups were 0 (corn
oil only), 36, 120, 360 and 1200 mg/kg/day 1,2,4-tr1chlorobenzene. All the
dams 1n the 1200 mg/kg/day group died by the third day of dosing, the 360
mg/kg/ day group were observed with a maternal mortality rate of 22% and
greatly reduced body weight gains. Maternal liver weights, liver/body
weight ratios and hepatic mlcrosomal protein content were not affected by
1,2,4-tr1chlorobenzene administration. 1,2,4-Tr1chlorobenzene was observed
to be a strong Inducer of hepatic enzymes at the 120 and 360 mg/kg/day dose
levels. Liver histology 1n the pregnant dams was unremarkable 1n the 36
mg/kg/day group, showed a slight degree of hepatocellular hypertrophy 1n 1
of 9 rats 1n the 120 mg/kg/day group and showed a moderate hepatocellular
hypertrophy 1n 7 of 8 rats 1n the 360 mg/kg/day group. The uteri from only
the 0 and 360 mg/kg/day groups were examined for 1,2,4-tr1chlorobenzene-
Induced embryonic effects. No statistically significant differences 1n
resorptlon, embryolethaHty or abnormalities were reported, although 3/12
treated Utters showed embryolethallty as compared to 0/12 1n the control
Utters. Several embryonic parameters were significantly decreased by
1,2,4-tr1chlorobenzene treatment. These parameters were embryonic head
length, crown-rump length, somite number and total embryo protein content
(reduced 23%).
9.4. INTERACTIONS
Several studies discussed 1n Section 9.3.1. on acute toxldty have
demonstrated that the Isomers of trlchlorobenzene are capable of affecting
9-28
-------
xenoblotlc metabolism by Inducing a variety of the hepatic drug-metabolizing
enzymes 1n rats. These Include cytochrome c reductase, cytochrome P-450,
glucuronyltransferase, benzpyrene hydroxylase, azoreductase (Carlson and
Tardlff, 1976; Carlson, 1977, 1978, 1981; Smith and Carlson, 1980), aceta-
nH1de esterase and acetanlUde hydroxylase, procalne esterase (Carlson et
al., 1979), arylesterase (Carlson, 1980), mlcrosomal proteins, phosphollplds
and amlnopyrene hydroxylase (Arlyoshl et al., 1975a,b,c). That trlchloro-
benzenes enhance xenoblotlc metabolism has been demonstrated by Smith and
Carlson (1980) and Carlson (1977a), who showed that administration of
1,2,4- or 1,3,5-tr1chlorobenzene to groups of 4 male Sprague-Dawley rats for
7 days Increased EPN detoxlcatlon. The administration of 1,2»4-tr1chloro~
benzene to pregnant rats was also reported to Induce hepatic levels of cyto-
chrome P-450, cytochrome c reductase, UDP glucuronyltransferase and
glutathlone S-transferase (KHchln and Ebron, 1983a).
Townsend and Carlson (1981) demonstrated that 1,2,4-tr1chlorobenzene,
administered by gavage 1n corn oil to groups of five male Swiss mice at
181.5 mg/kg (1 mmol/kg) for 7 days, Increased the ID™ and protected the
mice against the toxic effects of malathlon, malaoxon, parathlon and para-
oxon when graded doses of these Insecticides were administered on the day
following the last dose of trlchlorobenzene.
Experiments comparing the effects of trlchlorobenzenes with the effects
of phenobarbltal and 3-methylcholanthrene Indicated that the Inductions of
mlcrosomal enzymes by trlchlorobenzenes are of the phenobarbltal type
(Carlson, 1978).
9.5. SUMMARY
The trlchlorobenzenes appear to enter the systemic circulation readily
via Inhalation, ingestlon and dermal absorption; however, data were not
9-29
-------
available to quantltate the rates of these processes nor of any of the
pharmacoklnetlc processes. Initial distribution of the trlchlorobenzenes
and metabolites Is mainly to the liver, kidneys and adrenals, followed by
migration to adipose tissue or metabolism to polar compounds that are more
readily excreted. Metabolism appears to be Initially to arene oxides, and
then by different routes 1n different species, with different rates of ex-
cretion. Species differences are such that extrapolation of adverse effects
to humans probably requires the support of comparative metabolic data.
Human exposure to 1,2,4-tMchlorobenzene at 3-5 ppm causes eye and
respiratory Irritation (Rowe, 1975). The only other data on human exposure
are Individual case reports of aplastlc anemia of persons exposed occupa-
tional^ or domestically (Glrard et al., 1969).
The effects 1n mammals of acute exposure by various routes to trlchloro-
benzenes Include local Irritation, convulsions and death. Livers, kidneys,
adrenals, mucous membranes and brain ganglion cells appear to be target
organs with effects Including edema, necrosis, fatty Infiltration of livers,
Increased organ weights, porphyrln Induction and mlcrosomal enzyme Induction.
Quantitative data on the toxic effects of trlchlorobenzene following
subchronlc exposure by various routes were obtained 1n a variety of
species. In general, these studies Indicate that the Hver and kidney are
target organs. Inhalation of 1,2,4-tr1chlorobenzene at >74.2 mg/m3 (10
ppm) for 6 hours/day, 5 days/week for up to 26 weeks Induced hepatocyto-
megaly and hyaline degeneration 1n several species (Kodba et al., 1981;
Watanabe et al., 1978; Coate et al., 1977), although these effects may be to
some extent reversible. One study (Watanabe et al., 1978) Identified 22.3
mg/m3 (3 ppm) as a NOAEL 1n rats. Sasmore and Palmer (1981) reported that
some rats exposed by Inhalation to 1,3,5-tr1chlorobenzene at 7423 mg/m3
9-30
-------
(1000 ppm) for 13 weeks showed squamous metaplasia and focal hyperplasla of
the respiratory epithelium, which appeared to be reversible. Subchronlc
oral studies have also found that the trlchlorobenzenes Induce hepatic xeno-
blotlc metabolism {Carlson and Tardlff, 1976; Smith et a!., 1978) and por-
phyrla (Carlson, 1977b). Subchronlc dermal exposure resulted 1n mild to
moderate Irritation (Powers et a!., 1975; Rao et a!., 1982).
One chronic study, on the effects of trlchlorobenzene painted on the
skin of mice for 2 years, reported Increased mortality 1n females at the low
dose (30% solution 1n acetone) and 1n both sexes at the high dose (60% solu-
tion) (Yamamoto et al., 1957). While numbers of all tumor types appeared to
be Increased, no significant change was detected for any Individual tumor
type. Thus, the carcinogenic results of the only relevant study are
considered Inconclusive.
Results of two reports on mutagenlclty tests with Salmonella typh1mur1um
test strains were negative (Schoeny et al., 1979; Lawlor et al., 1979).
However, this test system 1s generally Insensitive to chlorinated compounds.
A multlgeneratlon study of the reproductive effects of oral exposure to trl-
chlorobenzene (Robinson et al., 1981) failed to show effects on reproduc-
tion. Teratogenldty studies after administration by the oral route 1n rats
(Black et al., 1983; KHchln and Ebron, 1983a) showed mild osteogenlc
changes 1n pups and significantly retarded embryonic development as measured
by growth parameters.
9-31
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10. TETRACHLORQBENZENES
Approximately 5 million pounds of the three tetraehlorobenzene Isomers
were produced annually 1n the United States 1n 1981 (Chlorobenzene Producers
Association, 1984). Recent Information Indicates that there 1s an Incident-
al annual "by-product" production of about 3 million pounds of the tetra-
chlorobenzenes (Chlorobenzene Producers Association, 1984). The 1,2,4,5-
Isomer 1s primarily used as an Intermediate 1n the synthesis of fungicides,
bacterlddes and herbicides (see Sections 4.1. and 4.2.) (U.S. EPA, 1977).
Tetrachlorobenzene Isomers have been detected 1n environmental samples as
well as 1n human tissues and breath, but no quantitative exposure assessment
has been completed (see Sections 4.3. and 4.4.). F1sh and other organisms
bloaccumulate the tetrachlorobenzenes, Indicating that human exposure from
the food chain 1s possible along with human atmospheric exposure (see
Section 4.4.).
10.1. PHARHACOKINETICS
No studies describing the absorption, distribution, metabolism or excre-
tion of 1,2,3,4-, 1,2,3,5- or 1,2,4,5-tetrachlorobenzene following Inhala-
tion exposure were located 1n the available literature. Several oral
studies describing the pharmacoklnetlcs of the three tetraehlorobenzene
Isomers in rats, rabbits and dogs are available and are discussed 1n detail
below.
10.1.1. Absorption. Jondorf et al. (1958) examined the absorption of
1,2,3,4-, 1,2,3,5- and 1,2,4,5-tetrachlorobenzene from the gastrointestinal
tract of female Chinchilla rabbits. Groups of three rabbits were given a
single dose of the tetraehlorobenzene Isomers by stomach tube at a dose
10-1
-------
level of 500 mg/kg as a 10% solution 1n arachls oil. Through 6 days post-
dosing, the percentages of the administered doses recovered 1n the feces as
the Intact compound were 5% for 1,2,3,4-tetrachlorobenzene, 14% for 1,2,3,5-
tetrachlorobenzene and 16% for 1,2,4,5-tetrachlorobenzene. Considering the
small amount of Isomers 1n the feces through 6 days postdoslng and that some
of this fecal content may have been due to biliary excretion, 1t can be
assumed that gastrointestinal absorption of the three tetrachlorobenzene
Isomers 1s a relatively efficient process 1n rabbits (Jondorf et al., 1958).
The percentages of the administered doses recovered unchanged 1n the "gut
contents" were 0.5, 1.4 and 6.2% for 1,2,3,4-, 1,2,3,5- and 1,2,4,5-tetra-
chlorobenzene, respectively, suggesting that the chlorine positions on the
molecule may Influence absorption.
10.1.2. Distribution. The tissue distribution patterns of 1,2,4,5-tetra-
chlorobenzene 1n beagle dogs (Braun et al., 1978) and of all three tetra-
chlorobenzene Isomers 1n Chinchilla rabbits (Jondorf et al., 1958) and rats
(Chu et al., 1983; Jacobs et al., 1977) have been described. None of these
Investigators speculated on comparisons between the animal species tested
and humans.
Braun et al. (1978) administered 5 mg/kg/day of 1,2,4,5-tetrachloroben-
zene 1n the diet to 2 male and 2 female beagle dogs for 2 years. The
resulting distribution of 1,2,4,5-tetrachlorobenzene was described 1n terms
of a two-compartment pharmacoklnetlc model, with clearance rate constants
(k ) of 6.64-»-0.82xlO~3 day"1 for plasma and 6.22+0.58x10~3 day"3-
e ~~ ~
for fat tissue. The half-lives for elimination from fat and plasma were 111
and 104 days, respectively. The authors concluded that steady-state was
approached at a faster rate 1n fat than 1n plasma. However, the steady-
10-2
-------
state profiles for both fat and plasma (Tables 10-1 and 10-2) appear to be
similar, and no statistically significant difference was reported. The
fat:plasma ratio (F/P) was -650 after 1 month of treatment, Indicating that
1,2,4,5-tetrachlorobenzene has a high affinity for fat. During the remain-
der of the study, F/P decreased steadily, reaching -280 by the end of the
study. Therefore, the fat was probably becoming saturated with each succes-
sive dose, and the 1,2,4,5-tetrachlorobenzene concentration 1n the plasma
Increased more rapidly over time than that 1n the fat. During the 20-month
observation period that followed treatment, F/P Increased rapidly, as the
available 1,2,4,5-tetrachlorobenzene 1n the plasma and other hypothetical
low affinity compartments was preferentially redistributed to the high
affinity fat compartment.
Jondorf et al. (1958) administered single dosages of 500 mg/kg each of
the three tetrachlorobenzene Isomers as a 10% solution 1n arachls oil by
stomach tube to groups of three Chinchilla rabbits. The animals were killed
6 days post-dosing, and the unchanged tetrachlorobenzene Isomers were
detected 1n the liver, brain, skin, depot fat, gut contents and other
unspecified parts of the body (cumulatively referred to as "rest of body").
The percentage of the administered dose measured as unchanged Isomer for
each of the above tissues 1s presented 1n Table 10-3.
Chu et al. (1983) administered l*C-labeled 1,2,3,4-, 1,2,3,5- and
1,2,4,5-tetrachlorobenzene as a single oral dose of 1 or 10 mg/kg to male
rats, and killed the treated animals 7 days postdoslng. At the higher dose
level, 1,2,4,5-tetrachlorobenzene was observed 1n all tissues examined,
Including fat (411 ppm), skin (33 ppm), kidney (23 ppm) and liver (22 ppm);
there was no Indication 1n the abstract whether these concentrations were
10-3
-------
TABLE 10-1
Percentage of 1,2,4,5-Tetrachlorobenzene Steady-State Reached
at Specific Times 1n Fat and Plasma of Dogs*
Percentage of Steady-State Profile
Time of Exposure
(days)
10
30
90
180
365
730
Fat
5.5
16
40
64
87
98
Plasma
4.8
14
35
58
83
97
*Source: Braun et al., 1978
2 Hale and 2 female beagle dogs were administered 5 mg/kg/day of 1,2,4,5-
tetrachlorobenzene 1n the diet.
10-4
-------
TABLE 10-2
Time Required to Reach Various Percentages of
1,2,4,5-Tetrachlorobenzene Steady-State 1n Fat and Plasma of Dogs*
Percentage of Steady State
99.9
98.0
90.0
50.0
Time
Fat
1220
691
407
122
(days)
Plasma
1418
803
473
142
*Source: Braun et al., 1978
2 Male and 2 female beagle dogs were administered 5 mg/kg/day of 1,2,4,5-
tetrachlorobenzene 1n the diet.
10-5
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TABLE 10-3
Unchanged Tetrachlorobenzene 1n RabbH Tissues
6 Days After Oral Dosing (500 mg/kg)*
Percentage
Tetrachlorobenzene
Isomer
1,2,3,4-
1,2,3,5-
1,2,4,5-
L1ver
0.1
<0.5
0.1
Brain Skin
0 2
<0.2 5
<0.1 10
Depot
Fat
5
11
25
of Dose
Gut
Contents
0.5
1.4
6.2
Rest of
Body
2.0
5.2
6.4
Total
10
23
48
*Source: Jondorf et al., 1958
10-6
-------
Intact compound or radioactivity. Similar tissue distribution patterns were
observed for animals given the higher doses of 1,2,3,4- or 1,2,3,5-tetra-
chlorobenzene, but tissue concentrations were much less; further detail
regarding target tissues and concentrations for these two Isomers were not
reported 1n the abstract. At the lower doses, a similar tissue distribution
pattern for all three Isomers was observed.
As reported 1n the summary of a German study, Jacobs et al. (1977)
continuously fed rats diets containing 1,2,4,5-tetrachlorobenzene (dose
level and duration not reported). Accumulation of 1,2,4,5-tetrachloroben-
zene and Its derivatives were greatest 1n adipose tissue. The maximum
concentrations 1n adipose tissue and blood were reached by 3 weeks after
Initiation of treatment, and steady state was attained 1n both adipose and
blood compartments by 5 weeks after Initiation of treatment.
Morlta et al. (1975d) analyzed adipose tissue samples of 15 residents of
the Tokyo metropolitan area for 1,2,4,5-tetrachlorobenzene. Residual tissue
levels of 1,2,4,5-tetrachlorobenzene ranged from 0.006-0.039 ng/g of fat,
with a mean residual tissue level of 0.019 yg/g of fat. The source and
route of exposure to 1,2,4,5-tetrachlorobenzene were not Identified.
10.1.3. Metabolism. KohH et al. (1976a) examined the metabolic fate of
the three tetrachlorobenzene Isomers In male rabbits following a single
1ntraper1toneal Injection of the compounds dissolved 1n vegetable oil at
dose levels of 60-75 mg/kg. The urine and feces of the treated animals were
collected for 10 days postdoslng and examined for major metabolites.
1,2,3,5-Tetrachlorobenzene was the most extensively metabolized Isomer,
yielding 2,3,4,5-, 2,3,5,6- and 2,3,4,6-tetrachlorophenol. 1,2,3,4-Tetra-
chlorobenzene was metabolized to 2,3,4,5- and 2,3,4,6-tetrachlorophenol,
10-7
-------
while 1,2,4,5-tetrachlorobenzene yielded the single metabolite, 2,3,5,6-
tetrachlorophenol. The authors proposed corresponding arene oxides as elec-
trophlllc Intermediate metabolites of all three tetrachlorobenzene Isomers,
with the ultimate tetrachlorophenol formation from 1,2,3,5- and 1,2,3,4-
tetrachlorobenzene Involving an NIH shift of a chlorine atom. The metabo-
lism of 1,2,4,5-tetrachlorobenzene to 2,3,5,6-tetrachlorophenol can be
achieved via the 2,3,5,6-tetrachlorobenzene oxide Intermediate without an
NIH shift of a chlorine atom. Further evidence of this metabolic pathway
was provided by Ar1yosh1 et al. (1974, 1975a,b}, who reported that all three
tetrachlorobenzene Isomers Increased the cytochrome P-450 enzyme activity 1n
the liver of rats, Indicating that oxldatlve metabolism with the formation
of the corresponding arene oxide Intermediate 1s a plausible pathway.
The metabolic fate of the tetrachlorobenzenes 1n rabbits was also Inves-
tigated by Jondorf et al. (1958). Single doses of 500 mg/kg tetrachloro-
benzene Isomers were given to groups of three rabbits by stomach Intubation
as a 10% solution 1n arachls oil. The metabolic products detected In the
urine through day 6 postdoslng, as summarized 1n Table 10-4, Included tetra-
chlorophenols and the glucuronlde, ethereal sulfate and mercapturlc add
conjugates. The authors suggested that the tetrachlorobenzenes were metabo-
lized via the competitive reactions Involving oxldatlve hydroxylatlon or
reductive dechlorlnatlon. In agreement with the results obtained by KohH
et al (1976a), Jondorf et al. (1958) also reported that 1,2,4,5-tetrachloro-
benzene was the least metabolized tetrachlorobenzene Isomer; 48% of the
administered dose of 1,2,4,5-tetrachlorobenzene was detected as the Intact
compound In the tissues of rabbits at 6 days after administration, as com-
pared to 10% for 1,2,3,4-tetrachlorobenzene and 23% for 1,2,3,5-tetrachloro-
10-8
-------
TABLE 10-4
Urinary Metabolites of Tetrachlorobenzene Isomers In Rabbits 6 Days After Oral Dosing (500 mg/kg)*
Percentage of Dose (Mean Values) Excreted as
Tetrachlorobenzene Tetrachlorophenol
Isomer Glucuronlde Ethereal Sulfate MercaptuMc Add
Free Total
1,2,3,4- 30 3 <1 8 43
1,2,3,5- 6 2 0 1.9 5
1,2,4,5- 4 1 0 1.3 2.2
*Source: Jondorf et al., 1958
-------
benzene. It was suggested by MoMta (1977) that the metabolism of 1,2,4,5-
tetrachlorobenzene via oxldatlve hydroxylatlon 1s partially Inhibited
because of steMc factors.
Chu et al. (1983) administered single oral doses of 1 or 10 mg/kg
i4C-labeled 1,2,4,5-, 1,2,3,5- and 1,2,3,4-tetrachlorobenzene each to male
rats, and killed the treated animals 7 days postdoslng. Urinary metabolites
of the tetrachlorobenzene Isomers detected Included tetrachlorophenols,
trlchlorophenols, dlhydroxylated tetrachlorobenzenes and trace amounts of
sulfur-containing metabolites; no distinction between Individual Isomers and
metabolites was made 1n the abstract.
The tetrachlorobenzenes have been reported as metabolites of Undane 1n
rats (Engst et al., 1976a), molds (Engst et al., 1979), hen pheasants,
wheat, lettuce and endives (KohH et al, 1976b,c; Saha and Burrage, 1976),
and of hexachlorobenzene 1n rats (Mehendale et al., 1975; Engst et al.,
1976a).
10.1.4. Excretion. Jondorf et al. (1958) administered single doses of 500
mg/kg each of the tetrachlorobenzene Isomers to groups of three rabbits by
stomach tube in a 10% solution 1n arachls oil. The tetrachlorobenzene
Isomers were excreted as phenols (primarily tetrachlorophenols) 1n the
urine, as Intact compound 1n the feces and breath and as other chloroben-
zenes 1n the expired air. Total excretion of the administered dose at 6
days postdoslng was 68% for both 1,2,3,4- and 1,2,3,5-tetrachlorobenzene,
and 83% for 1,2,4,5-tetrachlorobenzene. The excretion profiles for the
1somer1c tetrachlorobenzenes are summarized 1n Table 10-5, and the excretion
of the Intact compound 1n the expired air over 5 days postdoslng 1s summar-
ized 1n Table 10-6.
10-10
-------
TABlh 10-5
Summary of Excretion of the IsomeMc Tetrachlorobenzenes as Metabolites or as
Unchanged Compound 1n Rabbits Dosed Orally (500 mg/kg)*
Percentage of Dose Excreted as
Phenols 1n Urine Unchanged Tetrachlorobenzene 1n
_, Tetrachlorophenol
-------
TABLE 10-6
Excretion of Unchanged Tetrachlorobenzenes 1n the
Expired A1r of Rabbits After Oral Dosing (500 mg/kg)*
Tetrachlorobenzene
Isomer
1,2,3,4-
1,2,3,5-
1,2,4,5-
Percentage
Days after
113
1.9 2.2 1.6
2.1 2.1 1.2
1.2 0.2 0.2
of Dose
Dosing
4
0.2
2.9
0
1n Expired A1r
5_
0
2.6
0
Total
5.9
10.9
1,6
*Source: Jondorf et al., 1958
10-12
-------
Chu et al. (1983) administered single oral doses of 1 or 10 mg/kg each
of 14C-labeled 1,2,4,5-, 1,2,3,5- and 1,2,3,4-tetrachlorobenzene to male
rats, and killed the treated animals 7 days postdoslng. Animals receiving
the higher dose of 1,2,4,5-tetrachlorobenzene were observed to excrete 16.7%
of the administered dose 1n the urine and 4.8% 1n the feces. The percentage
of the administered dose excreted 1n the urine and feces of animals dosed
with 1,2,3,5- or 1,2,3,4-tetrachlorobenzene was greater than that for those
dosed with 1,2,4,5-tetrachlorobenzene; however, actual percentages were not
reported. Excretion of the lower doses of Isomers were similar to the per-
centage values observed with the higher doses, but quantitative results were
not presented 1n the abstract.
10.1.5. Summary. No studies describing the absorption, distribution,
metabolism or excretion of 1,2,3,4-, 1,2,3,5- or 1,2,4,5-tetrachlorobenzene
following Inhalation exposure were located 1n the available literature. The
pharmacoklnetics of the tetrachlorobenzene Isomers following oral admini-
stration 1s well characterized 1n rabbits, but not 1n other animal species.
The UpophlUc characteristics of the tetrachlorobenzene Isomers allowed
efficient transepHhellal absorption at the gastrointestinal and respiratory
surfaces. Once absorbed, the tetrachlorobenzene Isomers administered orally
to rabbits was rapidly accumulated 1n fat, metabolized primarily to tetra-
chlorophenols and their conjugates, partly as glucuronldes and ethereal
sulfates, or eliminated unchanged 1n the expired air or feces (Jondorf et
al., 1958).
No pharmacoklnetlc data were available for humans, except a report of
1,2,4,5-tetrachlorobenzene 1n adipose tissue (range of 0.006-0.039 mg/kg bw;
mean of 0.019 mg/kg bw) of 15 Tokyo residents (MorHa et al., 1975d).
10-13
-------
Although quantitative estimates of human exposure to the tetrachlorobenzene
Isomers via air, food or drinking water were not available, based on the
relatively limited Industrial use of the tetrachlorobenzene Isomers (U.S.
EPA, 1980b), human exposure may not be significant. The tetrachlorobenzene
Isomers are both 1_n vivo and _1n. vitro metabolites of the pesticides, Undane
and hexachlorobenzene (Mehendale et al., 1975; Engst et al., 1976a,b, 1979;
KohH et al, 1976b,c; Saha and Burrage, 1976); therefore, human exposure via
air, food and drinking water could occur from the environmental degradation
of these pesticides.
10.2. EFFECTS ON HUMANS
Only one ep1dem1olog1c study was available regarding the effects of the
tetrachlorobenzenes on humans. Klraly et al. (1979) examined peripheral
lymphocytes for chromosomal abnormalities 1n blood collected from Hungarian
workers engaged 1n the production of 1,2,4,5-tetrachlorobenzene. The
"normal control" group consisted of 49 nonfactory workers (ages, 26-52
years; average age, 38.2 years) who provided blood for chromosome examina-
tion at a genetic counseling clinic. The "factory employees control" group
consisted of 14 factory employees (ages, 28-47 years; average age, 35.4
years; duration of employment range of 10-30 years) not directly exposed to
the 1,2,4,5-tetrachlorobenzene manufacturing process, but with possible
Inadvertent exposure to other unspecified airborne pollutants. The "posi-
tive control" group contained 25 factory workers (ages, 31-59 years; average
age, 44.6 years) producing 1,2,4,5-tetrachlorobenzene; each had been
employed at that job for >6 months, working 8 hours/day, and wearing "Tucan-
type" face masks during work hours. Coded samples of peripheral lymphocytes
were cultured for 48 hours, and >50 metaphase cells were examined for each
sample. Factory air concentrations of 1,2,4,5-tetrachlorobenzene were not
determined.
10-14
-------
The group of workers exposed to 1,2,4,5-tetrachlorobenzene had a sig-
nificantly Increased (p<0.01) frequency of cells with <46 chromosomes when
compared with both the normal and factory employee control groups. Poly-
ploldy was observed 1n 2.94% (40 of 1360) of the mitoses examined from the
exposed group, compared with 0.59% (15 of 2523) in the normal control, and
2.50% (21 of 838) 1n the factory control group; statistical significance was
not Indicated. Inadvertent exposure to airborne pollutants may have
resulted 1n the relatively high percentage of polyploldy and chromosome
aberrations observed 1n the factory control group. The frequencies for
chromatld-type chromosome aberrations, labile chromosome-type aberrations
and stable chromosome-type aberrations for the three groups are listed 1n
Tables 10-7, 10-8 and 10-9, respectively. The authors concluded that
1,2,4,5-tetrachlorobenzene was mutagenlc (I.e., clastogenlc) to occupation-
ally exposed humans.
10.3. MAMMALIAN TOXICOLOGY
No animal studies on acute toxldty, subchronlc toxlclty, chronic toxic-
1ty, mutagenldty, carclnogenclty or reproductive and teratogenlc effects of
1,2,3,4-, 1,2,3,5- or 1,2,4,5-tetrachlorobenzene following Inhalation
exposure were located 1n the available literature. Several oral studies
describing some of the effects of the three tetrachlorobenzene Isomers 1n
animal species are available and are described below. A summary of sub-
chronic, chronic, reproductive and teratogenlc toxlclty studies on tetra-
chlorobenzenes can be found 1n Table 10-10.
10.3.1. Acute Toxlclty. The oral LD gQ for 1,2,4,5-tetrachlorobenzene
was reported to be 1035 mg/kg when given 1n sunflower oil and 2650 mg/kg
when given 1n 1.5% starch solution 1n mice and 1500 mg/kg when given 1n
10-15
-------
TABLE 10-7
Frequency of Chromat1d-type Chromosome
Aberrations 1n Peripheral Lymphocytes3
Parameter
No. of Mitoses Examined
(subjects)
Gap
Number
Percent
Isogapb
Number
Percent
Total (Gap + Isogap)
Percent
Break
Number
Percent
Isobreakb
Number
Percent
Total (Break + Isobreak)
Percent
Exchange
Number
Percent
Total Aberrations
Number
Percent
Normal
Control
2523 (49)
73
2.89
19
0.75
92
3.64
40
1.59
17
0.67
57
2.26
0
0
149
5.90
Factory 1
Control
838 (14)
46
5.48
2
0.23
48
5.71
26
3.10
18
2.14
44
5.24
0
0
92
10.97
,2,4,5-Tetrachlorobenzene
Exposed
1360 (25)
81
5.95
30
2.20
111
8.15
55
4.04
32
2.35
87
6.39
2
0.15
198
14.70C
aSource: Klraly et al., 1979
blsogap and Isobreak are aberrations Involving the same location on two
chromatlds
GStat1st1cally significant difference between exposed and each of the
control groups; test and p value not specified.
10-16
-------
TABLE 10-8
Frequency of Labile Chromosome-type Aberrations*
Parameter
No. of Mitoses Examined
(subjects)
Acentric Fragment
Number
Percent
Ring Chromosome
Number
Percent
D1centr1c Chromosome
Number
Percent
Total
Number
Percent
Normal
Control
2523 (49)
9
0.35
0
0
0
0
9
0.35
Factory
Control
838 (14)
8
0.95
0
0
0
0
8
0.95
1 ,2,4,5-Tetraehlorobenzene
Exposed
1360 (25)
19
1.40
2
0.15
2
0.15
23
1.69
*Source: Klraly et a!., 1979
10-17
-------
TABLE 10-9
Frequency of Stable Chromosome-type Aberrations3
Parameter
No. of Karyotypes Examined
Deletion
Number
Percent
Inversion
Number
Percent
Translocatlon
Number
Percent
Total
Number
Percent
Normal
Control
460
19
4.13
4
0.87
3
0.65
26
5.65
Factory
Control
144
10
6.94
1
0.69
2
1.38
13
9.02
1 ,2,4,5-Tetrachlorobenzene
Exposed
237
27
11.39
4
1.68
5
2.10
36
15.18&
aSource: Klraly et a!., 1979
^Statistically significant difference between exposed and normal controls
and factory controls (p<0.1, test not specified).
10-18
-------
TABLE 10-10
Summary of Toxlclty Studies on Tetrachlorobenzenes
Species
Rat
*
\
1
Rat
Rabbit
— '
o
i
io
Rat
!
!
j
Dog
Pregnant rats
i
! Pregnant rats
Pregnant rats
Route Dose
oral 0.5-500 mg/kg
of diet
1,2,4,5-TeCB
oral 0.001, 0.005,
0.05 mg/kg/day
1,2,4,5-TeCB
oral 0.001, 0.005,
0.05 mg/kg/day
1,2,4,5-TeCB
oral 75 mg/kg/day
1,2,4,5,-TeCB
oral 5 mg/kg/day
1,2,4,5-TeCB
oral 50, 100,
200 mg/kg/day
1,2,4,5-TeCB
oral 50, 100,
200 mg/kg/day
1,2,3,4-TeCB
oral 50, 100,
200 mg/kg/day
1,2,3,5-TeCB
Duration
28 or 90 days
8 months
8 months
2 months
2 years expo-
sure, 22 months
recovery
days 6-15 of
gestation
days 6-15 of
gestation
days 6-15 of
gestation
Effects
Increased liver and kidney weights and
hlstologlcal changes 1n liver and kidneys;
Increases 1n HFO activity, serum cholesterol
values
No effects observed 1n 0.001 mg/kg/day dose
group; 0.005 and 0.05 mg/kg/day doses caused
disruption In conditioned reflexes, Increases
1n liver weight coefficients and decrease 1n
serum SH groups
No effect observed 1n 0.001 mg/kg/day dose
group; 0.05 mg/kg dose caused disorder of
liver glycogen formation, altered serum SH
group levels, Increase In blood hemoglobin
and peripheral retlculocyte levels
Altered biochemical parameters Indicating
changes In hepatic and hematopoltlc homeo-
stasls
Mo controls used; elevated SAP and total
b111rub1n, returned to normal range 3 mo
after exposures ended
High-dose lethal to 9/10 of treated dams;
organ weight changes, elevated serum
cholesterol and liver metabolism enzymes,
no Indication of those changes were dose-
related
Induced maternal toxldty and Increased
lethality of pups at 200 mg/kg/day
Increased lethality 1n 200 mg/kg/day group
pups; one pup malformed and minor chondro-
genlc delay 1n other pups
Reference
VUleneuve et al.,
1983
Fomenko, 1965
Fomenko, 1965
Fomenko, 1965
Braun et al., 1978
Ruddlck et al., 1981
Ruddlck et al., 1981
Ruddlck et al., 1981
-------
TABLE 10-10 (cont.)
o
i
rvj
o
Species
Pregnant rats
Pregnant rats
Route Dose
oral 30, 100, 300,
1000 mg/kg/day
1,2,4,5-TeCB
oral 100, 300,
1000 mg/kg/day
1,2,3,4-TeCB
Duration
days 9-13 of
gestation ob-
served on day 14
days 9-13 of
gestation ob-
served on day 14
Effects
Only control and 1000 mg/kg/day group
examined for embryotoxIcHy and only
observed fewer Implantations than control,
slight hepatic centrolobular hypertrophy
In 1000 tng/kg/day group, hepatic enzymes
Induced at all doses.
Only control and 300 mg/kg/day group
examined for embryotoxlclty, significant
embryonic growth reduction was observed In
Reference
KHchln and Ebron,
1983b
Kltchln and Ebron,
1983c
the 300 mg/kg/day group, maternal lethality
In 300 (1/10 dams) and 1000 (7/19 dams)
mg/kg/day groups, minimal hepatocellular
hypertrophy 1n 300 mg/kg/day group, minimal
to moderate hepatocellular hypertrophy and
reduced body and liver weights 1n 1000
mg/kg/day group, hepatic enzymes Induced 1n
the 300 and 1000 mg/kg/day groups.
1,2,4,5-TeCB = 1,2,4,5-tetraehlorobenzene
1,2,3,4-TeCB » 1,2,3,4-tetraehlorobenzene
1,2,3,5-TeCB «= 1,2,3,5-tetrachlorobenzene
-------
apparently sunflower oil to rats and rabbits (Fomenko, 1965). VUleneuve et
al. (1983) reported an L05Q range of -1200-3000 mg/kg 1n rats for the
three tetrachlorobenzene Isomers with 1,2,3,4-tetrachlorobenzene > 1,2,3,5-
tetrachlorobenzene > 1,2,4,5-tetrachlorobenzene; further details regarding
doses and effects were not provided 1n the abstract.
R1m1ngton and Zlegler (1963) administered relatively large dietary doses
of 1,2,3,4-tetrachlorobenzene at a level of 660 mg/kg/day for 10 days or
1,2,4,5-tetrachlorobenzene at a level of 905 mg/kg/day for 5 days to rats.
1,2,3,4-Tetrachlorobenzene Induced weight loss, non-necrot1c Hver cell
degeneration and an Increase 1n porphyrln and hemoglobin metabolism, while
the only effect reported for 1,2,4,5-tetrachlorobenzene was non-necrot1c
liver cell degeneration.
No studies were available regarding the dermal toxlclty or sens1t1zat1on
reactions of the three tetrachlorobenzene Isomers.
10.3.2. Subchronlc Toxldty. As reported 1n an abstract, Vllleneuve et
al. (1983) administered dietary concentrations of 1,2,3,4-, 1,2,3,5- and
1,2,4,5-tetrachlorobenzene ranging from 0.5-500 ppm to both sexes of rats
for 28 or 90 days. Administration of 1,2,4,5-tetrachlorobenzene resulted 1n
Increased kidney and liver weights. Increased mixed function oxldase activi-
ties, Increased serum cholesterol values, moderate to marked hlstologlcal
Uver changes 1n both sexes, and marked hlstologlcal kidney changes 1n
males. The authors concluded that 1,2,4,5-tetrachlorobenzene was the most
toxic tetrachlorobenzene Isomer when administered 1n the diet to rats, and
that males appeared to be more susceptible than females. The authors did
not specify the number or strain of animals used, If the effects observed
were dose-related or only seen at the higher dose(s), the severity of
effects, or the type of hlstologlcal changes observed.
10-21
-------
Fomenko (1965) examined the subchronlc toxlclty of 1,2,4,5-tetrachloro-
benzene In rats and rabbits. Both species of animals were given the com-
pound dally by gavage In vegetable oil at dose levels of 0, 0.001, 0.005 or
0.05 mg/kg for 8 months. No treatment-related effects were observed In
either rats or rabbits at the 0.001 mg/kg dose level. Doses of 0.005 or
0.05 mg/kg to rats Induced a disruption of conditioned reflexes, Increased
liver weight coefficients, and decreased blood serum SH groups, while
Increased organ ascorbic acid was seen only 1n those rats given 0.05 mg/kg;
the author did not Indicate the statistical significance of these effects.
Rabbits given 0.005 mg/kg had a transient disorder of liver glycogen forma-
tion and a statistically significant (p=0.05) change 1n blood serum SH
groups during the last month of treatment. At the 0.05 mg/kg dose level,
rabbits were observed to have a disorder of liver glycogen formation during
the sixth month of treatment, Increased serum blood SH groups 1n the fifth
month that was followed by a decrease, a statistically significant (p=0.05)
Increase 1n the blood hemoglobin level during the third month of treatment,
an Increased level of peripheral retlculocytes at the end of the last month
of treatment, and an Increased retention of an Intravenous galactose load by
6 months of treatment.
In a 2-month oral study, rats were given dally doses of 0 or 75 mg/kg
1,2,4,5-tetrachlorobenzene 1n vegetable oil by gavage {Fomenko, 1965). No
treatment-related hlstologlc changes were observed, but several biochemical
parameters were affected, Indicating changes 1n hepatic and hematopoletlc
homeostasls. The blood chollnesterase activity Increased significantly
(p=0.01), and the prothrombln Index dropped by -1/3 of the control value (p
not reported). In addition, the number of peripheral retlculocytes
decreased significantly (p=0.02) but then Increased, the serum potassium
10-22
-------
levels were reduced (p not reported), and the number of peripheral large-
diameter erythrocytes was Increased (p not reported). The Incidence of
erythemla was significantly Increased (p=0.01). At cessation of treatment,
statistically significant (p=0.01) observed effects Included a decrease 1n
serum SH groups, adrenal hypertrophy and decreased adrenal ascorbic acid.
10.3.3. Chronic Tox1c1ty. Braun et al. (1978) fed two beagle dogs of each
sex diets containing doses of 5 mg/kg/day 1,2,4,5-tetrachlorobenzene for 2
years, and then observed them for a 20-month recovery period. The primary
goal of the study was to determine the uptake and elimination kinetics for
plasma and fat; therefore, no concurrent control animals were used. Histor-
ical control data, however, suggested that the elevations of serum alkaline
phosphatase and total bH1rub1n after 24 months of administration were
related to treatment. The elevated clinical chemistry values returned to
the normal range of values for the historical controls at 3 months Into the
20-month recovery period. Gross and hlstopathologlcal examinations of
tissues performed after the recovery period did not reveal any treatment-
related morphological changes 1n the animals. This study could not be used
to substantiate either a no-observed-effect level (NOEL) or a lowest-
observed-effect level (LOEL), because concurrent controls were not used, the
number of treated animals used was small, and only one dose level was tested.
10.3.4. Mutagenldty. Klraly et al. (1979) reviewed the chromosomal
effects of 1,2,4,5-tetrachlorobenzene 1n Hungarian workers and concluded
that 1,2,4,5-tetrachlorobenzene 1s mutagenlc 1n occupatlonally-exposed
humans. A more accurate conclusion from this data 1s that 1,2,4,5-tetra-
chlorobenzene 1s clastogenlc 1n the exposed humans. . This paper was
discussed 1n detail 1n Section 10.2.
10-23
-------
Paradl and Lovenyak (1981) reported that 1,2,4,5-tetrachlorobenezene did
not Induce an Increased frequency of sex-linked recessive lethals 1n Droso-
phlla melanogaster exposed by larval feeding at a dose less than the LC5Q
(actual dose not reported). Only an abstract of the original paper was
available and there was no Information regarding the number of chromosomes
assayed at each dose nor the doses used. This Information 1s essential
before any conclusions can be made as to whether or not 1,2,4,5-tetrachloro-
benzene can Induce sex-linked recessive lethal mutations 1n Drosophlla.
1,2,3,5- and 1,2,4,5-tetrachlorobenzene were tested for mutagenldty by
plate Incorporation with Salmonella typh1mur1um strains TA98, TA100, TA1535,
TA1537 and TA1538 at five unspecified dose levels (Lawlor et al., 1979).
Both Isomers gave negative results 1n the reverse mutation assay either 1n
the presence or absence of an S-9 metabolic activation system from rats
pretreated with Aroclor 1254. Because these results were reported 1n an
abstract, Insufficient experimental detail was provided to permit a critical
evaluation of the data and negative results 1n the Salmonella assay for
highly chlorinated compounds are not unexpected (Rlnkus and Legator, 1980).
10.3.5. Carc1nogen1c1ty. Pertinent data regarding the carc1nogen1c1ty of
1,2,3,4-, 1,2,3,5- and 1,2,4,5-tetrachlorobenzene were not located 1n the
available literature.
10.3.6. Reproductive and Teratogenic Effects. As reported 1n an abstract,
Ruddlck et al. (1981) administered 1,2,3,4-, 1,2,3,5- and 1,2,4,5-tetrachlo-
robenzene via gavage (vehicle not reported) at dose levels of 0, 50, 100 or
200 mg/kg to pregnant rats (10/dose level) on days 6 through 15 of gesta-
tion. 1,2,4,5-Tetrachlorobenzene was the most toxic Isomer, Inducing
lethality 1n 9 of 10 treated dams at the 200 mg/kg level. A dose-related
accumulation of compound residue was seen 1n dams and offspring with all
10-24
-------
three Isomers, but was greatest 1n those animals given 1,2,4,5-tetrachToro-
benzene. Other toxldty effects observed 1n dams treated with 1,2,4,5-
tetrachlorobenzene Included organ weight changes and significantly elevated
serum cholesterol, liver am1nopyr1ne-N-detnethylase and hepatic aniline
hydroxylase levels; 1t was unclear from the abstract whether these changes
were dose-related or occurred at a single dose level. 1,2,3,4-Tetrachloro-
benzene also Induced maternal toxldty, manifested 1n a significantly
lowered platelet count at the 200 mg/kg level. Fetotoxldty, as Indicated
by Increased lethality of pups, was observed at the 200 mg/kg level of
1,2,3,4- and 1,2,3,5-tetrachlorobenzene. One malformed pup and minor chon-
drogenlc delay were seen among the offspring of dams given 1,2,3,5-tetra-
chlorobenzene.
Kltchln and Ebron (1983b) conducted a maternal hepatic toxldty and
embryotoxlclty study 1n which they administered 1,2,4,5-tetrachlorobenzene
(>98% pure) suspended 1n 1.5% gum tragacanth (2 ml/kg) orally to pregnant
Sprague-Oawley (CO strain) rats on days 9-13 of gestation and the dams were
sacrificed on day 14 of pregnancy. The dosing groups were 0 (1.5% gum
tragacanth only), 30, 100, 300 and 1000 mg/kg/day 1,2,4,5-tetrachloroben-
zene. There were no maternal deaths 1n any 1,2,4,5-tetrachlorobenzene
treatment group. However, there was a significantly decreased body weight
gain In the 1000 mg/kg/day treatment group, Maternal liver weights, liver
to body weight ratios and hepatic mlcrosomal protein content were not
significantly affected by the 1,2,4,5-tetrachlorobenzene administration.
Normal liver histology was observed 1n the control, 100 and 300 mg/kg/day
dose groups. The 1000 mg/kg/day dose group was observed with 3/9 dams
showing slight hepatic centrolobular hypertrophy. The 1,2,4,5-tetrachloro-
benzene was found to Induce the cytochrome P-450 content at the 1000
10-25
-------
mg/kg/day dose level, amlnopyrene N-demethylase activity at the 300 and 1000
nig/kg/day dose levels, and ethoxyresoruf1n 0-deethylase activity at all dose
levels. The uteri from only the 0 and 1000 mg/kg/day groups were examined
for 1,2,4,5-tetrachlorobenzene-lndueed embryonic effects. No statistically
significant differences 1n resorptlon, embryonic deaths, abnormalities,
protein content, somite number, crown-to~rump length, head length or yolk
sac diameter were observed. The only effect seen after examining the 14 day
uteri were a slightly lower number of Implantations 1n the treated as
compared with the control group. It can be concluded from this study that
only the dams receiving 1000 mg/kg/day were adversely affected by the
1,2,4,5-tetrachlorobenzene treatment as Indicated by the parameters that
were studied.
KHchln and Ebron (1983c) conducted a maternal hepatic toxldty and
embryotoxldty study where they administered 1,2,3,4-tetrachlorobenzene
(>98% pure) suspended 1n 1.5% gum tragacanth (2 ma/kg) orally to pregnant
Sprague-Dawley (CO strain) rats on days 9-13 of gestation and the dams were
then sacrificed on day 14 of pregnancy. The dosing groups were 0 (1.5X gum
tragacanth only), 100, 300 and 1000 mg/kg/day 1,2,3,4-tetrachlorobenzene.
Phenobarbltal and B-naphthoflavone were also given to other pregnant rats by
1.p. Injection and used as positive hepatic controls. Maternal lethality
occurred only 1n the 300 mg/kg/day (1/10 dams) and 1000 mg/kg/day (7/19
dams) treated groups. A significant decrease 1n body weight and liver
weight was also observed 1n the 1000 mg/kg/day dose group. No effect on
maternal hepatic mlcrosomal protein content was observed 1n any dose group.
1,2,3,4-Tetrachlorobenzene was found to significantly Induce the levels of
hepatic cytochrome C-reductase and glutathlone S-transferase at the 1000
mg/kg/day dose level, and of hepatic cytochrome P-450, amlnopyrene
10-26
-------
N-demethylase and UDP-glucuronyUransferase at the 300 and 1000 mg/kg/day
dose levels. No hepatic lesions were observed In the 100 mg/kg/day dose
group. Minimal hepatocellular hypertrophy was seen In 2/9 dams In the 300
mg/kg/day dose group, and minimal to moderate hepatocellular hypertrophy was
seen In 9/13 dams 1n the 1000 mg/kg/day dose group. The uteri from the 0
and 300 mg/kg/day dose groups were examined for 1,2,3,4-tetrachlorobenzene-
Induced embryonic effects. No statistically significant differences 1n
embryonic resorptlons, lethality or abnormalities were seen. Embryonic
growth was found to be adversely affected by 1,2,3,4-tetrachlorobenzene
treatment. Head length and crown-to-rump length (embryonic growth param-
eters) were significantly reduced by maternal exposure. A significant
decrease 1n the day 14 yolk sac diameter was also observed. It 1s not known
1f these adverse effects seen at day 14 of gestation are reversible after
removal from 1,2,3,4-tetrachlorobenzene exposure.
10.4. INTERACTIONS
Tetrachlorobenzene 1s capable of Inducing the NADPH-dependent cytochrome
P-450 metabolizing enzymes, which are nonspecific for natural and xenoblotlc
substrates (Ar1yosh1 et al., 1974, 1975a,b). The substrate may either be
detoxified by such metabolism or become more hazardous (toxlfled) 1f con-
verted to a reactive Intermediate capable of binding to critical Intracellu-
lar macromolecules. In Itself, P-450 Induction 1s not a disadvantage, but
1t may become one when substrates are activated during metabolism (Neal,
1980). Thus, exposure to tetrachlorobenzene may enhance the toxlclty of a
compound that normally would be Innocuous. No studies were available,
however, to demonstrate the Interaction of tetrachlorobenzene with other
compounds.
10-27
-------
10.5. SUMMARY
No animal studies on pharmacok1net1cs» acute toxiclty, subchronlc
tox1c1ty» chronic toxiclty, mutagenldty, carc1nogen1c1ty or reproductive
and teratogenlc effects of 1,2,3,4-, 1,2,3,5- or 1,2,4,5-tetrachlorobenzene
following Inhalation exposure were located 1n the available literature.
Tetrachlorobenzenes are I1p1d-soluble compounds that bloaccumulate 1n
the fat of aquatic and terrestrial organisms. Although the Isomers were
preferentially distributed to adipose tissue, they did not cross the blood-
brain barrier of rabbits. Some Tokyo residents were found to have 1,2,4,5-
tetrachlorobenzene (mean of 0.019 mg/kg bw) 1n their adipose tissue.
The metabolism of the tetrachlorobenzene Isomers apparently follows
aromatic hydroxylatlon to tetrachlorophenols with an arene oxide Intermedi-
ate. Rabbits and rats treated with the tetrachlorobenzene Isomers excreted
unchanged compound 1n expired air and feces; the urine contained tetrachlo-
rophenols, trlchlorophenols, dlhydroxylated tetrachlorobenzenes, and the
glucuronldes and ethereal sulfates of those metabolites.
The tetrachlorobenzenes have been reported as metabolites of Undane 1n
rats, molds, hen pheasants, wheat, lettuce and endives, and of hexachloro-
benzene 1n rats.
Chromosome aberrations were observed 1n the lymphocytes of Hungarian
workers producing 1,2,4,5-tetrachlorobenzene; no airborne concentrations or
exposures were determined.
The only mammalian toxicology data available for tetrachlorobenzenes are
the result of oral exposures. The oral ID™ for 1,2,4,5-tetrachloroben-
zene was reported as 1035 mg/kg 1n mice and 1500 mg/kg 1n rats and rabbits
when administered 1n sunflower oil and 2650 mg/kg In mice when administered
1n 1.5% starch solution. Subchronlc oral exposure of rats and rabbits to
10-28
-------
1,2,4,5-tetrachlorobenzene resulted 1n statistically significant effects on
biochemical parameters, Including retlculocytosls, Increased blood cholln-
esterase activity, erythremla and an Indication that glycogen formation was
Impeded; at higher doses of 1,2,4,5-tetrachlorobenzene, rats also had
Increased kidney and liver weights, and renal and hepatic hlstologlc changes.
Reversible effects on serum alkaline phosphatase and total b1Hrub1n
were reported 1n dogs given 5 mg/kg/day 1,2,4,5-tetrachlorobenzene In the
diet for 2 years.
1,2,4,5-Tetrachlorobenzene was not mutagenlc 1n the sex-linked recessive
lethal assay with Drosophlla melanogaster. However, because only an
abstract of the Drosophlla study was available, experimental details were
too sparse to permit a critical evaluation of this negative result. Both
1,2,3,5- and 1,2,4,5-tetrachlorobenzene were negative 1n the reverse muta-
tion assay with Salmonella typhlmurlum strains TA98, TA1QQ, TA1535, TA1S37
and TA1538. These results were reported 1n an abstract with Insufficient
experimental detail. Also, a negative result 1n the Salmonella assay with
chlorinated compounds Is not unexpected.
No Information was available regarding the carc1nogen1c1ty of any of the
three tetrachlorobenzene Isomers In either animals or humans.
The tetrachlorobenzene Isomers Induced appreciable maternal toxlclty,
mild fetotoxldty and negligible teratogenlclty In rats following oral
administration.
10-29
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11. PENTACHLOROBENZENE
The annual production of pentachlorobenzene In the United States was
estimated to be 1-10 million pounds 1n 1977 (U.S. EPA, 1981a). Recent
Information Indicates that the production and Import of pentachlorobenzene
Into the U.S. 1s zero (U.S. EPA, 1983). The compound has been used as a
pesticide, a chemical Intermediate (Clement Associates, 1979; Ware and West,
1977) and as a flame retardant (Kw1atkowsk1 et al., 1976). Pentachloroben-
zene has been detected 1n surface waters (Barkley et al., 1980; Oliver and
Nlchol, 1982; Elder et al., 1981), drinking water contaminated by a toxic
waste site (Barkley et al., 1981), aquatic sediments (Elder et al., 1981),
fish and shellfish (Oliver and Nlchol, 1982; Ten Berge and Hlllebrand, 1974)
and 1n some edibles (U.S. EPA, 1980a).
11.1. PHARMACOKINETICS
11.1.1. Absorption. Pentachlorobenzene has UpophlUc characteristics and
1s therefore likely to be capable of crossing biological membranes. Several
studies were available on the absorption of pentachlorobenzene after oral
administration. One study discussed absorption after dermal application;
however, no studies were available on absorption via the Inhalation route.
No studies were encountered on the distribution of pentachlorobenzene after
Inhalation or dermal exposure.
Parke and Williams (1960) studied the absorption and metabolic fate of
pentachlorobenzene 1n rabbits. Three to four days after a 0.5 g/kg dose of
pentachlorobenzene suspended 1n an aqueous solution was administered by
gavage, 5% was recovered 1n the feces and 45% was found 1n the gut con-
tents. Biliary excretion was not measured; therefore, some of the penta-
chlorobenzene found 1n the gut and feces may have resulted from a portion of
the dose that was absorbed being excreted unchanged 1n the bile. Rozman et
11-1
-------
al. (1979) found that absorption 1n two male and two female rhesus monkeys
was very efficient. Four days following a single dose of 0.5 mg/kg penta-
chlorobenzene by gavage, at least 9554 was reported as being absorbed. Blood
and tissue levels of pentachlorobenzene and/or Its metabolites were found to
be similar to those measured for hexachlorobenzene, Indicating the Involve-
ment of the lymphatic system 1n the absorption process (latropoulos et al.,
1975). Other studies concerning the toxlclty and metabolism of pentachloro-
benzene (Under et al., 1980; Engst et al., 1976; Vllleneuve and Khera,
1975) also demonstrated that absorption occurs through the gastrointestinal
tract, but did not provide quantitative data.
In the only available study Involving dermal absorption of pentachloro-
benzene, Under et al. (1980) applied a single dose of 2500 mg/kg penta-
chlorobenzene dissolved 1n xylene to the shaved backs and shoulder areas of
two rats. No clinical signs of toxlclty were observed 1n males or females,
suggesting that percutaneous absorption of pentachlorobenzene was poor.
11.1.2. Distribution. Vllleneuve and Khera (1975) studied the distribu-
tion of pentachlorobenzene 1n dams and fetuses after dally administration by
gavage of pentachlorobenzene prepared 1n corn oil at levels of 40, 100 and
200 mg/kg to pregnant rats on days 6-15 of gestation. On day 22, the dams
were killed, fetuses removed and tissues analyzed by gas-Hqu1d chromato-
graphy for organohalogen residues. Recovery of pentachlorobenzene was >80%
for all tissues. In the tissues of the maternal animals, fat had the great-
est accumulation of pentachlorobenzene, followed by the Hver, brain, heart,
kidneys and spleen. In the fetuses, the levels detected 1n the brain were
equal to those measured 1n the whole fetus, while the levels 1n the liver
were double the whole fetus concentration. Tables 11-1 and 11-2 show these
distribution data. Both the maternal tissues and the whole fetuses appeared
to accumulate pentachlorobenzene 1n a dose-related manner.
11-2
-------
-------
TABLE 11-2
Distribution of Pentachlorobenzene Residues 1n the Tissues
of Fetal Rats after Oral Administration to Dams3
Whole Fetusb
Dose Level
(mg/kg)
50
100
200
{ mg/kg)
2.44+0.38
5.27+0.60
16.9 ±2.8
(total vg)
9. 65+1. 3
21.2 ±2.1
55.1 ±6.7
L1verc
(mg/kg)
4.37+0.69
10.4 ±1.31
40.4 ±6.02
Bra1nc
(mg/kg)
3.08±0.55
5.31±0,60
20.5 +2.64
aSource: Vllleneuve and Khera, 1975
^Represents the mean of two fetuses each from 15 Utters ± standard error
of the mean.
Represents the mean of five fetuses each from a different Utter ± stan-
dard error of the mean.
11-4
-------
Under et al. (1980) also reported that pentachlorobenzene accumulated
In the adipose tissue. Based on food consumption data provided by the
authors, groups of 10 male rats were fed 6-16 or 50-134 mg/kg/day (125 or
1QQQ ppm) for 100 days, and similar groups of females were fed 6-16, 16-31,
27-63 or 55-134 mg/kg/day (125, 250, 500 or 1000 ppm) for 180 days. The
results Indicated that pentachlorobenzene accumulated 1n adipose tissue
-1.5-2.2 times the dietary concentration, and the accumulation was dose-
dependent. Residues 1n males and females were similar, but could not be
compared directly because of the longer exposure period of the females and
the complicating factors of pregnancy and lactation. Suckling pups whose
mothers were fed >250 ppm pentachlorobenzene developed tremors, and at 1000
ppm, most died before weaning. Though no clinical signs of tremors were
observed In the parents, the authors stated that this result was presumptive
evidence for excretion of a toxic agent via the milk. Because pentachloro-
benzene accumulates In the fetus (Vllleneuve and Khera, 1975), prenatal
exposure of the pups may also have contributed to the observed effects.
Rozman et al. (1979) studied the distribution of pentachlorobenzene and
Us metabolites 1n four rhesus monkeys. Tissues of monkeys given a single
dose of 14C-labeled pentachlorobenzene (0.5 mg/kg) by gavage were analyzed
after 40 days. Quantitative determination of pentachlorobenzene and Its
metabolites was performed by gas chromatography. The highest concentrations
were found 1n the fat and bone marrow, followed by the thymus, lymph nodes
and adrenal cortex. Table 11-3 summarizes the distribution data for the 20
tissues examined.
Parke and Williams (1960) studied the distribution of pentachlorobenzene
1n rabbits and found that the compound was readily Isolated from the feces
and gut contents 3-4 days following administration by gavage of 0.5 g/kg.
11-5
-------
TABLE 11-3
Distribution of Pentachlorobenzene and/or Metabolites on the
40th Day 1n the Rhesus Monkey Following a Single Oral Dose
of 0.5 mg/kg Body Height*
Organ
Fatb
Bone marrow
Lymph nodesb
Thymus
Adrenal cortex
Adrenal medulla
Skin
Kidneys
Liver
Lungs
Spleen
Heart
B1le
Stomach
Duodenum
Cecum
Large Intestine
Small Intestine
Brain
Cerebellum
Male
(mg/kg)
1.86
1.10
0.35
0.50
0.31
0.18
0.26
0.09
0.19
0.06
0.04
0.07
0.09
0.06
0.11
0.24
0.31
0.17
0.05
0.05
Female
(mg/kg)
2.68
2.35
0.79
0.61
0.56
0.07
0.26
0.10
0.17
0.06
0.04
0.12
0.09
0.06
0.06
0.18
0.33
0.07
0.06
0.06
aSource: Rozman et al., 1979
bAverage value from five different parts of the body
11-6
-------
Subcutaneous Injections of 0,5 g/kg (10% w/v solutions 1n arachls oil)
resulted 1n concentrations of 47% 1n the pelt (mostly at the site of Injec-
tion), 22% In the fat, a total of 2% 1n the gut and feces, and 10% 1n the
rest of the body. Table 11-4 summarizes the distribution data for penta-
chlorobenzene for this study.
11.1.3. Metabolism. The metabolism of pentachlorobenzene has been studied
in male Wlstar rats by Engst et al. (1976) following administration by
gavage of 8 mg/kg pentachlorobenzene dissolved in 1 ma of filtered sun-
flower oil. The major metabolites detected 1n the urine were Identified as
2,3,4,5-tetrachlorophenol and pentachlorophenol. Pentachlorobenzene,
2,3,4,6-tetrachlorophenol and/or 2,3,5,6-tetrachlorophenol were present 1n
the free form. Trlchlorophenol (Isomer not specified), 2,4,6-trlchloro-
phenol and 1,2,3,4-tetrachlorobenzene were present In small concentrations.
Quantities of the metabolites obtained were not reported for this study.
Koss and Koransky (1977) reported pentachlorophenol, 2,3,4,5-tetra-
chlorophenol, tetrachlorohydroquionone and a hydroxylated chlorothlo com-
pound as metabolites of pentachlorobenzene 1n the urine and feces of three
female rats collected for 4 days after administering a single 1ntraper1to-
neal dose of 403 yM/kg (sic). Pentachlorophenol and other hydrophHlc
metabolites accounted for 9% of the eliminated dose.
Rozman et al. (1979) measured and Identified the metabolites of penta-
chlorobenzene 1n the rhesus monkey. Table 11-5 summarizes the metabolic
breakdown during 40 days following a single oral dose by gavage of 0.5 mg/kg
14C-labeled pentachlorobenzene. The major metabolites Identified 1n the
urine were pentachlorophenol, 2,3,4,5-tetrachlorophenol and 2,3,5,6-tetra-
chlorophenol. No significant differences were observed In the metabolism
patterns of male and female monkeys.
11-7
-------
TABLE 11-4
Distribution of Pentachlorobenzene In Chinchilla Doe Rabbits
Expressed as a Percentage of Administered Dose3
CO
Time
Dose/Route After
(g/kg) Dosing
(days)
0.5 oral 3
0.5 oral 4
0.5 s.c. 10
Urine
Tr1- or Penta- Other
chlorophenol Phenols
0.2 1
0.2 1
0.7 1
Feces Gut Pelt Depot
Contents Fat
5.0 45.0 1.0 15.0
5.0 31.0 5.0 9.0
1.5 0.5 47.0& 22. Ob
Rest of
Body
6.0
5.5
10.0
Expired Air
Unchanged Other Chloro-
hydrocarbons
0 9.0
0 21.0
0 <2.0
Total
Accounted
%
82
78
85
^Source: Parke and Williams. I960
^Located mainly at site of Injection
s.c. = subcutaneous
-------
TABLE 11-5
Percentage of Pentachlorobenzene and Its Metabolites Identified In Urine, Feces and
Various Organs of Rhesus Monkeys Dosed 0.5 rng/kg Body Weight Pentachlorobenzene*
Liver
Bile
Feces
Blood
lOdnpu
Urine
Pentachlorobenzene
99, OX
nonpolar compound(s)
99. OX
45. BX
51 3S
ND
1 ,2,3,4-Tetrachlorobenzene
1.0X
nonpolar compound(s)
l.OX
ND
ND
ND
Pentachlorophenol
ND
ND
ND
54. 2X
I
j ______
SB. IX
2,3,4,5-Tetrachlorophenol
ND
ND
ND
ND
32. 2X
2,3,5,6-Tetrachlorophenol
ND
ND
ND
ND
'
9.7X
•Source: Rozraan et al., 1979
ND = Not detected
-------
Similar results were obtained by KohH et al. (1976) 1n male rabbits.
Following 1ntraper1toneal Injection of 300 mg pentachlorobenzene dissolved
1n 10-15 mi vegetable oil, urinary metabolites were Identified as 2,3,4,5-
tetrachlorophenol and pentachlorophenol. Both were detected at yields of 1%
of the administered dose during the 10 days following administration of the
dose. Parke and Williams (1959) reported that <0.2% of the dose recovered
1n rabbit urine was pentachlorophenol.
The metabolic pathway of pentachlorobenzene was thought to Involve oxi-
dation and formation of an arene oxide Intermediate by hepatic metabolic
enzymes (KohH et al., 1976). Subchronlc feeding of 0.05% pentachloroben-
zene 1n the diets of female adult Wlstar rats for 60 days Induced hepatic
cytochrome P-450 content and enhanced the Q-dealkylatlon of 7-ethoxycoumar1n
(Goerz et al., 1978), suggesting the Involvement of the hepatic cytochrome
P-450 system 1n metabolism. However, Rozman et al. (1979) reported that
more phenolic Intermediates were present 1n the blood, kidney and urine of
monkeys than 1n the Hver, bile and feces 40 days after a single dose of
pentachlorobenzene. The evidence suggested that a metabolizing system other
than hepatic cytochrome P-450 was Involved In the hydroxylatlon of chlori-
nated benzenes. The authors proposed that two different hydroxylatlon path-
ways could be Involved, one Involving the oxidation of the pentachloro-
benzene to pentachlorophenol, and the other Involving nucleophlUc displace-
ment reactions of pentachlorobenzene to produce tetrachlorophenols.
Koss and Koransky (1977) suggested that a major consideration 1n the
toxldty of pentachlorobenzene Is Its metabolic transformation to penta-
chlorophenol. As previously stated, pentachlorophenol has been Identified
as a metabolite 1n the urine and excreta (Engst, 1976; Rozman et al., 1979;
11-10
-------
Kohll et al., 1976; Parke and Williams, 1960). Rozraan et al. (1979)
estimated that the elimination half-life of pentachlorobenzene 1n the rhesus
monkey was 2-3 months, and after 40 days pentachlorophenol accounted for
58,1% of the metabolites Identified In the urine.
11.1.4. Excretion. The excretion of pentachlorobenzene and Its metabo-
lites was described 1n rhesus monkeys following administration by gavage of
a single oral dose of 0.5 mg/kg (Rozman et al., 1979). Approximately 12% of
the administered dose was excreted In the urine after 40 days (see Table
11-5). Over the same period, ~24% of the dose was excreted via the feces,
of which 99% was unmetabollzed. Table 11-6 displays the cumulative urinary
and fecal excretion of pentachlorobenzene and Its metabolites. This study
Indicated that the metabolites of pentachlorobenzene were excreted primarily
via the urine, while the unabsorbed or unmetabollzed compound was excreted
via the feces. These results also Indicated that pentachlorobenzene was
eliminated very slowly, with an estimated excretion half-life 1n primates of
2-3 months.
Koss and Koransky (1977) Identified 3% of the administered dose of
pentachlorobenzene 1n Its unchanged form, pentachlorophenol, 2,3,4,5-tetra-
chlorophenol and a hydroxylated chlorothlo compound 1n the feces of rats 4
days after intraperltoneal administration of 403 pM/kg (sic) pentachloro-
benzene. Parke and Williams (1960) also Isolated 5% of the administered
dose of pentachlorobenzene after 4 days from the feces of rabbits given 0.5
g/kg pentachlorobenzene orally.
Under et al. (1980) fed pentachlorobenzene 1n the diet (250-1000 ppm)
to female Sherman rats with suckling pups and observed that the pups
developed tremors and most died before weaning 1n the 1000 ppm group. This
work provides presumptive evidence for excretion of pentachlorobenzene via
the milk.
11-11
-------
TABLE 11-6
Cumulative Urinary and Fecal Excretion of Pentachlorobenzene and
Metabolites During 40 Days Following a Single Oral Dose of
0.5 mg/kg 1n Male and Female Rhesus Monkeysat&
4
Hales
urine 1.9
feces 6.3
Females
urine 2.4
feces 4.4
Days After
10 20
4.8 8.6
11.5 19.3
4.3 7.8
8.3 16.4
Exposure
30
11.3
23.6
10.0
19.8
%
40 Total
Recovered
13.2 40.2
27.0
H*4 33.2
21.8
aSource: Rozman et a!., 1979
''Expressed 1n percent of the total administered dose
11-12
-------
11.1,5. Summary. Although studies of the absorption of pentachlorobenzene
Indicated that absorption does occur through the gastrointestinal tract, the
extent of absorption has not been determined. A study 1n rabbits Indicated
that up to 50% of a dose was absorbed within 3-4 days. Oral administration
to monkeys Indicated 95% absorption within 4 days. Absorption resulting
from Inhalation has not been studied, and absorption from dermal exposure
was found to be rather poor 1n rats. Once absorbed, pentachlorobenzene 1s
widely distributed to many tissues, with the highest levels appearing 1n fat
and bone marrow. A study 1n rats demonstrated that transport across
placental membranes occurred readily and that accumulation of pentachloro-
benzene 1n the fetus 1s highest 1n the Hver. No studies were encountered
that described the distribution of pentachlorobenzene after Inhalation or
dermal exposure.
The metabolism of pentachlorobenzene 1s not fully understood, but some
studies suggested that metabolic activity other than the hepatic cytochrome
P-450, xenoblotlc metabolizing system may be Involved. Metabolism appeared
to be primarily via oxidation to two major metabolites, pentachlorophenol
and 2,3,4,5-tetrachlorophenol, which were excreted 1n the urine. Metabolism
and excretion occurred at a slow rate; an estimated elimination half-life
for a single dose 1n primates was 2-3 months.
11.2. EFFECTS ON HUMANS
No epldemlologlc studies or case studies of effects 1n humans resulting
from exposure to pentachlorobenzene were available for review.
11.3. MAMMALIAN TOXICOLOGY
11.3.1. Acute Toxldty. Llnder et al. {1980J Investigated the acute and
subchronlc toxldty of solutions containing 99.1% pure pentachlorobenzene 1n
adult and weanling Sherman strain rats and adult Swiss-Webster mice. Wean-
ling rats (27-35 days of age; 10 animals/dosage level) and adult animals
11-13
-------
(90-120 days of age; 10 animals/dosage level) were administered by gavage a
single dose of 5.0-15.0 ma/kg pentachlorobenzene dissolved In peanut oil.
The oral ID™ values ranged from 1080-1125 mg/kg for adult rats, and
1175-1370 mg/kg for adult mice; for weanling rats the LD5Q was reported as
940 mg/kg (Table 11-7).
The characteristic toxic signs observed Included a decrease In activity,
hypersens1t1v1ty to touch, and tremors. The tremors started 1n mice ~24
hours after dosing and ~48 hours after dosing In rats. Death usually
occurred 1n rats 5-12 days after dosing; 1n mice the survival time was less,
with death usually occurring 2-4 days after the lethal dose was adminis-
tered. The authors reported many cases of rats with reddish stains around
the eyes, nose and mouth; no explanation of this phenomenon was given.
Ar1yosh1 et al. (1975) Investigated the effects of various chlorinated
benzenes, Including pentachlorobenzene, on the mlcrosomal drug metabolizing
enzymes, 5-am1nolevul1n1c add synthetase, mlcrosomal proteins and cyto-
chrome P-450 content. Groups of 2-6 female Wlstar rats were orally adminis-
tered 250 mg/kg pentachlorobenzene suspended 1n a 2% tragacanth gum solu-
tion, once a day for 3 days. The compound Increased the liver content of
cytochrome P-450 and Increased the activities of aniline hydroxylase and
amlnopyrlne demethylase. Significant Increases were also observed for
mlcrosomal protein and 6-am1nolevul1n1c add synthetase. Glycogen content
decreased markedly, and trlglycerlde content Increased 1n pentachloroben-
zene-treated rats.
In the only available study Involving acute dermal toxldty of penta-
chlorobenzene, Under et al. (1980) applied a single dose of 2500 mg/kg
pentachlorobenzene dissolved In xylene to the shaved backs and shoulder
areas of two rats. No clinical signs of toxldty were observed In either
male or female adult rats.
11-14
-------
TABLE 11-7
Acute Oral Toxlclty of Pentachlorobenzene*
Species/
Sex
Rat/M
Rat/F
Rat/F
Mouse/N
Mouse/F
Age
adult
adult
weanling
adult
adult
LD50
(rag/kg)
1125
1080
940
1175
1370
95% Confidence
Limits
(mg/kg)
1015-1247
952-1226
864-1023
1035-1334
1263-1487
Dosage
Range
Tested
{mg/kg)
750-1350
750-1500
600-1200
750-1500
1050-1500
Dose
Volume
(ml/kg)
7.5
7.5
5.0
15.0
15.0
*Source: Llnder et a!., 1980
Weanling animals were 27-35 days old. Adult animals were 90-120 days old.
Ten animals per each dosage group were given pentachlorobenzene dissolved 1n
peanut oil.
11-15
-------
11.3.2. Subchronlc Tox1c1ty. No studies of tox1c1ty resulting from
subchronlc Inhalation exposure to pentachlorobenzene were located 1n the
available literature. A summary of subchronlc, reproductive and teratogenlc
toxldty studies on pentachlorobenzene can be found 1n Table 11-8.
Under et al. (1980) studied the subchronlc toxldty of pentachloroben-
zene 1n rats as part of an Investigation of the compound's toxic effects on
reproduction. Groups of 10 female weanling rats were fed diets containing
0, 125, 250, 500 and 1000 ppm of pentachlorobenzene for ~180 days; while
groups of 10 male rats received 0» 125 and 1000 ppm for 100 days. Based on
food consumption data provided by the authors, 1t was estimated that the
female groups consumed an average of 11, 23, 46 and 99 mg/kg/day, respec-
tively (actual reported ranges of 7-16, 16-31, 27-63 and 55-134 mg/kg/day).
The male groups consumed -11 and 97 mg/kg/day, respectively (reported ranges
of 7-16 and 50-134 mg/kg/day). None of the animals died or exhibited clini-
cal signs of toxldty throughout the study. Food consumption and body
weight gain for the dosed groups were similar to the control groups. In
hematologlc parameters, erythrocyte count and hematocrlt were slightly lower
than the control group (p<0.05) for the 1000 ppm males, and hemoglobin was
reduced and leukocyte count Increased 1n both 1000 ppm groups (p<0.05).
Examination of the liver and viscera under ultraviolet light did not
reveal the presence of porphyrlns 1n males or females. Only tissues of the
female rats were analyzed quantitatively for porphyrlns. Total liver
porphyrlns were slightly higher 1n female rats fed 1000 ppm compared with
the control group (0.79 pg/g compared with 0.64 iig/g), but the differ-
ence was not judged to be a porphyrogenlc response and was of doubtful
consequence.
11-16
-------
TABLE 11-8
Summary of Subchronlc, Reproductive and Teratogenlc Toxlclty Studies on Pentachlorobenzene
Species
Rat (female)
Rat (male)
Rat
(offspring)
Mice
Rat
Route
oral
(diet)
oral
(diet)
oral
(diet)
oral
oral
Dose
125, 250, 500
or 1000 mg/kg
1n diet
125 or 1000
mg/kg In diet
125, 250, 500
or 1000 mg/kg
In mothers diet
50 or 100
mg/kg/gavage
50, 100 or 200
mg/kg/gavage
Duration
180 days
100 days
gestation and
during suckling
days 6-15 of
gestation
days 6-15 of
gestation
Effects Reference
Changes In hematologlc parameters In high- Under et al., 1980
dose group; Increase In Hver weights,
hepatic hypertrophy and vacuollzatlon 1n
500 and 1000 mg/kg groups; Increased kid-
ney weight In high-dose group
High-dose group Induced changes In hemato- Under et al., 1980
logic parameters; hepatic and renal
histology and Increase In Hver, kidney
and adrenal weights
Offspring treated with >25Q mg/kg/d1et were Under et al., 1980
adversely affected (reduced survival, body
weights and Increased liver weights, hepato-
cellular enlargement)
Increase 1n liver weights of dams; no Courtney et al., 1979
adverse effects on total development or
survival
No observed toxldty 1n adult rats; 1n- Khera and Vllleneuve,
creased total deaths at all doses, but not 1975
In dose-related manner; extra ribs In ex-
posed fetuses and sternal defects In 200
rag/kg group
-------
Tissues of 21-day-old weanlings appeared normal although relative
weights (organ/body weight ratio) of the livers were Increased 1n pups of
mothers fed >250 ppm for 180 days. The obvious change was hepatocellular
enlargement 1n all pups from the groups fed 500 and 1000 ppm.
At necropsy of the adult rats, no pathological changes were observed 1n
tissues of males fed pentachlorobenzene for 100 days or females fed penta-
chlorobenzene for 180 days. Weights of livers relative to body weight
Increased 35-4554 In the animals fed 500 or 1000 ppm. Relative weights of
the kidneys of both sexes and the adrenals of males Increased 1n the 1000
ppm groups. Microscopically, hepatic cell enlargement (hypertrophy) and
vacuollzatlon were observed 1n the female rats of the 500 and 1000 ppm
groups. Similar changes were apparent 1n the males fed 1000 ppm. In high-
dose groups, the kidneys of males showed hyaline droplet formation, atrophlc
tubules and lymphocytlc Infiltration. Results of this study Indicated that
the toxlclty of orally administered pentachlorobenzene was directed toward
the Hver and kidneys.
The ability of pentachlorobenzene to Induce porphyrla 1n Wlstar rats has
been Investigated by Goerz et al. (1978). Adult female rats were fed a diet
containing 0.0554 (~25.Q mg/kg/day or 500 ppm) pentachlorobenzene for 60
days. This treatment Increased the hepatic cytochrome P-450 content
(1.06i0.30 and 1.20+0.30 nMol/mg mlcrosomal protein for 10 and 60-day
exposures, respectively, for the controls compared with 2.25*1.10 and
2.06^0.65 nMol/mg mlcrosomal protein, for 10 and 60-day exposures for the
pentachlorobenzene-treated rats), but did not Increase the excretion of
porphyrlns 1n the urine.
11.3.3. Chronic Toxldty. No studies of toxldty resulting from chronic
exposure of pentachlorobenzene were located 1n the available literature.
11-18
-------
11.3.4. Hutagen1c1ty. The only Information available on the mutagenlcHy
of pentachlorobenzene was a study presented 1n abstract form on a plate
Incorporation assay for reverse mutation 1n h1st1d1ne-dependent strains of
Salmonella typhlmurlum (Lawlor et al., 1979). Five strains of Salmonella
typhlmurlum (TA98, TA100, TA1535, TA1537 and TA1538) were tested at five
unspecified concentrations of pentachlorobenzene 1n the presence and absence
of rat liver mlcrosomes Induced by Aroclor 1254. No detectable levels of
mutagenlc activity were found 1n the Salmonella tester strains (Lawlor et
al., 1979). Because these results were reported 1n an abstract, experi-
mental details were too sparse to permit a critical evaluation of this nega-
tive result. Also, this result 1s not unexpected because the Salmonella
test system 1s generally Insensitive to highly chlorinated compounds (Rlnkus
and Legator, 1980).
11.3.5. Carc1nogen1c1ty. The Ambient Water Quality Criteria Document for
Chlorinated Benzenes (U.S. EPA, 1980b) cited a study by Preussman (1975)
that was reported as alluding to the carc1nogen1c1ty of pentachlorobenzene.
The German text 1s now being translated and reviewed by the Carcinogen
Assessment Group of the U.S. EPA.
11.3.6. Reproductive and Teratogenlc Tox1c1ty. The reproductive toxlclty
of pentachlorobenzene was demonstrated In three studies. Of these studies,
Under et al. (1980) and Khera and Vllleneuve (1975) provided sufficient
data to estimate a NOEL and a LOAEL, respectively.
Llnder et al. (1980) tested 99.1% pure pentachlorobenzene for Us toxic
effects on reproduction In rats. No other chlorinated compounds were
detected by GC-EC analysis of the sample. Dietary concentrations of 0
(control), 125, 250, 500 and 1000 ppm (7.4-16, 16-31, 27-63 and 55-134
mg/kg/day) were fed by gavage to groups of ten 4- to 5-week-old weanling
female Sherman strain rats, while similar groups of males received diets
11-19
-------
containing 0 (control), 125 or 1000 ppm (6-16 or 50-134 mg/kg/day). Both
males and females were fed treated diets for 67 days before mating with
untreated males or females. Pregnant females continued to receive treated
diets until their Utters were weaned, for a total exposure of 180 days;
males were dosed for a total of 100 days before being sacrificed.
Litters sired by treated males showed no treatment-related effects.
Although clinical signs were not observed In the parents, litters from
treated females (>250 ppm) were adversely affected. Pup survival and body
weight at weaning were reduced In the two highest dose groups, and the off-
spring of the 250, 500 and 1000 ppm groups showed statistically significant
(p<0.05) Increases 1n Hver-to-body weight ratios. Survival decreased
dramatically from 88.6 to 28.0% during days 4-21 for pups whose mothers were
fed concentrations of 500 and 1000 ppm, respectively. Table 11-9 summarizes
the reproductive effects 1n Utters of treated females.
H1stolog1c examination of the livers of weanling rats revealed hepato-
cellular enlargement 1n all pups examined from the 500 and 1000 ppm groups,
and 1n 2 of 9 male pups from the 250 ppm group. The hepatotoxlc effects
were not seen In the offspring of the dams exposed to dietary concentrations
of 125 ppm. These data Indicated that pentachlorobenzene was transferred to
the offspring during gestation and/or lactation and had a toxic effect on
the pups 1n the 250, 500 and 1000 ppm groups. Therefore, this study sug-
gested a NOEL of 125 ppm 1n the diet for no toxic effects on the reproduc-
tion of rats.
In a study by Khera and VUleneuve (1975), pregnant Wlstar rats
(17-19/group) were administered pentachlorobenzene by gavage on days 6-15 of
gestation. The doses 1n mg/kg with percentage concentrations 1n corn oil
(1n parentheses) were 50 (0.5), 100 (1.0) or 200 (2.0). Uterine and viscera
11-20
-------
TABLE 11-9
Reproductive Effects 1n LHters of Female Rats Fed Diets Containing Pentaehlorobenzene3
Pentachlorobenzene 1n Diet (ppm)
Parameter
Dosage range (mg/kg/day)
Litters born
Pups per Utter (mean)
Litters weaned
Pup survival (%)
Days 0-4
Days 4-21
Pup body weight at Hale
wean1ngb Female
Liver/body weight Male
rat1oc (g/100 g) Female
0
NA
8
10.4
8
100
100
45 (5)
45 (8)
3.9 (0.1)
4.0 (0.1)
125
6-16
6
12.0
6
98.6
91.7
44 (3)
41 (3)
3.9 (0.1)
3.9 (0.1)
250
16-31
9
11.9
9
98.1
95.4
41 (4)
40 (3)
4.3 (0.1)d
4.2 (0.1)
500
27-63
9
13.2
9
98.3
88.6
40 (4)
38 (4)
5.1 (0.1)d
5.3 (0.1)d
1000
50-134
8
10.8
4
94.2
28.0
31 (4)
37 (4)
6.5 (0.1)d
6.5 (0.2)d
aSource: Llnder et a!., 1980
bValues are Utter means 1n grams (+_ standard deviation)
cValues are group means (i standard error of the mean)
^Significantly different from control; p=0.05 (statistical analysis performed on liver weights only)
NA = Not applicable
-------
contents were removed following sacrifice of the dams on day 22 of gesta-
tion. No overt signs of toxldty were observed 1n the adult rats; however,
the treatment Increased fetal death rate at all of the doses tested, but not
1n a dose-related manner (Table 11-10). This study demonstrated a lethal
effect of jm. utero exposure to pentachlorobenzene at doses to the dams as
low as 50 rag/kg/day, therefore Identifying a LOAEL for this study,
Khera and VUleneuve (1975) also reported that sternal defects were
observed 1n the fetuses of Wlstar rat mothers treated with 200 mg/kg/day.
In addition, all three doses Increased the Incidence of both un1- and
bilateral extra ribs (Table 11-11). The latter effect (Increase 1n extra
ribs), although not a gross malformation, occurred at an Incidence 5-9 times
greater than the controls, Indicating a potential for pentachlorobenzene to
alter fetal skeletal development. In addition, quantitative chemical
analysis of fetuses for pentachlorobenzene residues showed a dose-related
accumulation of the unchanged compound 1n the whole fetus, brain and liver
(Table 11-12) (VUleneuve and Khera, 1975). These results suggested that
the parent compound may have been responsible for the observed teratogenlc
and reproductive effects, but did not preclude metabolites as potential
causes of the observed effects.
In a study of possible reproductive and teratogenlc effects, Courtney et
al. (1977) reported that no reproductive toxldty occurred 1n Utters of
pregnant CD-I mice treated by gavage with 50 or 100 mg/kg of >91% pure
pentachlorobenzene 1n 0.1 mi corn oil on days 6-15 of gestation. There
were no teratogenlc effects observed 1n the 10 or 9 litters whose mothers
had been treated with 50 or 100 mg/kg, respectively, when compared with the
6 control Utters. There was, however, a significant Increase (p=0.01) 1n
the liver weight and the Hver-to-body weight ratio of the treated mice
11-22
-------
TABLE 11-10
Toxic Effects of Pentaehlorobenzene on Reproduction
In Rats Dosed on Each of Gestation Days 6-15a
Pentachlorobenzene
Number
Parameter
of rats pregnant at term
Live fetuses per Utter
Fetal
Fetal
death (54)b
mean body weight (g)
0
19
12.1
1.3
4.8
50
18
12.5
4.2
4.9
Dose (mg/kg/day)
100
19
11.5
3.1
4.8
200
17
10.7
3.2
4.4
aSource: Khera and Vllleneuve, 1975
^Percent fetal death = (no. dead plus dedduomas) x 100/total no. of Implants
11-23
-------
TABLE 11-11
Skeletal and Soft-Tissue Abnormalities Observed 1n Rat Litters of
Dams Treated with Pentachlorobenzene on Each of Gestation Days 6-15*
Pentachlorobenzene Dose (mg/kq/day)
Parameter 0 50 100 200
Skeletal Defects
No. of fetuses examined 127 129 . 122 100
Extra ribs: unilateral 2 18 10 17
bilateral 2 10 11 46
Fused ribs NA NA NA 2
Wavy ribs 52 NA NA
Sternal defects 5 4 NA 31
Exencephaly NA NA NA NA
Soft-Tissue Anomalies
No. of fetuses examined for
visceral defects 67 69 67 52
Runts 1 2 NA 2
Cleft palate NA 1 NA NA
Cardiac defects NA NA NA NA
Other Defects NA NA NA 2
*Source: Khera and VUleneuve, 1975
NA = No abnormality observed
11-24
-------
TABLE 11-12
Fetal Wlstar Rat Residues of Pentaehlorobenzenea»b
Maternal
Intubated
Dose Level
(rag/kg)
50
100
200
Whole Fetus
L1verd Brain0
(ppm) (Total ng) (ppm) (ppm)
2.44 + 0.38 9. 65 ±1.3 4.37 +_ 0.69 3.08 + 0.55
5.27+0.60 21.2 ±2.1 10.4 +1.31 5.31+0.60
16.9 +2.8 55.1 +6.7 40.4 +6.02 . 20.5 +2.64
aSource: Adapted from Vllleneuve and Khera, 1975
^Pregnant rats were exposed to pentachlorobenzene during days 6-15 of
gestation and the fetuses were removed and analyzed on day 22.
Represents the mean of two fetuses from 15 Utters +_ s.e.m.
^Represents the mean of five fetuses each from a different Utter +_ s.e.m.
11-25
-------
compared with the control mice. Pentachlorobenzene had no adverse effect on
fetal development or survival. One fetus 1n the 50 mg/kg dose group
displayed a cleft palate, but the occurrence was within the normal Incidence
for this strain of mice.
11.4. INTERACTIONS
Ar1yosh1 et al. (1975) and Goerz et al. (1978) demonstrated the ability
of pentachlorobenzene to Increase the activity of NAOPH-cytochrome P-450
dependent enzyme systems 1n rats. Induction of the cytochrome P-450
monoxygenase-catalyzed metabolism could result 1n an Increase or decrease 1n
the toxldty of the compound. Therefore, exposure to pentachlorobenzene
could result 1n the blotransformatlon and toxldty of drugs and other
chemicals. However, no studies were available to support this.
11.5. SUMMARY
Pentachlorobenzene 1s absorbed from the gastrointestinal tract; studies
Indicated that 50-95% of an administered dose 1s absorbed within 4 days.
One dermal study that Indicated absorption through the skin suggested that
pentachlorobenzene was poorly dermally absorbed. No studies were available
that measured absorption through the lungs.
Distribution 1s to many tissues, primarily the fat and bone marrow.
Transfer across placenta! membranes and excretion Into the milk probably
occur.
Metabolism 1s believed to be by oxidation to phenolic compounds, espe-
cially pentachlorophenol, that are excreted 1n the urine. Excretion appears
to occur slowly; an estimated half-life 1n primates was 2-3 months.
No data were available on the effects of exposure to pentachlorobenzene
1n humans, and no chronic or carclnogenlcHy studies were available for
review.
11-26
-------
Oral LDcg values were determined for adult rats (1080-1125 mg/kg) and
mice (1175-1370 mg/kg), and for weanling rats (940 mg/kg). No clinical
signs of toxiclty were observed 1n adult rats following dermal application
of 2500 mg/kg pentachlorobenzene. Also, 1t was demonstrated that penta-
chlorobenzene caused an Increase 1n the liver content of cytochrome P-450,
mlcrosomal drug metabolizing enzymes and mlcrosomal proteins.
A subchronlc feeding study Indicated that the primary toxic effects are
on the liver and kidneys, although slight changes 1n some hematologlc
parameters (e.g., decreased erythrocyte count, hemoglobin and hematocMt;
Increased leukocyte count) occurred 1n the high-dose groups. H1stolog1c
examination Identified pathologic changes 1n the livers of the female rats
fed 500 and 1000 ppm for 180 days and 1n the 1000 ppm male rats treated for
100 days. These data were sufficient to Identify a subchronlc LOAEL of 500
ppm (-27-63 mg/kg/day) and a NOEL of 250 ppm (-16-31 mg/kg/day).
No mutagenlc activity was detected 1n five strains of Salmonella typh1-
mur1 urn when tested at five unspecified concentrations of pentachlorobenzene
1n the presence and absence of rat liver mlcrosomes Induced by Aroclor 1254.
These results were reported 1n an abstract with few experimental details.
Also, a negative result for pentachlorobenzene 1s not unexpected, because
the Salmonella assay 1s generally Insensitive to chlorinated compounds.
Studies also have shown that pentachlorobenzene 1s capable of causing
reproductive and developmental effects. Female rats fed diets containing
pentachlorobenzene during mating and gestation produced Utters with reduced
pup survival and body weights at weaning, and Increased I1ver-to-body weight
ratios. No adverse effects were observed 1n the offspring of the dams
exposed to 125 ppm (6-16 mg/kg/day).
11-27
-------
Single oral doses of pentachlorobenzene given dally to pregnant rats
during gestation Increased the Incidence of fetal death at all doses tested,
Identifying a LOAEL of 50 mg/kg/day. Sternal defects and an Increase 1n the
Incidence of extra ribs also were observed at doses of 200 mg/kg/day and 50,
100 and 200 mg/kg/day, respectively.
In a study of possible reproductive and teratogenlc effects, doses of 50
and 100 mg/kg/day of pentachlorobenzene administered by gavage to pregnant
mice had no adverse effect on fetal development or survival.
11-28
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12. HEXACHLOROBENZENE
Hexachlorobenzene 1s not manufactured as a commercial product 1n the
United States, but an estimated 2-5 million pounds were produced each year
during the synthesis of several chlorinated chemicals (Mumma and Lawless,
1975). Hexachlorobenzene has been found as a trace contaminant 1n the
herbicides/fungicides Pentachloronltrophenol, Dacthal and Daconll, Hexa-
chlorobenzene Is also an Ingredient 1n a fungicide of which ~200,000 pounds
are Imported each year (IARC, 1979). Hexachlorobenzene 1s resistant to bio-
degradation, accumulates In the biological environment and has been detected
in ambient air, drinking and surface water, sediments, cropland and food
(see Section 4.3.). Hexachlorobenzene residues also have been found 1n
samples of human blood, fat and breast milk. The greatest degree of human
exposure 1s most likely to occur 1n the workplace and near manufacturing and
disposal sites, although the general population 1s likely to be exposed
through Inhalation of polluted air and the 1ngest1on of contaminated food
and water.
12.1. PHARMACOKINETICS
12.1.1. Absorption. Absorption of hexachlorobenzene from the gut has been
studied 1n detail; however, no Information has been found 1n the available
literature on hexachlorobenzene absorption through the lungs or skin.
Absorption of hexachlorobenzene from the Intestinal tract appears to depend
on the solvent vehicle used during test material administration. Thus, when
hexachlorobenzene 1s administered 1n olive oil, -80% of the dose 1s
absorbed; when 1t Is administered 1n an aqueous solution, 1n 1% methyl
cellulose, or 1n a solid crystalline form, relatively little (<20%) 1s
absorbed. Intestinal absorption of hexachlorobenzene occurs primarily
through lymphatic channels (latropoulos et al.» 1975), with only a minor
portion being absorbed Into the portal circulation.
12-1
-------
Ingebrlgtsen et al. (1981) Investigated the absorption of [14C]hexa-
chlorobenzene (10 mg 1n peanut oil) administered to male, blle-duct-cannu-
lated Wlstar rats by gastric catheter. Four days after dosing, 24.8% of the
administered 14C had been recovered 1n the feces, Indicating that at least
15% of the administered hexachlorobenzene was absorbed.
Albro and Thomas (1974) studied the absorption of hexachlorobenzene 1n a
squalane/cotton seed oil vehicle by male rats following administration of a
single dose by stomach Intubation. The results Indicated that at doses of
12 and 30 mg/kg, -82 and 72%, respectively, were absorbed within 96 hours.
Koss and Koransky (1975) compared the absorption rates of [14C]hexa-
chlorobenzene 1n female Wlstar rats following oral administration of olive
oil solutions or suspensions 1n 6% gum arable 1n water (4, 20, 50.5, 60 and
180 mg/kg). Approximately 80% of the dose was absorbed from the olive oil
solutions; however, only 6% was absorbed from the aqueous suspension.
Similarly, Zablk and Schemmel (1980) found that, when hexachlorobenzene
(32 mg/kg/day) was administered 1n the diet, high-fat (45.3% w/w) diets
resulted 1n greater accumulation of hexachlorobenzene 1n the tissues and
less hexachlorobenzene excreted 1n the feces than did high-carbohydrate
diets (67.7% w/w). The female rats received 32 mg/kg body weight hexa-
chlorobenzene/day for 6, 12 or 18 days. Although this study did not Include
a control group receiving a balanced diet, the data suggest that high fat
diets Increase the absorption of hexachlorobenzene.
Sundlof et al. (1982) administered seven consecutive dally oral doses of
10 or 100 mg crystalline hexachlorobenzene/kg body weight to male laboratory
beagles. The results from the 100 mg/kg group Indicated that hexachloro-
benzene can continue to be absorbed from the Intestines for up to 1 week
following the cessation of dosing.
12-2
-------
Bleavlns et al. (1982) fed female European ferrets (Hustela putorlus
furo) a single dose of 57.6 v»g hexachlorobenzene (14C-labeled) 1n 7,5 g
of standard mink diet (22% fat) and calculated that 98.5% of the hexachloro-
benzene dose was absorbed by the ferrets. They made this calculation based
on predicted hexachlorobenzene excretion as extrapolated from this study,
and owing to a food passage time 1n the female ferret of Just over 3 hours.
12.1.2. Distribution. Following Intestinal absorption, hexachlorobenzene,
which 1s UpophlUc, distributes to tissues that are rich 1n I1p1d content.
The adipose tissue accumulates the greatest concentrations of hexachloro-
benzene In all species studied, although bone marrow and skin, which contain
large amounts of I1p1ds, also accumulate hexachlorobenzene. The adrenal
cortex accumulates hexachlorobenzene at concentrations approaching those of
fat. Other tissues (e.g., liver, kidneys, lungs, heart, spleen and blood)
generally contain lower amounts of hexachlorobenzene. Intravenous Injection
of hexachlorobenzene results 1n a tissue distribution similar to that
following oral administration. Hexachlorobenzene 1s transported via the
placenta and 1s distributed 1n fetal tissue.
Mehendale et al. (1975) studied the disposition of 14C-hexachloro-
benzene by adult male rats following a single oral dose of 5 mg/kg.
14C-Hexachlorobenzene was mixed with arachls oil and administered by
stomach Intubation at a dose of ~5 mg/kg. The animals were sacrificed 7
days later and the tissues and organs radloassayed. Forty-three percent of
the total radioactivity administered was present 1n fat tissue 7 days after
14C-hexachlorobenzene administration. In addition, muscle and skin tis-
sues each contained -9% of the radioactivity, whereas the other 12 tissues
analyzed contained -5% combined (Table 12-1).
12-3
-------
TABLE 12-1
Storage and Excretion of i4C-HCB Administered Orally
1n Arachls 011 In Rats3
Organ or Tissue
Fatb
Muscle0
Skln^
Liver
Small Intestine
Bone6
Kidneys
Large Intestine
Stomach
Blood
Lungs
Testes
Heart
Brain
Spleen
Total 1n tissues
Excretion
Feces
Urine
Gut contents
Total recovery
Percent of Total
Radioactivity
Administered
42.81 + 6.14
9.41 + 1.17
8.64 + 1.21
3.01 + 0.23
2.43 * 0.47
1.04 + 0.09
0.76 + 0.11
0.43 + 0.08
0.36 + 0.04
0.24 + 0.04
0.24 + 0.04
0.21 + 0.04
0.18 + 0.03
0.17 + 0.03
0.04 £ 0.002
70.09 £ 5.48
16.02 + 2.31f
0.85 + 0.13f
2.48 £ 0.45
89.44 £ 10.57
aSource: Hehendale et al., 1975
bBased on 9% body weight as fat
cBased on 50% body weight as muscle
BBased on 16% body weight as skin
eBased on 10% body weight as bone
^Cumulative total for 7 days
Adult male rats were given 5 mg/kg of hexachlorobenzene.
HCB - Hexachlorobenzene
12-4
-------
When 14C-hexachlorobenzene was suspended 1n 1% methyl cellulose and a
single oral dose containing 150 yg of hexachlorobenzene was administered
to Sprague-Dawley rats, the absorption of 14C-hexachlorobenzene by the
walls of the stomach and duodenum 1 hour later was relatively low: ~1.0 and
0.6 ppm were found 1n the stomach and duodenum, respectively (latropoulos et
al., 1975). Increased radioactivity was found 1n the jejunum and Heum as
well as the lymph nodes and adipose tissues 3 hours after administration
(Table 12-2). Although the radioactivity also Increased 1n the Hver and
kidneys, this Increase was relatively low compared to that found 1n the
lymph nodes and adipose tissue. Moreover, the radioactivity In the Hver
and kidneys decreased within a 2-day period, whereas the radioactivity 1n
the lymph nodes and fat remained relatively constant. These results Indi-
cate that the portal venous transport of hexachlorobenzene to the liver
appears to be a minor pathway, whereas the major part of the Ingested hexa-
chlorobenzene 1s absorbed by the lymphatic system 1n the duodenum and
jejuno-lleum and deposited 1n the fat, bypassing the systemic circulation
and the excretory organs.
Knauf and Hobson (1979) Investigated the tissue distribution of hexa-
chlorobenzene In six female rhesus monkeys following the gastric administra-
tion of dally doses of hexachlorobenzene [0 (one monkey), 8 (one monkey), 32
(one monkey), 64 (one monkey), or 128 (two monkeys) mg/kg/day] 1n 1% methyl
cellulose for a period of 60 days. The highest concentrations of hexa-
chlorobenzene were located 1n tissues with high llpld content. Tissue
levels correlated more with body fat content than with dose, with the monkey
that had the least adipose tissue producing the highest nonfat tissue and
serum values (Table 12-3).
12-5
-------
TABLE 12-2
Tissue Concentration (ppm) of i4C-Hexaehlorobenzenea and Its Metabolites In Sprague-Dawley Rats'1
I
-------
TABLE 12-3
Tissue Levels of HCB (ppm) 1n Adult Female Rhesus Monkeysa«b
Monkey No.
Dose (mg/kg/day)
Body fat
Bone marrow
Adrenal cortex
Adrenal medulla
Liver
Kidney
Brain
Ovaries
Muscle
Serum
61 3C
128
930
460
150
12
20
18
25
6
4
2,5
618d
128
215
175
30
9
50
19
19
23
21
1.5
627e
64
540
1700
325
285
365
258
108
133
24
n.o
817
32
250
255
90
35
40
11
12
3
7
0.5
1163
8
580
350
50
4
30
3
8
1
2
3.3
1826
0
1.1
1.6
0.1
<0.1
<0.1
<0.1
<0.1
<0.1
<0.1
<0.1
aSource: Knauf and Hobson, 1979
bHCB was administered dally for 60 days 1n 1% methylcellulose (orally)
cMonkey was small and slight
^Monkey was obese
eMonkey had very little adipose tissue
HCB = Hexachlorobenzene
12-7
-------
The highest levels of hexachlorobenzene residues were found 1n fat tis-
sue (215-930 ppm) and bone marrow (175-1700 ppm), and selectively higher
levels were found 1n the adrenal cortex (30-325 ppm) than In the adrenal
medulla (4-285 ppm). Residues 1n semen, muscle, ovaries, brain, kidneys and
liver were relatively much lower (0.5-258 ppm).
Engst et al. (1976) reported the administration by gavage of 8 mg/kg of
hexachlorobenzene 1n 1 mi of sunflower oil to male Wlstar rats for a dura-
tion of 19 days. The animals were then sacrificed, and the liver, kidneys,
adrenals, heart, spleen and Intestinal fat were analyzed for hexachloroben-
zene residues. The following results were reported: fat tissue, 82 pg/g;
muscle, 17 yg/g; liver, 125 pg total; kidneys total 21 vg each; spleen
total 9 \ig; heart total 1.5 yg; and adrenals total 0.5 yg each. High
levels of hexachlorobenzene residues 1n fat tissues also have been reported
for rats receiving 50,0 mg/kg (177 ^moles/kg) of hexachlorobenzene every
second day for 10 weeks (Koss et al., 1980b).
Szymczynsk! and Wal1szewsk1 (1981) analyzed human semen and testlcular
and fat tissues, and Identified several chlorinated pesticides that Included
hexachlorobenzene. The compound was not detected 1n testlcular tissue, but
was present In semen and fat tissues at concentrations of 0.001 and 0.128
iig/g» respectively. Similarly, hexachlorobenzene was one of several
chlorinated compounds found 1n semen collected 1n 1979 from 132 college
students (Dougherty et al., 1981).
Sundlof et al, (1982) studied the distribution of 14C-hexachloroben-
zene or unlabeled hexachlorobenzene 1n male beagles following a single
Intravenous dose of 1 mg/kg 1n olive oil. Two dogs each were sacrificed
after 2, 4, 8, 16, 32 and 48 hours and after 12 weeks; hexachlorobenzene
concentrations were determined 1n 16 tissues and organs as well as In the
blood (Table 12-4). Two hours after dosing, the highest concentration
12-8
-------
TABLE 12-4
HC8 Concentrations 1n Tissues of Hale Beagles
Receiving Single Intravenous Doses of 1 mg/kg bw 1n Olive 011*
Tissue
Lungs
Adrenals
Subcutaneous fat
Perlrenal fat
Mesenterle fat
Spleen
Liver
Thyroid
Heart
Kidneys
Stomach
Pancreas
Brain
Duodenum
Colon
Small Intestine
Blood
2 hours
36.14
2.82
1.14
1.00
0.56
0.54
0.51
0.37
0.28
0.18
0.18
0.17
0.15
0.12
0.12
0.07
0.07
HCB Concentration (ppm)
Time Interval After Dosing
48 hours
0.08
0.38
3.38
3.24
2.40
0.01
0.04
NR
0.04
0.02
0.36
0.06
0.02
0.02
0.01
0.02
0.03
12 weeks
<0.01
0.06
0.37
0.46
0.41
<0.01
0.02
0.02
0.01
0.01
0.01
0.07
0.02
0.02
<0.01
0.01
0.01
*Source: Sundlof et a!., 1982
NR = Not reported
HCB = Hexachl,orobenzene
12-9
-------
was found 1n the lungs (36.14 ppm). This was considered to be a property of
the Injection vehicle rather than a property of hexachlorobenzene per se.
That 1s, 1t was believed that the olive oil vehicle formed mlcroemboll 1n
the blood which became trapped 1n the capillaries of the lung. Residue
levels 1n the lungs then dropped (4.4 ppm), and a concurrent Increase 1n
hexachlorobenzene was detected 1n fat tissues (10.32 ppm 1n subcutaneous,
perlrenal and mesenterlc fat) 4 hours postlnjectlon. Residues 1n all
tissues, organs and blood declined during the 48 hours postlnjectlon except
for fat tissue, which remained constant. Twelve weeks after dosing, tissue
concentrations were very low 1n all tissues, Including fat (>0.01-0.46 ppm),
Indicating significant excretion of the compound by that time.
Yang et al. (1978) studied the distribution of hexachlorobenzene 1n male
Sprague-Dawley rats and female rhesus monkeys following Intravenous Injec-
tion of 14C-hexachlorobenzene 1n 1,2-propaned1ol:plasma (1:8). Rats
received 0.1 mg of 14C-hexachlorobenzene and then were replaced 1n meta-
bolic cages for 48 hours before sacrifice. About 0.2 and 1.0% of the admin-
istered dose was excreted 1n the urine and feces, respectively. No radio-
activity was exhaled from the animals. Over 20 tissues from the rats were
analyzed and all were found to contain radioactivity. The highest levels
were 1n fat (~3 yg/g of tissue). The adrenal glands also contained a
relatively high level of radioactivity, whereas the other tissues contained
much lower levels, generally 1n the range of 1/12 to 1/300 of those 1n fat
tissue.
The tissue distribution of 14C-hexachlorobenzene 1n rhesus monkeys was
determined 1n Individual animals 100 days, 6 months and 1 year after Intra-
venous Injection of 14C-hexachlorobenzene at 0.38, 0.32 and 0.22 mg/kg,
12-10
-------
respectively. The results again Indicated that the highest levels were
present 1n fat (6069 ng/g on day 100 and 828 ng/g on day 365) and bone
marrow (1638 ng/g on day 100 and 373 ng/g on day 365) among the 30 tissues
analyzed 1n all three monkeys. The adrenal glands contained ~l/6 to 1/8 of
the levels present 1n fat, whereas the other tissues contained radioactivity
levels ranging between 1/10 to <1/800 of those 1n fat.
The transplacental transfer of hexachlorobenzene from pregnant mice,
rats and rabbits has also been reported. Brandt et al. (1980) conducted a
qualitative study on the distribution of 14C-hexachlorobenzene and several
of Its sulfur-containing metabolites 1n pregnant mice. The mice were
Injected l.v. and sacrificed at Intervals ranging between 20 minutes and 32
days after Injection. The animals were frozen, sectioned and submitted to
autoradlography. The autoradlograms showed a strong uptake of hexachloro-
benzene 1n the adipose tissues. This hexachlorobenzene was found to persist
1n the adipose tissues for more than 1 month after the administration.
Radioactive hexachlorobenzene was also found to penetrate the placenta,
resulting 1n the blood and Hver concentrations 1n the fetus which appeared
to equal those of the dams.
vmeneuve and Hlerlihy (1975) studied the placenta! transfer of hexa-
chlorobenzene In Wlstar rats and reported that hexachlorobenzene crosses the
placenta and accumulates 1n the fetus 1n a dose-dependent manner. The
females were dosed orally dally (5, 10, 20, 40 and 80 mg/kg) from gestation
day 6-16 and then sacrificed on day 22. Only Hver, brain and whole fetus
residue levels were determined In this study. Fetal liver residues
(1.8-35.8 vg/g) were much lower than those of the dams (9.3-86.0 wg/g).
The fetal brain and whole fetus levels were 1.1-17.5 pg/g and 1.5-18.9
Mg/g» respectively.
12-11
-------
VUleneuve et al. (1974) also reported that the transplacental transport
of hexachlorobenzene In New Zealand rabbits was dose-dependent. Rabbits
were mated and then treated orally with hexachlorobenzene from days 1-27
with subtoxlc doses of 0, 0.1, 1.0 or 10 mg/kg. On day 28 the dams were
killed for fetal and maternal tissue analysis for hexachlorobenzene. In
dams, the hexachlorobenzene residue concentrations were highest 1n fat,
followed by the Hver, heart, kidneys, brain, lung, spleen and plasma.
Hexachlorobenzene residues were higher 1n the fetal liver than 1n the
maternal liver.
Courtney et al. (1976) reported on the distribution of hexachlorobenzene
(assayed 90.4% hexachlorobenzene and 9.6% pentachlorobenzene) administered
via oral Intubation on days 7-11 of gestation at a dose of 50 mg/kg/day 1n a
corn oil acetone mix to five pregnant and two non-pregnant CD-I mice. They
found there were no remarkable differences 1n the hexachlorobenzene tissue
levels between the pregnant and non-pregnant animals sampled at day 12 of
pregnancy. The levels of pentachlorobenzene 1n sampled tissues were low as
compared to the very high hexachlorobenzene levels detected 1n the thymus,
skin, fat and urinary bladder. No detectable levels of hexachlorobenzene or
pentachlorobenzene were found In the control mice.
Courtney et al. (1979) studied the tissue distribution of hexachloroben-
zene 1n the maternal and fetal tissues of CD rats and CD-I mice and reported
that placentas and fetuses of both species demonstrated a dose-dependent
relationship for hexachlorobenzene residues, with levels 1n the fetuses
being higher than those 1n their corresponding placentas. The dams were
treated via oral Intubation with single or multiple oral doses (10, 50 or
100 mg/kg 1n corn oil) at different periods during gestation. The hexa-
chlorobenzene concentrations 1n mice and rat fetuses at mid-gestation were
12-12
-------
very similar. In mice, multiple low doses of hexachlorobenzene resulted 1n
higher concentrations of hexachlorobenzene 1n maternal and fetal tissues
than single doses of equivalent total doses. In another study, Courtney and
Andrews (1979) reported that 1n mice the fetus could be exposed to hexa-
chlorobenzene from maternal body burdens, established before fetal Implanta-
tion, and was not limited to maternal exposure during the postlmplantatlon
gestation.
Bleavlns et al. (1982) studied the tissue distribution and transfer of a
single dose of hexachlorobenzene given to female European ferrets (Hustela
putorlus furo). They gave a single 57.6 yg hexachlorobenzene (14C~
labeled) dose to each of three bred and five non-bred ferrets, 1n 7.5 g of
standard mink diet (22.2% fat). The dosed ferrets and offspring were
observed for 5 weeks after the kits were born, at which time they were
killed and tissue 14C-hexachlorobenzene levels were determined (Table
12-5). One ferret kit per Utter was also collected at birth and at weeks
1, 2, 3 and 4 for whole body residue determinations (Table 12-6). These
results Indicate that nursing mothers can significantly reduce their body
burdens of hexachlorobenzene, when compared to unbred female counterparts,
by transferring a large amount of the hexachlorobenzene to their offspring.
The mothers' milk contaminated with hexachlorobenzene seems to be a large
contributor to the kits' body burdens with a reported milk to placenta!
exposure ratio of 31:1. The distribution of hexachlorobenzene 1n ferrets
follows similar trends, as observed 1n the other mammals, where the highest
hexachlorobenzene levels were found 1n the I1p1d rich tissues.
The transfer of hexachlorobenzene to nursing Infant rhesus monkeys from
lactatlng mothers receiving via oral Intubation 64 mg/kg/day hexachloroben-
zene suspended 1n 1% methyl cellulose for 60 days was reported by Bailey et
al. (1980). M1lk concentrations were on the average 17-fold higher than
12-13
-------
TABLE 12-5
Mean (+SE) Hexachlorobenzene Radioactivity (dpm/g)
of Selected European ferret Tissues3*"
Tissues
Blood
Subcutaneous fat
Visceral fat
Muscle
Heart
Kidney
Spleen
Liver
Lung
Brain
Group I
(n=3)
49
4472
4429
53
34
105
13
248
1
61
i 34. 6<*
± 780. 5e
± 867. 6e
i 14. 4d
± 9.2d
± 31. le
± 7.5e
± 68. 9e
i 0.3e
± 30. Oe
Group II
(n=5)
166
19,525
19,704
384
310
611
180
1,445
241
395
1 26.8
i 1503.9
i 1666.0
± 64.0
± 56.8
± 80.4
1 24.8
± 145.2
1 18.4
i 48.5
K1tsc
(n=3)
—
11,678 + 712. 4f
—
561 i 204.8
—
209 + 37.2
—
1,420 ± 185.69
—
130 i 29.4
aSource: Bleavlns et a "I., 1982
62 days postdoslng from adult bred (group I) and unbred (group II)
female ferrets exposed to a single 57.6 wg dose of 14C-labeled hexa-
chlorobenzene and from offspring born to the bred females.
cK1t tissues, from 5-week-old offspring, were contrasted only with mater-
nal (group I) tissues.
^Significantly different (p<0.05) from group II tissue of the same type.
6S1gn1f1cant1y different (p<0.01) from group II tissue of the same type.
^Significantly different from maternal tissue (group I) at p<0.01,
9$1gn1f Icantly different from maternal tissue (group I) at p<0.05.
HCB = Hexachlorobenzene
12-14
-------
TABLE 12-6
Mean (±SE) HCB Radioactivity (dpm x 10s) of European Ferret K1tsatD
rvj
i
Weeks Postpartum
Per gram
Per whole
Increase
N1lk (per
Measure Number
of kit 3
kit 3
over previous week
ml) 3
0 1 2
3.0 + 0.19 2.7 tO.57 4.3 +
25.1 ± 1.43 76.7 ± 14.35 311.4 +
51.6 234,
6.1 +
0.67
63.39
.7
0.66
3
492
2
3
•9 ±
.5 ±
181.
.9 +
0.73
92.22
1
0.45
4
3.5 +
672.8 +
180.
1.8 t
0.50
117.63
3
0.17
5
2.7 *
805.7 *
132.
0.8 +
0.
54
8
0.
14
.25
20
aSource: Bleavlns et a!., 1982
''Born to female ferrets exposed to a single dose of 14C-1abeled hexachlorobenzene and the milk produced by those dams
HCB - Hexachlorobenzene
-------
maternal serum levels, whereas Infant serum levels were about 2- to 5-fold
higher than serum levels of their mothers. Similarly, the Infants had
higher tissue residues than their mothers and hexachlorobenzene was concen-
trated 1n the Infant fat, bone marrow, adrenals and lymph nodes.
Hexachlorobenzene residues also have been reported 1n human fat 1n the
United Kingdom (Abbott et a!., 1981, Japan (Curley et a!., 1973), and
Australia (Brady and S1yal1, 1972) and 1n human milk collected 1n Sweden
(Westoo and Noren, 1978; Hofvander et a!., 1981), Canada (Mes and Davles,
1979), Norway (Bakken and Se1p, 1976; Skaare, 1981), and Hawaii (Takahashl
et al., 1981).
12.1.3. HetaboHsm. The metabolism of hexachlorobenzene has been studied
1n male and female rats following oral administration, rhesus monkeys and
beagles following 1.v. Injection, and rabbits following 1.p. Injection
(Renner, 1981). Hexachlorobenzene 1s metabolized slowly Into other lower
chlorinated benzenes, chlorinated phenols and other minor metabolites and
forms glucuronlde and glutathlone conjugates. Tissues were found to contain
mainly unchanged hexachlorobenzene together with small amounts of metabo-
lites. Similarly, only small amounts of hexachlorobenzene metabolites were
detected 1n feces, whereas most of the metabolites were excreted 1n the
urine together with small amounts of unchanged hexachlorobenzene.
Hehendale et al. (1975) studied the metabolism of hexachlorobenzene 1n
male Sprague-Dawley rats 7 days after oral Intubation administration of a
single 5 mg/kg dose 1n arachls oil. The fat, liver, Intestines, kidneys,
lungs and brain were found to contain hexachlorobenzene primarily, along
with trace amounts of other chlorinated benzenes. Analysis of these chlori-
nated benzenes suggested the presence of pentachlorophenol, 2,4,5-trlchloro-
phenol, pentachlorobenzene and the tetrachlorobenzenes. Extraction and
12-16
-------
analysis of fecal radioactivity, which accounted for 16% of the dose, did
not reveal the presence of metabolites. Although urine contained only 0.85%
of the administered radioactivity, 1t provided the only evidence of hexa-
chlorobenzene metabolite excretion. Several unidentified metabolites were
evident following thin-layer chromatography (TIC) separation of urine, 1n
addition to 2,4,5-trlchlorophenol, pentachlorophenol and one spot was
reported to contain a mixture of chlorinated benzenes.
IE vitro metabolism studies with homogenates of the liver, lungs, kid-
neys and small Intestines produced trace amounts of chlorobenzene metabo-
lites when Incubated with [14C]-hexachlorobenzene in the presence or
absence of added cofactors. Liver mlcrosomal preparations produced amounts
of one or more chlorophenols when fortified with NAQPH; In the presence of
UDPGA, pentachlorophenol was reported to form the glucuronlde conjugate.
Fortification of kidney homogenates with glutathlone resulted 1n the appear-
ance of unextractable radioactivity 1n the aqueous phases, Indicating that
glutathlone conjugates of polar hexachlorobenzene metabolites might also be
formed (Mehendale et al., 1975).
The metabolism of hexachlorobenzene 1n male and female Sprague-Dawley
rats each receiving nine oral doses of 85.6 mg/kg hexachlorobenzene (99.7%
pure) 1n arachls oil over a period of 1 month was reported by Rlchter et al.
(1981). The animals were sacrificed 3, 24 and 52 days after the last dose,
and various tissues were analyzed for hexachlorobenzene and Its metabolites
by COE/GLC and GIG/MS. In addition to hexachlorobenzene, the following
metabolites were also detected: pentachlorobenzene (PCB), pentachlorophenol
(PCP), pentachlorothlophenol (PCTP) and 2,3,4,6- and 2,3,5,6-tetrachloro-
phenol (TCP). The results reported for the liver and kidneys for day 3
Indicated that the livers of the females contained significantly more PCTP,
12-17
-------
a derivative of a glutathlone conjugate, than those of the males (Table
12-7). However, 1t 1s not known whether this Increase 1s due to a higher
rate of PCTP production or to a lower rate of elimination.
R1zzard1n1 and Smith (1982) Investigated the sex differences 1n hexa-
chlorobenzene metabolism 1n young F344/N rats who had been Intubated every
other day for 103 days with 14 mg/kg hexachlorobenzene (analytical grade)
dissolved 1n arachls oil. Three hexachlorobenzene metabolites were analyzed
for: pentachlorobenzene, pentachlorothlophenol and 2,3,5,6-tetrachloroben-
zene~l,4-d1ol, and all three were found to be produced 1n larger concentra-
tions 1n the female rats during the first 10 weeks of hexachlorobenzene
treatment. The greater quantities of hexachlorobenzene metabolites being
formed 1n female rats was believed due to their body estrogen levels.
Engst et al. (1976) detected several urinary metabolites 1n male Wlstar
rats receiving by gavage 8 mg/kg of hexachlorobenzene dally dissolved 1n
sunflower oil for 19 days. The results of this study were presented quali-
tatively, and the authors reported that the major metabolic route for hexa-
chlorobenzene was to pentachlorophenol. In addition, the feces contained
mainly unchanged hexachlorobenzene together with traces of pentachloro-
benzene.
Koss et al. (1976) Investigated the metabolism of hexachlorobenzene 1n
female Wlstar rats given 2-3 1.p. doses of [i4C]hexachlorobenzene (260 or
390 mg/kg total dose). At the end of 4 weeks, ~l% of the administered radio-
activity was excreted 1n the urine, with >BO% of this amount contained 1n
the major metabolites (pentachlorophenol, tetrachlorohydroqulnone, and
pentachlorothlophenol). An Isomer of tetrachlorothlophenol was detected as
a minor urinary metabolite. Twenty-seven percent of the administered radio-
activity was excreted 1n the feces, of which 70% was Identified as unchanged
12-18
-------
TABLE 12-7
Concentrations of HCB and Us Metabolites {mg/kg}
1n the Liver and Kidneys of Male and Female Rats3*!3
Tissue/Sex
Liver
Males
Females
Kidneys
Males
Females
HCB
192
147C
127
111
PCB
0.05
0.03C
0.05
0.01
PCP
3.16
2.12°
5.79
3.69
PCTP
0.23
0.36C
0.24
0.10
TCP
0.02
0.04C
0.09
0.08
aSource: Rlehter et a!., 1981
bDeter«n1ned 3 days after the last of nine oral doses of 85.6 mg/kg HCB
given within 1 month 1n arachls oil
Cstat1st1cally significant from males (p<0.05)
HCB = Hexachlorobenzene; PCB = pentachlorobenzene; PCP = pentachlorophenol;
PCTP = pentachlorothlophenol; TCP = 2,3,5,6-tetrachlorophenol
12-19
-------
hexachlorobenzene. Only pentachlorophenol and pentachlorothlophenol were
Identified as fecal metabolites of hexachlorobenzene. In the tissues of the
animals, only pentachlorophenol was detected 1n measurable quantities,
accounting for 10% of the radioactivity 1n blood and <0.1% 1n body fat.
Total radioactivity contained 1n the metabolites detected 1n the animal
bodies and excreted at the end of the 4 weeks accounted for 1654 of the
administered radioactivity.
In follow-up studies, Koss et al. (1978a) compared the formation of
hexachlorobenzene metabolites 1n rats, mice, guinea pigs, Japanese quail,
laying hens and rainbow trout. The only metabolites detected were penta-
chlorophenol, tetrachlorohydroqulnone and pentachlorothlophenol; however,
the species tested differed greatly 1n their ability to metabolize hexa-
chlorobenzene (Table 12-8).
Gas-I1qu1d chromatography of urine, bile and fecal extracts from male
beagle dogs receiving a single 1.v, Injection of 14C-hexachlorobenzene at
1 rag/kg revealed that 96% of the fecal radioactivity occurred as the parent
compound. Hexachlorobenzene accounted for 4% of the biliary radioactivity,
but no parent compound was detected 1n urine (Sundlof et al., 1982).
Kohll et al. (1976) studied the metabolism of several chlorinated ben-
zenes, including hexachlorobenzene, 1n rabbits following 1.p. Injection.
The urine was collected for 10 days after Injection and analyzed for metabo-
lites following extraction and gas-l1qu1d chromatography, but no hexachloro-
benzene metabolites were found 1n the urine.
12,1.4, Excretion. The excretion of hexachlorobenzene from treated ani-
mals 1s slow and occurs mainly through the feces, with relatively little
being excreted 1n the urine. It 1s characterized by an Initial rapid phase
followed by a very slow phase. This slow phase of excretion can be enhanced
12-20
-------
TABLE 12-8
Hexachlorobenzene and Its Major Metabolites
1n the Excreta of Different Animal Species3
Spec1esb
Rat
Mouse
Guinea pig
Japanese quail
Laying hen
Rainbow trout
Total Amount
Total Dosec
(mMol/kg)
0.92
0.92
0.92
2.76
0.92
2.76
HCB
6. id
2.6
1.8
7.5
0.6
1.8
PCP
2.0
0.3
0.9
trace
0.1
0.4
of Substances
TCH
0.4
0.1
0.2
trace
0.07
ND
PCTP
1.8
ND
0.5
3.2
0.04
NO
aSource: Koss et a!., 1978a
^2-3 animals were used per each species Investigated
cHexachlorobenzene was dissolved 1n oil and administered 1ntraper1toneally.
dF1gures are given in viMol/kg bw/day
ND = Not detected. The lower detection limit of the metabolites was deter-
mined to be 0.03 nMol/mi urine or g feces.
HCB = Hexachlorobenzene; PCP = pentachlorophenol; TCH = tetrachlorohydro-
qulnone; PCTP = pentachlorothlophenol
12-21
-------
by the administration of mineral oil, paraffin and n-hexadecane. Both
biliary and Intestinal excretion contribute to fecal excretion. A three-
compartment mammalian model has been reported for the behavior of hexa-
chlorobenzene 1n beagles and rhesus monkeys following 1.v. Injection of a
single dose. Radioactivity was not detected 1n exhaled air following 1.p.
Injection of 14C-hexachlorobenzene.
Studies conducted by Hehendale et al. (1975) with rats receiving a
single oral dose Indicated that only 16.0 and 0.85% were excreted 1n the
feces and urine, respectively, 7 days after treatment (see Table 12-1).
Ingebrlgtsen et al. (1981) reported that 4 days after Intragastrlc admin-
istration of ^C-hexachlorobenzene, a total of 24.8 and 2.1% of the admin-
istered radioactivity were recovered 1n the feces and urine, respectively.
In addition, an average of 3.6% of the dose was recovered 1n the bile of
blle-duct-cannulated rats within 48 hours after dosing. Of the radioactiv-
ity excreted In the bile, only 2% was unchanged hexachlorobenzene, 1.8% was
pentachlorobenzene, 24% was pentachlorophenol and -72% was unidentified
metabolites.
Rozman et al. (1977) studied the excretion of hexachlorobenzene 1n
female rhesus monkeys receiving 110 yg 14C-hexachlorobenzene/day/monkey
via diet for 15 months. The excretion and storage patterns showed a very
slow approach to a saturation level, Indicating a high tendency for hexa-
chlorobenzene accumulation 1n rhesus monkeys. A total of 5.8 and 3.6% of
the administered dose was excreted 1n the urine of male and female monkeys,
respectively, after 15 months, of which 50% was pentachlorophenol, 25%
12-22
-------
pentachlorobenzene and the remaining 25% consisting of unidentified metabo-
lites with varying amounts of hexachlorobenzene, A total of 47.9 and 27,5%
of the dose was present 1n the feces of male and female monkeys, respec-
tively, of which 99% was hexachlorobenzene.
Koszo et al. (1978) administered hexachlorobenzene (0.2% 1n the diet) to
young male and female Wlstar rats for as long as 200 days and measured the
accumulation of hexachlorobenzene 1n the liver and fatty tissue and the
excretion of hexachlorobenzene and pentachlorophenol 1n the urine and feces.
The concentration of hexachlorobenzene 1n the liver and fat Increased stead-
ily throughout the treatment period. Pentachlorophenol appeared 1n both the
urine and feces 1n Increasing amounts throughout the treatment period, with
the excretion of other apolar and polar products being markedly enhanced
after 5-6 weeks.
R1zzard1n1 and Smith (1982) Investigated the sex differences 1n hexa-
chlorobenzene metabolism and excretion of hexachlorobenzene metabolites 1n
young F344/N rats. These rats were Intubated with 14 mg/kg analytical grade
hexachlorobenzene dissolved 1n arachls oil every other day for 103 days and
were analyzed for the three main hexachlorobenzene metabolites, pentachloro-
phenol, pentachlorothlophenol and 2,3,5,6-tetrachlorobenzene-l,4-d1ol, 1n
urine and feces. Results Indicated that the combined urinary excretion of
metabolites was greater 1n the female rats, especially during the first 10
weeks, with pentachlorothlophenol being particularly high 1n the females.
No wide variations between the sexes were seen 1n the analyzed feces hexa-
chlorobenzene metabolites after 103 days of treatment. Combined urine and
feces excretion of metabolites at the end of the study were found not to be
significantly different between males (2291ill6 nmole/ 24 hours/kg) and
12-23
-------
females (2425+.182 nmole/24 hours/kg). It was stated, though, that the total
excretion of pentachlorothlophenol was always significantly higher 1n the
female rats.
Koss and Koransky (1975) studied the metabolism of hexachlorobenzene 1n
rats when the compound was orally administered 1n an aqueous suspension or
1n olive oil. The animals received different amounts of 14C-hexachloro-
benzene 1n a single dose, and the feces and urine were collected at varying
time Intervals and radloassayed. When administered 1n water, hexachloro-
benzene was not readily absorbed; 76-97% of the dose was excreted 1n the
feces, and <0.1-0.4% was excreted In the urine 1 day after administration.
When administered In oil, only 45-46% of the dose was excreted 1n the feces
and 2.1-3.8% was excreted In the urine after 14 days of treatment. Rats
receiving 4 rag/kg of 14C-hexachlorobenzene administered 1.p. excreted a
total of 5 and 34% of the dose 1n the urine and feces, respectively, within
14 days. About 4 and 80% of the excreted radioactivity In the urine and
feces, respectively, was unchanged hexachlorobenzene. Animals Injected 1.p.
with 50.5 mg/kg [14C]hexachlorobenzene released no radioactivity 1n
exhaled air (Koss and Koransky, 1975).
Rozman et al. (1981) reported that administration of mineral oil or
n-hexadecane to female Sprague-Dawley rats or male or female rhesus monkeys
who were pretreated with 14C-hexachlorobenzene enhanced the fecal elimina-
tion of 14C-hexachlorobenzene. All animals were administered l4C-hexa~
chlorobenzene (100 mg/kg) 1n 1% methyl cellulose as a single oral Intubation
dose except for one monkey that received three consecutive dally doses and
two monkeys that received 14C-hexachlorobenzene (0.11 mg/kg) 1n sugar
pellets dally for 750 consecutive days. Aliphatic hydrocarbons were admin-
istered to the treated animals 11-40 days after hexachlorobenzene treatment.
12-24
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When mineral oil was added to the diet of the rhesus monkeys, fecal excre-
tion of hexachlorobenzene was enhanced 6- to 9-fold. Similarly, dietary
administration of hexadecane resulted 1n the same Increase 1n fecal excre-
tion of hexachlorobenzene 1n both the rhesus monkeys and rats. Residue
analyses Indicated an enhanced depletion of hexachlorobenzene from blood and
of stored hexachlorobenzene from adipose tissue. Enhanced fecal excretion
of hexachlorobenzene as a result of dietary administration of aliphatic
hydrocarbons was primarily due to Increased hexachlorobenzene elimination 1n
the large Intestine.
Rlchter and Schafer (1981) studied the Intestinal excretion of hexa-
chlorobenzene 1n male Sprague-Dawley rats using the pendular perfuslon
method. The animals were Injected 1.p. with hexachlorobenzene at 100 mg/kg
and, 1 and 4 weeks after treatment, various parts of the Intestines were
perfused with paraffin or squalane for 5 hours. The largest amount of hexa-
chlorobenzene excreted was Into the jejunum followed by the 1leum and
colon. The ratios of total hexachlorobenzene excreted during paraffin
treatment were: jejunum/1leum = 1.26 and jejunum/colon = 2.43. The authors
concluded that these results Indicate the Importance of Intestinal excretion
1n the elimination of hexachlorobenzene, and that paraffin treatment can be
one of the measures by which a long-term stimulation of hexachlorobenzene
excretion can be achieved.
Beagle dogs receiving a single 1.v. dose of 1 mg/kg excreted 44 and <6%
of the dose 1n the feces and urine, respectively, during a 12-week period
(Sundlof et al., 1982). Both biliary and Intestinal excretion contributed
to fecal excretion; however, the data Indicated that biliary excretion was
the major contributor to fecal excretion. A computer-assisted pharmaco-
klnetlc analysis of blood, urine and fecal radioactivity levels during a
12-25
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12-week period suggested a three-compartment model for the behavior of hexa-
chlorobenzene 1n beagles. The biological half-life values were calculated
for the three dogs used and ranged from 6 weeks to 3 years.
Yang et al. (1978) reported that the elimination rate of hexachloroben-
zene from male Sprague-Dawley rats and rhesus monkeys Injected l.v. with
hexachlorobenzene was slow because hexachlorobenzene 1s stored 1n the fat
tissue. The major route of excretion for the radlolabel 1n treated monkeys
was via the feces. About 17.1, 8.8 and 28.2% of the dose was excreted 1n
the feces after 100 days, 6 months and 1 year, respectively, after treatment
of Individual monkeys, with ~90% of the radioactivity determined to be
unchanged 14C-hexachlorobenzene. The cumulative urinary excretion of
hexachlorobenzene metabolites was determined to be 1.6% of the administered
dose after 1 year. An open system, three-compartment mammlllary model was
found to fit the data for plasma, fecal and metabolized hexachlorobenzene 1n
the rhesus monkey.
Koss et al. (1983) administered 100 mg/kg hexachlorobenzene 1n olive oil
every other day, via stomach tube, to female Wlstar rats for a period of 6
weeks and then observed the rats for an additional 18 months. At cessation
of hexachlorobenzene treatment they tried to assess the biological half-life
of hexachlorobenzene and determined a value of 8 days for the start of the
elimination phase, a value of 10 weeks when assessed 3 months later, and
finally a value of 1.5 years after 12 months. The authors then concluded
that It 1s not possible to establish a valid biological half-life for the
total elimination phase of hexachlorobenzene 1n rats.
Bleavlns et al. (1982) studied the excretion and transfer of hexachloro-
benzene given to female European ferrets (Hustela putorlus furo). Three
bred and five non-bred female ferrets were each given a single dose of 57.6
12-26
-------
pg ^C-hexaehlorobenzene In 7.5 g of standard mink diet (22.2% fat).
The Investigators Indicated that there were no significant differences 1n
the excretion of hexachlorobenzene metabolites, between bred and non-bred
groups, In urine for the entire 8-week study period or 1n feces during the
beginning of the study. The observed fecal excretion during the middle
weeks to the end of the study showed a leveling of the cumulative fecal
excretion 1n the bred females and a continued Increase 1n fecal excretion 1n
the non-bred female ferrets, although 1t was stated that this difference was
not statistically significant. Excretion of hexachlorobenzene or metabo-
lites 1n the milk was found to be an Important route of excretion for lac-
tatlng females, -20.3% of the Initial dose was eliminated by the fifth week
of lactation, and found to be a very Important route of exposure to nursing
offspring. The Importance of placenta! transfer and milk excretion Is fur-
ther presented by observing the time required for 50% of the Initial hexa-
chlorobenzene dose to be excreted. The bred females required 32 days to
excrete 50% while 41 days was required for the unbred females.
12.1.5. Summary. The pharmacoklnetlcs of hexachlorobenzene 1n a number of
mammalian species have been studied 1n detail following oral administration
and, to a lesser extent, following 1.v. or 1.p. Injection. No Information
was present 1n the available literature on hexachlorobenzene metabolism fol-
lowing Inhalation or topical application. Absorption of hexachlorobenzene
from the Intestinal tract appears to depend on the solvent vehicle used dur-
ing test material administration. Thus, when hexachlorobenzene 1s admin-
istered 1n olive oil, -80% of the dose 1s absorbed; when 1t 1s administered
1n an aqueous solution, 1n 1% methyl cellulose or 1n a crystalline form,
relatively little (<20%) 1s absorbed. Intestinal absorption of hexachloro-
benzene occurs primarily through lymphatic channels, with only a minor por-
tion being absorbed Into the portal circulation.
12-27
-------
Following absorption, hexachlorobenzene distributes to tissues that have
a high Upld content. The adipose tissue accumulates the greatest concen-
trations of hexachlorobenzene 1n all species studied, although bone marrow
and skin, which contain large amounts of Hplds, also accumulate hexachloro-
benzene. The adrenal cortex accumulates hexachlorobenzene at concentrations
approaching those of fat. Other tissues (e.g., liver, kidneys, lungs,
heart, spleen and blood) generally contain much lower amounts of hexachloro-
benzene. Intravenous Injection of hexachlorobenzene results 1n a tissue
distribution similar to that seen following oral administration. Hexa-
chlorobenzene 1s transported via the placenta and 1s distributed 1n fetal
tissue 1n rabbits, rats, mice, minks and ferrets.
Hexachlorobenzene 1s metabolized slowly Into other chlorinated benzenes,
chlorinated phenols and other minor metabolites and forms glucuronlde and
glutathlone conjugates. Tissues were found to contain mainly unchanged
hexachlorobenzene together with small amounts of metabolites. Similarly,
only small amounts of hexachlorobenzene metabolites were detected 1n feces,
whereas most of the metabolites were excreted 1n the urine together with
small amounts of unchanged hexachlorobenzene. There are Indications that
females produce and excrete more hexachlorobenzene metabolites than do males.
The excretion of hexachlorobenzene from treated animals 1s slow and
occurs mainly through the feces with relatively little being excreted 1n the
urine. It Is characterized by an Initial rapid phase followed by one or
more slow phases. This slow phase of excretion can be enhanced by the
administration of mineral oil, paraffin or n-hexadecane. Both biliary and
Intestinal excretion contribute to fecal excretion. A three-compartment
mammlllary model has been reported for the behavior of hexachlorobenzene 1n
beagles and rhesus monkeys following 1.v. Injection of a single dose.
12-28
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Radioactivity was not detected 1n exhaled air following 1.p. Injection of
14C-hexachlorobenzene. Hexachlorobenzene has been detected 1n the milk of
nursing mammals (see Sections 12,1.2. and 12.2.).
12.2. EFFECTS ON HUMANS
The effects of hexachlorobenzene on humans as a result of accidental or
occupational exposure have been reviewed by Courtney (1979) and Currier et
al. (1980). A few reports of data collected on occupatlonally exposed
workers have been reported with studies conducted 1n Turkey and 1n the
United States (I.e., Louisiana) on the general population following
accidental exposure to hexachlorobenzene. The exposure of humans to lexico-
logically significant levels of hexachlorobenzene 1n Turkey from 1955-1959
by Ingestlon of contaminated grain, as reported by Cam (1959, 1960}» Cam and
Nlgogosyan (1963) and Peters (1966), caused an epidemic of hexachloro-
benzene-lnduced porphyrla cutanea tarda (PCT), also known as porphyrla
turdca.
12.2.1. Ep1dem1olog1c Studies. Burns et al. (1974) found 0-310 ppb hexa-
chlorobenzene 1n blood samples from 20 vegetable spraymen. There were no
signs of PCT, and no correlations were observed between hexachlorobenzene
levels and urinary porphyrln excretion, serum glutamlc-oxaloacetic trans-
amlnase, serum glutamlc-pyruvlc transamlnase or lactate dehydrogenase.
Increased levels of urinary porphyMns were detected 1n 1 of 54 men occupa-
tlonally exposed to hexachlorobenzene (Horley et al., 1973).
A medical survey was conducted by Dow Chemical Company (Currier et al.,
1980) on 50 employees working at a chlorinated solvents plant 1n Louisiana,
to determine blood hexachlorobenzene levels and signs suggestive of PCT or
other adverse effects, as well as any changes 1n hematologlc, clinical chem-
istry and urlnalysls parameters. The results from this study are of limited
12-29
-------
value because the various parameters studied during the 4-year period were
analyzed by several laboratories using various methods and on different
Individuals. There was potential exposure to other substances also. During
various times of the study, the time-weighted-average airborne concentra-
tions of hexachlorobenzene ranged from <1-13 ppb, and wipe samples from
lurfaces 1n the control, laboratory and clerical work areas ranged from
0.03-1.24 pg/100 cm2.
The laboratory analyses and physical examinations performed on the 1977
study group and on a control group from a polyethylene plant did not reveal
any signs Indicative of PCT. Levels of hexachlorobenzene, urinary porphyMn
and coproporphyrln and the average years of exposure are listed 1n Table
12-9. A statistically significant (p<0.05) correlation was found between
hexachlorobenzene levels 1n blood and the number of years worked 1n the
plant. For the other studied parameters no statistically significant
differences were noted between the 44 chlorinated solvents workers and the
44 control workers for 1977, except for higher protein levels and lower
hematoerlt values 1n the former workers which were not considered to be bio-
logically significant. In addition, significantly lower levels of urinary
coproporphyrlns and albumin were detected 1n white male workers with hexa-
chlorobenzene blood levels >200 ppb than 1n those with hexachlorobenzene
levels <200 ppb.
Burns and MUler (1975) studied plasma hexachlorobenzene residues of 86
residents living and/or working 1n an area exposed to the production, trans-
portation and disposal of "hex" wastes (hexachlorobenzene and other chlorl-
4 *
nated hydrocarbons) 1n Louisiana. Plasma hexachlorobenzene levels were mea-
sured and correlated with demographic characteristics, occupational hazards,
12-30
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CO
TABLE 12-9
Results of Blood and Urine Analysis 1n Men Employed 1n a Chlorinated Solvents Plant, 1974-19773
Parameter
Blood HCB
(vg/D
Urinary
uroporphyrlns
(vg/l)
Urinary
coproporphyrlns
Age
(years)
Plant-years
1974
(n=50)
310.7 i 287. 7b
22.4 + 21.1
77.4 t 40.5
30.1 i 6.3
5.5 + 3.9
Study
1975
(n=49)
311.5 t 242. 9b
20.9 t 11.0
67.2 + 36.1
31.1 i 6.6
6.3 + 4-0
Group
1976
(n=49)
159.9 + 142. 7C
37.4 + 14.4
100.6 + 40.8
30.8 + 6.7
5.9 ± 4.5
1977
(n=44)
170.3 i 111.8C
26.2 + 14.3
95.2 i 48.9
31.7 + 7.1
6.6 + 4.8
Comparison Group
1977
(n=44)
0.1 ± 0.6
NR
NR
31.3 ± 6.8
6.6 t 4.4
aSource: Currier et a!., 1980; 1974-1975 results conducted by Blosdence Laboratories; 1976-1977 results
conducted by Pathology Laboratories (± Standard Deviation)
bln plasma
cln blood
N = Sample size
NR = Not reported
HCB = Hexachlorobenzene
-------
food sample analyses and house dust analyses. Average plasma levels of
hexachlorobenzene ranged from 2.4-3.6 ppb 1n exposed subjects as compared
with 0.5 ppb 1n controls (p<0.001; Table 12-10).
Higher levels of hexachlorobenzene residues, which were statistically
significant (p<0.05), were found 1n the male subjects (4.71 ppb) than 1n the
female subjects (2.79 ppb). These were not associated with race or exposure
to hexachlorobenzene through the consumption of homegrown vegetables and
animals. About 68% of the house dust samples contained an average hexa-
chlorobenzene concentration of 380 ppb as compared with 20 ppb 1n control
samples. When the hexachlorobenzene levels 1n dust were compared with the
mean plasma hexachlorobenzene residues for the same household, a significant
correlation was obtained (p<0.025). In addition, blood samples from 11
workers employed for an average of 4.8 years (10 months to 15 years) at the
chemical plant contained an average of 78.6 (14-233) ppb hexachlorobenzene.
12.2.2. Accidental Ingestlon 1n Turkey. The hexachlorobenzene-lnduced PCT
epidemic 1n Turkey, a result of exposure during 1955-1959 In Individuals who
used contaminated seed wheat for food, has been reviewed by Courtney (1979).
Cam and Nlgogosyan (1963) estimated that 0.05-0.2 g of hexachlorobenzene was
consumed per day. The method of estimation was not described. PCT 1s a
disease of disturbed porphyrln metabolism manifested by cutaneous lesions
and 1s commonly followed by hypertrlchosls (hairiness) and hyperplgmenta-
tlon. The Induction of porphyrla by hexachlorobenzene has been reviewed
(DeHattels, 1967; Granlck, 1965; Tschudy and Bonkowsky, 1972; Courtney,
1979). Porphyrlas are metabolic disorders of porphyrln metabolism that are
characterized by Increased excretion of porphyrlns and their precursors.
Normally, s-am1nolevu!1n1c add synthetase 1s the rate-limiting step 1n
porphyrln synthesis and heme acts as an end-product Inhibitor or an
12-32
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TABLE 12-10
HCB Plasma Levels 1n Exposed Individuals and Controls3
Parameter
Number of subjects
Age (years)
Black/white ratio
HCB plasma residues (ppb)
Range (ppb)
Percent positive
Percent >1 ppb
Exposed0
86
39.8 ± 19.1
1.0
2.4 + 2.3C
0-23
99
99
Controls*5
43
32.3 + 18.6
2,3
0.5
0-1.8
95
5
aSource: Burns and Hlller, 1975
^Values are mean i 1 SO
cLevel for random sample only, N=63 (3.6 +_ 4.3 for random and biased
samples, N=83)
HCB = Hexaclorobenzene
12-33
-------
end-product represser of a-am1nolevu!1n1c add synthetase. In hexachloro-
benzene-lnduced porphyrla, 4-am1nolevul1n1c add synthetase Is Induced but
heme does not suppress or Inhibit the enzyme. The activity of uropor-
phyrlnogen decarboxylase 1s decreased, consequently, porphyrln and Its pre-
cursors (e.g., uroporphyrlnogen, coproporphyrlnogen and occasionally series
I porphyrlns) are excreted mainly 1n the urine but also 1n the feces.
Increased levels of porphyrlns also can be measured 1n the liver, skin,
Intestinal tract and other tissues (Courtney, 1979). PCT appeared to occur
more frequently 1n children 4-16 years of age, whereas the number of adults
and children under 5 years of age reporting PCT was much lower (10-2454 of
cases were Individuals over 15 years of age and <5% were children below the
age of 4). A distinct disease described as "pink sore" was observed 1n
children under 1 year of age and achieved an epidemic scale. The clinical
symptoms were weakness and convulsions and usually death 1n children whose
mothers had clinical symptoms of PCT or who had Ingested contaminated bread
during gestation and/or lactation. The presence of hexachlorobenzene 1n the
milk of nursing mothers suggested that pink sore was a manifestation of
hexachlorobenzene toxldty. The reviewer states that there was a 95% mor-
tality 1n these Infants 1n addition to the very high Incidence of still-
births.
In a follow-up study, Crlpps et al. (1980) examined 32 patients 20 years
after the onset. Porphyrlns were determined 1n urine and stool specimens of
29 patients and clinically significant porphyrln levels were observed 1n 5
patients. Clinical features such as hyperpigmentation, scarring, pinched
fades, hypertrlchosls, enlarged thyroid and distinctive arthritis were
present 1n about half of the patients.
12-34
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A detailed follow-up study was also conducted with 161 Turkish patients
25 years after the Initial hexachlorobenzene Incident (Peters et al., 1982).
The patient group studied Included some of the patients previously examined
(Peters et al., 1966), Twenty-six patients were over 17 years of age at the
time of acute toxlclty, whereas the average age of the remaining patients
was 7.1 years. An evaluation of the clinical signs and symptoms 1s sum-
marized 1n Table 12-11.
The chronic disease state was manifested by generalized hyperplgmenta-
tlon and hypertrlchosls, scarring on the cheeks and hands, and tight sclero-
dermold changes of the nose with perloral scarring. The most striking clin-
ical features 1n those patients who developed signs of hexachlorobenzene
toxlclty at an average age of 7 years consisted of painless arthritic
changes with osteoporosis of carpal, metacarpal and phalangeal bones and
atrophy or failure to develop 1n the terminal phalanges. In addition,
neurologic symptoms Including weakness, parestheslas, myotonla, cogwheellng
and painless arthritic changes of the hands and feet, were observed 1n
50-70% of the patients examined. Since the signs and symptoms 20-25 years
later represented a continuum of signs and symptoms observed personally by
Peters and Gocmen (1959-1963), It was concluded that the symptoms repre-
sented the effects of both hexachlorobenzene toxlclty and changes caused by
the Induced mixed porphyrla. Control patients from the villages Inhabited
by these patients Included unaffected family members and demonstrated
clearly the uniqueness of this disorder which allowed for ready Identifica-
tion of affected patients. In addition the 60% Incidence of large thyroid
tumors 1n the females proved a sharp contrast to the 5% Incidence of thyroid
tumors 1n the geographical area. No conclusions were drawn as to the
12-35
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TABLE 12-11
Clinical Signs and Symptoms 1n Humans 25 Years After Exposure to
Low Levels 1n HCB 1n Turkey, 1955-19593
Clinical Signs/Symptoms
No. of Patients
with Symptoms^
Percent
Porphyrl a— Neurological
Weakness
Paresthesias
Sensory shading
Nervousness
Hyotonla
"Cogwheel 1ng"
Colic
Constipation
Recent red urine
Enlarged Hver
Dermatologlc
Hyper pigmentation
Scarring
H1rsut1sm
Pinched fades
Fragile skin
Thyroid enlargement
Total
Men
Women
Orthopedic and others
Arthritis
Small hands
Short stature
117 (161)
89 (161)
75 (125)
39 (60)
35 (76)
34 (125)
84 (161)
31 (161)
17 (161)
10 (161)
125 (161)
134 (161)
81 (161)
69 (161)
62 (161)
64 (161)
26 (98)
38 (63)
108 (161)
107 (161)
74 (161)
73
55
60
65
46
27
52
19
11
6
78
83
50
43
39
40
27
60
67
67
46
aSource: Peters et al., 1982
^Numbers 1n parentheses represent total number of patients examined for
this symptom
HCB = Hexachlorobenzene
12-36
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Incidence of cancer and mortality. Studies on these endpolnts are still 1n
progress and the length of time that has elapsed from the time of exposure
may not yet be adequate for drawing conclusions.
A boy and three women of the exposed Individuals treated In the early
1960's with 1.v. and/or oral edetlc add (the metal chelatlng agent EDTA)
showed no active symptoms when examined, and s.k1n pigmentation and scarring
were much less severe than 1n most of the other patients. Urine and/or
stool porphyrln studies showed that seven patients had clearly recognizable
Increases In porphyrln levels (Table 12-12). Clinical chemistry and milk
residue data are summarized 1n Table 12-13. Percent 6-am1nolevul1n1c add
values were found to be above the upper normal limit of 4 mg/8, 1n 32 of 55
patients. The average residue levels 1n human milk samples from Turkish
mothers with porphyrla was 0.5H0.75 ppm; 0.16+0.23 ppm was found 1n milk
samples from nonporphyrlc but hexachlorobenzene-exposed mothers.
12.2.3. Summary. A few epidemlologic studies with occupationally-exposed
workers have been reported, together with studies and surveys conducted 1n
Turkey and In the United States (I.e., Loulsana), on the general population
following accidental exposure to hexachlorobenzene. These studies qualita-
tively support the toxlclty of hexachlorobenzene but give little dose
response Information. Biological monitoring of plasma levels show clearly
more hexachlorobenzene In plasma of exposed compared to non-exposed Individ-
uals although no biologically significant adverse health effects were seen
during the observation periods.
The exposure of humans to hexachlorobenzene In Turkey from 1955-1959
caused an epidemic of hexachlorobenzene Induced PCT, also known as porphrya
turdca, which 1s manifested by disturbed porphyrln metabolism, cutaneous
12-37
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TABLE 12-12
Porphyrln Levels 1n Patients and Controls*
IV>
1
CO
CD
Controls
Turkey,
mean * SD
(N-33T
United States,
mean * SD
(N-40)
Hexachlorobenzene-Exposed patients
Patients with active porphyrla
(N-15)
Remainder
(N=146)
Stool (>iq/q dry yelqht
Coproporphyrln Protoporphyrln Uroporphyrln
4.80*3.2 7.65*9.83 1.41*1.57
6.1 * 4.7 21.1 i 11.6 2.8 + 2.7
70,14 12.19 25.8
(1.0-837.6) (0.7-61.8) (0.7-189.2)
5.74 9.02 1.19
(0.5-4.1) (0-103.4) (0-12.6)
Urine
Coproporphyrln
30.0 * 23.6
69.0 + 27.0
174.5
(32.6-779.3)
31.91
(0-198.4)
(ua/l)
Uroporphyrln
5.80 * 4.25
9.0 + 4.0
111.4
(16-1607)
7.25
(0-29.5)
*Source: Peters et al., 1982
-------
TABLE 12-13
Laboratory Test Results of Turkish Patients3
Test
Urine
6-Am1nolevul1n1c add, rng/8,
PorphobHlnogen, rng/l,
Copper, ppm
Z1nc, ppm
Serum
Copper, wg/dSt
Z1nc, yg/da
Creatlne klnase, units/8.
Iron, vig/da.
Thyroid function tests
Thyroxlne, yg/ds,
Tr11odothyron1ne uptake,
percent
Free thyroxlne Index
Blood
Lead, erythrocyte, yg/ds.
Uroporphyrlnogen synthetasec
M1lk hexachlorobenzene, ppm^
Patients with porphyrla
Patients without porphyrla
Normal Range
<4
<1
0.01-0.06
0.1-0.7
70-155
70-120
women, <120
men, <150
65-170
5-11
37-59
1.85-6.5
<35
>20
NA
NA
Patient Range
0.14-10.1
0.11-1.04
0.01-0.046
0.02-1.22
88-153
57-112
65-141
51-318
69-147
2.2-10.1
36-51.1
0.9-4.6
2-17
12.4-34.8
0.51 (0-3.12)
0.16 (0-1.26)
No. of
Abnormal
Results**
32 (55)
0 (56)
0 (31)
7 (31)
0 (30)
9 (29)
1 (8)
4 (11)
0 (29)
women, 5 (10)
men, 2 (9)
women, 1 (10)
men, 1 (9)
women, 4 (10)
men, 0 (9)
0 (11)
5 (30)
53 (56)
16 (77)
aSource: Peters, et a!., 1982
^Numbers 1n parentheses represent total number of patient specimens
analyzed.
cValues expressed 1n nanomoles formed per mill 111ter of RBCs per hour
dAllowable limit 1n United States for cow's milk 1s 0.02 ppm
NA = Not applicable
12-39
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lesions and hyperplgmentatlon. The authors estimated that 0,05-0.2 g/day
were Ingested. In children under 1 year of age, pink sore was observed as
well as 95% mortality 1n these Infants.
Follow-up studies conducted with patients 20-25 years after the onset of
porphyrla showed that a few patients stm had active porphyrla, whereas
>50% exhibited hyperplgmentatlon scarring as well as other dermatologlc,
neurologic and skeletal features of hexachlorobenzene toxldty. Hexachloro-
benzene residues were also found 1n the blood, fat or breast milk of some
patients.
A correlation was found between hexachlorobenzene levels 1n blood and
the number of years worked In a chlorinated solvents plant. The concentra-
tion of urinary uroporphyrlns and coproporphyrlns 1n workers ranged from
21-37 and 67-101 pg/8,, respectively, for the period between 1974 and
1977. An ep1dem1olog1c survey conducted with 86 residents In the vicinity
of this chlorinated solvents plant showed elevated hexachlorobenzene
residues 1n plasma. Higher levels of hexachlorobenzene residues were found
In males than 1n females, but these were not associated with race or food
consumption.
12.3. MAMMALIAN TOXICOLOGY
12.3.1. Acute Toxldty. Information on the acute toxldty of hexachloro-
benzene was limited to oral LDrn values determined with a few mammalian
DU
species. The following LD5Q values were reported 1n the available litera-
ture: rats, 3500-10,000 mg/kg; rabbits, 2600 mg/kg; cats, 1700 mg/kg; and
mice, 4000 mg/kg (IARC, 1979; NAS, 1977; Sax, 1979).
Graef et al. (1979) reported that hexachlorobenzene blocked the activity
of rat hepatic 3-hydroxystero1d dehydrogenase leading to the accumulation of
5B-H-stero1ds, which are known Inducers of porphyrln biosynthesis. Hexa-
chlorobenzene-lnduced porphyrla has also been reported to occur as a result
12-40
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of a deficiency 1n the uroporphyrlnogen decarboxylatlon process that 1s
catalyzed by porphyrlnogen carboxylase. This enzyme 1s the only one 1n the
herne pathway that exhibits a decrease 1n activity. The Inhibition of por-
phyrlnogen carboxylase 1n liver homogenates from female Wlstar rats with
severe porphyrla Induced by hexachlorobenzene was studied by R1os de Molina
et al. (1980). Hexachlorobenzene had no effect on enzyme activity at
10~3 M, whereas pentachlorophenol caused a 90K Inhibition at the same con-
centration. However, pentachlorophenol did not Inhibit the enzyme at a con-
centration of 1(T5 M. It was concluded that a concentration >10~s M of
pentachlorophenol, possibly together with other hexachlorobenzene metabo-
lites, was needed to cause enzyme Inhibition.
Hexachlorobenzene has also been reported to Induce the activity of
hepatic mlcrosomal enzymes 1n male or female rats following subchronlc
administration (Carlson, 1978; Carlson and Tardiff, 1976; Chadwlck et al.,
1977). Hexachlorobenzene produced a so-called "mixed-type" Induction of
cytochrome P-450 content 1n female rats resembling that produced by a com-
bination of phenobarbltal (cytochrome P-450) and 3,4-benzopyrene (cytochrome
P-448) (Goldstein et al., 1982; Debets et al., 1980a). In female rats,
hexachlorobenzene Increased the activities of 5-am1nolevul1n1c add syn-
thetase and amlnopyrlne demethylase (Ar1yosh1 et al., 1974), ethoxy-
resoruf1n-0-deethylase, amlnopyrlne demethylase, aryl hydrocarbon hydroxy-
lase,, p-n1trophenol glucuronyl transferase, and NADPH-cytochrome c reductase
(Goldstein et al,, 1982; Debets et al., 1980a). Similarly, 1n male rats,
hexachlorobenzene Increased the activities of hepatic ethyl morphine N- and
p-n1troan1sol 0-demethylases, aniline hydroxylase, and UDP glucuronyl trans-
ferase (Hehendale et al., 1975), acetanlUde hydroxylase, acetanlUde ester-
ase, procalne esterase, and arylesterase activities (Carlson et al., 1979;
Carlson, 1980).
12-41
-------
12.3.2. Subchronlc Tox1c1ty. Several oral subchronlc studies of hexa-
chlorobenzene have been reported, but no studies were located on the effects
of hexachlorobenzene following Inhalation. In several animal species, hexa-
chlorobenzene was found to cause alopecia and scabbing, decreased body
weight, Increased liver and kidney weights and Increased porphyrln levels 1n
the urine and 1n several organs. H1stopatholog1c changes were noted 1n the
liver and kidneys of rats, gastric lymphold tissue of dogs, and ovaries of
monkeys. When placed on untreated diets, rats were able to recover from
most of the toxic effects of hexachlorobenzene treatment. Hexachlorobenzene
was also reported to cause certain neurologic effects (ataxla, paralysis,
etc.) on rats, mice, hamsters and female beagles, and to Induce an Increase
1n hepatic mlcrosomal enzyme activity. Toxldty data for hexachlorobenzene
can be found 1n Table 12-14.
latropoulos et al. (1976) reported that five adult female rhesus monkeys
given dally gavage treatments of hexachlorobenzene suspended In 1% carboxy-
methylcellulose at 8, 32, 64 or 128 mg/kg/day for 60 days, showed extensive
morphologic changes 1n the ovaries. These changes were dose-related.
Subchronlc studies conducted by Koss et al. (1980a) with groups of four
female rats treated orally (probably by gavage) with 100 mg/kg of hexachlo-
robenzene 1n olive oil every other day, suggested that hexachlorobenzene
metabolites covalently bind to cytosollc proteins although no binding to
uroporphyrlnogen decarboxylase was specifically demonstrated.
EUssalde and Clark (1979) reported a significant Increase 1n the Vn
yljro metabolism of 3H-testosterone by liver mlcrosomes from male mice fed
diets containing 250 rag hexachlorobenzene/kg for 21 days. In addition,
decreases 1n the concentration of testosterone 1n the serum and 1n the
12-42
-------
TABLE 12-14
Summary of Toxkity Studies on Hexacfilorobenzene
Species
Route
Dose
Duration
Effects
Reference
Rat
(females)
Rat
oral
oral
(diet)
100 mg/kg every other
day
0.5 mg/kg/day
2.0 mg/kg/day
8.0 mg/kg/day
32.0 mg/kg/day
up to 43 days
15 weeks exposed and
held to 48 weeks
15 weeks exposed and
held to 48 weeks
15 weeks exposed and
held to 48 weeks
15 weeks exposed and
held to 48 weeks
I
CO
Rat oral 50 mg/kg every other
(females) (gavage) day
Rats oral 0.5 mg/kg twice
(females) (gavage) weekly
2.0 mg/kg twice
weekly
8.0 mg/kg twice
weekly
32.0 mg/kg twice
; weekly
Rat oral 100 mg/kg diet
(females) (diet)
15 weeks
29 weeks
29 weeks
29 weeks
29 weeks
98 days
Suggested covalent binding of hexachlorobenzene Koss et al.,
metabolites to cytosolk proteins 1980a
Transient Increases In liver porphyrln levels Kulper-Goodman
In females after termination of exposure et a!., 1977
Increases 1n liver porphyrln levels 1n females
after termination of exposure, Increased size
of centrHcbular hepatocytes
Increased liver weights, Increased liver,
kidney and spleen porphyrln levels In females
(porphyrla), centrllobular liver lesions espe-
cially In females at 48 weeks
Increased mortality In females, Intension
tremors 1n males and females and ataxla 1n a
few females, Increased liver, kidney and
spleen weights, Increased liver, kidney and
spleen porphyrln levels In females (porphyrla),
centrllobular liver lesions and splenomegaly
Increased liver, kidney, spleen and adrenal Koss et al.,
weights, porphyrla (Increased liver porphyrln 1978b
levels and Increased excretion of porphyrlns
and precursors}, tremors, hair loss and skin
lesions
Increase In relative liver weight
Boger et al.,
1979
Increase In relative liver weight, moderately
enlarged hepatocytes
Porphyrla, markedly enlarged hepatocytes,
Increase In relative liver weight
Porphyrla, markedly enlarged hepatocytes,
Increase In liver weights
Porphyrla (Increased liver lobe porphyrlns). Smith et al.,
decreased activity of uroporphyrlnogen 1980
decarboxylase
-------
TABLE 12-14 (cont.)
Species
Rat
Rat
Rat
Rat
(male)
Rat
(female)
Rat
(female)
Rat
Rat
(females)
Rat
(females)
Rat
Route
oral
(diet and
nursing)
oral
(diet?
oral
(diet)
oral
(diet)
oral
(diet)
oral
(gavage)
oral
(gavage)
oral
(gavage)
oral
(diet)
oral
(diet)
Dose
50 rag/kg diet
150 mg/kg diet
SOD, 1000 or 2000
rug/kg diet
2000 rag/kg diet
2000 nig/kg diet
3000 ng/kg diet
50, 100 or 200 mg/kg
14 mg/kg every other
day
100 mg/kg every
other day
6-8 mg/kg/day
75 mg/kg diet
(4-5 mg/kg/day)
Duration
gestation until
5 weeks of age
gestation until
5 weeks of age
3 weeks
10 weeks
100 days
11 weeks
120 days
103 days
6 weeks exposed and
held for additional
18 months
75-90 weeks
up to 2 years
Effects
Depressed resistance to L.. monocvtogenes and
T. spiral Is. enhanced thymus-dependent antibody
fesponse
Increased serum IgM and IgG, depressed resis-
tance to J... ironocytooenes and T. splralls,
enhanced thymus-dependent antibody response,
Increased liver and adrenal weights
Dose-related Increases In relative spleen,
lymph nodes, liver, adrenals, thyroid, testes
and kidney weights, dose-related Increase In
serum IgM levels, no change In serum IgG
levels, dose-related pathological changes In
liver, lymph nodes and spleen
Porphyrla found microscopically at 5 weeks and
grossly at 10 weeks using fluorescence
Elevated hepatic enzymes by 1 week and Increased
urinary porphyrln and ALA levels (porphyrla) as
early as 40 days
Decreased uroporphyrlnogen decarboxylase
activity and porphyrla after 4 weeks
Dose- and time-dependent Increase In liver and
urine porphyrlns (porphyrla)
Porphyrla In treated females, susceptibility of
females to porphyrla may be related to estrogen
levels
Porphyrla {liver uroporphyrln levels peaked 7
months postexposure and levels had not returned
to normal by 18 months), decreased liver proto-
porphyrln and coproporphyrln levels, Inhibition
of uroporphyrlnogen decarboxylase activity
until 18 months postexposure
Decline In body weights, porphyrla, enlarged
livers and liver tumors
Porphyrla, time-related appearance of severe
hepatic and renal pathologies, after 1 year 1n-
Reference
Vos et al..
1979b
Vos et al.,
1979a
Gralla et al.,
1977
Llssner
et al., 1975
Elder et al.,
1976
Carlson, 1977b
Rlzzardlnl and
Smith, 1982
Koss et al.,
1983
Smith and
Cabral, 1980
Lambrecht et
al., 1983a,b
150 mg/kg diet
(8-9.5 mg/kg/day}
creases 1n hepatomas, hepatocardnomas, bile duct
adenomas, renal adenomas and renal carcinomas
-------
TABLE 12-14 (cont.)
Species
Rat
Rat
Route
oral
(diet)
oral
(diet and
nursing}
oral
(diet)
Dose
0.32, 1,6, 8.0 or
40 mg/kg diet
0.32 or 1.6 mg/kg
diet
8.0 mg/kg diet
40 mg/kg diet
10 or 20 mg/kg diet
Duration
-130 days
gestation through
lifetime (130 weeks)
gestation through
lifetime (130 weeks)
gestation through
lifetime (130 weeks)
FO to F4 generations
Effects Reference
Hematologlcal changes at all dose levels In Arnold et al.,
males, Increases 1n liver and heart weights 1n 1985
males at 8.0 and 40 ppcn diets, no treatment-
related effects observed 1n bred females
Glycogen depletion 1n 1.6 mg/kg males; no
effects reported at 0.32 mg/kg
Increase In liver pathologies
Increased mortality as pups. Increase 1n liver
and kidney pathologies. Increase 1n adrenal
pheochromocytomas 1n females and parathyroid
tumors 1n males
No effects reported Grant et al.,
1977
i
-e»
tn
Rat
Rat
Rat
oral
(diet)
oral
(diet)
oral
(diet)
40 mg/kg diet
80 mg/kg diet
160 mg/kg diet
320 and 640 mg/kg
diet
60, 80, 100, 120 or
140 mg/kg diet
0 or BO mg/kg diet
80 mg/kg diet
to F4 generations
to F4 generations
FQ to F4 generations
to F4 generations
FO to Fia and
generations
gestation and
nursing or cross
nursed with controls
2 weeks prior to
mating to 35-36 days
after weaning
Increases 1n liver weights and aniline
hydroxylase activity
Decreased body weights, fj and F4 generations had
decreased lactation Index and postnatal viability
and Increased stillbirths
Increased mortality and decreased lactation
Index starting In F-) generation
20 and 50% mortality 1n FQ 320 and 640 mg/kg
groups, respectively, greatly reduced fertility
Index and litter size and Increase In still-
births, viability Index zero 1n f-\
Increased mortality 1n all groups at 21 days,
21-day LOjo values for pups were 100 and 140
mg/kg for F]a and FU, generations, respectively
Nursing exposure produced greater effects than
did gestatlonal exposure, effects noted were:
smaller brains, hearts, kidneys and spleens,
Increased Hver weights
Increased porphyrln levels and decreased liver
esterase activity 1n dams, no changes In
gestation Indices or neonatal survival
KHchln
et al., 1982
Mendoza
et al., 1978
Mendoza
et al., 1979
-------
TABLE 12-14 (cent.)
Species
Rat
Route
oral
(gavage)
Dose
10, 20, 40, 60, 80
or 120 rag/kg
Duration
days 6-21 of gesta-
tion
Effects
Haternal toxlclty (weight loss, tremors and
convulsions) and reduced fetal weights at 120
Reference
Khera, 1974
House
House
(male)
House
(male)
House
House
Hamster
Hamster
oral
(diet)
oral
(diet)
oral
(diet)
oral
(diet)
oral
(gavage)
oral
(diet)
oral
(diet)
Cats oral
(breeding (diet)
females)
2.5, 25 or 250
mg/kg diet
10 mg/kg diet (8.4
(mg/mouse/24 weeks)
or 50 Mg/kg diet
(35,3 tug/mouse/
24 weeks)
167 mg/kg diet
6, 12, 24 and 36*
mg/kg/day
100 mg/kg/day to
pregnant mice
200 or 400 mg/kg
diet
4, 8 or 16 mg/kg/day
3 or 8,7 mg/day/cat
21 days
24 weeks
3-6 weeks
101-120 weeks
*(15 weeks exposed
held until 120
weeks)
days 7-16 of
gestation
90 days
llfespan
142 days
and 80 mg/kg maternal doses, dose-related In-
crease In Incidence of unilateral and bilateral
14th Mb, sternal defects were also noted In
one experiment
Dose-related Increase 1n liver and decrease In Ellssalde and
prostate and seminal vesicle weights, dose- Clark, 1979
related alterations 1n testosterone metabolism,
altered hepatic enzyme levels
Dose-related reduction In weight gain, no tumor Shlral et al.,
pathology observed 1978
Impairment In host resistance as measured by Loose et al.,
Increased sensitivity to S. typhosa and £. 1978a,b
bergherl, and decrease In IgA levels
Reduced growth rate at all dose levels, short- Cabral et al.,
ened llfespan associated with tremors and con- 1979
vulslons In 24 and 36 mg/kg/day groups, dose-
dependent Increase In liver-cell tumors 1n the
12, 24 and 36 mg/kg/day dose groups
Increased maternal livers and decreased fetal Courtney
body weights, Increased Incidence of abnormal et al., 1976
fetuses per litter observed
Preclrrhotlc and drrhotlc hepatic lesions, Lambrecht
bile-duct hyperplaslas and hepatomas et al., 1982
Shortened llfespan In 16 mg/kg/day group, In- Cabral et al.,
crease In hepatomas at all dose levels, increase 1977
In liver haentangloendothellona In males and
females and an Increase In thyroid alveolar
adenomas 1n males In 16 mg/kg/day group
Height loss and Increased disease susceptibility Hansen et al.,
In bred females, dose-related decrease 1n Utter 1979
size and survival of offspring, hepatomegaly In
offspring
-------
TABLE 12-14 (cont.)
Species
H1nks
Dog
(female)
Dog
donkey
(female)
Monkey
Route
oral
(diet)
oral
(capsule)
oral
(capsule)
oral
(gavage)
oral
(nursing)
Dose
1 or 5 mg/kg diet
50 or 150 mg/kg/day
1, 10, 100 or 1000
mg/day/dog
8, 32, 64 or 128
mg/kg/day
7.51-186 ppm milk
Duration
during gestation
until 17 weeks of
age
21 days
1 year
60 days
60 days
Effects
Dose-related Increase In offspring mortality,
Induction of hepatic dFO enzymes in exposed
offspring
Liver and hepatocyte enlargement, dose-Induced
electroencephalogram dysrhythmlas
Increase In mortality, neutrophilla, and
anorexia 1n the 100 and 1000 mg dose groups,
dose-related nodular hyperplasla of gastric
lymphold tissue 1n all treated animals
Dose-related pathology in liver, kidney, ovaries
and thymus
2 of 3 Infants died as a result of exposures
Reference
Rush et al.,
1983
Sundlof
et al.. 1981
Gralla et al.,
1977
latropoulus
et al., 1976
Bailey et al.,
1980
-------
weights of seminal vesicles and ventral prostates were reported. Hexa-
chlorobenzene was also reported to cause certain neurologic lesions 1n male
and female rats, hamsters and mice fed diets containing varous levels of
hexachlorobenzene for 13 weeks. These Included hyperemla, edema, arboriza-
tion and hemorrhages 1n the brain and menlnges. The lesions extended to the
cerebrum, cerebellum, medulla, spinal cord and their menlnges. The severity
of these lesions was higher 1n males and was dose dependent 1n both sexes
(Headley et a!., 1981). Physiologic changes (electroencephalogram dys-
rhythmlas) 1n the central nervous system were reported 1n 10 female beagles
receiving gelatin capsules containing doses of 50 or 150 mg/kg of hexa-
chlorobenzene for 21 days (Sundlof et al., 1981).
Ku1per-Goodman et al. (1977) conducted a 15-week subchronlc feeding
study wherein groups of 70 male and 70 female COBS rats were fed diets pro-
viding 0, 0.5, 2, 8 or 32 mg/kg bw/day of hexachlorobenzene originally
dissolved 1n corn oil (5%) and mixed with the feed. Female rats were more
susceptible to hexachlorobenzene than males, as Indicated by all the param-
eters studied, and a NOEL of 0.5 mg/kg/day was suggested by the authors.
This NOEL may be better Interpreted as a NOAEL since a transient Increase 1n
liver porphyrln levels was observed 1n females 4 weeks after removal from
hexachlorobenzene. The 2 mg/kg/day dose may be Interpreted as a LOAEL since
this level caused Increases 1n Hver porphyrln levels 1n females even 33
weeks after removal from hexachlorobenzene, and Increases 1n the relative
observed severity of centrllobular Hver lesions as compared to control
rats. About 40% mortality occurred 1n females, but none 1n males at the
highest dose. Clinical signs Included Intention tremor, excessive Irrita-
bility, multiple alopecia, scabbing and ataxla, with hind leg paralysis at
the highest dose. There was a significant Increase 1n Hver and kidney
12-48
-------
weights at the higher doses. An Increase 1n liver weight was also found 1n
groups of 36 female Wlstar rats treated by gavage twice weekly with hexa-
chlorobenzene dissolved 1n olive oil at 32 mg/kg for 29 weeks (Boger et al.,
1979). Similarly, Koss et al. (1978b) reported a 1.5- to 2-fold Increase 1n
the weights of the liver, spleen, kidneys and adrenal glands from female
Wlstar rats treated orally (esophageal tube) with 50 mg/kg of hexachloroben-
zene dissolved In corn oil every other day for 15 weeks. When hexachloro-
benzene-treated rats were placed on untreated diets, they no longer showed
signs of hexachlorobenzene toxldty, such as dermal lesions, and body and
organ weights returned to normal (Kulper-Soodman et al., 1977; Koss et al.,
1978b). Enlarged livers were reported 1n subchronlc studies with female
beagles (Sundlof et al., 1981) and male mice (Sh1ra1 et al., 1978) adminis-
tered hexachlorobenzene In diet.
A dose-dependent enlargement of hepatocytes was observed 1n groups of 36
female Wlstar rats receiving gavage treatments of olive oil containing hexa-
chlorobenzene (99.8% pure) 0.5, 2.0, 8.0 and 32 mg/kg twice weekly for 29
weeks (loger et al., 1979). This effect was associated with the prolifera-
tion of the smooth endoplasmlc retlculum 1n the centrllobular area, and an
Increase 1n glycogen deposits; however, animals receiving 0.5 mg/kg did not
develop enlarged hepatocytes. In addition, atypical membrane complexes In
treated animals were noted and liver-cell mitochondria were moderately
enlarged and had irregular shapes. Kulper-Goodman et al. (1976) also
reported significantly enlarged hepatocytes 1n male and female COBS rats
receiving hexachlorobenzene in diets, containing 5% corn oil, at the 8.0 and
32.0 mg/kg bw dose levels for 15 weeks. They observed that this hepatocyte
enlargement consisted to a large degree of proliferation of the smooth
endoplasmlc retlculum. In males this proliferation was often associated
with large whorls of compacted membranes surrounding lipld droplets. The
12-49
-------
nuclei of enlarged hepatocytes were also enlarged while the mitochondria
were very small and sparse. They stated that this proliferation of smooth
endoplasmlc retlculum was related to the Increased drug metabolizing enzyme
activity of the liver and was considered an adaptive rather than toxic
response to the hexachlorobenzene, since the enzyme activity and liver
morphology returned to normal after exposures were discontinued. An
Increase 1n the size of centrllobular hepatocytes was also reported 1n male
and female rats receiving 2 mg/kg/day for 15 weeks, together with hlsto-
pathologlc changes 1n the spleen (Ku1per-Goodman et al., 1977).
Nodular hyperplasla of gastric lymphold tissue was reported In groups of
6 male and 6 female beagles receiving dally gelatine capsules containing 1»
10, 100 and 1000 mg hexachlorobenzene/dog/day for 12 months (Gralla et al,,
1977). Extensive dose-related hlstopathologlc changes were also observed 1n
ovaries from groups of two rhesus monkeys given dally methyl cellulose/
distilled water solutions containing doses of 8, 16, 32, 64 or 128 mg hexa-
chlorobenzene/kg of body weight by gavage for 60 days {Knauf and Hobson,
1979; latropoulas et al., 1976). Sh1ra1 et al. (1978) conducted a 24-week
study with male mice fed diets containing 10 or 50 ppm of hexachlorobenzene,
followed by a recovery period of 14 weeks. H1stolog1c examination revealed
no pathologic changes 1n the Hver or any other organ.
Lambrecht et al. (1982) fed male and female Syrian golden hamsters hexa-
chlorobenzene at doses of 0, 200 and 400 ppm 1n their diet for 90 days. The
hamsters were killed on day 91 and at 6-week Intervals through day 361. No
differences were seen 1n growth and food consumption between control and
exposed animals. The liver was reported as the most severely affected organ
exhibiting a variety of preclrrhotlc and drrhotlc lesions, bile-duct hyper-
plaslas and hepatomas. The Incidence of neoplasms found 1n this study will
be further discussed In Section 12.3.5.
12-50
-------
Hexachlorobenzene has been found to cause Increased porphyMn levels tn
the liver of male and female rats receiving the compound Incorporated Into
the diet at doses of 8 and 32 mg/kg/day for 15 weeks (Kulper-Goodman et al.,
1977). Koss et al. (1978b) reported that female rats treated orally with 50
mg hexachlorobenzene/kg every other day for 15 weeks still showed Increased
levels of porphyrln In the liver, 38 weeks after the last treatment. In
addition, porphyrln, &~am1nolevu!1n1c add, and porphoblUnogen levels In
the urine gradually Increased during the 15-week treatment period, but sub-
sequently decreased to normal levels. Smith et al. (1980) reported that the
lobes of livers from female Agus rats fed diets containing 0.01% hexachloro-
benzene developed porphyrla at different rates. During the Initial course
of treatment, porphyrla 1n the caudate lobe developed at a significantly
slower rate than the median, left or right sections of the liver, but event-
ually, all lobes became equally porphyrlc. In contrast, porphyrla was not
observed when viewed for hepatic fluorescence of porphyrlns 1n male and
female beagle dogs treated dally with 0, 1, 10, 100 or 1000 mg/dog/day for 1
year (6ralla et al., 1977), Gralla et al. (1977) observed that female CO
rats fed 0.2% hexachlorobenzene were porphyrlc using this fluorescence
method.
R1zzard1n1 and Smith (1982) clearly confirmed that female rats are more
susceptible to hexachlorobenzene-lnduced porphyrla than are male rats, and
that this difference In susceptibility Is probably associated with the
faster metabolism of hexachlorobenzene In females. They Intubated male and
female F344/N rats every other day for 103 days with 14 mg/kg (50 ymole/
kg) hexachlorobenzene dissolved 1n arachls oil and monitored the rats for
hexachlorobenzene metabolites and porphyrln levels. The results Indicated
that after 75 days of hexachlorobenzene treatment the excretion of urinary
12-51
-------
porphyrlns Increased rapidly 1n the females and after 103 days the females
had urine and liver porphyrln levels 40- and 310-fold higher, respectively,
than did the males. During this treatment period the females were found to
excrete greater quantities of hexachlorobenzene metabolites, especially
pentachlorothlophenol, than the males. Estrogen levels seem to play an
Important part 1n the Increased susceptibility of females to hexachloroben-
zene-1nduced porphyrla. When both male and female rats were pretreated
1ntraper1toneally with four doses of 20 jimole/kg of diethylstllboestrol
dlproplonate {an estrogenlc drug), both sexes had stimulated excretion of
hexachlorobenzene metabolites.
A better understanding of hexachlorobenzene-lnduced porphyrla was
provided by Koss et al. (1983). These researchers administered every other
day for 6 weeks, through stomach tube, 100 mg/kg hexachlorobenzene dissolved
1n olive oil to female Wlstar rats and then observed the rats for an addi-
tional 18 months. The rats were evaluated during both the exposure period
and the 18-month holding period for liver hexachlorobenzene levels, levels
of liver porphyrlns, and the activity of liver uroporphyrlnogen decarb-
oxylase. The results revealed a rapid Increase 1n hexachlorobenzene liver
levels which reached a plateau after 10 days of treatment and remained
constant until exposure was terminated at 6 weeks. The levels of liver
hexachlorobenzene then decreased over time with no valid biological half life
determinate. The liver porphyrln levels, however, started to rise slightly
after 3 weeks of hexachlorobenzene exposure and reached a maximum liver
porphyrln concentration ~7 months after the exposures had ceased (Table
12-15). The liver porphyrln levels decreased to a constant level -14 months
after ceasing hexachlorobenzene exposures. At 18 months after ceasing
exposures, the treated rats liver porphyrln levels were still substantially
12-52
-------
TABLE 12-15
PorphyMn Content and Uroporphyrlnogen Decarboxylase Activity
1n the Liver Cytosol of Female Rats Pretreated with 100 mg/kg HC8
Every Other Day for 6 Weeks3
Time After the
End of Treatment
1 day
7 months
14 months
18 months
Controls
Porphyrln Content
(nmol/6 ma, cytosol)''
14 + 3d
133 + 15
9 + 6
8 + 5
0.06 + 0.04
Enzyme Activity
{pmol • mg"1 » m1n~l)c
NDe
ND
ND
0.3 i 0.2d
0.5 + 0.1
aSource: Koss et a"!., 1983
^6 ma cytosol correspond with 1 g liver tissue
cpmol coproporphyrlnogen I (determined as coproporphyrln) formed from uro-
porphyrlnogen I 1n 1 m1n by 1 mg cytosol protein
dMean (+. SO) of three or four animals
eNO = Not detectable. The lower detection limit was determined at 0.02
pmol • mg"1 • mln*"1 coproporphyrln
HCB = Hexachlorobenzene
12-53
-------
higher than the levels 1n control rats. The distribution pattern of the
liver porphyMns was observed to be changed as early as after the second
hexachlorobenzene administration. The observed changes were Increases 1n
liver uroporphyrln levels and decreases 1n liver protoporphyMn and
coproporphyrln levels. The change 1n porphyrln patterns was traced to the
decreased activity of uroporphyrlnogen decarboxylase activity which was
found to be undetectable at the end of the 6-week exposure period and the
activity did not become detectable again until 18 months postexposure (see
Table 12-15). These data led the Investigators (Koss et al., 1983) to
propose that there are four phases of hexachlorobenzene-lnduced porphyrla:
During the first phase an almost constant content of hexa-
chlorobenzene and a gradual decrease of uroporphyrlnogen decarboxy-
lase activity 1s achieved. In the second phase a noticeable accu-
mulation of porphyrlns and a practically complete Inhibition of
decarboxylase activity are conspicuous. In the third phase, which
occurs after hexachlorobenzene administration has been discon-
tinued, a further accumulation of porphyrlns and a continuing Inhi-
bition of uroporphyrlnogen decarboxylase activity can be seen, even
after extensive elimination of hexachlorobenzene. During the
fourth phase a decrease 1n porphyrln content and a return of
decarboxylase activity are clearly observable.
A possible reason for the continued Inhibition of uroporphyrlnogen decarb-
oxylase activity, even after substantial elimination of hexachlorobenzene
has occurred, was also discussed 1n this report. Koss et al. (1983) pre-
sented the scenario that once hexachlorobenzene had caused an Inhibition of
uroporphyrlnogen decarboxylase activity and Increased liver porphyrln levels
that the accumulation of porphyrlns could themselves maintain the Inhibition
of the enzyme activity.
Hexachlorobenzene pretreatment has been reported to cause altered Immune
responses. Vos et al. (1979b) studied the effect of hexachlorobenzene on
the Immune system after combined pre- and postnatal exposure. Wlstar rats
12-54
-------
viere fed d\ets conta\n\ng 50 or 150 pg/kg hexachlorobenzene during preg-
nancy and lactation. The pups were weaned after 3 weeks and continued on
the test diets until 5 weeks of age, when their Immune system was function-
ally assessed. At the higher dietary level, hexachlorobenzene caused a
statistically slgnflcant Increase 1n serum IgH and IgG concentrations.
Hexachlorobenzene treatment also caused a decreased resistance to Infec-
tion with Usterla monocytogenes (Vos et al.» 1979b). The LDr_ values
3U
were reported to be 14xlOs, 7.1xlOs and 5.0xlOs bacteria 1n pregnant
Wlstar rats receiving diets containing 0, 50 and 150 mg/kg, respectively.
Similarly, decreased resistance of Tr1ch1nel1a splralls Infection, as Indi-
cated by an Increase 1n the number of larvae found 1n muscle tissue, was
noted. Hexachlorobenzene also enhanced the thymus-dependent antibody
response to J_^ splralls antigen and to tetanus toxold. No effects were
observed on allograft rejection, mltogenlc response of thymus and spleen
cells, thymus-lndependent IgM response to Escher1ch1a coll Upopolysac-
charlde, passive cutaneous anaphylaxls, and on the clearance of carbon par-
ticles and L^ monocytogenes. The authors concluded that hexachlorobenzene
suppressed cellular Immunity and enhanced humoral Immunity 1n both test
groups.
In contrast, hexachlorobenzene pretreatment of weanling rats did not
alter their cell-mediated Immunity, but did stimulate their humoral Immune
response and enhanced the in vitro responsiveness of spleen cells to dif-
ferent mltogens, which was mainly a result of an Increase 1n the number of
splenic lymphocytes. The rats received diets containing 1000 yg hexa-
chlorobenzene/g for 3 weeks after weaning, before assessing their Immune
system (Vos et al., 1979a).
12-55
-------
Loose et al. (1978a,b) found that hexachlorobenzene pretreatment also
resulted 1n Impaired host resistance. Hale BALB/c mice received diets con-
taining 167 pg hexachlorobenzene/g for 3 or 6 weeks before assessing their
Immune functions. The concentration of IgA was significantly decreased,
whereas those of IgG and IgH did not exhibit consistent significant altera-
tions as compared with the controls. Hexachlorobenzene-treated mice were
more sensitive to gram-negative endotoxln (Salmonella typhosa). showed a
decreased resistance to a malaria challenge (Plasmodlum bergheD. and
exhibited slgnflcantly depressed antibody synthesis.
12.3.3. Chronic Toxldty. Cabral et al. (1977) studied the tumor 1gen1 city
of hexachlorobenzene 1n 6-week-old Syrian golden hamsters given 0, 50 (4
mg/kg/day), 100 (8 mg/kg/day) and 200 (16 mg/kg/day) ppm hexachlorobenzene
1n their diets for their remaining Hfespan. Shortened Hfespan was
observed 1n the male and female 200 ppm dose groups after 70 weeks of expo-
sure along with marked weight reduction 1n the males. Neoplasms were
Increased by the hexachlorobenzene exposures and are reported 1n Section
12.3.5. No other pathologies were reported 1n this study.
Cabral et al. (1979) studied the tumor1gen1city of 6- to 7-week-old male
and female outbred Swiss mice given 0, 50 (6 mg/kg/day) 100 (12 mg/kg/day)
and 200 (24 mg/kg/day) ppm hexachlorobenzene for 101-120 weeks and 300 ppm
(36 mg/kg/day) hexachlorobenzene for 15 weeks and held until 120 weeks of
age. Results Indicated that shortened Hfespan occurred 1n the 200 and 300
ppm dose groups starting after the 30th week of the test and that this
reduced survival was associated with tremors and convulsions. Reduction 1n
the rate of growth was observed 1n female mice 1n the 50, 200 and 300 ppm
dose groups and more pronounced growth rate reduction was observed 1n male
mice 1n the 100, 200 and 300 ppm dose groups. An Increase 1n neoplasms were
12-56
-------
found as a result of hexachlorobenzene exposures and are discussed 1n
Section 12.3.5. No other pathologies were reported 1n this study.
Smith and Cabral (1980) fed young female Agus or MRC Wlstar rats 100 ppm
(6-8 mg/kg/day) hexachlorobenzene 1n a diet containing 2% arachls oil for 90
weeks. Hexachlorobenzene exposure resulted 1n a slower rate of body weight
gain over the study period and 1n the exposed rats possessing less hair than
the controls. Tremors or other nervous symptoms were not seen during this
study. Onset of porphyrla was observed 1n the hexachlorobenzene rats after
~3 months, as Indicated by urines fluoresdng red under UV light, and liver
porphyrla was confirmed at autopsy by a red fluorescence of the liver. The
livers were enlarged 2-fold 1n the hexachlorobenzene-exposed females and
were associated with multiple liver cell tumors. This neoplastlc Incidence
will be discussed 1n Section 12.3.5.
Male and female Sprague-Dawley rats were fed hexachlorobenzene diets for
2 years containing 0, 75 or 150 ppm hexachlorobenzene (Lambrecht et al.,
1983a,b). Four rats per group were killed at weeks 0, 1, 2, 3, 4, 8, 16,
32, 48 and 64 of the study and liver and kidney evaluations were made.
Times of appearance of lesions were as follows: 4 weeks — hepatic hyper-
emla, edema, parenchymal and hydropic degeneration, renal hyperemla, con-
gestion, swelling and parenchymal degenerations; 32 weeks — renal tubular
nephritis with hyaline casts, severe parenchymal degeneration, epithelial
necrosis accompanied by proximal convoluted tubular regeneration, and pre-
neoplastlc foci; and 36 weeks — hepatic preneoplastlc foci; and 64 weeks —
hepatic neoplasms and renal neoplasms. The Incidence of neoplasms will be
further discussed In Section 12.3.5.
12-57
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A two-generation hexachlorobenzene (analytical grade) feeding study was
conducted using Sprague-Oawley rats fed diets containing 0 (64 males, 64
females), 0.32 (40 males, 40 females), 1.6 (40 males, 40 females), 8.0 (40
males, 40 females), or 40.0 (66 males, 66 females) ppm hexachlorobenzene
(Arnold et al., 1985). The parental rats (FQ) received their respective
test diets for 90 days before mating and until 21 days after parturition (at
weaning), at which time they were killed and evaluated for hexachloro-
benzene-lnduced effects. The number of offspring (F, generation) from
these matlngs were reduced to 50 males and 50 females per dose group at 28
days of age and fed their respective parents' diets. Thus, the F, animals
were exposed to hexachlorobenzene and metabolites in utero. from maternal
nursing and from their diets for the remainder of their lifetime (130 weeks).
The results from this two-generation study Indicated no consistent
treatment-related effects upon growth or food consumption 1n either gener-
ation and no change 1n fertility, gestation or lactation Indices. A
decreased viability Index was noted 1n the 40.0 ppm group relative to con-
trols. No treatment-related effects were found In the F~ females. The
FO males were found to have significantly Increased Hver, heart and brain
absolute weights 1n the 8.0 ppm group and significantly Increased liver and
heart absolute weights 1n the 40.0 ppm group. The FQ males were observed
to have various statistically significant changes 1n hematologlcal param-
eters at all dose levels, but the authors felt that these changes were prob-
ably not biologically significant. Neoplasms were seen 1n the F, genera-
tion and are discussed 1n Section 12.3.S. In the F^ generation the
following changes were seen:
1) Centrllobular basophlllc chromogenesls showed a significant
dose-related trend 1n both males and females. Additionally, at
doses of 8.0 and 40.0 ppm the Increases were significant 1n
comparison with controls for both males and females.
12-58
-------
2) Increases In perlblHary lymphocytosls were statistically
significant 1n the 0.32, 1.6 and 40,0 ppm male groups, while
Increases In per1b1!1ary flbrosls were statistically significant
1n the 0.32 and 40.0 ppm male groups.
3) Increases 1n severe chronic nephrosls were observed which were
dose related, but statistically significant relative to controls
only for the 40.0 ppm male dose group.
In a second study conducted by Arnold et al. (1985), 50 male Sprague-
Dawley rats per group were fed hexachlorobenzene (0 or 40 ppm) and various
levels of vitamin A diet (0.1, 1 or 10 times normal control levels). The
test groups were as follows: control diet; control diet plus 40 ppm hexa-
chlorobenzene; 1/10 vitamin A diet; 1/10 vitamin A diet plus 40 ppm hexa-
chlorobenzene, 10 times control vitamin A diet; and 10 times vitamin A diet
plus 40 ppm hexachlorobenzene. Five rats per group were killed and evalu-
ated both at 25 and 49 weeks and the remaining animals were killed and
evaluated after 119 weeks.
Results revealed that the animals on the 1/10 vitamin A diet had sig-
nificantly reduced body weights and survlvablHty when compared with control
diet animals. The animals on 1/10 vitamin A plus 40 ppm hexachlorobenzene
diet had significantly decreased body weights and did not survive as long as
rats receiving the control diet plus 40 ppm hexachlorobenzene. Hematolog-
1cal evaluations revealed no consistent treatment-related effects. Neo-
plasms were observed 1n the test animals and are discussed 1n Section
12.3.5. No significant differences were found 1n the Incidence of any
pathological lesions between the test groups.
12.3.4. Mutagen1c1ty. In a dominant lethal mutation study, male rats
(strain not given) received by oral gavage 0, 70 or 221 mg hexachloroben-
zene/kg body weight dissolved In corn oil for 5 consecutive days. A dose-
dependent reduction 1n male reproductive performance was observed, but hexa-
chlorobenzene did not Induce dominant lethal mutations (Simon et al., 1979).
12-59
-------
Khera (1974) also reported a lack of dominant lethal mutations 1n Wlstar
rats following oral administration of 0, 20, 40 or 60 mg hexachloroben-
zene/kg 1n 0.25% aqueous gum tragacanth for 10 consecutive days. In 14
sequential mating trials, no significant differences 1n the Incidence of
pregnancies, corpora lutea, live Implants and dedduomas between the treated
and control groups were observed. Mutagenlc activity has been observed 1n a
yeast, Saccharomyces cerevlslae. assay (Guerzonl et al., 1976). The muta-
genlclty of hexaehlorobenzene was Investigated 1n three strains of S_^ cere-
vlslae using reversion from H1st1d1ne and methlonlne auxotrophy, and hexa-
chlorobenzene was reported to be mutagenlc at a minimum concentration of
100 ppm.
Lawlor et al. (1979) measured the activity of hexaehlorobenzene 1n the
Ames assay, strains TA98, TA100, TA1535, TA1537 and TA1538, at five unspeci-
fied dose levels both with and without metabolic ac1t1vat1on by Aroclor 1254
Induced rat liver mlcrosomes. Hexachlorobenzene possessed no detectable
levels of mutagenlc activity 1n any of the Salmonella strains used either
with or without mlcrosomal activation. These results were reported 1n an
abstract with few experimental details. In addition, this result Is not
unexpected because the Salmonella test system 1s generally Insensitive to
highly chlorinated compounds (Rlnkus and Legator, 1980).
12.3.5. Carc1nogen1c1ty. Studies on the carcinogenic potential of hexa-
chlorobenzene have been carried out on hamsters, mice and rats.
12.3.5.1 HAMSTER STUDIES --
12.3.5.1.1. Cabral et al. (1977) — In one study on Syrian golden
hamsters (Cabral et al., 1977) hexaehlorobenzene was administered 1n the
diet at 50, 100 or 200 ppm. These concentrations correspond to dosages of
4, 8 and 16 mg/kg/day based on body weight and food Intake averages. The
12-60
-------
hexachlorobenzene was prepared by dissolution 1n corn oil which was then
mixed with the feed. The feed was analyzed periodically to Insure that the
Intended level of hexachlorobenzene was maintained (Hollner, 1983). The
hexachlorobenzene preparation used 1n this study was 99.5% pure. Impurities
reported to be present 1n some hexachlorobenzene preparations Include
chlorinated dlbenzofuran and chlorinated d1benzo-p-d1ox1n, both members of
classes of compounds which are carcinogens (Vllleneuve et a!., 1974). The
dosages selected for this study were chosen 1n order to be comparable to
those believed to be consumed by victims of accidental hexachlorobenzene
ingestlon 1n Turkey.
In this study on hamsters 1t was difficult to determine from the pub-
lished report whether an MTD was reached or exceeded because the Information
on mortality and weight changes was not detailed enough for an unambiguous
evaluation. Although mortality was monitored, the Investigators only stated
that 71% of the treated animals were alive at 50 weeks and that at the
highest dose, 16 mg/kg bw/day, there was a reduced Hfespan among treated
animals after 70 weeks. The study was run for the lifetime of the animals,
but the actual duration 1n weeks was not given. Since the Investigators
also reported "marked weight reduction" 1n the highest dose group one could
conclude that the MTD may have been reached. However, In the absence of
weight data definite conclusions cannot be made.
The tumor Incidence among the hamsters 1s given In Table 12-16. The
Increased Incidence of hepatomas 1n males and females was statistically
significant 1n all treated groups. The Increased Incidence of liver haeman-
gloendothelloma 1n males and females was statistically significant 1n the
high dose groups and 1n males 1n the middle dose groups. There was a
significant dose-related trend for both tumor types. Three Instances of
12-61
-------
TABLE 12-16
Tumor Incidence 1n Hamsters Given HCB 1n the Diet*
no
i
Group
Control
50 ppm
(4 mg/kg)
100 ppm
(8 mg/kg)
200 ppm
(16 mg/kg)
Effective
No.
39 F
40 H
30 F
30 H
30 F
30 H
60 F
57 H
TBA
No.
5
3
16
18
18
27
52
56
No. of Tumors
*
12.8
7.5
53.3
60.0
60.0
90.0
86.6
98.2
No.
5
3
21
27
32
45
73
87
per
Hamster
0.13
0.08
0.70
0.90
1.06
1.50
1.21
1.52
More Than
One Tumor
No.
0
0
4
8
11
14
15
27
X
0
0
13.3
26.6
36.6
46.6
25.0
47.3
Thyroid
No.
0
0
2
0
1
1
3
8
X
0
0
6.6
0
3.3
3.3
5.0
14.0
Hepatoma
No.
0
0
14
14
17
26
51
49
X
0
0
46.6
46.6
56.6
86.6
85.0
85.9
Haemang1oendothel1oroas
Liver Spleen
No.
0
0
0
1
2
6
7
20
x
0
0
0
3.3
6.6
20.0
11.6
35.0
No.
1
0
0
1
3
3
4
4
x
2.5
0
0
3.3
10.0
10.0
6.6
7.0
Other
No.
4
3
5
11
9
9
8
6
X
10.2
7.5
16.6
36.6
30.0
30.0
13.3
10.5
*Source: Cabral et al., 1977
TBA = Tumor-bearing animals
HCB - Hexachlorobenzene
-------
metastases were found among the animals with liver haemangioendotneHoma. No
hepatoma metastases were found. One of the hepatomas 1n a female animal was
found at necropsy at 18 weeks; the Investigators did not Indicate which
dosage level this animal received.
Hamsters 1n the control groups showed no thyroid tumors but thyroid
alveolar adenomas were significantly Increased 1n the high dose males and
there was a significant dose-related trend. Thyroid tumors occurred 1n all
treated groups of females but were not significantly Increased.
Chemical Induction of thyroid tumors has not been Identified with chem-
ically related compounds except for toxaphene, which 1s a mixture of chlori-
nated camphene derivatives. Other chemicals associated with Induction of
thyroid tumors are thloureas, thlouradls, 3-am1no-4-ethoxyacetan1l1de»
amltrok, o-an1s1d1ne, 2,4-d1am1nan1sole sulfate, ethlonamide, 4,4'-methylene
bls(n.n'-dlmethyl) n,n'-d1methylbenzenam1ne, 1,5-naphthylened1am1ne, 4,4'-
oxydlanallne, pronetalol»HCl, 4,4'-th1od1anal1ne, lodoform, dlbromomethane
and dlchloroethane (Krayblll, 1983; Welsburger, 1983). Hexachlorobenzene 1s
1n a different chemical class from these agents.
Induction of thyroid tumors 1n the animal studies 1s of particular
Interest because a very high Incidence of enlarged thyroids was found among
victims of an accidental exposure to hexachlorobenzene In Turkey (Peters,
1983). The Incidence among females, examined over 25 years after the Inci-
dent, Is 61.4% whereas the background Incidence In that geographic area for
females 1s about 5% (Peters, 1983). The data and pathology reports have not
been made available yet, but 1t Is clear that the cohort exposed to hexa-
chlorobenzene has an unexpectedly high Incidence of enlarged thyroid. It
cannot be stated at present what percentage 1f any of the enlarged thyroids
1s the result of tumor 1 genesis.
12-63
-------
This hamster study provides strong positive evidence of tumor1gen1c1ty
and evidence of carc1nogen1c1ty of hexachlorobenzene, as Indicated by the
significant Increase In hepatomas, significant Increase of thyroid adenomas
1n males and the occurrence of metastaslzlng Hver haemangloendothellomas In
treated but not 1n control animals. Although not reported 1n detail 1n this
one page publication, the authors noted an Increase 1n adrenal neoplasms as
well. The data presented show that the tumor Incidence 1s positively dose-
dependent 1n most Instances and that this 1s true not only of the number of
animals with tumors of all sites but also for the number of tumors per
animal. The authors also Indicated that latency period was reduced, but
actual supporting data was not presented. Although strong evidence for
carc1nogen1c1ty was provided 1n the hamster study, a cautionary note should
be added regarding the results of this study and possibly other hexachloro-
benzene studies as well. The hexachlorobenzene used was reported to be
99.5% pure. However, chlorinated dlbenzofuran and chlorinated d1benzo-p-
dloxln, both very potent carcinogens, have been reported 1n the past to be
present 1n some samples of hexachlorobenzene. Very small amounts of such
contaminants could Influence results.
12.3.5.1,2. Lambrecht et al. (1982a) Hamster Study — Another study on
hamsters, carried out 1n a different laboratory, adds further suggestive
evidence for the tumor1gen1c1ty of hexachlorobenzene 1n hamsters (Lambrecht
et al., 1982a). This study, reported only 1n abstract form, was also
carried out 1n the Syrian golden hamster. In this study the animals were
exposed for only 90 days to the hexachlorobenzene. On day 91, half of the
Initial exposed 50 animals were sacrificed. The remaining animals were
sacrificed periodically until the end of the 1-year study. The exposure
levels used were 200 or 400 ppm hexachlorobenzene 1n the diet. Assuming
12-64
-------
that the hamsters from the Cabral (1977) study were comparable 1n weight and
dietary consumption,, these ppm figures would be approximately equal to and
twice those of the high dose used 1n the lifetime studies of Cabral et a!.,
1977). Lambrecht et al. (1982a) reported the Incidence of hepatoma at the
200 ppm level to be 7.7% 1n males and 6.7% 1n females; at the 400 ppm level
the Incidence was 5% 1n females and 14.3% 1n males. These figures are based
on the numbers of animals at risk at the time of the earliest observed
tumor. The time to first tumor was relatively late 1n the study, 276 days
for males and 255 days for females of the lower dose and 153 days for males
and 299 days for females at the higher dose. Since the test animals were
systematically sacrificed from 3 months onward8 the time to tumor figures
should be reasonably close to actual time to tumor. Table 12-17 shows the
results reported by Lambrecht et al. (1982a).
The tumor1gen1c1ty and carc1nogen1c1ty of hexachlorobenzene has been
demonstrated by one lifetime study 1n hamsters. Additional suggestive evi-
dence for tumorIgenlcity 1s found 1n a 90-day study In another laboratory.
In both cases hepatomas resulted. The longer period of exposure also
produced thyroid adenomas and metastatlc liver haemang1oendothel1oma.
12.3.5.2. HOUSE STUDIES —
12.3.5.2.1. Cabral et al. (1979) — Cabral et al. (1979) reported that
outbred Swiss mice were fed hexachlorobenzene (99.5% purity) 1n their diets
for up to 120 weeks. The hexachlorobenzene content of the diet was mon-
itored periodically during the study and the diet was found to be free of
aflatoxlns. The exposure levels used were 50, 100 and 200 ppm corresponding
to dosages of 6, 12 or 24 mg/kg/day based on body weight and food intake
averages. One other test group was given 300 ppm (36 mg/kg/day) for only 15
weeks and retained on an hexachlorobenzene-free diet for the remainder of
the study.
12-65
-------
TABLE 12-17
Effect of HC8 on Hamsters: Liver Tumors and Other Liver Lesions3
Sex
M
f
HCB
(ppm)
0
200
400
0
200
400
PC-i-Cb
Incidence
3/50
48/49
50/50
10/43
48/49
45/45
BDHC
Incidence
0
0
1/25
0
1/6
2/20
Day First
Observed
101
340
174
Hepatomas
Incidence
0
1/13
1/20
0
1/15
1/7
Day First
Observed
276
153
255
299
aSource: Lambrecht et al., 1982a
bpredrrhotlc + drrhotlc
cB1!1ary duct hyperplasla
HCB = Hexachlorobenzene
12-66
-------
Growth rates were monitored but not given 1n detail 1n the published
report. The Investigators stated that among female mice there was a reduced
growth rate for all doses except 1n the 12 mg/kg/day dose group and among
males for all doses except 1n the 6 mg/kg/day dosage group.
Survival times were reported 1n detail. Survival was essentially
unaffected 1n the two lower dosage level groups at 50 weeks, but at the high
dose only 60% of the females and 52% of the males survived 50 weeks. By 70
weeks on test the survival was decreased 1n the two lower dose groups as
well, and 1n the highest dose group 1t was down to 14% 1n females and 10% 1n
males. At 90 weeks there were only four surviving males out of the 50 and
no surviving females 1n the highest dosage group as compared with 96 and
100% survival 1n the female and male controls.
The yield of tumors 1n this study 1s given 1n Tables 12-18 and 12-19.
In Table 12-18, the effective number of animals 1s the number of animals
alive at the earliest time a liver cell tumor was observed 1n each group
while 1n Table 12-19 the effective number of animals 1s that number of
animals alive at the earliest appearing tumor for any site 1n the body
within that group. There was a statistically significant elevation 1n the
Incidence of liver cell tumors at the high dose 1n females and a marginal
Increase 1n high-dose males, with a positive dose-related trend 1n both
cases. There was also a dose-dependent decrease 1n latent period and a
dose-dependent Increase 1n the size and multiplicity of liver cell tumors
(see Table 12-18). The liver cell tumors were subsequently defined as
hepatomas (Cabral, 1983).
In this study there was a high Incidence of both lymphoma and lung
tumors 1n control mice. A dose-related decrease 1n the Incidence of lympho-
mas appears 1n the treated groups. The Investigators attributed this to the
12-67
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TABLE 12-18
Liver Tumor Incidence 1n Hlce Fed HCBa
Exposure Leve1b
(ppra diet)
100
_, 200
i\»
i
co 300
(15 weeks
exposure)
Initial
No. of
Animals
F 30
M 30
F 50
H 50
F 30
N 30
Mice with LOT
Effective^
No. Animals
F 12
H 12
F 26
N 29
F 10
N 3
No.
3
3
14
7
1
1
*
25
25
54
24
10
33
Node Size {mm)
<8
2
1
5
4
—
—
>8
1
2
9
3
1
1
HultlDl1c1tv
Single
1
2
3
2
1
—
Multiple
2
1
11
5
—
1
Aqe at Death (weeks)
Range
87-104
83-98
47-85
46-101
101
97
Average
98
89
67
73
101
97
asource: Cabral et al., 1979
b50 nice/group were used as controls while 30/sex/group were given 50 ppm. No liver tumors were detected In these groups.
cSurv1vors at time first LOT was observed 1n each group
LCT = Liver cell tumors
HCB = Hexachlorobenzene
-------
TABLE 12-19
Tumor Data on H1ce Fed HCBa
IVJ
I
Animals with Tumors
Lymphomas
Exposure
Level
(ppni diet)
Control
50
100
200
300
(15 weeks)
Initial
No.
Animals
F 50
H 50
F 30
H 30
F 30
H 30
F 50
H 50
F 30
H 30
Effective15
No.
Animals
49
47
30
30
30
29
41
44
26
16
Lunq
TBAC
No.
39
22
21
15
13
10
19
12
20
5
X
80
47
70
50
43
34
46
27
77
31
No.
21
12
16
13
5
7
5
4
8
3
X
43
26
S3
43
17
24
12
9
31
19
Average Age
at Death
(weeks)
89.6
80.8
69.8
73.7
94,4
70.4
58.2
53.2
97.7
68.6
No.
14
13
4
4
6
0
2
4
4
2
X
29
28
13
13
?0
0
5
9
15
13
Average Age
at Death
(weeks)
89.0
83.8
84.5
87.0
83.5
66.5
82.5
91.2
83.5
Liver-cell
No.
0
0
0
0
3
3
14
7
1
1
X
0
0
0
0
10
10
34
16
4
6
Gonads
No.
3
0
2
0
1
0
1
1
3
0
X
6
0
7
0
3
0
2
2
12
0
Other
No.
9d
46
2'
0
39
1"
11
0
83
0
X
18
9
7
0
10
3
2
0
31
0
aSource: Cabral et a1.» 1979
lumber of survivors at moment of appearance of first tumor at any site 1n each group
cln relation to the effective number
^Skln flbrosarcoma, uterine haernangloendothelloma, one skin haemangloendothelloma, two adrenal adenoma, two maranary adenoma
6Ur1nary bladder transition cell carcinoma, one liver haemang1oendothe11oma, one skin haemangloendotheHoma, one skin flbrosarcoma
fflne uterine haemangloendothelloma, one skin flbrosarcoma
9Two skin flbrosarcoma, one skin haemangloendothelloma
hflne skin squamous-cell carcinoma
10ne Intestinal lelomyosarcoma
3flne skin flbrosarcoma, two liver haemangloendothelloma, one cecum carcinoma, one stomach papHloma, one skin haemangloendothelloma, one
uterine adenoma, one mammary adenoma
HCB = Hexachlorobenzene
-------
decreased survival time of hexachlorobenzene-treated animals. This seems
reasonable but does not explain the reduction 1n lung tumors 1n the 50 ppm
(6 mg/kg/day) group when they are compared to controls, since there was not
an appreciable reduction of Hfespan 1n this low dose group.
This study by Cabral (1979) demonstrates the tumor1gen1c1ty of hexa-
chlorobenzene In Swiss mice by the significant Increase In liver cell tumors
1n both sexes and by the demonstration of dose-dependency 1n the response
with respect to tumor Incidence, tumor size, multiplicity and latent period
duration. Tumor1gen1c1ty was detected as low as 12 mg/kg bw/day (100 ppm)
for lifetime exposure but not at 6 mg/kg bw/day (50 ppm).
12.3.5.2.2. Lambrecht et al. (1982b) -- Swiss mice exposed to hexa-
chlorobenzene for only 90 days at levels of 100 and 200 ppm 1n the diet
showed degenerative changes of liver and kidneys when examined at various
Intervals after they were removed from the hexachlorobenzene-contalnlng diet
(Lambrecht et al., 1982b). Although liver tumors were not reported, treated
animals showed lymphosarcomas 1n both dosage groups 1n both sexes at levels
significantly above those of controls. Exposure to hexachlorobenzene 1n
this Instance produced leukemogenlc changes. The animals were not permitted
to live beyond selected Intermediate sacrifice dates, so 1t was not possible
to determine whether survivors would have developed liver or other tumors.
The method of preparation of the hexachlorobenzene-contalnlng diet may have
been different in the Cabral et al. (1979) and Lambrecht et al. (1982b)
studies, but detailed Information was not presented 1n the Lambrecht et al.
(1982b) abstract.
Mice may be somewhat less sensitive than hamsters to hexachlorobenzene
as evidenced by the difference In Incidence of hepatoma formation at various
doses. These animal species may differ 1n the distribution of the hexa-
chlorobenzene Into various tissue compartments (Lambrecht et al., 1981), and
12-70
-------
differ 1n rates of metabolism and absorption. Administration of the same
levels of hexachlorobenzene In the feed can be expected to give different
effective dosages.
12.3.5.2.3. Sh1ra1 et al. (1978) — Shlral et al. (1978) administered
hexachlorobenzene to male ICR mice (35 animals/group) at levels of 10 or 50
ppm 1n the diet for periods of 24 weeks. Polychlorlnated terphenyl was
given alone to another group at 250 ppm, and 1n combination with 50 ppm
hexachlorobenzene to a third group. Animals were examined h1stolog1cally at
40 weeks.
Final body weights were slightly lower 1n the hexachlorobenzene-treated
groups while liver weights were higher. Examination of the livers showed
that the hexachlorobenzene-treated groups had hypertrophy of the centri-
lobular area at both doses. No Hver tumors were found in either group.
The total Intake of hexachlorobenzene was calculated to be 8.4 and 35.3
mg/mouse over 24 weeks In the 10 ppm and 50 ppm groups, respectively.
Polychlorlnated terphenyl alone, at 250 ppm (total dose 207.4 mg/mouse)
gave 3/28 (10.7%) nodular hyperplasla. When this same level of polychlorl-
nated terphenyl was given along with hexachlorobenzene at 50 ppm (total dose
36.9 mg/ mouse) there were 23/26 (88.5%) nodular hyperplasla and 8/26
(30.8%) hepatocellular carcinoma. This response Indicates that hexachloro-
benzene can enhance the carcinogenic potency of polychlorlnated terphenyl.
The duration of administration, 24 weeks, 1n this mouse study and the
doses used were below those used in the Cabral (1979) study on Swiss mice
and also below the levels used 1n the 13-week study by Lambrecht (1982b) on
Swiss mice. Therefore, 1t 1s not surprising that hepatomas were not found
when hexachlorobenzene was given alone. The occurrence of liver lesions,
however, does Indicate the liver is a target organ.
12-71
-------
These three studies 1n mice demonstrate the tumor1gen1city of hexa-
chlorobenzene with respect to the Induction of hepatomas, the leukemogenlc
effect of subchronlc exposure and the ability of hexachlorobenzene to
enhance the carcinogenic effect of another compound.
12.3.5.3. RAT STUDIES —
12.3.5.3.1. Smith and Cabral (1980) —The carcinogenic potential of
hexachlorobenzene was tested 1n several different laboratories 1n rats. In
one study (Smith and Cabral, 1980) small numbers of female Agus rats, and
even smaller numbers of female Wlstar rats, were used. There were 12
control and 14 treated Agus rats and 4 control and 6 treated Wlstar rats.
The hexachlorobenzene was analytical grade (99.5% purity) dissolved 1n
arachls oil and mixed with the feed to give 100 ppm In the diet. This
dietary level supplied an average dally dose of 6-8 mg/kg/day to the rats.
In this study the Agus rats showed signs of porphyrla after 3 months
exposure to hexachlorobenzene, but other toxic manifestations were not
found. The Investigators stated that "there was a steady decline 1n body
weight to eventually 80% of control animals" (Table 12-20). Examination of
the weight data presented 1n the publication Indicates that this Interpreta-
tion 1s based upon comparison of "final" average weight 1n control
(286£I9 g) and treated (225+16 g) animals (see Table 12-20), representing a
21% difference 1n weight. This method of comparison can be misleading since
the final weights represent accumulated differences 1n growth rates and
varying composition of the groups because of animal deaths. An effect
produced, even transiently, at an early age, may persist 1n the figures,
even though all subsequent growth may be normal. Growth rates, rather than
absolute difference 1n weights provide a more suitable picture of the animal
response. Growth rates for the time Intervals reported were calculated
based on the data given 1n the publication and are shown 1n Table 12-21.
12-72
-------
TABLE 12-20
Body Weights of Female Agus Rats Fed Hexachlorobenzene for 90 Weeks3
Body Weight (g)
Weeks of Diet
0
10
30
50
90
Control
46 ± 6 (8)
191 + 5
236 i 13
257 ± 17
286 + 19 (8)
HCB
45 ± 24 (9)
180 ± 17
212 i 13b
221 t 19C
225 + 16 (7)c
% Difference
2
6
10
14
21
aSource: Smith and Cabral, 1980
^Significantly different from controls as assessed by Student's t-test
p<0.01
cp<0.001
Female Agus rats were fed HCB (100 ppm) 1n MRC 41B diet for 90 weeks and
then killed. Weights are means (no. of animals 1n parentheses) +_ S.D.
HCB = Hexachlorobenzene
12-73
-------
TABLE 12-21
Growth Rates for Female Agus Rats on a Diet Containing 100 ppm HCB*
Average Growth Rate %/week
Interval (on diet)
0-10 weeks
10-30 weeks
30-50 weeks
50-90 weeks
Control
31.5
1.2
0.45
0.28
Treated
30.0
0.89
0.22
0.05
*Source: Calculated from Smith and Cabral, 1980
HCB = Hexachlorobenzene
12-74
-------
The equation used was:
weight atend of Interval - weight at start of Interval ,QQ
weight at start of Interval
According to this calculation weight Increases occurred 1n both groups
during each time Interval, although the Increases were less In the treated
groups.
The survival of the treated Agus rats was good; one test animal was
sacrificed at 52 weeks and a second one died of pneumonia at 70 weeks. Both
of these animals had Hver cell tumors found by hlstologlc examination.
Another five treated animals were sacrificed at 75 weeks and the remaining
seven treated animals lived until the end of the experiment at 90 weeks.
Among controls, one was killed at 63 weeks and three more at 75 weeks. The
remaining eight were killed at 90 weeks.
No control animals had liver pathology. In contrast, 14/14 (100%) of
the treated Agus rats had liver tumors; the earliest of these was detected
at 52 weeks. The livers of the treated animals were grossly enlarged and
some of the tumors were 1.5-2 cm 1n diameter. Although one liver cell tumor
was described as pedunculated, hlstopathology detail was not given, except
to note the absence of metastases In all cases. Four of the six (67%)
Wlstar rats also had Hver cell tumors and none of the four controls showed
such pathology at 75 weeks.
In this rat study hexachlorobenzene was a potent Inducer of liver
tumors, causing a 100% Incidence with the earliest tumor observed at 52
weeks. It 1s Important to determine whether the magnitude of the effect 1s
all attributable to the hexachlorobenzene or whether contaminants, unusual
characteristics of the test animals, or procedural factors were operative 1n
this study. In this context the following points are noted.
12-75
-------
First, historical control data on tumor Incidence for Agus rats were not
available, but, according to Cabral (1983), the Agus rat 1s a strain partic-
ularly sensitive to porphyrla and hepatic tumors. In regard to the question
of contaminants, peanut oil 1s generally believed to be free of aflatoxlns
[they are destroyed 1n processing {MAS, 1977)] and the feed was analyzed for
both aflatoxlns and dlbenzofurans and found to be free of both {Cabral,
1983). Absorption 1s another factor to consider. The absorption of the
hexaehlorobenzene 1n these animals might be enhanced by dissolution 1n the
arachls oil.
12.3.5.3.2. Lambrecht et al. (1983a,b, 1984) — Another study on rats
was carried out by Lambrecht et al. (1983a,b, 1984). In this study 94
Sprague-Dawley rats of each sex for each dosage and control groups were
used. Four animals of each group were sacrificed at each of 10 Intervals:
0, 1, 2, 3, 4, 16, 32, 48 and 64 weeks. The remaining 54 animals of each
group were allowed to continue until they died, or to the end of the 2
years. The number of animals at risk was considered to be those that sur-
vived at least 12 months, since this was the earliest time to tumor. This
number would be, at minimum, 54 plus some animals from the last sacrifice
time.
The hexaehlorobenzene was highly purified and the prepared diet moni-
tored for hexachlorobenzene levels periodically. The preparation was also
analyzed for aflatoxlns and found to be negative. The test diet was
prepared by mixing the hexachlorobenzene with dextrose and Wayne laboratory
feed (1.5 g hexachlorobenzene * 98.5 g dextrose + 9.9 kg lab chow to give
150 ppm hexachlorobenzene). Half the amount of hexachlorobenzene was used
1n the mix for the 75 ppm hexachlorobenzene level. This oil-free vehicle 1s
different from the vehicle used by both Smith and Cabral (1980) and Arnold
12-76
-------
et al. (1985). The hexachlorobenzene was well absorbed as demonstrated by
progressive accumulation 1n fat which was measured 1n this study.
Based on an average food consumption of 22.6 g/rat/day for males and
16.5 g/rat/day for females, and on an average adult weight for females of
265 g and for males of 400 g, the low dose was calculated to be 4-5 mg/kg/
day and the high dose, 8-9.5 mg/kg/day. In order to compare the results
obtained 1n this study with those obtained 1n Sprague-Dawley rats by Arnold
et al. (1985), more detailed calculation of doses at different time periods
on test are given 1n Table 12-22.
The administration of hexachlorobenzene 1n the diet at these doses 1n
the Lambrecht et al. (1983a) chronic feeding study 1n rats resulted 1n liver
pathology just before the appearance of hepatoma or hepatocellular carci-
noma. Pathology observed at the early sacrifice time Included parenchyma!
degeneration, preneoplastlc foci and adenoma. At 48 and 64 weeks of the
test females had gross liver tumors which measured between 1 and 2 mm2.
Porphyrla was also detected.
Rats that lived 12 months or longer showed a significant Increase In
hepatoma Incidence In both sexes. A statistically significant Increase 1n
the Incidence of hepatocellular carcinoma was found at both doses 1n the
females, and 1n males a slight non-significant Increase was found. None of
the liver cell tumors metastaslzed. Table 12-23 summarizes the findings.
Renal cell adenoma was found to be significantly elevated 1n both sexes
but with greater frequency In males. In this study the control male group
had a high Incidence of renal cell adenoma which was not explained; never-
theless, the Increase 1n the hexachlorobenzene-treated animals was statis-
tically significant. The Incidence of renal cell carcinoma in treated
animals was not significantly increased over control animals 1n either males
or females.
12-77
-------
TABLE 12-22
Dosage Levels 1n the Chronic Feeding Study of Hexachlorobenzene
1n Sprague-Dawley Rats3
(mg/kg/day)
Time on D1etb
(weeks)
0
26
52C
79
99
Hales
75 ppm
19.5
3.2
3.3
3.4
6.2
150 ppm
37.0
7.1
6.4
6.7
10.0
Females
75 ppm
16.1
3.7
3.8
3.5
4.3
150 ppm
32,2
8.7
8.0
8.4
10.6
aSource: Calculations and data provided by Lambrecht, 1984
^The animals were 3 weeks old when placed on test
cAt 52 weeks on test the males consumed an average of 24.7 g of the diet/
day and weighed an average of 553.7 g. The females consumed an average of
16.0 g diet/day and weighed an average of 311.7 g.
12-78
-------
TABLE 12-23
Liver and Kidney Tumors 1n Sprague-Oawley Rats Given Hexachlorobenzene
1n the Diet for up to 2 yearsa»b
Exposure
Level
0
percentage
75 ppm
percentage
150 ppm
percentage
Hepatoma
M
0/54
0
10/52
19
11/56
20
F
0/52
0
26/56
46
35/55
64
Hepatocellular
Carcinoma
M
0/54
0
3/52
6
4/56
7
F
0/52
0
36/56
64
48/55
87
Renal Cell
Adenoma
M
7/54
13
41/52
79
42/56
75
F
1/52
2
7/56
13
15/54
28
Renal Cell
Carcinoma
M
0/54
0
0/52
0
0/56
0
F
1/52
2
2/56
4
2/54
4
aSource: Lambrecht et a!., 1983a,b; Lambrecht, 1983
bThe diet was prepared without solubH1zat1on of the hexachlorobenzene,
but by mixing 1t as a pulverized solid.
12-79
-------
In an updated report from this laboratory (Peters et al., 1983) hlsto-
pathology details were supplied. These data show that 1n addition to the
liver and kidney lesions there was an Increase 1n adrenal pheochromocytoma
1n female rats which was statistically significant at both 75 and 150 ppm.
Females also had elevated Incidences of adrenal cortical adenoma and
hemangloma 1n the treated groups. Among males the background Incidence of
adrenal pheochromocytomas 1s high (76.5%), making 1t difficult to determine
whether the 90.6% Incidence found 1n the 150 ppm group has any biological
significance. Other adrenal neoplastlc and non-neoplast1c lesions were
detailed: hyperemla and/or congestion, cortical hyperplasla, preneoplastlc
foci, cysts, Upoma and adenocardnoma; none of these were elevated 1n the
treated animals. The adrenal tumor Incidences are given In Table 12-24.
One point to consider 1n the Interpretation of the results, particularly
1n terms of their application to risk assessment, 1s the form 1n which the
hexachlorobenzene was administered 1n the diet. The absorption from a
partlculate form Introduces an additional possible exposure route, namely,
from the food preparation by Inhalation. This consideration does not
Invalidate the study, but raises the question of the actual exposure levels
If an additional route of exposure was occurring 1n the same experiment
simultaneously with oral Ingestlon. The effect of mixing the hexachloroben-
zene 1n the diet 1n an oil-free form may also affect absorption and thereby
the effective dose.
12.3.5.3.3. Arnold et al. (1985) — In this study hexachlorobenzene
(organic analytical grade) was administered to parental male and female
Sprague.-Dawley rats for 3 months. These animals were mated at that time and
the females continued to receive hexachlorobenzene-containlng diets during
pregnancy and throughout lactation. At weaning, 50 pups of each sex were
12-80
-------
TABLE 12-24
Adrenal Tumors 1n Sprague-Oawley Rats Given Hexachlorobenzene
1n the Diet for up to 2 Yearsa»b
MALES
Days on diet 400-599
Exposure ppm 0 75 150
hexachlorobenzene
Number of tissues 17 23 28
examined
Cortical adenoma 326
(X)
Pheochromocytoma 369
(X) (17.6) (26.1) (32.1)
Hemangloma (%) 000
0
34
6
26
(76.5)
0
600 +
75
25
3
17
(68)
0
150
23
4
21
(91.
0
3)
FEMALES
Days on diet 400-599
Exposure ppm 0 75 150
hexachlorobenzene
Number of tissues 12 5 13
examined
Cortical adenoma 032
(X)
Pheochromocytoma 002
(X)
Hemangloma (%) 0 0 2
0
35
2
(5.7)
5
(14.3)
3
(8.5)
600+
75
47
11
(23.4)
31
(66)
8
(17)
150
32
6
(18.
29
(90.
5
(15.
8)
6)
6)
aSource: Peters et al., 1983
bThe diet was prepared without solublllzatlon of the hexachlorobenzene,
but by mixing 1t as a pulverized solid.
12-81
-------
separated and fed for the remainder of their lifetime on hexachlorobenzene-
contalnlng diets. Controls were fed diets free of hexachlorobenzene. The
range of doses used 1n this study Is considerably lower than those used by
either Smith and Cabral (1980) or Lambrecht et al. (1983a,b). Table 12-25
shows the doses used 1n the Arnold et al. study at particular points 1n time
since the doses were not adjusted throughout the study. These doses repre-
sent a greater exposure to the test animals from the point of view of expo-
sure duration, since the F, animals were exposed Ijn utero and during
nursing 1n addition to their exposure from feeding on an hexachlorobenzene-
contalnlng diet. Total doses cannot be calculated since the actual dose
received during nursing 1s not known.
Arnold et al. (1985) found no differences 1n treated F, animals when
compared to controls with respect to growth rates, food consumption or hema-
tology. The only observed difference was a decreased viability Index for
pups 1n the 40.0 ppm dose group.
Hlstopathology showed that F, females had a significant elevation 1n
neoplastlc liver nodules and 1n adrenal pheochromocytoma 1n the high dose
females compared to controls (Table 12-26). There was also a significant
positive dose-related trend 1n the Incidence of these tumors In F.. females.
Among F, males, 1n the highest dose group parathyroid tumors were
significantly Increased: 25% (12/48) 1n the treated groups and 4.2% (2/48)
among controls. Females also showed a few parathyroid tumors 1n the two
highest dose groups but none 1n controls or 1n the two lowest dose groups.
The differences were not significantly different from controls. Table 12-26
gives the tumor Incidences. Although kidney tumors were not reported to be
elevated, there was an Increased chronic nephrosls 1n the F, treated
animals.
12-82
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TABLE 12-25
Intake of Hexachlorobenzene (mg/kg/day) In the Chronic Feeding,
2-generat1on Study of Hexachlorobenzene 1n Sprague Dawley Rats
Time on D1etb
(weeks)
1
30C
70
1
30C
70
0.32 ppm
0.04
0.01
0.01
0.04
0.02
0.01
Exposure
1 .6 ppm
MALES
0.12
0.06
0.05
FEMALES
0.17
0.08
0.06
Level
8.0 ppm
0.93
0.29
0.25
0.84
0.40
0.32
40.0 ppm
4.85
1.5
1.3
4.64
1.9
1.6
aSource: Calculations and data provided by Arnold, 1984
animals were placed on feed at 6 weeks of age.
cThe mean body weight of male controls was 663 g and for the highest dose
group males 653 g. The mean weekly food consumption for male controls at
that time was 178 g and for the highest dose group 169 g. Females of the
same age weighed 351 g for controls and 353 g for the highest dose treated
group and the mean weekly food consumption was 113 and 118 g, respectively.
12-83
-------
TABLE 12-26
Tumors In Organs that Showed Statistical Differences from Control In FI Sprague-Dawley Rats Treated with Hexachlorobenzene3
[Incidence (X)]
Parathyroid Adenoma
Dose at 30 weeks
(mg/kg bw/day)
Controls
0.01-0.02
0.06-0.08
_i 0.29-0.40
1°
00
"*" 1.5-1.9
Other statistical tests
IARC trend test
Armltage time-related
trend test
Fisher exact
treated vs. control
Hales
2/48 (4.2)
4/48 (8.3)
2/48 (4.2)
1/49 (2.0)
12/49 (24.5)
p<0.01
p<0.01
p<0.05
Females
0/49 (0)
0/49 (0)
0/50 (0)
1/49 (2.0)
2/49 (4.1)
p<0.05
p<0.05
Adrenal Pheochromocytoma
Hales Females
10/48 (20.8) 2/49 (4.1)
12/48 (25.0) 4/49 (8.0)
7/48 (14.6) 4/50 (8.0)
13/49 (26.5) 5/49 (10.2)
17/49 (34.7) 17/49 (34.7)
p<0.01 p<0.01
p<0.05 p<0.01
p<0.01c
Hepatocellular Carcinoma
Hales Females
0/48 (0) 0/49 (0)
2/48 (4.2) 0/49 (0)
1/48 (2.1) 0/49 (0)b
1/49 (2.0)b
2/49 (6.1) 0/50 (0)
1/49 (0) 0/49 (0)b
1/49 (2.0)b
NeoolasUc Liver Nodules
Hales Females
2/48 (4.2) 0/49 (0)
0/48 (0) 0/49 (0)
0/48 (0) 2/50 (4.0)
2/49 (4.1)b 2/49 (4.1)b
3/49 (6.1)b 3/49 (6.1)b
1/49 (2.0) 10/49 (20.4)b
9/49 (18.4)b
p<0.01
p<0.01
p<0.01c
aSource: Arnold et al.. 1985; Arnold, 1984
bD1fferent results of two different pathologlsts reading the same slides
cCompar1son of high dose group versus control
-------
12.3.5.3,4, Arnold et al. (1985) — In another study by Arnold et al.
(1985) which was related to the 2-generat1on study, the effect of vitamin A,
because of Its supposed ant1tumor1gen1c properties, was tested 1n conjunc-
tion with hexachlorobenzene. This was a l-generat1on study and the level of
hexachlorobenzene was the same as the highest dose of the 2-generat1on
study, 40 ppm. There were six separate groups of 50 animals each and the
experiment ran for 119 weeks. At 29 weeks and at 49 weeks five animals from
each group were sacrificed and evaluated hlstologlcally. The six groups are
shown 1n Table 12-27. The vitamin A did not apparently alter the effects of
hexachlorobenzene. The number of animals with parathyroid tumors and
adrenal, pheochromocytomas was somewhat elevated 1n all the cases 1n which
hexachlorobenzene was administered compared with the total cases with the
three levels of vitamin A and no hexachlorobenzene. The significance of
these tumor Incidences cannot be determined by simple comparison because 1t
was also found 1n the study that vitamin A had an effect on the background
level of some common tumors and these data have not yet been completely
analyzed.
12.3.5.4. DISCUSSION OF RAT STUDIES — It seems appropriate to compare
the findings of Smith and Cabral (1980) 1n Agus and Wlstar rats, Lambrecht
et al. (1983a,b) and Arnold et al. (1985) 1n Sprague-Dawley rats. None of
the three studies agree precisely on all four of the tumor target organs:
Smith and Cabral reported liver tumors, Lambrecht reported liver, adrenal
and kidney tumors and had some liver carcinomas not found by Smith and
Cabral. Arnold found adrenal and parathyroid tumors and neoplastlc liver
nodules but no Increase 1n kidney tumors. We find that, although differ-
ences do occur, the results are not contradictory for the following reasons:
12-85
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TABLE 12-27
Parathyroid and Adrenal Pheochromocytomas 1n Sprague-Dawley Rats
Maintained on Synthetic Diets of Varying Vitamin A Content and
With or Without Hexachlorobenzene*
Group No. with
Parathyroid Tumors
Controls on diet with normal
vitamin A content
Control diet + 40 ppm HCB
Diet with 0.1 times normal vitamin A
Diet with 0.1 times normal
vitamin A + 40 ppm HCB
Diet with 10X vitamin A
Diet with 10X vitamin A + 40 ppm HCB
Total without HCB
Total with HCB 40 ppm
3
4
0
0
1
3
4
7
No. with Adrenal
Pheochromocytoma
4
6
2
2
4
7
9
15
*Source: Arnold et al.f 1985
HCB = Hexachlorobenzene
12-86
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1. The dosages used In the Arnold et al. (1985) study were below those used
by either Smith and Cabral (1980) or Lambrecht et al. (1983a,b). The
range of doses used by Smith and Cabral was given as 6-8 mg/kg/day and
those used by Lambrecht were 3-9 rug/kg bw/day. Those of Arnold were, at
most, between 1.5 and 2.0 mg/kg bw/day.
2. There were notable differences 1n the animals used: 1n the case of
Smith and Cabral the Hver tumor susceptible strain of Agus rat was
used, although tumors were also found with Wlstar rats. We do not have
full data on historical tumor Incidences 1n these animals to allow for
more detailed evaluation.
3. The conditions of the Smith and Cabral study and those of Lambrecht were
both different from the 2-generat1on study of Arnold. Differences 1n
sensitivity due to prenatal exposure may occur because of rapid cell
division and/or differences In xenoblotlc metabolism compared with older
animals. The dose received transplacentally and from nursing 1s also
uncertain.
4. The method of preparation of the hexachlorobenzene 1n the diet was
different 1n that both Smith and Cabral and Arnold used arachls oil and
corn oil as hexachlorobenzene solvents while Lambrecht did not use an
oil vehicle. Absorption characteristics are known to depend upon the
vehicles used.
5. The Sprague-Dawley animals used by Arnold may have more fat than those
used by Lambrecht as they were somewhat larger. Distribution Into
different tissue compartments, especially Into fat where 1t Is likely
the hexachlorobenzene 1s at least temporarily stored, Is Hkely to alter
the effective concentration 1n target tissues. In this regard the hexa-
chlorobenzene 1s known to concentrate In adrenal tissue; the degree of
such concentration may well vary with strain or diet of the host animals.
12-87
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In summary, orally administered hexachlorobenzene has Induced hepato-
cellular carcinoma In male Sprague-Dawley (S-0) rats as well as hepatomas 1n
female Agus and Wlstar rats and 1n S-0 rats of both sexes. At the lowest
dose used 1n any of the studies (40 ppm 1n the diet or 1.5 mg/kg/day), neo-
plastlc nodules were Induced 1n S-0 rats, whereas hepatocellular carcinomas
occurred 1n the same strain at a higher dose (4-5 mg/kg/day). Adrenal pheo-
chromocytoma was significantly elevated 1n two separate studies 1n female
S-0 rats. In the same strain one Investigator reported parathyroid tumors
and a different Investigator reported kidney tumors; neither of these
findings has been repeated by other authors. Table 12-28 summarizes this
Information.
12.3.5,5. OTHER STUDIES — In addition to the studies described on
hamsters, mice and rats there are a few studies which cover specific kinds
of tests other than lifetime exposure and examination of all potential
target tissues for tumorlgenlc or carcinogenic response.
One such study was that of Thelss et al. (1977) 1n which the experiment
was designed to detect only pulmonary tumors following 1.p. Injection of
organic chemicals found as contaminants of drinking water. In this assay
hexachlorobenzene was one of the chemicals tested. Strain A mice were given
three dosage levels of hexachlorobenzene with the top level as the MTD. A
total of 24 Injections over a period of 8 weeks were given to 20 mice/group.
The total doses received were 190, 480 and 960 mg/kg. The lungs were the
only organ examined and hexachlorobenzene did not Increase tumor Incidence
1n that organ. The study ran for 32 weeks. Although this assay has proved
useful In detecting some pulmonary carcinogens, 1t 1s not designed to detect
other tumors.
12-88
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TABLE 12-28
Qualitative Comparison of Tumor Development 1n Rats Following Hexachlorobenzene Administration 1n Different Studies
(NJ
1
00
<£>
Stra1n/Sex
Ag us /Female
Ml star/Female
Sprague-Dawley/
Male and female
Sprague-Dawley/
Male and female
f-j animals of
2-generatlon study
Dosage
(lowest dose that
produced tumor)
100 ppm (6-8 mg/kg bw/day)
prepared by dissolving In
oil and mixing oil with food
75 ppm (3-4 mg/kg bw/day)
prepared 1n feed sans oil
vehicle
40 ppm (0.3-1.5 mg/kg bw/day)
prepared \n oil and mixing
oil with food at weaning —
animals exposed 1_n utero and
during nursing
Liver
liver -cell tumor
(F)
liver -cell tumor
(F)
hepatocellular
carcinoma (M&F)
hepatoma (M&F)
neoplastlc Hver
nodules (F)
Kidney Adrenal
m m
m NA
renal cell pheochromo-
adenoma (M&F) cytoma (F)
cortical
adenoma (F)
not found pheochromo-
cytoma (F)
Parathyroid Reference
NA Smith and
Cabral, 1980
NA Smith and
Cabral, 1980
NA Lambrecht,
1983a,b
adenoma (M) Arnold, 1983
NA = It 1s not known whether or not these tissues were examined.
-------
In another study on beagle dogs (Gralla et al., 1977} 1n which hexa-
chlorobenzene was given 1n dally gelatin capsules to 30 animals of each sex/
dosage group the duration of the study was only 1 year. Although this 1s
not a long enough period of time for a carc1nogen1c1ty study 1n dogs, 1t 1s
of Interest to note that the doses of 100, 10, 1 and 0.1 mg/kg bw/day
produced a number of toxic manifestations 1n the liver Including bile duct
hyperplasla, hepatomegaly and liver necrosis. This study 1s more appro-
priately considered under chronic toxldty.
Finally, Perelra et al. (1982) designed a study to determine whether
hexachlorobenzene Increased Y-9lutamyltranspept1dase-pos1t1ve foci 1n
rats. These foci are believed to be preneoplastlc 1n the Hver. The assay
was designed to test Initiation/promotion 1n this case by employing dlethyl-
N-n1trosam1ne (DENA) as the Initiating agent and hexachlorobenzene as the
promoter. Unfortunately, there are some errors 1n reporting of the results
1n the published paper and some Important controls were not Included
(Perelra, 1983). We have not yet received a corrected manuscript.
12.3.5.6. QUANTITATIVE ESTIMATION — Among the six chlorinated ben-
zenes reviewed 1n this document for their carcinogenic potential, only hexa-
chlorobenzene provides sufficient data for a risk estimate. This quantita-
tive section deals with estimation of the unit risk for hexachlorobenzene as
a potential carcinogen 1n air and water, and with the potency of hexachloro-
benzene relative to other carcinogens that have been evaluated by the U.S.
EPA Carcinogen Assessment Group (CAG). The unit risk for an air or water
pollutant 1s defined as the lifetime cancer risk to humans from dally expo-
sure to a concentration of 1 yg/m3 of the pollutant In air by Inhala-
tion, or to a concentration of 1 yg/a, 1n water by Ingestlon.
12-90
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The unit risk estimate for hexachlorobenzene represents an extrapolation
below the dose range of experimental data. There 1s currently no solid
scientific basis for any mathematical extrapolation model that relates expo-
sure to cancer risk at the extremely low concentrations. Including the unit
concentration given above, that must be dealt with 1n evaluating environ-
mental hazards. For practical reasons the correspondingly low levels of
risk cannot be measured directly either by animal experiments or by eplde-
mlologic study. Low dose extrapolation must, therefore, be based on current
understanding of the mechanisms of cardnogenesls. At the present time the
dominant view of the carcinogenic process Involves the concept that most
cancer-causing agents also cause Irreversible damage to DNA. This position
is based 1n part on the fact that a very large proportion of agents that
cause cancer are also mutagenlc. There 1s reason to expect that the quantal
response that Is characteristic of mutagenesls 1s associated with a linear
(at low doses) non-threshold dose-response relationship. Indeed, there 1s
substantial evidence from mutagenlclty studies with both Ionizing radiation
and a wide variety of chemicals that this type of dose-response model 1s the
appropriate one to use. This 1s particularly true at the lower end of the
dose-response curve; at high doses there can be an upward curvature,
probably reflecting the effects of multistage processes on the mutagenlc
response. The linear non-threshold dose-response relationship 1s also
consistent with the relatively few ep1dem1olog1c studies of cancer responses
to specific agents that contain enough Information to make the evaluation
possible (e.g., radiation-Induced leukemia, breast and thyroid cancer, skin
cancer Induced by arsenic 1n drinking water, liver cancer Induced by
aflatoxlns In the diet). Some supporting evidence also exists from animal
experiments (e.g., the Initiation stage of the two-stage cardnogenesls
model 1n rat Uver and mouse skin).
12-91
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Because Its scientific basis, although limited, 1s the best of any of
the current mathematical extrapolation models, the non-threshold model,
which 1s linear at low doses, has been adopted as the primary basis for risk
extrapolation to low levels of the dose-response relationship. The risk
estimates made with such a model should be regarded as conservative, repre-
senting the most plausible upper limit for the risk (I.e., the true risk 1s
not Hkely to be higher than the estimate, but 1t could be lower).
For several reasons, the unit risk estimate based on animal bloassays 1s
only an approximate Indication of the absolute risk 1n populations exposed
to known carcinogen concentrations. First, there are Important species
differences 1n uptake, metabolism and organ distribution of carcinogens, as
well as species differences 1n target site susceptibility, Immunological
responses, hormone function, dietary factors and disease. Second, the con-
cept of equivalent doses for humans compared to animals on a mg/surface area
basis 1s virtually without experimental verification as regards carcinogenic
response. Finally, human populations are variable with respect to genetic
constitution and diet, living environment, activity patterns and other
cultural factors.
The unit risk estimate can give a rough Indication of the relative
potency of a given agent as compared with other carcinogens. Such estimates
are, of course, more reliable when the comparisons are based on studies In
which the test species, strain, sex and routes of exposure are similar.
The quantitative aspect of carcinogen risk assessment 1s addressed here
because of Its possible value In the regulatory decision-making process,
e.g., 1n setting regulatory priorities, evaluating the adequacy of technol-
ogy-based controls, etc. However, the Imprecision of presently available
technology for estimating cancer risks to humans at low levels of exposure
should be recognized. At best, the linear extrapolation model used here
12-92
-------
provides a rough but plausible estimate of the upper limit of risk — that
1s, with this model 1t 1s not likely that the true risk would be much more
than the estimated risk, but 1t could be considerably lower. The risk esti-
mates presented In subsequent sections should not be regarded, therefore, as
accurate representations of the true cancer risks even when the exposures
Involved are accurately defined. The estimates presented may, however, be
factored Into regulatory decisions to the extent that the concept of upper-
risk limits 1s found to be useful.
12.3.5.6.1. Procedures for the Determination of Unit Risk —
12.3.5.6.1.1. Low Dose Extrapolation Model. The mathematical formula-
tion chosen to describe the linear non-threshold dose-response relationship
at low doses 1s the linearized multistage model (Crump and Watson, 1979).
This model employs enough arbitrary constants to be able to fit almost any
monotonlcally Increasing dose-response data, and 1t Incorporates a procedure
for estimating the largest possible linear slope (1n the 955C confidence
limit sense) at low extrapolated doses that 1s consistent with the data at
all dose levels of the experiment.
Let P(d) represent the lifetime risk (probability) of cancer at dose d.
The multistage model has the form
P(d) = 1 - exp [~(qQ + Qjd + q2d* + ...+ qkdk)]
where
q1 > 0, 1 = 0, 1, 2, .... k
Equlvalently,
Pt(d) = 1 - exp [-(qid * q2d2 + ... qRdk)]
where
Pt(d) BP(d)-P(0)
1 1 - P(0)
Is the extra risk over background rate at dose d.
12-93
-------
The point estimate of the coefficients q., 1 = 0, 1, 2, ..., k» and
consequently, the extra risk function, P*.(d), at any given dose d, 1s
calculated by maximizing the likelihood function of the data.
The point estimate and the 95% upper confidence limit of the extra risk,
P.(d), are calculated by using the computer program, 6LOBAL79, developed
by Crump and Watson (1979). At low doses, upper 95% confidence limits on
the extra risk and lower 95% confidence limits on the dose producing a given
risk are determined from a 95% upper confidence limit, q,*, on parameter
q,. Whenever q, > 0, at low doses the extra risk Pt(d) has approxi-
mately the form Pt(d) = q-i* x d. Therefore, q,* x d 1s a 95% upper
confidence limit on the extra risk and R/q,* Is a 95% lower confidence
limit on the dose, producing an extra risk of R, Let LQ be the maximum
value of the log-likelihood function. The upper-limit q * 1s calculated
by Increasing q, to a value q,* such that when the log-likelihood 1s
remaxlmlzed subject to this fixed value q-,* for the linear coefficient,
the resulting maximum value of the log-likelihood L, satisfies the equation
2 (LQ - L^ = 2.70554
where 2.70554 1s the cumulative 90% point of the chl-square distribution
with one degree of freedom, which corresponds to a 95% upper-limit (one-
sided). This approach of computing the upper confidence limit for the extra
risk P,(d) 1s an Improvement on previous models. The upper confidence
limit for the extra risk calculated at low doses 1s always linear. This 1s
conceptually consistent with the linear non-threshold concept discussed
earlier. The slope, q,*, 1s taken as an upper-bound of the potency of the
chemical 1n Inducing cancer at low doses. [In the section calculating the
risk estimates, Pt(d) will be abbreviated as P.]
12-94
-------
In fitting the dose-response model, the number of terms In the poly-
nomial is chosen equal to (h-l)» where h 1s the number of dose groups 1n the
experiment, Including the control group.
Whenever the multistage model does not fit the data sufficiently well,
data at the highest dose 1s deleted and the model 1s refH to the rest of
the data. This Is continued until an acceptable fit to the data 1s
obtained. To determine whether or not a fit 1s acceptable, the ch1-square
statistic
h
the
1s calculated where N, 1s the number of animals 1n the 1 dose group,
the
X. 1s the number of animals 1n the 1 dose group with a tumor
the
response, P. 1s the probability of a response 1n the 1 dose group
estimated by fitting the multistage model to the data, and h 1s the number
of remaining groups. The fit 1s determined to be unacceptable whenever X2
1s larger than the cumulative 99% point of the ch1-square distribution with
f degrees of freedom, where f equals the number of dose groups minus the
number of non-zero multistage coefficients.
12.3.5.6.1.2. Selection of Data. For some chemicals, several studies
In different animal species, strains and sexes, each run at several doses
and different routes of exposure, are available. A choice must be made as
to which of the data sets from several studies to use In the model. It may
also be appropriate to correct for metabolism differences between species
and for absorption factors via different routes of administration. The
procedures used 1n evaluating these data are consistent with the approach of
making a maximum-likely risk estimate. They are as follows:
12-95
-------
1. The tumor Incidence data are separated according to organ sites or tumor
types. The set of data (I.e., dose and tumor Incidence) used 1n the
model 1s the set where the Incidence 1s statistically significantly
higher than the control for at least one test dose level and/or where
the tumor Incidence rate shows a statistically significant trend with
respect to dose level. The data set that gives the highest estimate of
the lifetime carcinogenic risk, q *, 1s selected 1n most cases.
However, efforts are made to exclude data sets that produce spuriously
high risk estimates because of a small number of animals. That 1s, If
two sets of data show a similar dose-response relationship, and one has
a very small sample size, the set of data having the larger sample size
1s selected for calculating the carcinogenic potency.
2. If there are two or more data sets of comparable size that are Identical
with respect to species, strain, sex and tumor sites, the geometric mean
of q,*, estimated from each of these data sets, 1s used for risk
assessment. The geometric mean of numbers A,, A?, ..., A 1s
defined as
ffi v & v Y S ^
^ r? «| ft t* rt
-------
equivalent exposure. In an animal experiment, this equivalent dose 1s
computed 1n the following manner.
Let
Le = duration of experiment
le = duration of exposure
m = average dose per day 1n mg during administration of the agent
(I.e., during le), and
W = average weight of the experimental animal
Then, the lifetime exposure 1s
d =
le x m
LP x W2/3
ORAL: Often exposures are not given 1n units of mg/day, and 1t
becomes necessary to convert the given exposures Into mg/day. Similarly, 1n
drinking water studies, exposure 1s expressed as ppm 1n the water. For
example, 1n most feeding studies exposure 1s given 1n terms of ppm 1n the
diet. In these cases, the exposure In mg/day 1s
m = ppm x F x r
where ppm 1s parts per million of the carcinogenic agent 1n the diet or
water, F 1s the weight of the food or water consumed per day 1n kg, and r Is
the absorption fraction. In the absence of any data to the contrary, r 1s
assumed to be equal to one. For a uniform diet, the weight of the food
consumed 1s proportional to the calories required, which 1n turn 1s propor-
tional to the surface area, or two-thirds power of the weight. Water
demands are also assumed to be proportional to the surface area, so that
2/3
m a ppm x W x r
or
12-97
-------
As a result, ppm 1n the diet or water 1s often assumed to be an equivalent
exposure between species. However, this 1s not Justified 1n dose extrapola-
tion of laboratory animals to humans, since the ratio of calories to food
weight 1s very different In the diet of man as compared to laboratory
animals, primarily due to differences In the moisture content of the foods
eaten. For the same reason, the amount of drinking water required by each
species also differs. It 1s therefore necessary to use an empirically-
derived factor, f = F/W, which Is the fraction of an organism's body weight
that 1s consumed per day as food, expressed as follows:
Fraction of Body Weight Consumed as
Species W ffood fwater
Man 70 0.028 0,029
Rats 0.35 0.05 0.078
Mice 0.03 0.13 0.17
Thus, when the exposure Is given as a certain dietary or water concentration
2/3
1n ppm, the exposure In mg/W 1s
f x W ___ .. c x wl/3
= ppm x 1
When exposure 1s given In terms of mg/kg/day = m/Wr = s, the conversion 1s
simply
_JL_ = s x wl/3.
rW2/3
INHALATION: When exposure 1s via Inhalation, the calculation of dose
can be considered for two cases where 1) the carcinogenic agent Is either a
completely water-soluble gas or an aerosol and 1s absorbed proportionally to
the amount of air breathed 1n, and 2) where the carcinogen 1s a poorly
water-soluble gas which reaches an equilibrium between the air breathed and
12-98
-------
the body compartments. After equilibrium 1s reached, the rate of absorption
of these agents 1s expected to be proportional to the metabolic rate, which
1n turn Is proportional to the rate of oxygen consumption, which 1n turn 1s
a function of surface area.
Case 1: Agents that are 1n the form of partlculate matter or virtu-
ally completely absorbed gases, such as sulfur dioxide, can reasonably be
expected to be absorbed proportionally to the breathing rate. In this case
the exposure 1n mg/day may be expressed as
m - I x v x r
where I = Inhalation rate per day In m3, v = mg/m3 of the agent 1n air,
and r = the absorption fraction.
The Inhalation rates, I, for various species can be calculated from the
observations of the Federation of American Societies for Experimental Biol-
ogy (FASEB, 1974} that 25 g mice breathe 34.5 si/day and 113 g rats breathe
105 a/day. For mice and rats of other weights, W (1n kg), the surface
area proportionality can be used to find breathing rates In m3/day as
follows:
?/*\
For mice, I = 0.0345 (W/0.025) ' mVday
o/-3
For rats, I = 0.105 (W/0.113) ' mVday
For humans, the value of 20 mVday* 1s adopted as a standard breathing
2/3
rate (ICRP, 1977). The equivalent exposure 1n mg/W for these agents
can be derived from the air Intake data 1n a way analogous to the food
*From "Recommendation of the International Commission on Radiological Pro-
section", page 9. The average breathing rate 1s TO7 cm3 per 8-hour
workday and 2xl07 cm3 in 24 hours.
12-99
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Intake data. The empirical factors for the air Intake/kg/day, 1 = I/H,
based upon the previously stated relationships, are tabulated as follows:
Species W 1 = I/H
Han 70 0,29
Rats 0.35 0.64
Mice 0.03 1.3
Therefore, for participates or completely absorbed gases, the equivalent
2/3
exposure 1n mg/W 1s
d = _ffl_ = Ivf_ = IWvL . 1wl/3vr
W2/3 W2/3 W2/3
In the absence of experimental Information or a sound theoretical argu-
ment to the contrary, the fraction absorbed, r, 1s assumed to be the same
for all species.
Case 2: The dose 1n mg/day of partially soluble vapors 1s proportlon-
?/l
al to the Op consumption, which 1n turn 1s proportional to W and 1s
also proportional to the solubility of the gas 1n body fluids, which can be
expressed as an absorption coefficient, r, for the gas. Therefore, express-
Ing the Op consumption as Op
dent of species, 1t follows that
2/3
1ng the 0? consumption as Op = k W , where k 1s a constant Indepen-
p/q
m = k W ' x v x r
or
. m ,
d = = kvr
H2/3
As with Case 1, 1n the absence of experimental Information or a sound theo-
retical argument to the contrary, the absorption fraction, r, Is assumed to
be the same for all species. Therefore, for these substances a certain
concentration 1n ppm or yg/m3 1n experimental animals 1s equivalent to
the same concentration 1n humans. This 1s supported by the observation that
12-100
-------
the minimum alveolar concentration necessary to produce a given "stage" of
anesthesia 1s similar 1n man and animals (Drlpps et a!., 1977). When the
animals are exposed via the oral route and human exposure 1s via Inhalation
or vice versa, the assumption Is made, unless there 1s pharmacoklnetlc
evidence to the contrary, that absorption 1s equal by either exposure route.
12.3.5.6.1.4. Calculation of the United Risk from Animal Studies. The
? f\
risk associated with d mg/kg /day 1s obtained from GLOBAL79 and, for
most cases of Interest to risk assessment, can be adequately approximated by
P(d) = 1 - exp (-q-d). A "unit risk" 1n units X 1s simply the risk
corresponding to an exposure of X = 1. This value 1s estimated simply by
2/3
finding the number of mg/kg /day that corresponds to one unit of X, and
substituting this value Into the above relationship. Thus, for example, If
X Is 1n units of Mg/m3 1n the air, then for Case 1, d = 0.29 x 701/3 x
2/3
10 3 mg/kg /day, and for Case 2, d = 1, when pg/m3 Is the unit
used to compute parameters In animal experiments.
If exposures are given 1n terms of ppm in air, the following calculation
may be used;
1 ppm = 1.2 x Molecular weight (gas) mg/m3
molecular weight (air)
Note that an equivalent method of calculating unit risk would be to use
the
mg/kg for the animal exposures, and then to Increase the j polynomial
coefficient by an amount
(Wh/wa)3/3 j = l, 2, .... k,
and to use mg/kg equivalents for the unit risk values.
ADJUSTMENTS FOR LESS THAN LIFESPAN DURATION OF EXPERIMENT: If the
duration of experiment L Is less than the natural Hfespan of the test
c
animal L, the slope q,*, or more generally the exponent g(d), Is Increased
by multiplying a factor (L/L )3. We assume that 1f the average dose d
12-101
-------
1s continued, the age-specific rate of cancer will continue to Increase as a
constant function of the background rate. The age-specific rates for humans
Increase at least by the third power of the age and often by a considerably
higher power, as demonstrated by Doll (1971). Thus, 1t 1s expected that the
cumulative tumor rate would Increase by at least the third power of age.
Using this fact, 1t 1s assumed that the slope q * or more generally the
exponent g(d), would also Increase by at least the third power of age. As a
result, 1f the slope q * [or g(d)] 1s calculated at age L , 1t 1s
expected that 1f the experiment had been continued for the full Hfespan L
at the given average exposure, the slope q * [or g(d)] would have been
Increased by at least (L/L )3.
\7
This adjustment 1s conceptually consistent with the proportional hazard
model proposed by Cox (1972) and the t1me-to-tumor model considered by
Daffer et al. (1980), where the probability of cancer by age t and at dose d
1s given by
P(d,t) = 1 - exp [-f(t) x g(d)].
12.3.5.6.2. Unit Risk Estimates —
12.3.5.6.2.1. Data Available for Potency Calculation. Hexachloro-
benzene has been shown to Induce tumors 1n hamsters, mice and rats. The
primary target organ appears to be the liver 1n all three of these species.
Liver haemang1oendothel1omas 1n hamsters and hepatocellular carcinomas 1n
rats were significantly Increased 1n the hexachlorobenzene-treated animals.
The potency estimate calculated on the basis of hepatocellular carcinomas 1n
female rats 1s used to derive unit risk estimates for hexachlorobenzene In
air and water. This particular tumor site 1s selected for calculating unit
risks because 1t 1s a malignant tumor In the primary target organ and
results 1n the highest potency estimate.
12-102
-------
Increased Incidences of thyroid, parathyroid, adrenal and kidney tumors
were also observed among these species. Fourteen data sets showing signifi-
cant tumor Incidences have been used herein to calculate the carcinogenic
potency of hexachlorobenzene. These calculations provide a range of esti-
mates that, 1n part, reflect the uncertainties Inherent 1n the risk assess-
ment process. Tables 12-29 through 12-32 summarize the data used to calcu-
late the potency of hexachlorobenzene. These data have been discussed and
evaluated elsewhere 1n this chapter.
12.3.5.6.2.2. Choice of Low-Dose Extrapolation. In addition to the
multistage model currently used by CAB for low-dose extrapolation, CAG also
uses three other models, the problt, the Welbull and the one-hit models, to
estimate the risks from exposure to hexachlorobenzene using the data for
hepatocellular carcinoma 1n female rats. These models cover almost the
entire spectrum of risk estimates that could be generated from the existing
mathematical extrapolation models. These models are generally statistical
In character, and are not derived from biological arguments, except for the
multistage model which has been used to support the somatic mutation
hypothesis of carcinogenesls (Armltage and Doll, 1954; Whlttemore, 1978;
Whlttemore and Keller, 1978). The main differences among these models 1s
the rate at which the response function, P(d), approaches zero or P{0) as
dose, d, decreases. For Instance, the problt model would usually predict a
smaller risk at low doses than the multistage model because of the differ-
ence of the decreasing rate 1n the low-dose region. However, It should be
noted that one could always artificially give the multistage model the same
(or even greater) rate of decrease as the problt model by making some dose
transformation and/or by assuming that some of the parameters 1n the multi-
stage model are zero. This, of course, 1s not reasonable without knowing, a
12-103
-------
TABLE 12-29
Tumor Incidences 1n Hale and Female Hamsters Given
Hexachlorobenzene 1n D1eta
Doseb
(mg/kg/day)
0
4
9
16
Thyroid
Hale
0/40
0/30
1/30
8/57
Hepatoma
Hale
0/40
14/30
26/30
49/57
F ema 1 e
0/30
14/30
17/30
51/60
Liver HemangloendotheHoma
Hale
0/40
1/30
6/30
20/57
F ema 1 e
0/39
0/30
2/30
7/60
aSource: Cabral et a!., 1977
the equivalent dose between humans and hamsters 1s assumed to be on
the basis of body surface, the dose 1n mg/kg/day 1s multiplied by a factor
(0.1/70P/3, where 70 and 0.1 kg are, respectively, the average body
weights of humans and hamsters.
12-104
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TABLE 12-30
Incidence of Liver Cell Tumors 1n Male and Female Swiss Mice
Given Hexachlorobenzene Diet3
Doseb Hale0 Female0
(mg/kg/day)
0 0/47 0/49
6 0/30 0/30
12 3/12 3/12
24 7/29 14/26
aSource: Cabral et al . , 1979
the equivalent dose between humans and mice is assumed to be on the
basis of body surface area, the dose in mg/kg/day is multiplied by a factor
(0.035/70)1/3, where 0.035 kg and 70 kg are, respectively, the average
body weights of mice and humans.
°The number of animals that survived at the first observed liver cell
tumor Is used as the denominator.
12-105
-------
TABLE 12-31
Liver and Kidney Tumor Incidence Rates 1n Male and Female
Sprague-Dawley Rats Given Hexaehlorobenzene 1n D1eta
Sex
Hale
Female
Doseb
(mg/kg/day)
0
4.24
8.48
0
4.67
9.34
Hepatocellular
Carcinoma
0/54
3/52
4/56
0/52
36/56
48/55
Hepatoma
0/54
10/52
11/56
0/52
26/56
35/55
Renal Cell
Adenoma
7/54
41/52
42/56
1/52
7/56
15/54
aSource: Lambrecht, 1983a,b. Additional data from this study on adrenal
pheochromocytoma has recently become available (Peters et a!., 1983, summa-
rized 1n Table 12-24) but was not available when quantitative estimates
were made.
Nhe dosages are calculated by the Investigator based on the average food
consumption of 22.6 g/rat/day and an average body weight of 400 g for male
rats. For female rats, the average food consumption Is 16.5 g/rat/day and
the average body weight 1s 265 g. If the equivalent dose between humans and
mice 1s assumed to be on the basis of body surface area, the dose presented
1n the table Is multiplied by a factor (Wa/70}1/3, where Wa 1s the
body weight of male or female rats, and 70 kg 1s the human body weight.
12-106
-------
TABLE 12-32
Incidence Rate of Adrenal Pheochromocytoma 1n Female Sprague-Dawley
Rats (F-| generation) 1n a 2-Generat1on Feeding Study
Dosea
(mg/kg/day)
0
0.02
0.08
0.40
1.90
Incidence Rate^
(used In calculations)
2/48
4/50
4/50
5/49
17/49
Revised Incidence Ratec
2/49
4/49
4/49
alf the equivalent dose between humans and rats 1s assumed to be on the
basis of body surface, the dose 1n this table 1s multiplied by a factor
(0.35/70)1/3^ where 70 kg and 0.35 kg are, respectively, assumed to be
the body weight of humans and rats.
bSource: Arnold et al., 1985
cSource: Arnold, 1984. The amended 1984 data are presented 1n Table
12-26, but were not available when quantitative estimates were made.
12-107
-------
priori, what the carcinogenic process for the agent 1s. Although the multi-
stage model appears to be the most reasonable or at least the most general
model to use, the point estimate generated from this model 1s of limited
value because 1t does not help to determine the shape of the dose-response
curve beyond experimental exposure levels, furthermore, point estimates at
low doses extrapolated beyond experimental doses could be extremely unstable
and could differ drastically, depending on the amount of the lowest experi-
mental dose. Since upper-bound estimates from the multistage model at low
doses are relatively more stable than point estimates, 1t 1s suggested that
the upper-bound estimate for the risk (or the lower-bound estimate for the
dose) be used 1n evaluating the carcinogenic potency of a suspect carcino-
gen. The upper-bound estimate can be taken as a plausible estimate 1f the
true dose-response curve 1s actually linear at low doses. The upper-bound
estimate means that the risks are not likely to be higher, but could be
lower 1f the compound has a concave upward dose-response curve or a thresh-
old at low doses. Another reason one can, at best, obtain an upper-bound
estimate of the risk when animal data are used 1s that the estimated risk 1s
a probability conditional to the assumption that an animal carcinogen 1s
also a human carcinogen. Therefore, 1n reality, the actual risk could range
from a value near zero to an upper-bound estimate.
12.3.5.6.2.3. Calculation of the Carcinogenic Potency of Hexachloroben-
zene. Fourteen sets of tumor Incidences which show significant Increases
(see Tables 12-29 through 12-32) are used herein to calculate the carcino-
genic potency of hexachlorobenzene. Since preparing these calculations
additional data from the Lambrecht et al. (1983a,b) study (adrenal pheo-
chromocytoma) and from the Arnold et al. (1985) study (neoplastlc liver
nodules) have become available. Quantitative estimates have not been made
12-108
-------
using this data. Using the multistage model for low-dose extrapolation, as
shown 1n Table 12-33, the potency estimates calculated on the basis of these
data sets are approximately within an order of magnitude from each other,
with the exception of the thyroid tumor. These potencies provide a range of
estimates that reflects the uncertainties stemming from the differences in
species, tumor sites, solvent vehicles and composition of diet. The range
does not reflect uncertainty resulting from the use of different extrapola-
tion models.
To calculate the unit risks of hexachlorobenzene 1n air and water, CAG
used an estimate of carcinogenic potency based upon the data for hepatocel-
lular carcinoma 1n female rats. For comparison, three additional low-dose
extrapolation models, the problt, the Welbull and the one-hit models, are
also used to provide risk estimates at dose levels 0.01, 0.1 and 1 mg/kg/
day. These results are presented 1n Table 12-34. The maximum likelihood
estimate of the parameters for all four models are presented 1n Table A-l 1n
the Appendix. At 1 mg/kg/day, all four models predict comparable risks. At
lower doses, the multistage model predicts a higher risk than the probit
model, but a lower risk than the Welbull model.
12.3.5.6.2.4. Risk Associated with 1 pg/a, of Hexachlorobenzene 1n
Drinking Water. Under the assumption that dally water consumption for a 70
kg person 1s 2 a, the hexachlorobenzene Intake 1n terms of mg/kg/day 1s
d = (28,/day) x (lyg/a) x (lO'" mg/pg) x (1/70 kg) = 2.86 x 10~5 mg/kg/day.
Therefore, the risk from drinking water containing 1 pg/8, of hexachloro-
benzene 1s estimated to be
P = 1.7 x 2.86 x 10~5 = 4.9 x 10~5.
12-109
-------
TABLE 12-33
The Carcinogenic Potency3 of Hexachlorobenzene, Calculated on the Basis of 14 Data Sets,b
Using the Linearized Multistage Model
Dose 1s Assumed to be
Equivalent on the Basis of
Study Date Base Reference
Body Weight Surface Area
Hamster
Thyroid (male)
Hepatoma:
Male
Female
9.3 x 10~3
1.9 x lO'i
1.5 x IQ-i
8.3 x 10~2
1.7
1.3
Cabral
et al.,
1977
Hemangloendothel1oma:
Male 3.2 x 1(T2 2.8 x NT1
Female 1.1 x 10~2 1.0 x 10"1
Mice Liver cell: Cabral
Male 1.7 x 10~2 2.1 x NT1 et al., 1979
Female 1.4 x 10~2 1.8 x KT1
-------
TABLE 12-33 (cont.)
Study
Date Base
Dose 1s
Equivalent
Assumed to be
on the Basis of
Body Weight
Surface Area
Reference
rvs
i
Rats
Rats
2-generat1on
study
Renal cell:
Hale
Female
Hepatocellular carcinoma;
Hale
Female
Hepatoma:
Hale
Female
Adrenal
Pheochromocytoma
(female)
2.5 x ID"1
4.2 x 10"2
1.8 x 10~2
2.7 x KT1
4.7 x 10~2
1.5 x KT1
2.8 x 10"1
1.4
2.6 x 10"1
1.0 x 10"1
1.7
2.6 x 10"1
9.0 x NT1
1.6
Lambrecht,
1983
Arnold
et al., 1985
aq-|* (mg/kg/day)'1 1s the 95% upper confidence limit of the linear component 1n the multistage model.
bS1nce preparing these calculations, additional data from Lambrecht et al. (1983a,b) study (adrenal
pheochromocytoma) and from Arnold et al. (1985) study (neoplastlc liver nodules) has become available.
These data have not been evaluated.
-------
TABLE 12-34
Upper-Bound3 (Point) Estimation of Risk,
Based on Hepatocellular Carcinoma 1n Female Ratsb
Assumption of
Human Equivalent
Dose
On the basis of
body weight
On the basis of
surface area
Models
multistage
problt
Welbull
one-hit
multistage
problt
Welbull
one-hit
Risk at
0.01
2.7 x 10~3
(2.2 x 10~3)
3.6 x 10"9
(1.3 x 10~i0)
1.2 x 10~2
(2.5 x 10~3)
2.7 x 10~3
(2.2 x 10"3)
1.7 x 10~2
(1.4 x 10~2)
6.2 x 1Q~5
(4.1 x 10~6)
5.0 x 10"2
(1.3 x 10~2)
1.7 x 10~2
(1.4 x 10~2)
Dose Level (mg/kg/day)
0.1
2.7 x 10~2
(2.2 x 10~2)
1.0 x 10"3
(8.9 x 10"*)
8.4 x 10"2
(2.5 x 10~2)
2.7 x 10~2
(2.2 x 10~2)
1.7 x ID"*
(1.3 x 10~»)
1.3 x 10"i
(2.9 x 10~2}
2.9 x 10'1
(1.3 x ID'1)
1.7 X 10~1
(1.3 x 10-*)
1.00
2.4 x 10"1
(2.0 x 10'1
3.4 x 10'1
(1.2 x 10"1
4.3 x 10"1
(2.2 x 10~l
2.4 x 10"1
(2.0 x 10"1
B.OxlO'1
(7.4 x 10'1
8.2 x lO'i
(7.5 x 10"1
8.1 x 10-i
(7.4 x 10"1
8.0xlO"i
(7.4 x 10""1
)
)
)
]
>
)
)
)
a95% upper confidence limit
bSource: Lambrecht, 1983
12-112
-------
This calculation uses the carcinogenic potency q-,* = 1.7/(mg/kg/day),
based on the data on hepatocellular carcinomas 1n female rats, assuming that
dose per surface area 1s equivalent between rats and humans. If the equiva-
lent dose 1s assumed to be on the basis of body weight, the unit risk, P,
would be reduced to 7.6xlO~6.
12.3.5.6.2.5. Risk Associated with 1 yg/m3 of Hexachlorobenzene 1n
A1r. Since no Inhalation study has been performed on hexachlorobenzene,
the risk from Inhalation exposure can only be estimated by using the carci-
nogenic potency, q,* = 1.7/{mg/kg/day), as calculated from the dietary
study referred to elsewhere 1n this chapter. The assumption 1s made that
the hexachlorobenzene absorption rate is the same whether exposure 1s via
the oral or the Inhalation route.
Assuming the volumetric breathing rate of 20 mVday for a 70 kg
person, the rate in mg/kg/day corresponding to 1 yg/m3 hexachlorobenzene
1n air 1s
d = (20 mVday) x (10~3 mg/Vg) x (1/70 kg) = 2,86 x 10~« mg/kg/day.
Therefore, the risk due to inhaling air contaminated with 1 pg/m3 hexa-
chlorobenzene 1s
P = 1.7 x 2.86 x 10~4 = 4.9 x 10~4.
This estimation is based on the assumption that dose per surface area 1s
equivalent between humans and rats. If dose per body weight is assumed to
be equivalent, the unit risk would be reduced to 7.6xlO~5.
12.3.5.6.3. Comparison of Potency with Other Compounds — One of the
uses of quantitative potency estimates 1s to compare the relative potency of
carcinogens, figure 12-1 is a histogram representing the frequency distri-
bution of potency Indices for 54 suspect carcinogens evaluated by CAG. The
actual data summarized by the histogram are presented in Table 12-35.
12-113
-------
18
16
14
>- 12
o
z
UJ
§ 10
UJ
n:
"" 8
6
4
2
4th
QUARTILE
3rd
QUARTILE
2nd
1st
QUARTILE QUARTILE
V7
1 x 1Q*1 4 x 1Q*2 2 x 10*3
-2
246
LOG OF POTENCY INDEX
6
FIGURE 12-1
Histogram Representing the Frequency Distribution of the Potency Indices
of 54 Suspect Carcinogens Evaluated by the Carcinogen Assessment Group
12-114
-------
TABLE 12-35
Relative Carcinogenic Potencies Among 54 Chemicals Evaluated by the Carcinogen Assessment Group
as Suspect Human Cardnogensa'k«c
IV
I
Compounds
AcrylonHMle
Aflatoxin B,
I
AldMn
Allyl chloride
Arsenic
8[a]P
Benzene
Benzldene
Beryllium
Cadmium
Carbon tetrachlorlde
Chlordane
Chlorinated ethanes
l,2-D1chloroethane
Hexachloroethane
1 ,1 ,2,2-Tetrachloroethane
1..1 ,2-Tr1chloroethane
Chloroform
Chromium
Slope
(mg/kg/day)'1
0.24 (W)
2924
11.4
1.19 x 10~2
15 (H)
11.5
5.2 x 10~2 (W)
234 (W)
2.6
7.8 (W)
1.30 x TO"1
1.61
6.9 x 10~2
1.42 x 10~2
0.20
5.73 x ID"2
7 x 10~2
41 (U)
Molecular
Weight
53.1
312.3
369.4
76.5
149.8
252.3
78
184.2
9
112.4
153.8
409.8
98.9
236.7
167.9
133.4
119.4
100
Potency
Index
IxlQ*1
9xlO*5
4xlO+3
9x10'*
2xlO*3
3xlQ*3
4x10°
4xlQ*4
xl
2x10
+2
9x10
2X10+1
7xlO+2
7x10°
3x10°
3xlO+1
8x10°
8x10°
4xlO+3
Order of
Magnitude
(logio Index)
+1
+6
+4
0
+3
+3
+1
+5
+1
+3
+ 1
+3
+1
0
+ 1
+ 1
+1
+4
-------
TABLE 12-35 (cent.)
I
—1
««1
-------
TABLE 12-35 (cont.)
ro
I
Compounds
NHrosamlnes
Dime thy Inltrosamlne
D1ethyln1trosatn1ne
D1butyln1trosam1ne
N-n1trosopyrrol1d1ne
N-n1troso-N-ethylurea
N-n1 tr oso-N-methylurea
N-n1troso-d1phenylam1ne
PCBs
Phenols
2,4,6-TMchlorophenol
Tetrachlorod1benzo-p-d1ox1n
Tetrachloroethylene
Toxaphene
Trlchloroethylene
Vinyl chloride
Slope
(mg/kg/day)"1
25.9 (not by q-\*)
43.5 (not by qi*)
5.43
2.13
32.9
302.6
4.92 x 10~3
4.34
1.99 x 10~2
1.56 x 10s
3.5 x 10"2
1.13
1.9 x 10"2
1.75 x 10~2 (I)
Molecular
Weight
74.1
102.1
158.2
100.2
117.1
103.1
198
324
197.4
322
165.8
414
131.4
62.5
Potency
Index
2x10+3
4x10+3
9x10+2
2x10+2
4xlO+3
3xlO+4
1x10°
IxlO*3
4x10°
5x1 O*7
6x10°
5X10+2
2.5x10°
1x10°
Order of
Magnitude
(logio Index)
+3
+4
+3
+2
-1-4
+4
0
+3
+1
+8
+1
+3
0
0
3An1mal slopes are 95% upper-limit slopes based on the linearized multistage model. They are calculated
based on animal oral studies, except for those Indicated by I (animal Inhalation), W (human occupational
exposure, and H (human drinking water exposure). Human slopes are point estimates based on the linear
non-threshold model.
bThe potency Index 1s
slopes 1n (mg/kg/day)
a rounded-off slope 1n (mMol/kg/day) 3
1 by the molecular weight of the compound.
and 1s calculated by multiplying the
cNot all of the carcinogenic potencies presented 1n this table represent the same degree of certainty.
All are subject to change as new evidence becomes available.
-------
The potency Index 1s derived from q,*, the 95% upper bound of the linear
component 1n the multistage model, and 1s expressed 1n terms of (mHol/kg/
day)"1. Where no human data were available, animal oral studies were used
1n preference to animal Inhalation studies, since oral studies have consti-
tuted the majority of animal studies.
Based on data concerning hepatocellular carcinomas 1n female rats, the
potency Index for hexachlorobenzene has been calculated as SxlQ2. This
figure 1s derived by multiplying the slope q * = 1.7/(mg/kg/day) and the
molecular weight of hexachlorobenzene, 284.4. This places the potency Index
for hexachlorobenzene 1n the second quartHe of the 54 suspect carcinogens
evaluated by CA6.
The ranking of relative potency Indices 1s subject to the uncertainties
Involved 1n comparing a number of potency estimates for different chemicals
based on varying routes of exposure 1n different species by means of data
from studies whose quality varies widely. All of the Indices presented are
based on estimates of low-dose risk, using linear extrapolation from the
observational range. These Indices may not be appropriate for the compari-
son of potencies 1f linearity does not exist at the low-dose range, or 1f
comparison 1s to be made at the high-dose range. If the latter 1s the case,
then an Index other than the one calculated above may be more appropriate.
12.3.5.6.4. Summary of Quantitative Estimation — Data on hepatocellu-
lar carcinomas 1n female rats after oral 1ngest1on have been used to esti-
mate the carcinogenic potency of hexachlorobenzene and the risks associated
with one unit of the compound 1n drinking water and air. The upper bound
cancer risks associated with 1 yg/st of hexachlorobenzene 1n drinking
water and 1 yg/m3 of hexachlorobenzene 1n air are estimated to be
5xlO~s and 5xlO~4, respectively. These estimates are calculated on the
12-118
-------
basis of the assumption that dose per surface area 1s equivalent among
species. If the dose 1s assumed to be equivalent on the basis of body
weight, the corresponding risk would be reduced approximately by a factor of
6. The carcinogenic potencies of hexachlorobenzene are also estimated on
the basis of 13 other data sets, encompassing different tumor sites and
animal species. Except for the case of thyroid tumors, these potency esti-
mates differ from each other within a single order of magnitude. The range
of the estimates reflects the uncertainties due to differences 1n species,
tumor sites, solvent vehicles, composition of diet, etc.
12.3.5.7. CARCINOGENICITY SUMMARY — In a lifetime study of hexa-
chlorobenzene administration to hamsters, hepatoma was Induced 1n both males
and females. The response at a dose of 4-5 mg/kg/day dissolved 1n corn oil
and mixed 1n the feed was 47% for both sexes and controls had no hepatomas.
In addition to hepatomas, hamsters responded to hexachlorobenzene treatment
with malignant liver haemangloendotheHoma and thyroid adenoma. The Inci-
dence of haemang1oendothel1oma was 20% 1n males (versus 0 1n controls) at 8
mg/kg/day and 12% 1n females (versus 0 1n controls) at 16 mg/kg/day. The
thyroid adenoma occurred at 14% Incidence 1n males treated with 16 mg/kg
hexachlorobenzene (versus 0 1n controls).
Liver cell tumors, described as hepatomas, were also produced 1n both
sexes 1n Swiss mice. At 24 mg/kg/day the Incidence was 34% for females and
16% for males and the response showed a dose-dependency not only 1n the
number of tumor-bearing animals but also 1n the latent period, multiplicity
and size of tumors. In ICR mice, hexachlorobenzene administered concur-
rently with polychlorlnated terphenyl Induced hepatocellular carcinomas.
In rats target organs for hexachlorobenzene-lnduced tumors Included
liver, kidney, adrenal gland and parathyroid gland 1n various studies.
Liver tumors were found 1n three studies which Included three different
12-119
-------
strains of rat: Agus (a liver tumor sensitive strain), Wlstar and Sprague-
Dawley rats. These tumors were Induced with doses between 1.5 and 8 mg/kg/
day. The Incidence was as high as 100% 1n Agus rats but lower for the other
strains. Renal cell tumors were found 1n one study on Sprague-Oawley rats.
In two studies on Sprague-Oawley rats, significant Increases 1n adrenal
pheochromocytoma 1n females were found. In one of these studies the Inci-
dence of parathyroid tumors 1n males was significantly Increased as well.
Table 12-36 summarizes the tumor data for hamsters, mice and rats for
hexachlorobenzene experiments.
The data on hexachlorobenzene provide sufficient evidence of the card-
nogenlclty and tumor1gen1c1ty of hexachlorobenzene since there were In-
creased Incidences of malignant tumors of the liver 1n two species (haeman-
g1oendothel1oma 1n hamsters and hepatocellular carcinoma 1n rats) as well as
reports of hepatoma 1n mice, rats and hamsters.
The appearance of thyroid tumors 1n hamsters and adrenal pheochromocyto-
mas and parathyroid tumors 1n rats as a result of hexachlorobenzene exposure
1s particularly Interesting because of the clinical association of adrenal
pheochromocytomas with parathyroid and thyroid tumors In humans {Fraumenl,
1974; H111, 1974), and because follow-up of Individuals 1n Turkey, who were
accidentally exposed to hexachlorobenzene over 25 years ago, shows a marked
Increase 1n enlarged thyroids. Only a few of these subjects have had their
thyroids examined h1stolog1cally and the pathology reports are not yet
available.
If the IARC criteria for the classification of carcinogens were used,
this animal evidence would be considered "sufficient." In the absence of
human evidence of carc1nogen1c1ty, hexachlorobenzene would be classed 1n
IARC category 28, meaning that 1t has been demonstrated to be carcinogenic
in animals and 1s probably carcinogenic 1n humans.
12-120
-------
TABLE 12-36
Significantly Increased Incidence of Tumors 1n Animals Given Hexachlorobenzene 1n Diet
ro
i
X Treated/X Control
Animal
(strain)
Hamsters
Hamsters
Nice
Rats
(S.D.)
Rats
(S.D.)
Rats
(S.D.)
Rats
(Wlstar)
Rats
(Agus)
Rats
(S.D.)
Rats
(S.D.)
Rats
(S.D.)
Rats
(S.D.)
Hamsters
Organ
liver
liver
liver
liver
liver
liver
liver
liver
adrenal
adrenal
kidney
parathyroid
thyroid
Tumor
hepatoma
haemang1oendothel1oma
hepatoma
neoplastlc nodules
hepatoma
hepatocellular
carcinoma
hepatoma
hepatoma
pheochromocytoma
pheochromocytoma
renal cell adenoma
adenoma
adenoma
Hales
47/0
20/0
16/0
NS
19/0
NS
NS
NS
79/13
25/4
14/0
Females
47/0
12/0
34/0
20/0
46/0
64/0
67/0
100/0
35/4
91/14
13/2
NS
Lowest Dose
to Produce Tumor
(mg/kg bw/day)
4
8 1n males
16 1n females
24
1.5
4-5
4-5
6-8
6-8
1.5
4-5
4-5
1.5
16
Reference
Cabral, 1977
Cabral, 1977
Cabral, 1979
Arnold, 1983
Lambrecht
et al., 1983a
Lambrecht
et al., 1983a
Smith and
Cabral, 1980
Smith and
Cabral, 1980
Arnold, 1983,
1984
Peters et al.,
1983
Lambrecht
et al., 1983b
Arnold, 1983
Cabral, 1977
NS = Not stated
-------
A quantitative estimate of the carcinogenic potency of hexachlorobenzene
was made from data on the hepatocellular response 1n female rats. The unit
risk estimate for human exposure to 1 yg/m3 1n air 1s 5xlO~4 and for 1
pg/j, of drinking water Is 4.9xlO~s. The upper-bound slope of the
dose-response curve, q,*, 1s 1.7/{mg/kg/day), giving a potency Index which
is 1n the second quartlle of 54 suspect carcinogens evaluated by the
Carcinogen Assessment Group of the U.S. EPA. Corresponding estimates from
13 other data sets, encompassing different tumor sites and animal species,
fall within a factor of 10 of these estimates except for thyroid tumors 1n
hamsters, which give estimates of about 1/20 of the potency based on the rat
hepatocellular carcinoma response.
12.3.6. Reproductive and Teratogenlc Effects. Hexachlorobenzene has been
shown to cross the placenta Into fetal tissues and to be present 1n the milk
of nursing dams (see Section 12.1.2.). The NOEL 1n a 4-generat1on reproduc-
tion study with rats was reported to be 20 ppm of hexachlorobenzene 1n the
diet. Pups from treated dams (receiving diets containing 80 ppm hexachloro-
benzene} recovered from elevated liver weights when nursed by foster dams.
Hepatomegaly and reduced survival was reported 1n kittens from cats receiv-
ing 263 ppm of hexachlorobenzene 1n their diets. Infant rhesus monkeys
developed clinical signs of toxldty, but hlstologlc examination showed only
mild effects. Fetal mice from dams treated with 100 mg/kg/day during days
7-16 of gestation exhibited teratogenlc abnormalities.
Results from a 4-generat1on reproduction study with Sprague-Dawley rats
was reported by Grant et al. (1977). Weanling rats, 1n groups of 20 females
and 10 males, were fed diets containing 0, 10, 20, 40, 80, 160, 320 or 640
yg hexachlorobenzene/g and at 100 days of age the FO generation was
mated to produce the F, generation. The F, pups were weaned at 21
12-122
-------
days, and the Fn rats were rested for 14 days and again mated to produce
the second Utter, F,. animals. The F,. animals were then used to
Ib Ib
produce the next generation, and this sequence was followed to the F-.
generation. The two highest doses (320 and 640 yg/g) were toxic to the
mothers and resulted 1n 20 and 50% mortality, respectively, before the first
whelping and 25% 1n each high dose group before the second whelping. In
addition, the fertility Index 1n these rats was greatly reduced 1n these two
dose groups and the average Utter size was decreased 1n the F1K, F0
ID e.3,
and F?, generations. The pups exhibited no gross abnormalities, but there
was an Increased number of stillbirths and all pups born alive died within 5
days In the 320 and 640 pg/g diet groups.
At the 160 yg/g level, 55% of the pups survived to day 5 but survival
to day 21 was greatly reduced. The number of live births and survival was
normal for the first two generations at the 80 yg/g dietary level, but by
the third generation there were stillbirths and a low degree of postnatal
v1ab1l1lty. In addition, birth and weanling body weights were consistently
less than those of the control group. At 40 yg/g diet only the liver
weights of the 21-day-old pups were significantly Increased, while the
kidney, heart and brain weights were not affected. Tissue concentrations of
hexachlorobenzene were dose-related, with body fat having the highest
concentration. The NOEL was reported to be 20 ppm in the diet.
The effect of hexachlorobenzene on rat reproduction was also reported by
K1tch1n et al. (1982). Female Sprague-Dawley rats (10 animals/treatment
group) were fed diets containing 0, 60, 80, 100, 120 and 140 yg hexa-
chlorobenzene/g of diet. The females were mated with untreated males after
96 days and then bred a second time 12 days after weaning of the F,
I a
Utter. Fertility and fecundity of treated females were not affected by
12-123
-------
treatment; however, a dose-related 21-day Increase In mortality was observed
1n both Utters and the LD,-n values were determined to be 100 and 140
3U
(maternal dietary concentration) for the F,_ and F1K generation,
I a ID
respectively.
Hendoza et al. (1978) studied the effects of hexachlorobenzene on
preweanllng Wlstar rats after a reciprocal transfer between 5 treated and 5
control dams. A significant Increase 1n the liver weight over that of the
control was observed 1n pups nursed by dams fed diets containing 80 pg
hexachlorobenzene/g for 2 weeks before mating until birth, but this effect
did not persist after the treated pups were transferred to a control foster
dam. Similarly, the pups nursed by treated dams had smaller brains, hearts,
kidneys and spleens than the controls, and these organs were larger 1n
treated pups nursed by control dams. The authors concluded that hexachloro-
benzene transmission via the milk had greater effects on the pups than
transmission via the placenta.
Hendoza et al. (1979) placed female Wlstar rats on diets containing 80
pg hexachlorobenzene/g beginning 2 weeks before mating until 35-36 days
after weaning. Results Indicated that there were no marked differences 1n
the external appearance, body weight, liver weight, gestation, or neonatal
survival between the hexachlorobenzene treated and control females. In
addition, there were no differences 1n the number of Utters, average number
of pups/Utter, average number of pups at birth and gestation Index.
Hansen et al. (1979) studied the effects of hexachlorobenzene on repro-
duction 1n cats fed contaminated pork cakes for 142 days. These cakes con-
tained 90i51 pg hexachlorobenzene/g, equivalent to an Intake of 3 mg/day/
cat, and were obtained from gilts fed diets containing 100 pg hexachloro-
benzene/g for 6-8 weeks before slaughter. The positive and untreated
12-124
-------
control groups received pork cakes from gilts fed diets that did not contain
hexachlorobenzene, with the positive control group receiving hexachloro-
benzene-splked cakes {263^120 yg/g equivalent to 8.7 mg/day/cat). These
females were mated with untreated males and the resulting kittens did not
receive hexachlorobenzene-contalning cakes. Effects on survival were noted
1n kittens born to only those cats receiving hexachlorobenzene-splked cakes
and was apparently due to the kittens being too weak to survive the stress
of weaning. There was a tendency for reduced average Utter sizes and
Increased mortality of nursing positive control kittens, and statistically
significant hepatomegaly and reduction in positive control kitten survival
at weaning. Treated positive control females exhibited a net weight loss
and increased susceptibility to disease but no changes 1n relative organ
weights, hematologlc parameters, or fecal coproporphyMn excretion.
Rush et al. (1983) fed adult male and female standard dark minks
(Mustela vision) diets containing 0, 1 or 5 ppm hexachlorobenzene and then
mated the males to the females In each of the respective study groups. The
resulting mink kits were fed their parents respective diets after weaning
from their mothers. The effects of exposures to hexachlorobenzene in_ utero
and from nursing milk resulted 1n Increased mortality In the hexachloro-
benzene-treated weanlings with mortality 1n the 0, 1 and 5 ppm groups being
8.2, 44.1 and 77.454, respectively. The surviving kits from all three groups
had no observed alterations in whole body, kidney or liver weights and no
observed damage to the kidneys or livers at 17 weeks of age. Induction of
hepatic mixed-function oxldases was observed 1n the surviving hexachloro-
benzene-exposed kits without any observable frank hepatotoxldty.
12-125
-------
Bailey et al. (1980) studied the transfer of hexachlorobenzene to three
nursing Infant rhesus monkeys from three lactatlng mothers receiving by
gavage 64 mg/kg/day of hexachlorobenzene suspended 1n methyl cellulose for
60 days. The hexachlorobenzene concentrated 1n the mothers' milk ranged
from 7.51-186 ppm during the dosing schedule. One Infant, by day 22, had
developed symptoms of hypoactlvlty and lethargy which progressed to ataxla
and death 1 week later. Autopsy revealed severely congested lungs. A
second Infant died on day 38 and autopsy revealed a subdural hematoma and
bilateral hemorrhaglc pneumonia. This Indicated that the risk of exposure
to nursing Infants was greater than the risk to their mothers. Blood
(0.42-49.44 ppm) and tissue levels 1n the Infants were higher than 1n their
mothers (0.41-16.16 ppm blood), and the Infants developed clinical symptoms
of toxldty while the mothers were asymptomatic.
Studies on the placental transfer of hexachlorobenzene 1n Wistar rats
and New Zealand rabbits did not reveal any apparent adverse effects on fetal
development. The female rats were dosed dally with 5, 10, 20, 40 or 80
mg/kg from day 6-16 of gestation, whereas the rabbits were treated with 0,
0.1, 1.0 or 10 mg/kg from day 1-27 of gestation. The compound was dissolved
1n corn oil and administered by means of a stomach tube (Vllleneuve et al.,
1974; Vllleneuve and H1erl1hy, 1975).
Khera (1974) conducted a teratogenlclty study with groups of 7-16 female
Wistar rats given single oral doses of 0, 10, 20, 40, 60, 80 or 120 mg
hexachlorobenzene/kg suspended 1n corn oil or 0.25% aqueous gum tragacanth
during gestation days 6-21. Maternal toxldty and reduction 1n fetal
weights resulted from the two higher doses. Maternal toxldty was charac-
terized by loss 1n body weight, hyperesthesla, tremors and convulsions. A
significant Increase In the Incidence of unilateral and bilateral 14th Mb
12-126
-------
was observed and was related to the duration of treatment (days 10-13, 6-16
or 6-21 of gestation) and the dose. Sternal defects were observed 1n only 1
of 4 experiments, which lead the authors to conclude that 1t was doubtful
that hexachlorobenzene caused the observed sternal defects. There were no
hexachlorobenzene-related effects on external morphology. Visceral abnor-
malities were not observed, and microscopic examinations did not reveal any
treatment-related change 1n the histology of the fetuses. Values for live
and dead fetuses, resorptlon sites, and fetal weight were within the control
limits.
Courtney et al. (1976) studied the effects of 1ngest1on of 100 mg/kg/day
hexachlorobenzene on days 7-16 of gestation In 10 pregnant CD-I mice. This
study was undertaken to evaluate the possibility that hexachlorobenzene
could be responsible for fetal malformations seen 1n pregnant animals
exposed to hexachlorobenzene-contamlnated pentachloronltrobenzene. The
results showed that the hexachlorobenzene-treated mice had significantly
Increased maternal I1ver-to-body weight ratios and decreased fetal body
weights. Also, a significant Increase 1n the Incidence of abnormal fetuses
per Utter were observed as compared to control mice. The abnormalities
that were observed 1n these affected fetuses were cleft palates, one
straight leg, small kidneys, one renal agenesis, and enlarged renal pelvis.
They concluded from this study that the teratogenlc activity of contaminated
pentachloronltrobenzene was probably due to hexachlorobenzene.
12.4. INTERACTIONS
Certain chemicals have been shown to alter the toxldty and pharmaco-
klnetlcs of hexachlorobenzene 1n mammals. Pentachlorophenol and Iron
Increased the porphyrlnogenlc effect of hexachlorobenzene, whereas
decachloroblphenyl had no effect. Hexachlorobenzene pretreatment resulted
12-127
-------
1n Increased CC1. toxldty and altered Immune responses In hexachloro-
benzene-treated animals. In addition, hexachlorobenzene has been shown to
Induce hepatic xenoblotlc metabolism and thus has the potential to alter the
rate and extent of metabolism of other chemicals (see Section 12.3.1.).
Debets et al. (1980b) studied the effect of pentachlorophenol (PCP) on
hexachlorobenzene toxldty. Groups of female rats were fed diets containing
1000 yg hexachlorobenzene/g, 500 yg pentachlorophenol/g, or both chemi-
cals In the same amounts, and a fourth group served as the control. Penta-
chlorophenol accelerated the onset of hexachlorobenzene-induced porphyrla,
as Indicated by an Increase 1n urinary excretion of uroporphyrln and a
decrease of porphyrlns with two and three carboxyllc groups. This Increase
occurred ~3 weeks earlier 1n the hexachlorobenzene plus pentachlorophenol-
treated animals than 1n hexachlorobenzene-treated animals.
Razzard1n1 and Smith (1982) Investigated dlethylstHboestrol (DES) pre-
treatment on hexachlorobenzene metabolite excretion 1n young male and female
F344/N rats. The rats were Injected l.p. with four doses of OES dlproplo-
nate 20 ymoles/kg dissolved 1n arachls oil over a 24-day period and then
given 14 tug/kg hexachlorobenzene by oral Intubation for 7 days. The results
Indicated that the DES pretreatment stimulated the excretion, via urine and
feces, 1n both males and females (Table 12-37).
Blekkenhorst et al. (1980) reported that the simultaneous 1.m. adminis-
tration of Iron and hexachlorobenzene caused a marked potentlatlon of hexa-
chlorobenzene porphyrlnogenlc effect 1n rats. This was shown by a decrease
1n hepatic uroporphyrlnogen decarboxylase activity and Increased urinary and
fecal porphyrln excretion. Conversely, simultaneous bleeding of hexachloro-
benzene-treated rats diminished the porphyrlnogenlc effect of hexachloro-
benzene.
12-128
-------
TABLE 12-37
Analysis of the Excreta from Rats Administered Hexachlorobenzene
After an Initial Treatment with D1ethylst1lboestro1a»b
fv>
ID
Sex and Treatment
Pentachlorophenol
Tetrachlorobenzene-1,4-d1ol
(nmole/24 hours/kg bw)
Pentachlorothlophenol
Urine
Male + oil
Male + DES
Female + oil
Female + DES
Feces
Male + oil
Male + DES
Female + oil
Female + OES
151 + 19
190 + 22
174 + 17
453 + 105f
85 + 15
160 + 23f
116 + 35
279 + 80
3 + 1
17 + 2C
16 + 2d
35 + 9
Trace
Trace
Trace
Trace
23
158
142
176
74
166
65
149
+ 3
t 9C
+ 12e
±?f
+ 23
+ 33
+ 4
+ 13C
Source: Razzardlnla and Smith, 1982
bMale and female rats (52-54 and 71-73 days old, respectively) were given 20 ymole of DES dlproplo-
nate/kg dissolved 1n arachls oil (10 mg/ml) or oil alone by 1.p. Injection on days 1, 4, 14 and 24.
From day 25 all rats were given 14 mg of hexachlorobenzene/kg by oral Intubation dally for 7 days. After
the last dose 24-hour samples of urine and feces were collected, hydrolyzed and analyzed. Results are
means +_ S.E.H. (n=4/group).
€S1gn1f1cance of differences from rats not given OES, p<0.001
Significance of differences from males, p<0.005
e
Significance of differences from males, p<0.001
^Significance of differences from rats not given DES, p<0.05
Total excretions of these metabolites were: male,
female + OES, 1092+175 (p<0.025) nmole/24 hours/kg
336+57; male + OES, 691+70 (p<0.01); female, 513+62;
-------
Goldstein et al. (1978) studied the comparative toxldty of pure hexa-
chlorobenzene (purity >99%) and technical hexachlorobenzene (purity 32%)
which was known to contain 200 ppm of decachloroblphenyl and 4 ppm of octa-
chlorodlbenzofuran, 1n female CD rats fed diets containing 0, 30, 100, 300
or 1000 pg hexachlorobenzene/g for up to 15 weeks. Neither grade con-
tained other chlorinated dlbenzofurans or d1benzo-p-d1ox1ns. Both grades
resulted 1n comparable effects (porphyrla, cutaneous lesions, hyperexclta-
bmty, changes 1n liver enzymes and morphological Hver changes) In treated
rats, although the technical grade appeared to be slightly more potent than
pure hexachlorobenzene 1n its effects on the pulmonary endothellum. The
Impurities did not appear to have a synerglstlc effect.
Kluwe et al. (1982) reported that pretreatment of male Sprague-Dawley
rats with hexachlorobenzene resulted 1n Increased CC1. toxldty. The rats
received seven doses of hexachlorobenzene at 30 mg/kg 1n corn oil once every
72 hours followed by an 1.p. Injection of CC1. at 0.0, 0.03, 0.05, 0.25,
1.0 or 2.0 ml/kg 1n 4 ma/kg corn oil 24 hours after the last hexachloro-
benzene treatment. Hexachlorobenzene pretreatment Increased the CCld-
Induced acute growth retardation, renal tubular functional Impairment,
hepatocellular necrosis and further reduced the survival of the animals.
Variable results were reported 1n a study on the effect of hexachlorobenzene
pretreatment of male albino Sprague-Oawley rats on the jyn vivo blotransfor-
matlon, residue deposition, and elimination of l4C-aldr1n, 1-naphthol,
DDT, hexachlorobenzene or mlrex (Clark et al,, 1981a). There was no evi-
dence of qualitative changes 1n the blotransformatlon of any test compound
that could be attributed to hexachlorobenzene pretreatment. Analysis of
residue deposition gave mixed results: less 14C residues were found 1n
rats fed diets containing hexachlorobenzene and then treated with
12-130
-------
l4C-aldr1n, more 14C residues were found after 14C~DDT or l4C-m1rex
treatment, and no difference was evident after 14C-hexachlorobenzene or
14C-l-naphthol treatment. Hexachlorobenzene also potentiates the effects
of stress on male Sprague-Dawley rats (Clark et a!., 1981). Rats fed 250
ppm hexachlorobenzene resulted 1n an Increased severe loss of body weight
when placed Into crowded cages and compared to the weight loss of crowded
control rats. Crowded rats fed hexachlorobenzene had higher tissue residues
of hexachlorobenzene and higher mortality than the non-crowded hexachloro-
benzene-treated rats or the control rats.
12.5. SUMMARY
The acute oral toxldty of hexachlorobenzene has been found to be low,
with LO™ values ranging from 1700-10,000 mg/kg. Subchronlc oral toxldty
studies with a number of mammalian species Indicated a significant Increase
In liver and kidney weights 1n hexachlorobenzene-treated animals. Some
studies have shown Increases 1n other organs as well. The livers from
hexachlorobenzene-exposed animals have shown hlstologlc changes such as
Irregular shaped and moderately enlarged liver mitochondria and Increases 1n
the size of the centrllobular hepatocytes. Chronic oral toxldty studies
revealed similar effects to those seen In the subchronic studies plus
hexachlorobenzene-assodated life-shortening and various hepatic and renal
pathologies. These subchronic and chronic effects were usually dose-
related. Other effects Included multiple alopecia and scabbing, together
with neurologic effects 1n rats, mice and dogs. A dose-related hlstopatho-
loglc change In the ovaries of monkeys has also been reported.
Increased porphyMn levels 1n the liver and In urine have been reported
for all species studied except the dog, which does not exhibit Increased
porphyrln levels. Hexachlorobenzene was found to cause the accumulation of
12-131
-------
B~H-stero1ds which Induce porphyrln biosynthesis and to Inhibit uroporphy-
rlnogen decarboxylases. The Inhibition of uroporphyrlnogen decarboxylases
appears to be due to pentachlorophenol, a hexachlorobenzene metabolite.
Indications are that females are more susceptible to hexachlorobenzene-
Induced porphyrla than are males, which may be related to the female estro-
gen levels and greater hexachlorobenzene metabolism, Hexachlorobenzene was
reported to produce a mixed-type Induction of cytochromes resembling that
produced by a combination of phenobarbltal (P-450) and 3,4-benzpyrene
(P-448). In addition, the activities of several hepatic mlcrosomal enzymes
were found to be Induced by hexachlorobenzene.
Hexachlorobenzene did not Induce dominant lethal mutations 1n two
studies but was reported to be mutagenlc 1n a yeast, SL_ cerevlslae, assay at
a concentration of 100 ppm. Hexachlorobenzene possessed no detectable
levels of mutagenlc activity 1n the Salmonella hlstldlne reversion assay.
The chronic toxldty studies provide sufficient evidence of the cardno-
genlclty of hexachlorobenzene 1n animals since there was an Increased Inci-
dence of malignant tumors of the Hver 1n two species, haemangloendothelloma
1n hamsters and hepatocellular carcinoma 1n rats as well as confirmed
reports of hepatoma 1n both of these species. Hexachlorobenzene was found
to cause teratogenlc effects In fetal mice whose mothers were Ingesting 100
mg/kg/day of hexachlorobenzene during days 7-16 of gestation. Certain
chemicals were found to alter the toxldty of hexachlorobenzene 1n mammals,
whereas hexachlorobenzene pretreatment was reported to Increase CC1.
toxldty and alter the Immune responses of treated animals.
12-132
-------
13. OVERVIEW OF EFFECTS OF MAJOR CONCERN
A primary factor 1n Identifying the major effects of concern resulting
from exposure to the chlorinated benzenes 1s the extent and adequacy of the
available studies on mammalian and human toxicology. As Indicated 1n the
section on research needs (see Section 2.3.), several areas related to the
toxlcity of these chemicals either have not been or have been poorly Inves-
tigated. Except for hexachlorobenzene, few studies have been performed on
the carcinogenic, reproductive and teratogenlc toxlcity of chlorinated
benzenes. However, reasonable data are available on the subchronlc toxic
effects produced by the oral and Inhalation routes of exposure for most of
the chlorinated benzenes In several species. Studies that provide adequate
data on the consequences of chronic exposure or reproductive and teratologlc
effects of particular chlorinated benzenes do exist, but are more limited in
number. The absence of discussion or presentation of data on a particular
chlorinated benzene should not be equated with an absence of effects or
diminished need for concern; more likely, it reflects a lack of adequate
Investigation.
13.1. PRINCIPAL EFFECTS AND TARGET ORGANS
The data available for identifying the principal effects and sites of
toxicity for the chlorinated benzenes are derived mainly from studies of
subchronic tox'idty, reproductive and teratogenlc effects, and reports of
effects on humans accidentally or occupationally exposed to chlorinated
benzenes. In general, the main sites affected by short-term, high-level
exposures are the hepatic, renal and nervous systems. Inhalation and oral
toxicity studies 1n several species Indicate that chlorinated benzenes are
capable of inducing hepatic and renal degeneration and necrosis, disrupting
porphyrln metabolism, and depressing the short-term functioning of the
nervous system. Levels of exposure below those causing hepatic and renal
13-1
-------
toxldty for some of the chlorinated benzenes have adverse effects on the
long-term functioning of the nervous system and on the hematopoletie system.
In several studies, the administration of two of the chlorinated benzenes,
penta- and hexachlorobenzene, during gestation In rats resulted 1n Increased
fetotoxldty, postnatal mortality and Incidence of fetal skeletal malforma-
tions. Studies In rodents have also shown hexachlorobenzene to be a
carcinogen.
Monochlorobenzene, when administered to rats, rabbits and dogs at
moderate to high doses by Inhalation or oral routes caused hepatic and renal
toxldty manifested by Increased liver and kidney weights, hlstopathologlc
changes, elevated serum enzymes, and liver and kidney necrosis (Monsanto
1967a,b; Irish, 1963; Khanln, 1969; OHley, 1977}. At high doses, dogs
developed depression of bone marrow activity (Monsanto 1967a, 1978). Con-
tinuous exposure by Inhalation at low doses disturbed the proper chronaxy
correlation of the muscle antagonists and Increased blood chollnesterase 1n
rats (Tarkhova, 1965). Humans exposed occupatlonally to monochlorobenzene
Intermittently for up to 2 years displayed signs of neurotoxlclty Including
numbness, cyanosis, hyperesthesla and muscle spasms (Rozenbaum, 1947).
Subchronlc administration of dlchlorobenzenes by Inhalation to rats,
rabbits and guinea pigs caused liver and kidney toxldty and pulmonary
congestion (HolUngsworth et al., 1956). Oral administration produced
hepatic porphyrla, pathologic changes 1n the kidneys and liver, and Inhibi-
tion of erythropolesls and bone marrow activity (R1m1ngton and Zlegler,
1963; HolUngsworth et al., 1956; Varashavskaya, 1976a,b). Chronic adminis-
tration of 1,2-d1chlorobenzene by gavage to rats and mice possibly at less
than maximum tolerated doses, did not produce statistically significant
changes 1n tumor Incidences (NTP, 1982). Case studies of human exposures
report a range of effects Including liver necrosis, depression of erythro-
13-2
-------
polesls and leukemia. A study of 26 persons exposed to 1,2-d1chlorobenzene
for 4 work days reported Increased chromosomal aberrations 1n peripheral
leukocytes (Zapata-Gayon et al., 1982).
Studies of the subchronlc Inhalation toxlclty of 1,2,4-tr1chlorobenzene
have Identified hepatic porphyMa and cellular degeneration as effects 1n
rats but not In rabbits or monkeys (Coate et al., 1977; Watanabe et al.,
1978). PorphyMa was also Induced 1n rats after the dietary adminis-
tration of high doses of 1,2,3- or 1,2,4-tr1chlorobenzene for 7 days
(R1m1ngton and Zlegler, 1963). Three studies using dermal applications of
1,2,4-tr1chlorobenzene or a mixture of 1,2,4- and 1,2,3-tr1chlorobenzene to
rabbits and guinea pigs reported skin Irritation at doses as low as 30
mg/kg/day and some systemic toxlclty at higher doses (Brown et al., 1969;
Powers et al., 1975; Rao et al., 1982). In a reproductive study 1n rats,
25, 100 or 400 ppm of 1,2,3-tr1chlorobenzene, administered to the parental
animals 1n their drinking water, produced no reproductive, hematologlc or
neurologic effects (Robinson et al., 1981). Retarded embryonic development
was observed 1n pregnant rats receiving 1,2,4-tr1chlorobenzene 360 mg/kg/day
on days 9-13 of gestation (KHchln and Ebron, 1983a). Adrenal enlargement
occurred In both the parents and offspring at the highest dose level. In a
2-year mouse skin painting study (Yamamoto et al., 1957) a slight Increase
1n tumors of all sites was reported, but no conclusions can be drawn about
carclnogenlclty because of the lack of details 1n the English translation of
the text.
More limited data were available on the toxlclty of the tetrachloroben-
zenes. A single oral subchronlc study with 1,2,4,5-tetrachlorobenzene 1n
rabbits Indicated effects on blood chemistry and hematology at low doses
(Fomenko, 1965); a chronic study with the same Isomers 1n dogs suggested
adverse effects on liver metabolism (Braun et al., 1978). In a study of
13-3
-------
workers exposed to 1,2,4,5-tetraehlorobenzene, found an Increased Incidence
of chromosomal abnormalities (decreased chromosome number per cell, poly-
ploldy during mitosis, and chromosomal malformations) 1n the leukocytes of
the workers (Klraly et al., 1979).
Data on the toxlclty of pentachlorobenzene were also limited. High
levels 1n the diets of rats caused Increased excretion of porphyrlns (Goerz
et al.» 1978) and Induced hlstopathologlc changes 1n the kidneys and liver
(Under et al., 1980). Studies of the reproductive and teratologlc effects
of pentachlorobenzene 1n rats Indicated that the chemical Increased fetal
deaths, reduced postnatal survival of pups and Increased the Incidence of
extra ribs and sternal defects (Under et al., 1980; Khera and Vllleneuve,
1975). Teratogenlc effects were not seen 1n mice (Courtney et al., 1979).
The toxlclty of long-term dietary exposure of humans to hexachloroben-
zene was demonstrated by the epidemic of porphyrla cutanea tarda (PCT) 1n
Turkish citizens who accidentally consumed bread made from grain treated
with hexachlorobenzene (Cam, 1963; PeteYs et al., 1966; Peters et al.,
1982). In addition to the PCT-assoc1ated symptoms of skin lesions, hyper-
trlchosls, and hyperplgmentatlon, the exposure caused neurotoxlclty and
liver damage. Follow-up studies reported PCT symptoms, reduced growth, and
arthritic changes In the appendages of children who were directly or
Indirectly (I.e., through breast milk) exposed. Studies 1n rats have demon-
strated hexachlorobenzenes ability to Increase the Incidence of stillbirths,
decrease fetal growth and decrease postnatal survival (Grant et al., 1977;
Khera, 1974). A study 1n rats reported that administration of hexachloro-
benzene during gestation Increased significantly the number of fetuses with
extra ribs. A study 1n mice found that hexachlorobenzene given on days 7-16
of gestation resulted In an Increased Incidence of fetal abnormalities when
compared to controls (Courtney et al., 1976). Hexachlorobenzene has been
13-4
-------
shown to produce tumors In animals. Lifetime dietary administration of hex-
achlorobenzene to hamsters, rats and mice Increased the Incidence of thyroid
tumors 1n hamsters {Cabral et al., 1977), liver tumors 1n hamsters (Cabral
et al., 1977), mice (Cabral et al., 1979) and rats (Smith and Cabral, 1980;
Lambrecht, 1983; Arnold, 1984), kidney tumors 1n rats (Lambrecht, 1983) and
adrenal tumors 1n rats (Arnold, 1983; Peters et al., 1983).
13.2. ANIMAL TOXICITY STUDIES USEFUL FOR HEALTH ASSESSMENT AND ESTIMATED
TOXICITY THRESHOLDS
13.2,1. Animal Toxlclty Studies. The studies useful for health assessment
determinations of each of the chlorinated benzenes 1s presented 1n the res-
pective dose/effect Tables 13-1 through 13-7, extracted from the Hammallan
Toxlclty Sections of Chapters 7-12 of this document. These tables should
provide assistance 1n selecting the most useful and appropriate studies for
health assessment determinations.
Tables 13-8 through 13-12 attempt to compare a variety of toxic res-
ponses to chlorinated benzenes 1n rats, mice, rabbits, dogs and monkeys.
The recorded values reflect the lowest dosage reported for each listed
effect category for each species, taken from the subchronlc, chronic, car-
c1nogen1c1ty, reproductive and teratogenldty studies reported 1n Chapters
7-12 of this document. It should be noted that there 1s the potential for
similar responses 1n each species to occur at lower dose levels than
reported and that a blank entry does not necessarily mean that the effect
does not occur 1n that species Induced by the particular chlorinated
benzene. This 1s probably Indicative of the fact that lower dose levels may
not have been tested and/or that particular effect may not have been speci-
fically looked for by research investigators. In attempting to use these
tables to determine fine-line conclusions/interpretations about chlorinated
benzenes structure activity relationships, further complications arise
13-5
-------
TABLE 13-1
Suirraary of Subchronk ToxicHy Studies on Honochlorobenzene3
CO
I
Species
Dog
|!>p;iqle)
Rat
Rat
Rat
Rat
Rat
Rat
Rabbit
Route
Inhalation1*
Inhalation
Inhalation
Inhalation
Inhalation
Inhalation
Inhalation
Inhalation
Dose
0.75 mg/l, 6 hrs/day,
5 days/week (162 ppm)
1.50 mg/l, 6 hrs/day,
5 days/week (424 ppm)
2.00 mg/l, 6 hrs/day,
5 days /week
0.75, 1.50 or 2 mg/l
6 hrs/day, 5 days/week
0.1 or 1.0 mg/m3
(continuous)
0.1 nig/ffl3 (continuous)
1.0 mg/m3 (continuous)
0.1, 1.25 or 1.5 mg/l
0.1 rag/l, 3 hr/day
(alternate days)
75 and 250 ppm, 7 hrs/day
5 days/week
75 and 250 ppm, 7 hrs/day,
5 days/week
Duration
(days)
62
exposures
over 90 days
62
exposures
over 90 days
62
exposures
over 90 days
62
exposures
over 90 days
72-80
60
60
49-98
37 weeks
120
exposures
120
exposures
Effects Reference
None Monsanto, 197B
Height loss; conjunctivitis; moribund at
31 days
Weight loss; hypoactlvlty and conjunctivitis;
vacuolated hepatocytes; cytoplasmlc vacuolation
of renal collecting tubules; bilateral atrophy
of seminiferous tubules; lower total leukocyte
counts, elevated SAP, SGOT, SGPT; aplastlc bone
marrow; mortality in 5/8 dogs after 25-29 days
None Monsanto, 1978
Liver necrosis and regeneration; kidney Khanln, 1977
hyperplasla; encephalopathy; pneumonia
None Tarkhova, 1965
Inhibited chronaxla of antagonistic muscles
at 39 days; Increased blood chollnesterase
Chronaxlmetrk Inhibition Plslaru, 1960
Inhibition of extensor tiblalls 7-14 weeks; Gabor and Raucher,
normal by 20 weeks 1960
Focal lesions of adrenal cortex; lesions In Dllley, 1977
tubules of kidneys; congestion of liver and
kidneys; decreased SGOT
Decreased SGOT after 24 weeks of exposure Dllley, 1977
-------
TABLF 13-1 (conl.)
Species
Route
Dose
Duration
{days}
Effects
Reference
Mouse oral (gavage)
Rat
Oral {gavage}
60 rag/kg/day, 5 days/week 13 weeks
125 mg/kg/day, 5 days/week 13 weeks
250 rag/kg/day, 5 days/week 13 weeks
500 rag/kg day, 5 days/week 13 weeks
750 mg/kg/day, 5 days/week 10 weeks
60 mg/kg/day, 5 days/week 13 weeks
125 rag/kg/day, 5 days/week 13 weeks
250 mg/kg/day, 5 days/week 13 weeks
500 mg/kg/day, 5 days/Meek 13 weeks
750 nig/kg day, 5 days/week 13 weeks
one male with hepatic necrosis NTP, 1983
Increased liver weights 1n males one male
with hepatic necrosis
>50X reduction 1n weight gain, Increased
excretion of coproporphyrlns 1n females,
Increased liver weights, lesions of the
liver, kidney, bone marrow, spleen and
thymus
10W lethal to males within 1 week,
reduced body weight gains, polyurla
1n females, Increased liver weights,
lesions of the liver, kidney, bone
marrow, spleen and thymus.
100% lethal to male mice within 1 week
and to female mice within 10 weeks,
lesions of the liver, kidney, bone marrow,
spleen and thymus at death
Hone NTP, 1983
None
Minimal centrolobular hepatocellular
necrosis
Decreased body weights gain. Increased
GGTP and alkaline phosphatase In females,
Increased excretion of porphyrlns, con-
trolobular hepatocellular necrosis,
nephropathy 1n males, myelold depletion
of bone marrow.
Decreased body weight gain and survival
of animals, hematologlc effects. Increased
GGTP and alkaline phosphatase In females,
polyurla In males, Increased excretion of
porphyrlns, centrolobular hepatocellular
necrosis, nephropathy, lymphold depletion
of thymus and spleen, myelo'd depletion of
bone marrow.
-------
TABLE 13-1 (cont.)
CO
1
CD
Species Route Dose
Dog oral (capsule) 27.3 mg/kg/day
54.6 mg/kg/day
272.5 mg/kg/day
Rat oral (diet) 12.5 or 50 mg/kg/day
100 mg/kg/day
250 rag/kg/day
Rat oral (diet) 14.4 mg/kg/day
144 and 288 mg/kg/day
Duration
(days)
90
90
90
93-99
33-99
93-99
192
192
Effects
None
Diarrhea and vomiting; conjunctivitis
4/8 died In 3-5 weeks; Increased Inmature
leukocytes; elevated SGOT and SAP, blllrubln
and cholesterol; low blood sugar; hlstopatho-
loglc changes In liver, kidneys, spleen, and
seminiferous fubules.
None
Increased liver and kidney weights
Increased liver and kidney weights;
retarded growth In males
None
Increased liver and kidney weights;
Increased salivation and hair loss
Reference
Monsanto, 1967a
Monsanto, 1967b
Irish, 1963
asource: Updated from U.S. EPA, 1980a
bl ppm -4.60 mg/ni», 1 mg/S. -219 ppm (Irish, 1963)
-------
TABLE 13-2
Subchronlc ToxIcHy of 1,2-D1chlorobenzene*
Route Concentration
or Dose
Inhalation 560 mg/m3
290 mg/m3
455 mg/m"
Oral 376 mg/kg (tube)
188 mg/kg (tube)
co 18.8 mg/kg (tube)
i
Regimen
7 hours/day, 5 days/week,
6-7 months
7 hours/day, 5 days/week
6.5 months
dally up to 15 days
5 days/week, 138 doses
5 days/week, 138 doses
5 days/week, 138 doses
Subject
rat, guinea
pig, rabbit,
monkey
rat, guinea
pig
rat
rat
rat
rat
Effect
No effect on several parameters
except decreased spleen weights
1n male guinea pigs
No effect on several parameters
Hepatic porphyMa
Liver, kidney weight Increase;
cloudy swelling 1n liver.
Increase In liver and kidney
we 1 gh t
No effects noted
Reference
Holllngsworth et al. ,
1958
HolUngsworth et al, ,
1958
R1m1ngton and
Zlegler, 1963
Holllngsworth et al. ,
1958
HolUngsworth et al. ,
1958
Holllngsworth et al. ,
1958
0.01-0.1 mg/kg/day 5 months
500 mg/kg
5 days/week, 13 weeks
rat
rat
250 mg/kg
125 mg/kg
60 mg/kg
30 ing/kg
5 days/week, 13 weeks
5 days/week, 13 weeks
5 days/week, 13 weeks
5 days/week, 13 weeks
rat
rat
rat
rat
HematopoleMc system; altered .
conditioned reflexes; Increased
prothromb time and altered
enzyme activities
Increased liver weights; polyur.la
1n males; Increased urinary por-
phyrlns; hepatic necrosis and
degeneration; renal tubular
degeneration; thymlc lymphold
depletion; and hematologlc and
clinical changes
Increased liver weights; hema-
tologlc and clinical changes;
hepatic necrosis
Increased liver weights; hema-
tologlc and clinical changes;
some hepatic necrosis
Hematologlc and clinical changes
Hematologlc and clinical changes
Varshavskaya, 1967a
NTP, 1982
NTP, 1982
NTP, 1982
NTP, 1982
NTP, 1982
-------
1ABLF 13-2 (coot.)
Route
Concentration
or Dose
Regimen
Subject
Effect
Reference
w
i
Oral (cont.) 500 rag/kg
250 rag/kg
5 days/week, 13 weeks
5 days/week, 13 weeks
30, 60, 125 rag/kg 5 days/week, 13 weeks
Subcutaneous unspecified repeated
mouse
mouse
mouse
rabbit
Increased mortality; Increased
liver weights; Increased urinary
and liver porphyrlns; hepatic
necrosis and degeneration; heart
and skeletal muscle lesions;
lymphold depletion of thymus and
spleen
Hepatic necrosis and degeneration
In males; no effects 1n females
No effects
Blood dyscraslas, (agranulo-
cytosls)
NTP, 1982
NTP, 19B2
NTP, 1982
Ware and West, 197?
'Source: Hodlfed from U.S. EPA, 1980c
-------
TABLE 13-3
Subchronlc and Chronic Toxldty of 1,4-D1chlorobenzene*
Route Concentration Regimen
or Dose
Inhalation 105 mg/m3 0.5 hours/day, 5-9 days
4800 mg/m3 8 hours/day, 5 days/week,
up to 69 exposures
Subject
rabbit
rat, guinea pig,
rabbit
Effect
Granulocytopenla; Irritation; CNS
and lung toxldty; death (12/18)
Severe Irritation; CNS depression
and collapse; liver, kidney, lung
Reference
Zupko and Edwards,
1949
HolUngsworth et al. ,
1956
4600-4800 mg/m3
2050 mg/m3
1040 mg/m3
950 mg/m3
900 mg/m3
580 mg/m3
500 ppm
(-3000 mg/m3)
75 ppm
(-450 mg/m3)
8 hours/day, 5 days/week,
7 hours/day, 5 days/week,
6 months
7 hours/day, 5 days/week,
16 days
7 hours/day, 5 days/week,
157-219 days
8 hours/day, 2 weeks
7 hours/day, 5 days/weeks
6-7 months
5 hours/day, 5 days/week,
for 76 weeks followed by
36 weeks with no exposure
5 hours/day, 5 days/week,
for 76 weeks followed by
36 weeks with no exposure
rabbit
rat, guinea pig
rat, guinea pig
rat, guinea pig,
rabbit, mouse,
monkey
rat, guinea pig,
mice, rabbit,
monkey
rat
rat
pathology; deaths
Tremors, weakness, nystagmus;
some deaths
Growth depression, Increased liver,
kidney weight; liver pathology
(necrosis, fatty degeneration,
swelling, flbrosls)
Increased liver, kidney weight
(rat); lung, liver pathology
Growth depression (guinea pig);
Increased liver, kidney weight;
hlstologlcal liver changes
(cloudy swelling, granular
degeneration) In rats, no adverse
effects reported 1n rabbit, mouse
or monkey
Respiratory excitation; liver
pathology, deaths; at serum
concentration of 39 mg/i
No adverse effects on several
parameters
Slightly elevated protein and
coproporphyrln outputs. Increased
liver and kidney weights.
Some Increases 1n liver weights
P1ke, 1944
HolUngsworth et al.
1956
HolUngsworth et al.
1956
HolUngsworth et al.
1956
Ir1e et al., 1973
HolUngsworth et al.,
1956
Loeser and Utchfleld,
1983
Loeser and LHchHeld,
1983
-------
TABLE 13-3 (cont.)
Route Concentration
or Dose
Inhalation 500 ppw
(cent.) (-3000 ppro)
200 ppra
(-1200 rag/m')
75 ppm
(-450 mg/m*)
Oral 1000 rag/kg per
dose (tube)
to 770 mg/kg/day
i
500 mg/kg/day
(tube)
5000 mg/kg diet
500 mg/kg/day
(tube)
376 mg/kg/day
250 mg/kg/day
188 mg/kg/day
20-40 mg/kg/day
18.8 mg/kg/day
Regimen Subject
6 hours/day from days rat
6-15 of pregnancy
6 hours/day from days rat
6-15 of pregnancy
6 hours/day from days rat
6-15 of pregnancy
92 doses 1n 219 days rabbit
up to 5 days rat
5 days/week, 20 doses rat
up to 35 days Peking duck
5 days/week, 263 doses In rabbit
367 days
5 days/week, 138 doses in rat
192 days
3 days rat
5 days/week, 138 doses in rat
192 days
2 weeks rat
5 days/week, 138 doses in rat
192 days
Effect
5 dams out of 20 delivered litter
1 day early, one fetus with
agnathia and cleft palate
1 dam out of 20 delivered Utter
1 day early, one fetus with
gastroschisls and malrotation
of hindHmb
1 dam out of 20 delivered litter
1 day early, one fetus with
gastroschisls and malrotation
of hlndlin*
CNS depression; weight loss;
liver degeneration and necrosis;
deaths
Hepatic porphyria
Hepatic centrolobular necrosis;
cloudy swelling, renal tubular
epithelium, and casts
Death in 3/10. Retarded growth
CNS depression; weight loss; liver
pathology
Increased liver and kidney weight;
liver cirrhosis and focal necrosis
Induced liver metabolism enzyme
system
Increased liver and kidney weight
Induced liver metabolism enzyme
system
No adverse effects detected
Reference
Loeser and Litchfield,
1983
Loeser and Litchfield,
1983
Loeser and Litchfield,
1983
HolHngsworth et al.,
1956
Rimington and Zlegler,
1963
Hollingsworth et al..
1956
HolHngsworth et al. ,
1956
HolHngsworth et al..
1956
HolHngsworth et al. ,
1956
Arlyoshi et al.,
1975a,b
Hollingsworth et al.,
1956
Carlson and Tardiff,
1976
Hollingsworth et al..
1956
•Source: U.S. EPA, 1980c
-------
TABLE 13-4
Summary of Subchronlc and Chronic Toxldty Studies on Trlchlorobenzenes
CO
i
Species Route
Rat Inhalation
Rats, rabbits, Inhalation
two dogs
Rat Inhalation
Rat Inhalation
Rabbits, Inhalation
monkeys
Monkey oral
Rat oral
Rat oral
House oral
Dose
74.2, 742 or
7423 ntg/m'
of 1,3.5-TCB
223 or 742 mg/m»
of 1,2,4-TCB
22.3 or
74,2 mg/ms
of 1,2,4-TCB
186, 371 or
742 mg/m*
of 1,2,4-TCB
186, 371 or
742 mg/m*
of 1,2,4-TCB
1, 5, 25, 90,
125 or 173.6
mg/kg/day
of 1,2,4-TCB
50, 100 or
200 mg/kg/day
of 1,2,4-TCB
10, 20 or
40 mg/kg/day
of 1,2,4-TCB
600 ppm diet
{0.078 mg/kg/
day) of
1,2,4-lCB
Duration
6 hr/day, 5 day/wk
for up to 13 wk
7 hr/day, 5 day/wk;
total of 30 expo-
sures In 44 days
6 hr/day, 5 day/wk,
3 mo
7 hr/day, 5 day/wk,
26 wk
7 hr/day, 5 day/wk,
26 wk
30 days
30, 60, 90 or
120 days
90 days
6 mo
Effects Reference
No hepatotoxlclty; three high-dose rats had Sasmora and Palmer,
squamous metaplasia and focal hyperplasla 1981
of respiratory epithelium, believed to be
reversible
Increase In urinary excretion of porphyMa Koclba et al., 1981
1n exposed rats; Increase In liver weights
In high-dose rats and dogs; Increased kid-
ney weights In high-dose rats
Increase In urinary porphyrln excretion 1n Uatanabe et al., 1978
high-dose rats; no effects In 22.3 mg/m"
group
Enlarged hepatocytes and nondose-dependent Coate et al., 1977
hepatocytes vacuoHzatlon, liver granulance,
biliary hyperplasla and kidney hyaline de-
generation at 4 and 13 wk; no hUtopathology
evident at 26 wk
No treatment related changes at 26 wk Coate et al., 1977
<25 mg/kg/day - no effects observed; Smith et al., 197B
>90 mg/kg/day - observed toxldty and death
Increases 1n liver weights, liver porphyrlns Carlson, 1977b
and urine porphyrlns, dose and time related
Increase 1n I1ver-to-body weight ratio In Carlson and Tardlff,
high-dose group; changes 1n enzyme actlva- 1976
tlon at all doses
Mo effects Goto et al., 1972
-------
TABLE 13-4 (cent.)
CO
i
Species
Guinea pig
House
Rats
Rats
Route
dermal
dermal
oral
(drinking
water)
oral
Dose
0,5 ml/day
of 1,2,4-TCB
0.003 ml/paint-
ing of 30 and
60% solution 1n
acetone of
1,2,4-TCB
25, 100 or
400 mg/l
of 1,2,4-TCB
36, 120, 360 or
1200 mg/kg/day
of 1,2,4-TCB
Duration
5 day/wk, 3 wk
2 t1mes/wk, 2 yr
FO to FZ
generations
days 9-13 of
gestation
Effects
Death following extensor convulsion; livers
showed necrotlc foci
Painting Induced excitability, panting and
epidermal thickening, Inflammation and
keratlnlzatlon; Increased organ weights and
mortality
Enlarged adrenals In FQ and FI generations
1200 tng/kg dose all dead by the 3rd day,
360 rag/kg dose caused 22% mortality 1n
dams and moderate hepatocellular hyper-
Reference
Brown et a"!., 1969
Yamamoto et al., 1957
Robinson et al., 1981
K1tch1n and Ebron,
1983a
Rabbits
dermal
30, 150 or
450 mg/kg/day
of 1,2,3-TCB
5 day/wk, 4 wk
trophy and non-significant Increases In
embryonic lethality and significantly
retarded embryonic development, 36 and
120 mg/kg groups not observed for embryonic
effects, but slight hepatocellular hyper-
trophy was reported In one 120 mg/kg dam
Dose-related skin Irritation; Increase In
urinary coproporphyrln In high-dose males
and slight pallor of liver In males and
females
Rao et al., 1982
1,2,3-TCB = 1,2,3-trlchlorobenzene; 1,2,4-TCB = 1,2,4-trlchlorobenzene; 1,3,5-TCB = 1,3,5-trlchlorobenzene
-------
TABLE 13-5
Summary of Toxldty Studies on Tetrachlorobenzenes
Species Route
Rat oral
Rat oral
Rabbit oral
i
01 Rat oral
Dog oral
Pregnant rats oral
Pregnant rats oral
Pregnant rats oral
Dose
0.5-500 mg/kg
of diet
1,2,4,5-TeCB
0.001, 0.005,
0.05 mg/kg/day
1,2,4,5-TeCB
0.001, 0.005,
0.05 mg/kg/day
1,2,4,5-TeCB
75 mg/kg/day
1,2,4,5,-TeCB
5 mg/kg/day
1,2,4,5-TeCB
50, 100,
200 rng/kg/day
1,2,4,5-TeCB
50, 100,
200 mg/kg/day
1,2,3,4-TeCB
50, 100,
200 mg/kg/day
1,2,3,5-TeCB
Duration
2B or 90 days
8 mo
8 mo
2 mo
2 yr exposure,
22 mo recovery
days 6-15 of
gestation
days 6-15 of
gestation
days 6-15 of
gestation
Effects
Increased liver and kidney weights and
hlstologlcal changes In liver and kidneys;
Increases In HFO activity, serum cholesterol
values
No effects observed In 0.001 mg/kg/day dose
group; 0.005 and 0.05 mg/kg/day doses caused
disruption In conditioned reflexes, Increases
1n liver weight coefficients and decrease 1n
serum SH groups
No effect observed In 0.001 mg/kg/day dose
group; 0.05 mg/kg dose caused disorder of
liver glycogen formation, altered serum SH
group levels, Increase 1n blood hemoglobin
and peripheral retlculocyte levels
Altered biochemical parameters Indicating
changes 1n hepatic and hematopoHIc homeo-
stasls
No controls used; elevated SAP and total
b1!1rub1n, returned to normal range 3 mo
after exposures ended
High-dose lethal to 9/10 of treated dams;
organ weight changes, elevated serum
cholesterol and liver metabolism enzymes,
no Indication of those changes were dose-
related
Induced maternal toxklty and Increased
lethality of pups at 200 mg/kg/day
Increased lethality 1n 200 mg/kg/day group
pups; one pup malformed and minor chondro-
genlc delay In other pups
Reference
VUleneuve et al.,
1983
Fomenko, 1965
Fomenko, 1965
Fomenko, 1965
Braun et al., 1978
Ruddlck et al., 1981
Ruddlck et al., 1981
Ruddlck et al., 1981
-------
TABLE 13-5 (cent.)
CO
i
Species
Pregnant rats
Route Dose
oral 30, 100, 300,
1000 mg/kg/day
1,2,4,5-TeCB
Duration
days 9-13 of
gestation ob-
served on day 14
Effects Reference
Only control and 1000 mg/kg/day group KHchln and Ebron,
examined for embryotoxIcUy and only 1983b
observed fewer Implantations than control,
slight hepatic centrolobular hypertrophy
1n 1000 mg/kg/day group, hepatic enzymes
Induced at all doses.
Pregnant rats oral
100, 300,
1000 mg/kg/day
1.2,3,4-TeCB
days 9-13 of
gestation ob-
served on day 14
Only control and 300 mg/kg/day group
examined for embryotoxkUy, significant
embryonic growth reduction was observed 1n
the 300 mg/kg/day group, maternal lethality
1n 300 (1/10 dams) and 1000 (7/19 dams)
mg/kg/day groups, minimal hepatocellular
hypertrophy 1n 300 mg/kg/day group, minimal
to moderate hepatocellular hypertrophy and
reduced body and liver weights 1n 1000
mg/kg/day group, hepatic enzymes Induced 1n
the 300 and 1000 mg/kg/day groups.
KHchln and Ebron,
1983c
1,2,4,5-TeCB = 1,2,4,5-tetrachlorobenzene
1,2,3,4-TeCB = 1,2,3,4-tetrachlorobenzene
1,2,3,5-TeCB = 1,2,3,5-tetrachlorobenzene
-------
TABLE 13-6
Summary of Subchronlc, Reproductive and Teratogenlc Toxldty Studies on Pentachlorobenzene
OJ
i
Species
Rat (female)
Rat (male)
Rat
(offspring)
Nice
Rat
Route
oral
(diet)
oral
(diet)
oral
(diet)
oral
oral
Dose
125, 250, 500
or 1000 mg/kg
In diet
125 or 1000
mg/kg In diet
125, 250, 500
or 1000 mg/kg
In mothers diet
50 or 100
mg/kg/gavage
50, 100 or 200
mg/kg/gavage
Duration
180 days
100 days
gestation and
during suckling
days 6-15 of
gestation
days 6-15 of
gestation
Effects Reference
Changes In hematologlc parameters 1n high- Under et al., 1980
dose group; Increase 1n liver weights,
hepatic hypertrophy and vacuollzatlon In
500 and 1000 mg/kg groups; Increased kid-
ney weight In high-dose group
High-dose group Induced changes In hemato- Under et al., 1980
logic parameters; hepatic and renal
histology and Increase 1n liver, kidney
and adrenal weights
Offspring treated with >250 mg/kg/dlet were Under et al., 1980
adversely affected (reduced survival, body
weights and Increased liver weights, hepato-
cellular enlargement)
Increase In liver weights of dams; no Courtney et al., 1979
adverse effects on total development or
survival
No observed toxlclty 1n adult rats; In- Khera and Vllleneuve,
creased total deaths at all doses, but not 1975
1n dose-related manner; extra ribs 1n ex-
posed fetuses and sternal defects In 200
mg/kg group
-------
1ABLE 13-7
Summary of Toxlclty Studies on Hexachlorobenzene
Species
Route
Dose
Duration
Effects
Reference
Rat
(females)
Rat
oral
oral
(diet)
100 nig/kg every other
day
0.5 mg/kg/day
2.0 mg/kg/day
B.O mg/kg/day
32.0 mg/kg/day
Rat
(females)
oral
(diet)
weekly
100 mg/kg diet
up to 43 days
IS weeks exposed and
held to 48 weeks
15 weeks exposed and
held to 48 weeks
15 weeks exposed and
held to 48 weeks
15 weeks exposed and
held to 48 weeks
u
CO
Rat oral
(females) (gavage)
Rats oral
(females) (gavage)
50 dig/kg every other
day
0.5 mg/kg twice
weekly
?.0 mg/kg twice
week 1 y
8.0 mg/kg twice
week 1 y
32.0 mg/kg twice
15 weeks
29 weeks
29 weeks
29 weeks
29 weeks
98 days
Suggested covalent binding of hexachlorobenzene
metabolites to cytosollc proteins
Transient Increases 1n liver porphyrln levels
In females after termination of exposure
Increases In liver porphyrln levels In females
after termination of exposure. Increased size
of centrllobular hepatocytes
Increased liver weights, Increased liver.
kidney and spleen porphyrln levels 1n females
(porphyrla), centrllobular liver lesions espe-
cially In females at 48 weeks
Increased mortality 1n females. Intension
tremors In males and females and ataxla 1n a
few females, Increased liver, kidney and
spleen weights. Increased liver, kidney and
spleen porphyrln levels In females (porphyrla),
centrllobular liver lesions and splenomegaly
Increased liver, kidney, spleen and adrenal
weights, porphyrla (Increased liver porphyrln
levels and Increased excretion of porphyrlns
and precursors), tremors, hair loss and skin
lesions
Increase In relative liver weight
Increase In relative liver weight, moderately
enlarged hepatocytes
Porphyrla, markedly enlarged hepatocytes,
Increase In relative liver weight
Porphyrla, markedly enlarged hepatocytes,
Increase In liver weights
Porphyrla (Increased liver
porphyrlns).
Koss et al.,
198Qa
Kulper-Goodman
et al., 1977
Koss et al.,
1978b
B6ger et al.,
1979
decreased activity of uroporphyrlnogen
decarboxylase
Smith et al.,
1980
-------
TABLE 13-7 (cont.]
Species
Route
Dose
Duration
Effects
Reference
Rat
oral
(diet and
nursing)
Rat
to
i
Rat
Rat
(male)
Rat
(female)
Rat
(female)
Rat
Rat
(females)
Rat
(females)
Rat
oral
(diet)
oral
(diet)
oral
(diet)
oral
(diet)
oral
(gavage)
oral
(gavage)
oral
(gavage)
oral
(diet)
oral
(diet)
50 rag/kg diet
150 rag/kg diet
500, 1000 or 2000
rag/kg diet
2000 mg/kg diet
2000 mg/kg diet
3000 mg/kg diet
SO, 10.0 or 200 mg/kg
14 mg/kg every other
day
100 mg/kg every
other day
6-8 mg/kg/day
75 mg/kg diet
(4-5 mg/kg/day)
150 mg/kg diet
(8-9.5 mg/kg/day)
gestation until
5 weeks of age
gestation until
5 weeks of age
3 weeks
10 weeks
100 days
11 weeks
120 days
103 days
6 weeks exposed and
held for additional
18 months
75-90 weeks
up to 2 years
Depressed resistance to L_, monocytogenes and v"os et al.,
1- splralls. enhanced thymus-dependent antibody 1979b
response
Increased serum IgM and IgG, depressed resis-
tance to L_. monocytogenes and ]_. splralls,
enhanced thymus-dependent antibody response,
Increased liver and adrenal weights
Dose-related Increases 1n relative spleen, Vos et al,,
lymph nodes, liver, adrenals, thyroid, testes 1979a
and kidney weights, dose-related Increase 1n
serum IgM levels, no change 1n serum IgG
levels, dose-related pathological changes 1n
liver, lymph nodes and spleen
PorphyrU found microscopically at 5 weeks and Gralla et al,,
grossly at 10 weeks using fluorescence 1977
Elevated hepatic enzymes by 1 week and Increased Llssner
urinary porphyrln and ALA levels (porphyrla) as et al., 1975
early as 40 days
Decreased uroporphyrlnogen decarboxylase Elder et al.,
activity and porphyrla after 4 weeks 1976
Dose- and time-dependent Increase In liver and Carlson, 1977b
urine porphyrlns (porphyrla!
Porphyrla 1n treated females, susceptibility of R1zzard1n1 and
females to porphyrla may be related to estrogen Smith, 1982
levels
Porphyrla (liver uroporphyrln levels peaked 7 Koss et al.,
months postexposure and levels had not returned 1983
to normal by 18 months), decreased liver proto-
porphyrln and coproporphyrln levels, Inhibition
of uroporphyrlnogen decarboxylase activity
until 18 months postexposure
Decline 1n body weights, porphyrla, enlarged Smith and
livers and liver tumors Cabral, 1980
Porphyrla, time-related appearance of severe Lambrecht et
hepatic and renal pathologies, after 1 year 1n- al., 1983a,b
creases In hepatomas, hepatocardnomas, bile duct
adenomas, renal adenomas and renal carcinomas
-------
TABLE 13-7 (cont.)
Species
Rat
Route
oral
(diet)
oral
{diet and
nursing)
Dose
0,32, 1,6, 8.0 or
40 mg/kg diet
0,32 or 1.6 rag/kg
diet
Duration
-130 days
gestation through
lifetime (130 weeks)
Effects Reference
Hematologlcal changes at all dose levels In Arnold et al.t
males. Increases In liver and heart Heights In 1985
wales at 8.0 and 40 ppm diets, no treatment-
related effects observed 1n bred females
Glycogen depletion 1n 1.5 Dig/kg males; no
effects reported at 0.32 mg/kg
Rat
oral
(diet)
i
r\>
o
Rat
Rat
Rat
oral
(diet)
oral
(diet)
oral
(diet)
8.0 mg/kg diet
40 ng/kg diet
10 or 20 mg/kg diet
40 mg/kg diet
80 mg/kg diet
160 mg/kg diet
320 and 640 mg/kg
diet
60, 80, 100, 120 or
140 rag/kg diet
0 or 80 mg/kg diet
80 mg/kg diet
gestation through
lifetime (130 weeks)
gestation through
lifetime (130 weeks)
FQ to ^4 generations
FO to ?4 generations
FQ to F4 generations
FQ to F4 generations
FQ to F4 generations
F0 to Fla and F1b
generations
gestation and
nursing or cross
nursed with controls
2 weeks prior to
mating to 35-36 days
after weaning
Increase 1n liver pathologies
Increased mortality as pups. Increase In liver
and kidney pathologies, Increase In adrenal
pheochromocytomas 1n females and parathyroid
tumors In males
No effects reported
Increases In liver weights and aniline
hydroxylase activity
Decreased body weights, F3 and F4 generations had
decreased lactation Index and postnatal viability
and Increased stillbirths
Increased mortality and decreased lactation
Index starting 1n FI generation
20 and SOX mortality In F0 320 and 640 mg/kg
groups, respectively, greatly reduced fertility
Index and litter size and Increase 1n still-
births, viability Index zero In FI
Increased mortality 1n all groups at 21 days,
21-day LDsg values for pups were 100 and 140
mg/kg for F]a and FJJ, generations, respectively
Nursing exposure produced greater effects than
did gestatlonal exposure, effects noted were:
smaller brains, hearts, kidneys and spleens,
Increased liver weights
Increased porphyrln levels and decreased liver
esterase activity 1n dams, no changes 1n
gestation Indices or neonatal survival
Grant et al.,
1977
KHchln
et a!., 1982
Nendoza
et al., 1978
Hendoza
et al., 1979
-------
TABLE 13-7 (cont.]
CO
I
ro
Species
Rat
Mouse
House
(male)
House
(male)
Mouse
Mouse
Hamster
Hamster
Cats
(breeding
Route
oral
(gavage)
oral
(diet)
oral
(diet)
oral
(diet)
oral
(diet)
oral
(gavage)
oral
(diet)
oral
(diet)
oral
(diet)
Dose
10, 20, 40, 60, 80
or 120 mg/kg
2.5, 25 or 250
mg/kg diet
10 mg/kg diet (8.4
(mg/mouse/24 weeks)
or 50 mg/kg diet
(35.3 mg/inouse/
24 weeks}
167 mg/kg diet
6, 12, 24 and 36*
mg/kg/day
100 mg/kg/day to
pregnant mice
200 or 400 mg/kg
diet
4, 8 or 16 mg/kg/day
3 or 8.7 mg/day/cat
Duration
days 6-21 of gesta-
tion
21 days
24 weeks
3-6 weeks
101-120 weeks
*(15 weeks exposed
held until 120
weeks)
days 7-16 of
gestation
90 days
llfespan
142 days
Effects
Maternal toxldty (weight loss, tremors and
convulsions) and reduced fetal weights at 120
and 80 mg/kg maternal doses, dose-related In-
crease 1n incidence of unilateral and bilateral
14th rib, sternal defects were also noted 1n
one experiment
Dose-related Increase In liver and decrease 1n
prostate and seminal vesicle weights, dose-
related alterations 1n testosterone metabolism.
altered hepatic enzyme levels
Dose-related reduction 1n weight gain, no tumor
pathology observed
Impairment 1n host resistance as measured by
Increased sensitivity to S. typhosa and £.
berqher! , and decrease In IgA levels
Reduced growth rate at all dose levels, short-
ened llfespan associated with tremors and con-
vulsions 1n 24 and 36 mg/kg/day groups, dose-
dependent Increase 1n liver-cell tumors 1n the
12, 24 and 36 mg/kg/day dose groups
Increased maternal livers and decreased fetal
body weights. Increased Incidence of abnormal
fetuses per litter observed
Predrrhotlc and drrhotlc hepatic lesions,
bile-duct hyperplaslas and hepatomas
Shortened llfespan In 16 mg/kg/day group. In-
crease In hepatomas at all dose levels. Increase
In liver haemang1oendothel1oma 1n males and
females and an Increase 1n thyroid alveolar
adenomas 1n males 1n 16 mg/kg/day group
Weight loss and Increased disease susceptibility
1n bred females, dose-related decrease 1n Utter
Reference
Khera, 1974
EHssalde and
Clark, 1979
Sh1ra1 et al.,
1978
Loose et al,,
1978a, b
Cabral et al.,
1979
Courtney
et al., 1976
Lambrecht
et al., 1982
Cabral et al.,
1977
Hansen et al. ,
1979
females)
size and survival of offspring, hepatomegaly In
offspring
-------
1ABLE 13-7 (cont.)
w
I
Species
Hlnks
Dog
(female)
Dog
Honkey
(female)
Honkey
Route
oral
(diet)
oral
(capsule)
oral
(capsule)
oral
(gavage)
oral
(nursing)
Dose
1 or 5 mg/kg diet
50 or 150 mg/kg/day
1, 10. 100 or 1000
mg/day/dog
8, 32, 64 or 128
ing/kg/day
7.51-186 ppm milk
Duration
during gestation
until 17 weeks of
age
21 days
1 year
60 days
60 days
Effects
Dose-related Increase In offspring mortality,
Induction of hepatic HFO enzymes In exposed
offspring
Liver and hepatocyte enlargement, dose-Induced
electroencephalogram dysrhythmlas
Increase In mortality, neutrophlUa, and
anorexia In the 100 and 1000 mg dose groups,
dose-related nodular hyperplasla of gastric
lymphold tissue 1n all treated animals
Oose-related pathology In liver, kidney, ovaries
and thynus
2 of 3 Infants died as a result of exposures
Reference
Rush et al.,
1983
Sundlof
et al., 1981
Gralla et al.,
1977
latropoulus
et al., 1976
Bailey et al.,
1980
-------
TABLE 13-8
Comparison3'13 of Toxic Effects of Chlorinated Benzenes 1n Rats
Chemical
Mono-CB (I)
(0)
1,2-DCB (I)
(0)
_, 1,3-DCB (I)
co (0)
l
W 1,4-DCB (I)
(0)
1,2,3-TCB (I)
(0)
1,2,4-TCB (I)
(0)
1,3.5-TCB (I)
(0)
1,2.3,4-TeCB (I)
(0)
Organ/Body
Weight Changes
2000 mg/m3
(90)d
100 mg/kg/day
(99)
125 mg/kg (90)e
-450 mg/m5
(532)J
188 mg/kg
(192)6
186 mg/m3 (90)c
40 mg/kg/day
(90)
Altered
Enzyme
Levels
1.0 mg/m3
(60)
500
mg/kg/day
(90)
0.1
mg/kg/day
(150)
20
mg/kg/day
(14)
10
mg/kg/day
(90)
300
mg/kg/day
(5)
Porphyr1nogen1c Neurologic Hematopo1et1c
Effects Effects Effects
1 . 0 mg/m3
(60)
500 mg/kg/day 500 mg/kg/day
(90) (90)
455 mg/m3 (15)
500 mg/kg (90)e 0.01 0.1 mg/kg/day
mg/kg/day (150)
(150)
-3000 mg/m3 4800 mg/m3
(532)3 (97)f
770 mg/kg/day
(5)
74.2 mg/m3
(90)1
100 mg/kg/day
(30)
200 mg/kg/day
(10)
Renal
Effects
0.1 mg/m3
(80)
500
mg/kg/day
(90)
500 mg/kg
(90)6
4800
mg/m3
(97)f
500 mg/kg
(28)6
186 mg/m3
(90) =
Hepatic Adrenal Reproductive Carcinogenic
Effects Effects and Teratogenlc Effects
Effects
0.1 mg/m3 345 mg/m3C
(80) (168)
250
mg/kg/day
(90)
125 mg/kg
(90)6
950 mg/m3 -450 mg/m3
(219)C (10)1
376 mg/kg
(192)6
186 mg/m3
(90) =
120 33-56 360
mg/kg/day mg/kg/day mg/kg/day
(5) (95)9 (5)
300 200 mg/kg/day
mg/kg/day (10)
(5)
-------
TABU 13-8 (eont.)
I
tsj
Chemical
Organ/Body
Height Changes
Altered Porphyr1nogen1c Neurologic
Enzyne Effects Effects
Levels
HematopoleUc
Effects
Renal
Effects
Hepatic
Effects
Adrenal
Effects
1,2,3,5-TeCB (I)
(0)
Reproductive
and Teratogenlc
Effects
200 rag/kg/day
(10)
Carcinogenic
Effects
1,2,4,5-TeCB (I)
PCB
HCB
(0)
(I)
(0)
(I)
(0)
0.005 mg/kg/day
(240)
46 mg/kg/day
(180)
0.3 mg/kg/day
(130)
75
mg/kg/day
(60)
25
ng/kg/day
(10)
5 mg/kg/ 0.5 mg/kg/day
day (60) (105)
0.005
mg/kg/day
(240)
50 mg/kg
(105)h
75 mg/kg/day
(60)
97 mg/kg/day
(100)
0.01 mg/kg/day
(130)
97
mg/kg/day
(100)
2 rag/kg/
day (910)
1000
mg/kg/day
(5)
46
mg/kg/day
(180)
2 mg/kg/
day (105)
75
mg/kg/day
(60)
2 mg/kg/
day (910)
200 mg/kg/day
(10)
16-31 mg/kg/day
(100}
10 mg/kg/day
(10)
4-5 ing/kg/day
(730)
aAll values are the lowest dose level reported for each listed effect category, from the mammalian toxldty sections of Chapters 7-12, with dosing duration
listed In (days). A .blank Indicates that the effect has not been reported In this species for this Isoner,
From subchronlc, chronic, reproductive and teratogenldty studies
rag/m*. 7 hours/day, 5 days/week
mg/ms, 6 hours/day, 5 days/week
enig/kg, 5 days/week
ng/in3, 8 hours/day, 5 days/week
"in fn and F, generations: enlarged adrenals
h
mg/kg every other day
6 hours/day
Jmg/m3; 5 hours/day, 5 days/week
I = Inhalation exposure; 0 = oral exposure
Mono-OS = monochlorobenzene; DC8 » dlchlorobenzene; TCB = trlchlorobenzene; TeCB = tetrachlorobenzene; PCB = pentachlorobenzene; HCB = hexachlorobenzene
-------
TABLE 13-9
Comparison3'6 of Toxic Effects of Chlorinated Benzenes In Mice
Chemical
Organ/Body
Weight Changes
Altered
Enzyme
Levels
Porphyr1nogen1c Neurologic HematopoletU Renal
Effects Effects Effects Effects
Hepatic Adrenal Reproductive Carcinogenic
Effects Effects and Teratogenlc Effects
Effects
CO
I
ro
tn
Hono-CB (I)
(0) 125 mg/kg/day
(90)
1,2-DCB (I)
(0) 500 mg/kg
1,3-DCB (I)
(0)
1,4-DCB (I)
(0)
1.2,3-TCB (I)
(0)
1,2,4-TCB (I)
(0)
1,3,5-TCB (I)
(0)
1,2,3,4-TeCB (I)
(0)
1,2,3,5-TeCB (I)
(0)
1,2.4,5-TeCB (I)
(0)
250 mg/kg/day
(90)
500 mg/kg (90C
900 mg/m3
(14)d
500 mg/kg/day 250 mg/kg/
(90) day (90)
500 mg/kg
(90)c
60 mg/kg/
day (90)
250 mg/kg
(90)c
900 mg/m3
(14)1
-------
TABLE 13-9 (cont.)
to
Chemical
PCB
HCB
(I)
(0)
(I»
(0)
Organ/Body Altered PorphyMnogenlc Neurologic Hematopo1et1c
Weight Changes Enzyme Effects Effects Effects
Levels
50 mg/kg/day
(10)
0.01 mg/kg/day 0.01 mg/kg/ 24 mg/kg/
(21) day (21) day (840)
Renal Hepatic Adrenal Reproductive Carcinogenic
Effects Effects Effects and Teratogenk Effects
Effects
12 mg/kg/ 100 nig/kg/day 12 mg/kg/day
day (840) (10) {840)
aAll values are the lowest dose level reported for each listed effect category, from the mammalian toxldty sections of Chapters 7-12, with dosing duration
listed In days. A blank Indicates that the effect has not been reported 1n this species for this Isomer.
subchronlc, chronic, reproductive and teratogenicity studies
cmg/kg, 5 days/week
<% hours/day
I = Inhalation exposure; 0 = oral exposure
Mono-C8 = monochiorobenzene; DCS • dlchlorobenzene; TCB = trlchlorobenzene; TeCB = tetrachlorobenzene; PCB = pentachlorobenzene; HCB = hexachlorobenzene
-------
TABLE 13-10
Comparison3-11 of Toxic Effects of Chlorinated Benzenes 1n Rabbits
Organ/Body Altered Porphyr1nogen1c Neurologic Hematopoletlc Renal Hepatic
Chemical Height Changes Enzyme Effects Effects Effects Effects Effects
Levels
Adrenal Reproductive Carcinogenic
Effects and Teratogenlc Effects
Effects
OS
i
Ronp-CB
1,2-OCB
1,3-OCB
1,4-DCB
(I)
(0)
(I)
(0)
(I)
(0)
(I)
(0) 500 rag/kg
(367)s
345 mg/B3
1,2,3-TCB (I)
(0)
1.2.4-TCB (I)
(0) .
1,3,5-TCB (I)
(0)
1,2,3,4-TeCB (I)
(0)
1,2,3,5-TeCB (I)
(0)
4800 mg/m3
(97}d
500 rog/kg
(367)1
4800 mg/tn1
(97)d
4800 mg/is1
500 mg/kg
(367)g
-------
TABLE 13-10 (cont.)
w
1
ro
CO
Organ/Body Altered Porphyrlnogenlc Neurologic
Chemical Height Changes Enzyme Effects Effects
Levels
1,2,4.5-TeCB (I)
(0)
PCB (I)
(0)
HCB (I)
(0)
Hematopoletlc Renal
Effects Effects
0.05 mg/kg/day
(240)
Hepatic Adrenal Reproductive Carcinogenic
Effects Effects and Teratogenlc Effects
Effects
aAll values are the lowest dose level reported for each listed effect category, from the mammalian toxlclty sections of Chapters 7-12, with dosing duration
listed In days. A blank Indicates that the effect has not been reported In this species for this Isomer.
"From subchronlc, chronic, reproductive and teratogenlcHy studies
cmg/m>, 7 hours/day, 5 days/week
dmg/m3, 8 hours/day, 5 days/week
emg/kg, 5 days/week
I = Inhalation exposure; 0 = oral exposure
Hono-CB = monochlorobenzene; DCB = dlchlorobenzene; TCB = trlchlorobenzene; TeCB = tetrachlorobenzene; PCB = pentachlorobenzene; HCB = hexachlorobenzene
-------
TABLE 13-11
Comparison3"13 of Toxic Effects of Chlorinated Benzenes In Dogs
Organ/Body Altered Porphyrlnogenk Neurologic Hematopoletlc Renal Hepatic Adrenal Reproductive Carcinogenic
Chemical Weight Changes Enzyme Effects Effects Effects Effects Effects Effects and Teratogenlc Effects
Levels Effects
ro
U>
Hono-CB (I) 1500 mg/m*
(0)
2000 mg/m3
272.5 mg/
kg/day (90)
1,2-DCB
1,3-DCB
1,4-DCB
(I)
{0)
(I)
(0)
(I)
(0)
1,2,3-TCB (I)
(0)
1,2,4-TCB (I) 742 ng/m» (441s1
(0)
1,3,5-TCB (I)
(0)
1.2,3,4-TeCB (I)
(0)
1,2,3,5-TeCB (I)
(0)
2000 ing/m1
272,5 mg/kg/
day (90)
2000 mg/m' 2000 mg/m*
(90)c
272,5 mg/ 272,5 mg/
kf/day (90) kg/day (90)
2000 mg/m3
(90)c
272.5 mg/
kg/day (90)
-------
TABLE 13-11 (cont.)
CO
i
o
Organ/Body
Chemical Height Changes
1,2,4,5-TeCB (I)
(0)
Altered Porphyr1nogen1c Neurologic Hematopoletlc Renal
Enzyme Effects Effects Effects Effects
Levels
5 mg/kg/day
(730)
Hepatic Adrenal Reproductive
Effects Effects and Teratogenic
Effects
Carcinogenic
Effects
PCB
HCB
(I)
(I)
(0)
50 mg/dog/ 100 mg/dog/day
day (21) (365)
50 rug/dog/
day (21)
values are the lowest dose level reported for each listed effect category, from the mammalian toxldty sections of Chapters 7-12, with dosing duration
listed 1n (days). A blank Indicates that the effect has not been reported In this species for this Isomer.
bFrom subchronlc, chronic, reproductive and teratogenklty studies
ci»g/m*, 6 hours/day, 5 days/week
•'mg/m', 7 hours/day, 5 days/week
I = Inhalation exposure; 0 « oral exposure
Nono-CB = monochlorobenzene; DCB = dlchlorobenzene; TCB = trlchlorobenzene; TeCB = tetrachlorobenzene; PCB = pentachlorobenzene; HCB - hexachlorobeniene
-------
TABLE 13-12
Comparison3''' of Toxic Effects of Chlorinated Benienes 1n Monkeys
Organ/Body Altered Porphyr1nogen1e Neurologic Hematopoletlc Renal Hepatic Adrenal Reproductive Carcinogenic
Chemical Height Changes Enzyme Effects Effects Effects Effects Effects Effects and Teratogenlc Effects
Levels Effects
to
Mono-CB
1,2-DCB
1,3-DCB
1,4-OCB
d!
(0)
(I)
(0)
(I)
(0)
(I)
(0)
1,2,3-TCB (I)
(0)
1,2.4-TCB (I)
(0)
1,3,5-TCB (I)
(0)
1,2,3,4-TeCB (!)
(0)
1,2,3,5-TeCB (I)
(0)
1,2,4,5-TeC8
PCB
HCB
(0)
(I)
(0)
(I)
(0)
90 ng/kg/
day (30)
174 mg/kg/
day (30)
174 mg/kg/
day (30)
8 rag/kg/ 8 mg/kg/dav
day (60) (60)
3A11 values are the lowest dose level reported for each listed effect category, from the mammalian toxlclty sections of Chapters 7-12, with dosing duration
listed 1n (days). A blank Indicates that the effect has not been reported 1n this species for this Isomer.
From subchronlc, chronic, reproductive and teratogenlclty studies
I = Inhalation exposure; 0 = oral exposure
Hono-CB * monochlorobenzene; DCB « dlchlorobenzene; KB = trlchlorobenzene; TeCB = tetrachlorobenzene; PCB = pentachlorobenzene; HCB = hexachlorobenzene
-------
because the variety of studies used to glean the effects Information were as
follows: the studies were conducted under a wide range of experimental
conditions; employed different study durations; used a variety of animal
strains and different group sizes; and designed to assess different end-
points. Nevertheless, these tables do allow for a broad comparison of the
toxic effects Induced by the 12 different chlorinated benzenes 1n a variety
of species.
After reviewing Tables 3-8 through 13-12, It can be seen that large data
gaps exist for many of the chlorinated benzenes, especially for 1,3-
dlchlorobenzene, the trlchlorobenzenes and the tetrachlorobenzenes. Also,
except for the rat and possibly the mouse, the effects from subchronlc and
chronic exposure to the different chlorinated benzenes 1n a variety of
animal species have not been studied. The Interpretation of possible
chlorinated benzenes structure activity relationships will, for the most
part, be left to the document reader. The only Interpretation that will be
proposed from these comparison tables Is an apparent trend, for many of the
toxic effect categories, of Increased toxldty with Increased chlorlnatlon
of the benzene ring.
13.2.2. Estimated Toxldty Thresholds. Estimated toxldty threshold
levels as determined from the studies discussed In the respective mammalian
toxldty sections of Chapters 7-12 of this document are presented 1n Table
13-13.
13.3. CARCIN06ENICITY STUDIES
Adequate evidence of the carclnogenlclty of the different chlorinated
benzenes has only been found for hexachlorobenzene. The other chlorinated
benzenes either have not been studied for their carclnogenlclty or the
studies that have been conducted are Inadequate.
13-32
-------
TABLE 13-13
ToxkHy Data for Threshold Estimates
Compound
Honochlorobenzene
Monochlorobenzene
Honochlorobenzene
Honochlorobenzene
Monochlorobenzene
Honoc h 1 or oben zene
Honochlorobenzene
W Honochlorobenzene
i
co
w
1,2-Dlchlorobenzne
1,2-D1chlorobenzene
1 »2-D1chlorobenzene
1,2-Olchlorobenzene
1 ,2-D1chlorobenzene
1,2-Dlchlorobenzene
1,2-Dlchlorobenzene
l,4-D1chlorobenzene
1 , 4-OUhlorobenzene
l,4-D1chlorobenzene
Species
dog
rat
dog
rat
rat
rat
rat
mouse
rat, rabbH,
monkey
guinea pig
rat
rat
rat
mouse (female)
mouse (male)
rat, guinea pig,
mouse, rabbit,
monkey
rat
rat
Route
Inhalation
Inhalation
oral
oral
oral
oral
oral
oral
Inhalation
Inhalation
oral
oral
oral
oral
oral
Inhalation
Inhalation
oral
Dose
Concentration
0.75 mg/i (162 ppm),
6 hour/day, 5 day/week
2.0 mg/4, 6 hour/day,
5 day/week
27.3 mg/kg/day
50 rag/kg/day
14.4 mg/kg/day
125 mg/kg/day,
5 day/Meek
250 mg/kg/day,
5 day/week
60 mg/kg/day,
5 day/week
560 mg/m3,
7 hour /day, 5 day/week
290 mg/ra5,
7 hour/day, 5 day/week
18.8 mg/kg, 5 day/week
0.001 mg/kg/day
30 mg/kg, 5 day/week
250 mg/kg, 5 day/week
125 rag/kg, 5 day/week
580 mg/m3,
7 hour/day, 5 day/week
-450 flig/m3
5 hours/day, 5 days/week
18.8 mg/kg, 5 day/week
Dose
Duration
62 exposures
over 90 days
62 exposures
over 90 days
90 days
93-99 days
192 days
13 weeks
13 weeks
13 weeks
6-7 months
6.5 months
138 doses
5 months
13 weeks
13 weeks
13 weeks
6-7 months
76 weeks
138 doses
Effect
Level
NOEL3
NOEL3
NOEL3
NOEL3
NOAELb
NOEL3
LOAEL3
LOAEL3
NOEL3
NOEL3
NOELa
NOEL3
LOAEL3
NOELa
NOEL3
NOEL3
NOEL3
NOEL3
Reference
Honsanto, 1978
Monsanto, 1978
Honsanto, 1967a
Monsanto, 1967b
Irish, 1963
NTP,. 1983
NTP, 1983
NTP, 1983
HolUngsworth
et a!., 1958
HolUngsworth
et a!., 1958
HolUngsworth
et al., 1958
Varshavskaya,
1967a
NTP, 1982
NTP, 1982
NTP, 1982
HolUngsworth
et al., 1956
Loeser and
LHchfleld, 1983
Holllngsworth
et al., 1956
-------
TABLE 13-13 (cent.)
c*>
Compound
1,2,4-Trlchlorobenzene
1,2,4-Trlchlorobenzene
1 ,2,4-Tr1chlorobenzene
1 ,3,5-Tr1chlorobenzene
1 , 2,4,5-Tetrachlorobenzene
Pentachlorobenzene
Pentachlorobenzene
Pentachlorobenzene
Pentachlorobenzene
Hexachlorobenzene
Hexachlorobenzene
Hexachlorobenzene
Hexachlorobenzene
Species
rat
rabbit, monkey
monkey
rat
rat, rabbit
rat
rat
rat (offspring)
rat (offspring)
rat
rat
rat
rat
Route
Inhalation
Inhalation
oral
Inhalation
oral
oral
oral
oral
oral
oral
oral
oral
oral
Dose
Concentration
22." K^/n',
6 hour/day, 5 day/week
742 rag/m*.
7 hour/day, 5 day/week
25 rag/kg/day
74.2 mg/m»,
6 hour/day, 5 day/week
0.001 mg/kf/day
250 mg/kg diet
(-16-31 mg/kg/day)
500 mg/kg diet
(-27-63 ng/kg/day)
125 mg/kg diet
(-14-16 mg/kg/day)
50 mg/kg/day
0.5 mg/kg/day
2.0 mg/kg/day
0.32 rug/kg diet
(0.01-0.04 mg/kg/day )
20 mg/kg diet
Dose
Duration
3 months
26 weeks
30 days
13 weeks
8 months
180 days
180 days
gestation and
suckling
days 6-15 of
gestation
15 weeks
15 weeks
gestation-
lifetime
F0 to F4
generations
Effect
Level
NQAEL3
NOEL*
NOEL3
NOAEL3
NOEL3
MOELa
LOAELa
NOEL*
LOAEL3
NOAEL3
LOAEL3
NOEL3
NOEL*
Reference
Hatanabe et al.,
1978
Coate et al.,
1977
Smith et al.,
1978
Sasmore and
Palmer, 1981
. Fomenko, 1965
Llnder et al.,
1980
Llnder et al.,
1980
Linder et al.,
1980
Khera and
Villeneuve, 1975
Kul per -Goodman
et al., 1977
Ku1 per -Goodman
et al., 1977
Arnold et al.,
1985
Grant et al.,
1977
Estimated toxldty thresholds as determined 1n the respective Mammalian Tox1c1ty Sections of this document.
Estimated toxldty thresholds as found 1n U.S. EPA, 1980b.
NOEL = No-observed-effect level: That exposure level at which there are no statistically significant Increases 1n frequency or severity of
effects between the exposed population and Its appropriate control.
NOAEL = No-observed-adverse-effect level: That exposure level at which there are no statistically significant Increases 1n frequency or
severity of adverse effects between the exposed population and Its appropriate control. Effects are produced at this dose, but they are not
considered to be adverse.
LOAEL = Lowest-observed-adverse-effect level; The lowest exposure level 1n a study or group of studies which produces statistically signifi-
cant Increases 1n frequency or severity of effects between the exposed population and Its appropriate control.
-------
The chlorinated benzenes for which animal carc1nogen1c1ty studies were
available for review were hexachlorobenzene, 1,2,4-tMchlorobenzene,
1,2-dlchlorobenzene, 1,4-dlchlorobenzene and monochlorobenzene. One study
which Included both rats and mice, was available for monochlorobenzene and
for 1,2-d1chlorobenzene. The chronic studies which were available for
hexachlorobenzene Included two on hamsters, one on mice and four on rats as
well as a few studies which failed to qualify as carc1nogen1c1ty tests,
All of the hexachlorobenzene eardnogenldty studies, with the exception
of one, conducted at low doses, yielded one or more sites of significantly
Increased tumor Incidence. The primary target organ appears to be the
liver, but thyroid, parathyroid and adrenal glands also showed Increases and
1n one Instance kidney tumors were Increased.
Malignant liver tumors occurred 1n one experiment on hamsters (Cabral et
al., 1977) where the Incidence of haemang1oendothel1oma was 6/30 (20X) 1n
treated males at 8 mg/kg bw/day compared with 0/40 (OX) 1n controls and an
Incidence of 7/60 (12X) 1n treated females at 16 mg/kg bw/day compared with
0/39 (0%) 1n controls. In an experiment 1n rats Lambrecht (1983) obtained
hepatocellular carcinoma 1n hexaehlorobenzene-treated males with a frequency
of 3/52 (6%) compared to 0/54 (0%) 1n controls and of 36/56 (64X) 1n treated
females compared to 0/52 (0%) 1n controls. These effects were produced at
4-5 mg/kg bw/day.
*
Hepatoma was Increased as a result of hexachlorobenzene 1n two hamster
studies (Cabral et al., 1977; Lambrecht et al., 1982). In the latter report
i
the number of animals at risk was small and the Increase Just detectable.
In the Cabral study at a dose of 4 mg/kg bw/day both males and females had a
14/30 (47%) Incidence of hepatoma while controls for each sex had 0/40
(0%). The Incidence went as high as 51/60 (85%) at the largest dose used.
13-35
-------
Cabral (1979) also found an Incidence of 3/12 (25%) 1n both male and female
mice administered 6 mg/kg bw/day hexachlorobenzene compared with 0/50 (0%)
1n controls. The rat studies 1n which hepatomas were reported are those of
Smith and Cabral (1980) and Lambrecht et al. (1983). In the study of Smith
and Cabral (1980) an Agus rat strain was used that 1s particularly suscepti-
ble to porphyrla and Hver tumors. These animals, all female, gave 100%
yield of hepatoma 1n 14 animals compared to 0/12 (0%) 1n controls. Part of
that same study employed six female Wlstar rats 1n a treated group and 4 of
6 cases of hepatoma (67%) were reported compared with 0/4 (0%) 1n controls.
In Lambrecht's study at 4-5 mg/kg bw/day Sprague-Oawley rats developed
hepatoma 1n 19% of treated males and 46% of treated females while controls
for both sexes were 0/52 (0%). They also found hepatocellular carcinoma 1n
females with an Incidence of 36/56 (64%) at 4-5 mg/kg/day and 48/55 (87%) at
8-10 mg/kg/day.
The liver carc1nogen1c1ty and tumor1gen1city of hexachlorobenzene,
therefore, seems established by repeated experiments 1n rats and hamsters
and by a single study In mice. A high Incidence 1s Induced with doses as
low as 4-5 mg/kg bw/day. This dosage appears to be effective 1n three
rodent species 1n Inducing hepatoma.
Other tumors were reported as well as those which occurred 1n the liver.
In male hamsters, thyroid tumors were significantly elevated at 16 mg/kg
bw/day In males and 1n a 2-generat1on study, rats of the F, generation had
significant Increase In adrenal pheochromocytoma 1n females and parathyroid
tumors 1n males. These tumors may not be spurious for the following
reasons. The doses Involved did not produce significant toxldty and 1t 1s
13-36
-------
unlikely that nonspecific stress or systemic toxlclty evoked these re-
sponses. Also, one of the observations made on humans accidentally exposed
to hexachlorobenzene, Initially and 1n a 25-year follow-up, 1s thyroid
enlargement well above expected levels for that area (Peters et al., 1982).
In addition, rats exposed to monochlorobenzene had a significant decrease 1n
pituitary adenoma Incidence suggesting that the endocrine balance may be
affected by chlorinated benzenes.
There was one report of a significant Increase 1n renal cell adenoma 1n
rats of both sexes at 4-5 mg/kg bw/day.
The studies on 1,2-d1chlorobenzene and monochlorobenzene were conducted
at doses which may have been less than the MTD as estimated by subchronlc
range finding studies. In the case of 1,2-d1chlorobenzene 1n rats no
Increase In tumors or other pathology was found. In mice no tumor type was
significantly Increased compared with controls.
In the case of the monochlorobenzene a significantly Increased Incidence
of neoplastlc nodules 1n male rats was induced at a gavage dose of 120 mg/kg
bw/day. The data on the 1,2-d1ehlorobenzene and monochlorobenzene are In-
adequate to draw conclusions concerning the human carc1nogen1c1ty of these
compounds.
For hexachlorobenzene, the studies showing positive tumor responses are
summarized 1n Table 13-14. This compound has Induced Hver tumors 1n
hamsters, mice and rats, thyroid tumors 1n hamsters, and kidney and adrenal
tumors 1n rats. Using the IARC ranking system for classifying the evidence
of carc1nogen1c1ty, hexachlorobenzene would be a Group 2 chemical which IARC
describes as a probable carcinogen 1n humans.
13-37
-------
TABLE 13-14
Summary of Tumors Induced 1n Rodents by HCB
Hales
w
i
CO
00
Species
Hamsters
Hamsters
Hamsters
Hamsters
Nice
Rats
Rats
Rats
Rats
Lowest
to Produce Tumor
mg/kg bw/day
4
B male; 16 female
16
200 ppm
6
6-8
6-8
FI dose unknown
In utero, adult =
0.4
FI dose unknown
In utero, adult =
0.4
Tumor
Type
hepatoma
haeroangloendo-
thelloma of
Hver
thyroid adenoma
hepatoma
hepatoma
parathyroid
adrenal pheo-
chromocytoma
X
Treated/Control
47/0
20/0
14/0
8/0
25/0
25/4
35/23
Females
Tumor
Type
hepatoma
haemangloendo-
thelloma of
liver
thyroid adenoma
hepatoma
hepatoma
hepatoma
(Agus)
hepatoma
(Mlstar)
adrenal pheo-
chromocytoma
%
Treated/Control
47/0
12/0
6/0
8/0
25/0
100/0
67/0
35/4
Reference
Cabral. 1977
Cabral, 1977
Cabral, 1977
Lambrecht et
Cabral, 1979
Smith and
Cabral, 1980
Smith and
Cabral, 1980
Arnold et al.
Arnold et al.
al., 1982a
, 1985
, 1985
Rats VH. A content
varied. HCB = 0.4
none
none
Arnold et al., 1985
-------
TABLE 13-14 (cont.)
Hales
Species
_,
CO
1
CO
to Rats
Rats
Rats
Lowest
to Produce Tumor
imj/kg bw/day
4-5
4-5
4-5
Tumor
Type
hepatoma
hepatocellular
carcinoma
renal cell
adenoma
X
Treated/Control
19/0
6/0
79/13
Females
Tumor
Type
hepatoma
hepatocellular
carcinoma
renal cell
adenoma
X
Treated/Control
46/0
64/0
13/2
Reference
«
Lambrecht, 1983
Larnbrecht, 1983
Lambrecht, 1983
-------
A quantitative estimate of the carcinogenic potency of hexachlorobenzene
and an upper-bound estimate of the risks from continuous human exposure to 1
vtg/m3 1n air and 1 vg/9. In drinking water were made from data on the
hepatocellular carcinoma response 1n female rats. The upper-bound slope of
the dose-response curve, q,*, 1s 1.7/(mg/kg/day), giving a potency Index
which 1s 1n the second quartile of 54 suspect carcinogens evaluated by the
Carcinogen Assessment Group. The unit risks for air and water exposures are
4.9xlO~4 for 1 yg/m3 1n ambient air and 4.9xlO~5 for 1 vg/9, 1n
drinking water. Corresponding estimates from 13 other data sets, encompass-
ing different tumor sites and animal species, fall within a factor of 10 of
the above estimates, except for thyroid tumors 1n hamsters, which give esti-
mates of about 1/20 of the potency based on the rat hepatocellular carcinoma
response.
13.4. HUMAN STUDIES
Although animal studies Indicate that hexachlorobenzene 1s carcinogenic
1n hamsters, rats and mice, no adequate ep1dem1olog1c studies were available
to corroborate these findings 1n humans. However, the human data which has
been collected were not designed to detect human carc1nogen1c1ty, but rather
to provide a better understanding of hexachlorobenzene toxldty 1n Infants
(pink sore) and adults (porphyria cutanea tarda) (Cam, 1963; Crlpps et al.,
1981; Peters et al., 1966; Peters et al., 1982). In the studies of hexa-
chlorobenzene-lnduced toxldty, human consumption of hexachlorobenzene
through contaminated wheat was estimated at 50-200 mg hexachlorobenzene/
person/day (0.71-2.86 mg/kg bw/day for a 70 kg male); these doses were
sufficient to cause porphyria cutanea tarda and other effects 1n 3000-5000
people (Courtney, 1979). Ep1dem1olog1c studies with occupationally-exposed
workers or people living 1n the vicinity of a chlorinated solvents plant
13-40
-------
were not designed to detect carc1nogen1c1ty. The exposure Information
provided by those studies 1s not sufficient to relate dose level to effect
(Currier et al., 1980).
Two other chlorinated benzenes were reported to have effects 1n humans.
1,2-D1chlorobenzene (Zapata-Gayon et al., 1982) and 1,2,4,5-tetrachloroben-
zene (Klraly et al., 1979) each caused statistically significant Increases
in the frequency of chromosomal aberrations, but neither study reported the
ambient atmospheric concentration. Thus, these two chlorobenzenes are clas-
togenlc, but the critical exposure concentration 1s not known.
13.5. FACTORS INFLUENCING HEALTH HAZARD ASSESSMENT
13.5.1. Exposure. For an Individual or a population, exposure to poten-
tially toxic substances occurs on two levels. The first 1s exposure to
ambient environmental levels which occurs through food, drinking water and
air. Physiologic exposure 1s the second and more Important level and occurs
after the compound has been absorbed and 1s 1n a position to Interact
directly with critical cellular components. This Interaction 1s the basis
for toxlcologlc effects.
Chlorinated benzenes 1n the environment are resistant to blotransforma-
tlon and degradation and are, therefore, ecologically persistent compounds.
At the level of the Individual organism, these compounds are biologically
persistent because of their affinity for fatty tissues and their slow rate
of blotransformatlon or elimination (see Section 5.3.). Thus, biological
persistence and bloaccumulatlon 1n nonhuman organisms Increase the likeli-
hood of human exposure. Tables 13-15 and 13-16 present some useful proper-
ties and trends of chlorinated benzenes which Illustrate the differences
that exist between the chlorinated benzenes Isomers and their potential for
human exposure.
13-41
-------
TABLE 13-15
Comparison of Chemical and Physical Properties of Chlorinated Benzenes
Chemical
Honochlorobenzene
Olchlorobenzene
1,2-
1,3-
1,4-
_, Trlchlorobenzene
w 1,2,3-
i, 1,2,4-
NJ 1,3,5-
Tetrachlorobenzene
1,2,3,4-
1,2,3,5-
1,2,4,5-
Pentachlorobenzene
Hexachlorobenzene
Molecular
Height*
112.56
147.01
147.01
147.01
181.46
181.46
181.46
215.90
215.90
215.90
250.34
284.76
Melting
Point ("C)a
-45.6
-17.0
-24.7
53.1
52.6
16.95
63.4
47.5
54.5
139.5
86
230
Boiling
Point (°C)a
132
180.5
173
174
221
213.5
208.4
254
246
246
277
3229
Density
g/rac (20"C)a
1.1058
1.3048
1.2828 (25)
1.2475
1.69
1.4542
1.3865 (64)
HK
NA
1.858 (22)
1.8342 (16.5)
1.569 (23)
Log P"a
2.84
3.38
3.38
3.39
4.1
4.12
NA
NA
NA
4.93
5.63
5.8
Volatility 1n
Vapor Pressure
m Hg at 25*Cb
11.8
1.28
1.89
1.0
Likely to
be present as
* vapor In ambient air
0.07
0.29
0.15
0.04
0.07
0.05
-0
Not likely
1n ambient
likely to
condensed
etc.
1.68xlO~*''
to be present
air — more
be present In
state In soil
alncreas1ng trend
^Decreasing trend
NA = Not available
P" = Partition coefficient at 25*C
-------
TABLE 13-16
Comparison of Chlorinated Benzenes BCF and Water Concentrations
CO
I
CO
Chemical
Honoehlorobenzene
Dlchlorobenzene
1,2-
1,3-.
1,4-
Trlchlorobenzene
1,2,3-
1.2,4-
1.3,5-
Tetrachlorobenzene
1,2.3.4-
1,2,3,5-
1,2,4,5-
Pentachlorobenzene
Hexachlorobenzene
BCFa
(rainbow trout)
46
270-560
420-740
370-720
1,200-2,600
890-3,200
1,800-4,100
5,200-12,000
NA
5,300-13,000
13,000-20,000
5,500-20,000
Hean Chlorobenzene
Concentrations 1n
Drinking Waterb
(ng/l)
NA
3
1
13
0.1
2
<0.1
0,3
<0.05
0.2
0.04
0.1
Chlorinated Benzenes In
Various Wastewaters
Mean Concentration
(pg/D
667
141
21
79
NA
161
NA
NA
NA
NA
NA
NA
Increasing trend
^Decreasing trend
BCF = Bloconcentratlon factor; NA = not available
-------
Although toxic effects 1n humans have not been directly related to
ambient chlorinated benzene exposure, 1t 1s apparent from the residue levels
1n human tissues that humans receive physiologic exposures to the chlori-
nated benzenes (see Section 4.3.5.)- A comparison of human ambient exposure
levels and tissue concentrations confirms that humans bloaccumulate chlori-
nated benzenes (Burn et al., 1974; Currier et al., 1980). Prolonged
physiologic exposure and the uncertainty of the toxic effects of chronic
low-dose exposure to the chlorinated benzenes Increase the concern for human
exposures resulting from ambient levels of these substances.
The large number of locations at which chlorinated benzenes have been
detected Indicate their ubiquity 1n the environment and 1s a reflection of
their annual production volume, release rate, end uses (Sections 4.1.-4.3.),
and their environmental transport and fate (Sections 5.1.-6.3.). Human
exposure to these ambient concentrations depends on the chlorobenzene
concentration 1n, and absorption efficiency from, air, drinking water or
food. The relative contribution of each medium to the total human exposure
was estimated from monitoring data for several areas of the United States;
the limitations of these estimates are discussed 1n Section 4.4. The
estimated yearly exposures to the chlorinated benzenes from air are shown 1n
Table 13-17 and are based on the data for each chlorobenzene shown 1n Table
4-8. The available data Indicate that human exposure to chlorinated ben-
zenes through Inhalation may be greater than 1ngest1on exposure either
through drinking water or through foods. The relative contribution of food
to human exposure 1s less certain because food has not been extensively
monitored for chlorinated benzene residues; two studies estimated annual
hexachlorobenzene exposures of 0.026 mg/year and 0.145 mg/year, respectively
(IARC, 1979).
13-44
-------
TABLE 13-17
Estimated Yearly Exposure to Several
Chlorinated Benzenes Via Inhalation
Exposure (mq/yr)
Chemical
Honochlorobenzenes
1 ,2-D1chlorobenzene
1 ,3-D1chlorobenzene
1 ,4-D1chlorobenzene
Trlchlorobenzenes
Tetrachlorobenzenes
Mean Ambient Con-
centration (ng/m^)*
3087
1142
571
1563
136
3502
Adult
Han
25.9
9.6
4.8
13.1
1.1
29.4
Adult
Woman
23.8
8.8
4.4
12.0
1.0
27.0
Child
(10 yr)
17.0
6.3
3.1
8.6
0.7
19.3
Infant
(1 yr)
4.3
1.6
0.8
2,2
0.2
4.9
*Hean levels obtained from Table 4-8
13-45
-------
This paucity of data for food, however, does not preclude this medium as
a significant human exposure route. Trout from the Great Lakes, for
example, had detectable levels of all of the chlorobenzenes except mono-
chlorobenzene (Oliver and N1col, 1982), and It 1s conceivable that other
animals used as food sources also have tissue residues of chlorobenzenes.
13.6. REGULATIONS AND STANDARDS
The chlorinated benzenes are regulated under numerous United States and
foreign statutes. These have been grouped according to the type of activity
or medium being controlled.
13.6.1. Occupational Standards.
13.6.1.1. HONOCHLOROBENZENE -- The current OSHA standard for mono-
3
chlorobenzene levels 1n the workplace 1s 75 ppm (350 mg/m ). This thres-
hold limit value (TLV), established In 1974, Is not to be exceeded for an
8-hour time weighted average (TWA) for an employee's exposure 1n any 8-hour
shift of a 40-hour workweek (29 CFR 1910). This standard 1s Identical to
those recommended by the American Conference of Governmental Industrial
Hyg1en1sts (ACGIH, 1982). Occupational standards for monochlorobenzene have
also been established 1n four foreign countries. These are presented 1n
Table 13-18.
The Interagency Testing Committee (ITC) designated monochlorobenzene a
TSCA Section 4(e) priority chemical 1n Us Initial Report to the Administra-
tor of EPA (44 FR 70666). Additionally, all manufacturers and producers of
monochlorobenzene were required to report exposure, production and use
Information to EPA's Office of Toxic Substances In the form of a Preliminary
Assessment Information Manufacturers Report. The deadline for submission
was November 19, 1982 (40 CFR 712).
13-46
-------
TABLE 13-18
Occupational Standards for Honochlorobenzene*
TLV
Year
Country ppm mg/m^ Adopted
USSR 11 50 1972
German Democratic Republic — 50 1973
Czechoslovakia — 200 1969
Federal Republic of Germany 50 230 1974
USA 75 350 1974
*Source: Verschueren, 1977
13-47
-------
13.6.1.2. OICHLOROBENZENES — The OSHA standard for 1,2-d1chloroben-
3
zene 1n the workplace 1s set at a celling of 50 ppm (300 mg/m ). Levels
1n the workplace are at no time permitted to exceed this value (39 FR, No.
125). The 1982 AC6IH TLV for 1,2-dlchlorobenzene 1s Identical (ACGIH,
1982). Foreign standards for occupational exposure to l,2-d1chlorobenzene
are shown 1n Table 13-19.
In 1978, NIOSH classified l,2-d1chlorobenzene as a Group II pesticide (a
pesticide that poses "adverse acute health risks at moderate doses") and
recommended criteria for standards for occupations 1n pesticide manufactur-
ing and formulating (NIOSH, 1978). These standards rely on engineering
controls, work practices and medical surveillance programs, rather than
workplace air limits, to protect workers from adverse effects of pesticide
exposure 1n pesticide manufacturing and formulating. NIOSH specifically
chose not to establish scientifically valid environmental (workplace air)
limits for pesticides (except those already promulgated) because exposure
via other routes, especially dermal, had proven to be of critical Importance
for many pesticides and NIOSH believed that "Immediate action" was needed to
protect workers 1n pesticide manufacturing and formulating plants (NIOSH,
1978).
The current OSHA standard for l,4-d1chlorobenzene 1n the workplace 1s a
3
TLV of 75 ppm, 450 mg/m (39 FR, No. 125). In addition to recommending a
TLV Identical to the OSHA standard, ACGIH has recommended a short-term
exposure limit (STEL), the maximum concentration allowable 1n a 15-mlnute
period, of 110 ppm (675 mg/m ) for l,4-d1chlorobenzene (ACGIH, 1982).
NIOSH has also classlfed 1,4-dlchlorobenzene as a Group II pesticide and
recommended criteria for workplace standards In pesticide manufacturing and
formulating plants (NIOSH, 1978). Foreign standards for occupational expo-
sure to 1,4-dlchlorobenzene are presented In Table 13-20.
13-48
-------
TABLE 13-19
Occupational Standards for l,2-D1chlorobenzenea
Country
(Standard)
Level
ppm
Year
Adopted
USSR (TLV)b
German Democratic Republic (TLV)
USA (MAC)C
Federal Republic of Germany (TLV)
50
50
20
150
300
300
1972
1973
1974
1974
aSource: Verschueren, 1977
^Threshold limit value
cNax1mum allowable concentration
13-49
-------
TABLE 13-20
Occupational Standards for 1,4-D1chlorobenzene*
Country
USSR
German Democratic Republic
USA
Federal Republic of Germany
TLV
ppm mg/m3
20
200
75 450
75 450
Year
Adopted
1972
1973
1974
1974
*Source: Verscnueren, 1977
13-50
-------
There are no occupational workplace standards, either United States or
foreign, for 1,3-d1chlorobenzene. However, dlchlorobenzenes (no Isomer
specified) were designated by the ITC as TSCA Section 4(e) priority chemi-
cals (44 FR 70666). Separate Preliminary Assessment Information Manufac-
turers Reports on 1,2-, 1,3- and 1,4-d1chlorobenzene were to be submitted to
EPA by November 19, 1982 (40 CFR 712).
13.6.1.3. TRICHLOR08ENZENES — There are no United States workplace
standards for the tMchlorobenzenes.
3
The ACGIH has recommended a celling of 5 ppm (40 mg/m ) for 1,2,4-
tMchlorobenzene (ACGIH, 1982), and NIOSH classified 1t as a Group III
pesticide. Group III pesticides are less toxic than Group II pesticides and
the recommended criteria for workplace standards are less stringent than
those recommended for Group II pesticides (NIOSH, 1978). The British
Journal of Industrial Medicine reported a provisional operational limit of
25 ppm for 1,2,4-tr1chlorobenzene (Verschueren, 1977). The 1971 TLV for
3
1,2,3-tr1chlorobenzene 1s 1.3 ppm [10 mg/m (n.s.1.)] for the USSR
(Verschueren, 1977).
TMchlorobenzenes have been designated by the ITC as TSCA Section 4(e)
priority chemicals (44 FR 70666). Preliminary Assessment Information Manu-
facturers Reports were to be submitted to the EPA Office of Toxic Substances
by November 19, 1982, for each of the tMchlorobenzenes (40 CFR 712).
13.6.1.4. TETRACHLOROBENZENES AND PENTACHLOROBENZENE — There are no
occupational workplace standards or recommended criteria for standards,
United States or foreign, for the tetrachlorobenzenes or pentachlorobenzene.
These chlorobenzenes have been designated as TSCA Section 4(e) priority
chemicals (44 FR 70666). Preliminary Assessment Information Manufacturers
Reports were required on 1,2,3,4-tetra-, 1,2,3,5-tetra- and 1,2,4,5-tetra-
chlorobenzene and pentachlorobenzene (40 CFR 712).
13-51
-------
13,6.1.5. HEXACHLOROBENZENE -- Workplace standards have not been
established 1n the United States. The USSR has established a TLV of 0.08
3
ppm (0.9 mg/m ) (Verschueren, 1977). NIOSH classified hexachlorobenzene
as a Group II pesticide and recommended criteria for standards (NIOSH, 1978).
13.6.2. Transportation Regulations. The Department of Transportation
(DOT), the Coast Guard and the Departments of Commerce and Energy regulate,
1n varying degrees, the transport of the chlorinated benzenes.
All of the chlorinated benzenes are regulated under the Hazardous
Material Transportation Act (HHTA) as amended by the Comprehensive Environ-
mental Response Compensation and Liability Act (CERCLA), I.e., "Superfund"
Act (49 CFR 172.101, 46 FR 17738). The HMTA, administered by DOT, specifies
the requirements to be observed 1n the preparation for Interstate shipment
and transport of hazardous materials (46 CFR 171-179). CERCLA further
classified the chlorinated benzenes as hazardous substances and provides
that common carriers of hazardous substances may be held liable for releases
of hazardous substances 1n amounts equal to or greater than the reportable
quantity (RQ). The RQs for mono-, 1,2-dl- or l,4-d1chlorobenzene are set at
100 pounds (45.4 kg) (49 CFR 172.101). RQs for the remaining chlorinated
benzenes have been set at 1 pound pending establishment of different RQs by
EPA (46 CFR 17738).
DOT has designated monochlorobenzene as a flammable liquid. The maximum
net quantity permitted 1n 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/package (49 CFR 172.101).
The U.S. Coast Guard regulates the transport of hazardous materials
while aboard vessels. Title 46, Part 150, specifies the compatibility of
cargoes and operating requirements for bulk liquid hazardous waste cargoes;
13-52
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^.e,, tnonochlorobenzene, dlchlorobenzene (no Isomer specified) and 1,2,4-
trlchlorobenzene (46 CFR 150). Part 153 prescribes the safe handling
procedures for self-propelled vessels carrying hazardous liquids; I.e.,
mono- and dlchlorobenzene (no Isomer specified) (46 CFR 153). Part 151
details the minimum requirements for unmanned tank barges carrying bulk
dangerous cargoes; I.e., monochlorobenzene (46 CFR 151),
The International transport of hazardous materials 1s regulated by the
International Maritime Dangerous Goods Code (IHCO) and administered by DOT
1n this country. Mono-, l,2-d1- and l,4-d1chlorobenzene are regulated under
the IHCO code (46 FR 29392, 49 CFR 172.102).
The export of the chlorobenzenes (I.e., mono-, l,2-d1-, l,4-d1, 1,2,3-
tr1-, l,2,4-tr1- and hexachlorobenzene) 1s regulated by the DOT via the
departments use of the Commodity Control List (15 CFR 399). The DOE
regulates the Import of oil and petrochemicals, and mono-, d1~ and tetra-
chlorobenzene (no Isomers specified) are classified as petrochemicals under
the authority of the 011 Import Regulations (10 CFR 213).
13.6.3. Solid Waste Regulations. Under the Solid Waste Disposal Act as
amended by the Resources Conservation and Recovery Act (RCRA), EPA has
designated mono-, l,2-d1-» l,3-d1~, l,4-d1-, 1,2,4,5-tetra-, penta- and
hexachlorobenzene as hazardous wastes (40 CFR 261.33); subject to the
disposal and permit regulations of Title 40 Code of Federal Regulations,
Parts 262-265 and Parts 122-124 (40 CFR 261). All of the chlorinated
benzenes are designated as hazardous constituents of hazardous wastes from
specific sources subject to RCRA disposal regulations (40 CFR 261.32).
Table 13-21 shows these specific wastes 1n relation to the chlorinated
benzenes. Monochlorobenzene and l,2-d1chlorobenzene, as spent halogenated
solvents, and their still bottoms from the recovery of these solvents, are
13-53
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TABLE 13-21
The Chlorinated Benzenes as Constituents
of Hazardous Wastes from Specific Sources*
EPA Hazardous
Waste No.
Hazardous Waste
Hazard
Constituent
KOI 5
K016
KOI 8
K030
K042
K085
K105
St1-ll bottoms from the distillation
of benzyl chloride
Heavy ends or distillation residues
from the production of carbon
tetrachlorlde
Heavy ends from the fractlonatlon
column In ethyl chloride production
Column bottoms or heavy ends from
the combined production of tr1-
chloroethylene and perchloroethylene
Heavy ends or distillation residues
from the distillations of tetra-
chlorobenzene 1n the production of
the 2,3,5- Isomer
Distillation or fractlonatlon column
bottoms from the production of
chlorobenzene
Separated aqueous stream from the
reactor product washing step 1n the
production of chlorobenzene
Monoch1oroben zene
Hexach1oroben zene
Hexachlorobenzene
Hexach1oroben zene
l,2-D1chloro- and
hexachlorobenzene
Dlchlorobenzenes,
trlchlorobenzenes,
tetrachlorobenzenes,
pentachlorobenzene,
hexachlorobenzene
Monochlorobenzene,
dlchlorobenzenes
*Source: 40 CFR 261.32
13-54
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also regulated for disposal under RCRA (Hazardous Waste No. F002) provided
that the combined concentrations of the spent solvent 1n the resulting
mixture 1s no greater than 25 ppm (46 FR 56582, 40 CFR 261.31).
13.6.4. Food Tolerances. Food tolerances have been established for mono-
chlorobenzene and hexachlorobenzene,
13.6.4.1. HONQCHLOROBENZENE — Monochlorobenzene 1s exempted from the
requirement of a tolerance when used 1n accordance with good agricultural
practices as an Ingredient In pesticide formulation applied to growing crops
only. Permitted uses are as a solvent or cosolvent 1f monochlorobenzene
contains not more than 1% Impurities. Under FIFRA, use of monochlorobenzene
1s prohibited after edible parts of plants begin to form. The grazing of
livestock In treated areas 1s prohibited within 48 hours after application
[40 CFR 180,10Ql(d)].
The FOA permits the use of polysulfone resins and polycarbonate resins
as articles or components of articles for use 1n producing or holding food.
These resins are permitted to contain 500 ppm of monochlorobenzene as a
residual solvent 1n finished (basic) resin (21 CFR 177.1580, 21 CFR
177.2500). Monochlorobenzene as a component of adheslves used 1n the
packaging of food 1s also regulated by FDA (21 CFR 175.105).
13.6.4.2. HEXACHLOROBENZENE — USDA regulates the use of hexachloro-
benzene as a seed treatment for the control of wheat bunt (smut) under the
Federal Seed Act (7 CFR 201).
13.6.5. Water Regulations. Under Section 311(b)(2)(A) of the Federal
Water Pollution Control Act, EPA designated monochlorobenzene, dlchloroben-
zene (no Isomer specified), l,2-d1- and 1,4-d1chlorobenzene as hazardous
substances (40 CFR 116.4) and established an RQ of 100 pounds (45.4 kg) for
these chlorinated benzenes (40 CFR 117.3). Discharges equal to or greater
13-55
-------
than the RQ Into or upon United States waters are prohibited unless the
discharge 1s 1n compliance with applicable permit programs (40 CFR 117.11).
Under the Clean Water Act, Section 307(a), EPA has designated chlori-
nated benzenes (other than dlchlorobenzene) and dlchlorobenzenes (all
Isomers) as toxic pollutants, I.e., priority pollutants (40 CFR 401.15).
Effluent limitation guidelines, new source performance standards, and pre-
treatment standards have been developed or will be developed for the prior-
ity pollutants for 21 major Industries. Specific definitions for classes
and categories are set forth 1n 40 CFR, Parts 402 through 699.
Under the Clean Water Act, Ambient Water Quality Criteria for chlori-
nated benzenes have been developed (U.S. EPA, 1980a,b). These are summar-
ized 1n Tables 13-22 and 13-23. The USSR 1n 1971 established a drinking
water standard for monochlorobenzene of 0.02 mg/s,, and an organoleptlc
limit for l,2-d1- and 1,4-d1chlorobenzene has been set at 0.002 mg/l
(Verschueren, 1977).
13.6.6. A1r Regulations. Ambient air quality standards for the chlori-
nated benzenes have not been established 1n the United States. Maximum
1mm1ss1on concentration (MIC) and maximum emission concentration (NEC)
standards have been established 1n several European countries for mono-
chlorobenzene, 1,2- and 1,4-d1chlorobenzene.
Maximum emm1ss1on concentration standards; I.e., ambient air quality
standards, are used for calculating the minimum stack heights permitted by
law. Dispersion of emitted compounds must be such that the addition of
these compounds to ground level concentrations does not result 1n the
ambient air quality standard being exceeded more frequently than the allowed
percentage. The MEC 1s the maximum concentration of a specific pollutant 1n
emitted gases. MECs are derived from ambient air quality standards by
taking Into account the dispersion phenomena (Verschueren, 1977).
13-56
-------
TABLE 13-22
Ambient Hater Quality Criteria
for Chlorinated Benzenes—Aquatic L1fea
Chlorinated
Aquatic Life Benzenes0 Olchlorobenzenes
Freshwater aquatic life
Acute toxldty 250 vg/fcC 1120
Chronic toxldty — d»e 763 vg/ac
Saltwater aquatic life
Acute toxldty 160 vg/ic 1970 vg/stc
Chronic toxldty 129
aSource: U.S. EPA, 1980a»b
^Includes all of the chlorinated benzenes except the dlchlorobenzenes
Clox1c1ty would occur at lower concentrations among species that are more
sensitive than those tested.
dNo data available
6Tox1c1ty occurs at concentrations as low as 50 jig/a, for fish species
exposed for 7.5 days.
13-57
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TABLE 13-23
I
tn
oo
Monochlorobenzene
Dlchlorobenzenes
Trlchlorobenzenes
1,2,4,5-Tetra-
chlorobenzene
Pentachlorobenzene
Hexachlorobenzene
Ambient Water Quality Criteria for the
Chlorinated Benzenes for the Protection of Human Health3
From Toxic Properties
Ingested Through:
Water and Contaminated
Compound Contaminated Aquatic
Aquatic Organisms Organisms Alone
Based on
Available:
Tox1c1ty Organoleptlc
Data Data
From the
Potential:
Carcinogenic
Effects
400 yg/a
__c
38 yg/a
74 yg/a
2.6 mg/a
__c
48 yg/a
85 yg/a
488 yg/a 20 yg/ab
aSource: U.S. EPA, 1980a,b
bOrganolept1c data as a basis for establishing a water quality criteria have limitations and have no
demonstrated relationship to potential adverse human health effects.
C0ue to Insufficient data, a criterion was not derived.
dBased on the nonthreshold assumption, however, a zero level may not be attainable at the present time
and, therefore, levels that may result 1n Incremental Increases of cancer risk of the lifetime were esti-
mated at 10~s, 10~6 and 10~7. The corresponding recommended criteria are 7.2 ng/a, 0.72 ng/a
and 0.072 ng/a, respectively; 1f estimates are for consumption of aquatic organisms only, the levels
are 7.4 ng/a, 0.74 ng/a and 0.074 ng/a.
-------
13.6.6.1. MQNOCHLOROBENZENE — Ambient air quality standards for
monochlorobenzene have been established 1n five countries and are shown 1n
Table 13-24. In addition, HEC limits have been established 1n the Federal
Republic of Germany; should emissions exceed 3 kg/hour, then a concentration
3
of 150 mg/m cannot be exceeded (Verschueren, 1977).
13.6.6.2. DICHLOROBENZENES -- Maximum emission concentration limits
for 1,2- and l,4-d1chlorobenzene have been established 1n the Federal
Republic of Germany. The HEC limits set were the same as those established
for monochlorobenzene: 150 mg/m3 1f emissions are >3 kg/hour
(Verschueren, 1977).
13-59
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TABLE 13-24
Haxlmum Imm1ss1on Concentration Standards for Monochlorobenzene*
Country
USSR
German Democratic
Republic
Bulgaria
Federal Republic of
Germany-VDI (Assoc.
of German Engineers)
Yugoslavia
mg/m^
0.100
0.3
0.1
15.0
0.1
HICS
Average
ppm Time
20 m1n
30 m1n
0.02 20 m1n
3.0 30 mln
0.02 30 mln
MIC,
mg/rn^ ppm
0.100 —
0.1
0.1 0.02
5.0 1.0
0.1 0.02
Average
Time
24 hr
24 hr
24 hr
30 m1n
24 hr
*Source: Verschueren, 1977
13-60
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APPENDIX A
Comparison Among Different Extrapolation Models
Four models used for low-dose extrapolation, assuming the Independent
background, are:
Multistage: P(d) = 1 - exp [-(q^d + ... + qkdk)]
where q, are non-negative parameters.
A + B ln(d)
ProbU: P{d) = J f{x) dx
- CO
where f{.) 1s the standard normal probability density function.
Welbull: P(d) = 1 - exp [-bdk]
where b and k are non-negative paramters.
One-hit: P(d) = 1 - exp [-bd]
where b Is a non-negative parameter.
The maximum likelihood estimates (HLE) of the parameters 1n the
multistage and one-hit models are calculated by means of the program
GLOBAL82, which was developed by Howe and Crump (1982). The MLE estimates
of the parameters 1n the problt and Welbull models are calculated by means
of the program RISKS!, which was developed by Kovar and Krewskl (1981).
Table A-l presents the MLE of parameters 1n each of the four models.
A-l
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TABLE A-1
Maximum Likelihood Estimate of the Carcinogenic Risk for HCB Using the Four Extrapolation Models
Based on Hepatocellular Carcinomas 1n Female Rats*
(mg/kg/day)
Basis of Interspecles
Extrapolation
Body weights
Body surface area
Multistage
qi = 2.20 x 10"1
q2 = 5.01 x 10~5
qi = 1.35
q2 = 1.90 x 10~3
A
B
A
B
Problt
* -1.35
= 1.12
= 6.70 x NT*
- 1.12
b =
k =
b =
k =
Welbull One-hit
2.20 x ID'1 b = 2.20 x 10~i
1.00
1.35 b . 1.35
1.00
*Source: Lambrecht, 1983
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