United States
Environmental Protection
Agency
Environmental Criteria and
Assessment Office
Cincinnati OH 45268
EPA/600/8-86/018A
June 1986
Review Draft
Research and Development
&ERA
Health Assessment
Document for
Polychlorinated
Dibenzofurans
Review
Draft
(Do Not
Cite or Quote)
NOTICE
This document is a preliminary draft. It has not been formally
released by the U.S. Environmental Protection Agency and
should not at this stage be construed to represent Agency policy.
It is being circulated for comments on its technical accuracy and
policy implications.
-------
(Do Not EPA/600/8-86/018A
Cite or Quote) June 1986
Review Draft
Health Assessment Document for
Polychlorinated Dibenzofurans
NOTICE
This document is a preliminary draft It has not been formally
released by the U S Environmental Protection Agency and
should not atthis stage be construed to represent Agency policy
It is being circulated for comment son its technical accuracy and
policy implications
Environmental Criteria and Assessment Office
Office of Health and Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268
-------
DISCLAIMER
This report 1s an external draft for review purposes only and does not
constitute Agency policy. 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 has prepared this
Health Assessment Document on polychloHnated dlbenzofurans at the request
of the Office of Air Quality Planning and Standards.
In the development of this assessment document, the scientific litera-
ture has been Inventoried, key studies have been evaluated, and summary and
conclusions have been prepared such that the toxldty of polychlorlnated
dlbenzofurans 1s qualitatively and, where possible, quantitatively Identi-
fied. Observed effect levels and dose-response relationships are discussed
where appropriate In order to Identify the critical effect and to place
adverse health responses 1n perspective with observed environmental levels.
This document was reviewed by a panel of expert scientists during the
peer review workshop held at U.S. EPA, Cincinnati, OH, on May 29 and 30,
1986.
111
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DOCUMENT DEVELOPMENT
Debdas Mukerjee, Ph.D., Document Manager and Project Officer
Environmental Criteria and Assessment Office, Cincinnati
U.S. Environmental Protection Agency
Authors
Chapter 1: Introduction, D. Mukerjee, Ph.D.
Chapter 2: Physical and Chemical Properties, S.S. Que Hee, Ph.D.
Chapter 3: Sampling and Analysis, S.S. Que Hee, Ph.D.
Chapter 4: Sources 1n the Environment, S.S. Que Hee, Ph.D.,
S. Garattlnl, M.D. and A. JoM, Ph.D.
Chapter 5: Environmental Fate, Transport and Distribution,
S. Garattlnl, M.D., A. Jor1, Ph.D. and S.S. Que Hee, Ph.D.
Chapter 6: Environmental Levels and Exposure, S.S. Que Hee, Ph.D.,
S. Garattlnl, M.D. and A. Jorl, Ph.D.
Chapter 7: Toxlcologlcal Effects 1n Man and Animals
Sections 7.1. through 7.4. and 7.8. — S. Garattlnl, M.D.
A. JoM, Ph.D.
Section 7.5. — G.L. Klmmel, Ph.D.
Section 7.6. -- S. Rosenthal, Ph.D.
Section 7.7.1. — S. Garattlnl, M.D.
A. JoM, Ph.D.
C. Hlremath, Ph.D.
Section 7.7.2. — D. Bayllss, M.S.
Chapter 8: Effects of Major Concern and Health Risk Assessment,
D. Mukerjee, Ph.D.
1v
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PolychloMnated Dlbenzofurans Peer Review Panel Members
May 29 and 30, 1986
Cincinnati, Ohio
Co-chairmen:
Dr. Jerry F. Stara
Environmental Criteria and Assessment Office
U.S. Environmental Protection Agency
Cincinnati, Ohio
Dr. Debdas Mukerjee
Environmental Criteria and Assessment Office
U.S. Environmental Protection Agency
Cincinnati, Ohio
Members
Dr. Linda Blrnbaum
Systemic Toxicology Branch
Natl. Institute of Environmental Health
Sciences
Research Triangle Park, NC
Mr. David L. Bayllss
Carcinogen Assessment Group
Office of Health and Environmental
Assessment
U.S. Environmental Protection Agency
Washington, DC
Dr. G. Choudhary
Division of Physical Sciences and
Engineering
National Institute for Occupational
Safety and Health
Cincinnati, OH
Dr. Thomas A. Gas1ew1cz
Dept. of Radiation Biology and
Biophysics
University of Rochester
Rochester, NY
Dr. AUstalr W.M. Hay
Department of Chemical Pathology
Old Medical School
The University of Leeds
Leeds, United Kingdom
Dr. Charallngayya B. Hlremath
Carcinogen Assessment Group
Office of Health and Environmental
Assessment
U.S. Environmental Protection Agency
Washington, DC
Dr. Otto Hutzlnger
Lehrstuhl fur Okologlsche Chemle
und Geochemle
Un1vers1tat Bayreuth
Bayreuth, Federal Republic of
Germany
Dr. Armanda Jor1
InstHut d1 Rlcerche Farmaco-
loglche "Mario Negrl"
Mllano, Italy
Dr. Gary Klmmel
Reproductive Effects Assessment
Group
Office of Health and Environmental
Assessment
U.S. Environmental Protection Agency
Washington, DC
Dr. Edo D. PelHzzaM
Analytical and Chemical Sciences
Research Triangle Institute
Research Triangle Park, NC
Dr. Shane S. Que Hee
Ketterlng Laboratory
Department of Environmental Health
University of Cincinnati Medical
Center
Cincinnati, OH
Dr. Chrlstoffer Rappe
Department of Organic Chemistry
Umea Un1vers1tet
Umea, Sweden
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Members (cont.)
Dr. John J. Ryan
Health Protection Branch/Food Division
Health and Welfare — Canada
Ottawa, Ontario, Canada
Dr. Steven H. Safe
Department of Veterinary Physiology
and Pharmacology
College of Veterinary Medicine
Texas A&M University
College Station, TX
Dr. Arnold J. Schecter
Department of Preventive Medicine
Upstate Medical Center, College of
Medicine
Blnghamton, NY
Dr. RHa Schoeny
Environmental Criteria and
Assessment Office
U.S. Environmental Protection Agency
Cincinnati, OH
Dr. Stanlslaw Tarkowskl
World Health Organ1zat1on/EMRO
Copenhagen, Denmark
Dr. Michael L. Taylor
Brehm Laboratory
Wright State University
Dayton, OH
Only written comments were sent by the following reviewers:
Dr. Jerry N. Blancato
Exposure Assessment Group
Office of Health and Environmental
Assessment
U.S. Environmental Protection Agency
Washington, DC
Dr. David H. Cleverly
Office of A1r Quality Planning and
Standards
U.S. Environmental Protection Agency
Washington, DC
Mr. William G. Ewald
Environmental Criteria and Assessment
Office
U.S. Environmental Protection Agency
Research Triangle Park, NC
Dr. Fred Hlleman
Monsanto Company
St. Louis, MO
Dr. Charles H. Nauman
Exposure Assessment Group
Office of Health and Environmental
Assessment
U.S. Environmental Protection Agency
Washington, DC
Dr. Ellen Sllbergeld
Environmental Defense Fund
Washington, DC
Dr. David Stalling
Columbia National Fisheries Research
Laboratory
U.S. Fish and Wildlife Service
Columbia, MO
vl
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TABLE OF CONTENTS
Page
1. INTRODUCTION 1-1
2. PHYSICAL AND CHEMICAL PROPERTIES 2-1
2.1. SUMMARY ; 2-1
2.2. INTRODUCTION 2-2
2.3. PHYSICAL PROPERTIES 2-3
2.3.1. Molecular Structure 2-3
2.3.2. Melting Points 2-3
2.3.3. Solubility 2-15
2.3.4. Partition Coefficients 2-15
2.3.5. Boiling Points and Vapor Pressures 2-15
2.3.6. Adsorption and Desorptlon 2-15
2.3.7. Miscellaneous Properties 2-19
2.3.8. Spectroscoplc Properties 2-19
2.4. CHEMICAL REACTIONS 2-22
2.4.1. General 2-22
2.4.2. PhotodecomposHlon 2-28
2.4.3. Pyrolysls 2-33
2.5. SYNTHESIS 2-37
2.5.1. General Methods 2-37
2.5.2. Methods for PHDFs 2-38
3. SAMPLING AND ANALYTICAL METHODS 3-1
3.1. SUMMARY 3-1
3.2. SAMPLING METHODS 3-2
3.2.1. Incinerators 3-2
3.3. GC METHODS FOR THE PCDFs 3-4
3.3.1. Packed Column GC 3-4
3.3.2. Representative Sample Preparations and
Methods of Analysis 3-9
3.4. GC/MS METHODS FOR PBDFs 3-44
3.5. OTHER METHODS FOR PCDFs 3-46
vll
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TABLE OF CONTENTS (cont.)
Page
4. SOURCES IN THE ENVIRONMENT 4-1
4.1. SUMMARY 4-1
4.2. SOURCES OF PCDFs FROM CHEMICALS 4-1
4.2.1. Chlorinated Phenols 4-2
4.2.2. PCDFs 1n Polychlorlnated Blphenyls 4-10
4.2.3. PCDFs In Phenoxy Herbicides 4-19
4.2.4. PCDFs In Chlorodlphenyl Ether Herbicides
(PCDPE) 4-23
4.2.5. PCDFs 1n Hexachlorobenzene 4-25
4.2.6. PCDFs 1n Hexachlorophene 4-25
4.2.7. PCDFs In PBBs 4-25
4.2.8. PCDFs 1n Metal Salts 4-26
4.3. PHOTOCHEMICAL PRODUCTION 4-26
4.4. SOURCES OF PCDF FROM BURNING AND OTHER HIGH
TEMPERATURE PROCESSES 4-29
4.4.1. Thermal Degradation of Technical Products .... 4-29
4.4.2. Incineration of Municipal Waste 4-37
4.4.3. Sewage Sludge Combustion 4-50
4.4.4. Electrical Equipment Fires 4-51
4.4.5. Incineration of Hazardous Waste 4-51
4.4.6. Car Exhaust 4-54
4.4.7. Other Sources 4-54
5. ENVIRONMENTAL FATE, TRANSPORT AND DISTRIBUTION 5-1
5.1. SUMMARY 5-1
5.2. INTRODUCTION 5-1
5.3. ENVIRONMENTAL TRANSPORT 5-2
5.3.1. Air 5-2
5.3.2. Water 5-4
5.3.3. Soil 5-4
5.4. ENVIRONMENTAL TRANSFORMATION 5-5
5.4.1. Abiotic Transformation 5-5
5.4.2. Blotransformatlon 5-5
5.5. BIOACCUMULATION 5-6
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TABLE OF CONTENTS (cont.)
Page
6. ENVIRONMENTAL LEVELS AND EXPOSURE 6-1
6.1. SUMMARY 6-1
6.2. ENVIRONMENTAL LEVELS 6-1
6.2.1. Introduction. 6-1
6.2.2. In Soil and Sediments 6-2
6.2.3. In Plants 6-5
6.2.4. In Aquatic Organisms 6-8
6.2.5. In Birds 6-16
6.2.6. In Terrestrial Animals 6-17
6.3. EXPOSURE 6-18
6.3.1. Food 6-18
6.3.2. Air 6-20
6.3.3. Other Exposure Routes 6-20
7. TOXICOLOGICAL EFFECTS IN MAN AND EXPERIMENTAL ANIMALS 7-1
7.1. SUMMARY OF EFFECTS OF POLYCHLORINATED DIBENZOFURANS. ... 7-1
7.1.1. Tox1cok1net1cs 7-1
7.1.2. Toxlclty 7-3
7.1.3. Epidemiology 7-5
7.2. INTRODUCTION 7-6
7.3. TOXICOKINETICS 7-7
7.3.1. Introduction 7-7
7.3.2. Absorption 7-7
7.3.3. Distribution, Metabolism and Excretion 7-8
7.3.4. Metabolic Fate 7-42
7.4. ACUTE, SUBACUTE AND CHRONIC TOXICITY 7-47
7.4.1. Acute Toxlclty 7-47
7.4.2. Subacute Toxlclty 7-49
7.4.3. Chronic Toxlclty 7-53
7.4.4. Contribution of PCDF to the Toxlclty of
Industrial Products and Environmental
Contaminants 7-53
7.4.5. Some Particular Aspects of the Toxic Reactions. . 7-59
1x
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TABLE OF CONTENTS (cont.)
Page
7.5. REPRODUCTIVE/DEVELOPMENTAL TOXICITY 7-78
7.5.1. Introduction 7-78
7.5.2. Development 7-78
7.5.3. Reproductive Function/Fertility 7-84
7.5.4. Summary 7-85
7.6. MUTAGENICITY 7-86
7.7. CARCINOGENICITY 7-87
7.7.1. Animal 7-87
7.7.2. Human 7-88
7.8. EPIDEMIOLOGY -- SYSTEHIC TOXIC EFFECTS 7-89
7.8.1. The Japanese Incident (Yusho disease) 7-90
7.8.2. The Taiwan Incident ("Yu-Cheng" disease) 7-93
7.8.3. The Blrmlnghamton, NY Transformer Accident. . . . 7-95
8. EFFECTS OF MAJOR CONCERN AND HEALTH HAZARD ASSESSMENT 8-1
8.1. EXISTING GUIDELINES AND STANDARDS 8-1
8.2. CURRENT LEVELS OF EXPOSURE 8-1
8.3. HIGH RISK POPULATIONS 8-2
8.4. BASIS AND DETERMINATION OF RISK 8-3
8.4.1. Estimation of LOAELs 8-4
8.4.2. Derivation of Reference Dose (RfD) 8-6
9. REFERENCES 9-1
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LIST OF TABLES
No. Title Page
2-1 Chemical Abstracts System Registry Numbers (CAS-RN) for
PHDFs 2-4
2-2 X-Ray Crystallographlc Indices of Some Selected PHDFs .... 2-9
2-3 Melting and Boiling Points of Some PHDFs 2-10
2-4 The Logarithm of Octanol/Water Partition Coefficients (Kow)
of Some PHDFs from HPLC Methods 2-16
2-5 Vapor Densities and Pressures of PCDFs as Estimated from
a Gas Chromatographlc Method 2-17
2-6 Ultraviolet Absorption Maxima and Molar Absorptlvltles
for Some Substituted PBDFs 2-20
2-7 Absorption Maxima of Some PCDFs In Chloroform 2-21
2-8 Proton NMR Data for Some PCDFs 2-23
2-9 FMedel-Crafts Acetylatlon of Substituted PHDFs 2-29
2-10 ChloMnatlon of TrCDF with Antimony Pentachlor1de/CCl4
Reagent to Produce TCDFs 2-30
2-11 Photodegradatlon of TCDFs (1 yg/ml) at 254 nm After
4 Hours of Irradiation In Hexane or Tetradecane 2-34
2-12 Photodecomposltlon of PeCDFs In Hexane at 254 nm 2-35
2-13 Formation of PCDFs from the Pyrolysls of Specific PCBs. . . . 2-40
2-14 Formation of PCDFs from Pyrolysls of Chlorobenzenes 2-44
2-15 Synthetic Methods for PCDFs using Chlorinated Dlphenyl
Ethers (CDPEs) as Substrates 2-46
3-1 Relative Intensities of Ions Containing from One to Eight
Chlorine Atoms 3-11
3-2 Percent Intensities of the Major Ions In the Mass Spectra
of Isomerlc PCDFs 3-14
3-3 Exact Molecular Weights of the PCDFs 3-16
3-4 Characteristics of Some Glass Capillary Columns for
Separating PCDFs 3-22
x1
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LIST OF TABLES (cont.)
No. Title page
3-5 Relative Retention Times of PCDFs and PCDDs on the
Capillary Columns of Table 3-4 3-24
3-6 Relative Retention Times (RRT) and Relative Response
Factors (RRF) for PCDFs Synthesized as Reference Standards. . 3-29
3-7 Error 1n Estimation of PCDFs by Six Methods 3-42
4-1 Presence of PCDFs 1n Industrial Chemicals and
Their Entry Into the Environment 4-3
4-2 Estimated PCDD and PCDF Release to the Canadian
Environment from Chemical Sources 4-4
4-3 Chemical Analysis of a Typical Pure and Technical
Pentachlorophenol 4-5
4-4 Levels of PCDFs and PCDDs 1n Some Commercial Chlorophenols. . 4-6
4-5 Order of Abundance of PCDFs 1n European Chlorophenols .... 4-9
4-6 PCDF Content 1n Some PCBs and 1n Kaneml 011 4-14
4-7 Suspected Maximum Levels of Toxic PCDFs 1n Various
PCBs and In Kaneml 011 4-16
4-8 PCDFs 1n Commercial PCBs 4-20
4-9 PCDFs and PCDDs 1n Phenoxy Herbicides 4-22
4-10 Levels of PCDDs and PCDFs 1n Commercial Dlphenyl Ether
Herbicides 4-24
4-11 PCDFs and PCDDs In Several Metal Salts 4-27
4-12 Estimated PCDD and PCDF Releases to the Canadian
Environment from Combustion Sources 4-30
4-13 Pyrolysls Products of Aroclor 1254 1n a Quartz Tube for
Pyrolysls Time of 3 Seconds 4-32
4-14 Levels of Trapped PCDFs and PCDDs on Charcoal from
Burning Chlorophenolate-Impregnated Leaves and Wood Wool. . . 4-34
4-15 Production of PCDDs and PCDFs after 0.6 to 1.00 Second
Pyrolysls In Air of 2-Butoxyethyl-2,4,5-TMchloro-
phenoxyacetate 4-36
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LIST OF TABLES (cont.)
No. Title Page
4-16 PCDF Composition of Municipal Incinerator Fly Ash
Resulting from Electrostatic Precipitation 4-39
4-17 PCDF Composition of Stack Effluent of Municipal
Incinerators 4-41
4-18 Influence of Type of Waste on PCDF Formation 4-43
4-19 Annual Toxic Equivalent of CDBFs Emitted by Netherlands
Incinerators In Terms of 2,3,7,8-TCDD Toxic Equivalents . . . 4-47
4-20 Levels of PCDD and PCDF from MSW Incineration, Umea 4-48
4-21 PCDFs Formed During PCB Fires 4-52
6-1 Levels of PCDFs 1n Environmental Media 6-3
6-2 Ranges of 2,3,7,8-TCDD and PCDFs 1n Top Soil and
Biological Samples In ppt fresh weight In a Dutch
Hazardous Waste Dump 6-6
6-3 PCDF Residues In Wildlife 6-10
6-4 PCDF Distribution In Pond Known to be Contaminated
by Aroclor 1260 6-12
6-5 Levels of Toxic PCDF Isomers 1n Wild Animals 6-14
7-1 Percentages of the Total Intake of Original PCDF Isomers
Found 1n the Livers of Monkeys and Rats 7-10
7-2 Concentration of "C-2.3.7.8-TCDF 1n Tissues of Male
Fisher 344 Rats 7-11
7-3 Distribution of Radioactivity 3 Days After Administration
of 14C-2,3,7,8-TCDF to 200-250 g Male Fisher 344 Rats .... 7-13
7-4 Components for the Elimination of Radioactivity from
Tissues of Fisher 344 Rats 7-14
7-5 Excretion of 2,3,7,8-TCDF-Der1ved Radioactivity by
Three Mice 7-17
7-6 Distribution of 2,3,7,8-TCDF-Derlved Radioactivity 1n
Three Rhesus Monkeys 21 Days After Treatment 7-19
7-7 Distribution of 2,3,7,8-TCDF-Der1ved Radioactivity During
the First 24 Hours 1n Liver, Fat and Skin of Bulnea P1gs. . . 7-21
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LIST OF TABLES (cont.)
No. Title Page
7-8 2,3,7,8-TCDF Pharmacoklnetlc Parameters for Physiological
Model 7-24
7-9 Concentrations of PCBs, PCDFs and PCQs 1n the Rice 011s
from Japan and Taiwan 7-26
7-10 Concentrations of PCDF Congeners 1n the Tissues of the
Deceased Patient with Yu-Cheng 1n Taiwan 7-28
7-11 Structural Assignments by PCDF Congeners 1n the Toxic
Rice-Bran 011 Ingested by the Deceased Yu-Cheng Patient . . . 7-29
7-12 PCB and PCDF Concentrations In Liver and Adipose Tissue
of Deceased Yusho Patients 7-30
7-13 Levels of PCDF (pg/g) and PCB (ng/g) 1n Tissue Samples of a
Baby from a Woman from Taiwan Suffering from Yusho Disease. . 7-31
7-14 Relationship Between the Amount of R1ce 011 Used by Yusho
Patients, Amounts of PCBs and PCDFs Ingested, and Clinical
Severity 7-32
7-15 Concentrations of PCBs, PCDFs and PCQs 1n the Tissues of
Yusho Patients, Unexposed Individuals and in the M1lk Fat
of a Worker Occupatlonally Exposed to PCBs 7-34
7-16 Levels of PCDD and PCDF 1n Blood Samples from Workers 1n
the Saw Mill Industry After Exposure to 2,3,4,6-Tetra-
chlorophenolate 7-36
7-17 Levels of PCDD and PCDF 1n Blood Samples from Workers In
the Textile and Leather Industry After Exosure to
Pentachlorophenol (PCP) or PCP Derivatives 7-37
7-18 Levels of PCDF In Tissue Samples of a Worker Exposed to
Phenoxy Herbicides 7-38
7-19 Half-lives and Contents of PCDDs and PCDFs 1n
Human Adipose Tissue 7-40
7-20 PCDD and PCDF Levels 1n Blnghamton, New York Adipose
Tissue Levels (ppt) 1n Exposed and Unexposed Subjects .... 7-41
7-21 Levels (pg/g) of PCDDs and PCDFs 1n Human Samples from
Sweden, U.S.A. and Vietnam Subjects 7-43
7-22 2,3,7,8-ChloMne-Substltuted PCDDs and PCDFs 1n Human
Unexposed Subject 7-44
xlv
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LIST OF TABLES (cont.)
No. Title Page
7-23 Total PCDD and PCDF Levels (pg/g) 1n Tissue Samples from
an Unexposed Subject 7-45
7-24 Levels of PCDDs and PCDFs In Breast M1lk 7-46
7-25 Summary of Pathologic Effects of 2,3,7,8-Tetra-CDBF
1n Rhesus Monkeys 7-50
7-26 Acute Tox1c1ty of 2,3,7,8-TCDF 1n Several Animal Species. . . 7-51
7-27 Subacute Tox1c1ty of PCDFs 1n Laboratory Animals 7-54
7-28 Probable Contributions of PCDF In Aroclor 1260 and Pure
2,4,5,2',4',5'-Hexachlorob1phenyl to the Toxlclty 1n
Rabbits of PCBs Applied at 120 mg/50 cm2 7-57
7-29 Effects of Different PCDF Congeners on Rat Hepatic
Cytosollc Receptor Binding Avidities, AHH and EROD
Induction 1n Rat Hepatoma H-4-II E Cells 7-72
7-30 Signs and Symptoms of Yusho Disease In Adult Japanese .... 7-91
xv
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LIST OF FIGURES
No. Title Page
2-1 Possible Reaction Schemes to Produce PCDFs from the
Pyrolysls of 2,2' ,4,4' ,5,5'-Hexachlorob1phenyl ........ 2-42
3-1 Mass Spectral Behavior of PCOFs ............... 3-12
3-2 Low Resolution Mass Spectra of (A) TrCDF, (B) TCOF,
(C) PeCDF, (D) HxCDF and (E) OCDF .............. 3-17
3-3 Mass Chromatograms of (A) 2,3,4,6-Tetrachlorophenol and
(B) Pentachlorophenol Showing the Elutlon of PCDF ...... 3-19
3-4 Mass Chromatograms (OV-101 Glass Capillary Column at
197°C) of "Yusho" 011 Showing Elutlon of PCDF ........ 3-25
4-1 Comparison of Specific Ion Current Profiles after
Chromatography of "Yusho" 011 ................ 4-18
7-1 A plot of the -log ECso Values for AHH Induction 1n Rat
Hepatoma H-4-IIE Cells vs. the jm vivo -log £059 Values
for Thymlc Atrophy (top) and Body Weight Loss (bottom)
for Several PCDFs, 2,3,7,8-TCDD and a Reconstituted PCDF
Mixture ........................... 7-74
xv1
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1. INTRODUCTION
Polychlorlnated dlbenzofurans (PCDFs) are members of a group of the
widespread and environmentally stable halogenated tMcycHc aromatic hydro-
carbons. This group 1s structurally similar to polychlorlnated blphenys
(PCBs), polychlorlnated napthalenes (PCNs), polychlorlnated d1benzo-£d1ox1ns
(PCDDs), polybromlnated blphenyls (PBBs) and polychlorlnated blphenylenes.
PCDFs are not deliberately disseminated In the environment as primary Indus-
trial products, but enter the environment as unintentional trace Impurities
1n PCBs, chlorinated phenols, PCNs, 2,4,5-T formulations and as a result of
diverse combustion processes. PCDFs have been detected In fly ash and In
flue gas from municipal and Industrial Incinerators, and have been found to
bloaccumulate 1n seal, fish, turtle and human adipose tissue. Residues of
PCDFs have also been detected In human breast milk. As a result of PCDF
detection 1n milk from Swedish mothers 1t has been suggested that primary
PCDF exposure to this population 1s occurring from municipal Incinerators.
Consequently, 1n 1985 the National Swedish Environmental Protection Board
decided on a moratorium for new municipal Incinerators.
PCDFs are extremely toxic to animals and humans. Signs and symptoms of
toxlclty are very similar to those caused by 2,3,7,8-tetrachlorod1benzo-p_-
dloxln. Of the total 135 possible Isomer congeners so far determined for
PCDFs, 2,3,7,8-TCDF and 2,3,4,7,8-PeCDF seem to be the most toxic. However,
the relative toxlclty of all the congeners have not yet been fully studied.
The guinea pig Is the most sensitive species to toxic effects of PCDF. Mice
and rats are less sensitive than guinea pigs. Like PCDDs, these compounds
are strong Inducers of aryl hydrocarbon hydroxylase (AHH) and the most
acutely toxic Isomers are also the most potent Inducers of AHH activity.
1925A 1-1 06/23/86
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Mutagenlclty data on PCDFs are extremely limited and three Isomers
studied gave negative responses In short-term mlcroblal mutagenesls assay.
Recent observations Indicate that 2,3,7,8-TCDF 1s teratogenlc 1n mice
resulting 1n dose-related Increases 1n both Isolated cleft palates and
hydronephrosls 1n the fetus. Animal experimentation data Indicate that
PCOFs are capable of being transferred to the developing offspring, pre-
natally through transplacental migration and postnatally through the nursing
mother's milk. When these two rates of transfer were compared, 1t was found
that PCDF transported through nursing mother's milk appeared to be higher
than transplacental migration to the fetus.
Information on adverse health effects 1n humans has been observed from
the Japanese and Taiwanese population who consumed PCB- and PCDF-contam1-
nated rice oil In 1968 and 1979, respectively. Severe adverse toxic
effects, many of which are similar to those caused by 2,3,7,8-TCDD, were
observed 1n the affected population.
Though PCOFs have been detected 1n a number of chemicals as contaminants
and 1n the emissions from waste Incinerators, a chronic toxldty data base
for this class of compounds 1s not available. Primarily because of their
prevalence In the environment and structural similarities between PCDFs and
PCDDs, there Is concern as to the public health Impact of their presence 1n
the environment.
1925A 1-2 06/30/86
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2. PHYSICAL AND CHEMICAL PROPERTIES
2.1. SUMMARY
The dlbenzofurans (DBFs) are a group of organic compounds that contain
two benzene rings annulated to a central furan ring. The benzene rings can
be substituted at the 1,2,3,4,6,7,8,9 positions. For a single substHuent
like chlorine there are 135 congeners over the substitution range mono- to
octa-: these Include 4 mono-, 16 dl-, 28 tr1-, 38 tetra-, 28 penta-, 16
hexa-, 4 hepta- and 1 octa-.
The crystal structure of the 2,3,7,8-tetrachlorodlbenzofuran (2,3,7,8-
TCDF) Is monocllnlc. The latter 1s a structural analogue of the highly
toxic 2,3,7,8-TCDD. The two benzene rings are coplanar except for the
1,2- and 1,9-subst1tuted derivatives. Most polyhalogenated dlbenzofurans
(PHDFs), Including the polychlorlnated dlbenzofurans (PCDFs), polyfluoMnat-
ed dlbenzofurans (PFDFs), polybromlnated dlbenzofurans (PBDFs) and the poly-
lodlnated dlbenzofurans (PIDFs) are solids at room temperature and have
relatively high boiling points, which Increase with degree of substitution.
PHDFs are generally quite soluble In organic solvents but water solubility
decreases with chlorlnatlon. The octanol/water partition coefficients of
PCDFs Increase with chlorine number; log K values for 2,8-d1chlorod1ben-
zofuran (2,8-DCDF), 2,3,7,8-TCDF, and the octachlorodlbenzofuran (OCDF) are
5.30, 5.82i0.02, and 8.78, respectively. Vapor pressure data for some PCDFs
are available. The less polar the PHDF, the more readily 1t will adsorb to
organic matter.
The PHDFs are aromatic compounds, and thus there are at least two dis-
tinct UV absorption maxima near 217 nm (o* *=• a transition) and near 280
nm (tr*^-ir transition). The effect of ring substitution on absorption
maxima tends to be minor. The proton and 13C-nuclear magnetic resonance
1926A 2-1 06/23/86
-------
(NMR) spectra are characteristic of aromatic molecules. As expected of
aromatic molecules, the parent Ions In the mass spectra are Intense and are
suitable for specific Ion monitoring. The major fragment Ions are M-C1,
M-Clp, M-C1-CO, and M-C1--CO. The major cleavage patterns are determin-
ed by the great stability of the ring system so that the fragmentations tend
to be directed by the substUuents.
The PHDFs exhibit reactions directed by the substUuents of the most
substituted ring towards the least substituted ring.
Higher PCDFs can be degraded by ultraviolet (UV) light. These reactions
are dependent on solvent and wavelength. Degradation Is slow at 310 nm, but
rapid at 254 nm or at 310 nm 1n the presence of suitable triplet sensltlzers.
PHDF can also be produced by photodecomposltlon of higher PHDFs, by
photocycllzatlon or pyrolysls of halogenated dlphenyl ethers, by pyrolysls
of polyhalogenated blphenyls (PHBs) and halobenzenes, by halogenatlon of
lower PHDFs, by palladium acetate cycllzatlon of substituted dlphenyl
ethers, and by cycllzatlon of substituted hydroxylated PHBs, and substituted
dlazotlzed halophenoxy-o-an1lines.
2.2. INTRODUCTION
The accepted system of numbering of PHDFs 1s now as follows:
The ring that 1s most substituted has priority 1n numbering. For equally
substituted rings, the numbering that allows the lowest sum of the sub-
stituted positions 1s the correct one.
1926A 2-2 06/21/86
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There are 135 positional Isomers for a single substltuent: 4 mono-, 16
dl-, 28 tr1-, 38 tetra-, 28 penta-, 16 hexa-, 4 hepta- and 1 octa-. There
has been IHtle Investigation of the physico-chemical properties of such
compounds.
The Chemical Abstracts System Registry Numbers (CAS RN) for the PHDFs
that have been cited In the literature are provided 1n Table 2-1.
2.3. PHYSICAL PROPERTIES
2.3.1. Holecular Structure. The acutely toxic 2,3,7,8-TCDF belongs to
the monocl1n1c space group C2/c and the molecule 1s planar (Table 2-2). A
crystallographlc 2-fold axis bisects the bond between the rings and passes
through the oxygen atom. The longest carbon-carbon bond distance within the
benzenold rings joins the carbon atoms to which the chlorine atoms are
attached (Hubbard et al., 1978). Thus, the compound 1s very closely related
1n structure to the highly toxic 2,3,7,8-TCDD (Boer et al., 1972). No X-ray
data for the other PHDFs are available.
The electronic structures of 25 PCDFs were characterized by Cheney and
Tolly (1979). The powder X-ray diffraction patterns of 4 PCDFs have been
reported by Cantrell et al. (1986). An unspecified MCDF, 3,6-DCDF, 2,3,7,8-
TCDF, and OCDF were examined. The DCDF 1s easily distinguished from OCDF
and the TCDF by Us Intense diffraction at 24.35 2e, and the TCDF by Us
strong line at 15.20 2e. The OCDF can with difficulty be detected by Us
diffraction at 31.81 2e.
2.3.2. Heltlng Points. Table 2-3 gives the melting points of some
PHDFs. Norstrom et al. (1979) showed by gas chromatography/mass spectrome-
try (GC/MS) that the PCDFs for which they determined melting points were
pure. Kurokl et al. (1984) have published melting points for other PCDFs
(Table 2-3).
1926A 2-3 06/21/86
-------
TABLE 2-1
Chemical Abstracts System Registry Numbers
(CAS-RN) for PHDFs
Chemical Substitution
PCDF
Mono mono
2-
3-
4-
D1 d1
1,2-
1,3-
1,4-
1,6-
1,7-
1,8-
1,9-
2,3-
2,4-
2,6-
2,7-
2,8-
3,4-
3,6-
3,7-
4,6-
Tr1 tr1
1,2,3-
1,2,4-
1,2,6-
1,2,7-
1,2,8-
1,2,9-
1,3,4-
1,3,6-
1,3,7-
1,3,8-
1,3,9-
1,4,6-
1,4,7-
1,4,8-
1,4,9-
CAS-RN
42934-53-2
84761-86-4
25074-67-3
74992-96-4
43047-99-0
64126-85-8
94538-00-8
94538-01-9
74992-97-5
94538-02-0
81638-37-1
70648-14-5
64126-86-9
24478-74-8
60390-27-4
74992-98-6
5409-83-6
94570-83-9
74918-40-4
58802-21-4
64560-13-0
43048-00-6
83636-47-9
24478-73-7
64560-15-2
83704-37-4
83704-34-1
83704-38-5
82911-61-3
83704-39-6
64560-16-3
76621-12-0
83704-40-9
82911-60-2
83704-41-0
64560-14-1
70648-13-4
1926A
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06/21/86
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TABLE 2-1 (cont.)
Chemical
Substitution
CAS-RN
PCDF (cont.)
Tr1 (cont.) 2,3,4-
2,3,6-
2,3,7-
2,3,8-
2,3,9-
2,4,6-
2,4,7-
2,4,8-
2,4,9-
2,6,7-
3,4,6-
3,4,7-
3,4,8-
3,4,9-
Tetra tetra
1,2,3,4-
1,2,3,6-
1,2,3,7-
1,2,3,8-
1,2,3,9-
1,2,4,6-
1,2,4,7-
1,2,4,8-
1,2,4,9-
1,2,6,7-
1,2,6,8-
1,2,6,9-
1,2,7,8-
1,2,7,9-
1,2,8,9-
1,3,4,6-
1,3,4,7-
1,3,4,8-
1,3,4,9-
1,3,6,7-
1,3,6,8-
1,3,6,9-
1,3,7,8-
1,3,7,9-
1,4,6,7-
1,4,6,8-
1,4,6,9-
1,4,7,8-
57H7_34_7
57117-33-6
58802-17-8
57117-32-5
58802-18-9
58802-14-5
83704-42-1
54589-71-8
82911-59-9
83704-43-4
83704-43-2
83704-44-3
83704-45-4
83794-46-5
30402-14-03
24478-72-6
83704-21-6
83704-22-7
62615-08-1
83704-23-8
71998-73-7
83719-40-8
64126-87-0
83704-24-9
83704-25-0
83710-07-0
70648-18-9
58802-20-3
83704-26-1
70648-22-5
83704-27-2
70648-16-7
64126-87-0
83704-28-3
57117-36-9
71998-72-6
83690-98-6
57117-35-8
64560-17-4
66794-59-0
82911-58-8
70648-19-0
83704-29-4
1926A
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06/21/86
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TABLE 2-1 (cont.)
Chemical Substitution
PCDF (cont.)
Tetra (cont.) 2,3,4,6-
2,3,4,7-
2,3,4,8-
2,3,4,9-
2,3,6,7-
2,3,6,8-
2,3,7,8-
2,4,6,7-
2,4,6,8-
3,4,6,7-
Penta penta
1,2,3,4,6-
1,2,3,4,7-
1,2,3,4,8-
1,2,3,4,9-
1,2,3,6,7-
1,2,3,6,8-
1,2,3,6,9-
1,2,3,7,8-
1,2,3,7,9-
1,2,3,8,9-
1,2,3,6,7-
1,2,4,6,8-
1,2,4,6,9-
1,2,4,7,8-
1,2,4,7,9-
1,2,4,8,9-
1,2,6,7,8-
1,3,4,6,7-
1,3,4,6,8-
1,3,4,6,9-
1,3,4,7,8-
1,3,4,7,9-
1,3,4,8,9-
2,3,4,6,7-
2,3,4,6,8-
2,3,4,6,9-
2,3,4,7,8-
2,3,4,7,9-
CAS-RN
83704-30-7
83704-31-8
83704-32-9
83704-33-0
57117-39-2
57117-37-0
51207-31-9
57117-38-1
58802-19-0
57117-40-5
30402-15-4
83704-47-6
83704-48-7
67517-48-0
83704-49-8
57H7_42-7
83704-51-2
83704-52-3
57117-41-6
83704-53-4
83704-54-5
83704-50-1
69698-57-3
70648-24-7
58802-15-6
71998-74-8
70648-23-6
69433-00-7
83704-36-3
83704-55-6
70648-15-6
58802-16-7
70648-20-3
70872-82-1
57117-43-8
67481-22-5
83704-35-2
57H7_31_4
70648-21-4
1926A
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06/21/86
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TABLE 2-1 (cont.)
Chemical
Substitution
CAS-RN
PCDF (cont.)
Hexa
Hepta
Octa
"C
PBDF
Mono
D1
hexa
1,2,3,4,6,7-
1,2,3,4,6,8-
1,2,3,4,6,9-
1,2,3,4,7,8-
1,2,3,4,7,9-
1 23489-
1 , C , J,t,O ,3-
1,2,3,6,7,8-
1,2,3,6,7,9-
1,2,3,6,8,9-
1,2,3,7,8,9-
1,2,4,6,7,8-
1,2,4,6,7,9-
1,2,4,6,8,9-
1,3,4,6,7,8-
1,3,4,6,7,9-
2,3,4,6,7,8-
hepta
1,2,3,4,6,7,8-
1,2,3,4,6,7,9-
1,2,3,4,6.8,9-
1,2,3,4,7,8,9-
octa
1,2,3,4,6,7,8,9-
1,2,7,8-
2,3,7,8-
2,3,4,7,8-
1-
2-
3-
4-
2,7-
2,8-
3,7-
55684-94-1
79060-60-9
69698-60-8
91538-83-9
70648-26-9
91538-84-0
92341-07-6
57H7_44_9
92341-06-5
75198-38-8
72918-21-9
67562-40-7
75627-02-0
69698-59-5
71998-75-9
92341-05-4
60851-34-5
38998-75-3
67562-39-4
70648-25-8
69698-58-4
55673-89-7
3268-87-9
39001-02-0
78813-10-2
78813-09-9
87665-66-5
50548-45-3
86-76-0
26608-06-0
89827-45-2
65489-80-7
10016-52-1
67019-91-4
1926A
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06/21/86
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TABLE 2-1 (cont.)
Chemical
PBDF (cont.)
TM
Tetra
Penta
Hepta
PFDF
Mono
D1
TM
Tetra
Octa
PIDF
Mono
D1
Nixed halo
Substitution
1,2,8-
2,3,8-
1,2,7,8-
2,3,7,8-
penta
hepta
2-
3-
2,7-
2,8-
1,2,4-
1,2,3,4-
octa
2-
3-
4-
2,7-
2,8-
3,7-
tr1bromo-2,4,8-tr1chloro
tetrabromo-2,4,8-tr1chloro
CAS-RN
84761-81-9
84761-82-0
84761-80-8
67733-57-7
68795-14-2
62994-32-5
391-46-8
391-54-8
71277-77-5
10016-52-1
15950-24-0
15945-39-8
16804-47-0
5408-56-0
5896-29-7
65344-26-5
81050-43-3
5943-11-3
5914_49_8
54605-27-5
54605-26-4
1926A
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06/24/86
-------
2,3,7,8-TCDF
TABLE 2-2
X-Ray Crystallographlc Indices of Some Selected PHDFs*
Dlbenzofuran
Unit Cell Indices (A)
a b c
Density (g/m/i)
(exptl) (calc'd)
Bond Length
(A)
14.702(4) 12.886(4) 6.256(1)
1.72
1.74
(2,8)C-C1(1):1.725(2)
(3,7)C-C1(2):1.732(2)
€-0:1.385(2)
Benz C-C:1.366(2)
to 1.404(2)
*Source: Hubbard et al., 1978
00
-------
TABLE 2-3
Melting and Boiling Points of Some PHDFs
Dlbenzofuran
— , 1-MBOF
— , 2-MBOF
--, 3-HBDF
— , 4-MBDF
--, 2-MCDF
— . 3-MCDF
— , 2-MIDF
— . 3-MIDF
--. 2-MFDF
--, 3-MFDF
--, 2,8-DBDF
--. 2,8-DCDF
— . 3,7-DCDF
— , 2,8-DIDF
--, 2,3,8-
TrCDF
Melting Point Boiling Point
PC) (°C)
67
no
110 220 (40 mm Hg)
120
120 220 (40 mm Hg)
72
67
106
101
101-102
112
142
89
88.5
195
199-200
191.5-192.5
185
184-185
186-188
77
180-181
189-191
Reference
Coffey, 1973
Coffey, 1973
Weast, 1985
Coffey, 1973
Weast, 1985
Coffey, 1973
Weast, 1985
Coffey, 1973
Coffey, 1973
Kurokl et al., 1984
Coffey, 1973
Coffey, 1973
Coffey, 1973
Coffey, 1973
Coffey, 1973
Weast, 1985
Oshlma et al . ,
1968c
Gllman and Young,
1934
Kurokl et al., 1984
Norstrom et al.,
1979
Coffey, 1973
Norstrom et al.,
1979
Gray et al., 1976
1926A
2-10
06/23/86
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TABLE 2-3 (cont.)
Dlbenzofuran
Melting Point Boiling Point
Reference
-, 2,4,6-
TrCDF
-,2,4,8-
TrCDF
--, 1,2,4-
TrCDF
—,1.3,6,8-
TCOF
-,1,3,6,7-
TCOF
-.1,3,7,9-
TCDF
-.1,4,6,7-
TCDF
--,2,3,6,8-
TCDF
-.2,4,6,7-
TCDF
-,1,2,7,8-
TCDF
-.1,2,6,7-
TCDF
-1,2,7,9-
TCDF
-,2,3,7,8-
TCDF
-.2,3,6,7-
TCDF
-.2,4,6,8-
TCDF
116-117
155-156
100-101
177-178
176.5-177
206.5-207.5
180-181
197-198
202-203
164-164.5
210-211
199-200
184-185
219-221
226-228
227-228
195-196
192-192.5
198-200
Gray et al., 1976
Kurokl et al., 1984
Coffey, 1973
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Gray et al., 1976
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Norstrom et al.,
1979
Gray et al., 1976
Kurokl et al., 1984
Kurokl et al., 1984
Gray et al., 1976
1926A
2-11
06/21/86
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TABLE 2-3 (cent.)
Dibenzofuran
-.1.2,3,7-
TCDF
-.1.2,3,8-
TCDF
-,2,3,4,8-
TCDF
-.2,3,4,7-
TCDF
-.2,3,4,6-
TCDF
-.1.2,4,8-
TCDF
-1.2,3,4-
TCDF
-.1,2,4,6,8-
PeCDF
-,1,2,4,7,8-
PeCDF
-.1,2,4,7,9-
PeCDF
-.1,2,3,7,8-
PeCDF
-.1,2.3,6,7-
PeCDF
-,1,2,6,7,8-
PeCDF
-,2,3.4,7,8-
PeCDF
-.2,3,4,6,7-
PeCDF
Melting Point
167.5-168
197-198
176-177
172.5-173
153-154
191-193
168.5-169
169-170
204-205
236-238
225.5-226.5
234-235
196-197
225-227
205-207
220-221
196-196.5
201.5-202
Boiling Point Reference
Kurokl et al.,
Kurokl et al..
Kurokl et al.,
Kurokl et al.,
Kurokl et al.,
Kurokl et al.,
Kurokl et al.,
Coffey, 1973
Kurokl et al.,
1984
1984
1984
1984
1984
1984
1984
1984
Kurokl et al., 1984
Norstrom et al.,
1979
Gray et al., 1976
Kurokl et al.,
Kurokl et al.,
Kurokl et al.,
Kurokl et al.,
Kurokl et al.,
Kurokl et al..
1984
1984
1984
1984
1984
1984
1926A
2-12
06/21/86
-------
TABLE 2-3 (cont.)
Dlbenzofuran
Melting Point Boiling Point
Reference
-.2,3,4,6,8-
PeCDF
-,1,3,4,7,8-
PeCDF
-.1,3,4,6,7-
PeCDF
-.1,2,3,4,6-
PeCDF
-,1,2,4,6,7-
PeCDF
-.1,2,3,4,8-
PeCDF
-.1,2,4,6,7,8-
HxCDF
-,1,3,4,6,7,8-
HxCDF
-.1,2,4,6,8,9-
HxCDF
-,1,2,3,4,6,7-
HxCDF
-.1,2,3,4,7,8-
HxCDF
-,1,2,3.6.7.8-
HxCDF
-,1,2,3,4,6,7,8-
HxCDF
-,1,2,3,6,8,9-
HxCDF
-.1,2,4,6,7,9-
HxCDF
219-220
168-170
195-195.5
194-195
178.5-179.5
177-178
221-222
229-230
247-249
246-248
227-228
225.5-226.5
232-234
239-240
206-207
180-181
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Norstrom et al.,
1979
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
1926A
2-13
06/21/86
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TABLE 2-3 (cont.)
Dlbenzofuran
Melting Point Boiling Point
Reference
-.1,2,3,4.6,8-
HxCDF
-,1,2,3,4,6,9-
HxCDF
-,1,2,3,4,7,9-
HxCDF
-.1,2,3,4,6.7,8-
HpCDF
-.1,2,3,4,6,8,9-
HpCDF
-.1,2,3.4,7,8,9-
HpCDF
—, OFDF
233.5-234
196-197
216-217
236-237
211-212
221-223
100
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Kurokl et al., 1984
Coffey, 1973
1926A
2-14
06/21/86
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2.3.3. Solubility. PHDFs are less polar than dlbenzofuran and would be
expected to be less soluble 1n water and more soluble 1n organic solvents.
Research needs to be done 1n this area. No solubility data even for the
PCDFs have been reported. However, 2,3,7,8-TCOF 1s said to be slightly more
soluble In water than 1s the corresponding dloxln, 2,3,7,8-TCDD (Greenlee
and Poland, 1978), which has a solubility of -0.2 vg/a at 25°C (Helling
et al., 1973).
2.3.4. Partition Coefficients. Sarna et al. (1984) and Burkhard and
Kuehl (1986) have documented the octanol/water partition coefficients for
some PCDFs (Table 2-4). The disagreement for OCDF arises because of uncer-
tainties 1n the K values of reference compounds of high K . The par-
ow r 3 ow K
tHIonlng of organic chemicals between Upld and water 1s an Important
determinant of the bloconcentratlon potential of a toxicant and has some-
times been effectively used as an Indicator of the preferred degradatlve in
vivo pathways.
2.3.5. Boiling Points and Vapor Pressures. No vapor pressures of any of
the PHDFs at 25°C have been measured directly.
The vapor pressures for a series of PCDFs have been estimated by a gas-
chromatographlc method (Firestone, 1977a; Berg et al., 1985a) (Table 2-5).
Unfortunately, different Investigators have obtained different pressures by
this method for the esters of 2,4-d1chlorophenoxyacet1c acid (Jensen and
Schall, 1966; Flint et al., 1968). The vapor pressures, however, of the
PCDFs will certainly not be lower than the values In Table 2-5, and a
decrease In vapor pressure with Increasing chlorlnatlon 1s expected.
2.3.6. Adsorption and Desorptlon. It would be expected that the PHDFs
would adsorb very strongly to organic matter and nonpolar nonorganlc
material such as charcoal, but not so strongly to polar Inorganic material
1926A 2-15 06/21/86
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TABLE 2-4
The Logarithm of the Octanol/Water Partition Coefficients (Kow)
of Some PHDFs Using HPLC Methods
PCDF
2,8-d1chloro-
2,3,7,8-tetrachloro-
octachloro-
log Kow
5.95
5.30°
5.82+0.02
13.37
8.78
Reference
Sarna et al., 1984a
Burkhard and Kuehl, 1986C
Burkhard and Kuehl, 1986C
Sarna et al., 1984a
Burkhard and Kuehl, 1986C
aQuadrat1c equation treatment: Blorad B1os1l (10 ym) data
bQuadrat1c equation treatment: Unspecified "mlcrobore" HPLC column
cSarna et al. (1984) data recalculated from experimental data
1926A 2-16 06/23/86
-------
TABLE 2-5
Vapor Densities and Pressures of PCDFs as Estimated from a
Gas Chromatographic Method*
PCDF Isomer
Estimated
Vapor Pressure
(mm Hg x 10~«
at 25°C)
Estimated Vapor
Density at 25°C
(yg/m3)
D1-
2,4-
3,7-
2,8-
Tr1-
2,4,6-
2,3,8-
Tetra-
1,4,6,8-
2,4,6,8-
2,3,4,8-
2,4,6,7-
1,2,7,8-
2,3,7,8-
2,3,6,7-
3,4,6,7-
Penta-
1,3,4,7,8-
1,2,4,7,8-
1,2,3,6,7-
2,3,4,7,8-
Hepta-
unspedfled
unspecified
unspecified
Octa-
7.3
7.0
6.8
4.0
3.7
2.5
2.5
2.2
2.1
2.0
2.0
1.9
1.8
1.3
1.3
1.1
1.1
0.44
0.36
0.30
0.19
82
79
77
52
48
38
38
33
32
30
30
29
27
22
22
18
18
9.0
7.4
6.2
4.3
*Source: Firestone, 1977a
1926A
2-17
06/21/86
-------
such as alumina. The predicted behavior 1s borne out by the fact that alum-
ina 1s used generally for the column chromatography of PCDDs and PCDFs
(O'Keefe, 1978a). Strong binding 1s expected to soils of high organic con-
tent as observed for 2,3,7,8-TCOD (Isensee, 1978). This has been observed
for 2,3,7,8-TCDD In South Vietnam and northwestern Florida soils (Westing,
1978). Although both 2,3,7,8-TCDD and -TCDF are comparatively Insoluble In
water, the 2,3,7,8-TCDF appears to be slightly more water soluble. There 1s
a possibility that the TCDF will be more readily leached from soils than
TCDD. Experiments should be performed to test this hypothesis. The rela-
tion, Equation 2-1, has been shown to hold by Karlckhoff et al. (1979) for
nonpolar compounds.
log Koc = log Kow - 0.21 (2-1)
where
Koc 1s a constant that 1s characteristic of the compound under con-
sideration and
Kow Is the octanol/water coefficient
If the K values of Burkhard and Kuehl (1986) (see Table 2-4) are
ow
assumed, the log K values for 2,8-DCDF, 2,3,7,8-TCDF, and OCDF are 5.09,
\)\f
5.61 and 8.57, respectively.
If the PHDFs are adsorbed 1n the same manner as dibenzothlophene by
soils (Hassett et al., 1980), the FreundUch adsorption Isotherm should
apply as follows:
Cs = KdCw 1/n (2-2)
where
Cs Is the amount sorbed 1n yg/g of soil
Cw 1s the equilibrium solution concentration (yg/s.)
n Is an Integer characteristic of the process and
K(j Is the FreundUch constant
1926A 2-18 06/21/86
-------
If n = 1, as for dlbenzothlophene (Hassett et al., 1980),
Cs = KdCw (2-3)
Since
Kd = 100KOC/%OC (2-4)
where XOC 1s the percentage of son organic content. For an organic content
of 1%, K = 100 K . Thus, log Kd for 2,8-DCBF, 2,3,7,8-TCDF, and
OCDF are then 7.09, 7.61 and 10.57, respectively.
2.3.7. Miscellaneous Properties. The molar refractions of the DCDFs,
TrCDFs, TCDFs, PeCDFs, HpCDFs and OCOF have been estimated to be 60.2, 65.0,
69.8, 74.6, 84.2 and 89.0, respectively (Firestone, 1977a).
2.3.8. Spectroscoplc Properties.
2.3.8.1. ULTRAVIOLET ABSORPTION SPECTRA — There are few studies of
the UV absorption spectra of PHDFs. The existing data for MBOFs are provid-
ed by Weast (1985) In Table 2-6 and for PCOFs by Firestone (1977a) and Gray
et al., (1976) 1n Table 2-7.
The UV spectra of some 2-MHDFs have been reported by Kurokl (1968). The
Hammett p value for the 2-substltuted derivatives was 1.44. Thus, there
was not much effect on \ of substitution at this position (Kurokl,
1968).
Chloro-substltutlon does not affect the x „ of the parent compound
max
very much for the 2-MCDF and 3-MCDF (Cern1an1 et al., 1954). The UV spec-
trum of 2,8-DCDF Is also similar to that of unsubstltuted OF with a \max
at 290 nm 1n methanol (Crosby et al., 1973). Firestone (1977a) reported the
UV absorptions of several PCDFs and found the red shift on Increasing chlor-
1nat1on was small as shown In Table 2-7.
2.3.8.2. FLUORESCENCE -- No fluorescence data for the PHDFs have yet
been published.
1926A 2-19 06/21/86
-------
TABLE 2-6
Ultraviolet Absorption Maxima and Molar AbsorptlvHIes for
Some Substituted PBDFs*
MBDF
Solvent
Absorption Peak In nm
[log(molar absorptivity)]
2-Bromo
3-Bromo
Acetic acid
Ethanol
222(4.58), 251(4.19), 288(4.25),
310(3.67)
219(4.55), 254(4.31), 290(4.31),
300(4.22)
*Source: Weast, 1985
1926A
2-20
06/21/86
-------
TABLE 2-7
Absorption Maxima of Some PCDFs In Chloroform3
UV Absorption
PCDF Maxima Wavelengths (nm)
2,3,8-TrCDF 256, 302, 313
2,3,6,8-TCDF 250, 260, 285, 297, 314b
2,3,7,8-TCDF 257, 294, 310, 323
2,4,6,8-TCDF 259, 309, 316
1,2,4,7,8-PeCDF 256, 266, 297
1,3,4,7,8-PeCDF 263, 272, 297, 320
aSource: Firestone, 1977a
bGray et al., 1976
1926A 2-21 06/21/86
-------
2.3.8.3. PHOSPHORESCENCE — Few studies have been reported on the
phosphorescence of the PHDFs. Teplyakov et al. (1970) noted an Increase In
quantum efficiency on substitution of dlbenzofuran with chlorine atoms. The
use of phosphorescence techniques to determine PCDFs has been suggested by
Brownrlgg et al. (1972). The difficulties of low-temperature technology and
the Interference by Impurities are substantial.
2.3.8.4. NMR AND EPR SPECTROSCOPY —The proton and "F spectra of
1,2,3,4-TFDF and 1,2,4-TrFDF Implied that there was a large para F-F coup-
ling (Brown and Mooney, 1967). The proton NMR data of some PCDFs are given
In Table 2-8.
The 13C-NMR chemical shifts and assignments of carbon-proton coupling
constants have been reported for the 2-MBDF and 2,8-DBDF. Long range sub-
stltuent effects were observed In unsymmetMcal compounds (Huckerby, 1979).
2.3.8.5. MASS SPECTROSCOPY -- The spectra for the 2,3,7,8-TCDF and
OCDF (Curley et al., 1974) attest to the great stability of the parent 1on
since the latter 1s the largest peak 1n the spectrum.
The mass spectra of the PHDFs will be discussed more thoroughly 1n the
section on GC/MS (Section 3.4.2.2.2). The major fragment Ions are M-C1,
M-C12, M-C1-CO and M-C12-CO.
2.4. CHEMICAL REACTIONS
2.4.1. General. PHDF reactions have been reviewed by Coffey (1973);
Gllman's group was largely responsible for much of the early chemistry
(GUman et al., 1934a,b, 1935a,b, 1939a,b,c,d, 1940a,b,c, 1944; GUman and
Young, 1934, 1935; GUman and Ess, 1939; GUman and Abbott, 1943; Oilman and
Thlrtle, 1944; Gllman and Swiss, 1944; Gllman and Avaklan, 1945a,b; Gllman
and Esmay, 1953, 1954).
1926A 2-22 06/21/86
-------
TABLE 2-8
lO
ro
Proton NMR Data for Some PCDFs
PCDF
1-
2-
3-
4-
2.3-d1-
2,6-d1-
2
2
i
B 3
1
1
2
2
2
2
1
,7-dl-
,8-d1-
,7-d1-
,3,6-tr1-
,3,8-tM-
,3.4-trl-
,3.8-trl-
,4.8-tr1-
,6,7-trl-
,2.3,4-tetra-
Chemlcal Shifts
(ppm downMeld from tetramethylsllane)
Coupling Constants (J)
(Hertz)
ra, 7.20-8.10
m. 7.20-8.10
m, 7.20-8.10
m, 7.20-8.10
m. 7.20-8.10
m. 7.20-8.20
in, 7.20-8
m, 7.20-8
m- H3.4,6
HT = 7.98
H2 = 7.55
H2 = 7.55
m. H6 to
HT = 8.27
HT = 8.07
HI = 7.77
H! = 8.08
H6 = 7.65
.20
.10
.7 = 1
; H2 =
; H4 =
; H4 =
Hg = 7
; H4 =
; "6.7
;H3 =
; H3 =
; H7 =
.45-7.47; H-\
7.38; H4 =
7.85; H7 =
7.79; H6 =
.25-8.25; H-\
7.82; H6 =
= 7.54 (m);
7.49; H6 =
7.46; H4 =
7.58; H8 =
,9 =
7.68
7.68;
7.76;
= 8.
7.56;
Hg .
7.56;
7.63;
7.44;
7.87
H8 = 7.50; Hg = 8.27
H7 = 7.64; Hg = 8.27
39
H7 = 7.45; Hg = 8.09
8.24
H7 = 7.48; Hg = 7.87
H8 = 7.48; Hg = 7.99
Hg = 8.34
Jl,9
J1.2
J2.4
J8.9
J2.4
J7.9
J6.7
J1.3
J7.9
Jl,3
J8.9
J7.8
= 1.1;
= 8.30
= 1.6;
= 8.1;
= 1.6;
= 2.2
= 8.6;
= 2.0;
= 1.8
- 2.4;
= 8.4
; 6.7;
1.8
; J2>4 = 1.40
J7.8 • 8.1;
J7.g = 1.2
J&.7 • 8.4;
J7.9 • 2.2
36.7 • 8.8;
H3.4 • 8-2;
7.8= 7.0;
Reference
Safe and Safe,
Safe and Safe,
Safe and Safe,
Safe and Safe,
Safe and Safe,
Safe and Safe.
Safe and Safe,
Safe and Safe,
Kurokl et al..
Nor strom et al
Safe and Safe,
Safe and Safe,
Safe and Safe,
Safe and Safe,
Norstrom et al
Kurokl et al.,
Safe and Safe,
Bell and Gara,
1984
1984
1984
1984
1984
1984
1984
1984
1984
. . 1979
1984
1984
1984
1984
., 1979
1984
1984
1985
1,2,3,7-tetra-
CO
m, 8.29-8.40 for Hg
ro. 7.33-7.76 for H&>7>8
H4 = 7.63; He = 7.57; H8 7.38; Hg = 8.23
H4 = 7.65; H6 = 7.60; H8 = 7.41; Hg = 8.25
J6.8/7.9 = 2.0
J8.9 • 8.4; H6(8
Kurokl et al., 1984
Kurokl et al., 1984
-------
V0
ro
3*
PCDF
1.2,3,8-tetra-
1.2.4.7-tetra-
1,2.4.8-tetra-
1,2,6,7-tetra-
1.2.7,8-tetra-
1,2.7.9-tetra-
1,3.4.7-tetra-
ro 1.3,6.7-tetra-
i
£ 1.3.6,8-tetra-
1.3,7.9-tetra-
1.4,6.7-tetra-
2.3.4.6-tetra-
2,3.4,7-tetra-
2.3,4,8-tetra-
2,3,6.7-tetra-
-5 2.3,6.8-tetra-
r
(ppm
H4 =
H3 *
H3 •
«3 •
H3 •
H3 =
H3 •
H3 •
H2 =
H2 -
H2 =
H2 =
H2.8
H2 =
Hl =
H] =
Hl =
Hi -
»1 -
H] -
Hi -
H] =
Hi -
Chemical Shifts
downfleld from tetramethylsllane)
7.64;
7.59;
7.60;
7.76;
7.50;
7.52;
7.43;
7.43;
7.45;
7.36;
7.39;
7.39;
= 7.41
7.30;
7.95;
8.39;
7.91;
7.88;
7.92;
7.87;
8.39;
7.99;
7.98;
8.44;
H6>7 * 7.50; H9
H& = 7.67;
H6.7 = ">•
H6 = 7.71;
H4 = 7.57;
H4 = 8.20;
H4 = 7.57;
H4 = 7.61;
H6 = 7.64;
H4 = 7.54;
H4 = 7.56;
H4 = 7.56;
; H4>6 - 7
H3 = 7.43;
H7 = 7.55;
H7 = 7.69;
H6 = 7.64;
H6 = 7.62;
H6 = 7.57;
H6 = 7.54;
H6 . 7.78;
H4 = 7.77;
H4 = 7.76;
H4 = 8.06;
HB •
7.45-7
H7 .
H8 =
H8 =
H6 •
H6 •
H8 .
H8 =
H7 =
H7 =
.48
H8 =
HB -
H8 -
H8 =
HB •
H7 .
H7 =
H7 =
H8 -
H7 =
H7 =
= 8.
7.43
.56;
7.58
7.50
7.62
7.71
7.49
7.40
7.46
7.52
7.52
7.51
7.33
7.51
7.39
7.36
7.50
7.46
7.64
7.47
7.51
7.70
29
; H9
H9
; H9
; H9
; H9
; H9
; H8
: H9
; H9
; H9
: H9
; H9
; H9
; H9
; H9
; H9
; H9
; H9
; H9
; H9
- 8.28
= 8.31
= 8.25
= 8.17
= 7.49
- 8.44
= 7.43
= 8.19
- 8.06
= 8.16
- 8.16
= 8.14
= 7.79
= 8.18
= 7.79
= 7.77
= 7.87
= 7.82
= 8.18
= 7.71
= 7.77
= 8.23
Coupling Constants (J)
(Hertz)
'7.9 =
J8,9 =
J7,9 =
J3,4 *
'3.4 •
J3.4 •
J8.9 =
J8.9 =
'2.4 •
J2,4 -
J2.3 =
J8.9 '
J7,9 =
J8.9 =
J7,9 =
J8,9 =
Jfl.9 =
J6.7 -
J&.7 -
•>8,9 =
J7,9 =
^7,9 =
1.3
8.3;
1.1
9.0;
8.8;
8.7;
8.8;
8.8;
8.9;
8.5;
1.5;
1.8;
2.0;
8.4;
7.4;
1.5
7.8;
1.2
8.3;
7.9;
8.8;
8.8;
8.8;
8.5
1.65
2.0
'6.8 •
J7.9 -
J8,9 -
J8,9 =
J6,9 =
J6.8 -
J6,8 =
'2.4 -
J7,9 =
J7,9 =
J&.8 *
J8,9 =
J7.8 -
J7.8 =
J6.8 =
'6.8 =
J7.9 =
J7,9 =
J7.9 -
1.8
2.3
8.5
8.7
0.4
1.9
2.0
1.65
1.5
2.0
2.0
8.5
7.6
7.8
1.6
2.0
1.8
2.0
2.2
Reference
Kurokl et al..
Bell and Gara,
Kurokl et al..
Safe and Safe.
Kurokl et al.,
Bell and Gara.
Kurokl et al..
Kurokl et al..
Bell and Gara.
Kurokl et al..
Kurokl et al..
Bell and Gara,
Kurokl et al..
Kurokl et al..
Kurokl et al..
Safe and Safe,
Kurokl et al..
Bell and Gara,
Kurokl et al..
Bell and Gara,
Safe and Safe,
Kurokl et al..
Kurokl et al..
Bell and Gara,
1984
1985
1984
1984
1984
1985
1984
1984
1985
1984
1984
1985
1984
1984
1984
1984
1984
1985
1984
1985
1984
1984
1984
1985
2,3.7.8-tetra-
CO
cr>
H! 9 = 7.90; H4 6 = 7.70
Hi'9 = 7.94; H4'fc = 7.67
Hi'9 = 7.89; H4'fc = ?
Kurokl et al.. 1984
Bell and Gara. 1985
Norstrom et al.. 1979
-------
INJ
TABLE 2-8 (cont.)
PCOF
2,4.6-7-tetra-
2.4,6.8-tetra-
3,4.6,7-tetra-
i
tn
o
CP
2
CO
1,2,3,4
1,2.3.4
1.2,3,6
1.2,3,6
1.2,3,7
1.2.3,8
1.2.4,6
1.2.4.6
1.2.4,6
1.2,4.7
1,2,4.7
1.2,4.8
1.2.6,7
1.3.4.6
1.3.4.6
1.3.4,6
,6-penta-
,8-penta-
,7-penta-
,9-penta-
,8-penta-
,9-penta-
,7-penta-
,8-penta-
,9-penta-
,8-penta-
,9-penta-
,9-penta-
,8-penta-
,7-penta-
,8-penta-
,9-penta-
(ppm
Hi -
Hi -
Hi. 9
Hi. 9
H7 .
H6.7
H4 =
H4 =
H4 .
H4 =
H4 =
H4 .
H4 =
H3 =
H3 =
H3 =
H3 =
H3 =
H3 =
H3 =
H3 =
H2 =
H2 =
H2 =
Chemical Shifts
downfleld from tetramethylsllane)
7.79; H3 = 7.51;
8.23; H3 = 7.73;
= 7.77; H3j * '
1 H8
L53
= 7.48; H9 = 7.72
= 7.70; H9 = 8.18
= 7.70; H2?8 = 7.46
7.58;
= m;
7.89;
7.72;
7.71;
7.71;
7.65;
8.06;
7.69;
7.65;
7.63;
7.60;
7.65;
7.69;
7.63;
7.68;
7.69;
7.52;
7.51;
7.48;
7.51;
H8 = 7.38;
7.55-7.56;
H7 = 7.74;
HB = 7.52;
H8 = 7.45;
H7 = 7.43;
H6 = 7.71;
H6 = 8.08;
H6 = 7.64;
H6 = 7.64;
H8 = 7.54;
HB = 7.51;
H7 = 7.59;
H7 = 7.46;
H6 = 7.79;
Hj = 7.58;
H6 = 7.67;
H4 = 7.61;
H8 = 7.53;
H7 = 7.54;
H7 = 7.44;
;«4
H9 =
; H9
; H9
; H9
; H8
; H9
; H9
; H9
! H7
; H9
; H9
; H9
; H8
; H9
; H8
I **~J
| no
; H9
; H9
;H8
* 8.24
8.27
= 8.37
= 8.15
= 8.13
= 7.34
= 8.40
= 8.52
= 8.39
= 7.43
= 8.19
= 8.15
= 8.25
= 7.37
= 8.45
= 7.47
- 7.52
= 8.38
= 8.11
- 8.14
= 7.34
Coupling Constants (J)
(Hertz)
J8,9 °
J8,9 =
Jl.3; ;
J8.9; 1
^7.8 =
J7.9 =
J7.9 -
J6.7 •
J8,9 =
J8,9 =
J7.8 •
J&.7 •
Ja!9 =
J7,9 =
-"7.8 •
J6.8 -
J&.7 =
J3.4 •
J8,9 «
J7.9 =
J7.8 -
8.45; J] 3 = 2.1
8.3; Ji>3 = 1.9
f,9 = 1-8
1.2 = 8.7
7.75; J8 9 = 7.55;
1.4
1.3
8.6; J7>9 = 2.2
8.6
8.5
8.5
8.6
8.7
8.6
2.1
8.5
2.0
8.B
8.85
8.4
2.0
8.9
Reference
Kurokl et al..
Bell and Gara,
Kurokl et al..
Bell and Gara,
Kurokl et al..
Kurokl et al..
Safe and Safe,
Kurokl et al..
Bell and Gara,
Bell and Gara,
Kurokl et al..
Safe and Safe,
Bell and Gara.
Bell and Gara,
Kurokl et al..
Bell and Gara,
Kurokl et al..
Bell and Gara,
Kurokl et al..
Kurokl et al..
Bell and Gara,
Kurokl et al.,
Kurokl et al..
Bell and Gara,
Bell and Gara,
1984
1985
1984
1985
1984
1984
1984
1984
1985
1985
1984
1984
1985
1985
1984
1985
1984
1985
1984
1984
1985
1984
1984
1985
1985
-------
vO
TABLE 2-8 (cont.)
«i^
rvj
I
O>
O
\
—1
^
1.3.4,7
1.3.4.7
2,3.4,6
2,3.4.6
2.3,4,6
2.3,4,7
1.2.3.4
1,2.3,4
1,2,3,4
1.2,3,4
1.2.3,4
1.2,3,4
1,2,3.6
1,2.3,6
1,2,3,6
1.2.3.7
1,2.4,6
PCDF
,8-penta-
,9-penta-
,7-penta-
,8-penta-
,9-penta-
,8-penta-
,6.7-hexa-
6 8-hexa-
6 9-hexa-
7 8-hexa-
,7,9-hexa-
,8,9-hexa-
7 8-hexa-
7 9-hexa-
R 9-hexa-
,8,9-hexa-
,7,8-hexa-
Chemlcal Shifts
(ppm downfleld from tetramethylsllane)
H2 = 7.46;
H2 = 7.49;
H2 = 7.51;
H! = 7.89;
H! = 7.93;
H! = 7.92;
H! = 8.32;
H! = 7.91;
HT = 8.43;
HT = 7.93;
H8 = 7.55;
H8 = 7.53;
H7 = 7.59;
1 '
H7 = 7.59;
H7 = 7.50;
H7 = 7.48;
H6 = 7.79;
H6 * 7.77;
H6 = 7.59;
H6 = 7.58;
H6 = 7.68;
H4 = 7.74;
H4 = 7.72;
H4 = 8.16;
Hi = 7.73;
^ •
Hi 7 = 7.71
^ ( 1
H4.6 = 7-65
H3 = 7.65;
H6 =
H& •
H6 •
H8 =
H8 -
H7 .
H7 =
H& •
H& •
H& •
H9 .
H9 .
H9 =
H9 =
HB -
o
HB -
H9 =
H9 .
HB -
HB -
H7 =
H9 =
H9 =
H9 .
HR =
O
or
H9 =
7
7
7
7
7
7
7
7
8
7
8
8
8
8
7
7
8
8
7
7
7
8
8
8
7
7.
8
.73; H9 = 8.31
.77; H9 = 8.36
.56; H8 = 7.44
.47; H9 = 7.68
.50; H9 = 7.71
.54; H9 = 7.77
.45; H8 = 7.30
.77; H9 = 7.97
.09; H9 = 8.46
.73; H9 = 7.87
.16
.14
.22
.21
.40
.38
.41
.40
.48
.47
.52
.35
.34
.51
.55
74
.36
Coupling Constants
(Hertz)
36.8 =
Ja.9 -
Ja.9 =
J7.9 =
J7.8 -
J8.9 =
J8.9 =
J7 9 =
J7|9 •
J7,8 =
J6.8 •
J&.8 =
J6,7 =
1
8
8
2
8
8
8
1
1
8
8
1
1
8
.9
.4
.2
.0
.4
.25
.5
.8
.9
.7
.5
.8
.9
.8
(J) Reference
Bell and Gara,
Kurokl et al.,
Bell and Gara,
Bell and Gara,
Kurokl et al..
Kurokl et al..
Bell and Gara,
Kurokl et al.,
Safe and Safe,
Bell and Gara,
Kurokl et al..
Bell and Gara,
Kurokl et al..
Bell and Gara,
Kurokl et al..
Bell and Gara.
Kurokl et al..
Bell and Gara,
Kurokl et al.,
Bell and Gara,
Bell and Gara,
Kurokl et al..
Bell and Gara,
Safe and Safe,
Bell and Gara,
Kurokl et al..
Bell and Gara,
Kurokl et al..
1985
1984
1985
1985
1984
1984
1985
1984
1984
1985
1984
1985
1984
1985
1984
1985
1984
1985
1984
1985
1985
1984
1985
1984
1985
1984
1985
1984
ca
7.99; H9 = 8.54
Safe and Safe, 1984
-------
TABLE 2-8 (cont.)
I
ro
PCDF
Chemical Shifts
(ppm downfleld from tetramethylsllane)
Coupling Constants (3)
(Hertz)
Reference
1.2
1.2
1 3
2 3
1.2
1 ?
1.2
.4
.4
4
4
.3
3
.3
.6
.6
f>
f,
.4
4
.4
.7
.8
7
7
.6
f.
.7
,9-hexa-
,9-hexa-
8-hexa-
8-hexa-
,7,8-hepta-
8 9-hepta-
,8,9-hepta-
H3
H3
H3
H3
H2
H2
Hi
1
Hi
Hi
Hi
H9
H7
/
H&
_
=
,7
_
=
.
,9
.9
.9
=
_
=
7
7
-
8
7
7
=
=
=
=
8
7
7
.71
.69
7.
.02
.52
.51
7.
7.
8.
8.
.35
.75
.74
; H8 = 7.57
; H8 = 7.54
73
; Hg = 8.30
; H9 = 7.51
92
91
24
32
Kurokl et al..
Bell and Gara.
Kurokl et al..
Nor strom et al
Kurokl et al..
Bell and Gara.
Kurokl et al..
Be 1-1 and Gara,
Nor strom et al
Safe and Safe,
Kurokl et al.,
Kurokl et al..
Kurokl et al..
1984
1985
1984
., 1979
1984
1985
1984
1985
.. 1979
1984
1984
1984
1984
o
\
ro
\
o>
-------
The 2-carboxyl1c add of DBF has been made by the Grlgnard reaction from
2-MBDF. Since BDBFs yield the llthlo compounds by Interchange, all four
Hthlo-DBFs are accessible and the lithium atoms can be replaced by car-
boxyl, methyl (using dimethyl sulfate), halogen or amlno groups (using
0-methylhydroxylam1ne).
OFDF reacts with sodium methoxlde 1n methanol to yield the 3-mono- and
3,7-dlether. Similarly, 1,2,3,4-TFDF undergoes nucleophlUc substitution
reactions with sodium hydrogen sulflde, sodium methlolate and lithium tetra-
hydrldoalumlnate, In which the 3-fluorlne atom 1s replaced by SH, SMe and H,
respectively.
The directing effects of functional groups at the 2-pos1t1on appear to
be mainly toward the 8-pos1t1on regardless of whether the group Is elec-
tron-attracting or electron-withdrawing (Keuml et al., 1972a). This 1s not
so for the 3-pos1t1on since 3-halo- groups give substantial amounts of the
7-subst1tuted compounds (Table 2-9).
2-MIDF reacts with ammonia over cuprous bromide at 200-210°C to yield
95% 2-am1no-DF (Brown and Coleman, 1973a). 3-MBDF directs the nltronlum Ion
to the l-pos1t1on; 1-MBDF 1s nitrated at the 3-pos1t1on (Grlnev et al.,
1973); the TrCDFs are monochlorlnated by SbCl5/CCl4 reagent to produce
TCOFs (Table 2-10).
2.4.2. Photodecomposltlon. Irradiation of various lower chlorinated PCBs
In aqueous suspensions of 1 and 10 mg/ft. at 254 nm (mercury arc) caused no
appearance of PCDFs. Irradiation at 310 nm resulted 1n PCDFs from 2,5-d1-
chloro- and 2,2',5,5'-tetrachloro-PCB to the extent of -0.2% after Irradia-
tion times of up to 200 hours. This suggests that PCDFs may be destroyed at
254 nm but only very slowly at 310 nm (Hutzlnger et al., 1973). The pres-
1926A 2-28 06/21/86
-------
TABLE 2-9
Friedel-Crafts Acetylatlon of Substituted PHOFs*
PHDF Conditions Acetylatlng Agent Derivative Percentage
Yield
-(2-Br)
-(2-C1)
-(3-Br)
-(3-C1)
8-
8-
N1trobenzene/AlCl3 Benzoyl chloride 8-
(50-60°C/1 hr) 7-
8-
7-
84
84
56
31
60
3
*Source: Keuml et al., 1972a
1926A 2-29 06/23/86
-------
TABLE 2-10
Chlorlnatlon of TrCDF with Antimony Pentachlor1de/CCl4 Reagent to Produce TCDFs*
Produced Original TrCDF
TCDF
123 124 126 127 128 129 134 136 137 138 139 146 147 148 149 234 236 237 238 239 246 247 248 249 346 347 348 349
1234 XXX X
1236 X X X X
1237 XXX X
1238 X X X X
1239 XXX X
1246 XX X X
1247 XX X X
1248 XX X X
1249 XX X X
2346 XX X X
2347 XX X X
2348 XX X X
2349 X X X X
1346 XXX X
7» 1347 XXX X
La "48 XXX X
<=> 1349 XXX X
1267 XX XX
1268 XX XX
1269 XX XX
1278 XX XX
1279 XX XX
1289 XX
1367 XX XX
1368 XX XX
1369 X XXX
1378 XX XX
1379 X X
1467 XX XX
1468 XX XX
1469 X X
1478 XX X X
2367 XX XX
SB ' , !
O 2467 xx XX
a* 2468 X x
£ 3467 XX
CO
:: Adapted from Hazer et al., 1983a,b
-------
ence of the triplet sensltlzer, 4,4'-d1chlorod1benzophenone, Increased
photodecomposltlon of 2,8-DCDF (Crosby and Mollanen, 1973).
Choudhry and Hutzlnger (1982a,b) reviewed the photochemical formation
and degradation of PCDFs. PCDFs could be produced 1n yields up to 10% from
polychlorlnated dlphenyl ethers (0.8 mM 1n methanol) bearing 2- or 2'-chloro
substHuents after 88 hours Irradiation at 310 nm {Choudhry et a!.,
1977a,b). The addition of the triplet sensltlzer acetone at 0.45 M markedly
Increased the PCDF yield while suppressing reductive dechlorlnatlon.
Greater than 50% yields of 2-MCDF, 2,3-DCDF and 2,3,8-TrCDF were obtained 1n
the presence of acetone at 310 nm after 10-15 hours Irradiation of 2,4'-d1-
chloro-, 2,4,5-trlchloro- and 2,4,4',5-tetrachlorod1phenyl ether, respec-
tively. The PCDFs can also be produced In solvents such as n-hexane and
ethanol (Choudhry and Hutzlnger, 1982a,b).
The photodegradatlon of OCDF Irradiated at 310 nm In n-hexane 1s more
rapid than that of 2,8-DCDF similarly Irradiated (Hutzlnger et al., 1972a,b,
1973). The half-lives were -1.5 and 2.5 hours, respectively. There was
also a pronounced solvent effect. The half-life for OCDF In methanol was
-15 minutes. This 1s the reverse of results with OCDD (Crosby et al.,
1971). Irradiation of both PCDFs In hexane for 20 hours caused formation of
a "yellow gum," probably a polymer, which showed no recognizable chlorine
1sotop1c patterns 1n the mass spectra. At shorter Irradiation times, reduc-
tive dechlorlnatlon was postulated to be the major photoprocess although no
photoproducts were Identified unambiguously. When water was used as a sol-
vent and thin films of the two PCDFs were Irradiated at 310 nm, the same
products were formed although much more slowly. The 2,8-DCDF also produced
a TrCDF, probably because the viscous film allowed free radical "cages" to
exist for comparatively long times. This proves that -Cl 1s produced
1926A 2-31 06/21/86
-------
during the Irradiations. The OCDF 1n a thin film or 1n solution appeared to
produce PCDFs to HpCDFs.
Crosby and Mollanen (1973) confirmed the previously stated results for
2,8-DCDF (10-100 mg/a) 1n methanol after Irradiation 1n a solar simulator
and 1n addition, unambiguously Identified 2-MCDF as the first reductive
dechlorlnatlon product. As expected, the half-life was longer than reported
for OCDF above (-75 hours). The 2-MCDF was almost unaffected by further
Irradiation (the half-life was -1 day), whereas 95% of the 2,8-DCDF was
degraded within 48 hours. The photodecomposHlon rate depended also on the
purity of the methanol, technical grade methanol containing acetone (a trip-
let sensltlzer) decreasing substrate half-life to near 25 hours. The
absence of lower chlorinated blphenyls 1n most environmental samples was
thought to Indicate a potential for the occurrence of the hydroxylatlon and
PCDF photochemical routes 1n the environment (Crosby et al., 1973).
Buser (1976b) reported that 4 hours of UV Irradiation (mercury arc) of
25 yg/ml OCDF In 95% hexane/benzene caused extensive dechlorlnatlon,
-90% of the compound being degraded. The major products were HxCDFs and
HpCDFs with trace amounts of PeCDFs. After 24 hours of Irradiation, HxCDFs
predominated (40%) along with PeCDFs and HpCDFs, and also trace amounts of
TCDFs. Only 2% of OCDF remained. At least 13 HxCDFs were formed from the
prolonged Irradiation. The dechlorlnatlon was not as selective as for
OCDD. These events were also confirmed by Firestone (1977b). Preferential
reductive dechlorlnatlon (mostly to HpCDFs) was Induced by 4 hours of
Y-1rrad1at1on (60Co; 1.4 Mrad/hr). Unfortunately, specific Isomers were
not Identified 1n any of these experiments.
Mazer and Hlleman (1982) and Mazer et al. (1983a,b) Irradiated eight
TCDFs (2,3,6,8-, 2,4,6,7, 1,2,7,8-, 1,4,6,8-, 1,2,4,8-, 2,4,6,8-, 2,3,7,8-
1926A 2-32 06/21/86
-------
and 1,2,3.4-) at 1 yg/mj. In n-tetradecane or n-hexane with 254 nm light
for 4 hours. GS/MS after HPLC showed the TrCDFs were mostly formed (Table
2-11). DCDFs were also produced towards the end of the photolysis experi-
ments. The order of photodegradatlon for chlorines 1n 1,2,3,4-TCDF was 3- >
2- > 1- = 4-. For 1,2,4,8-TCOF, the 1- and 2-pos1t1ons were most reactive.
Thus, 1t was found that chlorines on the same aromatic ring stabilize the
loss of a chlorine from that ring, that adjacent chlorines around a given
chlorine will facilitate preferential cleavage of that chlorine, and that 1f
there are an equal number of vicinal chlorines, the 3-chlorlne will be lost
before the 2-chlor1ne.
Mazer and Hlleman (1982) and Hlleman et al. (1985) also reported on the
photodegradatlon of PeCDFs (1,2,3,6,7-, 1,2,3,4,8-, 2,3,4,6,9-, 1,3,4,6,7-,
2,3,4,6,8- and 2,3,4,6,7-) at 254 nm 1n hexane (Table 2-12). The same gen-
eral principles appear to hold as for the TCDFs, that 1s, the most substi-
tuted ring tends to lose chlorine, especially 1f adjacent chlorines are pre-
sent. A quantitative analysis of the data 1s not yet available.
Ballschmlter et al. (1985) showed that the dechlorlnatlon of OCDF by UV
light favored expulsion of the chlorine In the 9-pos1t1on and to a lesser
extent the 8-pos1t1on. The 1,8-congeners dominated among the HpCDF and
HxCDFs formed.
2.4.3. Pyrolysls. Very few studies of pyrolysls of pure PHDFs have been
reported. Thermal degradation of OCDF at 400°C for 15 hours In a sealed
silica tube resulted 1n expulsion of chlorine 1n the 9-pos1t1on (Ball-
schmlter et al., 1985), and OCDF was thermally much less stable than OCDD.
HpCDFs mainly resulted with the 1,2,3,4,6,7,8-HpCDF dominating.
1926A 2-33 06/21/86
-------
TABLE 2-11
Photodegradatlon of TCDFs (1 pg/mft) at 254 nm After
4 Hours of Irradiation In Hexane or Tetradecane3
TCDF
4:0C
1,2,3,4-
3:1
1,2,4,8-
2:2
2,3,6,8-
1,4,6,8-
2,4,6,8-
2,4,6,7-
2,3,7,8-
1,2,7,8-
Major
1,2,4-
1.0
2,4,8-
1.0
2,4,8-
1.0
2,4,6-
1.0
2,4,8-
1.0
2,4,7-
1.0
2,3,8-
1.0
1,2,8-
1.0
TrCDFs and Their
Formed bv
Second
1,3,4-
0.38
1,4,8-
0.4
2,4,7-
0.59
1,4,6-
0.59
2,4,6-
0.40
2.4,6-
0.45
2,3,7-
0.52
2,3,8-
0.94
Relative Rat1osb
Photolysis
Third
2,3,4-
0.13
1,2,8-
0.1
2,3,8-
0.09
2,4,9-
0.60
-
3,4,8-
0.1
-
1,2,7-
0.75
Fourth
1,2,3-
0.12
1,2,4-
ND
2,3,6-
0.03
1,4,8-
0.21
-
3,4,6-
ND
-
2,3,9-
0.37
Source: Mazer and Hlleman, 1982
Assuming equivalent mass spectrometer responses.
C4:0 Substitution Indicates four chlorines on one aromatic ring and no
chlorines on the other aromatic ring.
ND = None detected at the point at which DCDFs began to form.
1926A
2-34
06/21/86
-------
TABLE 2-12
|5 PhotodecomposIt Ion of PeCDFs In Hexane at 254 nm*
rsj
CO
in
o
<3>
ro
_j
-v
00
Produced TCDF
1234
1236
1237
1238
1239
1246
1247
1248
1249
2346
2347
2348
2349
1346
1347
1348
1349
1267
1268
1269
1278
1279
1289
Original PeCDF
*^«o«.,-....o,;SSS|5|ESgS8S!5S8!o8S{5S!5
X X X X
X XXX
X XXX
X XXX
XX XXX
X XXX
X X XX
XXX X
XXX X
X XXX
X XXX
X XXX
X XXX
X XXX
X XXX
X XXX
X XXX
XX XX
XXX X
XXX X
XXX X
X XX X
X X
-------
VO
ro
TABLE 2-12 (cont.)
Original PeCDF
Produced TCDF
oo
1367
1368
1369
1378
1379
1467
1468
1469
1478
2367
2368
2378
2467
2468
3467
fXlCUCVJCsJCXlCNjCNJCNJCVICXlCXlCNJCNJeVlC^CVlCOC^COCOCOOCOCOCOCOCOCXl
X XX X
XX X X
XX XX
XX X X
X X
X XXX
X XXX
X X
XX XX
X XXX
XX XX
X X
X XXX
X X
X X
*Source: Adapted from Mazer et al., 1983a,b
o
c»
-V
ro
CO
CK
-------
2.5. SYNTHESIS
Synthesis of all 135 Isomers 1n large quantities 1s still an Important
research priority.
2.5.1. General Methods. Coffey (1973) has evlewed the older general
synthetic methods leading to the production of PCDFs.
PHDFs are produced by the following methods:
a. Eliminating water or hydrogen hallde from substituted 2,2'-d1-
hydroxy- or 2-halo-2'-hydroxyblphenyls, respectively:
Zn Clj
48% HBr/CH, COOH
OH OH
(Rij»90p«l»n end GiiMpmi. 1942)
Futad KOH
Cl OH
(Schimmelschmidt, 1950)
b.
Both types of reactions occur more easily under alkaline condi-
tions when halogen groups are present ortho- or para- to the
hydroxy and halogen groups to be eliminated (Alphen, 1932; Case
and Schock, 1943):
Decomposing blsdlazonlum salts of substituted 2,2'-d1am1nob1-
phenyls, although phenazones are frequent contaminants (Coffey,
1973):
O
H20
D1azot1zat1on of substituted o-am1nob1phenyl ethers (HcComble
et al.. 1931):
1926A
2-37
where Ph = CeHs-
06/21/86
-------
d. PHDFs have been prepared 1n poor yield by the pyrolysls of substi-
tuted dlphenyl ethers (Choudhry and Hutzlnger, 1982a,b) and chloro-
phenols (Bell, 1936). This route may be of environmental signifi-
cance since Incineration Is often used to dispose of phenol-Impreg-
nated woods and papers (Chapter 4). The reaction of phenol and HC1
at 550°C for 5 minutes for an Initial HC1 concentration >1 mM caused
the production of trace amounts of PCDFs and PCDOs (Eklund et al.,
1986).
2.5.2. Methods for PHDFs. Gllman et al. (1934a) reacted dlbenzofuran and
chlorine in carbon tetrachlorlde to yield 30% 2,8-DCDF. The 2-MCDF was pro-
duced In a similar fashion In ethanol as solvent at 60°C. The 2-MCDF,
3-MCDF-, 2,8-DCDF and 3,8-DCDF were synthesized by Shlbata et al. (1952) and
procedures were reviewed by Olta et al. (1955). The 1,3-cycloadd1t1on of
3,4,5,6-tetrachlorobenzene-2-d1azo-l-ox1de In chlorobenzene at 130°C yields
1,2,3,4-TCDF (Hulsgen et al., 1964).
The presence of higher PCDFs as reaction by-products was noted by Clark
(1940) after the chlorlnatlon of dlbenzofuran, and also by Cox et al. (1965)
after vacuum pyrolysls of various dlbenzofuran-based polymers and copoly-
mers. Cox et al. (1965) observed that the chlorinated polymers were more
stable than the bromlnated ones except for the tetra-bromlnated ones. R1ed
and Eng (1969) found that o-qu1none dlazldes added benzene with nitrogen
elimination to yield the 1,2,3,4-TCDF, 1,2,3-TrCDF and 1,3-DCDF. OCDF was
produced by pyrolysls of pentachlorophenol (Sanderman et al., 1958).
Further Interest In the synthesis of PCDFs had to await more Interest In
their toxlcologlcal properties and those of commercial PCBs.
Specific PCDFs have been synthesized by direct chlorlnatlon of unsubstl-
tuted dlbenzofuran (Gray et al., 1976), photolysis of chlorinated dlphenyl
1926A 2-38 06/21/86
-------
ethers (Choudhry et al., 1977a,b; Norstrom et al.. 1976a, 1977; Llndahl et
al., 1980), by Ullman reaction (MorHa et al., 1977b), photodechlorlnatlon
of OCDF (Buser, 1976b) and by thermal oxldatlve cycllzatlon of specific PCB
Isomers (MorHa et al., 1978; Buser et al., 1978a,d; Buser and Rappe, 1979),
or chlorobenzenes (Buser, 1979). The mechanisms Involved In the thermochem-
Ical generation of PCDFs have been reviewed by Choudhry and Hutzlnger
(1982b). Nearly all of these methods result 1n complex mixtures which
require extensive chromatography and purification before pure compounds can
be obtained, In general, with final yields <10%.
A synthetic route with environmental Implications 1s the thermal produc-
tion of PCDFs from PCBs. Buser et al. (1978a,d) and Buser and Rappe (1979)
showed that when specific PCBs were pyrolyzed 1n quartz ampules between 500
and 700°C, PCOF yields 1n the 1-10% range could be obtained, though they
were accompanied by many other products, Including chlorinated benzenes,
naphthalenes and hydroxy PCBs. Buser et al. (1978a) described the products
of pyrolysls at 600°C, Identified by GC/MS. Yields were as high as a few
percent In favorable cases. The decompositions are summarized 1n Table
2-13. There appear to be four major paths for production of PCDFs from
PCBs: loss of two ortho chlorines, loss of ortho hydrogen as well as chlor-
ine, loss of an ortho hydrogen as well as chlorine but Involving a shift of
chlorine from the 2- to the 3-pos1t1on and loss of two ortho hydrogens.
These paths are summarized In Figure 2-1 for 2,2',4,4',5,5'-hexachlorob1-
phenyl. Such paths are environmentally Important 1n the origin of toxic
PCDFs and PBDFs from various PCB and PBB formulations, respectively. Mazer
and HUeman (1982), Mazer et al. (1983a,b) and Hlleman et al. (1985) have
utilized the technique to produce 110 pure TCDFs to act as chromatographlc
standards after HPLC separation and confirmation by GC/MS and Kovats Indices.
1926A 2-39 06/21/86
-------
TABLE 2-13
Formation of PCDFs from the Pyrolysls of Specific PCBsa
PCB Pyrolyzed
PCDFs Formed
Tetra:
2,3,4,5-
2,3,5,6-
2,6,2',6'-
Penta:
2,3,4,5,6-
2,4,5,2',5'-
2,4,5,3',4'-
2,4,6,2',4'-
2,3,6.2'.5'-
Hexa:
2,4,5,2'.4',5'-
2,4,6,2',4',6'-
2,4,5,2',4',6'-
2,3,4,2',3',4'-
2,3,5,2',3',5'-
2,3,6,2',3',6'-
3,4,5,3',4',5'-
2,3,4-tr1-b; 1.2,3.4-tetra-
l,2,4-tr1-
l,9-d1; 1.4,9-trl-
l,2,3,4-tetra-b; small amounts of 1,2,4-tM-;
2,3,4-trl-;
2,3,8-tr1-b; small amounts of 2,3,6,8-tetra-
plus three other tetra; 1,3,4,6,9-penta-;
2,3,7,8-tetra-b; 2,3,6,7-tetra-;
1,3,4,7,8-penta;
l,3,7-tr1-b; 1,3,6.7-tetra-; 1,3,7,9-tetra-;
1,3,4,7-tetra-;
1,4,8-trl-; 1.2,8-trl-; 1,4,6,8-tetra-;
1,2,6,8-tetra-; 1,2,4,8-tetra-b;
l,2,6,9-tetra-;l,4,6,9-tetra-;
2,3,7,8-tetra-b; 2,3,4,7,8-penta-;
1,3,4,7,8-penta
1,3,7,9-tetra; 1,3,4,7,9-penta-;
l,3,7,8-tetra-b; 1,3,4,7,8-penta-;
1,3,4,7,9-penta
3,4,6,7-tetra-b; 1,2,3,6,7-penta-:
2,4,6,8-tetra-; 1,2,4,6,8-penta-;
l,2,4,8-tetra-b; 1,2,8,9-tetra-b;
l,2,4,6,9-penta(?); 1,2,4,8,9-penta-;
2,3,4,6,7,8-hexa-; small penta-
1926A
2-40
06/21/86
-------
TABLE 2-13 (cont.)
PCB Pyrolyzed PCDFs Formed
Hepta:
2,3,4,5,2',3',4'- -2,3,4,6,7-penta-; 1,2,3,4,6,7-hexa-b;
1,2,3,6,7,8-hexa-; 1,2,3,4,7,8,9-hepta-;
1,2,3,4,6,7,8-hepta-; 1,2,3,4,6,7,9-hepta-;
2,3,4,5,2',4',5'- 2,3,4,7,8-penta-b; 1,2,3,4,7,8-hexa-;
l,3,4,6,7,8-hexa-b; 2,3,4,6,7,8-hexa-;
1,2,3,4,6,7,9-hepta-;
Octa:
2,3,4,5,2',3',4',5'- 2,3,4,6,7,8-hexa-b; 1,2,3,4,6,7,8-hepta-b
aSource: Buser and Rappe, 1979
bMost abundant PCDF
1926A 2-41 06/21/86
-------
AT
loss of two
Ortho Cl
loss of ortho H
and Cl with
Cl shift
loss of ortho
H and Cl
loss of two
ortho H
2,3.7.8-tetra-CDBF
Cl Cl
Cl O
2.3,4,7,8-penta-CDBF
1.3.4,6,7,9-hexa-CDBF
FIGURE 2-1
Possible Reaction Schemes to Produce PCDFs from the Pyrolysls of
2,2', 4,4', 5,5'-Hexachlorob1phenyl
Source: Adapted from Buser and Rappe, 1979
1926A
2-42
06/21/86
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The formation of PCDFs by pyrolysls of commercial chlorobenzenes 1s
similarly Important (Buser, 1979). PCDDs, chlorinated naphthalenes, chlor-
inated styrenes and chlorophenols are also produced. Samples that were
pyrolyzed at 620°C 1n quartz ampules 1n the presence of air contained the
PCDF levels shown 1n Table 2-14. The PCDF content was much higher than the
PCDD content In all cases, especially for the trlchlorobenzenes for which
the ratio of PCDF to PCDD content was >40. The amounts of the toxic PCDF
Isomers (2,3,7,8-TCDF-; 1,2,3,7,8-PeCDF-; 2,3,4,7,8-PeCDF-) were small.
Llndahl et al. (1980) reported that pyrolysls of 2,2',4,4',5,5'-hexa-
chlorodlphenyl ether In quartz ampules at 500, 600 and 700°C for 1 minute
yielded 0.1, 1.3 and 0.1%, respectively, of PCDFs. The products were
1,2,4,6,8,9-HxCDF, 1,2,4,7,8-PeCDF and some PCDDs. More PCDFs than PCDDs
were formed when chlorodlphenyl ethers were pyrolyzed under these condi-
tions. Between 3.5 and 4.5% PCDFs were produced when the 3,5,4'-tr1-,
2,4,5,4'-tetra-, 2,4,6,3',5'-penta- and 2,3,4,5,6,2' ,3',4'-octa-chloro-d1-
phenyl ethers (CDPEs) were pyrolyzed at 600°C compared with up to 0.6% of
PCDDs. PCDF production was not so favored from the hexa-chlorodlphenyl
ethers (2,3,4,2',3',4'-, 2,3,4,2',3',5'-, 2,3,4,2',4',5'- and
2,4,5,2',4',5'-), PCDF and PCDD contents both ranging from 0.8-1.3%. The
same mechanisms depicted In Figure 2-1 for production of PCDFs from PCBs
also prevail from chlorinated dlphenyl ethers except that loss of 2-ortho-H,
ortho-H and -Cl are favored. Loss of two ortho-H appears to be favored for
the tr1- and tetra-CDPEs, but loss of ortho-H and -Cl Is predominant for
higher CDPEs. The octa-CDPE also yielded HxCDFs that can be formed when two
chlorines are lost by rearrangement, one of which was the 1,2,3,4,6,7-HxCDF
formed by loss of two ortho-Cl. Buser (1986) found that pyrolysls of three
polybromlnated dlphenyl ethers used as flame retardants resulted In yields
1926A 2-43 06/23/86
-------
1C
no
a*
TABLE 2-14
Formation of PCDFs from Pyrolysls of Chlorobenzenes*
Total Amount Percentage of PCDFs Formed Relative to Chlorobenzene
Chlorobenzene
Compound
Tri-
ll, 2, 3-; 1,2,4-; 1,3,5-)
Tetra-
Penta-
Mlxture
(tri-; tetra-; penta-)
Pyrolyzed (yg)
(ratio of Isomers) Tetra-
200 0.20
(1:1:1)
200 <10~4
200 <10~4
500 0.040
(1:1:1)
Penta-
0.55
2.5xlO~3
<2.5xlO"3
0.12
Hexa-
0.28
0.080
<2.5xlO~3
0.22
Hepta-
0.025
0.23
<2.5xlO~a
0.12
Octa-
<2.5xl03
0.10
<1.5xlO"3
1.20xlO~4
*Source: Recalculated from Buser, 1979
00
-------
of PBDFs and PBDDs of up to 10% at 510-630°C 1n quartz ampules. The mechan-
ism of formation appeared to be the same as that for the chlorinated deriva-
tives. The 2,3,7,8-TBDF was not a major product from pyrolysls of penta-,
octa- or decabromodlphenyl ethers.
Perchlorlnatlon has also been Investigated as a synthetic route to the
PCDFs (Williams and Blanchfleld, 1972; Hucklns et al., 1974; Crist and Hose-
man, 1977; Ballschmlter et al., 1985). The perchloMnatlon of 2-hydroxyb1-
phenyl with thlonyl chloride/antimony pentachlorlde/lodlne produced 8% of
pure OCDF (Gara et al., 1980). No lower PCDFs were formed from the reac-
tion. Perchlorlnatlon with BMC Reagent (SO Cl /S Cl /A1C1 Is
C C C C.
-------
10
ro
TABLE 2-15
Synthetic Methods for PCDFs Using Chlorinated Dlphenyl Ethers (CDPEs) as Substrates
i
£
o
NS
CO
a*
Starting COPE PCDF
3',4,4'-Tr1- 2,3,8-TM-
3.3'.4.4'-Tetra- 2,3.7.8-Tetra-
3,3'-D1- 3.7-D1-
2.2',4,4'.5,5'-Hexa- 1 ,2,4,6,8,9-Hexa-
2.2'.3,3',4,41-Hexa- 2,3.4,6.7.8-Hexa-
2.2'.4,4'-Tetra- 2,4,6.8-Tetra-
2,2'.4.4l.5-Penta- 1, 2.4,6, 8-Penta-
2,31,4.4',5-Penta- 1 ,2.4.6, 8-Penta-
2,2',3,4,4'-Penta- 2,3.4,6.8-Penta-
2,2',3,4,4',5-Hexa- 1,2,4,6.7,8-Hexa-
2.2'.3.3',4,4'.5-Hepta- 1 ,2.3,4.6.7,8-Hepta-
aM1th column chromatography
bWHh preparative TLC
Reaction Time
(hours)
0.50
1.0
0.66
14
3.5
2.0
2.0
2.0
2.0
2.0
2.0
Yield Recrystalllzlng
(X) Solvent
46 n-Hexane
43 n-Hexane
47 MeOH
36 Chloroform
55 Chloroform
cci4
67 CCl4a
54 CCl4a
49 CCl4a
84 CCV
33 CC14
Reference
Sanderman et al.,
Sanderman et al..
Norstrom et al..
Norstrom et al..
Norstrom et al.,
Gara et al., 1979
Crist and Moseman
Crist and Moseman
Crist and Hoseman
Crist and Hoseman
Crist and Hoseman
1958
1958
1977
1977
1977
. 1977
. 1977
, 1977
. 1977
. 1977
-------
tography or preparatlve-TLC. Bell and Gara (1985) have synthesized all 87
TCDFs, PeCDFs, HxCDFs, HpCDFs and OCTA using this synthetic route.
Another method 1s the cycllzatlon of dlazotlzed chlorophenoxy-o-an1l1nes
(Gray et al., 1976; Kurokl et al., 1984). Commonly, a nltro-chlorlnated
dlphenyl ether 1s obtained by reaction of the appropriate potassium halo-
phenolate and the halonltrobenzene at 120°C for 15 hours. The nltro group
Is converted to an amlno group (CH-COOH/Fe for 15 hours), dlazotlzed and
O
cycllzed with Isoamyl nitrite 1n tetrachloroethylene (80°C for 15 hours).
The crude product 1s purified by column chromatography and recrystalUzed.
Overall yields are typically -6-10%.
A modification of the above 1s the base-catalyzed cycllzatlon of
hydroxy-PCBs In DMSO/KOH (Safe and Safe, 1984). The hydroxy-PCB can be pre-
pared from coupling of a chlorinated anlsldlne and 1-substltuted symmetrical
benzene at 120°C 1n Isoamyl nitrite for 18 hours followed by purification
and demethylatlon (BBrg/CHpCK at 20°C for 24 hours). It can also
arise from coupling of a 2-chloro-an1l1ne with a chlorinated anlsole In Iso-
amyl nitrite followed by purification and demethylation. The yields Includ-
ing the Initial coupling reaction range from 3-15%. The major advantage of
the latter modification 1s that only single Isomers are usually produced.
Bell (1983) has reported on the synthesis of uniformly labeled
13CTCDF, using the Ullman ether procedure or by phenoxlde displacement on
an lodonlum salt. Yields commonly exceeded 30%. Synthesis of
14C-2,3,7,8- TCOF was first reported by Gray (1981).
Pyrolysls of octafluorodlbenzothlophene-5,51-dioxide affords perfluoro-
dlbenzofuran (Chambers et al., 1968).
1926A 2-47 06/21/86
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1,2,3,4-TFDF 1s prepared from hexafluorobenzene and o-!1th1oan1sole.
The o-methoxy blphenyl was first formed, then the free phenol (by reaction
with aluminum chloride) and finally the TFDF by refluxlng In with potassium
carbonate and dimethylformamlde (Brown et al., 1967).
1926A 2-48 06/21/86
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3. SAMPLING AND ANALYTICAL METHODS
3.1. SUMMARY
The major thrust of research In analytical methods for the PHDFs has
been toward specific synthesis, separation and sensitive detection of cer-
tain toxic PCDFs and PBDFs, especially the 2,3,7,8 derivatives. One such
analytical method Is the cap1llary-GC/MS technique developed by scientific
teams headed by Buser and Rappe (1979); many groups have now used this tech-
nique.
Earlier, the lack of analytical standards severely hampered the quantl-
tatlon and Identification of specific Isomers, though methods of quantHat-
Ing the total Isomers containing a given number of halogens (GC/MS) and
total PCDFs (by perchlorlnatlon) have been developed. Since many compounds
have been shown to co-elute with PCDFs on gas chromatographlc columns, and
compounds (for example, chlorinated dlphenyl ethers) may Interfere with the
mass spectral analysis, relatively specific cleanup techniques have been
developed, often based on the use of aluminum oxide column chromatography
and Judicious choice of solvents, usually various combinations of n-hexane
and methylene dlchlorlde. Techniques have been developed to Identify PHDFs
1n PCBs, Yusho oil, chlorophenols, fly ash, Agent Orange, 2,4,5-T, wood
dust, animal tissues, gelatin, human and fish tissues, mothers' milk and
bovine milk, wastewater and PBBs. Other methods are not as sensitive or as
specific as caplllary-GC/MS. Of some utility as a screening test before
GC/MS 1s a radlolmmunoassay technique. A cytosol receptor bloassay also
shows promise.
Sampling of PHDFs Is comparatively underdeveloped. Charcoal and XAD-2
resin have been used to sample vapors of PCDFs. Modified EPA-5 sampling
trains are used to sample stack emissions.
1927A 3-1 06/23/86
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3.2. SAMPLING METHODS
Few sampling methods have been developed specifically for PHDFs since
Interest has been focused on the analysis of various environmental and com-
mercial samples obtained by the usual procedures. A review has been pre-
sented by Tlernan (1983). The federal procedures governing custody, safety,
transportation, data review, sampling and the analysis of PCDDs and PCDFs
have been summarized by Elly (1985).
3.2.1. Incinerators.
PCDFs and PCDDs In the vapor phase have been collected by adsorption
onto charcoal (Norlt Pk, 0.5-1 mm or Amberllte XAD-2, 20/50 mesh) contained
In a glass filter (Rappe et al., 1978b). The vapors were drawn through a
glass funnel Into the filter by means of a vacuum pump. Both adsorbents had
to be cleaned by prior Soxhlet extraction with methylene dlchlorlde for 24
hours. The PCOFs and PCDDs were desorbed by Soxhlet extraction with methy-
lene dlchlorlde for 10 hours. Partlculate PCDD or PCDF can be trapped on
glass wool filters that can be eluted with toluene (Ahllng et al., 1977).
PCDFs and PCDDs In the vapor phase were also collected after this filter by
adsorption onto Chromosorb W coated with 30% Aplezon M; the columns can be
eluted with hexane and toluene. PCDF and PCDD partlculates and vapors have
also been collected 1n Implngers similar to a U.S. EPA method 5 sampling
train (OHe et al., 1977, 1982; Cavallaro et al., 1980; G1zz1 et al., 1982;
Redford et al., 1983; Wang et al., 1983; Benfenatl et al., 1983; Chlu et
al., 1983; Taylor et al., 1983; Ballschmlter et al., 1984; Brocco et al.,
1984; Clement et al., 1985a; Halle et al., 1985; Weeraslnghe et al., 1985a;
Scheldl et al., 1985). Only three groups of Investigators have attempted to
characterize the collection of PCDFs In the segments of their sampling
train. The closest approximation to the EPA-5 sampling train (Clement et
1927A 3-2 06/23/86
-------
al., 1985a) was the addition of two Flor1s1l (10 g) cartridges after the
third Implnger. In general, 95% of the total PCDD/PCDF was found In the
Implngers, -5% on the filter before the first Implnger, and 0.3% on the
FloMsIl cartridges. The backup FloMsIl cartridge contained 5% of PCDF
found In the front cartridge. The volatile PCDFs were relatively enriched
on the Florlsll filter compared with the prefllter and Implngers. However,
the extraction recovery of spiked 1,2,3,4-TCDD, 2,3,7,8-TCDD, 2,3,7,8-TCDF,
a PeCDD, a HxCDD, a HpCOD and the OCDD averaged only 43%.
In another sampling train (Ballschmlter et al., 1984), the sequence of
components was a glass fiber filter, a condenser at 15-18°C, and two Implng-
ers containing methoxyethanol at 0°C. The PCDD/PCDF ratio on the filter and
1n the condensate were similar, but PCDFs were much more concentrated than
PCDDs 1n the Implngers with no HpCDD/HpCDF and OCDD/OCDF present here. This
confirmed the vapor phase nature of the condensable fraction. The overall
collection efficiency was not determined.
The sampling train utilized by Tlernan et al. (1985) comprised, 1n
order, a heated sampling probe, a heated cyclone, a heated glass fiber fil-
ter, and a series of eight Implngers at 0°C, the first two containing water
and with XAD-2 traps Intervening between empty Implngers 3, 4 and 5. Sodium
arsenlte solution was placed 1n Implngers 5 and 6, Implnger 7 was empty, and
Implnger 8 was charged with silica gel. Known amounts of 1,2,3,4-TCDD (1.3
vg) and 1,2,4,8-TCDF (1 yg) were vaporized and collected with 92 and 97%
efficiency, respectively. Most of the PCDDs and PCDFs (-62%) were col-
lected 1n two XAD-2 traps. The recovery of spiked 37Cl4-2,3,7,8-TCDD
was <80% for the acetone back rinse, the cyclohexane back rinse and the
front XAD-2 trap.
1927A 3-3 06/23/86
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The sampling method recommended by the Swedish EPA Is also based on a
modified EPA-5 train (Beryvall and Jansson, 1986). The sampling equipment
Is fortified before sampling with known amounts of 13C-surrogate compounds
1n the filter, condensate or adsorbent (XAD-2) to act as Internal standards.
Fly ash samples have been collected from different points In flue gas
streams by electrostatic precipitation (Buser et al., 1978b). Since the
highest PCDD and PCDF levels were found from samples collected at flue gas
temperatures of 200-260°C, gas stream temperature should be measured during
sampling. Procedures to assure representative analysis of the collected
samples have not been developed. It 1s obviously desirable, however, to
analyze as much of the sample as possible. Soot type samples are generally
sampled with Kleenex tissues (Rappe et al., 1983c).
Liquid samples should be thoroughly shaken or ultrasonlcated before
cleanup and analysis. If liquid samples contain solid matter. It 1s prob-
ably advisable to filter the solution and perform analysis on the filtrate
and retain the dry solid. The choice of solvent to elute any adsorbed PCDFs
depends on the nature of the solid. Probably methylene dlchlorlde will be
adequate for most situations; recovery tests should be performed to choose
the correct solvent.
Organ and tissue samples are usually preserved In 4 or 10% buffered
formalin (Nagayama et al., 1977). Both tissue and formalin should be ana-
lyzed for PCDFs. Another much-used alternative 1s to freeze the tissues at
-70°C (Albro et al., 1985).
3.3. GC METHODS FOR THE PCDFs
3.3.1. Packed Column GC. Because certain PCDF Isomers are extremely
toxic to animals (Chapter 7), Interest In the analysis of these compounds
has Increased. Much of the early work was hampered by losses Incurred dur-
1927A 3-4 06/24/86
-------
ing sample cleanup and analysis, and by the fact that the PCDF peaks often
coincided with the peaks of chlorinated dlphenyl ethers containing two more
chlorine atoms (Crummett and Stehl, 1973). Thus, a confirmative spectro-
scoplc technique, such as mass spectroscopy using multiple 1on monitoring,
must be used to prove the specificity of the separation. QuantHatlon with-
out such confirmation w1lT yield maximum possible values of the contaminant
rather than the true value. The contaminant must be Identified before valid
quantification can occur. These problems have been reduced by the use of
capillary GC, which gives more specific separation.
The presence of PCDFs 1n PCBs was first deduced by Vos et al. (1970), on
the basis of GC/HS evidence. A 2 mg sample of PCB 1n hexane was placed on a
Flor1s1l column and eluted with hexane followed by 5% ethyl ether 1n hexane
until all PCBs were eluted. The PCDFs were eluted with 25% ether In hexane
and the eluate was evaporated. The residue was then chromatographed on a
Pyrex column (5 ft x 1/8 1n) containing 10% DC 200 on Gas Chrom Q (80/100
mesh) held at 200°C, with nitrogen as carrier gas at 50 mi/mln and the
separated peaks subjected to mass spectroscopy. Only the Clophen A-60 and
Phenoclor DP-6 PCBs contained PCDFs since they were the only ones that
caused a toxic response In the chick edema assay. The Investigators could
not separate or Identify the specific Isomers Involved. A similar procedure
was also utilized by Bowes et al. (1973) to determine PCDFs 1n PCBs. Before
GC/MS analysis, however, the three eluates described 1n Vos et al. (1970)
were subjected to further fractlonatlon on an aluminum oxide column with an
Initial elutlon with 1% methylene chlorlde/hexane to collect the PCBs and
then with 20% methylene chlorlde/hexane to elute the PCDFs. Bowes et al.
(1973) also analyzed the eggs, fat and liver of sea gulls for PCDF content.
Samples were freeze-drled, ground with sodium sulfate (2:1) In a heavy glass
1927A 3-5 06/23/86
-------
mortar, extracted using the Soxhlet procedure with a hexane/acetone azeo-
trope, the solution concentrated and cleaned up by passing over sulfurlc
acid-Impregnated CelHe before concentrating and continuing the cleanup on
the FloMsIl column mentioned 1n the Vos method. No account of recoveries
were given.
The lack of analytical standards for the PCDFs hampered further analy-
sis, but still did not prevent Roach and Pomerantz (1974), Bowes et al.
(1975a,b), Nagayama et al. (1975, 1976) and Kuratsune et al. (1976) from
establishing the presence of PCDFs 1n many PCBs, In Kaneml oil and 1n Yusho
patients exposed to Kaneml oil, from the mass spectrum alone, without stan-
dards. Morlta et al. (1977a) postulated the presence of the 2,3,7,8-TCOF on
the basis of GC retention-time data for an authentic sample, but the levels
quoted were maximal because of the relatively low resolving power of the
packed GC column used. Morlta et al. (1977a) eluted the activated Florlsll
column with hexane and then 95:5 hexane/acetone to remove the PCBs and col-
lected the PCDFs by elutlng with pure acetone. Recoveries were >90%. This
procedure had also been recommended by Vos et al. (1970). Morlta et al.
(1977a) then used the MS technique to quantltate the PCDFs using the parent
Ions.
The analysis of PCDFs In chlorophenols also was similarly Impeded by
lack of authentic standards. Nevertheless, PCDFs were found 1n chlorophe-
nols (Firestone et al., 1972; Pllmmer et al., 1973; Schwetz et al., 1974a,b;
Buu-Hol et al., 1972) 1n the crude base-Insoluble organic extract of the
chlorophenol. Buu-Ho1 et al. (1972) showed that consideration of the entire
spectrum allowed discrimination from other Interfering compounds, whereas
reliance on specific Ions led to errors. For example, the mass spectrum of
2,2',3,3',4,4',5,5',6-nonachloro-6'-hydroxyd1phenyl ether showed loss of two
1927A 3-6 06/23/86
-------
chlorine atoms to give a peak that coincides with the molecular 1on of
HxCDFs (m/e 372). The presence of such ethers can be verified by methyl-
atlon and subsequent GC/MS (Pllmmer et al., 1973). Typically (Firestone et
al., 1972), chlorophenol was converted to the phenolate by reaction with
sodium hydroxide, the PCOFs extracted In petroleum ether, the organic layer
washed, concentrated and transferred to an aluminum oxide column, the PCDFs
being eluted by the original procedure of Vos et al. (1970) plus a final
dlethyl ether elutlon. The concentrates were then partitioned between
petroleum ether and sulfurlc add, the organic layer neutralized with sodium
bicarbonate and the solutions evaporated and taken up 1n Isooctane for GC/MS
and EC/GC analysis. Recovery of 2,3,7,8-TCDD was 33%; omission of the sul-
furlc acid step Increased recovery to 63%. No recoveries were cited for
PCDFs. Firestone et al. (1972) used a 6 ft x 1/4 Inch Pyrex column packed
with 5% OV-101 on 80/100 mesh Chromosorb W (HP) under temperature programing
at 10°C/m1n to 250°C. The sulfurlc acid step was subsequently eliminated
(PUmmer et al., 1973). Capillary GC/MS Is discussed 1n Section 3.4.2.3.
Vlllanueva et al. (1974) found HpCDFs and OCDF 1n hexachlorobenzene by
using aluminum oxide column chromatography before GC/MS and using the
cleanup technique of Firestone et al. (1972). The Pyrex GC column
(10 ft x 6.3 mm OD) was packed with 3% SE-30 on 80/100 mesh Chromosorb W
(AW/DMCS) at 230°C, using a helium carrier (70 mn/mln). The Injector
temperature was 230°C. The recovery of the OCDF from the cleanup procedures
was -72%, much lower than for the OCDD.
Similarly, PCDFs were Identified In 2,4,5-T and Agent Orange. The
2,4,5-T ester formulation was diluted with chloroform and placed on a PX-21
foam-charcoal column. The nonpolar components In the formulation were
eluted with chloroform. Moderately polar compounds were then eluted with
1927A 3-7 06/23/86
-------
benzene. The PCDDs and PCOFs were finally recovered by elutlon of the
column with 1:1 toluene/benzene and then the eluate concentrated before
aluminum oxide chromatography. The aluminum oxide column was washed with
petroleum ether before the concentrate was placed on the column. The PCDOs
and PCDFs were eluted with 1:4 methylene dlchlorlde/petroleum ether. The GC
column was a 0.91 m x 4 mm ID Pyrex column packed with OV-7 on Chromosorb W
(HP) at 200°C. The temperature of the Injector was 250°C. The average
recovery for the 2,3,7,8-TCDD was >91%. Although the presence of PCDFs was
Inferred from the MS data, again no specific Isomer Identification was made
and no recoveries of PCDFs found (Hucklns et al., 1978). AhUng et al.
(1977) used the same technique but eluted the PCDDs and PCDFs with toluene
and confirmed the existence of PCDFs using the available authentic samples
and standards.
PCDF standards were probably first used by Hutzlnger et al. (1973) and
by Crosby et al. (1973) to Identify the reductive dechlorlnatlon products of
2,8-DCDF Irradiated In hexane solution by UV light. DCDFs, TrCDFs, TCDFs
and OCDF were not found 1n various tissues of wildlife 1n the Bay of Fundy/
Gulf of Maine area by ZHko (1972). Tissue samples were ground In anhydrous
sodium sulfate and Soxhlet extracted with hexane; the solution was evapo-
rated and applied to an aluminum oxide column. The column was eluted with
hexane, the effluent concentrated and then applied to a silicic acid column,
eluted with hexane and then with 10% ether/hexane. The hexane eluate was
further chromatographed on aluminum oxide and then eluted by 20% methylene
dlchlorlde/hexane. The 6 ft x 4 mm GC column was packed with 4% SE-30 on
Chromosorb W (AW; 60/80 mesh) at 250°C. The estimated recoveries of DCDFs,
TrCDFs and TCDFs were each 94%, and that of OCDF was 83% over the spiking
1927A 3-8 06/23/86
-------
range of 0.23-1.41 ng/g. These column chromatographlc conditions, when com-
pared with those cited previously, appear too mild to produce such good
recoveries.
3.3.2. Representative Sample Preparations and Methods of Analysis.
3.3.2.1. INTRODUCTION — In this section, the various optimized ana-
lytical treatments of selected samples will be presented with an Indication
of how the method evolved. In all the methods, the final step 1s GO/MS.
Thus, the mass spectrometrlc characteristics of the PCOFs are discussed
first, ending with the various treatments for selected samples.
Reviews of the analytical methodology to determine PCDFs have been
authored by Crummett (1983), Albro and Parker (1980), Tlernan (1983), Rappe
et al. (1983a), Buser (1985) and Buser et al. (1985). A step-by-step pro-
cedure for extract1on/cleanup/GC-MS of a typical sample Is provided by
Taylor et al. (1983). Albro et al. (1985) have presented a system to ensure
a valid Interlaboratory comparison of PCDFs and PCDDs 1n human adipose
tissue.
3.3.2.2. MASS SPECTROMETRY OF CHLORODIBENZOFURANS —Mass spectro-
metry, using an electron Impact source, Involves the fragmentation of a
molecule Into positively or negatively charged Ions (usually singly charged)
that are collected quantitatively so that the resultant plot of normalized
lon^current versus mass/charge ratio Is an Identifying fingerprint for the
compound. The technique can be made quantitative by external and Internal
standards techniques.
3.3.2.2.1. Chlorine Isotope Patterns — Before discussing the mass
spectra of PCDFs, It will be helpful to review the patterns of 1on Intensi-
ties that result from the two chlorine Isotopes at 35 and 37 amu. Since
these Isotopes have natural abundances of 75.5 and 24.5%, respectively (a
1927A 3-9 06/23/86
-------
ratio of 3.08), any Ion containing one chlorine will give peaks 1n the mass
spectrometer at two mass numbers separated by two mass units; If the Inten-
sity of the first peak is 100%, the Intensity of the second (higher) mass
peak will be 32% (100%/3.08). If more than one chlorine atom Is present,
the binomial distribution Is convoluted with the Isotoplc abundance to give
the appropriate peak pattern. Table 3-1 shows the relative intensity dis-
tributions for ions containing 1-8 chlorines. Thus, by matching a pattern
in an unknown mass spectrum with a pattern given 1n Table 3-1, it is usually
possible to determine the number of chlorine atoms present 1n an unknown
chlorinated aromatic compound.
3.3.2.2.2. Mass Spectra of PCDFs — The mass spectra of PCDFs are
dominated by the presence of many chlorine Isotope patterns and by an abun-
dant molecular 1on. In fact the molecular 1on 1s the most Intense peak 1n
the mass spectrum. The only fragmentation of the PCDFs is the loss of up to
thr^e chlorine atoms and the loss of CO (Buser, 1975). The fragment ions
are summarized 1n Figure 3-1, which also gives the average relative Inten-
sity of each 1on. Each Ion Is accompanied by the Isotoplc cluster of peaks,
each of which has a relative Intensity dependent on the number of chlorines
in it, their relative intensities given by the values in Table 3-1.
The complete mass spectra of TrCOFs to OCDF have appeared In the litera-
ture; they are given in Figure 3-1. For example, HxCDF has an intense mole-
cular 1on cluster at m/e 372. By convention the mass of the lightest mass
peak In an Isotoplc cluster Is referred to as the mass of that ion. The
loss of one and two chlorines gives peaks at m/e 337 and 302. The loss of
CO from each of these Ions gives peaks at m/e 309 and 274, and the loss of
another chlorine from the latter gives an ion at 239. The ion caused by the
loss of CO-C1 1s the daughter ion used in MS-MS (Ryan et a!., 1985a,b,c).
1927A 3-10 06/23/86
-------
TABLE 3-1
Relative Intensities of Ions
Containing from One to Eight Chlorine Atoms
(Ions of relative Intensity 1% or less have been omitted)
Number of Mass of
Chlorines Fragment
1
M
2
M
M
3
M
M
M
4
M
M
M
5
M
M
M
M
6
M
M
M
M
7
M
M
M
M
M
8
M
M
M
M
M
M
4- 2
M
4- 2
4- 4
M
4- 2
4- 4
4- 6
M
4- 2
4- 4
4- 6
M
4- 2
4- 4
4- 6
4- 8
M
4- 2
4- 4
4- 6
4- 8
M
4- 2
4- 4
4- 6
4- 8
4- 10
M
4- 2
4- 4
4- 6
4- 8
4- 10
Relative
Intensity, %
100
32
100
65
10
100
97
31
3
77
100
49
11
62
100
65
21
3
51
100
81
35
8
44
100
97
52
17
3
34
88
100
65
26
7
1927A
3-11
06/19/86
-------
M+
(100XJ-
-e
•*-(20X)
-Cl
N-C1.
H-35
<5X)
-Cl
M-C12
H-70
(5X)
-CO
-CO
M-COC1
M-63
(25X)
-Cl
M-COC12
M-98
(10%)
-Cl
H-COC13
M-133
(25X)
FIGURE 3-1
Mass Spectral Behavior of PCDFs
Percent Relative Intensities of Each Ion Are Given In Parentheses
Source: Buser, 1975
1927A
3-12
06/23/86
-------
The Isotoplc patterns associated with each of these Ions agrees with the
expected number of chlorine atoms as given by the relative Intensities of
the clusters In Figure 3-1; for example, the cluster at m/e 239-245 has
peaks of relative Intensities characteristic of three chlorines (see Table
3-1). Since three chlorines have been lost from a molecule containing six
chlorine atoms to form this 1bn, It should Indeed have a 3-chlor1ne pattern.
The fragment Ions of the MCDFs to OCDF are summarized In Table 3-2. In
addition, the number of chlorine atoms 1n each 1on Is given In parentheses
so that the appropriate Isotoplc pattern can be noted. Clearly, mass num-
bers of Ions characteristic of the various PCDFs can be specified from data
such as these. In addition, using such data, PCDFs can be Identified with
accuracy 1n environmental samples.
3.3.2.2.3. Mass Spectra of Isomerlc PCDFs — Characterizing 1somer1c
PCDFs by electron Impact MS alone 1s not possible with present technology
(Buser, 1985). For example, 2,3,8-TrCDF and 2,4,6-TrCDF have Identical mass
spectra (Gray et al., 1976). This 1s to be expected 1f the fragmentation
mechanisms operating In these two compounds are not sensitive to the
positions of the chlorine atoms. Table 3-2 summarizes the Intensities of
the four major Ions of various PCDFs (Gray et al., 1976). All Isomers,
except 2,3,6,8-TCDF, regardless of the position or number of chlorine atoms,
have the same losses of COC1 and COCK from their molecular Ions within an
experimental error of ^20% of the observed peak Intensity. For 2,3,6,8-
TCDF, the reported Intensities of the M-COC1 and M-COC1- Ions are much
greater than for the other compounds. The spectrum should be remeasured.
Parker et al. (1983) have shown that the molecular Ion carries 52*5% of the
total Ion current for PeCDFs to OCDF 1n magnetic sector MS.
1927A 3-13 06/23/86
-------
TABLE 3-2
Percent Intensities of the Major Ions In the
Mass Spectra of Isomerlc PCOFsa
2,3,8-Cl3
2,4,6-Cl3
2.3.7-Cl3
2,3,9-Cl3
2,3,6,8-Cl4
2,4,6,8-Cl4
2,3,7,8-Cl4
1,2,7,8-014
1,2,4,7,8-C15
1,3,4,7,8-C15
*• J
Average
M+
100
100
100
100
100
100
100
100
100
100
100
M-COC1
23
23
25
25
41b
22
19
17
22
18
22 ± 3
M-COC13
14
14
24
22
56b
22
16
22
26
22
20 ± 4
M++
11
11
22
20
23
22
12
20
-
24
18 i 5
aSource: Adapted from Gray et al., 1976
bOut!1ers
1927A 3-14 06/23/86
-------
Even though MS alone cannot distinguish among Isomers, GC can separate
many of them from one another. In fact, many GC retention values for spe-
cific PCOF Isomers have been published (Section 3.3.2.3.).
3.3.2.2.4. High Resolution Mass Spectrometry of PCDFs — Using double
focusing mass spectral Instrumentation, H Is possible to measure the mass
of an 1on to 6 or 7 significant figures. Because the various Isotopes have
masses that differ slightly from their nominal value (I.e., chlorlne-35 has
a mass of 34.9689 amu), such high mass accuracy permits calculation of the
exact elemental composition of an 1on from Us exact mass. The exact mole-
cular weights of the various PCDFs are given 1n Table 3-3.
Because of the high aromatlcHy and the large number of chlorine atoms,
these exact masses are up to 0.25 amu below the normal mass (see
C-^ClgO). Thus, these Ions can easily be distinguished from an 1on at
the same nominal mass that may come from a relatively saturated compound.
For example, at m/e 440, a hydrocarbon 1on may be C00H..,, which has an
•3d DO
exact mass of 440.4382; this Is 0.6924 amu higher than the corresponding
OCDF molecular Ion. When taken together with the chlorine 1sotop1c patterns
and adequate chromatographlc separation (see Table 3-1), these exact masses
(Table 3-3) are almost specific Indicators of the presence of PCOFs 1n
samples. The chlorinated dlphenyl ethers lose 2 chlorines 1n the Ion source
of a mass spectrometer to form a compound with the same elemental composi-
tion as a PCDF. The molecular 1on of the ether must be monitored to prove
whether the Interference Is present or not.
3.3.2.2.5. Specific Ion Monitoring Mass Spectrometry — Buser (1975)
based his quantHatlon of PCOFs In commercial chlorophenols on selected 1on
monitoring of the molecular Ions (304, 338, 372, 406 and 440; see Table
3-2). Figure 3-2 1s an example of such data; the peaks attributable to the
1927A 3-15 06/23/86
-------
TABLE 3-3
Exact Molecular Weights of the PCDFs
(masses used to calculate these molecular weights:
C = 12.000000, H = 1.007825, Cl = 34.968855 and 0 = 15.994915)
Compound Molecular Weight
C12H7C10 202.0185
C12H6C120 235.9796
C12H5C130 269.9406
C12H4Cl40 303.9016
C12H3C150 337.8627
C12H2C160 371.8237
C12HC170 405.7847
C^ClgO 439.7458
1927A 3-16 06/23/86
-------
B
0)
o
CO
"O
c
3
.£}
I
J2
DC
100
•0
ao
40
20
o
100
•0
ao
40
20
0
100
•0
20
0
100
K>
20
0
100
•0
207
' I ' ' T 1 I ' ' • ' I '
160 200 260
300 360 400
460
*•'
x» || m
160
1 . .*? fr '
200
2SO
300
360
400
460
206
160
1 ' I
200
I ' ' ' ' 1 '
250 300
350
r I"
400
"I
450
X»
I
160
200
250
SCO
350
1 I '
400
460
I
440
"1^
160
177
*70
200 260 300
FIGURE 3-2
ITOJ 4O6
I i Fi i l'i
360
400
Low resolution mass spectra of (A) TrCDF, (B) TCDF, (C) PeCDF, (D)
HxCDF and (E) OCDF. In all cases the chlorine substitution pattern Is not
known. The fragmentation patterns are explained In Figure 3-1.
Source: (A) HHes and Lopez-Avlla, 1979; (B.C.D) Bowes et al.. 1973; (E)
Hutzlnger et al., 1972b.
1927A
3-17
06/23/86
-------
HxCDF through OCDF are Indicated. Figure 3-3 shows that m/e 372 (Cl&)
gives a response for the OCDF Isomer (see retention time 12 m1n). This Is
because OCDF has an M-C1- 1on at m/e 370, which has a more Intense first
Isotope peak of 372. The specific and multiple 1on monitoring modes allow pg
sensitivity to be attained and 1s the best technique to use for quantHatlon
of PCDFs after prior separation on a capillary GC column (Buser et al.,
1985). For example, Rappe et al. (1977) showed that the 2,3,7,8-TCDF Isomer
was probably present In Yusho oil (see Figure 3-3 and Chapter 4).
3.3.2.2.6. Other Types of Mass Spectroscopy of PCDFs — Electron-
Impact (El) MS (50-70 eV at 250°C) 1s the preferred analytical mode for
TCDFs with sensitivity 1-10 pg by specific Ion monitoring. Decreasing sen-
sitivity occurs for the higher chlorinated species (Rappe et al., 1983a;
Buser et al., 1985). Negative chemical lonlzatlon (NCI) with methane as
reagent gas (0.35 torr, 180°C) 1s very sensitive (1-2 orders of magnitude
more sensitive than electron Impact MS) for all PCDF from tetra- to octa-,
but requires more cleaning of the Ion source, or a specially designed 1on
source, and 1s not as reproducible as an electron Impact source (Buser et
al., 1985; Rappe et al., 1983c; Mass et al., 1981; Kuehl et al., 1981). The
base peak 1s usually M" with (M%H«-C1») Ions, both singly and mul-
tiply charged. As In electron Impact MS, NCI spectra of Isomers are Identi-
cal (Buser et al., 1985). The Ions used for specific 1on monitoring are
M or M~+2. Rappe et al. (1983c) have reported that Isomers have dif-
ferent response factors In the NCI mode. The NCI/El response ratios range
from 0.86 for 1,2,7,9-TCDD to 11 for 2,3,6,8-TCDF. The El response Is low-
est for 1,2,6,7-TCDF being 59% of the highest response for a tetra-Uomer,
1,3,6,8-. The NCI response 1s lowest for 1,2,7,9-tetrabelng 6.8% of that
for the most responsive tetrachloro-lsomer, 2,3,6,8-. For PeCDF, the
1927A 3-18 06/24/86
-------
m/e 440
m/e 406
m/e 372
m/e 338
octa-
X20
hepta-
X100
hexa-
X50
m/e 304
6
minutes
12
m/e 304
6
minutes
12
FIGURE 3-3
Mass chromatograms of (A) 2,3,4,6-tetrachlorophenol and (B) pentachloro-
phenol showing the elutlon of PCDF (m/e 304 = tetra-, m/e 338 = penta-, m/e
372 = hexa-, m/e 406 = hepta-, and m/e 440 = OCDF).
Source: Buser, 1975
3-19
06/23/86
-------
2,3,4,6,8-lsomer has an 81% response relative to the most El responsive Iso-
mer, 1,2,4,6,8-; and 48% In the NCI mode. For HxCDF, the 1,2,3,4,7,8- 1so-
mer Is 65% of the El response of the most responsive Isomer, 2,3,4,6,7,8-.
The corresponding NCI data for the 1,2,3,4,7,8- Isomer Is 73% of the most
responsive 1,2,4,6,7,8-lsomer. For two hepta-1somers, the El responses were
equivalent, but for the NCI responses, the 1,2,3,4,6,8,9- Isomer was 77% of
the response of the 1,2,3,4,6,7,8-lsomer (Rappe et al., 1983a). A Townsend
discharge Ion source (negative oxygen chemical 1on1zat1on) seems to allow a
better discrimination of Isomers 1n each class (Mass et al., 1981; Ch1u et
a>., 1983; Lao et al., 1985). Negative methane/oxygen chemical 1on1zat1on
Is still being evaluated (Mass et al., 1981). The negative 1on atmospheric
pressure lonlzatlon mass spectrometry for 2,3,7,8-TCDF has been reported
(Korfmacher et al., 1983). Though the detection limit was 0.5 pg, there are
no significant mass spectral differences In the TCDF Isomers having 4:0, 3:1
or 2:2 chlorine ring substitution patterns.
Another technique of promise Is MS/MS (Voyksner et al., 1983; Sakuma et
al., 1985; Shushan et al., 1985, Ryan et al., 1985a,b,c). The first ana-
lyzer provides nominal mass separation of the analyte 1on (usually the
molecular Ion) from the matrix, the selected 1on 1s then fragmented by col-
lision (He, 2xlO~* mm Hg) Into specific daughter Ions (M-C1, M-COC1) thus
eliminating background noise. However, though cleanup problems are mini-
mized, specific Isomers cannot be distinguished through MS/MS alone, thus
necessitating prior GC separation. MS/MS results appear to be consistently
high with respect to GC/MS or GC/MS/MS for 2,3,7,8-TCDF (Voyksner et al.,
1983). An MS/MS arrangement preceded by a short capillary column allows
fast screening of TCDF congeners at 20-400 fg levels (Sakuma et al., 1985),
with the SP-2340 column used for separating specific Isomers.
1927A 3-20 06/23/86
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3.3.2.3. CAPILLARY GAS CHROMATOGRAPHY -- The environmental chemistry
groups headed by Buser, Bosshardt and Rappe have been largely responsible
for the development of capillary GC techniques as applied to PCDFs.
*. The GC/MS analyses noted 1n Section 3.3.2.1. were ultimately deficient
because the various PCDF Isomers were not resolved on packed GC columns, and
often only the total PCOFs could be estimated. Since certain PCDFs are more
toxic than others, separation of Isomers Is essential.
In view of the fact that each chlorinated class of PCDFs Includes sev-
eral congeners (with the exception of OCDF), the separation and quantHatlon
of other Isomers constituting that chlorinated class (for example quantHa-
tlon of 2,3,7,8-TCDF 1n the presence of the 37 other TCDFs) Is difficult.
Conditions for Isomer-speclf1c quantHatlon are developed using calibration
standards containing all of the Isomers that may co-elute with the Isomer of
Interest. A valid Isomer-speclf1c analysis will be accompanied by data (a
mass chromatogram) that demonstrate the appropriate conditions that permit
the separation of the Isomer of Interest from all other possible co-elut1ng
Isomers. If such calibration data 1s lacking, the Identification of spe-
cific Isomers must be regarded as tentative. Concentrations of Isomers cal-
culated on the basis of unvalldated 1somer-spec1f1c data should be regarded
as upper limits and therefore the concentrations are approximate.
Buser (1976a) was the first to use Pyrex capillary columns to produce
the desired separation of the PCDF Isomers. These columns with different
stationary phases (OV-101, OV-17 and Sllar IOC) allowed lower operating
temperatures (205-225°C) as well as Increased resolution compared with con-
ventionally packed columns. The column characteristics are provided 1n
Table 3-4. The average film thicknesses were 0.1 ym. The OV-101 Is a
nonpolar methyl slUcone; the OV-17 1s a semlpolar methyl phenyl slUcone;
1927A 3-21 06/23/86
-------
TABLE 3-4
Characteristics of Some Glass Capillary Columns
for Separating PCDFs*
Characteristic
Length (m)
I.D. (mm)
Temperature (°C)
Retention time
OV-101
22
0.32
225
33.4
Column
OV-17
22
0.34
225
31.8
S1lar IOC
22
0.34
205
26.9
(OCDD) (m1n)
Retention Index
(OCDD)
3055
3420
3645
Theoretical plates
HETP (mm)
Carrier (atm)
42,500
0.52
He(0.5-0.6)
43,400
0.51
He(0.5-0.6)
34,000
0.65
He(0.5-0.6)
*Source: Buser, 1976a
1927A
3-22
06/19/86
-------
and the Sllar IOC 1s a highly polar cyanopropyl slUcone. The capillary
column was linked to the MS by a fused 0.15 mm ID platinum capillary at
250°C leading directly Into the Ion source of the MS. The compounds were
detected by MS of the molecular Ions. Isothermal conditions were found to
be suitable for the separation of the PCDFs, and the reproduclblllty 1n
quantHatlon was better than that obtained with temperature-programmed GC.
The chromatography of Aroclor 1268 [which contains the most highly chlorin-
ated PCBs (up to Cl,_)3, under the same conditions given In Table 3-4,
produced complete PCB elutlon within 10 minutes on the Sllar IOC column; the
peaks were also better resolved than they were with a support-coated open
tubular column. On this column, however, there was some overlapping of PCDD
and PCDF Isomers that did not occur on the others and there was also some
reversal of elutlon order. When the same sample of PCDDs/PCDFs was Injected
on all columns, the OV-101 column always resolved more peaks. Better reso-
lution of the less chlorinated Isomers occurred on the OV-101 column, but
the Sllar IOC was best for the most highly chlorinated Isomers. The OV-17
column gave the best resolution of the Cl, Isomers. Relative retention
times of PCDFs and PCDDs on these columns are presented In Table 3-5. All
samples were Introduced by an Isothermal splltless technique.
Rappe et al. (1977) used the technique to analyze for PCDFs 1n Yusho oil
but used 22 m x 0.36 mm ID glass capillary columns with OV-101 and OV-17
stationary phases and specific mass detection of the molecular Ions (Figure
3-4). It was shown that the toxic 2,3,7,8-TCDF was probably one of the main
PCDFs 1n Kanem! oil causing Yusho disease, constituting 30% of the total
TQDFs. The 2,3,4,8-TCDF co-elutes with 2,3,7,8-TCDF on this column (Mazer
et al., 1983a) and thus the Identification Is ambiguous. Buser et al.
(1978a) used OV-17 capillary columns of 25 and 50 m length (0.36 mm ID) to
1927A 3-23 06/23/86
-------
TABLE 3-5
Relative Retention Times of PCDFs and PCDDs on
the Capillary Columns of Table 3-4*
Compounds
TCDDs
TCDFs
PeCDDs
PeCDFs
HxCDDs
HxCDFs
HpCDDs
HpCDFs
OCDD
OCDF
Relative
OV-101
0.13-0.18
0.10-0.14
0.20-0.27
0.17-0.24
0.32-0.41
0.29-0.42
0.56-0.63
0.51-0.65
1.00
0.978
Retention Times
OV-17
0.10-0.15
0.09-0.12
0.16-0.24
0.14-0.23
0.28-0.38
0.25-0.40
0.52-0.60
0.47-0.66
1.00
0.025
on Column
Sllar IOC
0.12-0.22
0.11-0.19
0.18-0.30
0.15-0.25
0.31-0.43
0.24-0.50
0.54-0.62
0.44-0.66
1.00
0.933
*Source: Buser, 1976a
1927A
3-24
06/19/86
-------
INJ
2,3,7,8-tetra-CDBF
CO
I
en
m/e 372
1
20 min
1 1
10
l~
0
CO
CO
cr<
FIGURE 3-4
Mass Chromatograms (OV-101 Glass Capillary Column at 197°C) of "Yusho" Oil Showing Elutlon of PCDF
(m/e 270 = TrCDF, m/e 304 = TCDF, m/e 338 = PeCDF and m/e 372 = HxCDF)
Source: Rappe et al.. 1977
-------
resolve the PCDF Isomers produced by PCB pyrolysls, quantHating by GC/MS
(Chapter 4). To achieve the best resolution, a complex temperature pro-
gramming scheme to achieve splltless Injection was adopted: 100°C, 2-mlnute
Isothermal; 15°C/m1nute to 145°C; 5°C/m1nute to 230°C for the 25 m column;
100°C, 2-mlnute Isothermal, 20°C/m1nute to 230°C for the 50 m column, with
helium carrier at 0.80 and 1.40 atmospheres, respectively. The samples were
Introduced splltlessly at 260°C. Optimal separations were achieved on the
50 m column. Similar techniques were utilized to analyze for PCDOs 1n fly
ash (Buser et al., 1978b; Chapter 4). PCDDs and PCDFs produced by pyrolysls
of chlorophenolates were detected (Rappe et al., 1978b), and In 2,4,5-T
esters as well as Agent Orange (Rappe et al., 1978c). PCDFs have been ana-
lyzed In fly ash and In pyrolyzed PCBs (Buser et al., 1978c; Chapter 4); 1n
Yusho oil and used Japanese PCB (Buser, et al. 1978d); 1n commercial chloro-
phenols (Rappe et al., 1978a); In the pyrolysates of specific PCB Isomers
(Buser and Rappe, 1979) and of chlorobenzenes (Buser, 1979); In tissues of
patients who suffered from Yusho disease (Rappe et al., 1979b); 1n the pro-
ducts of the palladium acetate synthetic method for PCDFs (Gara et al.,
1979); fish samples (Rappe et al., 1981); human adipose tissue, mothers'
milk, bovine fat and bovine milk (Nygren et al., 1986).
Thus, H appears that optimum separations are achieved with a 50 m x
0.36 mm ID glass capillary column coated with OV-17, operated with helium
carrier 1n the temperature programmed mode given by Rappe et al. (1978c).
Sometimes a combination of capillary columns, however. Is necessary. Rappe
et al. (1984) reported that a 60m Supelco SP-2330 capillary column could
separate most PCF Isomers. The toxic 1,2,3,7,8-PeCDF does co-elute with the
1,2,3,4,8-lsomer as does the toxic 1,2,3,4,7,8-HxCDF with the 1,2,3,4,7,9-
Vwrnier. These Isomers can be separated on less polar columns like OV-17 and
DB-5.
1927A 3-26 06/23/86
-------
Farrell (1980) attempted unsuccessfully to produce the thinly coated
columns mentioned previously (Rappe et al., 1978c). He did produce 25 m x
0.25 mm ID columns with film thicknesses of 0.4-0.5 ym OV-17 and OV-101,
but analyses took 4-5 hours at 220-260°C and at 34 cm/sec linear velocity of
nitrogen carrier. The problem may have been that the helium carrier was not
used, since when the hydrogen carrier was used a mixture of DCDFs, TrCDFs,
and TCDFs was resolved, the 2,3,7,8-TCDF being eluted In 6 minutes on a
50 m x 0.25 mm ID OV-101 column (0.5 wm liquid phase film) at 240°C and
100 cm/sec hydrogen carrier average velocity. Another deficiency of the
Farrell (1980) study was that he did not use the splHless Injection tech-
nique recommended by Buser (1976a). The splHless technique has now been
successfully used by other laboratories (Mazer and Hlleman, 1982; Mazer et
al., 1983a,b; Hlleman et al., 1985). Sources of PCOFs 1n the United States
have been summarized by Cantrell et al. (1986).
The kinds of Interferences expected 1n GC/MS of PCDFs are reviewed 1n
Smith and Johnson (1983), Rappe (1984) and Lau et al. (1985). The quality
assurance and control procedures to meet U.S. EPA requirements are embodied
In RCRA method 8280 (Donnelly et al., 1986). Here a SP-2250 or equivalent
column 1s recommended with multiple 1on monitoring and Internal standard
quantHatlon. The peaks to be monitored Include the following: TCDF,
3M.897, 321.894, 327.885, 256.933 and 258.930; PeCDF, 353.858, 355.855;
HxCDF, 389.816 and 391.813 for the PCDFs themselves, their (M-CO-C1) Ions,
and the molecular Ions for the Interfering chlorinated dlphenylethers as
well as the molecular Ions of chlorinated blphenylenes, PCBs and DDE
Isomers. All PCDF Isomers should be on hand.
1927A 3-27 06/23/86
-------
The most comprehensive recent 11st of retention times, flame lonlzatlon
and electron capture sensitivities for TCDFs to OCDF Is provided 1n Table
3-6 (Bell and Gara, 1985). Other lists can be found In Hazer and Hlleman
(1982); Firestone (1977a), Mazer et al. (1983a,b); Kurokl et al. (1984);
Safe and Safe (1984) and Llgon and May (1984). Hale et al. (1985) have
explained the retention behavior of all PCDFs In terms of number and posi-
tion of chlorines, relations between chlorines, and ring Interactions.
Keith et al. (1983) have described a thermally modulated electron-capture
detector that has higher sensitivity, better linear range and specificity to
PCDFs than the conventional electron-capture detector.
3.3.2.4. TREATMENT OF SPECIFIC SAMPLES — In the following methods,
the methods were generally validated by the spiking of known amounts of PCDF
and then measuring recovery. Such methodology does not necessarily measure
efficiency 1f an Initial pretreatment step 1s necessary to release PCDFs, as
fjw fly ash samples. The quality assurance and control necessary to achieve
valid results 1n the cleanup and column chromatographlc steps are provided
at length by Donnelly et al., (1986) 1n regard to an evaluation of RCRA
Method 8280. The experimental conditions for elutlon of compounds from the
"open" alumina mlcrocolumn are critical. A 60% methylene dlchlorlde 1n hex-
ane-elut1ng solvent was much more reproducible than a 10% when neutral or
basic alumina was used. Careful control of the water content was necessary
since deactlvatlon of neutral alumina caused weaker solvents to elute the
PCDFs and PCDDs thus leading to losses and the presence of unwanted Inter-
ferences.
3.3.2.4.1. PCB Formulations — The PCB (100 mg) was placed on an
aluminum oxide mlcrocolumn (basic aluminum oxide, Woelm, In a disposable
Pasteur pipette, 15 cm x 5 mm ID) and the bulk of the PCBs eluted with
1927A 3-28 06/23/86
-------
TABLE 3-6
Relative Retention Times (RRT) and Relative Response Factors (RRF)
for PCDFs Synthesized as Reference Standards3*".
The gas chromatographlc detectors were flame
1on1zat1on (FID) and electron capture (ECD).
Congener
1,2,3,4-
1,2,3,6-
1,2,3,7-
1,2,3,8-
1,2,3,9-
1,2,4,6-
1,2,4,7-
1,2,4,8-
1,2,4,9-
1,3,4,6-
1,3,4,7-
1,3,4,8-
1,3,4,9-
2,3,4,6-
2,3,4,7-
2,3,4,8-
2,3,4,9-
1,2,6,7-
1,2,6,8-
1,2,6,9-
1,2,7,8-
1,2,7,9-
1,2,8,9-
1,3,6,7-
1,3,6,8-
1,3,6,9-
1,3,7,8-
1,3,7,9-
1.4,6,7-
1,4,6,8-
Gas
SP-2330
RRT
±0.003
1.389
1.415
1.306
1.396
1.794
1.290
1.179
1.258
1.608
1.252
1.150
1.217
1.466
1.949
1.802
1.900
1.382
1.561
1.294
1.854
1.482
1.569
1.198
1.188
1.000
1.314
1.128
1.131
1.401
1.163
Chromatoqraphlc Column and Detector Usedc
ECD
RRF
±0.05
1.79
0.81
1.06
1.00
0.72
0.51
0.96
1.15
0.87
0.70
0.69
0.84
1.22
0.61
0.92
0.97
1.16
0.79
0.55
0.78
0.93
0.83
0.87
0.63
1.00
1.05
0.62
0.84
0.55
0.75
DB-5
RRT
±0.003
1.161
1.150
1.129
1.148
1.280
1.201
1.063
1.100
1.203
1.070
1.063
1.080
1.186
0.201
1.200
1.197
1.153
1.189
1.100
1.258
1.170
1.208
1.363
1.079
1.000
1.123
1.064
1.084
1.108
1.026
FID
RRF
±0.05
1.26
0.70
1.03
0.87
0.48
0.85
1.25
1.12
1.08
1.06
1.10
1.01
0.82
0.85
0.85
0.96
1.84
1.49
0.65
0.85
0.93
0.89
1.21
0.88
1.00
1.13
0.53
0.70
1.08
1.10
SP-2340
RRT
±0.003
1.413
1.452
1.338
1.446
1.870
1.313
1.194
1.283
1.608
1.278
1.165
1.244
1.515
2.074
1.915
2.006
1.444
1.637
1.336
1.953
1.551
1.633
2.420
1.216
1.000
1.345
1.150
1.145
1.459
1.189
ECD
RRF
±0.05
1.58
0.83
1.05
0.99
0.72
0.46
0.95
1.24
0.84
0.68
0.64
0.78
0.87
0.60
0.92
0.93
1.26
0.76
0.52
0.74
0.92
0.93
0.90
0.60
1.00
1.10
0.60
0.82
0.48
0.77
1927A 3-29 06/19/86
-------
TABLE 3-6 (cont.)
Congener
1,4,6,9-
1,4,7,8-
2,3,6,7-
2,3,6,8-
2,3,7,8-
2,4,6,7-
2,4,6,8-
3,4,6,7-
1,2,3,4,6-
1,2,3,4.7-
1,2,3,4,8-
1,2,3,4,9-
1,2,3,6,7-
1,2,3,6,8-
1,2,3,6,9-
1,2,3,7,8-
1,2,3,7,9-
1,2,3,8,9-
1,2,4,6,7-
1,2,4,6,8-
1,2,4,6,9-
1,2,4,7,8-
1,2,4,7,9-
1,2,4,8,9-
1,3,4,6,7-
1,3,4,6,8-
1,3,4,6,9-
1,3,4,7,8-
1,3,4,7,9-
1,3,4,8,9-
2,3,4,6,7-
2,3,4,6,8-
2,3,4,6,9-
2,3,4,7,8-
2,3,4,7,9-
2,3,4,8,9-
Gas
SP-2330
RRT
±0.003
1.586
1.304
1.978
1.609
1.866
1.706
1.390
2.149
1.997
1.837
1.961
2.524
2.058
1.659
2.394
1.960
2.023
3.017
1.809
1.471
2.111
1.706
1.759
2.330
1.760
1.427
1.923
1.660
1.615
2.327
3.074
2.418
1.843
2.906
1.568
2.127
Chromatographic Column and Detector Usedc
ECD
RRF
±0.05
0.94
0.79
0.72
1.15
1.05
0.73
0.67
0.68
3.94
4.17
7.27
4.03
2.90
5.02
4.31
3.70
2.24
3.99
3.84
3.66
4.77
6.13
4.64
2.72
2.67
2.58
3.76
3.79
5.43
3.28
3.79
4.08
3.10
7.36
3.50
4.36
DB-5
RRT
±0.003
1.152
1.105
1.234
1.071
1.198
1.137
1.035
1.254
1.573
1.569
1.596
1.801
1.636
1.487
1.726
1.612
1.664
1.901
1.493
1.360
1.569
1.490
1.533
1.720
1.491
1.357
1.544
1.493
1.506
1.723
1.746
1.550
1.512
1.716
1.474
1.653
FID
RRF
±0.05
1.30
1.02
0.93
1.09
1.15
1.04
0.86
1.0
0.67
0.68
1.25
0.63
0.54
0.76
0.72
1.20
0.34
0.61
0.87
0.63
0.74
1.07
0.77
0.41
0.81
0.54
0.71
0.71
1.33
0.52
0.81
0.76
0.64
1.09
0.66
0.88
SP-2340
RRT
±0.003
1.643
1.352
2.128
1.686
1.989
1.820
1.447
2.350
1.186
2.104
2.262
2.907
2.391
1.905
2.758
2.278
2.331
3.501
2.087
1.665
2.419
1.963
2.005
2.704
2.041
1.618
2.213
1.914
1.843
2.702
3.709
2.869
2.149
3.477
1.806
2.495
ECD
RRF
±0.05
0.91
0.79
0.73
1.23
1.02
0.68
0.74
0.63
3.87
4.21
7.25
__
3.05
4.10
3.74
—
1.84
3.36
4.12
3.40
4.10
5.86
3.98
2.31
2.94
2.58
3.20
3.66
4.40
2.79
4.04
4.17
3.07
7.27
3.45
4.63
1927A 3-30 06/19/86
-------
TABLE 3-6 (cont.)
Congener
Gas Chromatographlc Column and Detector Usedc
SP-2330
RRT
+0.003
ECD
RRF
+0.05
DB-5
RRT
+0.003
FID
RRF
+0.05
SP-2340
RRT
+0.003
ECD
RRF
+0.05
1,2,3
1,2,3
1,2,3
1,2,3
1,2,3
1.2,3
1,2,3
1,2,3
1,2,3
1,2,3
,4,6,7-
,4,6,8-
,4,6,9-
,4,7,8-
,4,7,9-
,4,8,9-
,6,7,8
,6,7,9-
,6,8,9-
,7,8,9
1,2,4,6,7,8-
1,2,4,6,7,9-
1,2,4,6,8,9-
1,3,4,6,7,8-
1,3,4,6,7,9-
2,3,4,6,7,8-
1,2,3,4,6,7,8-
1,2,3,4,6,7,9-
1,2,3,4,6,8,9-
1,2,3,4,7,8,9-
1,2,3,4,6,7,8,9-
.949
.320
.364
.800
.814
.251
.845
.039
3.376
4.016
1
2.
3.
2.
2.
4.
2,
3.
4,
4,
4.
5.
434
603
893
355
381
4.499
025
699
699
640
.36
,42
.44
.14
.20
.84
3.76
18
66
2.79
2.72
95
69
57
76
5.06
2,
2.
2.
2.
2.
2.
2.
2.
2.
3.
4.
316
066
426
320
378
771
345
442
488
2.748
2.101
2.174
2.217
2.099
2.140
2.500
3.333
3.440
507
002
7.809
5.708
0.18
0.57
0.45
0.54
0.44
0.34
0.34
0.63
0.51
0.33
0,22
0.59
0.53
0.60
0.34
0.46
2.61
0.61
1.35
1.32
0.43
3.
2.
4.
793
943
276
3.592
3.573
5.426
3.670
3.874
4.276
5.112
108
295
631
037
025
5.996
708
917
501
849
2.35
11.854
4.29
2.64
2.82
2.88
9.35
2.50
4.58
4.98
1.68
aSource: Bell and Gara, 1985
Detention times and response factors all given as ratios to those for
1,3,6,8-TCDF.
cColumns: SP-2330 fused silica 60 m x 0.25 mm ID at 240°C; DB-5 bonded
phase fused silica, 60 m x 0.25 mm ID at 240°C; SP-2340 bonded fused silica
50 m at 220°C. The ECD was a 63N1-electron capture detector. The FID
was a H2/a1r flame lonlzatlon detector. The nitrogen carrier flow rate
was 1-2 ml/mln.
1927A
3-31
06/19/86
-------
10 ml of n-hexane. PCDFs were recovered by elutlon with 6 ml of
methylene chloride; the eluate evaporated and was subjected again to cleanup
on a second Identical aluminum oxide mlcrocolumn. To achieve optimal
removal of PCBs, 10 ma of 4% methylene d1chlor1de/n-hexane was then used
as the first eluent; PCDFs, lower PCBs, chlorinated tMcyclIc aromatlcs were
then eluted with 10 ml of 50% methylene d1chlor1de/n-hexane. This final
eluate was concentrated and Injected Into the GC/MS. Recoveries of both
2,3,7,8-TCDF and TCDD were 90% at the 0.1-1 ppm level (Buser et al.,
l£78d). This 1s the basis of U.S. EPA method 613 except that 3% methylene
dlchlorlde/n-hexane and 20% methylene d1chlor1de/n-hexane are used for the
first and second eluents, respectively. RCRA Method 8280 also utilizes this
step. Either basic or neutral alumina can be used with consistent results
being obtained with a 3% methylene dlchlorlde/hexane wash (10 ml), a 20%
methylene dlchlorlde/hexane wash (15 ml) with elutlon of PCODs and PCDFs
with 50% methylene dlchlorlde/hexane (15 ml). If further cleanup Is
necessary a charcoal cleanup 1s recommended (Smith et al., 1984).
3.3.2.4.2. Yusho 011 -- Yusho oil (100 mg) was applied to a silica
mlcrocolumn (0.5 g silica gel, 70/230 mesh, Merck, In a 15 cm x 5 mm dispos-
able Pasteur plpet). The PCBs and PCDFs were eluted with 6 ml of n-hexane
with almost complete retention of rice oil on the column. The eluate was
concentrated and applied to an aluminum oxide mlcrocolumn (1.0 g basic
aluminum oxide, Woelm, In a disposable Pasteur plpet, 15 cm x 5 mm). Most
of the PCBs were eluted with 10 ml of 2% methylene d1chlor1de/n-hexane and
the PCDFs (plus other chlorinated trlcycllc aromatlcs and lower chlorinated
blphenyls) collected by elutlon with 10 ml of 50% methylene dlchlorlde 1n
n-hexane. The latter fraction was concentrated to 50 yi and 2 pi
Injected for GC/MS analysis. Recoveries of both 2,3,7,8-TCDF and -PCDD were
1927A 3-32 06/23/86
-------
90% at the 0.1-1 ppm level (Buser et al., 1978d). This procedure differs
slightly from the one quoted by Rappe et al. (1977) since the latter tech-
nique resulted In losses of some of the less polar PCDFs. This technique
has also been used by Cull and Dobbs (1984), Rappe et al. (1985b) and
Paaslvlrta et al. (1985).
PCB with soot or on wipes 1s generally treated with 1 M HC1 for 1 hour
by shaking, the slurry filtered by suction, washed with water, dried and
then Soxhlet extracted with toluene and further treated as 1n Section
3.3.2.4.4. (Rappe et al., 1983a).
3.3.2.4.3. Chlorophenol Formulations — A 4 g sample of the chloro-
phenol formulation was added to 30 ma. of methanol 1n a 250 ml separatory
funnel. Ten mil of 2.5 M lithium hydroxide was added and then 100 ml of
water. The neutral fraction was extracted with 40 mil of light petroleum
ether at pH 12. If the separation was difficult, 2-5 g of anhydrous sodium
sulfate was added to break the emulsion. The organic phase was washed with
100 ml of 0.05 M lithium hydroxide, and then with 50 mil of distilled
water, which should be neutral on separation. The organic phase was dried
over anhydrous sodium sulfate and a 20 ml aliquot concentrated to 2 ml
1n a stream of nitrogen. The procedure for PCB formulations was followed,
beginning at the second aluminum oxide column step, given above In Section
3.3.2.4.1. This method Is a combination of the published methods of Buser
and Bosshardt (1976) and Buser et al. (1978d). Recoveries of PCDFs were
90%. The method has been used also by Paaslvlrta et al. (1982) and Singh et
al. (1985).
3.3.2.4.4. Fly Ash Samples — Fly ash (25 g) was stirred 1n 1 M
hydrochloric add (200 ml). After centrlfugatlon the residual fly ash was
washed with delonlzed water 1n a Buchner funnel and dried at room tempera-
1927A 3-33 06/23/86
-------
ture. The dried fly ash was then extracted for 35 hours with toluene using
a Soxhlet apparatus. Approximately 98% was extracted In the first 24 hours,
and 1n the next 24 hours 1.2-1.6% more was recovered (Kooke et al., 1981).
This Is the crucial step since other solvents like methylene dlchlorlde,
acetone/hexane or chloroform are much less efficient. Also, use of HF
Instead of HC1 drastically lowers the extraction efficiency (Lustenhouwer et
al., 1980; Kooke et al., 1981). Wang et al. (1983) showed that 12N HC1 or
12N H_SO. Increases OCDD extraction by a factor of 2. The extract was
then concentrated to 1 ma and loaded onto a chromatographlc column con-
sisting of 4 g of silica loaded with 40% (weight) concentrated sulfurlc
acid. This column was connected to another, containing layers of 2 g of
sodium hydroxide treated silica gel (1 part by weight) of 1 M sodium hydrox-
ide solution on 2 parts (by weight of silica gel), 2 g of silver nitrate on
silica gel (10% by weight), and 1 g of silica gel. The columns were eluted
with redistilled n-hexane. The eluent was concentrated and then chroma-
tographed by the method described 1n Section 3.3.2.4.2. Recoveries of PCDDs
and PCDFs were reported to be 100% (Lustenhouwer et al., 1980; Kooke et al.,
1981). A quick method by Shushan et al. (1985) has not been adequately
evaluated yet, but HPLC cleanup caused a large PCDF loss.
PCDF recovery from EPA-5 type sampling trains 1s extremely variable
(Clement et al., 1985a; Wang et al., 1983; Tlernan et al., 1985); a multiple
rinse scheme Is necessary to achieve reasonable precision (Tlernan et al.,
1985). An Interlaboratory comparison 1s documented by Brocco et al. (1984),
the results sometimes vary by an order of magnitude.
3.3.2.4.5. Agent Orange and 2,4,5-T Ester Formulations — Ester for-
mulation (1 g) was added 1n n-hexane to a silica gel column (15 cm x 11 mm;
silica gel, Merck) and eluted with 30 ma of 5% methylene dlchlorlde In
1927A 3-34 06/23/86
-------
n-hexane. The eluate was concentrated 1n a stream of nitrogen. The concen-
trate was subjected to the technique given In Section 3.3.2.4.1. starting at
the second aluminum oxide column step. Recoveries of 20-50 yg of added
TPCODs and TCDFs were 90% (Rappe et al., 1978c).
3.3.2.4.6. Wood Dust — A published method by Levin and Nllsson
(1977) recommended that the wood dust be extracted with ether and the ether
concentrate subjected to TLC or silica gel, and the PCDFs and PCDDs eluted
with chloroform.
3.3.2.4.7. Gelatin Samples — A 15 g gelatin sample was weighed Into
a 100 ma round bottom flask. Ethanol (20 ma) and 40% aqueous potassium
hydroxide (40 ml) were added and the sample shaken for 1.5-2 hours at 180
strokes/minute. The solution was then extracted with hexane (1x20 ma;
3x18 ma) In a separatory funnel. The halogenated hexane extract was then
extracted first with 1 M potassium hydroxide (40 ma) and then with 40 ma
portions of concentrated sulfurlc add until the acid phase was colorless.
Heavy emulsions at this last step were broken by swirling the separating
funnel. Moderate emulsions were successfully broken by addition of a few
crystals of anhydrous sodium carbonate. The washed hexane extract was fur-
ther washed with water (10 ma) and saturated aqueous sodium carbonate (10
ma) and filtered through 5 cm (15 g) of anhydrous sodium carbonate In a 19
mm ID x 30 cm chromatographlc tube. The hexane eluent was concentrated to 3
ma on a steam bath, the containing flask being filtered with a modified
mlcro-Snyder column and also containing three 20-mesh carborundum boiling
chips. Although Firestone (1977b) then utilized a slightly different method
to achieve separation of PCDDs and PCDFs from other chlorinated compounds to
obtain >90% recoveries of PCDDs (tetra- to hepta-) and 2,3,7,8- TCDF, the
more modern procedure described 1n Section 3.3.2.4.4. should be followed
before GC/MS.
1927A 3-35 06/23/86
-------
3.3.2.4.8. Tissues — Four methods are current: a Japanese method
(Nagayama et al., 1977), one advocated by Albro and Corbett (1977), one by
Stalling et al. (1981), and others compared by Albro et al., (1985).
The Japanese method, which was also used by Kurokl and Masuda (1977,
1978) and Rappe et al. (1979b), Involves tissue homogenlzatlon 1n n-hexane
and sodium sulfate In a Waring blender followed by saponlfIcatlon with 1 N
potassium hydroxide In ethanol. The alkaline solution was extracted with
n-hexane, concentrated and then chromatographed on a silica gel column (2 g)
elutlng with n-hexane. The concentrate was further fractionated on a column
of aluminum oxide (3 g) using first n-hexane/carbon tetrachloMde (4:1 v/v)
and finally 25% methylene d1chlor1de/n-hexane to elute the PCDFs and PCDDs.
The final eluate was evaporated, so It could be Injected Into the GC. Albro
and Corbett (1977) and Albro (1979) showed that there were extensive losses
of OCDD and OCDF (up to 60%) at room temperature and 90% under reflux, dur-
ing the potassium hydroxide saponlfIcatlon step. These losses are Important
1f all PCDFs are to be quantltated or 1f these compounds take part In syner-
glstlc/antagonlstlc toxic reactions. The recovery of 2,3,7,8- TPCDD was
88-90% at room temperature.
Albro and Corbett (1977) described methods to extract PCDFs from liver,
blood, urine, milk, serum and adipose tissues. Liver samples were blended
1n 200 ml of chloroform/methanol (2:1 v/v); the solution was filtered
under suction through a glass fiber filter paper; and the filter paper and
cake was then blended with 100 ml of chloroform/methanol (2:1 v/v). The
solution was again filtered as above and the filtrates combined. A solution
of 1.2% aqueous potassium chloride (52 ml) was added and mixed. The
phases were allowed to clear, the lower phase collected, and evaporated just
to dryness at 40°C by rotary evaporation under reduced pressure. Blood,
1927A 3-36 06/23/86
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urine, milk and serum (100 ml) were extracted as described by Kates (1972)
and the resulting chloroform phase was concentrated by rotary evaporation.
Adipose tissues were ground In a mortar and pestle with 8 times their weight
of anhydrous sodium sulfate. The powder was extracted with chloroform In a
Soxhlet, allowing at least eight cycles. The chloroform extracts were con-
centrated by rotary evaporation. All the chloroform concentrates obtained
above were then further treated by the same cleanup procedure. The llpld
residue from 10 g of liver, 100 ml of biological fluid or 1 g of adipose
tissue was leached Into 15 ml of carbon tetrachlorlde. Concentrated sul-
fuMc add (15 ml) was added, the mixture shaken and centrlfuged at 2000
rpm for 30 minutes. The organic layer was removed, the volume noted and the
layer dried by passing H through anhydrous sodium carbonate In a glasswool
plugged funnel. The sodium carbonate was then rinsed with fresh CC1.
(2 ml). The filtrate was concentrated just to dryness by rotary evapora-
tion at 40°C. The residue was leached with 1.5 ml of n-hexane/methylene
dlchlorlde (97:3 v/v). The leachate was loaded onto a dry-packed column of
Fisher A-540 aluminum oxide (3 g) of diameter 0.5-0.7 cm. The flask was
rinsed with 1 ml of n-hexane:methylene dlchlorlde (97:3 v/v) and added to
the column. Fraction I was eluted with 28 ml of n-hexane:methylene
dlchlorlde (97:3 v/v). Fraction II was eluted with 30 ml of n-hexane:
methylene dlchlorlde (4:1 v/v). Fraction II was evaporated to near dry-
ness. An aliquot was then taken for GC/MS analysis.
The recoveries of dlbenzofuran, PCOFs and PCDDs, were reported to be 95%
(using the method of standard additions). The sulfuMc acid step was suffi-
cient to handle 150 mg of trlglycerlde/mi of acid, but only about 50 mg of
total liver Upld/ml of add. Emulsions were routinely broken by centrl-
fugatlon. Fisher A-540 aluminum oxide best retained I1p1ds 1n the solvent
1927A 3-37 06/23/86
-------
system described; other aluminum oxide types allowed partial elutlon of
sterol esters, waxy pigments and various trlglycerldes. The final concen-
trate was suitable for analysis at the ppt level by GC/MS.
The methods for PCDF quantltatlon In adipose tissues recommended by
Stalling et al. (1981) [see also Rappe et al. (1981)] Involved Initial pro-
cedures similar to those described by Albro and Corbett (1977). Thus, adi-
pose tissues were blended with 4 times their weight of anhydrous sodium sul-
fate and the column extracted with 100 mft volumes of methylene dlchlorlde
for every 10 g of tissue. After removal of solvent by rotary evaporation,
the weight of the oil residue was noted. This residue was dissolved 1n 1:1
(v/v) cyclohexane/methylene dlchlorlde to achieve an approximate concentra-
tion of 0.2 g/ml. AHquots (5 ma) were applied to a gel permeation col-
umn containing 69 g SX-3 BloBeads resin (2.5 cm ID x 48 cm) and eluted at 5
mi/mln. The 165-300 ma. eluent fraction of each analysis was collected,
amalgamated and then reduced to near dryness by rotary evaporation. The
extract was then passed through a column of potassium hydroxide-treated
silica gel topped with a small amount of cesium hydroxide-treated gel layer-
ed on top of a small band of sulfuMc acid-Impregnated silica gel. The
eluent was then run Immediately through a column containing carbon (Amoco
PX-121; <40 yiti) (Rappe et al., 1981) and dispersed on glass fibers (Stal-
ling, et al. 1981). Planar molecules (such as, toxic PCDDS and PCDFS) were
collected by reverse elutlon with toluene (30 ml), whereas nonplanar mole-
cules could be eluted with 1:1 (v/v) benzene/ethyl acetate (30 mj.) In the
usual manner.
PCOOs and PCDFs were then specifically collected by the technique des-
cribed In Section 3.3.2.4.2. before GC/MS. The reported recoveries were
1927A 3-38 06/23/86
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>70%. A slightly revised method was used by Nygren et al., (1986) for human
adipose tissue, mother's milk, bovine fat and bovine milk. The recovery of
13C,?-labeled surrogates 1s normally found to be >60%.
Albro et al. (1985) evaluated 6 different approaches 1n an Interlabora-
tory study for 3 PCDFs (2,3,7,8-TCDF; 2,3,4,7,8-PeCDF; 1,2,3,7,8,9-HxCDF)
added to human adipose tissue extracts at levels of 5 to 50 ppt. The adi-
pose tissue (600g) was combined with 4.8 kg of pesticide grade anhydrous
sodium sulfate In a clean food-grinder. After 3 processings, the powder was
packed Into two 20 In. x 4 1n. diameter pyrex columns. The packing was
eluted with 3ft. of ethyl acetate containing 0.001% butylated hydroxy-
toluene, and then 68. of methylene dlchlorlde. The extracts were combined,
concentrated 1n vacuo at 37°C, and the Upld diluted with chloroform to 900
mft. to stop solidification. The yield was equivalent to soxhlet extraction
with chloroform for 16 hours. The PCDFs 1n toluene were added and the adi-
pose tissue solution (60g adipose tissue equivalent) stirred magnetically.
The chloroform was reduced (water aspirator) In a dlsslcator over 24 hours.
Six extraction methods were then evaluated.
(I) 13C-2,3,7,8-TCDF and 37Cl-2,3,7,8-TCDF were added as Internal
standards. The sample was passed In cyclohexane/CHLCl,, through two
potassium silicate/silica gel columns and then through carbon adsorbent
(Stalling et al., 1981). The extracts were concentrated at 55°C 1n vacuo
and then passed through a H?SO./s1l1ca gel column In o-xylene. The
Interference of chlorinated naphthalenes can be further removed by a final
alumina chromatography step. This method 1s derived from Smith et al.
(1984).
(II) 13C-2,3,7,8-TCDF was added as Internal standard and the Upld
solution partitioned between hexane/concentrated FLSO. for 13-54 hours.
1927A 3-39 06/23/86
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The organic phase was washed through silica, NaOH on silica, silica,
HpSO. on silica, and silica, with 5% benzene 1n hexane. After concen-
tration, the residue was dissolved 1n hexane. This solution was then passed
through AgNO» on silica and trapped on basic alumina. After washing with
CCl^/hexane (4:1), PCDF/PCDDs were eluted with CH^/hexane (3:1),
the eluate evaporated under nitrogen and redlssolved 1n chloroform. After
reverse phase HPLC, the appropriate collected fractions were further purl-
fled by normal phase HPLC. This method 1s derived from Lamparskl and
Nestrlck (1980).
(Ill) The 37C1-TCDF, PCOF, and HxCDF were added as Internal stan-
dards. The samples were digested 1n concentrated HC1 for 1 hour, extracted
twice with hexane and dried over anhydrous sodium sulfate. The solution was
washed through a H SO./slUca gel column with hexane; the eluate was
washed with water, dried and concentrated. The concentrate was bonded onto
grade I alumina and washed with 5% CH2Cl2/hexane; the PCDFs/PCODs were
eluted with CHpClp/hexane (1:1), concentrated to 1 ma, and then loaded
on Carbopack C/Cellte treated as per Stalling et al. (1981). The extracts
were concentrated to 10 yL with dodecane entralner. This was an original
method.
(IV) a7Cl-2,3,7,8-TCOF was added as Internal standard. The solution
was then partitioned between hexane/H-SO. until the H0SO. was color-
24 24
less. The hexane phase was washed with ^% aqueous NaOH and water, dried
over NapSO./hexane, and eluted with CHpClp. The eluate was dried
under nitrogen, and the residue redlssolved In toluene or Isooctane. This
1s a method derived by Ryan et al. (1985a).
(V) 37Cl-2,3,7,8-TCOF was added as an Internal standard. The solution
was diluted and partitioned against H2S04 and neutralized through
1927A 3-40 06/23/86
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Ha2C03 and KOH/s1l1ca. After concentration, the PCDFs were exchanged
Into hexane from cyclohexane, loaded onto acidic alumina, washed with 3%
CH2Cl2/hexane, and eluted with CH2Cl2/hexane (1:1) onto charcoal/
cellte. This was then washed with 10% benzene/hexane and back-eluted with
5054 xylene/hexane onto neutral alumina, washed with 3% CHpCU/hexane,
eluted with CH2Cl2/hexane- (1:1) and then concentrated. This 1s an
original method.
(VI) iaC-2,3,7,8-TCDF was added as Internal standard. After dilution
with CH2Cl2/cyclohexane, the solution was passed through slUca/potas-
slum s1l1cate/Na2SO./potass1um sH1cate/s1l1ca/carbon column combina-
tions. The last column was washed with cyclohexane CH2C12 and
CH2Cl2/methanol/benzene (15:4:1), and back-eluted with toluene. The
extracts were concentrated at 35°C, passed through potassium silicate/
H2$04 on silica with hexane onto acidic alumina, washed with hexane and
2% CH2Cl2/hexane. PCDFs/PCDDs were eluted with CHC1 /cyclohexane
(1:1) and concentrated. This method was derived by Rappe et al. (1984).
The molecular Ions of all 13C/35C1 and 12C/37C1 PCDFs were monitored
except 1n Method IV where the (M -COC1)* 1on was monitored by MS/MS. The
Interferences from chlorinated dlphenyl ethers, PCBs, chlorinated methoxy
blphenyls, p.p'-DDE and o,p'-DDE were accounted for. Various GC columns
were used with DB-5 and SP-23SO fused silica capillary columns were most
utilized. Total analysis/report writing time varied between 48 and 140
days, though each method took between 6 and 26 working days.
The Isotoplc Internal standard method was definitely superior to the
external standard method (Table 3-7); the absolute quantHatlon 1s extremely
variable. Only methods II, III and VI had relative standard deviations for
unsplked samples of lower than 25%. As Table 3-7 shows, method II was most
1927A 3-41 06/23/86
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TABLE 3-7
Error In Estimation of PCDFs by Six Methods3
% Relative Error Compared With "Correct" Level In Method
Compound
I II III IV V VI
2,3,7,8-TCDDD -50 +10
a,3,7,8-TCDFc -(1+3) -(lj
2,3,4,7,8-PeCDFd +(13+6) *
l,2,3,7,8,9-HxCDFe -73 -18
leaked -90 -73 +70
+(2±3) (0.5+11) +(10+8) +(3.5+2.1)
+(12+7) -(3.5+3.5) +(27+16) +(24±34)
BO -100 NM +9
aSource: Albro et al. (1985)
b!0 ppt: (1 sample)
cll ppt: (average of 2 samples)
d!6 ppt: (average of 2 samples)
ell ppt: (1 sample)
BD = Below detection
NM = Not measured
1927A
3-42
06/23/86
-------
accurate (within 20% of the correct answer) for spiked PCDF and 2,3,7,8-TCDD
at -10-16 ppt. This method took 26 working days for sample processing.
There appeared to be no correlation between recoveries of PCDFs and PCDDs.
The results of this blind study Included certain commonly encountered
environmental contaminants that can Interfere with the quantltatlon of PCDDs
and PCDFs. Six laboratories using 6 different methods (all entailing GC-MS
for quantltatlon of PCDFs) participated In the study. Comparison of the
results Indicated the data obtained were qualitatively reliable; however
quantltatlon was problematical for some laboratories.
Another published report (Schecter et al., 1985a) describes the results
of analyzing blood and adipose tissues from patients exposed to residues
from a PCB-contalnIng transformer fire. The method of analysis Implemented
In the latter study entailed GC-MS quantltatlon of not only 2,3,7,8-TCDD and
2,3,7,8-TCDF but the sum of the congeners constituting each chlorinated
class of PCDDs and PCDFs, Including the tetrachlorlnated through
octachlorlnated congeners. The results of this study Indicate that the
tissue from patients with no known exposure to PCDDs or PCDFs had, In
several cases, several hun- dred parts-per-trllHon levels of higher
chlorinated PCDDs and PCDFs. Other publications by Schecter et al.
(.1986a,b) and Ryan et al. (1986) have Included the above approach.
3.3.2.4.9. Water — There Is currently no specific method for the
analysis of PCDFs 1n water. The procedure used for urine In Section
3.3.2.4.8. but scaled up to accommodate 1 8. of water In the Initial
extraction stage should be sufficient. A simpler method would be to extract
1 I (or a larger volume) of acidified water (pH 2) with methylene
dlchlorlde and then concentrate the extracts. The Yusho oil method (Section
3.3.2.4.2.) could then be applied. Obviously, further research Is required
1n this area.
1927A 3-43 06/23/86
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3.3.2.4.10. Sediment — Sediment samples (10 g) have been analyzed by
mixing and blending with 20 g anhydrous sodium sulfate, followed by Soxhlet
extraction for 48 hours with 100 ml of toluene with subsequent concentra-
tion of the extract, before cleanup and GC/HS analysis (Petty et al.,
1983a,b). The extraction efficiency 1s >74%. An article describing analyti-
cal methodology and the application of this method to the assessment of
environmental contamination by PCDOs and PCDFs as a result of Improper dis-
posal of chemical wastes has been published (Tlernan et al., 1985). The
analytical methodology employed entailed the use of GC-MS to quantHate
parts-per-tr1H1on to parts-per-b1H1on levels of PCDFs as well as PCDDs In
municipal refuse-fired boiler effluents and effluents from an RDF-flred
boiler as well as soot from a PCB-contalnIng transformer fire.
3.4. GC/HS METHODS FOR PBDFs
GC methods for the PBDFs are not as plentiful as those for PCDFs.
O'Keefe (1978b) has published a method for separating PBDFs from PBBs. The
PBB (50 mg) was dissolved In 2 ml of benzene and the solution was filtered
through a sodium carbonate column (40 m x 6 mm) 1n a disposable Pasteur
pipette, followed by two 5 ml benzene washings. The benzene eluate was
concentrated to 1 ml, 10 ml of cyclohexane added, and the solution evap-
orated to 1 ml. This replacement procedure with cyclohexane was repeated
twice. Methylene dlchlorlde (15 ml) was then added, and the solution
loaded onto a column of graphltlzed charcoal (Carbopack AHT; Supelco, Inc;
40 m x 6 mm).
Nonplanar aromatlcs were eluted with 50 ml benzene/dlethyl ether (4:6)
followed by pyrldlne (70 ml) to elute the planar aromatlcs. The pyrldlne
fraction was reacted with 140 ml of 1% hydrochloric add and then extract-
ed with hexane (3x30 ml). The hexane fractions were combined, washed with
1927A 3-44 06/23/86
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100 ml water, dried (sodium sulfate) and then concentrated. The concen-
trate was placed onto an activated aluminum oxide column (Woelm Neutral;
40x6 mm) 1n a disposable Pasteur plpet. The first eluate, 10 ml CC14>
contained PBBs, and the PBDFs were eluted with 7 ml methylene dlchlorlde.
GC/MS was performed on this eluate. O'Keefe (1978b) used a 6 ft x 1/8 Inch
Pyrex column, packed with 3% OV-1 on 100/200 mesh Gas Chrom Q temperature
programmed from 260-290°C at 6°C/m1n to separate any PBDFs before analysis
by MS. The molecular weight of the TBDF was 479.6996, and It was the 10054
peak (M). Other peaks were M-2Br (30%), M-COBr-2Br (20%), H-COBr (6%), M-Br
(654), Mf* (354) and (M-2Br)2f (weak). PeBDFs showed peaks at the molecu-
lar 1on 561.606. The other major Ions were M-COBr, M-Br, M-2Br and
M-COBr-Br Ions. This procedure allowed 5054 recovery of the 2,3,7,8-TBDF and
was applied to analyze pyrolyzed PBB.
No PBDFs (<0.5 ppm) were found In Flremaster BP-6 by Mass et al. (1978)
using a technique Involving Florlsll fractlonatlon of a 10 g sample dis-
solved In 150 ml of 354 methylene dlchlorlde 1n hexane eluted first with
500 ml of 354 methylene dlchlorlde/hexane and then with 500 ml of 5054
methylene dlchlorlde/hexane. The latter fraction contained 0.5 ppm PBDFs on
concentration and analysis by GC/MS, using a 1 m x 2 mm ID stainless steel
column packed with 354 OV-101 on 100/120 mesh Gas Chrom Q at 300°C. No
recoveries of PBDF standards were cited and analytical conditions were not
optimal with the use of the stainless steel GC column and the high tempera-
ture. A methyl-BDF was subsequently found 1n the polar fraction of a Fire-
master of unspecified type (Moore, 1977).
Buser (1986) used a 25 m x 0.31 mm ID SE-54 glass capillary column
(splHless) 1n conjunction with temperature programming. As expected, the
1927A 3-45 06/23/86
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retention times are longer and the elutlon temperatures are higher than the
corresponding PCDF analogs. The electron Impact mass spectra of TBDF and
PeBDFs agreed with those found by O'Keefe (1978b).
3.5. OTHER METHODS FOR THE PCDFs
It 1s clear from Section 3.3. that no one technique will suffice to
separate, quantify and confirm the presence of the PHDFs. Sometimes It Is
necessary to use all the available methods.
Hutzlnger et al. (1973) found that TLC on Merck Silica Gel F-254 of 0.25
mm thickness allowed 2,8-DCDF and OCDF to be separated 1n n-hexane solvent
with Rf values of 0.55 and 0.75, respectively.
A gel permeation technique for the cleanup of PCDDs and PCDFs was pro-
posed by Stalling et al. (1975). The system used BloBeads S-X_ with ethyl
o
acetate and ethyl acetate/toluene as solvents. This system has been alluded
to In Section 3.3.2.4.8. PCDFs, PCDDs and chloronaphthalenes were retained
on a charcoal column subsequent to gel permeation chromatography and were
eluted with ethyl acetate/toluene.
Perchlor1nat1on techniques have been used to obtain total PCDF content
(Masuda et al., 1976), but for reasons outlined previously are not useful In
Identifying Individual toxic Isomers. Ballschmlter et al. (1985) have used
BMC reagent to achieve perchlorlnatlon. Perbrom1nat1on Is not possible
because of steMc hindrance (Rlchtzenhaln and Schrage, 1978).
The Flremaster BP-6 metabolite, 6-hydroxy-2,2',4,4',5,5'-hexabromob1-
phenyl decomposes on OV-101 GC columns at 230-260°C to form two PeBDFs
(Gardner et al., 1979). This demonstrates that PBDF detection alone does
not prove PBDFs were originally present In the sample. If phenolic metabo-
lites are separated, there Is no lack of specificity. There 1s also the
need for a good HPLC method along the lines of the separation of PCDD
1927A 3-46 06/24/86
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Isomers (Nestrlck et al., 1979). The latter step should complement the GC
step of GC/MS. Substantial HPLC losses especially for lower PCDFs have been
reported (Shushan et al., 1985). The losses vary from 2- to 5-fold.
Clearly this warrants Investigation. Mazer et al. (1983a,b) have success-
fully used a reverse phase column (C,0) with 75% acetonltrlle/25% water at
lo
0.75 mfc/mln with detection at 235 nm. They also utilized a normal phase
Zorbax NH~ column with hexane as the eluent at 1.5 mi/mln. Both columns
were used In cleanup steps to produce pure Isomers rather than to provide
quantitative recovery. Cull and Dobbs (1984) have also used this tech-
nique. The quality control and assurance required for HPLC analysis of
PCOOs and PCOFs has been presented by Donnelly et al. (1986) 1n connection
with RCRA Method 8280.
A promising radlolmmunoassay for 2,3,7,8-tetra-PCDF 1n PCBs and 1n
environmental samples has been recently developed (Luster et al., 1980) as a
screening technique.
Biological monitoring by Induction of arylhydrocarbon hydroxylase activ-
ity (Poland et al., 1979; Bradlaw and Casterllne, 1979) 1s considered 1n
Chapter 7. Hutzlnger et al. (1981) and Wang et al. (1983) showed that the
toxldty of fly ash samples 1n the cytosol receptor assay could not solely
be explained In terms of 2,3,7,8-TCDD (72 pg 2,3,7,8-TCDD equivalents was
the ED _). The Induction and cytosol receptor assay tests often disagreed
(Sawyer et al., 1983). See Chapter 7 for a discussion of these tests.
Helder et al. (1982) and Helder and Selnen (1985) have used a biological
assay with rainbow trout yolk sac fry to show that the toxldty of fly ash
cannot be solely attributed to the 2,3,7,8-TCDD level but to the cumulative
presence of other PCDDs and PCDFs. The fry suffered hemorrhages, edema, fin
1927A 3-47 06/23/86
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necrosis and finally death at days 4-12. 2,3,7,8-TCDF also caused this
behavior. The raw fly ash was as toxic as the cleaned-up PCDDs/PCDFs from
the same fly ash sample reconstituted 1n the same volume.
A bloassay based on humoral .antibody production has been utilized by
Rlzzardlnl et al. (1983). 2,3,7,8-TCDF at 10 ng/g caused an Immunosuppres-
slon of 25%. Administration of both 2,3,7,8-TCDD and 2,3,7,8-TCDF caused
much less Immunosuppresslon than the TCDD alone.
1927A 3-48 06/23/86
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4. SOURCES TO THE ENVIRONMENT
4.1. SUMMARY
PCDFs occur as contaminants of a number of chemical products such as
phenoxy herbicides, polychloMnated phenols, hexachlorobenzene, polychloM-
nated blphenyls and chlorodlphenyl ethers. PCDFs are also associated with
thermal Insulation material's. Thus, both PCDFs and PBDFs are adventitious
copollutants released Into the environment together with economic poisons or
Industrial wastes. There Is experimental evidence that PCDFs and PBDFs may
be formed 1n the environment by photolytlc and pyrolytlc reactions from
chemical products and combustion processes. Many of the products contain a
larger percentage of PCDDs than PCDFs. However, the PCDFs 1n these products
Include more of the tetra- and penta- (the most acutely toxic) compounds
than do the PCDDs. The levels of PCDFs containing 4 and 5 halogen atoms are
generally <20 ppm for most chlorophenols, <0.40 ppm 1n most phenoxy herbi-
cides, <5 ppm 1n PCBs and <0.5 ppm 1n the PBB, Flremaster BP-6. The lower
chlorinated phenols and PCBs are most highly contaminated with toxic PCDFs.
The levels of the 2,3,7,8-TCDF and 2,3,4,7,8-PeCDF, the most toxic Isomers,
are generally <1 ppm In most PCBs.
PHDFs are formed by the pyrolysls of various materials. They have been
detected 1n Incinerator fly ash (<1.5 ppm), 1n pyrolyzed PCB (up to 104
ppm), 1n PCB that had been used for 2 years as heat exchanger fluid (<16
ppm), In pyrolyzed PBB (up to 10* ppm), In pyrolyzed (2-butoxyethanol)
ester of 2,4,5-T (<20 ppm), 1n some pyrolyzed commercial chlorophenols and
1n pyrolyzed chlorinated dlphenyl ethers.
4.2. SOURCES OF PCDFs FROM CHEMICALS
The early work on the detection of PCDFs was hampered by losses during
sample cleanup and analysis and by the fact that many PCDF gas chromato-
graphlc peaks were frequently superimposed on those of chlorinated dlphenyl
1928A 4-1 06/23/86
-------
ethers (Crummett and Stehl, 1973). The Interfering chlorinated dlphenyl
ether usually contained two chlorine atoms more than the PCDF. In spite of
these difficulties, OCDF and a HpCDF were detected 1n dipping wastes
containing pentachlorophenolate used as a wood preservative attributable to
side-reactions 1n manufacture (Jensen and Renberg, 1973).
PCOFs are therefore not commercially produced but are formed as trace
unwanted Impurities 1n the manufacture of other chemicals such as chlori-
nated phenols and their derivatives, chlorinated dlphenyl ethers, hexa-
chlorobenzene and polychlorlnated blphenyls, which are released Into the
environment either as economic poisons or as Industrial wastes (Table 4-1).
An estimate of the extent of release Into the Canadian environment of
known sources of PCDFs Is provided 1n Table 4-2 (Sheffield, 1985a,b).
4.2.1. Chlorinated Phenols. Chlorophenols are Industrially produced by
two methods, namely by the direct chlorlnatlon of phenol (Hutzlnger et al.,
1985b) and by alkaline hydrolysis of chlorobenzene 1n different solvents
(Ahlborg and Ihunberg, 1980). PCDFs appear as contaminants 1n both pro-
cesses with more formed In the latter process. The annual world production
In 1979 was -150,000 tons (Rappe et al., 1979a).
Chlorinated phenols are used as fungicides, herbicides, s!1m1c1des, wood
preservatives and bacterlddes; they are also used In the synthesis of
chlorinated phenoxy herbicides.
Table 4-3 provides the composition of two typical commercial PCPs.
Table 4-4 summarizes some of the PCDF and PCDD levels found 1n various
chlorinated phenols. As expected, the higher PCDDs and PCDFs predominated
with the PCDDs usually more abundant. However, the TCDFs and PeCDFs 1n all
chlorophenols were greater than the content of the corresponding PCDD
congeners. Since the HpCDDs, HpCDFs, OCDDs and OCDFs have lower toxlclty
1928A 4-2 06/23/86
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TABLE 4-1
Presence of PCDFs 1n Industrial Chemicals
and Their Entry Into the Environment
Industrial Products
Containing PCDFs
Use
PCDFs (total concentrations
1n Industrial products)*
Concentration
1n mg/kg
(m1n/max)
Reference
Phenoxy alkanolc
compounds
herbicides
Chlorinated phenols wood preservative
Hexachlorobenzene
Polychlorlnated
blphenyls
Chlorides of Iron,
aluminum and copper
fungicide and
Industrial waste
Industrial use
Industrial use
0.008-0.15
59.8-790
0.35-58.3
0.8-13.6
Rappe et al.,
1978b, 1979a;
AhUng et al.,
1977
Rappe et al.,
1979
Vlllaneuva
et al., 1974
CNRC, 1978b
0.0003-0.060 Helndl and
Hutzlnger, 1986
*0nly positive data are considered
1928A
4-3
06/23/86
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TABLE 4-2
Estimated PCDD and PCDF Release to the Canadian Environment
from Chemical Sources*
Release (q/year)
Source
PCDD
PCDF
Chlorophenol production
Air emission
Wastewater
900
13
600
13
Processes using pentachlorophenol
Wood preservation
Wastewater
Waste disposal
Leather tanning
Sludge
1000
>500
negligible
600
>300
Chemical products
2,4-D formulations
2,4,5-T
Pentachlorophenol
PCBs
100
5
[l.lxlO6]
[0.6x10*]
[750,000]
*Source: Sheffield, 1985a,b
1928A
4-4
06/23/86
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TABLE 4-3
Chemical Analysis of a Typical Pure and
Technical Pentachlorophenol3
Source
Pure
(AldMch, lot 120,717}
Technical
(Monsanto, lot KA578)
Phenols'5
Pentachlorophenol
Tetrachlorophenol
Nonphenollcs (ppm)D
ppm
84.654
3%
D1benzo-p-d1ox1ns
Tetrachloro- <0.1 ppm
Pentachloro- <0.1
Hexachloro- <0.1
Heptachloro- <0.1
Octachloro- <0.1
Dlbenzofurans
Tetrachloro- <0.1
Pentachloro- <0.1
Hexachloro- <0.1
Heptachloro- <0.1
Octachloro- <0.1
<0.1 ppm
<0.1
8
520
1380
<4
40
90
400
260
aSource: Goldstein et al., 1977a
^Samples were analyzed by GC/MS. The lower detection limit was 0.1 ppm.
1928A
4-5
06/20/86
-------
TABLE 4-4
CD
3>
Levels of PCDFs and PCDDs In Some Commercial Chlorophenols
Levels of PCDFs and PCDDs (mq/kq)
Chloropheno"
Penta-
Penta-
Penta-
Penta-
er* Penta-
Penta-
Penta-
Penta-
Penta-
Penta-
c Penta-
c Penta-
cc
cr
1 Formulation Tetra- Penta-
Type
CDF CDD CDF CDD
(Monsanto) <4 <0.1 40 <0.1
(Dow) 0.20 <0.02 0.20 <0.02
(Dowlclde 7)
(Dowlclde 7) <0.2 <0.2 <0.2 <0.2
(Dowlclde EC-7) -
(Dowlclde EC-7) 0.45 - 0.03
(Dowlclde EC-7) <0.02 - <0.03
(Dow) 0.20 - 0.20
(Dow) 0.07 - 0.20
(Monsanto) -
(Monsanto) -
(European) 0.9 ? 4 ?
Hexa- Hepta-
CDF CDD CDF CDD
90 8 400 520
13 10 70 130
30 4 80 125
39 9 280 235
<1 1 1.8 6.5
0.30 0.15 0.50 1.1
<0.03 0.03 0.5 1.1
13 10 70 130
9 5.4 60 130
19 11 81 199
90 8 400 520
32 ? 120 ?
Octa-
CDF
260
55
80
230
<1
0.5
0.2
55
65
137
260
130
CDD
1380
210
2500
250
15
5.5
5.5
210
370
1170
1380
?
Total Reference
CDF CDD
794 1908 Goldstein et
al., 1977a
138 350 Rappe et al.,
1979a
190 2629 Firestone.
1977a
549 494 Buser, 1975
3.8 22.5 Firestone,
1977a
1 .8 6.8 Buser and
Bosshardt,
1976
<0.08 6.6 Buser and
Bosshardt,
1976
135 350 Buser and
Bosshardt,
1976
134 3.5 Firestone,
1977a
237 1300 Firestone.
1977a
750 1980 Firestone,
1977a
280 1000 Rappe et al .,
1978a. b
-------
ro
CO
3*
^
1
*"J
O
a*
NJ
to
Chlorophenol
Penta-
Penta-
Penta-
Penta-
Penta-
Penta-
Penta-
Penta-
Tetra-
(2.3,4,6-)
Tetra-
(2.3,4.6-)
Tetra-
Trl-
Formulatlon
Type
(Fluka)
(Fluka)
Na Salt
(Fluka)
Na Salt
(Dow)
Na Salt
(Fluka)
K12
K,
8 samples
(Dowlclde 6)
(Finnish)
Kymnene
Ky-5
Ky-5
(Swedish)
1 nULL f~f I IUM L . J
Levels of PCDFs and PCDDs (raq/kq)
Tetra- Penta- Hexa- Hepta- Octa- Total
CDF CDO CDF CDD CDF CDD CDF CDD CDF CDD CDF CDD
<0.21 - 0.05 ? 15 9.5 95 125 105 160 220 295
0.05 - 0.25 - 36 9.1 320 180 210 280 566 469
<0.02 0.12 0.03 0.03 0.7 <0.03 1.3 0.3 2.1 1.5 4.23 2.0
<0.02 - 0.005 - 11 3.4 50 40 24 115 85 158
<0.02 0.05 0.04 <0.03 11 3.4 47 38 26 110 84 151
ND ND 5.9 92 3.0 1921 29 204 11
ND ND ND ND 98 3.5 39 1.1 1.5 -
4.5 62 35 130 610
e
<0.2 <0.2 <0.2 <0.2 30 6 500 55 135 39 865 100
<0.5 <0.7 10 5.2 70 9.5 70 5.6 10 0.7 160 22
2.0 2.1 2.8 - 21 - 30 0.6 4.0 0.8
1.5 <0.02 18 <0.03 36 <0.03 4.8 <0.1 - <0.1 60 <0.3
Reference
Buser and
Bosshardt,
1976
Buser and
Bosshardt,
1976
Rappe et al . ,
1978a,b
Rappe et al . ,
1978a.b
Buser, 1976a
Paaslverta
et al., 1982
Paaslverta
et al., 1982
Cull and
Oobbs, 1984
Buser, 1975
Rappe et al . ,
1978a.b
Paaslverta
et al., 1982
Rappe et al . ,
1977
CO
-------
(McConnell et al., 1978b; Poland and Glover, 1977), 1t may seem more appro-
priate to concentrate on the TCDFs to HxCDFs. This can be misleading, how-
ever, since 2,3,7,8-TCDD 1s 1000-10,000 times more toxic than 1,2,3,8-TCDD
{McConnell et al., 1978b; Poland and Glover, 1977). Nevertheless, the prob-
ability of encountering a toxic compound may be much greater for PCDFs than
for a PCDD since there are more PCOFs containing 4 or 5 chlorine atoms 1n
the vicinal positions, where toxldty 1s conferred. Firestone (1977a)
reported that the major PCDFs In Dowldde 7 and EC-7 formulations were the
2,3,6,7- and 2,4,6,7-tetra-; the 2,3,4,6,7-, 1,2,4,7,8-, 2,3,4,7,8-penta-;
and the 1,2,3,6,7,8-hexa-lsomers. Rappe et al. (1978a,b) found the major
components 1n an American PCP to be those given 1n Table 4-5. The Isomers
present were very different from those reported by Firestone (1977a). As
shown In Table 4-5, the European trlchlorophenol formulation contained
traces of the toxic 2,3,7,8-TCDF and 2,3,4,7,8-PeCDF.
In the United States, PCP 1s manufactured by the chlorlnatlon of phenol
(Firestone, 1977a). In Europe, both this process and the alkaline hydroly-
sis of hexachlorobenzene (135-275°C) have been used. The product from
chlorlnatlon of phenol tends to have comparatively nontoxlc, higher chlori-
nated Isomers. This process Is conducted at atmospheric pressure using two
reactors. In the first, phenol 1s chlorinated at 65-130°C until 3-4
chlorines have been added and until the mp of the product has reached 95°C.
The temperature Is then adjusted slowly Into the 195-205°C range, a process
typically complete 1n 5-15 hours. Aluminum chloride Is often added to
accelerate the final chlorlnatlon. Typically, the final product consists of
80-88% PCP and 12-20% 2,3,4,6-tetrachlorophenol 1n addition to hydroxypoly-
chlorlnated dlphenyl ethers, PCDDs, PCDFs, chlorodloxlns (1000-3000 ppm),
1928A 4-8 06/23/86
-------
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1928A
4-9
06/20/86
-------
chlorofurans (200-600 ppm) and chlorinated dlphenyl ethers (Firestone,
1977a). Poor temperature control 1n this final phase has been postulated to
account for the formation of PCDFs (and PCDDs) from the chlorinated dlphenyl
ethers (Firestone et al., 1973). The length of time that chlorlnatlon con-
tinues undoubtedly has some Influence, since refluxlng of PCP In benzene for
6 hours with chlorine bubbling can produce OCDF 1n 52% yield (Pllmmer, 1973).
Commercial sodium 2,4,5-trlchlorophenolate tends to contain more PCDDs
and PCDFs than the chlorophenol Itself (Firestone et al., 1972). Most of
the PCDFs possessed 3-6 chlorine atoms. While 2,3,4,6-tetrachlorophenol
formulations did contain TCDFs to OCDFs (see Table 4-4), no PCDFs or chloro-
ethers were found 1n samples of 2,4- or 2,6-dlchlorophenol.
Levin and Nllsson (1977) reported that Swedish sawmill workers were
exposed to PCDFs (levels up to 6 ppm) 1n Finnish 2,3,4,6-tetrachloro-
phenolate In the sawdust.
PCP has been detected 1n urban air (at concentrations of 0.25-0.93 and
5.7-7.8 ng/m3), In waters from various manufacturing and processing
plants, 1n sewage-plant effluent (1-5 vg/it) In rain, snow and lake
waters (2284, 14 and 10 ng/9.), 1n creek water containing Industrial wastes
(at levels of 0.1-10 mg/l); soil residues after application of 15 and 45
kg/ha PCP were 20.4 and 69.1 mg/kg (WHO, 1979). Trlchlorophenol (unspeci-
fied Homers) has been Identified In river water, tap water (2.4 mg/i),
landfill leachate (40 mg/8.), and In effluent from sewage treatment plants
(WHO, 1979). Other contaminants such as PCDDs and polychlorlnated dlphenyl-
ethers have been Identified In technical grade PCP (Rappe et al., 1978a,
1979a; Nllsson and Renberg, 1974; Buser, 1975).
4.2.2. PCDFs In Polychlorlnated Blphenyls. PCBs are mixtures of chlori-
nated blphenyls widely used 1n a number of Industrial applications, such as
1928A 4-10 06/23/86
-------
heat exchange, dielectric fluids, hydraulic and lubricating fluids, plastl-
dzers, printing Inks and flame retardants (WHO, 1976). Large amounts of
PCBs have been released Into the environment since 1929. Because of the
stability of some Isomers to chemical and biological attack, they are today
ubiquitous and persistent pollutants of ecosystems (Wassermann et al., 1979).
The presence of PCDFs In PCBs has been detected by several authors 1n
American, European and Japanese commercial PCB mixtures (Vos et al., 1970;
Nagayama et al., 1976; Roach and Pomerantz, 1974; Bowes et al., 1975a).
Because of production differences the PCDF concentrations and Isomers vary
with PCB type and origin. Bowes et al. (1975b) found levels of 0.8 and 2
ppm 1n American-made Aroclor 260 and 1248, respectively; 8.4 and 13.6 ppm of
PCDF were estimated In German Clophen A-60 and French Phenoclor DP-6
(Nlsbet, 1976). In the late 1960s, a significant difference 1n toxlclty to
chick embryos was discovered among three supposedly Identical PCB formula-
tions originating from three different countries {Vos and Koeman, 1970).
Large amounts of comparatively more polar contaminants were detected 1n a
German PCB (Clophen A-60) and a French PCB (Phenoclor DP-6) but not 1n an
American PCB (Aroclor 1260). Two of the contaminants 1n the German PCB were
Identified by GC/MS as a TCDF and a PeCDF (Vos et al., 1970); these com-
pounds apparently accounted for the observed acute toxlclty of the whole PCB
formulation. PCDFs were found also 1n American PCBs but not as much as In
the European formulations (Bowes et al., 1973). Kanechlors from Japan were
also found to contain PCDFs (Bowes et al., 1975a; Roach and Pomerantz,
1974). The average PCDF content 1n Aroclors 1248, 1254 and 1260 from the
United States was then put between 1 and 2 ppm. Less than ppt levels were
detected In Aroclor 1016 (Bowes et al., 1975a,b). The 2,3,7,8-TCDF and
2,3,4,7,8-PeCDF were detected 1n Aroclors 1248 and 1254, and In Kanechlors
1928A 4-11 06/23/86
-------
200 and 500 (Bowes et al., 1975a,b); Clophen A-60 contained ~8 ppm PCDFs,
and Phenoclor DP-6 contained -14 ppm. The PCBs with the highest PCDF
contents were toxic to chick embryos.
In 1968, some 1200 Japanese people consumed a rice oil (Kaneml) contami-
nated with 800-1000 ppm of a Japanese PCB formulation, Kanechlor 400, which
had leaked from a heat exchanger (Nagayama et al., 1975). This caused Yusho
disease. Several months before this Incident, 0.50 million chicks were
killed by a crude bran oil from the same origin but the warning went
unheeded (Kohanawa et al., 1969). The composition of Kanechlor 400
resembled that of Aroclor 1248. Of the GC peaks assigned to PCDFs six made
up 2 ppm of the PCB. However, reports of the total PCDF content varied
greatly from Investigator to Investigator. For example, Roach and Pomerantz
(1974) detected 1 ppm, Nagayama et al. (1975) 18 ppm and Kuratsune et al.
(1976) reported 33 ppm. These values were obtained on packed GC columns and
so are expected to be maximal figures. Later capHlary-GC work confirmed a
level of 2-5 ppm (Nagayama et al., 1977; Rappe et al., 1977), the major
PCDFs being the toxic 2,3,7,8-TCDF (0.45 ppm) and the 2,3,4,7,8-PeCDF (-0.2
ppm). It was calculated that the contaminating Kanechlor 400 PCB contained
a PCDF level 250 times greater than the value found for an unused Kanechlor
400 formulation (Nagayama et al., 1975). A 4-fold Increase of PCDFs (15-20
ppm) occurred when a PCB was used for 2 years 1n a heat exchanger 1n a situ-
ation similar to the suspected source of rice oil contamination (Morlta et
al., 1977a). More than 40 Isomers were found; of those found, the 2,3,7,8-
TCDF was the most abundant (1.25 ppm). Later work (Mazer et al., 1983a,b;
Rappe et al., 1984) showed that the 2,3,4,8-TCDF co-eluted. The chroma-
tographlc profile of the TCDF fraction was very similar to that of Kaneml
1928A 4-12 06/23/86
-------
oil (Buser et al., 1978d; Rappe et al., 1977). A summary of the levels of
PCDFs In various PCB formulations and 1n Kaneml oil 1s given In Table 4-6.
Table 4-7 gives the suspected maximum levels of the toxic 2,3,7,8-TCDF
and 2,3,4,7,8-PeCDF In these formulations (Morlta et al., 1977a). The
levels of the 2,3,7,8-derlvatlves were Identified by retention time of the
pure standard and were subsequently confirmed by Buser et al. (1978d) using
the GC/MS. If the same order of elutlon Is followed on the GC columns
utilized by these workers, probably peaks 6-10 and 13-19 of Buser et al.
(1978d) correspond to peaks 2 and 3, respectively, of Morlta et al. (1977a)
(Figure 4-1). Similarly, peaks 23-33, 37-40, 44-47, 51-52, 53-56, 58-66,
68-70, 71-73, 77-78 and 85 denoted by Buser et al. (1978d), correspond to
peaks 3, 5, 6, 9, 10, 11, 12, 14, 16 and 17 of Morlta et al. (1977a). The
2,3,7,8-TCDF-der1vat1ve, therefore, constitutes >90% of peak 6 as observed
by Morlta et al. (1977a), accounting for the near equivalence of the levels
of the 2,3,7,8-TCDF found 1n Kaneml oil by both sets of workers. Later It
was found that the 2,3,4,8-TCDF co-eluted (Mazer et al., 1983a,b; Rappe et
al., 1984). The actual level of the 2,3,4,7,8-PeCDF Is probably -70% of the
levels quoted 1n Table 4-7, since peak 68 of Buser et al. (1978d) 1s 70% of
the combined height of peaks 68, 69 and 70 corresponding to peak 12 of
Morlta et al. (1977a). The figures quoted 1n Table 4-7 are therefore
unlikely to have an error >30% provided that the elutlon profiles on the two
GC columns are comparable. Thus, these two toxic Isomers may have made up
between 5 and 34% of the total PCOFs 1n the PCBs. The low values for the
Aroclors T-1242 and T-1260 are to be noted. The low toxldty of Aroclor
1260 can thus be correlated with the low level of toxic CDBF (Vos and
Koeman, 1970).
1928A 4-13 06/24/86
-------
10
CO
3>
TABLE 4-6
Content in Some PCBs and 1n Kanemi Oil3
o
\
r\>
•«^
00
PCDF Content (ppm)
Substrate
Yu-Cheng oil5 (3)
Kanpmi oilb (4) (Yusho)
Unused Kanechlor 400
Used Kanechlor 400b (3)
Kanemi oil (Yusho)
Kanemi oil (Yusho)
Used Japanese
PCB (Mltsublshl-
Monsanto T-1248)
Kanemi oil
Kanechlor 300
Kanechlor 300
Kanechlor 400
Kanechlor 400
Kanechlor 400
Kanechlor 500
Kanechlor 500
Kanechlor 600
Kanechlor 600
Phenoclor DP-4
Phenoclor DP-5
Trl- Tetra- Penta- Hexa-
0.15 1.4 2.5 1.6
-
4.2 4.5 5.5 1.4
0.02 0.52 1.3 0.81
6.7 1.6
1.3
0.3 12.2 10.4 0.9
-
1 .
0.2 1.7 1.1 3.1
..
0.2 0.5 0.4
-
1.7 1.6 0.5
4.6 2.7 2.6
Total
0.14-0.18
2-7
33
20-510
5.7
5.0
16
2.7
8.3
1.3
24
18
--
6.1
3.3
1.1
4.0
3.8
9.9
Reference
Mlyata et al., 1985
Mlyata et al.. 1985
Mlyata et al.. 1985
Mlyata et al.. 1985
Buser et al., 1978d
Nagayama et al . , 1976
Buser et al., 1978d
Morlta et al., \977a
Morlta et al., 1977a
Nagayama et al . , 1976
Morita et al., 1977a
Nagayama et al . , 1976
Roach and Pomerantz,
1974
Morlta et al., 1977a
Nagayama et al . , 1976
Morlta et al., 1977a
Nagayama et al . , 1976
Morlta et al., 1977a
Morlta et al., 1977a
-------
VO
ro
CO
3>
TABLE 4-6 (cont.)
o
cr
no
CO
GO
PCDF Content (ppm)
Substrate
Phenoclor DP-6
Phenoclor DP-6
Aroclor
Aroclor
Aroclor
Aroclor
Aroclor
Aroclor
Aroclor
Aroclor
Aroclor
Aroclor
Aroclor
Clophen
Clophen
Clophen
Clophen
aNo data
T-1200
T-1241
T-1242
T-1248
T-1248C
T-1254
T-1254
T-1254
T-1260
T-1260
T-1264
A-30
A-40
A-50
A-60
found for hepta-
Trl- Tetra-
0.2 2
0
0
2
2
0
0.3 5
0
0
0
0
0
4
1.6 2
1.5 5
0.7 8
1
.1
.7
.1
.4
.3
.5
.8
.1
.2
.1
.2
.8
.8
.3
.4
.3
.4
Penta-
2.6
10
0.4
2.7
2.3
2.3
5.6
0.2
0.4
3.6
0.3
0.9
9.4
1.0
6.9
4.1
5.0
Hexa-
5
2
0
0
.6
.9
.5
.8
-
Total
11
14
1
5
4
.0
.9
.5
2.8
0
1
0
1
0
0
2
1
2
.7
.4
.9
.9
.3
.5
.0
•-
--
.8
.2
12
1
1
5
0
2
16
4
14
15
8
.7
.5
.6
.8
.2
.9
.4
Morlta
Bowes
Bowes
Morlta
MorUa
Morlta
MorUa
Bowes
Bowes
MorUa
Bowes
Morlta
Morlta
MorUa
MorUa
MorUa
Bowes
Reference
et al.,
et al..
et al.,
et al.,
et al.
et al.
et al.
et al.,
et al.,
et al.
et al.,
et al.
et al.
et al.
et al.
et al.
et al.,
, 1977a
1975a,b
1975a,b
, 1977a
, 1977a
. 1977a
, 1977a
1975a,b
1975a,b
, 1977a
1975a,b
, 1977a
, 1977a
, 1977a
, 1977a
, 1977a
1975a,b
bYu-Cheng oil also contained 22-113 ppm PCBs, 9-38 ppm PCQs. Kaneml oil also contained 151-968 ppm
PCBs, 490-866 ppm PCQs. Kanechlor 400 also contained 999,800 ppm PCBs, 209 ppm PCQs. Used Kanechlor 400
also contained 961,900-999,000 ppm PCBs, 690-31,000 ppm PCQs.
cUsed PCB
-------
TABLE 4-7
Suspected Maximum Levels of Toxic PCDFs 1n Various PCBs and In Kaneml 011
(Kaneml oil also contained 900 ppm PCBs and 800 ppm PCQs;a
Taiwan rice oil also contained 60-100 ppm PCBs and 90-180 ppm PCQs)
Formulation
PCDF Levels (ppm)
2,3,7,8-b'c 2,3,4,7,8-b'd
TCOF PeCDF
Total
Percentages of
Total PCDFs
for These Two
Derivatives
Taiwan rice
oil (2)a
Kaneml oil3
Kaneml o11b
Phenoclor3
DP -4
DP-5
DP-6
Kanechlorb
KC-300
KC -400
KC-500
KC-600
Aroclorb
T-1241
T-1242
T-1248
T-12486
T-1254
T-1260
T-1264
0.001-0.005
0.2
0.28
0.7
2.2
0.9
2.2
1.6
0.7
0.1
1.1
0.2
0.2
1.1
--
-_
2.4
0.02-0.70
0.7
0.42
0.4
0.8
0.6
0.6
0.9
0.7
0.1
0.4
0.1
0.8
1.4
1.6
0.1
2.3
0.08-0.10
2.02
2.68
3.8
9.9
10.5
8.3
23.8
6.1
1.1
5.9
4.5
2.8
12
5.6
2.2
16
20-25
45
26
29
30
14
34
11
23
18
25
7
36
20
29
5
29
1928A
4-16
06/20/86
-------
TABLE 4-7 (cont.)
PCDF Levels (pom)
Formulation
Clophenb
A-30
A -40
A-50
2,3,7,8-b'c
TCDF
1.0
2.1
3.6
2,3,4,7,8-M
PeCDF
0.1
0.7
0.6
Total
4.9
14
15
Percentages of
Total PCDFs
for These Two
Derivatives
22
20
28
aMasuda et al., 1982
Calculated from MorHa et al., 1977a
cBased on GC retention time, but subsequently confirmed by Buser et al.,
1978d
^Assumed the order of elutlon obtained by Buser et al., 1978d, Is followed
on the GC column utilized
eUsed PCB
-- Below detection limit
1928A
4-17
06/20/86
-------
I 17
V f
\ I1,
B
FIGURE 4-1
Comparison of Specific Ion Current Profiles
after Chromatography of Kaneml 011
Source: (A) Morlta et al. (1977a); (B) Buser et al. (1978d)
1928A
4-18
06/20/86
-------
A similar Incident occurred 1n Taiwan 1n 1978 affecting nearly 2000
people. This has been dubbed the "Yu-Cheng" episode (Chen et al., 1981;
Kashlmoto et al., 1981). The total PCDF content Is provided In Table 4-6,
the congener concentrations still not being available. The concentrations
of the toxic 2,3,7,8-TCDF and the 2,3,4,7,8-PeCOF are given 1n Table 4-7.
Masuda et al. (1982) also found 0.01-0.02 ppm of the 2,3,4,6,7-PeCDF, -0.01
ppm HxCDF and 0.04-0.05 ppm of an unidentified TCDF. The PCDF Isomers 1n
the rice oils exposing people 1n both Yusho and Yu-Cheng episodes were
Identical but the concentration 1n the Taiwan rice oil was at most 5X of the
Japanese concentration. Rappe et al. (1983) detected PCDF at levels of 1-18
ppm In Kanechlor mixtures 300-600. More recently, using a complete set of
PCDF standards and an Isomer-speclflc analytical method, Rappe et al. (1984)
reported on levels of 2,3,7,8-substHuted PCDF congeners found 1n a large
number of commercial PCB products (Table 4-8).
Concentrations of PCDFs 1n PCBs are Indicative 1n some cases of the
original concentrations of PCDFs. However, Buser et al. (1978a) demonstrat-
ed that after 2 years of using M1tsub1sh1-Honsanto PCB as a heat exchanger,
a 4-fold Increase 1n PCDF concentration occurred, the toxic 2,3,7,8-TCDF
being the major component.
4.2.3. PCDFs 1n Phenoxy Herbicides. Phenoxy herbicides are widely used
for the control of wood and herbaceous weeds by spraying from the air or
from the ground. They reach aquatic systems directly when applied for
macrophyte control 1n lakes, ponds and Irrigation ditches or Indirectly
during application near water, or through runoff, reaching concentrations as
high as 1-3 mg/l of water (CNRC, 1978a). A 50:50 mixture of the n-butyl
esters of 2,4-D and 2,4,5-T (Agent Orange) was used for military purposes
(defoliation or crop destruction) 1n South Vietnam (NAS, 1974). The phenoxy
1928A 4-19 06/23/86
-------
CO
TABLE 4-8
PCDFs In Comnerclal PCBs (ng/g)*
PCB-typ
Pyralene
A1254
^ A1260
g A30
A40
A50
A60
T64
Clophen C
Blank
Blank
Trl-
Total
700
63
10
500
1300
7400
770
47
710
ND
ND
Tetra-
2378
53
19
13
35
180
3300
840
23
54
ND
ND
Total
630
1.400
110
573
2.600
20,000
6,900
360
1,200
ND
ND
12348
12378
10
690
48
14
96
760
1100
97
34
ND
ND
Penta-
23478
T
490
56
28
8
1100
990
122
30
ND
ND
Total
35
4000
260
160
1700
8000
8100
840
270
ND
ND
123479
123478
ND
2500
500
50
79
700
1600
520
Nd
ND
ND
123678
ND
2100
120
59
68
360
330
390
T
ND
ND
Hexa-
123789
ND
190
190
ND
ND
18
170
58
ND
ND
ND
Heota-
234678
ND
130
27
ND
T
98
330
41
ND
ND
ND
Total
ND
10.000
1.500
220
310
3.100
6.800
2,600
T
ND
ND
Total
ND
960
1300
T
ND
75
2000
220
ND
ND
ND
Rec 2378-
TCDD-X
79
78
88
79
79
95
95
72
79
90
87
'Source: Rappe et-al., 1984
ND = Not detected
oo
er
-------
herbicides, 2,4-D, 2,4,5-T, MCPA, 2-(2,4,5-tr1chlorophenoxy)prop1on1c add
(Sllvex) and 2-(2,4-d1chlorophenoxy)prop1on1c acid (Dlclorprop) are synthe-
sized by condensing the appropriate haloalkanolc acid with the appropriate
chlorophenol, usually under alkaline conditions during reflux. Any PCDD and
PCDF Impurities remain 1n the product unless the latter Is purified. In
addition, since PCDOs and PCDFs are formed from chlorophenols under hot
alkaline conditions, these contaminants may also be formed 1n the presence
of excess chlorophenol.
Most attention has been accorded to the content of 2,3,7,8-TCDD 1n these
formulations, but PCDFs have also been detected 1n these products (Rappe et
al., 1978c; Hucklns et al., 1978; Ahllng et al., 1977). The analytical
techniques, however, were not refined enough to permit Identification of
specific Isomers. This has now been accomplished (Rappe et al., 1978c;
Cochrane et al., 1983). The major PCDDs present In 2,4-D are the compara-
tively nontoxlc 2,7-DCDD-, 1,3,7-TrCDD-, 1,3,4,8-TCDD and 1,3,6,8-TCDD.
Various 2,4,5-T formulations of European origin 1n the 1960s contained
2,3,7,8-TCDD at levels ranging from 0.1-1 vg/g as the predominant PCDD
(Table 4-9). Two samples showing the presence of TCDFs did not contain the
2,3,7,8-TCDD. In the Agent Orange samples, the major PCDD was the 2,3,7,8-
TCDD (0.12-5.1 v9/g levels). Only one sample contained PCDFs; they were
one tr1-, four tetra- and one penta-CDFs at a total level of 0.7 yg/g.
The 2,3,7,8-TCDF was not present. All samples, however, showed the presence
of chlorinated dlphenyl ethers, which are known precursors of PCDFs In
pyrolytlc reactions. The chlorinated dlphenyl ether content may range as
high as 1% (Rappe et al., 1978c). The PCDFs may be formed from a metal
Ion-promoted condensation of two 2,4,6-trlchlorophenolate Ions, by a mechan-
ism similar to the palladium (II) acetate synthetic method discussed
1928A 4-21 06/23/86
-------
vO
rsj
CD
TABLE 4-9
PCDFs and PCDDs In Phenoxy Herbicides
PCDf/PCOD Amounts (mq/kq)
Phenoxy
Herbicide
D1-
Trl-
Tetra-
Penta-
Hexa-
Hepta-
Octa-
Total
CDF CDD CDF CDD CDF CDD CDF CDD CDF COD CDF CDD CDF CDD CDF
CDD
Reference
2.4.5-T Esters
Butyl
Isooctyl
Butoxyethyl
Butoxyethyl
Agent Orange
0.01 - 0.95
0.15 0.18
0.96
0.15 0.18
0.11 0.22
a a a 0.13 0.018 0.008 0.007 <0.005 <0.0 <0.005 <0.005 <0.005 <0.005 <0.153 <0.40
0.15 - 0.02 0.12 - - - - a a a a - 0.29
0.15 0.10 0.50 0.40 1.1 0.20 0.05 - - a a a a 0.70 1.80
0.02 -5.1-- - -a a a a - 5.12
a = Not analyzed for; - = Below detection limit
1 = Rappe et al., 1978c
2 = Ahllng et al.. 1977
o
*v
00
-------
previously. In fact, H Is known that the chlorinated dlphenyl ethers and
PCDFs are formed at the beginning of the reaction before most of the
1,2,4,5-tetrachlorobenzene reacts, which Is used as starting material for
the preparation of 2,4,5-tMchlorophenol 1n the European manufacturing pro-
cess; PCDDs tend to be formed near the end of the process when 2,4,5-tM-
chlorophenolate formation 1s favored (Rappe, 1978; Adamoll et al., 1978).
The synthesis of PCDFs from chlorobenzenes Is also discussed In Section
2.5.2.2.
The concentration of the 2,3,7,8-TCDD 1n technical 2,4,5-T used 1n Agent
Orange manufactured between 1958 and 1969 ranged from 1-32 ppm (Young et
al., 1978). Levels In Agent Purple (n-butyl 2,4-D: n-butyl 2,4,5-T: 1so-
butyl 2,4,5-T:: 5:3:2) formulated 1n the mid-1950s appear to be even higher
than those In Agent Orange. Levels of 2,3,7,8-TCDD In the technical 2,4,5-T
used for Its formulation have been estimated to be as high as 90 ppm (Young
et al., 1978). Other herbicides containing 2,4,5-T used In Vietnam, namely,
Agent Pink (n-butyl 2,4,5-T/1sobutyl 2,4,5-T:: 3:2), Agent Green (n-butyl
ester of 2,4,5-T), Dlnoxol (1:1 butoxyethanol esters of 2,4,5-T and 2,4-D),
and TMnoxol (40% butoxyethanol ester of 2,4,5-T), have never been Inten-
sively studied for their PCDD or PCDF contents. However, only minor amounts
of PCDF and of other PCDD 1n relation to the major contaminant TCDD, could
be found 1n these herbicides. In >450 samples analyzed, TCDD had the mean
value of 1.98 vg/g (Young et al., 1975, 1983). As a result of govern-
mental regulation, efforts have been made to minimize the formation of
2,3,7,8-TCDD and now the products are estimated to contain <0.1 vg/g.
4.2.4. PCDFs In Chlorodlphenyl Ether Herbicides (PCDPE). Yamag1sh1 et
al. (1981) reported on the occurrence of PCDD and PCDF 1n the commercial
dlphenyl ether herbicides CNP, NIP and X-52. The levels of the various
Isomers are reported In Table 4-10.
1928A 4-23 06/23/86
-------
TABLE 4-10
Levels of PCDDs and PCDFs 1n Commercial Dlphenyl Ether Herbicides*
TrCDDs
TCDDs
PeCDDs
HxCDDs
MCDFs
DCDFs
TrCOFs
TCDFs
PeCDFs
HxCOFs
CNP
ND
14.00
37.00
0.80
ND
0.35
0.41
0.40
1.00
0.20
NIP
0.15
0.38
0.05
ND
0.34
0.12
0.47
0.29
ND
ND
X-52
0.03
0.03
0.01
ND
0.48
0.21
0.45
0.32
0.08
ND
*Source: Yamaglshl et al., 1981
ND = Not detected
1928A 4-24 06/20/86
-------
4.2.5. PCDFs In Hexachlorobenzene. Hexachlorobenzene Is a contaminant 1n
the production of many chemicals, such as tetrachloroethylene, trlchloro-
ethylene, carbon tetrachlorlde, chlorine, vinyl chloride, dlmethyltetra-
chloroterephthalate, Atrazlne®, S1maz1ne®, pentachloronltrobenzene and
Mlrex®. Hexachlorobenzene Is also used as an economic poison for the con-
trol of wheat bunt and fungi. As an environmental contaminant 1t has been
found In river water samples, drinking water, sewage treatment plants,
effluent waters from various chemical plants, urban rain water runoff (up to
339 ng/a) and surface waters (2.5 mg/a) (WHO, 1979). Vlllaneuva et al.
(1974), analyzing three commercial hexachlorobenzene preparations, Identi-
fied OCDD, HpCDF and OCDF. This last compound was present 1n concentrations
ranging from 0.35-58.3 ppm. These data suggest that PCDFs may also be found
1n association with hexachlorobenzene.
4.2.6. PCDFs 1n Hexachlorophene. The bacterldde, hexachlorophene, 1s
prepared commercially from 2,4,5-tMchlorophenol (Nllsson et al., 1978). It
1s purified, however, during this process and levels of 2,3,7,8-TCDD are <30
vg/kg. Nevertheless, hexachlorophene can contain 100 ppm of the
1,2,4,6,8,9-hexachloroxanthene (Gothe and Wachtmelster, 1972).
4.2.7. PCDFs 1n PBBs. The PBBs gained their greatest notoriety as a
result of a severe contamination of meat and dairy products In Michigan 1n
October, 1973 (Cordle et al., 1978). The contaminant, Flremaster BP-6
manufactured by the Michigan Chemical Corporation, contained 2% tetrabromo-
blphenyls, 10.6% pentabromoblphenyls, 62.8% hexabromoblphenyls, 13.8%
heptabromoblphenyls and 11.4% other bromoblphenyls. As a result of error,
Flremaster BP-6 was added to cattle feed Instead of magnesium oxide. The
levels of contamination Included the following: lot 405, 2.4 ppm; lot 410,
1928A 4-25 06/23/86
-------
1790 ppm; and lot 407, 4300 ppm. Milk from affected herds contained from
2.8-271 ppm on a fat basis. Contaminated butter (1-2 ppm), cheese (1.3-15
ppm) and canned milk (1.2-1.6 ppm) were later seized and destroyed.
PBDFs were found In pyrolyzed PBBs by O'Keefe (1978b) and Buser et al.
(1978a). PBDFs were not found 1n one study on unheated Flremaster FF-1
(Hass et al., 1978). Hass et al. (1978) concluded that the amount of BDBFs
must be <0.5 ppm, and that penta- and hexabromonaphthalenes were present to
the extent of 150 and 70 ppm, respectively. A metabolite of Flremaster BP-6
1n dogs, 6-hydroxy-2,2',4,4',5,5'-hexabromob1phenyl, decomposed at 230-260°C
on an OV-101 column 1n a gas chromatograph to form two PeBDFs (Gardner et
al., 1979). Thus, PBDFs can be formed from pyrolysls on GC columns as well
as being originally present 1n PBB formulations. Such artifacts need to be
Investigated before PBDF contamination of PBB formulations Is considered
confirmed.
4.2.8. PCDFs 1n Metal Salts. Salts of metals often used 1n Industrial
processes may contain PCDFs and PCDDs (Helndl and Hutzlnger, 1986). Table
4-11 shows that the chlorides of Iron, aluminum and copper contain between
0.02 and 60 ppb PCDFs and 0.02-0.63 ppt PCDDS, both mostly as OCDF and HpCDF.
4.3. PHOTOCHEMICAL PRODUCTION
Another potential source of PCDFs derives from the photodegradatlon
reactions of a number of chemicals present 1n the ecosystem as pollutants.
Perhaps the most Important potential source of environmental contamination
Is the photochemical production of PCDF from PCBs. Hutzlnger (1972)
observed 0.2% 2-PCDF production after 7 days of ultraviolet Irradiation (310
nm) of aqueous solutions of 2,5,2'5'-tetrachloro- and 2,5-d1chloro-b1phenyls
(5 mg/l). These products were confirmed by Crosby and Mollanen (1973)
(see Section 2.4.2. for more details of laboratory photochemistry).
1928A 4-26 06/23/86
-------
TABLE 4-11
PCDFs and PCDDs In Several Metal Salts*
(ppb)
Salt
FeCl3
A1C13 (A)
A1C13 (B)
CuCl2
CuCl
T1C14
S1C14
OCDF
42
<0.02
34
0.5
0.2
<0.02
<0.02
HpCDF
12
<0.02
0.1
0.1
0.08
<0.02
<0.02
OCDD
<0.02
<0.02
<0.02
0.6
0.03
<0.02
<0.02
HpCDD
<0.02
<0.02
<0.02
0.03
<0.02
<0.02
<0.02
*Source: Helndl and Hutzlnger, 1986
1928A 4-27 06/23/86
-------
However, Hutzlnger (1972) found no PCDFs after some aqueous PCB samples (167
mg/l) had been exposed to sunlight for >2 months. This discrepancy may
have arisen because of the different Irradiation times and different
wavelengths used. The yield of the products was wavelength dependent.
Irradiation at 254 nm (mercury arc) decomposed PCDF photoproducts but only
slowly at 310 nm or at the wavelengths encountered at sea level for
sunlight, or for a solar simulator. Triplet sensltlzers (for example,
4,4'-dichlorobenzophenone) Induced faster decomposition as observed In
methanollc solution alone for the photodecomposltlon of 2,8-DCDF (Crosby and
Mollanen, 1973). Thus, the presence of other compounds may be Important.
After ultraviolet Irradiation of some chlorophenols, Crosby et al.
(1973) detected OCDD but PCDFs were not sought. Chlorinated o-phenoxy-
phenols (predloxlns), which are common Impurities In chlorophenols (1-5%),
also produce PCDDs after ultraviolet Irradiation (NHsson et al., 1974) but
again PCDFs were not sought specifically. Another source of PCDFs through
photodecomposltlon Is from PCDPEs, which are often present at levels of 100
ppm In commercial chlorophenols. Irradiation of 100 ppm solutlos In
methanol, ethanol or n-hexane with light of 248-579 nm caused cycllzatlon In
dlphenyl ethers containing at least one ortho-chlorine. Reductive dehalo-
genatlon was enhanced In hexane. Although no yields were explicitly given,
the degradation was characterized by an Induction period and then decompo-
sition, obeying a first-order kinetic law (Norstrom et al., 1976a, 1977;
Choudhry et al., 1977a,b). Photodegradatlon of polychlorobenzenes (PCBZ)
can also be a source of PCDF production. Laboratory Irradiation at x >285
nm of a solution of PCBZ In the presence of phenol yielded PCDF (<1%)
containing two Cl atoms less than the starting PCBZ. This reaction probably
1928A 4-28 06/23/86
-------
proceeds through the Intermedlacy of PCDPE (Choudhry et al., 1983). The
relevance of these laboratory photodecomposHlons to environmental sources
of PCDFs Is unknown.
Another photochemical process of potential environmental Importance 1s
dechlorlnatlon of the higher PCDFs. This topic will be considered 1n
Chapter 5.
4.4. SOURCES OF PCDF FROM BURNING AND OTHER HIGH TEMPERATURE PROCESSES
In 1978 Dow Chemical scientists proposed the "Trace chemistries of fire"
hypothesis. This stated that many contaminants arise In trace amounts
during chemical reactions during combustion. PCDF and PCDD are Included
among these compounds. To assess the relative Importance of these sources,
Sheffield (1985a,b) estimated the environmental release of PCDF and PCDD
from combustion sources 1n Canada (Table 4-12) using the small data base at
his disposal.
4.4.1. Thermal Degradation of Technical Products. PCDFs can arise from
pyrolysls of specific PCBs, chlorinated benzenes and chlorinated dlphenyl
ethers, and PBDFs from the analogous bromlnated precursors. The occurrence
of these compounds must be taken Into consideration since they are released
Into the environment with their co-contaminants and may produce PCDFs upon
Incineration.
The levels of PCDFs 1n PCBs Increase with the length of time 1n service
at high temperatures as heat exchange media (Morlta et al., 1977b; Buser et
al., 1978d), as originally suggested by Kuratsune et al. (1976). A contrary
finding has been reported by Cull and Dobbs (1984). Aroclor 1254 on pyroly-
sls also contained the major toxic PCDFs, but the relative amounts of the
products differed somewhat from those obtained from the Mitsubishi-Monsanto
T-1248 (Buser and Rappe, 1979). The major PCDFs Identified In the used
1928A 4-29 06/23/86
-------
TABLE 4-12
Estimated PCOD and PCDF Releases to the Canadian Environment
from Combustion Sources3
Sources
Release (q/year)
PCDD
PCDF
Municipal Incinerator
Fly ash
A1r emission
Sewage sludge Incineration
A1r emission
Coal-fired utility boilers
Fly ash
Air emission
Fuelwood combustion
A1r emission
Residential oil combustion
A1r emission
Residential gas combustion
Source: Sheffield, 1985a,b
bPCDD + PCDF release
ND = Not detectable
2900-7100
250-13,700
1400-3300
ND-300
300
1800
4900-15,600
550-21,700
1500-6500
30-1300
700
not estimated
not estimated
Air emission
Wigwam burners/Wood waste boilers
Air emission
Railway ties
Air emission
Forest fires
Air emission
Slash burning
Air emission
Cigarette smoke
Air emission
Motor vehicles
A1r emission
900
0-30, 200b
6000°
58,700
3300
2-4
200
not estimated
not estimated
not estimated
not estimated
not estimated
1928A
4-30
06/23/86
-------
MHsublshl-Monsanto T-1248 were the 2,3,7,8-TCDF (1.25 ppm), the 2.3,4,7,8-
PeCDF (the pyrolysls product of 2,4,5,2',4',5'-hexachloroblphenyl),
1,2,3,7,8-PeCDF, 2,3,4,6,8-PeCDF, 1,2,3,4,8-PeCDF, 1,3,4,7,8-PeCDF (also
from pyrolysls of 2,4,5,2',4',5'-hexachlorob1phenyl) and the 2,3,4,6,7,8-
HxCDF. The 2,3,4,8- and 2,3,7,8-TCDFs were later shown to co-elute (Mazer
et al., 1983a,b; Rappe et al., 1984). Whereas the used MHsublshl-Monsanto
T-1248 was exposed for years to elevated temperatures 1n the liquid phase,
the laboratory pyrolyses of Aroclor 1254 and 1260 were performed In the gas
phase for a few seconds up to a maximum temperature of 700°C (Buser and
Rappe, 1979). WHh Aroclor 1254 (tr1- to hepta-PCBs), mostly MCDFs to
PeCDFs were formed at a level of -2%. WHh Aroclor 1260 (penta- to octa-
PCBs), mostly TrCDFs to HpCDFs were produced at a similar level. The toxic
2,3,7,8-TCDF was the most abundant TCOF, most likely derived from
2,4,5,2',4',5'-hexachlorob1phenyl or 2,4,5,3',4'-pentachlorob1phenyl. The
acutely toxic 1,2,3,7,8- and 2,3,4,7,8-PeCDFs were the major PeCDFs. The
former probably Is produced from pyrolysls of 2,3,4,2',4',5'-hexachlorobl-
phenyl , and the latter from 2,4,5,2',4',5'-hexa- or 2,3,4,5,2',4',5'-hepta-
chloroblphenyl. Pyrolyzed Aroclor 1254 also contained significant amounts
of 1,3,4,7,9-PeCDF. These results were confirmed by Paaslvlrta et al.
(1985) who also found small amounts of chlorinated phenols, naphthalenes,
and MCDFs and DCDFs (Table 4-13) produced between 500 and 700°C.
The polybromlnated blphenyl, Flremaster FF-1, when pyrolyzed for 20
minutes at 380-400°C 1n open glass tubes, produced 40 ppm TBDFs and 4 ppm
PeBDFs based on PBB content. Only trace amounts (~1 ppm) were found 1f the
pyrolysls was redone In a nitrogen atmosphere (O'Keefe, 1978b). It was
postulated that since 2,4,5,2',4',5'-hexabromoblphenyl was the major hexa-
bromoblphenyl present, nearly all of the tetrabromo-lsomer was the 2,3,7,8-
Isomer. The larger yield 1s expected from the weaker ring carbon-bromine
1928A 4-31 06/23/86
-------
rs>
CO
TABLE 4-13
Pyrolysls Products of Aroclor 1254 In a Quartz Tube for Pyrolysls Time of 3 Seconds*
The Carbon Filter Adsorbed any Volatilized PCB
[relative amounts (X)]
INJ
Pyrolysls Temperature
(X)
500
600
700
Filter
cii
0.1
0.3
0.01
52
C12
14
13
5.2
44
PCDF
"3
39
39
29
3.9
C14
40
36
49
ND
C15
7.4
11
15
ND
Cl6
ND
0.8
1.5
ND
"2
17
0.7
4.6
NA
PCDD
"3
10
18
14
NA
C14
73
57
66
NA
Cl5
ND
24
17
NA
*Source: Paaslverta et al., 1985
ND = Not detected
NA = Not analysed
ro
o
CO
-------
bond relative to that of carbon-chlorine. The production of PBDFs from
various PBBs In sealed ampules was also studied by Buser et al. (1978a) and
has been discussed In Section 4.2.5.1. The 10-mlnute pyrolysls of
2,4,6-trIbromophenol, pentabromophenol, tetrabromoblsphenol A and tetra-
bromophthallc anhydride, all used as flame retardants, resulted In PBDD and
PBDF between 700 and 900°C, except for the anhydride that did not form
PBDFs/PBDDs at any of the three temperatures. In general, the optimum
temperature to form PBDDs and PBDFs was at 800°C; 2,4,6-trlbromophenol
pyrolysls produced 8950 ppm TBDFs, pentabromophenol formed 7042 ppm HpBDFs
with no detectable amounts of the other PBDF congeners. Small amounts were
noted at 900°C, although lower congeners did appear at 700°C. The tetra-
bromoblsphenol A still produced relatively high PBDF concentrations at 900°C
and PBDFs predominated over PBDDs unlike at other temperatures and for the
other two phenols when they were pyrolyzed. The absence of any PBDDs or
PBDFs from pyrolysls of the anhydride Indicates that such compounds will be
safer flame retardants.
Chlorophenolates burned for 15 minutes on birch leaves or wood wool or
heated at 280°C for 30 minutes (Table 4-14) evolved PCDDs, but not many
PCDFs (Rappe et al., 1978b). Originally, both types of formulations studied
(Servarex Teknlsk and Kymmene KY-5, both containing 5X 2,4,6-trl-, 50X
2,3,4,6-tetra- and 10% pentachlorophenols as the sodium salts) contained 10
ppm each of tetra-, penta- and octa-CDFs and 70 ppm each of the hexa- and
hepta-CDFs, with 40-50 Isomers present (see Table 4-4). The burnt samples
produced only 10 Isomers and levels were lower than the original levels (see
Table 4-14). Nevertheless, the major TCDF (unknown) Increased 100-fold
during burning (from 0.04-5 ppm) and two others that were not 1n the
original formulation were detected. The 2,3,7,8-lsomer was a very minor
1928A 4-33 06/20/86
-------
VD
ro
CD
TABLE 4-14
Levels of Trapped PCDFs and PCDDs on Charcoal from Burning
Chlorophenolate-Impregnated Leaves and Wood Wool3
uQ PCDF
Commercial
Chlorophenol
£ 2,3,4,6-Tetra-
•*" (Servarex)
(Pure)
2,4,6-Trlchloro-
Pentachloro-
Substrate Tetra-
CDF COD
Birch leaves <10b 35
Wood wool <10b 96
<10b 30
Birch leaves — 2100
Birch leaves — 5
Penta-
CDF
<10b
<10b
<10b
—
—
COD
90
120
84
5
14
or PCDD/q Chlorophenate
Hexa-
CDF
<70b
<70b
<70b
--
—
COD
80
110
82
1
56
Hepta-
CDF
<70b
<70b
<70b
—
—
COD
8
65
8
3
172
Octa-
CDF COD
<10b 0.3
<10b 1.2
<10b 0.4
6
710
aSource: Rappe et al., 1978b
bS1gn1f1es the amount In the original formulation
ro
O
\
CO
-------
component 1n the starting material (0.1% of the total PCDFs). When the pure
phenols were mlcropyrolyzed, no PCDFs could be detected. Thus, the newly
formed PCDFs had to arise from Impurities 1n the technical products (for
example, PCBs or probably PCDPEs) (Llndahl et al., 1980). K12 formula-
tions were particularly contaminated by PCDF/PCDD contaminants, but KI and
Ky-5 types contained lower levels (Paaslverta et al., 1982). Polychlorl-
nated phenoxyblpheny dlols and polychlorlnated phenoxyphenols were also
detected. Ky-5 burnt 1n an open fire caused enhanced levels of all PCDDs/
PCDFs 1n the residual ash and 1n smoke vapor with disappearance of PCDEs
from the Ky-5 formulation. The PCDDs probably arose from the polychlorl-
nated phenoxyphenol and chlorophenol contaminants.
Samples of ash from two airtight wood burning stoves, one open fire-
place, and after outside open-air burning and from associated chimney ash
contained traces of PCDDs and PCDFs (total PCDD or PCDF never >15 ng/g).
PeCDFs to HpCDFs dominated. Unfortunately, the recovery of 1,2,3,4-TCDD
varied between 15 and 80% and of OCDD between 41 and 92%. The past history
of the wood was not documented fully (Clement et al.. 1985b).
The production of PCDDs and PCDFs from the 2-butoxyethyl ester of
2,4,5-T was studied by pyrolyzlng the ester for 0.6-1 second when the ester
(1 kg) was mixed with leaves or sawmill wood chips (Ahllng et al., 1977).
The PCDF levels usually exceeded PCDD levels (Table 4-15) but not between
100 and 625°C, even though PCDFs were more concentrated In the original
formulation (which also contained 47 mg/g 2,4,5-T, 230 vg/g of 2,4,5-tr1-
chlorophenol and 12 wg/g PCP 1n addition to the PCDFs and PCDDs cited In
Table 4-15). It was thought that the chlorophenols could not be the source
of the PCDDs or PCDFs but that a complex mechanism was Involved.
1928A 4-35 06/23/86
-------
10
INJ
oo
I
to
TABLE 4-15
Production of PCDDs and PCDFs after 0.6 to 1.00 Second Pyrolysls In Air
of 2-Butoxyethyl-2,4f5-Tr1chlorophenoxyacetate*
PCDF/PCDD Amounts(mg/kq)
Temperature
CO
25
100
500
625
675
800
Tetra-
CDF
0.13
1.0
0.4
0.2
8.0
2.1
CDD
0.018
2.5
2.2
0.4
2.3
0.5
Penta-
CDF
0.008
0.5
1.5
<0.3
7.1
3.0
CDD
0.007
<0.5
<0.5
<0.3
<0.5
<0.3
Hexa-
CDF
<0.005
<0.4
2.0
<0.3
2.8
3.3
CDD
<0.005
<0.5
1.3
<0.3
<0.5
<0.3
Hepta-
CDF
<0.005
2.3
<0.7
<0.3
<0.7
2.4
CDD
<0.005
<0.5
<0.5
<0.3
<0.5
<0.3
Octa-
CDF
<0.005
<0.4
<0.7
<0.3
<0.7
2.3
CDD
<0.005
<0.5
<0.5
<0.3
<0.5
<0.8
Total
CDF
0.36
4.6
5.3
1.4
19.3
13.1
CDD
0.040
4.5
5.0
1.6
4.3
1.9
'Source: Adapted from Ahllng et al.. 1977
CO
-------
Recently direct evidence has been obtained of the formation of PCOF and
PCDD from polyvlnyl chloride (PVC) (Marklund et al., 1986a), as previously
suggested by Ahllng et al. (1978). In laboratory conditions pyrolysls of
PVC results In the formation mainly of TCDF and HpCDF. The pattern of
Isomers appeared similar to those found 1n municipal and hazardous waste
Incinerators. The same Isomerlc pattern was also found 1n emissions from a
smoke generator burning hexachloroethane (Marklund et al., 1986a), and from
leaded gasoline containing 1,2-d1chloroethane (Marklund et al., 1986b).
4.4.2. Incineration of Municipal Waste.
4.4.2.1. PRESENCE OF PCDFs IN INCINERATOR EMISSIONS — Ol1e et al.
(1977) detected the presence of trace amounts of PCDDs and PCDFs In fly ash
and flue gas of some municipal Incinerators 1n The Netherlands. The
presence of PCODs and PCDFs In fly ash from a municipal and an Industrial
Incinerator was later confirmed by Buser and Bosshard (1978) and by EIceman
et al. (1979). Reports and reviews from many countries (Samuelson and
Llndskog, 1983; Ch1u et al., 1983; BallschmHer et al., 1983; Janssens et
al., 1982; Sheffield, 1985a,b) have since been published, confirming the
previous results. The average PCDF concentration for total stack effluents
1s -2.8 yg/m3. However, the terms of total levels Including those of
congeners may not be correlated to potential toxldty.
Buser and Bosshardt (1978) and Buser et al. (19786) suggested that the
presence of PCDFs In the fly ash of municipal Incinerators could be
explained by the pyrolysls of PCBs. Fly ash samples from an Industrial
heating facility at Aarau and a municipal Incinerator 1n Zurich, both In
Switzerland, were analyzed. PCDDs and PCDFs were found In both locations
with levels of 0.3 and 0.1 ppm for the Aarau and Zurich samples, respec-
tively, and were 0.33-0.50 the total PCDDs present. The levels of PCBs were
1928A 4-37 06/23/86
-------
the same as for PCDFs. PolychloMnated naphthalenes were also present 1n
smaller quantities. The PCDF congener profile from fly ash was very similar
to that from pyrolyzed commercial PCBs, suggestive of a common source.
PCDFs were detected as follows: 18 tr1-, 21 tetra- (mainly the 2,3.7,8-),
17 penta- (Including the acutely toxic 1,2,3,7,8- and 2,3,4,6,8-), 7 hexa-
and 4 hepta- (1,2,3,4,6,7,8- most abundant). Fly ash samples contained
large amounts of 2,3,4,6,8- and 1,2,3,4,8-penta- and other penta-lsomers
relative to pyrolyzed Aroclor.
Ol1e et al. (1977) also detected large amounts of chlorobenzenes and
chlorophenols 1n the fly ash of municipal Incinerators In The Netherlands,
but provided no quantitative data. The Dow Chemical Company has suggested
that PCDDs are ubiquitous products of the combustion of all chlorinated
material, Including that found In municipal Incinerators (Bumb et al., 1980;
Crummett and Townsend, 1984).
A summary of the large amount of work done since the Initial findings Is
provided 1n Tables 4-16 and 4-17 for Incinerator fly ash and stack effluent,
respectively. Lustenhouwer et al. (1980) postulated that polyvlnyl chloride
pyrolysls produced chlorobenzenes In small amounts, and these could further
pyrolyze to form PCDDs and PCDFs. Olle et al. (1983) found that PCDDs and
PCDFs did not Increase when hexachlorobenzene, wood treated with penta-
chlorophenol, paper treated with hypochlorHe, old painted wood or Hgnln
sulfonate/polyv1nylchlor1de were burnt. However, Hgnln sulfonate/HCl
burning caused a marked Increase. Nevertheless, L1bert1 and Brocco (1982)
showed that decreased levels of PCDDs/PCDFs occurred for wastes pretreated
by the Dano process or 1f paper, plastic and vegetable matter were removed.
Table 4-18 depicts the Influence of type of waste on PCDD/PCDF content.
1928A 4-38 06/20/86
-------
TABLE 4-16
vO
CO
1
to
VO
o
cr
l\3
O
CO
PCDF Composition of Municipal Incinerator Fly Ash Resulting from
Origin of Fly Asha
Arnhem, The Netherlands (1)
Alkmaar, The Netherlands (1)
Unknown, Switzerland (1)
Ontario Canada (1 )
Japan (1)
Japan (1)
The Netherlands (1)
Unknown (25)
One Incinerator over 4 weeks
Milan, Italy (1)
Milan, Italy (1)
Busto. Italy (1)
Deslo, Italy (1)
Unknown (1)
Unknown (2)
Bologne, Italy (1)
Milan, Italy (1)
Alkmaar, The Netherlands
(17 samples over 5 months)
Amsterdam, The Netherlands
(14 samples over 18
months)
Zaanstad, The Netherlands
(24 samples over 20
months)
(8 samples every 20
minutes over 2.5 hours)
West Germany (1)
Recovery Cl3
(X)
NS
NS
NS
100
100
NS
NS
NS
NS
100
100
NM
80b
NS
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
PCDF
C14
trace
trace
1
3272
789
ND
ND
13-223
50-147
NM
NM
NM
NM
169
223-315
112
51
76-410
12-184
20-471
118-291
0.8-28
(nq/q)
C15
trace
trace
4
5697
3691
3827
ND
42-510
153-239
NM
NM
NM
NM
364
510-560
205
115
165-364
29-122
31-792
222-568
0.3-8
Cl6
trace
trace
30
1415
1069
ND
ND
109-870
282-361
NM
NM
NM
NM
555
840-870
235
177
160-1260
29-287
82-1054
566-1310
0.4-8
Electrostatic Precipitation
C17
trace
trace
40
967
2389
2555
ND
89-407
177-291
NM
NM
NM
NM
287
407-495
345
310
108-300
71-89
47-714
397-699
0.6-10
C18
trace
ND
10
ND
1120
2341
ND
20-26
18-31
166
6-22
0-18
70
32
84-89
425
547
0-126
2-97
4-223
37-145
0.3-14
Reference
Olle et al., 1977
Buser et al., 1978d
Elceman et al., 1979
Lustenhouwer et al., 1980
Cavallaro et al., 1980
Kooke et al., 1981
Llbertl and Brocco, 1982
Olle et al., 1982
Karasek and Vlau. 1983
(rotary
samples)
-------
00
TABLE 4-16 (cent.)
Origin of Fly Asha
USA (1)
Florence, Italy (1)
(3 laboratories)
U.S. cyclone ash (1)
U.S. fly ash (1)
Ontario. Canada (1)
U.S. (1) (2 samples)
Vienna, Austria (1)
Ehlme, Japan
8 cities
cinders
Ontario, Canada
USA
PCDF (nq/q)
Recovery
(X)
50-100
NS
75
75
90
NS
NS
50
50
NS
NS
..,
NM
NM
NM
NM
NM
520-580
NM
0.7-32
0.4-14
NM
NN
C14
419
3.2
50
50
294
350-460
(Cl! =
39
0.4-31
0.1-84
965
26-150
C15
107
22
75
100
508
1860
40-42; C12 =
60
ND-180
trace
to 12
1865
52-390
...
263
35
170
400
420
75-91
69-110)
154
trace
to 56
ND-25
1990
120-880
C17
287
9-51
80
300
428
9-11
156
0.1-11
ND-0.8
850
72-380
Reference
25
10-230
5
20
101
0.57-2
168
ND-23
ND-0.9
ND
8-31
Taylor et al.
Brocco et al.
. 1983
, 1984
Czuczwa and Hltes, 1984
Tong et al .,
Halle et al..
Scheldl et al
Waklmoto and
1985
Shushan et al
Clement et al
1984
1985
.. 1985
Tatsukawa,
.. 1985
., 1985a
aNumber In parenthesis
>>1,2.3.4-TCDD
NS = Not stated
NM = Not measured
ND = Not detected
00
-------
CO
TABLE 4-17
PCDF Composition of Stack Effluent of Municipal Incinerators
o
CO
Origin of Study
Emission/Number
Unknown, Switzerland
Unknown
Milan, Italy (5 samples)
Milan. Italy (4 samples)
Busto, Italy
Deslo, Italy
Valmadrera, Italy (16
samples over 9 months)
Florence, Italy
Zaanstad, The Netherlands
(14 samples over 6 months)
Unknown, U.S.A.
Ontario, Canada (1)
(2)
Eksjo. Sweden
(? samples)
Florence, Italy
(3 samples)
Recovery
(X)
NS
100a
NSa
c/
NSa
£/
NSa
£/
NSa
£/
85d«e
NSd
£/
80d
81d
NSa
c/
NSa
£/
NSa
£/
NSa
c/
«i,
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
300
NM
NM
NM
NM
NM
NM
NM
NM
PCDF
C14
trace
460b
NM
NM
NM
NM
NM
NM
NMN
NM
17-2846
309 f
175b
ND
46-556
90
46
8
279
5
25-37
68-525
NM
NM
(nq/ma)
Cl5
trace
960b
NM
NM
NM
NM
NM
NM
NM
NM
17-2261
250f
240b
ND
109-702
NM
153
15
265
5
4.3-10
5.7-87
NM
NM
«u
trace
1600b
NM
NM
NM
NM
NM
NM
NM
NM
22-2928
314f
185b
125
174-1437
62
1712
ND
392
ND
4.3-25
10-80
NM
NM
C17
NM
1130b
NM
NM
NM
NM
NM
NM
NM
NM
17-1414
215f
400b
105
155-504
7.5
47
ND
104
ND
8.3-25
7.3-15
NM
NM
Reference
C18
NM Buser et al., 1978c
140b Lustenhouwer et al., 1980
20-47b Cavallaro et al.. 1980
270-1080
0.9b
90
4-70b
5-10
10b
51
17-749 Glzzl et al., 1982
124^
570b Llbertl and Brocco, 1982
31
0-201 Olle et al., 1982
0.8 Redford et al.. 1983
ND Chlu et al.. 1983
ND
ND
ND
8.3-9 Rappe et al.. 1983a
1.3-5.7
168-498 Brocco et al., 1984
411-546
-------
TABLE 4-17 (cont.)
r\>
00
PCOF (nq/m3)
Origin of Study
Emission/Number Recovery Cl3
Germany (1) NSa
c/
a/
c/
a/
c/
a/
c/
a/
c/
a/
£/
£ U.S. (1) (5 days) d/
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
1100-3300
CU
2h
98h
3^
98h
18^
82h
59h
4]h
9h
9]h
2^
98»>
480-2000
S (Cl^ = 300-420; C12 =
Vienna. Austria (1) d/
Quebec, Canada (3) d/
Ontario, Canada (2) d/
NH
NM
NM
100
ND-142
53-4850
C15
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
1300-15,000
400-700)
120
ND-124
64-3960
Cl6
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
170-1800
150
ND-1392
152-2120
C17
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
NM
83-380
110
ND-123
20-880
Reference
"8
7h Ballschmlter et al., 1984,
93h 1985
63h
37h
81^
19h
67h
33^
100h
<1 h
3^
97h
8-24 Halle et al.. 1985
30 Scheldl et al., 1985
ND-3 Sheffield, 1985a,b
10-170
PO
CO
3Part1culate
bng/g
C51gn1f1es "condensate"
^Combined a and c
e2,3,7,8-TCOD
fArithmetic means
91,2,3,4-TCDD
nln % of Homeric class
NS = Not stated; NM = not measured; ND = not detected
-------
00
TABLE 4-18
Influence of Type of Waste on PCDF Formation
PCDF (nq/q)
Waste
Untreated
Urban Waste
Agricultural
Waste
Recycled waste3
Unfermented waste
surviving the
Dano process
Coal-fired power
plant, U.S.A.
Coal-fired power
plant, Canada
Low temperature
Incinerator
Pentachlorophenol
Impregnated waste
Plasma Pyrolysls of
Substrate
bottom ash
fly ash
fly ash
sludge
sludge
partlculate
stack emission
sludge
stack emission
fly ash
fly ash
bottom ash
partlculate
condensate
condensate
"3
NM
NM
NM
NM
NM
NM
NM
ND
NM
NM
NM
NM
NM
C14
NM
NM
NM
NM
NM
NM
NM
ND
8
0.3
ND
ND
0.9
"5
NM
NM
NM
NM
NM
NM
NM
ND
32
1.1
ND
ND
4.8
"6
NM
NM
NM
NM
NM
NM
NM
ND
18
0.4
ND
ND
9.1
C17
25
70
ND
ND
ND
ND
ND
ND
15
0.1
ND
ND
18
C18
50
50
ND
ND
ND
ND
ND
ND
ND
0.1
25
ND
18
Reference
Llbertl and
Brocco, 1982
Halle et al.,
1983
Redford
et al., 1983
Chlu et al..
1983
CD
Aroclor 1254
-------
TABLE 4-18 (cont.)
ro
CO
.
rss
CO
Waste
Pentachlorophenol
Contaminated waste
PCB Incinerator
rotary cement
Low sulfur coal
Hazardous waste
Municipal waste
Peat 1
2
Coal (Canada)
In
kiln
1
2
3
1
2
3
Substrate
bottom ash
baghouse ash
partlculate
condensate
fly ash
stack emission
stack emission
stack emission
stack emission
stack emission
stack emission
stack emission
stack emission
fly ash
"3
NM
NM
NH
NH
NH
NH
NH
NH
NH
NH
NH
NM
NM
NM
NM
C14
ND
900
ND
ND
ND
ND
22b
62b
<0.4b
4.5b
4b
21b
<0.0006b
0.15b
ND-0.9
PCDF
"5
ND
1500
ND
ND
ND
ND
NM
NH
NM
NM
NM
NH
NM
NM
ND-
2.8
(nq/q)
cis
ND
150
ND
ND
2
0.1
NM
NM
NH
NH
NH
NH
NH
NH
ND-
0.25
C17
ND
60
ND
ND
4.2
0.2
NH
NH
NH
NM
NM
NH
NH
NH
ND-
0.19
C18
ND
6
ND
ND
0.1
ND
NH
NH
NH
NH
NH
NH
NH
NH
ND-
0.13
Reference
Rappe et al.,
1983a
Czuczwa and Kites,
1984
Oberg and Bergstrom,
1985
Sheffield, 1985a,b
aW1th paper, plastics and vegetable matter removed
bng/m3
NH = Not measured
ND -- Not detected
-------
Thus, no PCOFs/PCDDs were found 1n stack emissions, fly ash or coal at four
coal-fired power plants (Halle et al.. 1983; Redford et al., 1983). Some
trace PCDDs/PCDFs were found In the stack emissions when municipal waste was
burnt. The results are probably caused by optimum combustion conditions,
1200°C for the coal-fired plant and 650°C with a 20-m1nute residence time
for the municipal waste. Levels up to a few pg/m3 were reported from a
large peat Incinerator 1n Sweden (Marklund et al., 1986a).
There Is still controversy over whether the vapor or partlculate phase
contains the most PCDDs/PCDFs 1n the stack emissions of municipal Inciner-
ators (see Table 4-17). The vapor generally appears the preferred locali-
zation phase (Cavallaro et al., 1980; Ballschmlter et al., 1984, 1985;
Benfenatl et al., 1986). A 13C12-labeled surrogate added to the filter
before sampling was found preferentially 1n the condensate and adsorbent
(Rappe et al., 1986). Combustion In the municipal Incinerators sampled
occurred 1n the temperature range 500-1000°C with an optimum formation
temperature of ~500°C (Glzzl et al., 1982).
Higher temperatures (>1400°C) may solve the PCDF residue problem as
Indicated by a study of the PCB, PCDF and PCDD content of the residual ash
of a cement kiln. Residual PCB of -10 mg/kg was detected for a chamber
pyrolysls time of 5 seconds at 700-1000°C (AhUng and Llndskog, 1978), but
this was drastically lowered at 1400-1450°C (AhUng, 1979). No PCBs were
found In the flue gases, and no PCDDs or PCDFs found 1n the kiln residues,
although peaks corresponding to the expected retention time for OCDF were
noted. Municipal Incinerators operating from 320-900°C favored PCDF forma-
tion over PCDD {Benfenatl et al., 1983; Taylor et al., 1983). This 1s so
for all Isomer classes except for hepta- and octa- (Taylor et al., 1983).
1928A 4-45 06/23/86
-------
The highest emissions also appeared to be related to the presence of HC1,
but not with participate matter (Benfenatl et al., 1983). The degree of
temperature control appears to be the dominating factor (r=0.839) and not
absolute efficiency of combustion/precipitation since the lowest temperature
reached Is the most highly correlated parameter for PCDF/PCDD formation for
a given waste {Benfenatl et al., 1983). Ol1e et al. (1982) recognized that
the presence of 2,3,7,8-TCDD could not alone explain the toxldty of fly
ash. They have calculated an Index called the "2,3,7,8-TCDD toxic equiva-
lents" to describe the toxldty based on the presence of toxic 2,3,7,8-sub-
stltuted Isomers (Table 4-19). Marklund et al. (1986a) reported the toxic
TCDD equivalents (Eadon, 1982) based on the concentrations of the toxic
2,3,7,8-substHuted PCDFs and PCDDs found 1n the stack emissions of a
Swedish municipal Incinerator (Table 4-20).
4.4.2.2. ORIGIN OF PCDF IN INCINERATOR EMISSIONS — The origin of
PCDF and PCDD In Incinerator emissions has not been clarified. The hypothe-
sis that the contaminants are already present 1n waste and are not com-
pletely transformed during combustion Is generally not accepted. However,
Ozvaclc et al. (1984) found 1n raw waste an average concentration of 2-3
ng/g PCDF and 19.8 ng/g PCDD to 4 through 8 Cl-subst1tuted. Although con-
centrations of the refuse are low, Toslne et al. (1985) suggest that levels
of PCDD and PCDF In feedstock should not be Ignored 1n studies designed to
determine mechanisms of formation of these substances.
A second hypothesis suggests PCDF and PCDD formation from precursors
such as PCB, chlorophenols and chlorobenzenes present In the raw refuse.
Laboratory studies have shown that pyrolysls of a variety of chlorinated
chemicals can lead to the occurrence of PCDF and PCDD (see Section 4.4.1.).
1928A 4-46 06/20/86
-------
TABLE 4-19
Annual Toxic Equivalent of CDBFs Emitted by Netherlands
Incinerators In Terms of 2,3,7,8-TCDD Toxic Equivalents*
PCDF Isomers
Parameter
Substrate (kg)
Fly ash amount/year
toxic equivalent
Stack effluent amount/year
toxic equivalent
C14
10.4
0.2
2.3
0.048
"5
18.7
0.9
3.8
0.19
"6
27.5
2.1
7.4
0.56
"7
18.8
0.9
4.1
0.21
2,3,7,8-
TCDD
0.2
0.026
'Source: Ol1e et al., 1982
1928A
4^47
06/20/86
-------
ro
CO
TABLE 4-20
Levels of PCDO and PCDF from HSU Incineration, Umea*
(ng/Nm3 dg 10% C02)
-^
1
CO
o
cr>
\
o
\
CO
?n
Experiment
Season
Number of
experiments
2,3,7,8-TCDF
Total TCOF
2,3,7,8-TCDO
Total TCDD
1,2,3,7,8-7
1,2,3,4,8-PeCOF
2,3,4,7,8-PeCDF
Total PeCOF
1,2,3,7,8-PeCDO
Total PeCDD
1,2,3,4,7,8-7
1,2,3,4,7,9-HxCDF
1,2,3,6,7,8-HxCDF
Normal
Fall
3
2.5
86
0.5
43
9.0
6.1
97
2.5
53
3.6
3.7
Normal
Chips
Fall
2
2.3
75
0.6
45
8.3
7.3
100
3.6
70
4.6
4.6
Normal
Oil
Fall
2
2.4
68
0.7
52
9.8
7.8
120
3.6
76
5.6
5.5
Low
Temperature
Fall
3
2.6
87
0.4
54
8.3
7.4
110
3.2
80
5.2
5.0
Low
Temperature
Oil
Fall
3
2.1
75
0.3
47
7.1
6.5
87
3.6
70
3.6
3.4
Start
Fall
1
9.5
260
1.3
100
52
40
520
14
280
48
40
Start
Oil
Fall
1
2.3
80
0.7
49
9.0
9.0
120
3.9
90
5.7
5.7
Normal
Spring
3
0.85
19
<0.1
<10
2.5
3.9
43
2.4
49
4.5
4.6
-------
kO
CO
3>
TABLE 4-20 (cont.)
1,2,3,7,8,9-HxCDF
2,3,4,6,7,8-HxCDF
Total HxCDF
1,2,3,4,7,8-HxCDD
1,2,3,7,8,9-HxCDD
1,2,3,7,8,9-HxCDD
£ Total HxCDD
* Total HpCDF
Total NpCDD
OCDF
OCDD
TCDD equivalents
0.8
2.6
33
1.6
3.7
1.3
32
34
18
10
12
9.1
0.8
4.4
46
2.9
5.6
2.4
53
73
29
52
19
10
1.2
4.3
51
3.5
6.5
3.0
72
94
37
50
31
11
1.2
5.1
50
5.1
9.5
3.8
82
67
54
23
14
11
1.4
4.2
37
3.1
6.6
2.6
57
40
36
25
20
10
52
36
380
31
56
20
400
380
380
180
130
53
2.3
5.4
54
4.4
7.9
3.5
70
51
38
41
27
12
3.6
4.3
43
2.3
5.8
2.0
55
49
56
33
52
5.6
*Source: Marklund et al., 1986a
IV)
o
00
-------
However, In experiments 1n which known precursors were either left out or
added to Incinerator material, results were contrasting. Olle et al.
(1983), comparing the emission from burned PCP-treated wood and 60-year-old
painted untreated wood, detected no significant difference In PCDF (0.2
vg/g) and In PCDO formed. Hexachlorobenzene added 1n the proportion of
400 g to 2 m3 municipal raw waste raised PCDF and PCDD emission (Ol1e et
al., 1983). Chlu et al. (1983) burned PCB-treated wooden boxes and
discovered PCOD congeners In the range of 0.4 yg/g and only OCDF at 0.025
yg/g. On burning pine wood with HC1, Tlernan et al. (1983) detected total
PCDF (3 yg/g) that was 20 times more abundant than PCDD, while no PCDF
formation was found on the combustion of untreated wood. Conversely,
NestMck and Lamparskl (1982) detected PCDF and PCDD 1n soot and ash of
Incinerated untreated wood. Therefore, a third mechanism has been postu-
lated 1n which PCDF and PCDD are formed (te novo from nonchlorlnated organic
compounds 1n the presence of chlorine donors (Choudhry et al., 1982).
Marklund et al. (1986b) have discussed the striking similarities 1n congener
profiles of PCDFs and PCDDs found 1n emissions from municipal Incinerators
and pyrolysls of PVC, dlchloroethane and hexachloroethane. They postulate a
common mode of formation.
4.4.3. Sewage Sludge Combustion. Few data have been published on the
production of PCDF from combustion of sewage sludges. However, sludges have
been found to contain much synthetic chlorinated compounds so they could be
sources of PCDF contamination. Lamparskl et al. (1984) has reported PCDD In
dried sewage sludge samples. Surprisingly small differences have been found
between samples collected In 1981 and 1982 and dried samples collected In
1933 and sealed In glass until analysis. More recently, Weeraslnghe et al.
(1985a) analyzed samples of sewage sludge from two towns for PCDD and PCDF.
1928A 4-50 06/23/86
-------
They detected several congeners of PCDD at both and, except for OCDF (1300
ng/g In one sample), no congeners of PCDF.
4.4.4. Electrical Equipment Fires. PCBs are used as dielectric fluid In
two types of equipment: transformers containing mixtures of PCS and chloro-
benzenes (PCBZ) and capacitors containing only PCB. Accidental fires
Involving transformers have been reported to produce PCDF and PCOD; when
capacitors were Involved only PCDF were formed.
A summary of PCDFs found after PCB fires (Rappe, 1983a,b,c, 1985a,b,c;
M1lby et al., 1985; EMckson et al., 1985; Williams et al., 1985; Hutzlnger
et al., 1985c) Is provided In Table 4-21. The toxic 2,3,7,8-TCDF consti-
tutes -20-25% of the TCDF content, except for the Blnghamton fire where H
constituted 50% (Rappe et al., 1983a). The Blnghamton fire concerned a
dielectric fluid that contained 65% PCB and 35% chlorinated benzenes. This
may explain why the Blnghamton data are anomalous. The 2,3,4,7,8-PeCDF and
1,2,3,7,8/1,2,3,4,8-PeCDF were 18 and 9% of the total PeCDF, respectively
(Rappe et al., 1983a). In Blnghamton soot the 1,2,3,7,8- and 2,3,4,7,8-
PeCDF, and 1,2,3,4,7,8-, 1,2,3,6,7,8- and 2,3,4,6,7,8-HxCDF were 46, 7.2,
32, 16 and 10%, respectively, of their Isomerlc class.
4.4.5. Incineration of Hazardous Waste. PCDFs have been reported In
emission of hazardous waste Incinerators (Tlernan et al., 1983; Rappe et
al., 1983a). A Swedish waste Incinerator was monitored weekly for PCDF and
PCDD emissions that were strongly correlated with the levels of total
chlorine Input (Oberg et al., 1984). Table 4-18 contains other examples of
PCDFs produced during the Incineration of hazardous wastes.
1928A 4-51 06/23/86
-------
00
TABLE 4-21
PCDFs Formed During PCB Fires
PCDF Congener9
_^
i
en
PO
o
c*
CO
00
c*
Site
Stockholm. Sweden.
1978
Stockholm. Sweden.
1981
Blnghamton. NY. 1981
Surahammar. Sweden,
1982
Imatra, Finland, 1982
Hallstahammar.
Sweden. 1982
Railway locomotive.
Sweden. 1982
Klsa. Sweden. 1983
Sample
Liquid on capacitor
PCB after explosion
Floor wipe
Floor soot
Celling soot
Floor soot
Air
Air
Room floor
Wipe 1
Nlpe 2
Mall wipe 1
Mall wipe 2
Soot 1
Soot 2
Soot 3
Metal bar
Mlpe 1
Wipe 2
Floor wipe
Floor wipe 1
Floor wipe 2
Floor wipe 1
Floor wipe 2
"3
3-7b
lib
NN
NN
NN
NN
NN
NN
NN
NN
NN
NN
NN
NN
200
NN
NN
NN
NN
NN
NN
NM
C14
2b
3°
1600
0.028
162
100
4-195b
150°
4000
90
1250
480
1
16
20
1600
5000
50
6900
4900
100.000
500
Cl5
NM
NN
175
0.670
197
120
N0-60b
50b
3300
25
355
210
0.2
1.0
1
360
1800
12
29.000
1500
NM
NM
C16
NN
NN
<0.5
0.965
110
70
2-9b
2-3b
1800
17
150
140
0.04
0.3
NN
600
850
10
14.000
400
NN
NN
C17
NN
NN
<1
0.460
36
20
NN
NN
1500
17
65
60
0.02
0.2
NN
800
550
30
6200
170
NN
NN
C18
NH
NM
<1
0.040
14
4
NM
NM
300
4
13
30
0.01
0.1
NH
1340
440
20
140
40
NM
NM
Reference
Jansson and Sundstrom.
1982
Rappe et al.. 1982,
1983a, 1985b
Rappe et al., 1983a
O'Keefe et al.. 1985a
Schecter and Tlernan,
1985
Rappe et al., 1985b.c
Rappe et al.. 1985b
Elo et al.. 1985
Rappe et al., 1985b,c
Rappe et al.. 1985b.c
Rappe et al.. 1985b
-------
vO
rsj
00
TABLE 4-21 (cont.)
PCDF Congener*
Site Sample
^
i
os
Skovde. Sweden. 1982 Floor wipe
Floor wipe
Hall wipe
Bench wipe
London. U.K.. 1983 Overheated
Fresh PCB
1
2
PCB
Turin. Italy Concrete floor wipe
"3
NN
NN
NN
NN
NN
NN
NN
C14
100
600
<1
<1
50-90
ND-70
3852
Cl5
40
100
<1
10
460
150
296
"6
40
60
<1
<1
530-590
130-160
<9.5
C17
a
8
Not detected
INJ
CO
-------
4.4.6. Car Exhaust. BallschmHer et al. (1986) Identified a series of
PCDFs and PCDDs 1n used motor oil from automobiles. The congener profile
was similar to that from Incinerators burning urban waste. It was suggested
that chlorinated additives In the oil were Involved. Marklund et al.
(1986b) found that the appearance of PCDFs/PCDDs depended on the presence of
l,2-d1chloroethane scavenger. Contaminants added to the motor oil were not
responsible. The average emission from a car running on leaded gasoline was
30-540 pg 2,3,7,8-TCOD equivalents/km, equivalent to 10-100 g 2,3.7,8-TCDD
equivalents/year.
4.4.7. Other Sources. PCDFs have also been found 1n the emissions from
steel mills and copper mills, baths using recycled material contaminated by
PVC and polychlorlnated paraffins {Marklund et al., 1986a). Bergman et al.
(1984) also reported on the formation of PCDFs from chlorinated paraffins.
PCDFs have been mentioned 1n several patents for use In electrical Insula-
tion (British Thompson-Houston Co. Ltd., 1939), In capacitors (Clark, 1940),
flame retardants (Dazzl et al., 1974; Boyer, 1971) and as bacterlddes
(Shlbata et al., 1952). Commercial usage of the products Is not available.
1928A 4-54 06/20/86
-------
5. ENVIRONMENTAL FATE, TRANSPORT AND DISTRIBUTION
5.1. SUMMARY
PCDFs are ubiquitous contaminants. They are associated with the small
partlculate fraction and vapor of flue gas and can thus be transported In
the atmosphere over very wide areas. Weather conditions can greatly affect
their transport. Photodegradatlon by sunlight may occur 1n the presence of
triplet sensltlzers 1n aqueous solution; photodecomposltlon In the vapor
phase has not been Investigated.
In aquatic media, since PCDFs have poor solubility and tend to be
adsorbed, they are expected to be persistent In sediments or on partlculate
matter. Their presence has been associated with atmospherically transported
waste Incineration-derived PCDF. In soil they have been detected In top
soil layers after Improper disposal of wastes or after dissemination of
herbicides, but quantitative data are lacking. As for translocatlon poten-
tial from soil to vegetation 1s concerned, PCDFs tend to be adsorbed on
external surfaces. Little data on bloconcentratlon factors exist.
5.2. INTRODUCTION
Since PCDFs are trace Impurities In various chlorinated chemicals (see
Section 4), they will accompany these chlorinated chemicals when they are
released Into the environment. As concentrations are generally below ppm
levels, the quantity released determines the extent of the pollution. The
content of toxic Isomers also determines whether toxic effects may occur.
PCDFs have also been Identified 1n effluents from various combustion
processes (Section 4). Municipal solid wastes and Industrial waste Inciner-
ations have been found to emit fly ash and flue gas containing PCDF In the
range of yg/m3. As PCDF may be associated with the small partlculates
1929A 5-1 06/23/86
-------
and wHh flue gas, PCDFs could be distributed over large areas. This com-
bustion may be responsible for the ubiquitous PCDF pollution of the environ-
ment.
Very little data for the PCDFs are available, so they are expected to
act like PCDDs 1n the environment. The U.S. EPA (1985) gives an excellent
summary of the environmental fate, transport and distribution of 2,3,7,8-
TCDD.
5.3. ENVIRONMENTAL TRANSPORT
5.3.1. Air.
It 1s very difficult to assess the quantity and the IsomeMc distribu-
tion of PCDFs In the atmosphere as many variables Interfere. Emission from
Incinerators Is dependent on the refuse, combustion design, operating condi-
tions, composition of the fuel, operating temperatures and degree of emis-
sion control. The great heights of stacks contribute to disperse PCDF over
large areas and reduce concentrations near the source. Weather conditions,
height of the stack 1n relation to the size and position of natural obstruc-
tions or buildings, relative distribution of PCDFs 1n flue gas between par-
tlculate fraction and particle-free gas phase (see 4.4.2.1.), and particle
size are all factors that greatly modify the transport of PCDF In the atmos-
phere, as has been suggested for the other pollutants (Somers, 1971).
PCDFs may theoretically undergo photodecomposltlon In the atmosphere,
but the environmental relevance of this process Is not known. Photochemical
degradation has been demonstrated under laboratory conditions for the high-
est chlorinated congeners, but these reactions are unlikely to occur In sun-
light (Section 4.3.). Thus PCDFs are probably carried directly to the soil
or to water compartments such as lakes and oceans.
1929A 5-2 06/23/86
-------
A method to collect PCDFs and PCDDs In both the partlculate and vapor
phase 1s now available (Oehme et al., 1986). This method Involves a fiber
glass filter and a polyurethane foam plug from which the PCDFs were desorbed
by Soxhlet extraction with toluene, followed by liquid chromatographlc
separation, and GC/MS confirmation and quantltatlon. The recovery from the
filter 1s -75-8554 for 13Cno-l,2,3,4-TCDD, 37C1B-OCDD, and
\c o
13C,2-PeCDF; the recoveries from the foam are >80%. Typically, the PCDF
congeners below HxCDF were found on the foam plug following the filter,
while the rest were found adsorbed to particles on the filter. As soot
loading Increased, more of the lower congeners were found on the filter
adsorbed to soot particles. Samples near an Industrial city (Essen, Federal
Republic of Germany) reflected the congener patterns of nearby municipal
Incinerator stack emission patterns. Concentrations decreased with distance
away from the source. Samples 1n suburbia and In remote areas contained
mostly PCDFs. Of the 60 samples taken, the highest levels of 5-10 pg PCDDs
fPCDFs/m3 were found In a heavily Industrialized area. In suburban areas
the levels were 5-10 times lower. Thus people living near Industrial areas
may be exposed. The highest 2,3,7,8-TCDD equivalent level found was 0.1
pg/m3 with 2,3,7,8-TCDD always below detection. Ballschmlter et al.
(1986) have shown that PCDFs can be emitted from leaded automobile exhaust
but not from unleaded exhaust (Chapter 4 also has a discussion of this topic
from the perspective of accumulation of PCDDs and PCDFs In leaded gasoline
through use. The 1,2,-dlchloroethane additive was postulated to be the
chlorine donor). Ballschmlter et al. (1986) showed through pattern recogni-
tion that both automobile exhaust and emissions from municipal Incinerators
were correlated with lake sediments around the world. The municipal Incin-
erator component was postulated to arise mostly from pentachlorophenol
1929A 5-3 06/23/86
-------
Incineration (Cruczwa and Hltes, 1984, 1985, 1986; Cruczwa et al., 1984,
1985). The striking uniformity of the patterns of Isomers 1n a given
congener class suggested a more global source of PCDFs/PCDDs like automobile
exhaust. Three basic emission sources were postulated (Ballschmlter et al.,
1985) based on congener profiles.
5.3.2. Hater.
PCOFs are UpophlUc and practically Insoluble 1n water (Section
2.3.3.). Thus 1n aquatic media PCDF are present 1n the adsorbed state to
partlculate matter rich 1n organic content. Rappe et al. (1984) found con-
centrations (100 ng/ml) of PCDF 1n a suspension of soot/dust In wash water
from a PCB fire to be only 1n the adsorbed state, since when the soot
settled the water did not contain detectable PCDF. Abiotic degradation
(photoreactlons, hydrolysis, etc.) and blodegradatlon are not likely to
occur In this compartment. Thus PCDFs are expected to be very persistent 1n
aquatic media where they show high affinity for sediments. Czuczwa et al.
(1984), Czuczwa and Hltes (1984) and Czuczwa et al. (1985) found that fluxes
of PCDF to lake sediments In the United States and Switzerland were -0.1-0.3
ng/cnf Vyear"1.
5.3.3. Soil.
PCDFs are rapidly and strongly adsorbed Into most soils and sediments
where they are expected to be Immobile. Quantitative data on PCDFs are
lacking, but studies on 2,3,7,8-TCDD Indicate that these compounds are prac-
tically Immobile (Helling et al., 1973), the apparent half-life for TCDD 1n
soil exceeding 10 years (D1 Domenlco et al., 1980a). PCDF can be transport-
ed from soil to air or the water compartment through contaminated airborne
dust particles or waterborne eroded soil.
1929A 5-4 06/23/86
-------
Volatilization 1s probably not Important because of the low vapor pres-
sures of such compounds. However, D1 Domenlco et al., (1980b) suggested
that volatilization could be at least partially responsible for the observed
disappearance of TCOD from the topmost soil layers of the polluted Seveso
area, 1 year after the accident. Freeman and Schroy (1985) modeled the
volatilization of 2,3,7,8-TCOD adsorbed to Times Beach soil based on moni-
toring the volatilization over a period of 1 year. Thus H Is likely that
volatilization will occur also for the PCDFs, and this should be demon-
strated.
In general, higher chlorinated PCDDs and PCDFs volatilize more slowly
than lower chlorinated dloxlns since their vapor pressures decrease with
Increasing chlorlnatlon (Table 2-5).
5.4. ENVIRONMENTAL TRANSFORMATION
5.4.1. Abiotic Transformation.
PCDFs are chemically quite stable and are not likely to be degraded by
hydrolytlc reactions under environmental conditions (Section 2.4.1.).
Photochemical degradation was discussed In Section 2.4.2. Photodegrada-
tlon In organic solvents occurs for the higher PCDFs 1n organic solvents at
310 nm (Hutzlnger et al., 1972a,b, 1973; Buser, 1976b; Firestone, 1977b;
Mazer and Hlleman, 1982; Mazer et al., 1983a,b,; Hlleman et al., 1985). The
degradation 1s much slower In water at 310 nm (Crosby and Mollanen, 1973).
Direct evidence of PCDF decomposition In the environment Is lacking and
the Importance of sunlight In their environmental fate remains to be
evaluated.
5.4.2. B1otransformat1on.
Currently virtually nothing Is known about the blodegradatlon of PCDFs.
However H 1s conceivable that they are relatively resistant to blodegrada-
1929A 5-5 06/24/86
-------
tlon, like the corresponding 2,3,7,8-TCDD (U.S. EPA, 1985). Only 5 of the
100 soil microblal strains tested have shown some ability to metabolize
2,3,7,8-TCDD (Matsumura and Benezet, 1973). Ward and Matsumura (1978)
studied the blodegradatlon of 2,3,7,8-TCDD 1n Wisconsin lake waters and
sediments and found a half-life of 550-590 days In sediments, while about
70X of 2,3,7,8-TCDD 1n lake water samples alone remained unaltered after 590
days. Moreover, Matsumura et al. (1983) estimated the half-life of
2,3,7,8-TCDD 1n an aquatic ecosystem model to be ~1 year. The occurrence of
a polar metabolite of 2,3,7,8-TCDD with a probable hydroxy group 1n position
1 was found by PhlUppI et al. (1982) In several microbiological cultures
after long Incubation. Bumpus et al. (1985) demonstrated that
14C-2,3,7,8-TCDD at a level of 14 pmol was blodegraded to the extent of
~25% by the white rot fungus, Phanerochaete chrysosporlum. It was hypothe-
sized that the extracellular I1gn1n-degrad1ng enzyme system was responsible.
14CO_ was the metabolic product monitored. PCDFs are expected to cause
the same behavior.
5.5. BIOACCUMULATION
High correlations have been observed between the octanol/water partition
coefficient (K ) of many organic compounds and bloaccumulatlon In fish.
Two equations have been proposed to relate the two measures. Lu et al.
(1978) found that their data were fitted by the following equation:
log [bloaccumulatlon (fish)] = 0.7 log (partition coefficient) + 0.8
Therefore, high K values would Indicate high concentration factors.
K values were reported 1n Table 2-4. Using the Burkhard and Kuehl
1929A 5-6 06/23/86
-------
(1986) recalculated values, the values of the log [bloaccumulatlon (f1sh)J
defined by Lu et al. (1978) would be 4.51, 4.87 and 6.95 Indicating high
bloaccumulatlon for 2,8-DCDF, 2,3,7,8-TCDF and OCDF, respectively. The
value for the 2,3,7,8-TCDF agrees with that for 2,3,7,8-TCDD (Burkhard and
Kuehl, 1986). PCDFs have not been studied 1n model ecosystem.
1929A 5-7 06/23/86
-------
6. ENVIRONMENTAL LEVELS AND EXPOSURE
6.1. SUMMARY
PCDFs have been found 1n wastewaters and rivers, and also 1n the fat of
animals, mostly as acutely toxic 2,3,7,8-TCDF and 2,3,4,7,8-PeCDF, respec-
tively. Plants do not appear to bloaccumulate PCDFs. The similarity of
lake sediments with respect to PCDF and PCDD congeneric patterns Implies
that there may be a common source of PCDFs and PCDDs with air as the trans-
port medium. Both the emissions from municipal Incinerators and from leaded
automobile exhausts are correlated to PCDFs and PCDDs. This does not sup-
port Dow's trace chemistries of fire hypothesis. PCDFs In milk and from
Industrial accidents are Important routes of exposure also.
6.2. ENVIRONMENTAL LEVELS
6.2.1. Introduction. PCDFs have not been monitored systematically In the
environment. Zltko (1972) and Zltko et al. (1972) Investigated the presence
of PCDFs In several wildlife samples with negative results, but the analyti-
cal methodology used In these studies had a sensitivity several orders of
magnitude less than that required for detection of the probable environ-
mental concentrations (ppt) of PCDFs. In a similar study, Bowes et al.
(1973) Investigated the presence of PCDFs 1n wildlife populations with high
rates of embryonic death. Again, no PCDFs were Identified, either In
herring gull egg samples containing a total of -0.9 g PCBs or a sea lion
sample containing ~1 mg of PCBs.
The first positive report concerning the presence of PCDFs 1n the envi-
ronment was by Jungclaus et al. (1978) who Investigated the presence of
anthropogenic organic chemicals In the wastewater of a chemical manufactur-
ing plant and using GC/MS Identified a TrCDF among >100 compounds. The same
compound was Identified 1n the water of a river receiving the industrial
wastewater.
1930A 6-1 06/23/86
-------
Table 6-1 contains a summary of the levels found In environmental media
to 1986. PCDFs have been found 1n wastewater, river water, sediments and
air samples.
6.2.2. In Soil and Sediments. PCDFs deposited 1n soil strongly adhere to
the upper soil layers. Thlbodeaux (1983) stated that PCDFs and PCDDs
applied to the surface of soils having a high organic content generally stay
1n the upper 6-12" layer, while 1n more sandy soils they can migrate 3 feet
or more. However sediments seem to be the ultimate sinks of PCDFs. Czuczwa
et al. {1984, 1985) studied the presence of PCDFs and PCDDs In sediments and
sediment cores from the Great Lakes 1n the United States (Czuczwa and HHes,
1984, 1985, 1986), and annually In laminated sediments from lakes In
Switzerland (Czuczwa, et al. 1985). The accumulation of PCDDs and PCDFs
paralleled the rapid growth of the production of chlorinated chemicals since
1940. In fact PCDFs and PCDDs were absent from sediments before 1945, but
Increased thereafter. In all cases OCDD predominated (>1000 ppt); HpCDD and
HpCDF were the next most abundant congeners (1-15 ppt). The congener
profiles from Switzerland and from U.S. sediments were similar.
The absence of PCDFs from the sediments before 1940 Indicates that wood
burning, coal combustion and probably leaded automobile emissions, which
were prevalent before 1940, are not Important sources of PCDFs. The simi-
larity 1n the PCDF/PCDD profiles from the sediments In Switzerland and the
United Stages suggests a common source Involving atmospherically-transported
combustlonderlved PCDFs (Chapter 5). The PCDDs and PCDFs In the sediments
from Slsklwlt Lake located 1n Isle Royale In Northern Lake Superior could
only have been deposited from the atmosphere (Czuczwa et al., 1984). The
congener profile of PCDFs and PCDDs 1n sediments was similar to that 1n
airborne partlculate samples from two locations remote from S1sk1w1t Lake.
1930A 6-2 06/23/86
-------
TABLE 6-1
Levels of PCDFs In Environmental Media
Site
Sample
TCDF Concentration
Reference
Water
Soil/Sediments
CO
Mastewater. U.S.
Rlverwater, U.S.
Narragansett Bay, RI.
Water and suspended particles
Hudson River Sediment, U.S.
Woods Pond Sediment, MA
Saglnaw River/Lake Huron:
Surface Sediments (6)
Sediment Cores to 12 cm
Buffalo River Sediment, IL
Sediments:
Albany, Hudson River, NY
Tappan Zee Bridge, Hudson
River. NY
Slsklwlt Lake Sediment, MI
Cores 0-6 cm
8-9 cm
NBS Urban Dust
Surface Sediments from
lakes In Switzerland
Soil, Amsterdam, The
Netherlands
Trace of a TrCOF
Trace of a TrCDF
2,4,8-TrCDF found; 0.013-0.25 ng/ml
C14. <3-200; C15. <3-193; C16, <3-377;
Cl7, <3-2436; C18, <3-1010 pg/g
C14. <5; C15, 9; C16, 150; C17, 920;
C18, 270 pg/g
C14, 0.1-3; C15. 0.01-4; C16, 0.13-12;
C17, 0.2-30; C18, 0.05-7 ng/g
Cl4, 10-30; Cls, 100-200; Clfc, 100-200;
C17, 50-400; C18, 5-10 pg/g
2 TCDFs, OCOF detected
5 ng/g 2.3.7.8-TCDF
29-46 ng/g 2,3,7,8-TCDF
C14, C15, 2-5; C16, 2; C17, 17-20;
C18, 3-4 pg/g
Cl4, <0.4; Cls, <0-*: C16. <0-*:
C17, <0.4-1.6; C18, 1.1 pg/g
C14, 0.03-0.10; Cls, 0.31-1.3; Cl
C17, WO-5.8; C18, ND-1.9 ng/g
C14, 13-170; Cls, <10; C1&, 5-60;
C17, 100-240; C18, 10-510 ng/g
ND-1.6;
Jungclaus et al., 1978
Lake et al., 1981
Petty et al., 1983a
C14, 1413-24,000;
C16, ND-115 ng/g
, NO-5650;
Czoczwa and Kites, 1984.
1986
Kuehl et al., 1984b
O'Keefe et al.. 1984
Czuczwa et al.. 1984. 1985;
Czuczwa and Kites, 1986
L1 et al., 1985
Czuczwa et al., 1985;
Czuczwa and Kites, 1985
He Ida et al.. 1985
-------
TABLE 6-1 (cont.)
Site
Sample
TCOF Concentration
Reference
So11/Sediments
(cont.)
Air
60 Vapor and partlculate air
samples. Federal Republic of
Germany
Surface sediments from:
Lake Michigan (2)
Lake Erie (2)
Lake Ontario
Air Partlculate
Washington, DC (NBS)
St Louis, MO (NBS)
Air Sample, New York
U.S. Incinerator Plant Air
5-10 pg/m3 for sum of PCDDs and PCOEs
C14, <10; Cls, <1
C18, 0.5-24 ng/g
C14, <10; C15, <1
C18, 30-60 ng/g
C14. <10; C15, <1
Clg, 3600 ng/g
; C16. 8-33; C17. 70-240;
; Clb, <10; C17. 40-80;
; C16, 200; C17. 1200;
C14, 1.3; C15. 1.2; C16, 0.8; C17, 18;
Clg, 6.2 ppb
C14, 0.2; Cls, 0.2; C16. 0.3; C17. 12;
C18, 0.5 ppb
<0.1-1.9 pg/m» 2,3,7.8-TCDF
Total PCDF was 3 ng/m3
Oehme et al.,
Czuczwa and Hltes, 1986
Czuczwa et al., 1984, 1985;
Czuczwa and Hltes, 1986
O'Keefe et al., 1985b
Halle et al.. 1985
NO = Not detected
W
\
00
-------
They showed that the pattern of PCDDs/PCDFs 1n lake sediments resembled that
of the contaminants 1n chlorophenols and could be detected 1n sediments to a
depth of 12 cm. This does not support Dow's trace chemistry of fires
hypothesis. Reviews have been written by Hutzlnger et al. (1985a,b) and
Weeraslnghe and Gross (1985). A mathematical model for the transport/fate
of organlcs 1n soils has been published by Undstrom and Plver (1985).
6.2.3. In Plants. Few data on the concentrations of PCDFs 1n plants and
vegetables exist. Wild fruits collected from a Dutch area seriously contam-
inated with organochloMne waste compounds showed no presence of PCDFs and
PCDDs (Table 6-2) which, however, were present In topsoll, In fish and 1n
animals (Helda and OHe, 1985). Since there Is little bloaccumulatlon of
PCDDs 1n plants, probably PCDFs will not bloaccumulate to any extent either
(U.S. EPA, 1985). Plants' aerial parts may be contaminated through deposi-
tion of airborne particles or through volatilization or translocatlon of the
compounds present In soil. When 2,3,7,8-TCDD was added to soil, only 0.15%
accumulated 1n oats and soya beans (Isensee and Jones, 1971). Aerial
portions of plants growing In contaminated Seveso areas contained less
2,3,7,8-TCDD than roots, which had lower levels than the surrounding soil.
At a soil concentration of 10,000 ppt, fruits were found to contain no more
than 37 ppt, with 95% 2,3,7,8-TCDD being located In the peel. This strongly
suggests that contamination was due to surface dust deposition and not to
plant uptake (Wlpf et al., 1982). Underground tissues of some plants such
as carrots can take up UpophlUc chemicals from the soil (Cocuccl et al.,
1979; Pocch1ar1 et al., 1983). The 2,3,7,8-TCDD concentration 1n root
vegetables ranged from 3-10%, and In the aerial parts from 0.3-1% of the
surrounding highly contaminated soil (PocchlaM et al., 1983).
1930A 6-5 06/23/86
-------
TABLE 6-2
uO
CO
o
3>
cr
o
cr>
no
CO
Ranges
Material
Milk fat
Top soil (dry weight)
Worms
Mice/liver
/reference liver
Rabbits/liver
/reference liver
Pheasants/liver
/reference liver
Eel-Inside dump/liver
/muscle and fat
Eel-outs1de dump/liver
/muscle and fat
/reference liver
Pike-Inside dump/liver
/muscle
P1ke-outs1de dump/liver
/muscle
of 2,3,7,8-lCUl) and
1n ppt fresh weight
Number of
Samples Analyzed
6
9
9
4
1
14
5
8
2
2
2
3
1
1
1
3
2
pcui-s in lop sol I am
In a Dutch Hazardous
2,3,7,8-TCDD
<17
<20-1929
<1 5-889
<10-5237
<68
<10-50
<2-<50
<6-33
96-97
104-144
NA
30-42
584
21
<50-103
<30
d Biological s<
Waste Dump3
Total TCDFs
NA
1413-23,920
ND-3020
ND-13809
ND
ND-168
ND
ND
ND
2350-2765
4276-5193
NA
41-47
NA
959
41
ND
ND
ampies
Total
PeCDFs
NA
ND-5650
ND-300
1408-23,097
ND
ND-1121
ND-124
ND
ND
ND-169
182-558
NA
134-147
NA
975
154
ND-1419
263-268
Total
HxCDFs
NA
ND-115
ND-2694
ND-584
ND
ND-1081
ND-132
ND
ND
ND
ND
NA
ND
NA
ND
ND
ND
ND
CO
-------
WO
o
TABLE 6-2 (cont.)
I
—I
Material
Pike -reference/liver
/muscle
Minnow-inside dump/liver'3
Minnow-outside dump/liver
Blackberries
Elderberries
Pumpkin
Number of
Samples Analyzed
1
2
1
1
8
2
1
2,3,7,8-TCDD
<9
593
<20
ND-<10
<2-<17
<20
Total TCDFs
ND
ND
5808
ND
ND
ND
ND
Total
PeCDFs
ND
ND
ND
ND
ND
ND
ND
Total
HxCDFs
ND
ND
ND
ND
ND
ND
ND
aSource: Heida and Olie, 1985
^Only one specimen analyzed
ND = Not detectable
0 NA = Not analyzed
ro
CO
00
-------
6.2.4. In Aquatic Organisms. Evidence for aquatic wildlife contamination
by PCDFs was reported by Stalling et al. (1981), Kuehl et al. (1981) and
Rappe et al. (1981). Samples of fat from snapping turtles, (Chelyda serpen-
tlna). from the Hudson River and Baltic gray seal, (Hallchoerus grupus), and
from the Gulf of Bothnia were examined using a direct-probe negative
chemical lonlzatlon mass spectrometrlc technique of sensitivity 0.2 pg for
2,3,7,8-TCDF. The turtle fat containing >750 ppm of PCBs was found to be
contaminated by PeCDFs and HxCDFs while the seal fat, whose concentration 1n
PCBs was -100 ppm, contained trace amounts of PeCDFs (Stalling et al., 1981;
Rappe et al., 1981).
The same two samples were analyzed by Rappe et al. (1981) using high
resolution GC/MS In order to Identify the single PCDF Isomers. The 1somer1c
PCDF composition of the two samples showed a good correspondence despite the
differences between the two species and the different origins. The turtle
fat sample had a total PCDF concentration of 3 ppb distributed among 18 dif-
ferent Isomers; among these the toxic 2,3,7,8-TCDF and 2,3,4,7,8-PeCDF were
present at a concentration of 45 and 620 ppt, respectively. HpCDFs, PeCDFs
and HxCDFs predominated In that order (1000, 820 and 700 ppb, respectively).
In the seal fat, the total amount of PCDF was 40 ppt with the 2,3,7,8-
TCDF at 1 ppt and the 2,3,4,7,8-PeCDF at 15 ppt. The composition of the
major Isomers found In these biological samples corresponded fairly well to
the pattern present 1n commercial PCBs while a poor correlation was found
with the Isomer composition of other PCDF sources such as polychlorlnated
phenols and Incinerator fly ash samples. This suggested that the presence
of PCDFs 1n these samples was correlated with PCB pollution.
1930A 6-8 06/24/86
-------
Yamaglshl et al. (1981) also detected the presence of unspecified TCDFs
In Japanese fish (Zacco platypus), and Kuehl (1981) reported that fish taken
In U.S. rivers before 1978 contained a total estimated PCOFs ranging from
0.5-5 pg/g. Lake et al. (1981) quantHated a TrCOF 1n Rhode Island aquatic
life. In 1978, the mussel (Mytllus edudls) contained 643 ppb and six
samples in 1979 were found to have 52-1180 ppb. Clams (Hercenarla mercenla)
also contained 26 ppb, whereas lobster (Homorus amerlcanus) hepatocytes had
1260 ppb. Kuehl et al. (1986) also detected TrCDFs, TCDFs, PeCDFs, HxCDFs
and HpCDFs In fish from the Great Lakes.
These discoveries ushered 1n many Investigations (Table 6-3) Involving
the major IsomeMc PCDF classes 1n tissues of fish living 1n lakes, rivers
and seas.
PCDFs were Identified but not quantHated In fish from 18 major water-
sheds near the Great Lakes collected 1n 1979-1981 (Kuehl et al.. 1984a).
They have been Identified 1n fish from the Great Lakes (O'Keefe et al.,
1983; Ryan et al., 1983; Stalling et al., 1983). PCDDs were not detected In
fish samples from Lake Superior while a series of PCOFs could be Identified
In the same samples, Indicating more widespread background levels for the
PCOFs than for PCDDs (Stalling et al., 1983). Stalling et al. (1983) could
detect very little 2,3,7,8-TCDF In sediments but larger amounts In fish;
OCDF was abundant 1n sediments but low 1n fish. Similar results (Table 6-4)
were obtained 1n examining sediments and yellow perch from Woods Pond, which
was known to be contaminated by Aroclor 1260 (Petty et al., 1983a,b).
Stalling (1982) found carp, largemouth bass and striped bass from the lower
Hudson River contained 2,3,7,8-TCDF at concentrations ranging from 14-18
pg/g. The presence of 2,3,7,8-TCDF 1n striped bass was confirmed by O'Keefe
(1982). Since the upper section of the Hudson River was highly contaminated
1930A 6-9 06/23/86
-------
TABLE 6-3
PCOF Residues In Wildlife
Wildlife
Species
PCDF Level (pg/g)
Reference
Animals
Fish
I
__l
o
00
Fat from Snapping Turtle, U.S.
(Chelyda serplntlna)
Fat from grey seal, Sweden
(Hallchoerus grupus)
Seal. Sweden
Yellow Perch, NA
Bloater, Lake Superior
Lake Trout (Salvellnus naraavcush from:
Lake Superior (2)
Lake Ontario (2)
Lake Michigan (3)
Lake Huron
Brook Trout, (Salvellnus tontlnalls)
In Lake Ontario
Walleye (Stlzostedlon v. vltreum):
Lake Michigan
Lake Erie
Common carp (Ctenopharynqodon Idella):
Lake Erie
Lake Huron
Lake Michigan (2)
Rainbow Trout (Salmo galrdnerl).
Lake Ontario
C14, 45; C15, 820; C16, 700; C17, 1000;
C1B, 350
C14, 1; C15, 15; C16, 8; C17, 10; C18, 3
C14. 1; C15. 16; C16, 8; C17. 10; C18, 3
C14. 1.1; Clc, 0.64; Clf,, 0.44;
C17, <0.005; C18, <0.005
C14, 26; C15, 9; C16. 2; C17. 1; C18, 2
C14, 10-19; C15. 5; C16. ND-5; C17, ND-4;
C18, trace-36
C14, 34-40; Cls, 24-48; C1&, 10-29;
C17. ND-6; C18, MD-2
C14, 19-35; C15. 41-61; C16. 8-16; C17, 1-4;
C18, 1-6
C14. 19; C15, 9; C16. 242; C17. <1; C18. <1
C14, 19; C15, 4; C16, 3; C17. 3; C18, 3
C14, 4; C15. 4; C16, 0.7; C17. 0.7; C18, 3
C14. 18; C15, 9; C16, 6; C17, 5; C18, 2
C14. 5; C15, 5; C16. 2; C17. 4; C18. 2
C14, 27; C15, 44; C16. 34; C17, 44; C18. 4
C14, 5-37; C15, 12-73; C16, 5-145; C17, 6-31;
C18, 1-4
C14, 39; C15, 10; C16, 2; C17, ND;
C18, NO
Rappe et al., 1981;
Stalling et al., 1982
Buser et al., 1985
Petty et al., 1983a,b
Petty et al., 1983b;
Stalling et al., 1983
Stalling et al., 1983
Stalling et al., 1983
Stalling et al., 1983
Stalling et al., 1983
-------
TABLE 6-3 (cont.)
10
co
o
Wildlife
Bird
Ecosystem
ro
CO
CD
Species
PCDF Level (pg/g)
Herring, Sweden
Guillemot. Sweden
Herring, Sweden
Herring, Sweden
Guillemot, Sweden
Herring gull (Larus arqentatus).
Lake Huron
Contaminated animals and fruits 1n a
Netherlands Dump:
Worms (9)
Mouse liver (4)
Rabbit liver (14)
Pheasant liver (8)
Eel liver (2)
Eel muscle/fat (2)
Reference eel muscle/fat (3)
Pike liver
Pike muscle
Pike liver 2 (3)
P1ke muscle 2 (2)
Minnow liver 1
Minnow liver 2
Blackberries (8)
Elderberries (2)
Pumpkin (1)
C14, 50; C15, 430; C16, 420; C17, 1400;
C18, 100
C14, 10; C15, 800; C16, 1000; C17, 2500;
C18. 250
C14. 5; C15. 8; C16, 2
C14, 4; C15, 8; C16, 2
C14, 2; C15. 184; C16, 41
C14, 15-16; C15, 28-50; C16. 40-57;
C17, 7-17; C18. 3-5
C14, ND-3020; C15, ND-300; C16, ND-2694
C14, NO-13,800; C15, 1408-23,097; C16, ND-584
C14, ND-168; 015, ND-1121; C16, ND-1081
C14. C15. C16, NO
C14, 2350-2765; C15, ND-169; Clfc, ND
C14, 4276-5193; C15, 182-558; C16, ND
C14, 41-47; C15, 134-147; C16. ND
C14, 959; C15, 975; C16. ND
C14, 41; C15, 154; C16, ND
Cl5, ND-1419; C14, Clfe, ND
C15. 263-268; C14, C16. ND
C14, 5808; Cls, C1&. ND
C14, C15, C16, ND
C14, Cls, Cle, ND
C14. C15, C16. ND
C14. C15, C16, ND
Reference
Fish (cont.) Lake Ontario Fish:
Smelt (4)
Catfish
White Perch
Lake Trout (2)
Rainbow Trout
Ontario Salmon
3-112 C14
99 Cl4
15 C14
8.5-24 C14
200 C14
153 C14
Ryan et al., 1983
Buser et al., 1985
• Rappe et al., 1985d
Stalling et al., 1983
Helda and 011e, 1985
ND = Not detected
-------
TABLE 6-4
PCDF Distribution In a Pond Known to be
Contaminated by Aroclor 1260*
TCDF
PeCDF
HxCDF
HpCDF
OCDF
Total PCOF
PCB (vg/g)
PCDF/PCB
Ratio
Sediment
0.005
0.009
0.15
0.92
0.27
1.35
60
22.5xlO~6
PCDF Concentration (pq/q)
Yellow perch
1.06
0.64
0.44
0.005
0.005
2.14
170
12.6xlO~6
Aroclor 1260
290
1330
1810
780
29
4240
neat
4.24x10"*
*Source: Petty et al., 1983a,b
1930A
6-12
06/23/86
-------
by PCBs from Industrial discharges (Horn et al.. 1979), O'Keefe et al.
(1984) compared the PCOD and PCDF concentrations 1n striped bass from this
basin and from two other locations along the Atlantic coast. They found
2,3,7,8-TCOF 1n the fish from all three locations with concentrations vary-
ing from 6 ppt 1n Atlantic coast to 78 ppt In Hudson River. Analysis of
sediments and of nonmlgratory fishes confirmed the source of contamination.
Among fishes living Inside and 1n the near surroundings of a dump con-
taminated with waste organochloMne compounds, eels accumulated the highest
levels of PCDFs (see Tables 6-2 and 6-3). These amounted to 5,000 ppt 1n
the whole body and 2500 ppt In the liver. The levels 1n topsoll were
1500-2400 ppt (Helda and Ol1e, 1985) (see Table 6-2).
A search for toxic 2,3,7,8-conta1n1ng Isomers 1n wildlife has also been
conducted by many Investigators (Table 6-5). As expected, the more highly
chlorinated members bloaccumulated more than the TCDFs. The levels quoted
are lower than the actual levels because recoveries varied widely between 50
and 100% (Smith et al., 1984). Stalling et al. (1985a,b) have also utilized
principal components analysis to ascertain the sources of PCDFs 1n environ-
mental media. The toxic 2,3,7,8-substHuted Isomers predominate In all
wildlife with residues, these also being the predominant Isomers found In
Yusho victims. Norstrom et al., 91986) reported that crustaceans can
contain Isomers other than the 2,3,7,8-substHuted.
ZHko and Choi (1973) and ZHko et al. (1973) provided the only avail-
able Information concerning accumulation and metabolism of PCDFs In fish.
ZHko and Choi (1973) fed a group of 35 juvenile Atlantic salmon (Salmo
salar) several times dally for 140 days with dry fish food spiked with a
mixture of 2.1, 4.4, 2.2 and 9.7 yg/g food, respectively. An analysis of
the muscle and gut of fish that died between the 81st and 135th day and of
fish that survived for 140 days Indicated that only OCDF could be detected.
1930A 6-13 06/23/86
-------
TABLE 6-5
Levels of Toxic PCDF Isomers In Wild Animals
cr>
i
*
o
\
w
00
CT>
Wildlife Sampling Site/Spec les
Animal Hudson River, U.S./
Snapping turtle fat
(Chelyda serpentlna)
Gulf of Bothnia, Sweden/
Seal fat (HaHchocrus
grupus)
Seal fat
Fish Lake Ontario, Canada
Smelt (4)
Catfish
White perch
Lake trout (2)
Rainbow trout
Ontario salmon
Pacific salmon
Eel (2)
Hudson River. Water ford. NY
Carp (5), 1981
Goldfish (5). 1981
Hudson River, Albany, NY
Goldfish (3), 1981
Hudson River, Tappan Zee Bridge
Striped bass (9). 1981
Striped bass (4). 1983
Hudson River, Poughkeepsle
Striped bass (2), 1983
PCDF (pq/q)
«v
- 1
1 1 1 ooeo
, , coco co co f"- —
«o _ r»___ r~.r"j; ' — >*— •**>*
45 620
1 15
1 1 15 4
3.2-34
54
15
9-24
12
79
<10
NO
5
<9
<5
54
56
74-78
ci ,1
r- a
*7: *•£ Reference
Rappe et al.,
1981
Rappe et al.,
1981
5 5 Buser et al.,
1985
Ryan et al..
1983
O'Keefe
et al., 1984
O'Keefe
et al.. 1984
O'Keefe
et al.. 1984
O'Keefe
et al.. 1984
O'Keefe
et al.. 1984
-------
TABLE 6-5 (cont.)
vO
CO
o
3»
I
0
^
ro
Jj*
CD
Wildlife Sampling Site/Species
Fish Bayonne Bridge. Newark Bay
(cont.) Striped bass. 1983
Chesapeake Bay, MD
Striped bass (4). 1983
Long Island, NY
Striped bass (4), 1983
Rhode Island Coast (4), 1983
Striped bass (5), 1982
Common carp 1
2
Grass carp
Sweden
Herring
Guillemot
Herring
Herring
Guillemot
Bird Hudson River, Catsklll. NY
Black duck (6), 1980
Mallard duck, (12), 1980
Wood duck. 1980
Green Bay and Lake Michigan, WI
Night heron (Nyctlcorax
nyctlcorax)
1978
1982
Cormorant (Phalacrocorax aurlth)
1983
Herring Gull 1
2
PCOF (Da/a)
X.
COCO CO 0
00 r*> p- r» — » r* — » r- •*—
• — • •.— • • » irt « \ft U"»
ntj oo CM co *— • CM -— n-—
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50 100 80 250
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The detection limit was -0.02 yg/g wet weight for PCOF, TrCDF and TCDF.
The concentration of OCDF 1n muscle was 0.03 yg/g 1n dead fish and 0.01
yg/g In surviving fish. The concentrations in gut were 0.21 yg/g In
dead fish and 0.02 yg/g In living fish. The percent Hpid In muscle and
gut of dead and surviving fish was also measured and found to be higher in
gut. Concentrations of OCDF 1n I1p1d were calculated to be 3.53 yg/g and
1.30 yg/g In muscle of dead and alive fish, respectively, and 17.36 and
0.99 yg/g, respectively, in gut of dead and live fish. Although the
concentration of OCDF was higher In dying fish than 1n surviving fish, it 1s
unknown whether OCDF caused the toxic effects. The data indicate, however,
that the concentration of OCDF in fish was considerably lower than 1n the
spiked food.
Zitko et al. (1973) also showed that only a small portion of orally
administered 2,8-DCDF was accumulated 1n the tissues of brook trout (Salve-
linus fontinalis). Several fish were fed a cumulative dose (In gelatin
capsules) of 107-361 yg DCDF/g wet fish weight over variable exposure
periods. The tissue residues ranged between 0.052 and 0.340 yg/g
(0.01-0.13% of administered dose) 1n muscle and between 0.146 and 1.04
yg/g (0.08-0.4% of administered dose) 1n liver. Mass spectral analysis of
the organic compounds Isolated from the water containing excreta from one of
the fish indicated the presence of a DCDF and of a hydroxy-DCDF metabolite.
The results of Zitko et al. (1973) and ZHko and Choi (1973) indicate
that PCDFs are poorly absorbed 1n the gut of fish. The limited metabolic
information provided by Zitko et al. (1973) suggests that DCDFs are
hydroxylated by aquatic organisms.
6.2.5. In Birds. Tables 6-3 and 6-5 summarize the present data. O'Keefe
et al. (1984) determined the presence of PCDF in three duck species
1930A 6-16 06/23/86
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collected near the Hudson River. Concentrations of 2,3,7,8-TCDF ranged from
13-79 ppt 1n fat tissues. PCDFs and PCDDs were also detected In birds and
bird eggs In Wisconsin (Stalling et al., 1985a). 2,3,7,8-Chlorlnated
congeners were the major constituents. The pattern of PCDF and PCDD contam-
ination was compared In birds and fishes living In the same Great Lakes
area. The ratio of PCDFs to PCDDs 1n bird eggs were lower than In fish of
the Great Lakes, Indicating that 1n birds the rate of metabolism of PCDF 1s
probably higher than that of PCDDs.
6.2.6. In Terrestrial Animals. These data are summarized 1n Tables 6-3
and 6-5. TCDFs were determined In tissues of a horse grazing close to a
wire reclamation Incinerator; TCDF (unspecified Isomers) concentrations were
165 pg/g In the fat and 57 pg/g In the liver with unspecified TCDD levels of
45 and <6 pg/g 1n the fat and liver, respectively (Hryhorczuk et al., 1981).
Recently Rappe et al. (1985) found 2,3,7,8-chlorlnated PCDFs and PCDDs 1n
milk from cows that had grazed near an Incinerator 1n Switzerland. The
levels were found to be related to the distance between the grazing area and
Incinerator.
Helda and OHe (1985) analyzed soil and biological samples collected
from a household waste dump near Amsterdam, which appeared to be seriously
contaminated with organochlorlne compounds (2,4,5-T, dlchlorobenll, Undane,
tetrachlorodlfon). Fish, game animals, worms and mice contained measurable
amounts of the most toxic PCDDs and PCDFs (see Tables 6-2 and 6-3). The
highest levels were detected In fish, especially eel (see Table 6-2). The
worms tended to have the same congeneric pattern as soil (see Table 6-3),
while 1n mice the toxic Isomers prevailed probably as a result of differen-
tial blodegradatlon and excretion rates.
1930A 6-17 06/23/86
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Recently, Ryan et al. (1985a) found PCDDs and PCDFs 1n 40 chicken and
pork tissue samples also positive for pentachlorophenol. HxCDDs, HpCDDs and
OCDDs were present In 50, 62 and 46% of the samples at mean levels of 27, 52
and 90 ppt, respectively. Similar levels of HxCDF and HpCOF were found In
some of these samples. TCOFs, PeCOFs and OCOF were not detected. Compari-
son of the congeneric patterns of the chicken samples with those of penta-
chlorophenol -treated wood Indicated that pentachlorophenol was probably the
main source of contamination. Residues 1n humans are considered 1n
Chapter 7.
6.3. EXPOSURE
6.3.1. Food. Food may be an Important route of exposure since PCOFs have
high bloaccumulatlon coefficients (Section 5.5.). Since their concentra-
tions 1n PCBs 1s of most concern, It 1s Instructive to calculate the maximum
PCDF exposure levels associated with PCS exposure.
The present dietary Intake 1n the United States of PCBs from fish Is
-175 yg/day (Cordle et al., 1978). If the amount of 2,3,7,8-TCDF Is the
same as In a used Kanechlor 400 formulation (similar to Aroclor 1248), I.e.,
1.25 ppm, the amount of this Isomer stored 1f there 1s no excretion would be
-0.2 ng/day.
Workplace air levels of Aroclor 1242 can range as high as 2.22 mg/m3
(Ouw et al., 1976). If 1t 1s present as airborne partlculate, It would
contain ~3 ng/m3 of the 2,3,7,8-TCDF. If the same assumptions are used to
estimate the levels of this Isomer 1n fish, the levels of PCBs reported for
various environmental compartments are as follows: ambient air (<100
ng/m3), surface waters (10-50 ng/i), foundry effluent (12-335 ppb),
paper mill effluents (0.01-25 ppb), snow (0.17-0.24 ppb), wastewater (<520
ng/a) and sewage sludge (16 mg/kg) (Chapter 4). PCB residues 1n human
1930A 6-18 06/23/86
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adipose tissues 1n the United States are near detection limits: below
detection limit (34.2%), <1 ppm (33.3%), 1-2 ppm (27.3%) and >5.2 ppm (5.2%)
(Yobs, 1972). Human milk can contain up to 100 ppb of PCBs (Savage et al.,
1973). HxCDFs, HpCDFs and OCDFs have been detected In cow's milk, blood and
tissue fat, after pentachlorophenol was Ingested 1n the feed by a cow
(Firestone et al., 1979). Thus cow's milk may be a source of exposure to
animals exposed to PCDFs/PCDOs. Nygren et al. (1986) also reported low ppt
levels of PCDFs and PCDDs In samples of bovine fat, bovine milk, and 1n
commercial cream from Sweden and Scotland. Rappe et al. (1985e) also docu-
mented the PCDFs and PCDDs In bovine milk from Switzerland, with the highest
levels being found In samples collected 1n proximity to municipal and other
Incinerators. Human milk 1s also another possible source (Chapter 7).
Concern has been expressed over the formation of PCDDs during the
cooking of food. Liver, steak and hamburger from female Holstelns have been
examined, but only raw liver contained significant amounts of PCDDs; levels
decreased when the liver was cooked. Variation from sample to sample was
high so that no firm conclusions could be reached (Zablk and Zablk, 1980).
The situation 1s rather different for the PCDFs, especially If the tissues
contain PCBs. The PCDFs might possibly be formed from PCBs during cooking,
by the mechanisms cited 1n Section 2.5.2.1., and verification of this Is
Important. Cooking appears to decrease PCB levels 1n foods such as poultry
and milk as well as PBB levels 1n poultry (Zablk et al., 1978). The decline
appears to be related to the amount of fat rendered In the cooking process,
although a complete mass balance should be made using GC/MS. Firestone
(1977b) found 0.4 ppb of HpCDFs and 0.4 ppb of OCDFs In 2 of 15 commercial
U.S. gelatins. PeCDFs, HxCDFs and HpCDFs were found 1n one Mexican sample.
1930A 6-19 06/23/86
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It was thought that the PCDOs/PCDFs In fly ash would not be bloavallable
on Ingestlon (Hutzlnger et al.. 1985b), but rats, guinea pigs and hamsters
fed fly ash admixed with their food showed 2,3,7,8-substltuted Isomers 1n
their livers (Berg et al., 1985a). These results Imply that fly ash-
contaminated food, or particles 1n the air whose size 1s greater than
resplrable (15 vm) and which are cleared eventually to the stomach, will
be partially absorbed by animals and probably humans.
6.3.2. A1r. The Inhalation of fly ash particles containing PCDDs and
PCDFs 1s likely around municipal Incinerators 1n addition to PCDFs emitted
In automobile exhaust (Ballschmlter et al., 1986). PCDFs and PCDDs In both
vapor and partlculate form have been detected 1n Germany (Oehme et al.,
1986). The vapor and resplrable particles will be absorbed through the lung
alveoli, and the nonresplrable partlculate fraction may be partially
Ingested.
Another source of PCDF exposure Is during and after electrical fires
where PCBs were originally present (Chapter 4). Risk of exposure to PCDFs
Is high for fireman fighting electrical fires and the extent of exposures
should be Investigated as should workplaces Involving PCBs Including the
following: chemical plant handling or manufacturing halogenated aromatlcs;
factories making or repairing transformers or capacitors, using casting
waxes or having heat-exchange systems; In offices utilizing carbonless copy
paper containing PCBs; and during Incineration of halogenated aromatlcs.
6.3.3. Other Exposure Routes. Analysis of latex nipples (Gorskl, 1981)
revealed PCDDs and PCDFs to be present. TCDFs and PeCDFs were not found,
but HxCDFs (0.02-0.8 ppb), HpCDFs (0.1-3.1 ppb) and OCDFs (0.3-28 ppb) were
confirmed. These were not extractable by water, unlike pentachlorophenol.
1930A 6-20 06/24/86
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Gamma-Irradiation of the nipples for sterilization purposes was suggested as
a possible way that toxic Isomers could be produced. Exposure through
drinking water 1s unlikely.
1930A 6-21 06/23/86
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7. TOXICOLOGICAL EFFECTS IN HAN AND EXPERIMENTAL ANIMALS
7.1. SUMMARY OF EFFECTS OF POLYCHLORINATED DIBENZOFURANS
7.1.1. Tox1cok1net1cs. PCDFs are easily absorbed by passive diffusion
across cell membranes. For the highest chlorinated congeners I1poph1l1dty
can result 1n such Insolubility that absorption may be strongly Impeded.
The congeners with 2,3,7,8 chlorine-substitution are retained In tissues of
animals and humans. The presence of two adjacent unsubstltuted carbon atoms
predispose towards metabolism and hence excretion; therefore these congeners
have not been detected 1n the tissues of animals and humans even when
exposed to high levels. Broad species variations have been observed 1n
metabolism and excretion; the position that humans occupy regarding meta-
bolic fate and excretion relative to other species 1s still undetermined.
Physiological pharmacoklnetlc models developed 1n some animal species do not
permit extrapolation to man since little metabolic Information about the
pattern of the different Isomers within species Is available. Some of the
most relevant features are summarized here.
Studies on the absorption, metabolism and elimination of labeled
2,3,7,7-TCDF and of a mixture of PCDFs consisting of tetra-, penta- and
hexa-CDFs In laboratory animals (rats, guinea pigs, monkeys, mice) show that
CDFs preferentially accumulate In the liver and fat. The 2,3,4,7,8-PeCOF
accumulated more than other Isomers In the livers of monkeys and rats dosed
repeatedly with the mixture. Accumulation of the same Isomer several years
after accidental contamination was also observed In the liver of human
patients with Yusho or Yu-Cheng disease. Accumulation of PCDFs across the
placenta 1n the rat fetus Is limited, but transfer to suckling offspring Is
much greater. In the rat, the whole-body half-life of 2,3,7,8-TCDF 1s <2
days after a single 1.v. dose of 30.6 yg/kg bw. In contrast, the guinea
1931A 7-1 06/30/86
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pig has a minimum whole-body half-life of 20 days after a single 1.v. dose
of 6 vg/kg. A maximum whole-body half-life of 40 days was calculated In
guinea pigs manifesting no toxic effects. Intermediate behavior 1s apparent
1n the monkey with a whole-body half-life of 8 days after a single 1.v. dose
of 30.6 vg/kg. In all 3 species, excretion of unmetabollzed 2,3,7,8-TCDF
1s negligible and Us metab6Hsm appears to represent a detoxification pro-
cess. Excretion 1s predominantly by feces.
Highly chlorinated Homers accumulate In the liver and adipose tissue of
humans and persist for many years. 2,3,4,7,8-PeCDF and 1,2,3,4,7,8-HxCDF
are retained by the liver at higher concentrations than other Isomers. They
have been detected 1n the Hver and the adipose tissue of Yusho and Yu-Cheng
patients many years after Intoxication. PCDFs and PCDDs have been assayed
In tissues of patients 2 years or more after exposure In a building contam-
inated with these compounds; levels of PCOFs were elevated, especially those
Isomers found In the building. Many of these were similar to those found 1n
Yusho and Yu-Cheng patients. Surprisingly, high levels of 2,3,7,8-chlorlne
substituted congeners have been found In adipose tissue and 1n breast milk
of the general population from highly Industrialized countries of Europe
and North America. Lower levels have been detected In populations from
North Vietnam that was considered not contaminated by phenoxy add herbi-
cides and that had a low level of Industrial contamination. Therefore
levels of PCOFs In fat biopsies should be considered as a useful biological
Indicator of levels of exposure for ep1dem1olog1cal studies.
Studies establishing the human metabolism and the kinetics of these
compounds are still needed. The kinetics of the more persistent hexa- and
hepta-1somers warrant Intensive Investigations. When the general population
Is exposed to unknown contamination through the food chain and air, subjects
1931A 7-2 06/30/86
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with a high body burden can transmit toxic PCDFs to their progeny through
placenta and milk, Indicating that such babies are a population at high risk.
7.1.2. Tox1c1ty. Some PCDFs have high acute toxlclty. -.The 2,3,7,8-
TCDF, and 1,2,3,7,8- and 2,3,4,7,8-PeCDFs are some of the most acutely toxic
Isomers. The acute oral LD5Q of 2,3,7,8-TCDF 1s 5-10 vg/kg In guinea
pigs (2-4 times higher than 2,3,7,8-TCDD), >6000 yg/kg In the mouse and
the rat (20 times higher than 2,3,7,8-TCDD), and 1000 yg/kg In the rhesus
monkey. The symptoms of toxlclty were very similar to those reported for
2,3,7,8-TCDD; for example, thymlc atrophy 1n all species; liver pathology 1n
rabbits, rats and mice; hydroperlcardlum 1n chicks; alteration of skin and
adnexa, liver bile duct and gastrointestinal tract 1n addition to hyper-
plastic and metaplastlc changes In the melbomlan glands of the eyelid and
the cerumlnous glands of the ear canal 1n the rhesus monkey. The eyelid and
ear canal are the most sensitive Indicators of PCDF toxlclty 1n the monkey.
There Is a lag of several days between administration and the appearance of
clinical toxic responses. Two of three monkeys died on a diet containing 5
vg/kg (food) administered over 6 months. The acute LD™ values, In
yg/kg, for Hartley strain guinea pigs over 30 days are as follows:
2,3,7,8-TCDF (7); 2,3,4,7,8-PeCDF (<10); 2,3,7,8-TBDF (<15); and
2,3,4,6,7,8-HxCDF (120).
It Is highly probable that continuous exposure at low levels causes
cumulative toxic effects, especially In view of the lesions observed 1n
monkeys at an acute dose, which was <4% of the LD,0.
Few studies relate to the subacute toxlclty of PCDFs. The most complete
data concern the effect of 2,3,7,8-TCDF on mice, chicken, guinea pigs and
rhesus monkeys. Common signs for various species seem to be a decrease In
body weight and Involution of the thymus. In some species (rodents and mon-
1931A 7-3 06/22/86
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keys) "hver morphological lesions were observed such as changes of endo-
plasmlc retlculum, cytoplasmlc I1p1d droplets and mltochondrlal altera-
tions. These ultrastructural effects should be considered with particular
attention and carefully controlled 1n planned studies using single Isomers,
as such alterations have been recently observed 1n the Hver of patients
known to be exposed to PCDFs. Ultrastructural alterations of Hver mito-
chondria In Hver biopsies, together with the monitoring of the contaminants
1n fat biopsies, are useful biological markers for exposure to PCDF and
related chemicals.
It appears likely that PCDFs contribute substantially to the toxlclty of
commercial PCBs and polychlorophenols and are responsible for the symptom-
atology of Yusho and Yu-Cheng diseases.
Dermal toxlclty Is a common effect In animal species Including humans.
However, the different species develop different types of skin lesions.
After treatment with various mixtures of PCBs, rabbits and rats develop
lesions resembling the chloracne observed 1n Yusho and Yu-Cheng patients.
Information on PCOF effects on the Immune system 1s stm limited,
although reduction 1n thymus weight and humoral antibody production has been
observed 1n all the animal species tested. The results with pure PCDF
Isomers show that the pattern of 1mmunotox1c1ty for PCDF-active Isomers 1s
similar to that of the corresponding PCDD Isomers, provided that higher dos-
ages are given to laboratory animals. Data obtained with 2,3,7,8-TCDD In
mouse strains eliciting different sensitivities to Induction of aryl hydro-
carbon-hydroxylase (AHH), suggest that the Individual genetic background may
play a role 1n the degree of PCDF-1nduced Immunotoxlclty.
1931A 7-4 06/22/86
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A number of PCOF Isomers show the capacity to cause a relatively high
degree of Induction of some liver mixed function oxygenases, such as AHH and
ethoxyresoruf1n-o-deethylase (EROO) both In yjvo and ±n vitro. AHH Induc-
tion activity has two structural requirements: first, halogen atoms must
occupy at least 3 of the 4 lateral ring positions (2, 3, 7 and 8); second,
at least 1 vicinal carbon position must be unsubstHuted. Structureactlvlty
studies conducted in vitro demonstrated the existence of a broad difference
In potency of PCDF Induction. The lack of kinetic studies In rodents does
not allow precise comparisons to be made between In vitro and in vivo
results. However, recently good correlations were obtained by plotting the
Inducing capacity of AHH of a single PCDF congener versus the rat body
weight loss or thymlc atrophy. This approach seems to be particularly
useful for testing mixtures of PCDFs with an unknown exact composition.
Biochemical studies have shown that for the manifestation of their
Inducing effect PCDFs must bind to a cytosollc protein binding site. The
occupancy of the receptor seems to be crucial. This event not only causes
AHH and EROD Induction but (more Important) represents the first step trig-
gering the toxic effects displayed by these compounds. Recent results on on
the Interaction of TCDD with EGF receptor or with receptor protein present
In the thymlc epithelial cells reinforce this point of view. So far there
are no Indications that AHH Induction could be a cause of toxldty per se,
but this fact could be an Indication of the potential susceptibility of this
organism to more profound toxic signs.
7.1.3. Epidemiology. Information on the toxic effects of PCDF In man
comes mostly from Japan and Taiwan where 1n 1968 and 1979, respectively,
many people ate rice oil that was later found to be contaminated with PCBs,
PCDFs and PCQ. More than 4000 people 1n the two episodes were Intoxicated.
1931A 7-5 06/22/86
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Some general symptoms Included: retarded growth, abnormal I1p1d metabolism,
liver disturbances, acneform eruption, skin pigmentation and cutaneous
lesions. The clinical profile of this syndrome has been well defined but
long-term toxldty, carcinogenic risk and mortality rate consequent to PCDF
exposure have not yet been assessed.
7.2. INTRODUCTION
The toxlclty of PCDFs 1n laboratory animals has received attention as a
consequence of their Identification In widely used Industrial chemicals,
such as the PCBs and polychlorophenols. Evidence 1s accumulating that many
of the toxic effects of various commercial chemicals may actually result
from traces of PCDFs. The critical observation leading to the hypothesis
that a trace contaminant might be the major toxic factor was made by Vos and
Koeman (1970) who found significant differences In toxlclty between three
commercial PCB preparations. Later, Vos et al. (1970) showed that most
acutely toxic preparation contained TCDFs and PeCDFs. Out of 135 possible
Isomers of PCDFs only 10-12 are expected to have significant acute toxlc-
lty. The most acutely toxic Isomers appear to be 2,3,7,8-TCDF, 1,2,3,7,8-
PeCDF and 2,3,4,7,8-PeCDF (McKlnney et al., 1976; Poland et al., 1976; Moore
et al., 1979). Dlbenzofuran (DF) and OCDF have low acute toxlclty (Gold-
stein et al., 1978b). Toxlcologlcal studies of PCDFs reveal marked similar-
ities to the effects of PCDDs (Moore et al., 1979; WHO, 1978b). The estim-
ated difference 1n toxlclty between 2,3,7,8-TCDO and 2,3,7,8-TCDF Is at
least 10-fold (Klmbrough, 1974; Klmbrough et al., 1978; Moore et al., 1979),
but It 1s consistently dependent on the animal species considered and on the
specific parameter Investigated. The earlier studies were essentially
devoted to Identifying the acute toxic effects. The series of Incidents
Involving a large number of people from the general population or from
1931A 7-6 06/22/86
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exposed workers In which PCDFs have been Implicated as the major class of
compounds responsible for the toxic effects have prompted researchers to
design more extensive studies on the kinetics, subchronlc toxlclty and bio-
chemical mechanisms. Therefore, studies on long-term effects of these chem-
icals are warranted.
7.3. TOXICOKINETICS
7.3.1. Introduction. The disposition of the PCDFs In animals depends on
the physlcochemlcal properties of the single Isomers. Their relative lipld
solubility, which Increases with Increasing halogenatlon, promotes their
passive absorption from the gastrointestinal tract but Impedes their excre-
tion directly 1n the urine and the bile. In addition, since PCDFs are Upo-
phlUc, animals with high adipose weight/body weight ratios tend to retain
them. Thus, metabolism 1s the rate-limiting factor 1n the fate of PCDFs In
animals. A particular aspect that must be considered In assessing the b1o-
avallablHty Is the Influence of environmental matrices, route of adminis-
tration and vehicles when PCDFs come from environmental pollution or from
contamination of chemicals (Kamlnsky et al., 1985).
Metabolism Is also dependent on molecular characteristics, since the
less highly halogenated Isomers and those with unsubstHuted vicinal carbon
atoms are more readily metabolized. In all animal species studied, the
metabolites, and to some extent the parent compound, are excreted preferen-
tially 1n the feces, through the bile and to a lesser extent 1n the urine.
Data on distribution, metabolism and excretion of 2,3,7,8-TCDF are
available for several different animal species. Recently, a physiological
pharmacoklnetlc model was developed for 2,3,7,8-TCDF (King et al., 1983).
7.3.2. Absorption. PCDFs are I1p1d soluble compounds and are absorbed
a
from the gastrointestinal tract by passive diffusion across cell membranes.
1931A 7-7 06/22/86
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In rats (Blrnbaum et al., 1980, Decad et al., 1982), mice {Decad et al.,
1981a; 1982) and guinea pigs (Decad et al., 1981b; 1982) 2,3,7,8-TCDF
appears to be absorbed to a very high degree. In guinea pigs, absorption of
TCDF was estimated at >95% 1n the gut after an oral exposure.
As for other halogenated compounds (Damstra et al., 1982) the absorption
tends to decrease with Increasing halogenatlon above a certain point. For
the 1,2,4,6,8,9-HxCDF the degree of absorption seems to be lower than for
TCDF (Blrnbaum, 1985). That highly halogenated PCDFs could be less easily
absorbed has been suggested but not Investigated because of the lack of
appreciable amounts of the various congeners needed for these studies.
Absorption can also be strongly Influenced by the vehicle. In the
guinea pig acute toxlclty of 2,3,7,8-TCDD (LD™) 1s approximately one-
eighth of Its LD In corn oil (Sllkworth et al., 1982) and 3-5 times
higher levels of TCDD are needed to cause toxic effects when they are admin-
istered mixed with dirt (McConnell et al., 1984). It has been suggested
that similar differences may also apply to PCDF (SUkworth et al., 1982).
However, direct evidence Is lacking for PCDF. Berg (1985b, 1986) report
that the bloavallablUty of PCDF absorbed on fly ash of a municipal Inciner-
ator and administered with the diet was very scant In different animal spe-
cies.
7.3.3. Distribution, Metabolism and Excretion. Disposition has been
studied 1n various animal species. However, a critical comparative analysis
of the results Is difficult as PCDF mixtures of variable (and In some cases
unknown) composition have been used and the experimental conditions, doses,
routes of administration, duration and schedules of data recording differ.
Only a few trials used single purified PCDF Isomers. However, from all
o
these data, which are summarized 1n the following sections, 1L appears that
1931A 7-8 06/22/86
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liver and adipose tissue are the preferential tissues for localization, that
elimination 1s by metabolism, and that metabolites, once formed, are excret-
ed without delay.
7.3.3.1. RAT — Kurokl et al. (1980) used a PCDF mixture (14%
1,2,7,8-TCDF; 35% 2.3.7.8-TCOF; 1% 1,2,4,7,8-PeCDF; 49% 1,2,3,7,8-PeCDF; 1%
2,3,4,7,8-PeCDF; and 1% unspecified HeCDF) to Investigate the persistence of
PCDF In the liver of rats and monkeys. Four male Wlstar rats (-100 g and 4
weeks of age) were given a single 1.p. dose of PCDF 10 mg/kg bw. The rats
were killed after 5 days and their livers were removed for GC/MS assay of
PCDF. The data are presented In Table 7-1. The 1,2,7,8-TCDF and
1,2,4,7,8-PeCDF were not detected, and the 2,3,7,8-TCDF was present at
0.6-2/5% of the total dose. The amounts of 2,3,4,7,8-PeCDF and HeCDFs
retained 1n the liver were 76-82% and 100% of the total dose, respectively.
Thus, 2,3,7,8-TCDF was eliminated from the liver 1n these rats with a resi-
dence half-time of ~1 day.
Blrnbaum et al. (1980) and Decad et al., 1982 Investigated the absorp-
tion, distribution and excretion of radlolabeled 2,3,7,8-TCDF In 200-500 g
male Fisher 344 rats. The compound was administered at oral doses of 30.6
(0.1 ymol/kg) and 306 ymol/kg (1.0 yg/kg bw), and at an 1.v. dose of
30.6 yg/kg bw. About 90% of both oral doses was absorbed. Table 7-2
shows that the adrenals, liver, lungs and kidneys contained, the highest
level of radioactivity 15 minutes after the 1.v. Injection (0.1 pmol/kg
bw). Subsequent redistribution to adipose tissue followed, the concentra-
tion was maximum there 7 hours after administration. The pattern of distri-
bution of radlolabel 1n liver fat muscle, skin and blood, and the amounts
excreted In urine and feces 3 days after administration were essentially
Independent of the dose and route of administration (Table 7-3). No evl-
1931A 7-9 06/22/86
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TABLE 7-1
Percentages of the Total Intake of Original PCDF Isomers Found In the Livers of Monkeys and Ratsa.b
Mixture Administered
1,2,7,8-
(14%)
2,3,7,8-
(35%)
CDF
1,2,4,7,8-
(IX)
1,2,3,7,8-
(49%)
2,3,4,7,8-
(1%)
Hexa-
(1%)
Male Hlstar Rat
(100 g)
1
2
3
Mean * S.D.
ND
ND
ND
ND
ND
1
1.5
2.5
0.6
1.4*0.8
ND
ND
ND
ND
ND
18
17
21
11
18*3
81
80
82
80*3
109
104
108
107
107*2
i
o
Rhesus Monkey
1
2
3
Mean * S.D.
ND
ND
ND
ND
0.02
0.09
0.04*0.05
0.4
0.3
0.2*0.2
0.2
0.2
0.2
0.2
5.2
1.7
0.8
2.6*2.3
0.8
0.1
ND
0.3*0.4
o
V.
ro
CO
aSource: Kurokl et al., 1980
bThree monkeys were fed Kanechlor 400 (0.25; 0.5 mg/kg bw) and PCDF (1.25; 2.5 vg/kg bw) for up to 32
days. Rats were Injected l.p. with 10 mg PCDF/kg bw and killed after 5 days. The total Ingested
Kanechlor 400 was 86, 88 and 30 mg In monkeys designated 1, 2 and 3, respectively; the total Ingested
PCDF was 430, 440 and 151 wg, respectively. Levels of Kanechlor 1n the liver were 1.7, 0.8 and 0.04
ppm, and of PCDF 5.4, 4.2 and 1.1 ppb, respectively. Levels of Kanechlor In fat were 8.6, 25 and 1.2
ppm, and of PCDF 0.9, 26 and 0.2 ppb, respectively.
ND = Not detectable
-------
TABLE 7-2
Concentration of "C-2.3.7.8-TCOF 1n Tissues of Male Fisher 344 Ratsa»b
Tissue
Blood
Liver
Fat
Muscle
Skin
Kidneys
Adrenals
Thyrus
Spleen
Testes
Brain
Lungs
Heart
15 Minutes
0.12±0.04
4.4 ±0.2
0.20±0.03
0.25±0.01
0.17±0.02
0.67±0.04
7.4 ±6.9
0.52±0.12
0.37±0.07
0.09±0.01
0.25±0.01
1.08^0.08
0.66i0.03
% Total Label (q/tlssue basis)
180 Minutes
0.04±0.01
5.1 ±0.4
0.44±0.07
0.06±0.00
0.20i0.01
0.17±0.03
4.7 tl.3
0.54±0.13
0.08±0.02
0.09±0.01
0.15±0.03
0.24±0.02
0.1HO.OO
1 Day
0.03*0.01
2.2 ±0.4
0.64±0.11
0.30±0.02
0.07±0.01
0.08±0.01
0.34±0.14
0.07±0.03
0.02±0.00
0.06±0.02
0.02±0.00
0.07±0.02
0.02±0.02
aSource: Blrnbaum et al., 1980
bDosed with 0.1 ymol/kg bw 1.v.
1931A
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dence of enterohepatlc circulation was found Examination of tissue
extracts showed that nearly all radlolabel 1n blood and tissues, at least up
to 1 day postadmlnlstratlon, was present as unchanged 2,3,7,8-TCDF. Thus,
the half-time of radlolabel loss from selected tissues showing a blphaslc
rate (Table 7-4) represent half-times of loss of 2,3,7,8-TCDF. The loss
from skin was blphaslc, the early component having a half-time comparable to
other tissues (see Table 7-4), followed by the slow component with a half-
time of -11 days, which accounted for <1% of the total dose.
In contrast with blood and tissues, >99% of the radlolabel excreted 1n
the feces and urine was associated with metabolites of 2,3,7,8-TCDF. Since
the metabolites appeared to be excreted as soon as they were formed, the
half-times of radlolabel 1n the urine (1.3 days) and feces (1.8 days) repre-
sent the half-time of elimination (by metabolism) of 2,3,7,8-TCDF. These
values agree with the half-times 1n blood and liver (see Table 7-4). The
feces was the primary route of excretion, accounting for -85% of the total
dose, while <6% was excreted In the urine. The ratio of kidney to bile
clearance was 0.03.
Yoshlmura et al. (1984) studied the tissue and subcellular distribution
of 2,3,4,7,8-PeCDF given orally to the rat. More than 60% of the dose was
accumulated In the Hver after 5 days and persisted over a period of 3
weeks. The small percentage of radioactivity distributed In all other tis-
sues was eliminated quickly. The subcellular liver distribution of PeCDF
was parallel to that of cytochrome P450. Excretion of the unchanged com-
pound 1n the feces amounted to -32% of the dose during the first 24 hours
with progressively smaller amounts during the following 3 weeks. No excre-
tion was recorded In urine up to day 21. The fecal excretion of PeCDF can
be Increased by treatment with activated charcoal beads (Yoshlmura et al.,
1931A 7-12 06/22/86
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TABLE 7-3
Distribution of Radioactivity 3 Days After Administration of
to 200-250 g Male Fisher 344 Rats*
Percentage Total Dose
Tissue
Liver
Fat
Skin
Total excreted
In feces
In urine
Oral
1.0 pfflol/kg
3.9±2.2
9.8+3.6
1.H0.6
68 ±6
1.5+0.3
0.1 timol/kg
5.0+0.4
4.6+0.8
1.H0.2
70 ±4
1.76+0.01
l.v.
0.1 ymol/kg
5.9±0.3
12 t2
1.2±0.3
63 ilO
2.0±0.4
*Source: Blrnbaum et a!., 1980
1931A 7-13 06/22/86
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TABLE 7-4
Components for the Elimination of Radioactivity
from Tissues of Fisher 344 Rats3
Tissue
Blood
Liver
Fat
Muscle
Skin
Component
1
2
1
2
1
1
2
1
2
Pool Size
(X total dose)
1.3+0.4
0.9+0.2
29 +6
31 ±3
18 +1
25 +2
6.7il.5
6.8*0.5
1.2+0.4
Decay Rate
(day-*)
32 +22
0.6 ±0.3
7.0 +2.7
0.55+0.08
0.19+0.04
29 +6
0.96^0.50
1.6 +0.3
0.06+0.41
Mean Half-Time
(days)
0.02
1.1
0.10
1.3
3.8
0.024
0.7
0.45
nb
aSource: Blrnbaum et al., 1980
DComplex redistribution occurs making the calculation of an S.D. tenuous.
1931A
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06/22/86
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1986). Such treatment in rats for 3 weeks stimulated excretion -3-fold and
significantly suppressed PeCDF toxlclty (thymus atrophy and liver hyper-
trophy). PeCDF levels In blood, adipose tissue and, to a lesser extent, In
liver, tended to decline.
Recently, Brewster and Blrnbaum (1986) found that the disposition of
2,3,4,7,8-PeCDF 1n the rat was similar in animals given either an oral or
1.v. dose; they confirmed the greater accumulation of the administered chem-
ical 1n liver. Fat, skin and muscles were found to be minor depots. Excre-
tion by feces was <1% of the administered dose per day and <0.5 in the urine
within 3 days. No PCDF was detected 1n expired air.
Data from Blrnbaum (1985) Indicate that 1,2,3,6,7,9-HxCOF Is not easily
metabolized by rats, compared with the TCDFs and PeCDFs and Is excreted in
feces less than TCOF (ratio feces/urlne 13 for HxCOF and 31.4 for TCOF).
The half-life for elimination has been calculated at 13 days. These results
confirm the data obtained by Kurokl et al. (1980) who showed that rats elim-
inate 2,3,7,8-TCDF and retain PeCDFs and HxCDFs.
The few data available Indicate that in the rat PCDFs concentrate In
liver more than in fat with a tendency for higher levels of halogenatlon to
result in higher liver concentrations (liver/adipose % total dose 1.30 for
2,3,7,8-TCDF, 1.8 for 1,3,4,5,7,9-HxCDF) (Blrnbaum, 1985).
7.3.3.2. MOUSE — Morita and Olshl (1977) gave male ICR mice a single
i.p. dose of a mixture of 0.5 g PCDF 1n corn oil. The mixture contained
unspecified 2-TCDFs, 4-PeCDFs and 4-HxCDFs. The less chlorinated PCDFs were
eliminated from tissues faster than the highly chlorinated ones. The half-
times 1n tissues were a week or less.
Decad et al. (1982) studied the distribution and excretion of 2,3,7,8-
TCDF in two strains of mice given 30.6 yg 1.v. radlolabeled 2,3,7,8-
1931A 7-15 06/22/86
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TCDF/kg bw. Higher concentrations of radlolabel were observed 1n the liver
than In any of the other 12 tissues sampled up to 10 days postadmlnlstra-
tlon. DBA/2J mice had -1.6 times as much adipose tissues as C57B1/6J mice,
and the nature of adipose tissue radlolabel kinetics differed considerably
In the two strains, with the concentration of radlolabel peaking later and
at a higher level 1n the adipose tissue of DBA mice. Nearly all the tissue
radlolabel was unmetabollzed 2,3,7,8-TCDF, while essentially all of the
radlolabel In urine and -80% of the radlolabel In feces, after the first
postadmlnlstratlon day, represented metabolites. The whole-body residence
half-time, calculated from the rates of excretion of radlolabel In the urine
and feces, was ~2 days 1n C57B1/6J mice and ~4 days 1n DBA/2J mice
(Table 7-5).
Nagayama et al. (1980) studied the transfer of PCDFs to the fetuses and
offspring of ddN mice (25 g bw) fed a diet containing 0.6 ppm of a mixture
of PCDF (48% TCDFs, 49% PeCDFs and 3% HxCOFs) for 18 days after mating and
14 days after delivery. Accumulation of PCDF 1n the fetuses across the
placenta was only 0.003% of the total amount Ingested by the dam. There was
much greater transfer of PCDF to suckling offspring (1.2% of the total
amount Ingested by the dam). PCDF accumulated more 1n the dams' livers
(5.1% of the total amount Ingested) than 1n other tissues. Data from Weber
and Blrnbaum (1985) confirmed that <0.1% of the total dose appeared In the
fetuses 1 day after acute exposure on day 11 of gestation.
7.3.3.3. MONKEY — The PCDF mixture reported In Section 7.3.3.1. was
given orally, In combination with a commercial PCB mixture, to 1 female and
2 male monkeys dally for 26-32 days, after which the monkeys were killed and
adipose tissue and liver analyzed for their content of PCDF Isomers (Kurokl
1931A 7-16 06/22/86
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TABLE 7-5
Excretion of 2,3,7,8-TCDF-Derlved Radioactivity by Three Mice*
Mouse
Strain
C57B1/6J
DBA/20
Elimination
Route
Urine
Feces
Urine +• feces
Urine
Feces
Urine * feces
Mean +_ S.D. Cumulative
Excretion of
Dose by Day 10 (%)
12. 6+ 0.1
81.9±13.0
97. 0± 9.2
19.9+ 4.6
55.8+ 4.8
75.7 + 5.8
Mean
Half-Time
(days)
2.8
1.8
2.0
4.9
5.4
4.0
*Source: Decad et al., 1982
1931A
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06/22/86
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et al., 1980). As In the rat, PeCDFs and HxCDFs were retained at higher
concentrations than TCDFs (see Table 7-1).
In the only kinetic study using a single Isomer (Blrnbaum et al., 1981),
three male rhesus monkeys were given 30.6 yg 2,3,7,8-TCDF/kg bw l.v. Loss
of radlolabel from blood was monitored only for 18 minutes In order to avoid
an extended period of sedation (Ketaset 50 mg 1.m.) and to minimize stress.
The monkeys were killed 21 days after treatment, and the pattern of radio-
label distribution In the tissues was determined (Table 7-6). Less than 10%
of the Injected dose remained In the body at this time. As with the rat,
all radlolabel extractable from liver, fat, skin and muscle was unmetabollz-
ed 2,3,7,8-TDCF, but about one-third of the small amount of extractable
radlolabel In blood did not co-chromatograph with the parent compound and
may have been associated with metabolites. Only metabolized 2,3,7,8-TCOF
was extracted from the urine, and nearly all radlolabel extracted from the
feces represented metabolites. Thus, the half-times of radlolabel 1n urine
(6.2 days) and 1n feces (10.3 days) represent estimates of the whole-body
half-time. Eight percent of the dose was excreted 1n the urine and 4354 1n
the feces.
King et al. (1983), using the data for the monkey (Blrnbaum et al.,
1981), rats (Blrnbaum et al., 1980), and mice (Decad et al., 1982), compared
the tissue to blood distribution ratios (Section 7.3.3.5.).
7.3.3.4. GUINEA PIG -- The extremely high toxlclty of the PCDFs In
guinea pigs (Moore et al., 1979) makes 1t difficult to obtain reliable
Information concerning their disposition. Decad et al. (1981b) reported
that the distribution of 2,3,7,8-TCOF radlolabel In guinea pigs during the
first 24 hours after either oral or l.v. administration of 6 yg/kg bw
(near the LD ) was similar to that 1n other species, with liver, fat,
1931A 7-18 06/22/86
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TABLE 7-6
Distribution of 2,3,7,8-TCDF-Der1ved Radioactivity In Three
Rhesus Monkeys 21 Days After Treatment3
Tissue
Blood
Liver
Skin
Fat
Percentage Total Doseb
0.37±0.02
1.0 ±0.8
2.4 ±1.6
3.7 ±2.8
pmol TCDF/g T1ssuec
4.8± 0.3
47 ±30
12 ± 6
47 ± 0.2
aSource: Blrnbaum et al., 1981
b1.v., 0.1 vmol/kg bw
cMean + standard deviation
1931A 7-19 06/22/86
-------
muscle and skin containing much of the radlolabel, but the ratio of liver to
adipose tissue was lower than 1n the rat (Table 7-7). During the second
24-hour period, however, redistribution to the liver began. Subsequently,
the amount of radlolabel 1n the liver rose to over 50% of the total adminis-
tered amount by the ninth day postadmlnlstratlon. This redistribution and
accumulation was at least partly the consequence of mobilization of body
fat, a toxic effect of 2,3,7,8-TCDF. In this study a minimum half-life of
20 days has been Indicated, but 1n view of this fat loss with release of
radlolabel, no reliable half-life can be calculated from these data. In
fact In a more recent study by loannou et al. (1983) the half-life calculat-
ed on data from animals showing little or no significant effects of TCDF
Intoxication was -40 days. In a multiple oral treatment the concentration
In adipose tissue was proportional to the dose. With Increasing time after
dosage, the distribution starts to shift from adipose tissue to liver.
The amounts of radlolabel excreted In the urine and feces of these
guinea pigs were comparable, only -6.6% being excreted by each route by the
ninth day after 1.v. Injection. While radlolabel was excreted In the urine
as metabolites, >90% of the radlolabel excreted In the feces was unmetabol-
1zed 2,3,7,8-TCDF. Thus, although urinary excretion by these guinea pigs
appeared comparable with that In other species at exposures that do not
cause toxldty, fecal excretion was markedly lower compared with the other
species and was restricted largely to parent 2,3,7,8-TCDF.
Attention has been paid 1n recent years to the b1oava1labH1ty of PCDF
from different matrices. Berg et al. (1985b) studied Us availability from
fly ash of a municipal Incinerator 1n the rat, hamster and guinea pig. Fly
ash was mixed with the standard laboratory diet and animals were allowed to
eat ad libitum for 3 months. Contamination of the PCDF Isomers was 1n the
1931A 7-20 06/22/86
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TABLE 7-7
Distribution of 2,3,7,8-TCDF-Derlved Radioactivity During
the First 24 hours In Liver, Fat and Skin of Guinea P1gsa»b
Time After
1.v. Injection
(hours)
1
3
7
24
Percentaqe
Liver
43
23
17
15
.1*1.
.6*3.
.8*0.
.4*3.
4
8
8
7
27.
31.
68.
55.
of Total Dosec
Fat
4* 7
4* 0
5*11
2*19
.6
.7
.3
.8
Skin
14.6*0
22.5*0
20.9*1
19.4*3
.9
.1
.8
.3
aSource: Decad et al., 1981b
bHartley guinea pigs received a single 1.v. dose of (1*C)-2,3,7,8-TCDF
0.02 ymol/kg. Three animals were killed at each time 1, 3, 7 and 24
hours, respectively.
cMean * S.D.
1931A 7-21 06/22/86
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range of 0.5-3.6 yg/g food. Rats and hamsters retained mainly the Isomers
with 2,3,7,8-substltutlon patterns whereas guinea pigs retained also a num-
ber of other substituted Isomers probably because of less effective metabol-
ism. Usually recovery 1n the liver was below 10% for all three species. In
a subsequent paper Berg et al. (1986) gave rats and hamsters oral treatments
with fly ash at higher concentrations (15% 1n food compared with the earlier
concentrations of 2.9%); they found higher liver retention, from 20-60% for
penta and hexa congeners. In rats, retention was highest for 1,2,3,7,8-
PeCDF (41%) and very low for 2,3,7,8-TCDF (1.1-2.8%); 1n hamsters the high-
est retention was for 2,3,4,7,8-PeCDF (71%) and -38% for 2,3,7,8-TCDF. The
authors suggested that the hamster probably metabolizes TCDF less efficient-
ly than the rat. As a whole, these results Indicate that the bloavallabll-
Hy for PCDF, when absorbed on fly ash, 1s very low.
7.3.3.5. PHYSIOLOGICAL DISPOSITION MODEL — When uptake, accumulation
and disposition parameters are known, pharmacoklnetlc models can be useful
to predict the overall distribution and to describe the time course of tis-
sue concentrations and excretion of metabolites. These models can provide a
means of estimating the metabolic and excretory clearance, difficult 1n a
complex In vivo system.
The disposition data for 2,3,7,8-TCDF 1n the mouse (two strains) (Decad
et al., 1982), rat (Blrnbaum et al., 1980) and monkey (Blrnbaum et al.,
1981) were compared by King et al. (1983) with simulated data generated by a
physiological pharmacoklnetlc model. In developing the model, King et al.
(1983) used t1ssue-to-blood concentration ratios from the studies previously
discussed 1n Section 7.3.3.3. to estimate partition coefficients. Metabolic
clearances were calculated from the data sets as cumulative total radlolabel
excreted (urine and feces) divided by the total area under the curve of con-
1931A 7-22 06/22/86
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centratlon In hepatic venous blood. Values of these parameters are given In
Table 7-8.
Concentrations predicted by the physiological model generally compared
well with measured concentrations In all three species. Metabolic clearance
displayed the dependence on body weight to the power of 0.7 that 1s charac-
teristic of many physiological functions. Extrapolation to man on this
basis yields a whole-body half-life estimate of 5-12 days for 2,3,7,8-TCDF.
This Indication that TCDF would not be expected to persist for long periods
1n man does not disagree with the values determined 1n tissues of patients
exposed to PCDF. In fact 2,3,7,8-TCDF was less persistent than the higher
chlorinated hexa and hepta congeners. However, the whole body half-life
seems to be greatly underestimated. In general the possibility of extrapo-
lating these pharmacoklnetlc data Is limited by the fact that metabolism
varies between species and particularly as far as PCDF metabolism 1s con-
cerned, the differences and similarities 1n the metabolic fate 1n humans and
laboratory animal species are not yet satisfactorily distinguished.
7.3.3.6. MAN — Pharmacok1net1cs of PCDFs 1n man (Including absorp-
tion, metabolism, excretion and half-lives) are unknown. However, PCDFs
have been detected 1n tissues of subjects exposed 1n different ways to these
compounds. Measurements of tissue levels of the various Isomers over time
provide a gross estimate of the extent of exposure and can help to elucidate
the structural rules governing the metabolism and elimination of these
classes of compounds. As methods are now available to detect dlbenzofuran
Isomers In human tissues In the ppt range (Albro et al., 1985), measurement
of PCDFs 1n fat biopsies can be considered a biological marker of exposure.
PCDFs have been detected In man after different types of exposure to recog-
nized sources of contamination and, very recently, also 1n the general popu-
1931A 7-23 06/22/86
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TABLE 7-8
2,3,7,8-TCDF Pharmacoklnetlc Parameters for Physiological Model3
House
C57 DBA
Distribution ratio5
Liver 100 100
Fat 25 40
Skin 8 12
Muscle 2 4
Clearances
Metabolism5 0.07 0.06
km(mi/m1n)
Metabolites excretion ratio 0.14 0.27
kk/ke
Rat Monkey
100 30
35 30
4 7
2 2
1.0 2.25
0.03 0.19
aSource: King et al., 1983
bFor parent TCDF
1931A
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latlon that 1s evidently exposed to unknown contaminations. Depending on
the higher detection limit of the method used In the first reports, 1n many
cases not Indicated, the concentrations 1n these studies are not comparable,
particularly the levels 1n control subjects, but the earlier results are
reported here when they are significant 1n the context of the scientific
merit of recent observations.
PCDFs have been found 1n tissues of Yusho and Yu-Cheng patients Intoxi-
cated In Southwest Japan (1968) and 1n Taiwan (1979), respectively by con-
sumption of a commercial rice oil contaminated with high concentrations of
PCBs, PCDFs and PCQs. Although the PCDF concentrations In the Taiwan rice
oil were lower than In the Japanese oil, (Table 7-9) It has been roughly
estimated that the average Intake during the whole Intoxication period was
similar: 973, 3.8 and 586 mg In Taiwan (Chen et al., 1985a) and 633, 3.3
and 596 mg In Japanese patients (Hayabuchl et al., 1979) for PCBs, PCDFs and
PCQs, respectively. An example of the tissue PCDF distribution with the
relative concentrations of the various Isomers detected was obtained by the
analysis of samples from a Taiwanese patient who died ~2 years after the
onset of poisoning (Chen and H1tes, 1983). The PCDF congeners retained 1n
all the tissues were essentially the same, with higher levels*1n liver. The
relative amounts of the Individual PCDF congeners differed 1n liver and
other tissues (Table 7-10). The major congeners detected were 1,2,4,7,8-
and 2,3,4,7,8-PeCDFs; 1,2,3,4,7,8-HxCDF. Minor amounts of 2,3,7,8-TCDF and
1,2,3,4,6,7,8-HpCDF were found. All the PCDFs retained have at least 3
chlorine atoms at 2,3,7,8 positions and no vicinal hydrogens in the dibenzo-
furan ring. As stated in Section 7.4., these bioaccumulative PCDFs also
strongly Induce benzo(a)pyrene hydroxylase and DT-dlaphorase and cause
severe atrophy of the thymus and significant hypertrophy of the liver in
1931A 7-25 06/22/86
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TABLE 7-9
Concentrations of PCBs, PCDFs and PCQs In the R1ce 011s from
Japan and Taiwan3
Concentration (ppm)
PCBs
PCDFs
Unknown tetra-
2,3,7,8-tetra-
2,3,4,7,8-penta-
2,3,4,6,7-penta-
hexa-
TOTAL
PCQs
Japan R1ce Oil
900
0.65
0.20
0.70
0.35
0.12
2.02
800b
Taiwan
A
60
0.04
trace
0.02
0.01
0.01
0.08
90b
R1ce Oil
6
100
0.05
trace
0.02
0.02
0.01
0.10
180b
aSource: Masuda et al., 1982
^Approximate concentration
A and B = two different oil samples
Trace = 0.001-0.005 ppm
1931A
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06/22/86
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rats (Yoshlhara et al., 1981; Nagayama et al., 1983); and these Isomers can
be considered the most toxic compounds, at least 1n animals.
No congeners with adjacent unsubstltuted carbon atoms have been detected
1n tissues of the Yu-Cheng patient (see Table 7-10). By contrast these
unhalogenated v1c1nal-C-atom congeners were present In the rice-bran oil
that the patient had Ingested (Table 7-11). A similar pattern of Isomers
had been detected by Rappe et al. (1979b) In a Japanese Yusho patient who
died 18 months after the exposure. Thus, It has been suggested that congen-
ers with at least two vicinal hydrogens In the ring are metabolized or
excreted during the Interval. Some other autopsy results (Kurokl and
Hasuda, 1978) are provided In Table 7-12. Preferential accumulation In the
liver, followed by adipose tissue sequestration for the Isomers with Cl sub-
stitution 1n the 2,3,7,8-posltlons, was also observed (Rappe et al., 1983b)
1n tissues of a Taiwan Yusho baby (Table 7-13).
A correlation between the severity of clinical symptomatology In Yusho
patients and the.estimated contaminated oil Ingestlon (PCBs + PCDFs + PCQs)
was reported promptly (Kuratsune et al., 1972; Nagayama et al., 1976) (Table
7-14). However there 1s much evidence to support the hypothesis that PCDFs
and not PCBs are responsible for the disease. Analysis of the concentra-
tions of PCDF and PCB In the liver and adipose tissue of Yusho patients and
of control subjects killed In traffic accidents revealed comparable PCB con-
centrations In tissues of the two groups, but PCDF (1n the range of ppb)
only In the organs of Yusho patients (Masuda and Kurokl, 1982). Other evi-
dence of the Importance of PCDF and PCB In determining Yusho and Yu-Cheng
syndrome has been obtained more recently. Kashlmoto et al. (1985) compared
the blood levels of Yusho (11 years after the outbreak) and Yu-Cheng
patients with that of occupationally PCB exposed workers (19 years after
1931A 7-27 06/22/86
-------
TABLE 7-10
Concentrations of PCDF Congeners 1n the Tissues of the Deceased
Patient with Yu-Cheng 1n Taiwan*
Tissue
Liver
Intestinal fat
Bronchus
Large Intestine
Heart
Stomach
Small Intestine
Kidney
Lung
Brain
Spleen
1,2,4,7,8-
3.4
0.9
0.4
0.3
0.2
0.05
0.05
0.04
0.01
0.01
0.01
Level of PCDF Conqener
2,3,4,7,8-
6.3
4.0
1.8
1.2
0.8
0.23
0.21
0.18
0.06
0.06
0.08
(ppb)
1,2,3,4,7,8-
25.4
7.8
3.2
2.3
1.4
0.40
0.34
0.32
0.12
0.15
0.10
*Source: Chen and Hltes, 1983
1931A
7-28
06/22/86
-------
TABLE 7-11
Structural Assignments of PCDF Congeners 1n the Toxic
Rice-Bran 011 Ingested by the Deceased Yu-Cheng Patient*
Structure
2,3,6,8-TCDF
2,3,4,8-TCDF (major)
2,3,7,8-TCDF (minor)
1,2,4,7,8-PeCDF
1.2,3,4,8-PeCDF
1,2,6,7,8-PeCDF
2,3,4,7,8-PeCDF
2,3,4,6,7-PeCDF
1,2,3,4,7,8-HxCDF
1,2,3,4,6,7-HxCDF
1,2,3,4,6,7.8-HpCDF
*Source: Chen and Hltes, 1983
1931A 7-29 06/22/86
-------
10
CO
TABLE 7-12
PCB and PCDF Concentrations In Liver and Adipose Tissue of Deceased Yusho Patients3
(Exposure was In May-June 1968)
1
co
o
06/22/8
Case
1
2
3
4
5
aSource
bA = 2,
B = 2,
C = 1,
D = 2,
E = 1,
Time of
Death
July 1969
July 1969
May 1972
April 1975
March 1977
Tissue
liver
liver
adipose
liver
adipose
adipose
liver
adipose
PCB
Concentration
(ppm)
0.14
0.20
2.8
0.03
4.3
0.2
0.006
1.2
PCDF Concentration (ppb)b
A B C D E Total
0.7 0.3 7.1 6.9 2.6 18.0
0.08 0.02 0.4 1.2 0.3 2.0
0.6 0.3 1.0 5.7 1.7 9.3
0.03 0.005-0.01 0.09 0.3 0.03 0.45
0.08 <0.005 0.2 0.8 0.2 1.3
0.4 <0.005 0.8 0.1 0.5 1.8
<0.005 <0.005 0.02 0.1 0.04 0.16
<0.005 <0.005 0.2 0.5 <0.005 0.7
: Kurokl and Masuda, 1978
3,6,8-TCDF
3,7,8-TCDF
2,4,7,8-PeCDF
3,4,7,8-PeCDF
2,3,4,7,8-HxCDF
+ 1,2,3,6
,7,8-HxCDF
-------
TABLE 7-13
Levels of PCDF (pg/g) and PCB (ng/g) 1n Tissue Samples of a Baby
from a Woman from Taiwan Suffering from Yusho Disease*
2,3,7
1.2,4
1,2,3
2,3,4
1,2,3
PCDF
PCB
Isomer
,8-TCDF
,7,8-PeCDF
,7,8-PeCDF
,7,8-PeCDF
,4,7,8-HxCDF
Adipose
Tissue
17
14
44
68
88
231
316
Liver
60
42
194
91
193
580
27
Muscle
ND
ND
ND
ND
ND
ND
38
Omen turn
ND
ND
ND
ND
ND
ND
64
Diaphragm
ND
ND
ND
ND
ND
ND
46
*Source: Rappe et al., 1983b
ND = Not detected
1931A
7-31
06/22/86
-------
CO
»
—J
I
CO
ro
TABLE 7-14
Relationship Between the Amount of Rice 011 Used by Yusho Patients,
Amounts of PCBs and PCDFs Ingested, and Clinical Severity3
Consumed**
Rice 011
Consumed
(no*)
<720
720-1440
>1440
PCBs
(g)
<0.7
0.7-1.4
>1.4
PCDFs
(mg)
<3.6
3.6-7.2
>7.9
Unaffected
10 (12.0)
0
0
Number of People0
Light Cases
39 (49.0)
14 (31.0)
3 (14.0)
Severe Cases
31 (39.0)
31 (69.0)
18 (86.0)
Total
80
45
21
aSource: Kuratsune et al., 1972; Nagayama et al., 1976
Calculated on the basis of the concentrations of PCBs (1000 ppm), PCDFs (5 ppm) found In the contami-
nated rice oil by Nagayama et al. (1976)
Percentage of cases Is given 1n parentheses.
o
a*
>x
no
00
a*
-------
termination) and unexposed people. In spite of high levels of PCBs 1n all
the samples, detectable amounts of PCDFs were only found 1n blood of
Yu-Cheng patients. In 113 Yu-Cheng patients there was a clear correlation
between the blood PCDF concentration and the severity of dermatologlcal
symptoms. PCQs were present In blood of all the Yu-Cheng patients 6 months
after exposure and In 54 of the 56 living Yusho patients 11 years after the
outbreak. So the presence of PCQs In blood can be considered a good marker
of past 1ngest1on of contaminated oil.
In the blood of Yu-Cheng patients there was a distinctive PCB pattern,
very different from the original pattern (Masuda et al., 1985) and richer 1n
the more chlorinated Isomers (for example, 2,3,4,5,3,4-hexa-CBs a PCB with
high bloaccumulatlve properties). This distinctive chromatogram has now
been adopted as one of the criteria for Identification of Yu-Cheng disease.
An example of the levels of three halogenated congeners 1n tissues of dif-
ferent categories subjects 1s given 1n Table 7-15.
Preliminary data In the three Taiwan Yusho patients Indicated that 1n
the first year after exposure blood concentrations of penta- and hexa-CDF
dropped 20 and 15%, respectively. Thus, the half-time of highly-chlorinated
PCDF In man appears to exceed 1 year (Rappe et al., 1983c). Rappe et al.
(1979b) actually detected reliable levels of PCOF In blood of Yusho patients
10 years after the Intoxication. Of particular Interest 1s the detection of
high concentrations (100-500 ppt) of 2,3,4,7,8-PeCDF and 1,2,3,4,7,8-HxCDF
In the placenta of Yusho women 5 years after exposure. The Implications of
these results In relation to the Increased mlcrosomal enzyme activity
detected 1n the placenta of these patients Is discussed under Section
7.4.5.5.2. (Wong et al., 1985).
1931A 7-33 06/22/86
-------
TABLE 7-15
GO
3>
ton
Case
centrations or v
and
Age Sex
IBS, Kturs ana HH/S in ine (issues or TUSPO
1n the Milk Fat of a Worker Occupatlonally
Time After Tissue
Exposure (or fluid)
(years)
caiienis,
Exposed to
PCBs
unexposeu inai
PCBs3
Concentration
PCDFs
IV lUUd 15
(ppb)
PCQs
Yusho patients
-j
CO
4*
o
^
1
2
3
4
5
Unexposed
Avg.
PCB worker
25 H
46 F
72 M
59 M
59 N
Individuals
53 M,F
37 F
1 Adipose tissue
Liver
4 Adipose tissue
Liver
7 Intestine
Liver
9 Intestine
Liver
9 Intestine
Liver
Adipose tissue
Liver
11 Milk fat
5090
226
6091
68
3472
114
3630
64
1273
18
803b
33b
6241
5.3
70.1
9.6
15.5
2.5
6.2
2.2
6.1
0.2
0.02
0.019b
0.006b
0.02
2400
218
1444
144
1770
51.7
416
6.3
24.5
0.97
1.53b
0.41b
0.2
ro
ro
CO
aSource: Hlyata et al.. 1985
bAverage concentration from 14 persons
-------
Rappe and Buser (1981) reported measurable levels of PCOF In the blood
of workers exposed to 2,3,4,6-tetra-chlorophenol and in workers exposed to
pentachlorophenol or pentachlorophenol laurate. Tables 7-16 and 7-17,
respectively, report the levels of chlorophenols found 1n urine samples and
of PCDF and PCDD In whole blood. The pattern of PCDD and PCDF 1n the blood
differed In workers exposed to 2,3,4,6-tetrachlorophenol or pentachloro-
phenol. This parallels the difference In the concentrations of contaminants
In the two products: more PCDFs were present in 2,3,4,6-tetrachlorophenol
than 1n pentachlorophenol.
Recently, Rappe et al. (1983b) analyzed samples of liver and kidney
tissue of a worker who had been employed 1n the production of phenoxy acid
herbicides. He died of a pancreatic tumor 3-4 years after the exposure
ceased. The findings confirm the prolonged storage of the hlghly-halogen-
ated PCDF Isomers and their preferential accumulation In the liver (Table
7-18). In a subject hospitalized for granulocytopenla, probably caused by
long-term exposure to PCP, levels of highly chlorinated CDF and CDD were
detected In adipose tissue (Gorskl et al., 1984). The Interest of this case
consists 1n the two adipose tissue biopsies made at a 2-year Interval. A
rough estimate of the half-lives of these congeners has thus been suggest-
ed. Data reported In Table 7-19 confirm, although only preliminarily, that
the half-lives of PCDF 1n humans exceed 1 year.
Schecter et al. (1985a,b) reported a high level of PCDF residues In adi-
pose tissue of a patient who had been Involved In the Incident of the elec-
trical transformer In the Blnghampton State Office Building (BSOB) 1n NY.
This transformer contained 65% PCB and 35% chlorinated benzenes. The soot
samples contained a mixture of PCDFs and PCDDs 1n which the most toxic
2,3,7,8 substituted congeners were present. From Table 7-20 H appears that
1931A 7-35 06/22/86
-------
10
co
TABLE 7-16
Levels of PCDD and PCDF 1n Blood Samples from Workers In the Saw Mill Industry
After Exposure to 2,3,4,6-Tetrachlorophenolatea
Number Profession
Blank A
Blank B
1 Loader0
1 Loader6
2 Cleanerd
g 2 Cleaner6
3 Loaderd
3 Loader6
5 Packerd
5 Packer6
7 Control
aSource: Rappe and Buser
bMajor Isomer 1,2,3,4,6,
o cMajor Isomer 1,2,3,4,6,
v! dSamp!1ng 6 months after
K 6Sampl1ng after 1 month
CD
Chlorophenols
In Urine
(vg/mi)
—
—
0.04
5.2
<0.02
0.23
0.03
0.83
<0.05
0.11
<0.01
, 1981
7,8-HpCDD
7,8-HpCDF
latest exposure
exposure
PCDD (pq/q blood)
octa heptab hexa penta
7 <2 <3
<2 <1 <1 <1
5 <2 <3
7 <1 <1 <1
5 2 <3 —
22 8 <1 <1
18 10 3
3 <1 <1 <1
<3 <2 <3
4 <1 <1 2
3 <1 <1 <1
octa
<3
<2
<3
<2
<3
<2
<3
<2
<3
<2
<2
PCDF (pq/q blood)
heptac hexa
2 <3
<1 <1
40 <3
22 <1
30 <3
17 <1
18 <3
17 <1
7 <3
12 <1
3 <1
penta
__
<1
—
<1
--
5
__
1
—
-------
TABLE 7-17
Levels of PCDD and PCOF In Blood Samples from Workers In the Textile and Leather Industry
After Exposure to Pentachlorophenol (PCP) or PCP Derivatives3
vO
3»
Number
3
4
5 M
6
7
53 10
15
16
17
Profession
Textile
Textile
Textile
Textile
Tannery0"
Tannery''
Tanneryd
Tanneryd
Chlorophenols
1n Urine
(vg/ml)
3.12
—
<0.01
0.42
0.16
0.55*
0.04e
0.03e
—
PCDD (pq/q blood)
octa
304
3
10
105
30
20
80
12
7
heptab hexa penta
59 <1 <1
<1 <1 <1
1 <1 <1
15 <1 <1
6 <1 <1
7 <3
30 3
4 <3
2 <2
PCDF (oq/q blood)
octa heptac
10 33
<2 <1
<2 <1
<2 <1
<2 <1
<3 7
7 18
<3 3
<3 3
hexa penta
<1 <1
<1 <1
<1 10
<1 <1
<1 1
<3
3
<3
<3
aSource: Rappe and Buser, 1981
bMajor Isomer 1,2,3,4,6,7,8-HpCDD
cMajor Isomer 1,2,3,4,6,7,8-HpCDF
o dBlood sampling 8 months after last exposure
\
£ eUr1ne sampling 6 months after last exposure
CD
-------
TABLE 7-18
Levels of PCDF 1n Tissue Samples of a Worker Exposed to
Phenoxy Herbicides3
Isomer
Concentration
Liver
(pg/g organ)
Kidney
(pg/g organ)
2,3,7,8-TCDF
2,3,4,7,8-PeCDF
1,2,3,4,7,8- + 1,2,3,6,7,8-HxCDF
1,2,3,4,6,7,8-HpCDF
OCDF
10 (2)
55 (2)
100 (4)
<3
7 (2)
8 (2)
6 (4)
<3
aSource: Rappe et al., 1983b
''The detection limits are In brackets
1931A
7-38
06/22/86
-------
the Isomers that were high 1n the patient's sample were also high In the
soots to which the patient was exposed. Hence, In comparison to the levels
of unexposed subjects, It would appear that these high levels arose through
exposure to the soot. In this case the persistence of the 2,3,7,8 substi-
tuted Isomers 1s noteworthy. On the other hand, the pattern of high levels
of hexa- to octa- chlorinated dloxlns 1n adipose tissue of both exposed and
unexposed subjects Is consistent with a more general environmental contamin-
ation.
Recent findings document the presence of PCDF and PCDD In the general
population. Ryan et al. (1985a) reported the presence of 2,3,4,7,8-PeCDF In
adipose tissues of Canadian patients at a level of 17 pg/g, confirming earl-
ier data of Mlyata et al. (1977a) who found the same Isomer 1n tissues of
controls In a study on Yusho rice oil poisoning. More recently Ryan et al
(1985b) found 8-10 PCDDs and PCDFs, all with 2,3,7,8 chlorine substitution,
In 46 adipose tissue samples from accident victims (1n Canada). Major con-
stituents were OCOO 800 pg/g), PeCDF, PeCDD, HxCDD and HxCDF. No TCDF and
OCDF were detected. Levels ranged between 17 and 39 ppt, 2-3 orders lower
than those associated with adverse effects 1n Yusho and Yu-Cheng patients.
Similar patterns of congeners were found by Rappe et al. (1984) from samples
of human Swedish control tissues. 2,3,7,8 Cl-subst1tuted PCDFs have been
detected In the general population from New York State by Schecter et al.
(1985a,b) who compared tissues of subjects unexposed and exposed to chemical
contamination (see Table 7-20). Schecter et al. (1986a) also found PCDD and
PCDF at detectable levels In the general population of South and North Viet-
nam, areas of respectively high and low potential dloxln exposure. In both
populations PCDFs were present, the North Vietnam samples showing levels
1931A 7-39 06/22/86
-------
TABLE 7-19
Half-lives and Contents of PCDDs and
PCDFs 1n Human Adipose Tissue3
1,2,3
1,2,3
OCDD
1,2,3
OCDF
Compound
,6,7,8-HpCDD
,4,6,7,8-HpCDD
,4,6,7,8-HpCDF
Content In
(ppb, on
April 1981b
0.8
4.4
5.2
0.8
1.5
Adipose Tissue
a fat basis)
November 1983C
0.5
2.6
4.0
ND
0.4
Half-Life
(years)
3.5
3.2
3.7
<1.7
1.8
aSource: Gorskl et a!., 1984
bOeterm1ned by HR 6LC-EC
Determined by GLC-EC
ND = Not detected (<0.2 ppb)
1931A
7-40
06/22/86
-------
TABLE 7-20
PCDD and PCDF Levels In Blnghamton, New York Adipose Tissue Levels (ppt) In Exposed and Unexposed Subjects
Soot Sample Levels (ppm) from the Transformer F1rea
PCDDsb
2.3.7.8-TCDO
Other (4)
1,2,3,7,8-PeCDO
Others (7)
,2.3.4.7,8-HxCDD
,2,3.6.7,8-HxCDO
,2,3,7,8,9-HxCOD
,2,3,4,6,8/1,2,4,6,7.9/1,2,4,6,8,9-HxCOO
,2,3.6,8.9/1.2,3,6,7.9-HxCDD
,2,3,4,6,7-HxCDO
,2.3,4.6,7,9-HpCDO
,2.3,4,6.7.8-HpCDD
OCDD
TOTAL
Soot
0.6
0.6
2.5
2.5
0.7
0.6
0.4
1.2
1.3
0.5
4.0
3.0
2.0
19.9
Exposed Unexposed PCDFsb
(n=l) (n=4)
11.6 7.7 2.3,7,9-TCOF
1,3,7,9-TCDF
Others
15 10
1,2,3,7,8-PeCOF
2.3,4.7,8-PeCOF
1,2,4,7.8-PeCOF
72.6 55.4 1,2.4,7.9-PeCDF
7.3 7.3 1,3,4,7,8-PeCDF
2,3.4,8,9-PeCDF
1,2,3,6,7-PeCDF
2.3,4,6.7-PeCDF
Others (12)
9.6 2
209 86 1.2.3.4.7.8-HxCOF
1.2,3,6,7.8-HxCDF
690 602 2.3,4.6,7,8-HxCDF
1,2,3,4,6,8-HxCDF
1.2,3,6,8,9-HxCDF
1,2.4,6,7,8-HxCOF
1,3,4,6,7,8-HxCOF
Others (8)
1,2,3.4,6.7.8-HpCDF
1,2,3,4,6,7.9-HpCDF
1.2,3,4,6.8,9-HpCDF
1.2.3,4,7,8,9-HpCDF
OCDF
TOTAL
Soot Exposed
(n=l)
12.0 ND
1.0
15.0
310.0
48.0 74.7
25.0
22.0
65.0
25.0
60.0
12.0
110.0
310.0 149
150.0 112
10.0
30.0
38.0
50.0
125.0
250.0
230.0 39.3
120.0
55.0
55.0 25.9
40.0 1.6
2168
Unexposed
(n=4)
ND
14.2
14.2
8.9
13.6
10
0.7
•SJ
^
CO
aSource: Buser, 1985; Schecter et al.. 1985a,b
bNumber In parentheses refer to the number of Isomers
ND = Not detected
-------
lower than In other more Industrialized countries. Comparative data are
reported 1n Table 7-21.
Further Investigations prompted Ryan et al. (1985b) to seek Information
on the distribution and levels of PCDF 1n different tissues from the same
subjects. From samples of two deceased elderly patients, they found that
PCDF levels decreased from adipose tissue to liver, muscle and kidney 1n wet
tissues, but the values were closer when calculated on the basis of I1p1d
content. In a more complete study Ryan et al. (1986) showed that between 8
and 10 PCDDs and PCDFs with 2,3,7,8 chlorine substitution occur In all tis-
sues tested, the relative amounts and specific Isomers being the same (Table
7-22). Fat shows the highest levels and 1s probably a storage tissue but on
a Upld basis, liver and other tissues have the highest levels, suggesting
these may be target organs (Table 7-23).
Recently PCDDs and PCDFs have been detected 1n breast milk from human
populations of different countries. Particularly high levels of 2,3,7,8-
TCDD In milk of a South Vietnamese population were found In 1973 samples
(Table 7-24) but 1n five 1983 breast milk samples the values were below the
detection limits (Schecter et al., 1986a). However, milk of the general
population also appears to contain highly chlorinated PCDDs and PCDFs (see
Table 7-24).
7.3.4. Metabolic Fate. No satisfactory metabolic pathway for halogenated
DF has yet been reported although the metabolism has been regarded as a
route of detoxification for PCDF (Matthews and Blrnbaum, 1983). The more
readily metabolized PCDF have adjacent unsubstHuted carbon atoms. Data Is
scant on the Identification of metabolites of PCDF Isomers; quantitative
studies are lacking. One of the few published Investigations on the metab-
olism of PCDF reports that In salmon, an unidentified hydroxylated metabo-
1931A 7-42 06/22/86
-------
TABLE 7-21
10
CO
3>
2.3.7.8-TCDO
1.2,3,7,8-PeCDO
1.2.3,4,7,8-HxCDO
1,2,3,6,7,8-HxCDD
1.2,3.7,8.9-HxCOD
T1 1.2.3.4.6,7,8-HpCDD
" OCOO
2,3,7.8-TCDF
2,3,4,7,8-PeCDF
1.2.3,4.7,8-HxCDF
1,2.3,6.7.8-HxCDF
2.3,4.6.7,8-HxCDF
1,2,3.4.6,7.8-HpCOF
OCDF
Levels (pg/g) of PCDDs and PCDFs In Human Samples from Sweden. U.S.A. and
Adipose Tissueb
Sweden [31] Canada [10] U.S.A. [6]
C A B C
3 (0-9) 10 10/10 6.4 (3.7-8.3)
10 (3-24) 13.2 10/10 9.7 (7.5-13.8)
ND
15 (3-55) 90 10/10 57.8 (46.2-64.2)
4 (3-5)
97 (12-380) 116 10/10 95.2 (30.4-119)
414 (90-763) 611 10/10 585 (428-695)
4 (0.3-11)
54 (9-84) 18.4 9/9 14.7 (10.9-17)
6 (1-15) 17.3 8/9
5 (1-13) 28.7 (15.1-52.8)
2 (1-7)
11 (1-49) 39.4 7/9 16.4 (12.5-23.8)
4
Vietnam Subjects3
South [15] North [9]
A B A B
27.9 12/15 ND
15.4 15/15 3.8 1/9
99.8 15/15 11.4 6/9
178 15/15 28.8 6/9
1326 15/15 104 8/9
21 15/15 14.7 6/9
58.3 15/15 13.3 7/9
28.8 15/15 7 3/9
CO
aSource: Nygren et al., 1986; Ryan et al., 1985b; Schecter et al., 1986a
bNumbers In brackets refer to number of samples
A = Hean of positives
B = Number of positives
C = Ranges
-------
lO
>
TABLE 7-22
2,3,7.8-ChloMne-SubstHuted PCODs and PCDFs In Human Unexposed Subject*
(Values In pg/g on a wet tissue basis)
Fat
Compound
2,3,7,8-TCDO
2.3,4,7,8-PeCDF
1.2.3.7.8-PeCOD
1.2,3.4,7.8-/
1.2,3.6,7,8-PeCDF
1,2,3,6,7,8-HxCDD
1.2.3,4.6.7.8-HpCOF
1.2,3,4,6.7,8-HpCDD
1,2,3,4,6.7,8,9-OCDD
X L1p1d
Abdominal
5.7
17
7.8
52
64
15
110
680
75
Subcutaneous
6.0
17
8.2
22
60
12
120
700
75
Adrenal
3.8
4.9
3.1
8.5
35
3.5
55
600
28
Bone
Narrow
ND
4.4
12
9.4
30
2.7
48
540
28
Liver
ND
ND
ND
4.2
6.5
2.1
22
220
6.0
Muscle
ND
1.1
1.2
2.1
7.9
ND
14
170
9.0
Spleen
ND
ND
11
ND
1.9
ND
13
46
1.8
Kidney
ND
ND
ND
2.1
2.5
ND
5.2
31
3.0
Lung
ND
ND
ND
ND
1.4
ND
2.9
21
2.2
*Source: Ryan et al., 1986
ND = Not detected
IV
IV
CO
-------
TABLE 7-23
Total PCDD and PCOF Levels (pg/g) In Tissue Samples
from an Unexposed Subject*
Tissue
Fat, abdominal
Fat, subcutaneous
Adrenal
Bone marrow
Liver
Muscle
Spleen
Kidney
Lung
%
Llpld
75
75
28
26
6.0
9.0
1.8
3.0
2.2
Total
Wet
Basis
870
890
690
630
250
190
72
39
25
PCDDs
Llpld
Basis
1160
1180
2440
2440
4220
2160
3980
1290
1150
Total
Wet
Basis
84
51
17
17
6.3
3.2
3.2
2.1
--
PCDFs
Llpld
Basis
110
68
60
63
110
36
180
70
.._
*Source: Ryan et al., 1986
1931A
7-45
06/22/86
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TABLE 7-24
to
>
2,3,7,8-TCDD
1,2,3,7,8-PeCDO
Total HxCDOs
1,2,3,4,6,7,8-HpCDD
^ OCDD
2,3,7,8-TCDF
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCOF
Total HxCOFs
1,2,3,4,6,7,8-HpCDF
OCDF
Levels of
Germany8
n=53
NDd
11.0 (<1-40)
46.3 (9-168)
48.8 (11-174)
142.8 (13-664)
4.2 (
aSource: WHO, 1986
bSource: Nygren et al., 1986
cSource: Schecter et al., 1986a
dfielow detection level (5 ppt)
eBelow detection level (2 ppt)
fflelow detection level (0.5 ppt)
QBelow detection level (10 ppt)
-------
lite Is formed from 2,8-DCDF (Z1tko et al., 1973). Veerkamp et al. (1981)
reported Investigations on the metabolism of various PCDFs 1n the rat. No
metabolites were detected In urine, feces and tissues of rats receiving
OCDF. Conversely, 2-MCDF, 2,8-DCDF and 2,3,8-TrCDF 1n rats form mono- and
dl-hydroxylated metabolites. Moreover, metabolites containing sulfur (sul-
fone, thloether) could originate from 2-MCDF and 2,8-DCDF. Hydroxylatlon
can take place at various positions 1n the rings. Five monohydroxy deriva-
tives have been found from 2,8-DBCF, the most Important probably being
2-chloro-8-hydroxy-DF, that can originate from the 1,2 or 2,3 corresponding
epoxlde.
A molecular model developed by Veerkamp et al. (1983) predicted PCDF
metabolites. In contrast to the corresponding PCDD 1n which hydroxylatlon
takes place only In the 2 and 3 positions PCDFs can give rise to many
products depending on the chlorine substitution pattern. Recently the
metabolism of 2,3,7,8-TCDF was studied by Polger et al. (1984) In the rat,
unfortunately using a compound containing considerable amounts of other
PCDFs. B1le was collected for 48 hours; no TCDF or other PCDFs were detect-
ed 1n the extract. From the 13 chlorinated metabolites present, which
accounted for 8.6% of the total dose, four methoxylated compounds that prob-
ably originated from the corresponding hydroxylated species were present 1n
the major amounts. They were Identified as tr1chloromethoxy-DF, tetra-
chloromethoxy-DF and two tr1chlorod1methoxy-DF Isomers. The remaining 9
compounds wee detected In traces and their origin (from the parent compound
or Impurities) remains unclear.
7.4. ACUTE, SUBACUTE AND CHRONIC TOXICITY
7.4.1. Acute ToxIcHy. PCDFs are all toxic and some are acutely toxic.
The toxlclty depends on the number of Cl substltuents and on their posl-
1931A 7-47 06/22/86
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Hon. The tetra-, pentaand hexaderlvatlves are the most toxic. The most
acutely toxic PCOFs are the PCOF congeners substituted In the lateral 2, 3,
7 and 8 positions.
There Is a long latent period between administration and the first signs
of toxlclty; the LD,.- must be established 30 days after treatment. The
DU
main signs 1n monkeys and guinea pigs were severe weight loss, atrophy of
the thymus and spleen, debilitation of the lymphatic system, hemorrhage of
the adrenals and urinary bladder, and single-cell necrosis 1n the liver.
Hlstologlcal examination of tissues revealed loss of lymphold cells 1n the
thymlc cortex and hyperplasla of epithelial cells In the renal pelvis,
ureter and urinary gladder. These lesions, together with a lack of liver
pathology, hypocellularlty of bone marrow, seminiferous tubules, lymphold
elements 1n spleen and Peyer's patches, were similar to those described for
guinea pigs given large doses of PCDD (McConnell et a!., 1978b; McConnell
and Moore, 1979).
The toxic effects of PCDFs are markedly species-dependent, as are those
of PCODs. The guinea pig Is the most sensitive species followed by the mon-
key, rabbit, rat and mouse. In the guinea pig, the ID™ for 2,3,7,8- TCOF
1s between 5 and 10 yg/kg bw (McKlnney and McConnell, 1981). The same
authors showed that there was no significant different In acute toxlclty
(LDcn) between 2,3,7,8-TCDF, 2,3,4,7,8-PeCDF (< 10 v9/kg) and 2,3,7,8-
Du
TBDF, while the LD50 of 2,3,4,7,8-HxCDF was 120 vg/kg (Moore et al.,
1979; McKlnney and McConnell, 1981). The rat Is quite resistant to the
acute toxic effects of 2,3,7,8-TCDF. The monkey manifests Intermediate sen-
sitivity.
1931A 7-48 06/22/86
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These differences depend on the species' ability to metabolize and
excrete the parent compound. Since 2,3,7,8-TCDF has a whole-body halftlme
of <2 days 1n the rat, this species Is quite resistant to the acute toxic
effect. The guinea pig, on the other hand, 1s extremely sensitive (an
LD__ between 5 and 10 yg/kg) because the compound has a wholebody half-
time of at least 20 days. The monkey has Intermediate sensitivity because
the whole body half-time Is ~8 days. The pathological effects of acute
treatment with 2,3,7,8-TCDF 1n the rhesus monkey are summarized 1n Table
7-25. Table 7-26 sets out the LD™ for 2,3,7,8-TCDF 1n several animal
species.
Mixtures of PCDFs were often used 1n toxlclty studies. Mixtures of
TrCDFs and TCDFs given orally to rabbits at doses of 0.4 and 1 mg/kg (Bauer
et al., 1961) caused severe liver necrosis and often proved fatal.
7.4.2. Subacute Toxlclty. The most complete data relate to 2,3,7,8-TCDF.
Mice given 22 oral doses over 30 days of 30, 200, 300 yg/kg bw of 2,3,7,8-
TCDF did not develop clinical signs of toxlclty (Moore et al., 1979).
Autopsy showed "liver weights and the liver we1ght/bw ratio elevated, and the
thymus weight/bw ratio was low.
When 2,3,7,8-TCDF was fed to chicks for 21 days at doses of 1 and 5
vg/kg bw/day, mortality was 16 and 100%, respectively (McKlnney et al.,
1976). 2,3,7,8-TCDF at 5 yg/kg bw/day affected the liver moderately and
caused thymlc Involution and edema.
2,3,7,8-TCDF caused sickness and some deaths In two groups of three
rhesus macaques, fed for 6 and 2 months with 5 and 50 yg/kg diet (corres-
ponding to 5 and 50 ppb) (McNulty et al., 1981). The principal pathological
changes were trophy or squamous metaplasia of the sebaceous glands, Involu-
tion of the thymus and hypoplasla of the bone marrow. In animals that did
1931A 7-49 06/22/86
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TABLE 7-25
Summary of Pathologic Effects of 2,3,7,8-TCOF 1n Rhesus Monkeys*
Tissue
Gross Effects
Microscopic Effects
Skin and adnexa
Fat
Lymphold
Liver
B1le duct
Intestine
Clinical
pathology
Facial edema, dry skin,
loss of eyelashes, loss
of finger and toe nails,
streaks on ocular surface
of eyelids, and occlusion
of external ear canal
Moderate to severe loss
None
No effect
Markedly thickened
Hyperemla, petechlal
hemorrhage, focal ulcera
tlon and mucosal cysts
Hyperkeratosls; squamous
metaplasia, cystic
dilation of glands
None
Thymlc atrophy from
lymphold cell loss;
moderate atrophy of
lymphold cells In spleen
and lymph nodes
No lesions observed
Hypertrophy of biliary
duct and gall bladder
epithelium. Increased
mucous secretion
Loss of parietal cells,
Increased mucous cells,
mlcrocystlc dilatation
of crypts
Anemia, low serum
cholesterol, mild
lymphopenla, neutro-
phH1a, low serum
protein and albumin,
elevated SGOT, BUN
*Source: Moore et al., 1979
1931A
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06/22/86
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TABLE 7-26
Acute Tox1c1ty of 2,3,7,8-TCDF 1n Several Animal Species
Animal
LD50
Ug/kg bw)
Interval Between
Administration
and Death
(days)
Reference
Chicken
Guinea pig
ND = No data
<5
>5
21
9-20
McKlnney
et al., 1976
Moore et al.,
1979
Rat
Monkey
Mouse
6000
1000
>6000
ND
14-31
ND
Moore, 1975
Moore et al.,
1979
Moore, 1975
1931A
7-51
06/22/86
-------
not die during the feeding, recovery was complete after 3 months of a diet
free of 2,3,7,8-TCOF. It was suspected that the toxic effects of 2,3,7,8-
TCDF might accumulate above a certain critical dose and might be similar
whether the substance was administered acutely or Ingested gradually over
weeks or months.
Male guinea pigs given 2,3,7,8-TCDF 1n six or more weekly doses of 1
yg/kg bw showed Irreversible toxldty when a body burden of -5.6-6.6
yg/kg bw was attained, resulting In progressive weight loss and death
(Decad et al., 1981b). A later paper from loannou et al. (1983) reported
that a single dose of 4 yg/kg or multiple doses of 1 yg/kg for 4 weeks
did not produce any observable toxldty by day 36; when animals were killed
the authors estimated the body burden of these animals to be 2.1 and 2.8,
respectively. A body burden two or three times higher resulted 1n the deaths
of three of the four animals treated. These data Indicate that, despite
differences 1n Individual sensitivity, PCDF toxlclty appears only after a
critical body burden 1s reached. Similar results were obtained by Luster et
al. (1979) In female guinea pigs.
Inbred C57B1/6N mice, treated 1.p. with 2,3,4,7,8-PeCDF (30 yg/kg once
a week for 6-12 weeks) developed hlstopathologlcal signs 1n the liver, thy-
mus and spleen. In an "unresponsive" strain, the changes 1n these organs
were generally mild (Nagayama et al., 1985a).
Administration to rats of an unspecified mixture of PCDF (TCDFs, PeCDFs
and HxCDFs) at a concentration of 10 ppm In the diet (corresponding to 0.5
mg/kg bw) for 4 weeks Induced chloracne-Uke lesions on the ears, loss of
body weight, low hematocrlt and hemoglobin values, Increased serum choles-
terol and liver llplds, and low serum trlglycerldes. At 1 ppm In the diet
these effects were less marked (01sh1 et al., 1978).
1931A 7-52 06/22/86
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A summary of the subacute toxic effects of PCDF 1n various animal spe-
cies 1s provided 1n Table 7-27.
7.4.3. Chronic Toxlclty. Planned studies of toxic effects of chronic
treatment with PCDFs are lacking. McNulty et al. (1981) reported that
2,3,7,8-TCDF administered to three male rhesus macaques In the diet (5 ppb)
for 6 months caused sickness and two deaths. The major pathological changes
were atrophy or squamous metaplasia of the sebaceous glands, mucous meta-
plasia of the gastric mucosa, Involution of the thymus and hypoplasla of the
bone marrow. Animals that did not die during TCDF dosing recovered com-
pletely after 3 months of TCDF-free diet (see Section 7.4.2.). Because of
the small number of animals employed, these findings are preliminary. How-
ever, It 1s apparent that some cumulative action does occur, and H may be
reversible to some extent. Death 1n the monkeys was preceded by weight
loss, anorexia and depression, but no specific chemical lesion was Identi-
fied.
A study was made using monkeys fed a 20-week diet containing a mixture
of PCBs, PCQs and PCOFs with congeners similar to those found In Yusho oil,
or a mixture of PCBs or PCQs, or commercial PCB (Kanechlor 400). Only the
first group developed hair loss, edema of the eyelids, acneform eruptions,
dermal hyperkeratosis and pigmentation (Kunlta et al., 1985). These data
confirm PCDFs Involvement 1n the Yusho disease.
7.4.4. Contribution of PCDF to the Toxldty of Industrial Products and
Environmental Contaminants. PCDFs are released In the environment as con-
taminants of Industrial products and must be considered as adventitious pol-
lutants. Several studies have compared the toxlclty of Industrial products
contaminated with PCDFs with that of pure active Ingredients. Vos and Koe-
man (1970) found significant differences in the toxlcltles of three commer-
1931A 7-53 06/22/86
-------
TABLE 7-27
Subacute Toxlclty of PCDFs In Laboratory Animals
-. J
1
o
N:
CO
cr
Compounds
2,3.7.8-TCDF
2-1somers-TCDF
4-1somers-PeCDF
4-1somers-HxCDF
2.3.7,8-TBDF
2,3.4.7.8-PCDF
2,3.7,8-TCDF
PCDFs
2.3,7,8-TCDF
2.3,7,8-TCDF
2,3,7,8-TCOF
Species/
chick
Sprague-
Dawley
rat
C57B1
mouse
mouse
adult
rhesus
macaque
(3)
mature male
guinea
pig («)
Immature
female
guinea pig
Dose
(pg/kg/day)
1 (21 days)
5 (21 days)
50 (28 days)
500 (28 days)
30-300
(30 days)
50
(10 weeks)
90
(6 months)
300
(2 months)
1 pg/kg bw week
(6-7 weeks)
1 pg/kg bw week
(6 weeks)
Mortality
W
16
100
none
none
none
none
67
33
50
30
Route
gavage
1 ppm
10 ppm
diet
gavage
0.6 ppm
diet
5 ppb
50 ppb
diet
oral
oral
Intubation
Clinical Pathology
Edema; thymlc Involution; slight
or no liver damage; no porphyrla;
no ALA Induction
Thymlc Involution; dermal lesions;
hypercholesterolemla; hyper 11 pi d-
emla; hemolytlc anemia; thrombopenla
Thymlc Involution; hepatomegaly
Dermal lesions; hyper keratosls;
dilated hair follicles with kera-
tlnous material; damaged hepatocytes
Atrophy or squamous metaplasia
sebaceous glands, mucous metaplasia
and hyperplasla of the gastric
mucosa; Induction of the thymus;
hypoplasla of the bone marrow
Bald patches, dehydration, swelling
around eyes, no hepatomegaly
Hepatomegaly and thymlc atrophy
Reference
HcKlnney
et al., 1976
Olshl et al.,
1978
Hoore et al.,
1979
Nagayama
et al., 1979
McNulty
et al., 1981
Decad et al.,
1981
Luster
et al., 1979
-------
clal PCBs with Clophen A-60 and Phenoclor DP-6 causing the highest, and Aro-
clor 1260 the lowest mortality 1n chicks fed for 60 days diets containing
400 ppm of each product. Subcutaneous and abdominal edema and centrllobular
liver necrosis were recorded only In chicks fed Clophen or Phenoclor.
Hydroperlcardlum was recorded In nearly all chicks fed Clophen or Phenoclor
but only occasionally In chicks fed Aroclor. Chemical porphyrla was attrib-
uted to a direct effect of the PCB. Chemical analyses revealed the presence
of polar compounds In the 25% dlethyl ether fraction of Clophen and Pheno-
clor. The polar fraction of Clophen was toxic 1n the chick embryo assay,
and the differences In toxldty of the three PCB preparations were confirmed
by this assay. Mass spectrometry analyses Indicated the presence of
tetra- and penta-CDF 1n the polar fraction of Clophen and Phenoclor (Vos et
al., 1970).
Later, Vos and Notenboom-Ram (1972) compared the toxic effects In rab-
bits of Aroclor 1260 (which was shown to contain 1 yg PCDF/g) and of a
pure PCB Isomer, 2,4,5,2',4',5'-hexachloroblphenyl. The rabbits were given
120 mg of Aroclor and the hexachloroblphenyl dermally 5 times a week for 28
days. The animals receiving Aroclor 1260 showed more severe skin lesions,
hyperplasla and hyperkeratosls of the folUcular and epidermal epithelium
than those receiving the pure hexachloroblphenyl. Liver damage (subcapsular
and zonal necrosis, hydropic degeneration, peripheral and perlnuclear shift
of cell organelles, focal cytoplasmlc hyaline degeneration, proliferation of
smooth endoplasmlc retlculum) and Increased fecal coproporphyrln levels were
similar 1n both groups. From these studies, the probable contributions of
PCOF and pure PCB to the toxldty of crude preparations were assessed
(Table 7-28).
1931A 7-55 06/22/86
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Goldstein et al. (1978a) showed that porphyrla, cutaneous lesions,
changes 1n liver enzymes and morphological changes 1n the liver were Identi-
cal 1n female CD rats (Charles River) fed pure hexachlorobenzene contaminat-
ed with 4 ppm OCDF and decachloroblphenyls for 4 months, Indicating that
these changes were produced by hexachlorobenzene rather than by the contami-
nants.
Klmbrough and Under (1978) compared the toxic effects of purified and
technical grade pentachlorophenol (PCP) In Sherman rats given dietary con-
centrations of up to 500 ppm of each compound for 8 months. At 500 ppm,
technical PCP had severe effects on the livers of female rats, mainly caus-
ing degeneration of liver cells and bile duct proliferation. In male rats
the alterations were less marked. At 100 ppm, similar but less pronounced
effects were observed; only mild alterations were noted at 20 ppm. At 500
ppm, purified PCP 1n the diet caused slight enlargement of liver cells with
occasional eosinophlUc cytoplasmlc Inclusions; no alterations were observed
1n the livers of rats fed 100 and 20 ppm. These results suggest that most
of the toxicity associated with feeding technical PCP to rats at these con-
centrations stems from toxic contaminants rather than from PCP itself.
Technical PCP has been shown to be contaminated with PCDD and PCDF (Gold-
stein et al., 1978b) but the contributions of the various Isomers to the
toxic effects of PCPs have yet to be established.
The toxicity of a soot that contaminated an office building in Bingham-
ton (NY) after a fire Involving an electric service transformer and its
dielectric fluids that was composed of Aroclor 1254 (65%) and chlorinated
benzenes (35%) with some trace additives, was studied; the soot was found to
contain PCDO (2.8 ppm) and 2,3,7,8-TCDF (124-273 ppm) (Smith et al., 1981)
and a tentative estimate of the total amount of PCDF was In the range of
1931A 7-56 06/22/86
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TABLE 7-28
Probable Contributions of PCDF 1n Aroclor 1260 and Pure
2,4,5,2',4',5'-Hexachlorob1phenyl to the Tox1c1ty In Rabbits
of PCBs Applied at 120 mg/50 cm2*
Dose Chloracne Edema Liver Hepatic
Damage Porphyrla
Aroclor (with PCDF) n- t+ tt
PCB (PCDF-free) + - +• n-
*Source: Vos and Notenboom-Ram, 1972
1931A 7-57 06/22/86
-------
0.5%. In guinea pigs the oral LD5Q of the soot (0.75% aqueous solution)
was 410 mg/kg with an equivalent of 0.5 and 20 vg/kg of 2,3,7,8-TCDD and
TCDF, respectively. On the basis of the reported respective LD5Q (Huff et
al., 1980) the toxlclty could not arise only from these halogenated congen-
ers but probably came from other pollutants as well (SUkworth et al.,
1982). After acute treatment with the soot serum, trlglycerldes were ele-
vated; alkaline phosphatases decreased; body, thymus and kidney weight
decreased; and hyperplasla of the pancreatic duct developed. Guinea pigs
fed soot developed liver steatosls and smooth endoplasmlc retlculum prolif-
eration and mltochondrlal ultrastructural alterations (Turner and Collins,
1983, 1985). Dermal application to the rabbit at a dose equivalent to 500
mg/kg for 24 hours caused no overt toxlclty, although hepatic centrllobular
hypertrophy was observed. For subchronlc studies soot was administered to
guinea pigs for 90 days, mixed Into food at five dose levels between 0.2 and
231.5 ppm. Mortality reached 35% at 231.5 ppm and 30% at 46.3 ppm. No
effect attributable to soot exposure was noted In animals receiving 0.2 ppm
soot. Toxic effects were similar but rose at slightly lower total doses
than with acute exposure (DeCaprlo et al., 1983).
The l.p. toxlclty In Immature Wlstar rats (lOOg) of a PCDF composition
the same as that found 1n Yusho oil was compared (Bandlera et al., 1984a)
with that of the corresponding PCB composition; 20% weight loss and 50%
reduction In thymus weights relative to controls was caused by 398 mg PCB/kg
bw. The PCDF mixture was as follows: 2,3,7,8-TCDF, 7.4%; 1,2,4,7,8-PeCDF,
6.1%; 1,2,3,7,8-PeCDF, 19%; 2,3,4,7,8-PeCDF, 29.4% and 1,2,3,4,7,8-HxCDF,
39.1%.
The PCQ contaminants In Yusho oil were as toxic as PCBs In eliciting
Yusho symptoms (Hor1 et al., 1978, 1982). Thus, PCDFs are the most probable
1931A 7-58 06/22/86
-------
causal agents of Yusho. This was also shown (Kunlta et al., 1985) explldty
by experiments on rats and monkeys.
7.4.5. Some Particular Aspects of the Toxic Reactions.
7.4.5.1. DERHAL TOXICITY — Chloracne can be produced by a number of
chlorinated aromatlcs and has been observed among workers In contact with
these compounds. Chlorinated naphthalenes, PCB, PCDD and PCDF can all
Induce chloracne In man. Chloracne Is described as the formation of come-
dones with or without cysts and pustules. In chloracne the folUcular ori-
fices become clogged with sebaceous and keratlnous material. Hyperkeratosls
of the epidermis, cystic dilatation of the hair follicles, and an Increase
In melanin pigment are fairly common findings. Chloracne can be Induced by
external contact with these chemicals or by systemic absorption.
Investigations In animals have shown that different species develop dif-
ferent types of skin lesions on exposure to chloracnegenlc compounds. Rab-
bit ear skin provides a good test for the development of acneform derma-
titis. Application of a mixture of IrCDF and TCOF to the rabbit ear result-
ed In hyperplasla and hyperkeratosls (Bauer et al., 1961). Yos and Beems
(1971) reported that application to rabbit skin of a 25% d1ethylether-ex-
tracted polar fraction of Phenoclor and Clophen (Identified as consisting of
TCDFs and PeCOFs) caused hyperplasla and hyperkeratosls of the folUcular
epithelium. The fraction from Phenoclor was more active than that from
Clophen.
Vos and Notenboom-Ram (1972) compared the skin toxldty of Aroclor 1260
with 2,4,5,2',4',5'-hexa-CB. Dermal application of 120 mg PCB/50 cm2 5
times a week for 4 weeks to the shaved backs of rabbits resulted 1n early
macroscopic skin lesions In the Aroclor group, but not In the group given
the pure PCB (see Table 7-28).
1931A 7-59 06/22/86
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TCDFs and PeCDF Induced hyperkeratosls of the skin when applied to rab-
bit pinna for 3 days. When dally doses of 4 yg of 2,3,7,8-TCDF were
applied to the rabbit ear for 5 days, the animals developed hyperkeratosls
of the treated ear (Klmbrough et al., 1978). Hyperkeratosls was also pro-
duced 1n hairless mice fed rice oil contaminated with Kanechlor 400 (Inagaml
and Koga, 1969). Ear lesions resembling chloracne were reported within 3
weeks In Sprague-Dawley rats fed a mixture of two tetra- , four PeCDFs, and
four HxCOFs (Olshl et al., 1978) at a dose of 10 ppm PCDF 1n the diet and 1n
mice fed 0.6 ppm PCDF (Nagayama et al., 1979). No acnelgenlc response In
the rabbit ear bloassay was reported with 2,8-DCDF and 2,4,8-TrCDF (Kodba
and Cabey, 1985).
The dermatologlcal severity In Yu-Cheng patients 0.5 years after expo-
sure was related (Kashlmoto et al., 1985) to the presence of PCDF, PCQ and
PCB In blood, but workers exposed to similar amounts of fresh PCBs showed
only mild clinical signs. Investigations on 10 female Cynomolgus monkeys
(Kunlta et al., 1985) Indicated that dermal symptoms characteristic of Yusho
could be Induced after 16-20 weeks by oral administration of PCDF from Kane-
chlor 400 (20 vg/day), or PCB (5 mg/day) plus PCDF (20 yg/day) but not
by PCB alone (5 mg/day), or by PCQs (5 mg/day).
7.4.5.2. ULTRASTRUCTURAL LIVER ALTERATIONS — A number of character-
istic ultrastructural alterations 1n hepatic cells have been seen 1n rats
and monkeys In response to dloxlns and PCBs and In guinea pigs fed with BSOB
soot (Klmbrough et al., 1972; Burse et al., 1974; Sllkworth et al., 1982;
Turner and Collins, 1985). They consisted of I1p1d droplet formation,
Increased smooth and rough endoplasmlc retlculum and mitochondria! altera-
tions. Mitochondria! lesions In animals seem to be reversible and tend to
return to their usual structure with the passage of time.
1931A 7-60 06/22/86
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In humans similar mltochondrlal structural alterations have been report-
ed after exposure to chlordecone (Guzellan et al., 1980), to PCDFs + PCB 1n
the Yusho Incident (Yamamoto et al., 1971; Hlrayama et al., 1969), to PCB,
PCDD, PCDF, naphthalenes and blphenylenes 1n the BSOB electrical transformer
fire (Schecter et al., 1984).
From these studies mltochondrlal alterations (mltochondrlal pleomorph-
1sm, giant mitochondria, arrangement of the mltochondrlal crlstae parallel
rather than perpendicular to the mitochondria! axis) seem the most specific
ultrastructural lesions for humans exposed to PCB and furans, while changes
In endoplasmlc retlculum and Intracytoplasmlc llpld droplets are frequently
seen after toxic damage following exposure to nonchloMnated hepatotoxlc
chemicals (Schecter et al., 1984). Therefore, ultrastructural analysis of
livers, together with the analysis of fat biopsies for their chlorinated
chemical levels, may provide a useful biological marker 1n evaluating human
exposure to dloxln and related compounds (Schecter et al., 1985b). The role
of PCDF 1n these ultrastructural morphological lesions remains to be clari-
fied.
7.4.5.3. PORPHYRIA ~ In birds and mammals, exposure to PCB and PCP
causes changes 1n hepatic porphyrln synthesis, producing a form of hepatic
porphyMa similar to human prophyMa cutanea tarda (Goldstein et al., 1973,
1975, 1977a, 1978a). Disturbances of porphyrln biosynthesis have been con-
nected with an Increase In the activity of the rate limiting enzyme, ALA
synthetase (Goldstein et al., 1976). PCDFs have been shown not to cause
hepatic porphyMa and to Induce only slight ALA synthetase activity In
laboratory animals. Goldstein et al. (1976) found that 2,3,7,8-TCDF did not
Induce ALA synthetase In chicks after 21 dally doses of 1 yg/kg bw and did
not produce porphyrla even at the lethal dose of 5 pg/kg bw/day, whereas
1931A 7-61 06/22/86
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specific PCB produced both effects. In rats, a mixture of PCDF (2 TCDFs, 4
PeCDFs and 4 HxCDFs) at dietary concentrations of 10 and 100 ppm for 4 weeks
only slightly Induced ALA synthetase activity and slightly Increased uro-
porphyMns (Olshl and Hlraga, 1978). 2,3,7,8-TCDF did not cause porphyrln
accumulation In livers of mice up to 28 days after a dose as high as 4 mg/kg
bw (Goldstein et al.. 1974).
The lack of effect of PCDF on porphyrln accumulation 1n the liver had
already been emphasized by Yos et al. (1970), who observed that three com-
mercial 60% chlorinated PCB mixtures (Phenoclor DP-6, Chlophen 1-60, and
Aroclor 1260) Induced a similar degree and type of porphyrla although the
degree of contamination by PCDF varied for these formulations. Chlophen and
Phenoclor contained TCDFs and PeCDFs 1n amounts responsible for other toxic
effects. These findings suggest that PCDFs, at least In the experimental
conditions tested, do not play an Important role 1n the porphyrogenlc
effects of the Industrial chemicals they contaminate.
Based on some common mechanism of action of PCDF with PCDD and other
porphyrogenlc chlorinated compounds, the hypothesis that PCDF can also
elicit porphyrla cannot be ruled out, although this has not yet been experi-
mentally established.
7.4.5.4. IMMUNOSUPPRESSION -- Few studies have aimed at finding
alterations of the Immune system by PCDFs 1n laboratory animals. Toxic
doses of 2,3,7,8-TCDF and 2,3,4,7,8-PeCDF and 2,3,7,8-TBDF have been shown
to cause severe thymlc atrophy 1n the guinea pig and mouse (Moore et al.,
1979). Lymphopenla, atrophy of the thymlc cortex, and reduction In the num-
ber of germinal centers In the spleen appeared In chicks dosed with 1-5
ug/kg bw for 21 days (McKlnney et al., 1976). Studies 1n the female
guinea pig (Luster et al., 1979) showed that the effects of 2,3,7,8-TCDF on
1931A 7-62 06/22/86
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the Immune system resemble those of 2,3,7,8-TCDD. Depression of cell-medi-
ated responses was found only at higher dosages tested (0.5-1 yg/kg bw
weekly for 6 weeks). In this animal species the LD5Q Is 5-10 vg/g.
Humoral Immunity was only slightly depressed. These results were Interpret-
ed by the authors as Indicating possible marked effects of PCDFs on the
Immune system. As for 2,3,7,8-TCDD, consequences would be severe If
2,3,7,8-TCDF were administered during the developmental phase of the Immune
response.
Single doses of 2,3,7,8-TCDF have been tested 1n mice. Reduction In
thymus weight and splenocyte numbers were observed with relatively low doses
of 2,3,7,8-TCDF given either orally or 1.p. (100 vg/kg bw) to C57B1/6
mice, where the LD5Q for 2,3,7,8-TCDF Is reported to be >6000 vg/kg bw
(Moore et al., 1979). Humoral antibody production was significantly
Inhibited with a dose-response curve similar to that of 2,3,7,8-TCDD, but
2,3,7,8-TCOF was -30 times less active than 2,3,7,8-TCDD, the latter being
as suppresslve after 6 weeks as after 1 week, while recovery from the
effects of 2,3,7,8-TCDF was complete by 6 weeks (Vecchl et al., 1983a).
The effects of a mixture of TCDD and TCOF on Immune responses (thymus
weight and humoral antibody production) were Investigated using the toxins
at different ratios (TCDD:TCDF = 1:1; 1:10; 1:100 pg/kg) to cover differ-
ent "natural" conditions of exposures (Vecchl et al., 1985). TCDF given
alone at each of the three doses used had no effect on thymus weight; added
to the TCDD, 1t did not modify Us effect on thymus weight. Work now In
progress shows that an additive depressive effect appears at the 1:100 ratio
1n antibody production.
1931A 7-63 06/22/86
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Increased susceptibility to endotoxln has been described In mice treated
orally with a 1CDF (12%) and PeCOF (88%) mixture once a week for 4 weeks
(Oishi and Hiraga, 1980).
2,3,7,8-TCDF-1nduced thymlc atrophy and Immunosuppresslon were more
marked 1n mouse strains sensitive to Induction of AHH by 3-MC or by
2,3,7,8-TCDO than In strains genetically more resistant to Induction, as
described for 1CDD (Vecchl et al., 1983b), suggesting that the Individual
genetic background may play a role In the degree of PCDO- and PCDF-1nduced
1mmunotox1c1ty.
Recent data have shown a positive correlation between AHH Inducing
ability (see Section 7.4.6.) of different PCDFs given as single doses, and
thymlc Involution (Yoshlhara et al., 1981). Studying jn vivo and in vitro
structure-activity relationships, Mason et al. (1985) found a good linear
correlation (v = 0.88) for 15 PCDF between thymlc atrophy and ]n vitro AHH
Induction. Thus, toxlclty of DFs to the Immune system could be related 1n
some way to their Inducing potential.
Interestingly, there are also reports (Kunlta et al., 1985; Hor1 et al.,
1981) that, In female monkeys, commercial PCB preparations (Kanechlor 400)
specifically treated to eliminate contaminant PCDF, given In the diet for 20
weeks, were more Immunosuppresslve on humoral antibody production than the
commercial PCB with PCDF Impurities or normal commercial PCB preparations.
An antagonistic effect regarding Immunosuppresslon and enzyme 1nduc1b1l1ty
In mice was reported by Rlzzardlnl et al. (1983) who administered to C57B16
mice an artificial mixture of 2,3,7,8-TCDD and 2,3,7,8-TCDF at the ratio of
about 1:8 and found that the mixture elicited a significantly lower effect
on antibody production (50% Inhibition) than TCDD alone (80% Inhibition).
These data seem to be preliminary and need further Investigation. Converse-
1931A 7-64 06/22/86
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ly, other reports suggest that the simultaneous presence of PCDF and PCDD In
BSOB soot with the approximate composition of 1.2 ppm TCDD, 48 ppm TCDF and
0.5% PCB may have synerglstlc effects with regards to toxldty In guinea
pigs (Sllkworth et al., 1982).
Male Sprague-Dawley rats (Nakan1sh1 et al., 1985a) given 1.14 mg PCDF/kg
bw by gastric Intubation 3 times/week or 6 t1mes/2 weeks showed necrosis of
the Clara cells in the Itmg bronchioles and marked atrophy of the thymus,
whereas similar treatment with 11.4 mg Kanechlor 400/kg bw had only mild
effects. Male Sprague-Dawley rats and female Cynomolgus monkeys (Kunlta et
al.. 1985) given 67 vg PCDF from Kanechlor 400/kg bw and 8 vg PCDF/kg
bw/day, respectively, over 22-day and 20-week study periods presented Immun-
osuppresslon and thymlc atrophy. The PCBs from Kanechlor 400 were much less
potent than the PCDFs.
Recently Kochman et al. (1985) found, In human subjects exposed to PCDF
as a consequence of a PCB transformer fire and of the dispersion of the soot
1n the environment, an Increase In a specific subpopulatlon of T suppressor
cells, although the total suppressor as well as the total helper cells were
similar 1n control and 1n exposed subjects.
7.4.5.5. ENZYME INDUCTION —
7.4.5.5.1. Introduction —The mechanism of toxlcity of PCDFs has
been extensively studied but has not yet been elucidated. These compounds
are potent Inducers of one of the hepatic mixed function oxldases, AHH, that
metabolized precarclnogens such as benzopyrene. Investigations both jji
vitro and jjn vivo to clarify the biochemical basis of the Inductive effect
of TCDD and related polyhalogenated congeners have led to the Identification
of a macromolecular species 1n hepatic cytosol of rodents that binds these
compounds specifically with high affinity (Poland et al., 1976). The cyto-
1931A 7-65 06/22/86
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sollc protein 1s known to act as a receptor for AHH Inductive activity. No
evidence 1s currently available of direct Involvement of the Induction of
AHH with any sign of toxlclty Induced by these compound.s; however, the bind-
ing to the Ah receptor as a mediator of toxlclty 1s now accepted from the
data available on structure/activity studies and genetic segregation of
toxlclty with the Ah locus (Roberts et al., 1985).
7.4.5.5.2. AHH Induction — PCDFs are potent Inducers of the hepatic
mlcrosomal n-monooxygenase system (Goldstein et al., 1978b; Poland et al.,
1976; Kawano and Hlraga, 1978). This enzyme complex (also called hepatic
mlcrosomal drug-metabolizing enzymes) metabolizes drugs and other foreign
compounds. It consists of NADPH-cytochrome P-450 reductase and cytochrome
P-450, the terminal component that contains the active site of the enzyme
and determines substrate specificity. There are several distinct Isoenzymes
of cytochrome P-450 1n liver mlcrosomes that have different substrate speci-
ficity and are Induced differently by a number of compounds.
Compounds that Induce hepatic mlcrosomal monooxygenase activities can be
divided Into two major classes: one 1s typified by phenobarbltal that
Induces several subspecies of cytochrome P-450 and associated monooxygenase
activities directed toward a wide variety of substrates; the other 1s repre-
sented by 3-methylcholanthrene (3-MC) that Induces distinct Isoenzymes of
cytochrome P-450 termed respectively 1n the rat cytochrome P-450C or P-450d
(Thomas et al., 1983) and In the mouse P,- and P -P450 (Nebert and
I O
Neglshl, 1982). These Isoenzymes are associated with enzyme activities for
a more limited group of substrates and AHH activity Is one of them. The
PCDFs belong to the same class of Inducers as 3-MC.
1931A 7-66 06/22/86
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In the rat (female CD strain, Charles River), 2,3,7,8-TCDF (89% pure and
2% unspecified PeCDF) given for 3 days at doses of 0.1-2.5 vg/kg bw/day
was a potent Inducer of hepatic drug-metabolizing enzymes (Goldstein et al.,
1978b). The approximate ED5Q for AHH Induction In the rat (0.5 yg/kg bw
x 3 days) Is roughly double that reported for 2,3,7,8-TCDD (as a single
Injection); which 1s 30,000 times more potent that 3-MC (Poland and Glover,
1974) 2,3,7,8-TCDF reduced N-demethylase activity at all doses (Goldstein et
al., 1978b) and Increased P-448 [Previous terminology for cytochrome P-450
Isoenzymes Induced by 3-MC was "cytochrome P-448." This term will be used
In this paragraph to denote all forms of cytochromes Induced by 3-MC class
of Inducers as simplified expression.] Kawano and Hlraga (1978) Investigat-
ed the effect of a mixture of tetra- (12%) and penta-CDF (88%) In male
Wlstar JCL rats (105-165 g bw) at doses of 10, 100, 1000 yg/kg bw/day for
3 days. They found effects similar to those elicited by 3-MC (20 mg/kg bw)
on mlcrosomal drug-metabolizing enzymes. Both drugs Increased p-n1troan1-
sole demethylase activity, moderately Increased aniline hydroxylase activ-
ity, and produced Uttle change 1n amlnopyrlne demethylase activity. Fur-
thermore, both drugs Increased cytochrome P-448.
The AHH Inducing potencies of PCDF, TCDD and PCB were compared In rats
by Nagayama et al. (1983). A mixture of PCDF (13% 1,2,7,8-TCDF; 35%
2,3,7,8-TCOF; 1% 1,2,4,7,8-PeCDF ; 49% 1,2,3,7,8-PeCDF ; 1% 2,3,4,7,8-PeCDF;
1% unspecified hexa-CDF) and TCDD at the dose of 5 yg/kg bw and a PCB
mixture (Kanechlor-500) at the dose of 50 mg/kg bw were given 1.p. TCDD only
enhanced AHH activity 1n the prostate, thymus and spleen. In the kidney,
lung and liver the order of AHH InduclbllHy was TCDD > PCDF » PCB. PCDF
caused the greatest Induction In the lung. Thus, the lung and kidney seem
more sensitive than the liver to the Inductive properties of these compounds.
1931A 7-67 06/22/86
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Recently, Bandlera et al. (1984a) treated rats with a reconstituted mix-
ture of PCDF and PCB that reproduced the approximate composition of the PCDF
and PCB persisting 1n the liver of Yusho patients according to the most
recent data (Rappe et al., 1983a; Masuda and Kurokl, 1982; Nagayama et al.,
1983). The mixture of PCDF, Including 2,3,7,8-TCDF; 1,2,4,7,8-PeCDF;
1,2,3,7,8-PeCDF; 2,3,4,7,8-PeCDF and 1,2,3,4,7,8-HxCDF (7.4, 6.1, 19.0, 24.9
and 38.1%, respectively), was Injected 1.p. at various doses. The dose-re-
sponse pattern on AHH Induction showed that the mixture elicited, 4- and
40-fold Increases, respectively, In the liver enzymatic activity of rats, at
doses of 22 and 63 yg/kg bw. All the PCB mixtures were significantly less
active as AHH Inducers than PCDF. From an analysis of the PCBrPCDF ratios
1n the contaminated rice oils In Taiwan (100:0.32) and In Japan (100:0.16)
and of the ratios of the respective EC5Q for AHH Induction (at least
1:700), the authors suggest that PCDF must be considered the major etlologlc
agent In the contaminated rice oils and In Yusho poisoning of victims still
suffering from this syndrome (Bandlera et al., 1984a; Sawyer and Safe, 1985).
Induction of AHH activity by 3-MC Is expressed almost exclusively as a
single autosomal dominant trait regulated by the Ah locus (Nebert et al.,
1975). In Inbred strains of mice AHH 1nduc1b1l1ty Is genetically separate.
Eight PCDF were given 30 vg/kg 1.p. to four Inbred strains of mice with
different phenotypes of Ah locus (Nagayama et al., 1985b). In responsive
mice (C57) with the exception of 1,2,3,6,7-PeCDF and 1,2,3,4,6,7-HxCDF all
the PCDFs were active; In unresponsive strains (DBA and ODD) no Induction
was elicited by the 8 PCDFs tested. Results Indicate that AHH responsive-
ness 1n mice segregates with the AHH Inductive activity by PCDF and may also
segregate with the toxic potency of the Isomers as In fact has been demon-
strated for the 1mmunotox1c1ty (Vecchl et al., 1983b).
1931A 7-68 06/22/86
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Human tissue appear to have Ah receptor molecules with properties simi-
lar to those observed In animals (Nebert and Jensen, 1979; Roberts et al.,
1985; Gaslewlcz and Rued, 1984). Recently Wong et al. (1985, 1986) con-
ducted a study on pregnant Chinese women 1n Taiwan exposed 4-5 years earlier
to contaminated rice oils containing PCOFs, PCBs and PCQs. Placental mlcro-
somal fractions from exposed subjects presented significantly high AHH and
ethoxyresoruf1n-o-deethylase (EROD) activities. In the same subjects pla-
centa! mlcromes were found to contain a protein that cross-reacted with the
antibodies against cytochrome P-450, Isoenzyme 6. This Isoenzyme 1s known
to be Induced by polychlorlnated hydrocarbons.
The effect of PCDF was studied 1n human lymphoblastold cell lines with
different AHH 1nduc1b1l1ty for 3-MC (Nagayama et al., 1985b). Degrees of
the enzyme 1nduc1b1lH1es of PCDFs Increased proportionally with those for
3-MC, Indicating that 1n humans also, AHH 1nduc1b1l1ty for PCDF reflects the
genetic susceptibility of the cells. Induction with 2,3,4,7,8-PeCDF;
1,2,3,4,6,7-HxCDF and 1,2,3,4,7,8-HxCDF was comparable to that with TCDD and
higher than with TCDF, Indicating a species difference compared with the
laboratory animals.
7.4.5.5.3. Structure-Activity Relationship for Induction — The
structure/activity relationship of PCDFs to AHH activity was studied using
the chick embryo by Poland et al. (1976) who found that 2,3,7,8-TCDF and
2,3,4,7,8-PeCDF, as well as the corresponding CDD derivatives, were the most
active Isomers. These studies showed that both the number and the position
of the chlorine atoms are Important. The Isomers that Induce AHH activity
have two common properties: halogen atoms should occupy at least three, and
for maximal potency, four of the lateral ring positions (positions 2, 3, 7
1931A 7-69 06/22/86
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and 8); at least one ring perl-position (positions 1, 4, 6, 9) should be
unsubstUuted.
Nagayama et al. (1983) demonstrated In rat lung and liver mlcrosomal
preparations that chlorlnatlon at position 1 reduced AHH InduclblHty. In
fact 2,3,7,8-TCDF and 2,3,4,7,8-PeCDF are more active on liver AHH Induction
than 1,2,3,7,8-PeCDF and 1,2,3,4,7,8-HxCDF. A similar reduction was observ-
ed for Induction In the lung by 2,3,6,7-TCDF and 2,3,4,6,7-PeCDF In relation
to 1,2,3,6,7-PeCDF and 1,2,3,4,6,7-HxCDF. Moreover, an additional Cl sub-
stitution at the C-4 (or C-6) position Increases the AHH Induction potency
In relation to the corresponding C-l or C-9 substitutes.
Several data suggest that species differences may change the order of
PCDF Isomers for AHH-1nduc1ng activity. In mice 2,3,7,8-TCDF elicited the
highest AHH Induction (Nagayama et al., 1985b,d), In rats 2,3,7,8-TCDF for
some authors (Yoshlhara et al., 1981; Nayayama et al., 1983) In liver and
lung mlcrosomes but 2,3,4,7,8-PeCDF for others (Bandlera et al., 1984b;
Mason et al., 1985) 1n liver.
The small number of in vivo kinetic data available on this class of com-
pounds make 1t difficult to compare in vivo and in vitro results. The role
of tissue distribution, blotransformatlon and elimination of PCDFs should be
carefully considered for each compound under study. Only 2,3,7,8- TCDF has
been studied 1n some detail 1n this regard (Decad et al., 1981a,b; Blrnbaum
et al., 1980, 1981; Blrnbaum, 1985).
The most extensive structure/activity relationship studies have been
made using the highly (AHH) Induclble rat hepatoma H-4-11-E cell lines
(Bradlaw and Casterllne, 1979; Bandlera et al., 1984a,b; Mason et al., 1985).
Mason et al. (1985) correlated the in vitro AHH-1nduc1ng potency of
PCDFs with the thymlc atrophy or body weight loss caused by each congener 1n
1931A 7-70 06/22/86
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rodents. Although this 1s an obvious oversimplification of the problem It
may be accepted as a good model when a rapid Indication of the potential
tox1c1ty of new congeners 1s needed. This point will be discussed In more
detail In other Sections.
Bradlaw and Casterllne (1979), working with rat hepatoma cell culture
[as a rapid screen test for detecting minute (pg) amounts of certain classes
of compounds, e.g. PCDFs, PCDDs and PCBs], found that AHH activity was
Induced by PCDF containing at least 3 or 4 lateral ring positions substitut-
ed by chlorine atoms In the ring positions at 2,3,7,8; congeners with two to
fewer chlorine atoms In the lateral positions showed no Inductive effect up
to a dose of 5 or 10 mg/kg bw.
Table 7-29 summarizes the quantitative jji vitro data reported by
Bandlera et al. (1984b) and Mason et al. (1985) concerning the relationship
between the different TCDFs In their AHHand EROD-1nduc1ng potency. Bandlera
et al. (1984b) showed that the AHH Inductive ratios of the 2,3,4/1,3,8,
2,6,7/1,3,8, and 2,3,4,6/1,2,4,8 pairs of PCDF Isomers were 128, 7 and 9,
respectively.
Figure 7-1 reports a plot of the -log EC5Q values for In vitro AHH
Inducing potencies versus the -log ED5Q values for body weight loss and
thymlc atrophy for a series of PCDF congeners, a PCDF mixture with a compo-
sition similar to that found 1n the liver of Yusho patients and TCDD. Sta-
tistical analysis of both plots showed an excellent correlation, corroborat-
ed by the fact that the compounds tested differed widely In both jm vivo and
In vitro biological and toxic potencies. 2,3,7,8-TCDF was the most active
AHH Inducer in vitro and the most powerful and toxic congener in vivo.
1931A 7-71 06/22/86
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TABLE 7-29
Effects of Different PCDF Congeners on Rat Hepatic Cytosollc Receptor
Binding Avidities, AHH and EROD Induction In Rat Hepatoma H-4-II E Cells3
PCDF
Congeners
Dlbenzofuran
2-
3-
4-
2,3-
2,6-
2,8-
1,3,6-
1,3,8-
2,3,4-
2,3,8-
2,6,7-
1,2,3,7-
1,2,3,6-
2,3,4,7-
2,3,4,6-
2,3,4,8-
2,3,6,8-
2,3,7,8-
1,2,4,8-
1,2,4,6,7-
1,2,4,7,9-
1,2,3,4,8-
1,2,3,7,8-
1,2,4,7,8-
1,2,3,7,9-
Receptor Binding Avidities
(ED50) M
<10
2.8x10'*
4.2±0.6xlO~5
<10"3
4.72xlO"6
2.46xlO"4
2.57xlO"6
4.40xlO~6
8.50xlO~5
1.9xlO"5
1.0±0.1xlO~6
4.5xlO"7
1.12xlO"7
3.54xlO~7
2.51xlO~e
3.5xlO"7
2.0xlO"7
2.2xlO~7
4.H0.6xlO~8
>10"5
6.77xlO'5
2.0xlO~5
1.2xlO~7
7.45±2.04xlO~ed
1.3xlO"6
3.98xlO~7
Enzyme Induction
(EC50)
AHH (M)C
ND
ND
ND
l.OxlO"5
2.19xlO"6
6.17xlO~s
3.95xlO~5
2.53xlO"6
1.94xlO"5
1.51xlO"7
2.49xlO~6
2.80xlO~6
2.70xlO~5
>10"*
1.79xlO"8
1.32x10"*
4.14xlO~8
1.04xlO~6
3.91xlO~9
1.20xlO~5
3.25xlO~7
3.77x10"°
2.09xlO"7
2.54xlO~9
1.06X10"7
8.60x10'°
Potencies
EROD (M)C
ND
ND
ND
1.71xlO~5
4.84x10"*
6.31xlO~5
4.0xlO"5
3.37x10'*
3.02X10"5
2.48xlO"7
1.56x10"*
3.13x10'*
6.30xlO~5
>10~4
1.48x10"°
1.13x10"*
3.76x10'°
7.79X10"7
2.02xlO"9
9.26xlO~5
3.48xlO"7
3.84x10"°
1.63xlO"7
3.06xlO~9
1.48X10"7
8.60x10'°
1931A
7-72
06/22/86
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TABLE 7-29 (cont.)
PCDF
Congeners
1,2,4,6,8-
1,3,4,7,8-
2,3,4,7,8-
2,3,4,7,9-
1,2,3,4,7,8-
1,2,3,6,7,8-
1,2,4,6,7,8-
2,3,4,6,7,8-
2,3,7,8-TCDD
Receptor Binding Avidities
(ED50) M
3.09xlO~6
2.00xlO~7
1.50±0.1xlO"8<1
2.00xlO~7
2.3xlO~7
8.3xlO"6
4.7i0.4xl08d
l.OxlO"8
Enzyme Induction
(EC50)
AHH (M)c
l.OOxlO'5
1.60xlO~9
2.56xlO"10
7.90xlO~»
3.56xlO~10
1.47xlO"9
4.24X10"8
6.87xlO~10
7.23X10'11
Potencies
EROD (M)c
1.20xlO~5
1.40x10"'
3.79xlO"10
1.24xlO~9
3.79xlO~10
1.24x10"'
2.93X10"8
5.75xlO"10
1.85xlO"10
aSource: Bandlera et al., 1984b; Mason et al., 1985
bAHH = Aryl hydrocarbon hydroxylase
CEROD = Ethoxyresorufln o-deethylase
dose-response competition experiments were carried out In triplicate and
Illustrate the reproduclbUHy of the assay with the PCDF.
ND = Not detected
1931A
7-73
06/22/86
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10
I 6
.,
\
fCOf m,.
l.J.1 •'.»--
•
. [ •— 2 3 76-TCOO |
• —J 34 ;.»-
-J.J.4.«.7«-
34567
Thymic Atrophy
10
—J.i.7« rcoo
r • — PCOf
-12 3.6 r e
2.3 4 7»
1 J47»-
1.2 3 7 « —.
34567
Body Wf Lota (-tog CO JO'
FIGURE 7-1
A plot of the -log £659 values for AHH Induction In rat hepatoma
H-4-IIE cells vs. the ^n vivo -log ED5Q values for thymlc atrophy (top)
and body weight loss (bottom) for several PCDFs, 2,3,7,8-TCOD and a recon-
stituted PCDF mixture
Source: Mason et al., 1985
1931A
7-74
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In human lymphoblastold cells Nagayama et al. (1985c) showed that chlor-
Inatlon 1n position 1 has an activating effect. In these cells the HxCDFs
and PeCDFs (1,2,3,4,7,8-, 1,2,3,4,6,7- and 2,3,4,7,8-) were more potent than
TCDF and a sex effect appeared, with females consistently presenting higher
1nduc1b1l1ty. As far as species differences 1n AHH Inducing activity are
concerned, In rats and mice PCDFs were always less active than TCDDs but In
human cells the most active Isomers were as potent as TCDDs (Nagayama et
al., 1985c). This must be taken Into account In relation to their reported
high persistence 1n tissue of humans (Rappe et al., 1983a) and undomestlcat-
ed animals In evaluating their toxic potency.
A structure-activity relationship similar to that on AHH Induction was
reported by Nagata et al. (1981) for the effect of PCDFs on the mlcrosomal
metabolism of progesterone. Both 2,3,7,8-TCDF and 2,3,4,7,8-PeCDFs selec-
tively Increased 7a-hydroxylat1on, but strongly suppressed 2a, 60 and
16a hydroxylatlon and 5a reduction of progesterone. This change In the
progesterone metabolic pattern results In a marked depression of total
metabolism of the steroids. 2,8-DCDF; 1,3,6,7-TCDF; 1,3,6,8-TCDF and
1,2,4,6,8-PeCDF do not affect progesterone metabolism.
7.4.5.5.4. AHH Induction and Cytosol Receptor Binding — The signifi-
cance of the Induction of drug-metabolizing enzymes by PCDFs and chlorinated
aromatic hydrocarbons 1n general In the Interpretation of their toxldty Is
still an open question (Poland et al., 1979; Yoshlmura et al., 1979; Poland
and Kende, 1976). An Important observation was the correlation between AHH
Induction and binding affinities for the hepatic cytosol receptor protein of
a certain number of PCDF congeners (Nagayama et al., 1983; Poland et al.,
1979; Yoshlhara et al., 1981; Bradlaw and Casterllne, 1979). Recently the
effect of structure on rat hepatic cytosol receptor affinity and AHH 1nduc-
1931A 7-75 06/22/86
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tlon potency was studied for 26 pure PCDFs (Bandlera et al., 1984b) (see
Table 7-29) that were synthesized according to a new reported procedure
(Safe and Safe, 1984). As for AHH Induction, the most active PCDFs for
receptor binding were substituted In all four lateral positions. Moreover,
compounds with additional Cl groups at C-4 or C-6 bind with higher affinity
to the receptor than Isomers with substHuents at C-l or C-9. However,
there 1s no linear correlation between the receptor binding affinity and AHH
Induction potencies for the 26 Isomers. This suggests that after Initial
binding of PCDF Ugands with the receptor some molecular features (such as
conformation of the llgand-receptor complex, differential Ugand/receptor-
nuclear Interactions) distinct from those required for attachment, affect
events In the complex enzyme Induction process.
Structure/activity relationship (SAR) studies have been recently con-
ducted (Mason et al., 1985) with a number of pure PCDFs and a mixture of
PCDFs resembling the compositions of the mixture persisting 1n Yusho
patients (Bandlera et al., 1984a) 1n an attempt to correlate their AHH
Inductive properties and toxic effects (thymlc atrophy and weight loss).
Results Indicated an excellent linear correlation between jji vitro and In
vivo activity for the congeners that are substituted 1n both phenyl rings
and do not contain vicinal unsubstltuted carbon atoms. The effect of chlor-
ine substitution at the four different positions of the dlbenzofuran ring
can be summarized as follows: 3,7 > 2,8 > 44,6 > 1,9. This differs from
PCDDs, 1n which all the lateral positions are equivalent because of the sym-
metry of the d1benzo-p-d1ox1n ring system. Inspection of these results and
of the suggested hypothetical structural features Indicate that a molecule
can be hypothesized with high affinity for the receptor but scant Inductive
1931A 7-76 06/22/86
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power, thus eliciting no toxic effects. Such a molecule might be an anti-
dote for the most toxic PCDF.
Keys et al. (1986) have recently reported that two TCOF congeners
2,4,6,8-TCDF and 1,3,6,8-TCDF possess the peculiarity of being poor In vitro
Inducers of AHH and EROD activities but have relatively high binding avidi-
ties to the cytosollc receptor protein. These compounds competitively dis-
placed TCDD from the receptor and their coadm1n1strat1on with TCDO was sig-
nificantly was reduced In the expected additive Induction of AHH and EROD
activities. On the other hand, 2,3,7,8-TCDF administered together with
2,3,7,8-TCDD gave additive enzyme Induction responses.
Cheney (1982) and Cheney and Tolly (1979) developed a theoretical model
suggesting that four structural factors, Including the fused trlcycHc aro-
matic system, were Important for the attachment and stimulation of AHH
Induction, while the electronegative substHuent of the lateral ring did not
Influence Induction and binding In the same manner. More precisely, the
latter appears to Increase Induction to an extent that Is greatly out of
proportion to Us effect on binding. The lateral ring substitution seems
particularly Important to switch the receptor-complex to an active state to
promote AHH Induction.
In conclusion, the toxldty of PCDFs seems to correlate well with their
potential capacity to Induce AHH; however, toxldty Is not bound directly to
AHH activity, but both AHH Induction and toxic effects are to be considered
secondary to the Initial binding to the cytosollc protein receptor.
In the last 2 years a link has been found between TCDD, which 1s the
most powerful and the most widely studied of the polychlorlnated hydrocar-
bons, and the regulation of epidermal growth factor (EGF) binding 1n human
keratlnocyte lines was demonstrated (Hudson et al., 1985, 1986; Osborn and
1931A 7-77 06/23/86
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Greenlee, 1985; Greenlee et al., 1985a). These authors demonstrated that
TCDD can modulate the proliferation and differentiation of epidermal kera-
tlnocytes both j_n vivo and in vitro. This was explained by the competition
of TCDD with EGF 1n binding to the Ah receptor present In basal cells.
Another Important example of the relationship between the binding to a
specific receptor and manifestation of a toxic effect was reported by Green-
Ice et al. (1985b) who showed that TCDD-lnduced thymlc atrophy was mediated
by a receptor protein In thymlc epithelial cells. The major consequence of
this fact appears to be altered thymus-dependent maturation of T-lymphocyte
precursors, mediated through direct cell-cell contact between thymocytes and
thymlc epithelial cells.
7.5. REPRODUCTIVE/DEVELOPMENTAL TOXICITY
7.5.1. Introduction. The literature base on the potential reproductive
and developmental toxldty of the PCDFs 1s limited, and prior assessments
have relied largely on the similarity of these agents with the dloxlns (U.S.
EPA, 1983, 1984a). Recently, however, several laboratories have conducted
studies that focus on the dlbenzofurans. This chapter will pay particular
attention to these studies, as well as summarize those studies that have
been reviewed 1n previous U.S. EPA documents. In general, the greatest
focus has been on prenatal development, with little Information on other
aspects of the reproductive process. Most laboratory studies have concen-
trated on 2,3,7,8-TCDF, since the TCDD homolog 1s known to be highly toxic
and teratogenlc.
7.5.2. Development. The studies by Nagayama et al. (1980) Indicate that
polychlorlnated dlbenzofurans (PCDFs) are capable of being transferred to
the developing mouse, both prenatally through the placenta and postnatally
through the breast milk. Exposure of the dams to a mixture of tetra-,
1931A 7-78 06/24/86
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penta- and hexachlorodlbenzofurans was through the diet during the first 18
days of pregnancy or during the first 2 weeks of postnatal (PN) life. Low,
but measurable, levels of PCDFs were found both 1n fetuses at gestation day
18 and In neonates at 1 and 2 weeks of PN life. Whole body concentrations
were highest In the neonates, with the percentage of total Intake at PN week
2 double those at PN week 1, suggesting an accumulation of the PCDFs during
the early postnatal period. Weber and Blrnbaum (1985) also examined the
transplacental transfer of radioactive 2,3,7,8-TCDF 1n the C57B1/6N mouse.
Exposure was by gavage on gestation day 11 at a dose known to cause a high
level of cleft palate. Low levels of radioactivity were found 1n the
embryos at gestation day 12, but radioactivity 1n the embryos was below
detectable limits by gestation days 13 and 14. Together, these two studies
Indicate that the PCDFs are transferred at low levels to the embryo and
fetus transplacentally, and at higher levels to the nursing neonatal animal
during early PN life.
Weber et al. (1984) studied the effect on prenatal development of single
and multiple exposures to 2,3,7,8-TCDF 1n C57B1/6N mice. The 2,3,7,8-TCDF
was 98% pure (primary contaminant was PeCDF). Administration was by oral
gavage 1n corn oil (10 mil/kg/treatment). Single exposure was on gestation
day 10 (plug day = gestation day 0) at 0 (vehicle control), 250, 500 and
1000 jig 2,3,7,8-TCDF/kg bw. Multiple exposures were on gestation days
10-13, with dally exposure levels at 0, 10, 30, 50 and 100 v9 2,3,7,8-
TCDF/kg bw. The maternal animals were monitored for body weight changes and
signs of toxlclty during gestation, and the uterine contents were examined
on gestation day 18 before parturition. The fetuses were examined for gross
and visceral malformations, but were not examined for skeletal anomalies.
There were no signs of maternal toxlclty, except for changes 1n absolute and
1931A 7-79 06/22/86
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relative liver weight at the highest dose under both exposure regimens. The
authors Indicate that this Is probably a metabolic response of the liver to
2,3,7,8-TCDF exposure, although they note that a toxic effect on the liver
cannot be ruled out.
In terms of developmental toxldty, the most significant effects were on
the kidney and palate. Following single exposure, there was a significant,
dose-related Increase In Utters with hydronephrosls at all doses (250, 500
and 1000 yg/kg) and litters with cleft palate at the highest dose (1000
yg/kg). Following multiple exposure a similar pattern was observed, with
an Increase In Utters with hydronephrosls starting at 30 yg/kg/day and an
Increase 1n litters with cleft palate starting at 50 yg/kg/day. The
authors also reported an Increase 1n fetal mortality above control levels at
all doses following single exposure (0% In control; 12.6% at 250; 14.7% at
500; 21.IX at 1000 yg/kg). However, even though this was statistically
significant, the finding must be viewed with caution, since a total absence
of fetal mortality 1n mice, even 1n the control population, Is uncommon.
For example, the control group for the multiple exposure design In this
study (Weber et al., 1984), as well as a control group In another study by
this laboratory (Weber et al., 1985), exhibited 11.7% and 10.5% fetal
mortality, respectively, at a level comparable to the 2,3,7,8-TCDF-exposed
groups. There were no other gross or visceral malformations observed and
there was no effect on fetal weight. The authors do not Indicate why the
ICDFs would not be expected to produce skeletal changes. However, the
exclusion of this major class of anomalies, which 1s often quite sensitive
to Insult, may have limited the effects that were seen.
These results demonstrate that 2,3,7,8-TCDF Is a developmental toxicant
1n the mouse at exposure levels that are not maternally toxic. Based on the
1931A 7-80 07/01/86
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Utter, which 1s generally considered the experimental unit 1n this type of
study design, the LOAEL was at 250 yg/kg/day following a single exposure
and 30 yg/kg/day following the 4-day exposure. A NOEL was not established
for a single exposure; for multiple exposures the NOEL was 10 yg/kg/day.
Several additional points should be recognized In analyzing the data.
First, the number of animals/dose group was low. The current U.S. EPA
recommendation Is 20 Utters/dose group (U.S. EPA, 1984b); however, 1n this
study, the number of Utters ranged between 6 and 11. This does not reduce
the Importance of the observations, but had the number been Increased the
power of the study to detect change may have been Increased, with effects
being seen at lower doses. Second, while the Utter 1s considered the
experimental unit In this study design, the Individual fetal data should not
be discounted. Kidney changes as a mean percentage of fetuses were signifi-
cantly Increased following multiple exposure at the 10 yg/kg/day dose.
Since these changes were consistent with the dose-related response of the
other endpolnts on both a Utter and Individual fetus basis, 1t Is possible
that 10 yg/kg/day would be the LOAEL following multiple exposure. Third,
hydronephrosls can be a reversible alteration 1n development; and the
authors note that preliminary studies suggest this may be the case for the
kidney changes observed following TCDF. However, without Information on the
level and duration of exposure and the postnatal age at observation, 1t 1s
not possible to determine the significance of these findings relative to the
developmental toxldty of the agent. Moore et al. (1973) examined the
effect of pre- and postnatal exposure combinations to TCDD on the postnatal
observation of kidney lesions and concluded that the Incidence of hydro-
nephrosls Is probably a function of dose and length of exposure. Moreover,
the developmental significance of transient changes Is not well-understood
1931A 7-81 06/22/86
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but should not be arbitrarily discounted. Thus, these three points should
be kept 1n mind In the overall assessment of the toxlclty of the PCDFs, as
they may Influence the final determination of the effect levels.
Recently, Blrnbaum et al. (1986) have reported, 1n abstract form, pre-
liminary data from their studies on the relative teratogenlcHy of three
PCDFs. C57B1/6N pregnant mice were exposed to 2,3,4,7,8-PeCDF; 1,2,3,7,8-
PeCDF; or 1,2,3,4,7,8-HxCDF on gestation days 10-13 and examined for terato-
genlc effects on gestation day 18. All three PCDFs were teratogenlc, as
evidenced by the occurrence of cleft palate at doses that did not produce
maternal toxlclty. The ED5Qs were 40, 100 and 400 pg/kg for 2,3,4,7,8-
PeCDF, 1,2,3,7,8-PeCDF and 1,2,3,4,7,8-HxCDF, respectively. The NOEL for
2,3,4,7,8-penta-CDF was 1 vg/kg, apparently lower than that for the
2,3,7,8-TCDF. Hydronephrosls occurred at even lower doses. Further analys-
is of this study will be done as the Information becomes available since the
apparent Increased toxlclty of 2,3,4,7,8-PeCDF may have an effect on the
reported NOEL.
Weber et al. (1985) reported on the teratogenlc potency of TCDF and TCDD
when exposure to these two agents 1s combined. Individually, each agent
showed a steep dose-response curve for the Induction of cleft palate, with
TCDD -30 times more potent than TCDF. In combination, the two agents were
additive 1n their contribution to cleft palate Induction, with no Indication
of a synerglstlc or antagonistic effect of combined exposure.
Hassoun et al. (1984) also examined the developmental effects of
2,3,7,8-TCDF 1n mice of the C57B1 strain, using an 1.p. route of exposure.
Purity of the agent was 90%; 8% was hexachloroterphenyls, which the authors
Indicate "are probably less toxic and teratogenlc" than TCDF. Exposure was
by a single Injection on gestation days 10, 11, 12 or 13 to the dloxane
1931A 7-82 06/22/86
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vehicle or TCDF at concentrations ranging from 0.1-0.8 mg/kg bw. The ani-
mals were examined on gestation days 16 or 17 for Implantation sites,
resorptlons, and live and dead fetuses. Fetuses were examined for cleft
palate and hydronephrosls. The number of treated dams In each dose group
ranged from 5-14 (average = 8.8), which 1s less than optimal. Furthermore,
the use of the l.p. route Is generally not recommended since a mechanical or
acute chemical Insult can occur 1n the vicinity of the uterus. Neverthe-
less, this study supports the findings of Weber et al. (1984) and Indicates
that the most sensitive exposure period during gestation for the Induction
of cleft palate and hydronephrosls Is gestation days 11-12.
In relation to human exposure to the PCDFs, Kuratsune (1976, 1980) has
reviewed epidemlologic studies on Yusho (oil sickness), the Incident 1n
Japan 1n which humans were exposed to rice oil contaminated with PCBs con-
taining a high level of PCDFs and other organochlorlne compounds. Of 13
women who were exposed during pregnancy (11 showing signs of Yusho; 2 with
husbands showing signs), 9 offspring had unusual grayish/dark brown pigmen-
tation of the skin, glnglvae and nails, and most showed a discharge from the
eyes. Since similar signs were almost never seen elsewhere In Japan, the
findings suggest an association between prenatal exposure and adverse out-
come at birth. However, Kuratsune (1976, 1980) pointed out that the low
number of cases does not allow for a clear cause and effect relationship to
be drawn. Kuratsune (1976, 1980) also noted that 12 of the 13 babies were
small by national standards, but this difference disappeared with age and
the children had no apparent physical or mental handicap. In relation to
postnatal exposure, Kuratsune (1976, 1980) summarized a report suggesting
that boys of school age exposed to the contaminated oil showed a significant
decrease In both height and weight relative to unexposed boys. Exposed
1931A 7-83 07/01/86
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girls did not show any such decrease relative to unexposed girls. Finally,
from one case report, a child exposed only by nursing from the exposed
mother began showing signs of Yusho, Indicating transfer of the toxic agents
through the breast milk.
7.5.3. Reproductive Function/Fertility. Studies designed to monitor the
effect of the dlbenzofurans on male and female reproductive function and
fertility, apart from the developmental toxldty studies noted above, are
minimal. Olshl et al. (1978) compared the toxldty of a mixture of TCDFs,
PeCDFs and HeCDFS with that of a commercial polychlorlnated blphenyl mix-
ture. Male rats (100-130 g) were exposed through the diet to 0, 1 or 10 ppm
PCDF for 4 weeks. In relation to the reproductive system, they found that
the relative testes weight was Increased at both the 1 and 10 ppm doses.
There were other signs of general toxlclty at these doses, Including
decreased body weight and food consumption, and 1t 1s likely that the effect
on relative testes weight was associated with these changes. At 10 ppm,
there were reductions In the absolute and relative weights of the seminal
vesicles and ventral prostates, as well as a decrease 1n the concentration
of testlcular testosterone. However, because of the other general and sys-
temic toxlclty seen at this dose level, 1t Is not possible to determine 1f
PCDFs had a direct effect on the male reproductive system. In reviewing the
Yusho Incident, Kuratsune (1980) noted a report Indicating that Irregular
menstrual cycles and abnormal patterns of basal body temperature were
observed In 60 and 85% of the exposed women patients, respectively. Unfor-
tunately, other reproductive effects of the PCDFs must be assumed from stud-
ies on similar agents such as dloxln. A recent Health Assessment Document
for Polychlorlnated D1benzo-£-D1ox1n (U.S. EPA, 1985) has summarized this
literature and 1t will not be repeated here. However, 1n the few studies
1931A 7-84 06/22/86
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that have examined reproductive endpolnts not associated with developmental
toxldty, there has been no clear Indication of an adverse effect of the
dloxlns on reproduction.
7.5.4. Summary. The data clearly support the developmental toxldty of
2,8,7,8-tetrachlorodlbenzofuran 1n the C57B1/6N mouse at relatively low
levels of exposure. The -study of Weber et al. (1984) provides the most
direct support of this toxlclty, demonstrating an adverse effect following a
single exposure to 250 yg/kg on gestation day 10 and following a multiple
exposure to 30 yg/kg/day on gestation days 10-14. A NOEL could not be
determined from the results following a single exposure. Following multiple
exposure, no effect was observed at 10 yg/kg/day when the data were anal-
yzed on a Utter basis. Considering the additional points that are noted
above on study design and data Interpretation, It Is possible that biologic-
ally significant changes would occur at even lower levels of exposure.
The potential human developmental toxldty of the PCDFs has not been
systematically studied. However, the findings associated with the Yusho
Incident are suggestive that the PCDFs can have an effect on the fetus and
on the developing child.
Studies on other reproductive effects associated with PCDF exposure do
not provide sufficient data to be more than suggestive. Effects have been
reported on specific aspects of both the male and female systems. However,
additional study would be required to clearly delineate any possible direct
effect of the PCDFs on reproductive function and fertility.
As noted 1n Section 7.5.1., the literature base on the chlorinated
dlbenzofurans In the area of reproductive and developmental toxlclty Is very
limited. Many laboratories have focused on the dloxlns, with a considerable
extension of that data base, at the expense of the dlbenzofurans. Since the
data base on the dloxlns 1s more extensive, 1t would be useful to continue
1931A 7-85 06/30/86
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and expand reviews that address the similarities and differences In the
action of these two classes of agents. More directly, however, studies
should be designed to more adequately assess the low end of the PCOF effect
range relative to developmental toxlclty, and studies on fertility and
reproductive function should be Initiated. In addition, a clearer picture
should be developed regarding the effect of chemical species (since the
majority of studies have examined 2,8,7,8-TCDF) and animal species and
strain (since most studies have used the C57B1/6N mouse). Finally, a better
understanding of the pharmacoklnetlcs and metabolism of these agents, espe-
cially as related to transplacental transfer, transfer through the breast
milk, and localization/duration of exposure In the embryo/fetus and neonate
would help In establishing appropriate experimental models and extrapolating
data from animal to the human condition.
7.6. MUTAGENICITY
Schoeny (1982) tested four chlorinated dlbenzofurans (2,8-DCDF,
3,6-DCDF, 2,3,7,8-TCDF, and OCDF) In the Salmonella hlstldlne reversion
assay. Four doses (0.1-10 yg/plate) of 2,8-DCDF were assayed In Salmonel-
la. strains TA1535, TA100, TA1975, TA92, TA2322, TA1537, TS24, TA2637,
TA1538, TA98 and TA1978. 3,6-DCDF was tested Identically except that strain
TA1538 was not used. Five doses (0.1-20 yg/plate) of 2,3,7,8-TCDF were
assayed 1n strains TA1535, TA100, TA1537, TA98 and TA1978; three doses
(0.1-5 yg/plate) of octachlorodlbenzofuran were tested 1n strains TA100,
TA1975, TA92, TA2322, TS24, TA1537 TA2637, TA98 and TA1978. All four PCDFs
were tested In the absence or presence of Aroclor-lnduced rat S9 mix.
2,3,7,8-Tetrachlorodlbenzofuran was also tested In the presence of S9 mix
from rats induced with 3-methylcholanthrene, phenobarbltal and 2,3,7,8-TCDF;
OCDF was also tested 1n the presence of S9 mix from rats Induced with
2,3,7,8-TCDF.
1931A 7-86 06/30/86
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The results were negative (mutagenldty was reported as none). Toxlclty
was not observed except for 2,3,7,8-TCDF at 1 vg/plate 1n the absence of
S9 mix. Schoeny (1982) commented that the Salmonella hlstldlne reversion
assay Is relatively Insensitive to chlorinated hydrocarbons, perhaps because
the S9 preparations cannot effect dechlorlnatlon or some other reaction
necessary to produce active metabolites.
2,3,7,8-TCDF was also studied (along with several chlorophenols and
chlorophenol Impurities) In Saccharomyces cerevlslae strain MP-1 and was
negative for forward mutation, mltotlc crossing over and mltotlc gene con-
version at concentrations up to 1.0 mg/ml (FahMg et al, 1978). Survival
was 91X at 1.0 mg/ma. Stationary phase cells were tested In the absence
of exogenous activation. The control values varied widely among the tests
of the Individual chemicals.
In summary, two studies (both negative) on the mutagenlc potential of
chlorinated dlbenzofurans have been reported. Additional studies are needed
before any conclusion can be reached concerning the mutagenldty of these
compounds.
7.7. CARCINOGENICITY
7.7.1. Animal. Close structural similarity of PCDFs to PCDDs, especially
2,3,7,8-TCDD, which Is a proven animal carcinogen, raises concern as to the
potential carclnogenlcHy of 2,3,7,8-TCDF. However, no animal cardnogenlc-
Ity bloassay data on PCDFs are currently available 1n the literature.
However, as PCDFs are contaminants of commercial PCB and might have
contributed to the toxic effect 1n animals and man (Olshl et al., 1978; Vos
et al., 1970; Cordle et al., 1978), It can be hypothesized that they play a
role (e.g., promotion, cocarclnogenesls, Initiation) In the suspected
carcinogenic effect of PCB.
1931A 7-87 06/30/86
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PCB mixtures (Aroclor 1254 and 1260, Kanechlor 400 and 500) reportedly
produce hepatic tumors In mice and rats (WHO, 1978b; Klmbrough, 1974;
Klmbrough et al., 1972). Recently the hepatocarclnogenlc potential of
Aroclor 1260 was confirmed In Sprague-Dawley rats (dose of 100 yg/g In the
diet for 16 months), a strain with a low Incidence of spontaneous hepato-
cellular neoplasms (Norback and Weltman, 1985). After 18 months, In the
PCB-exposed group hepatocellular neoplasms were present 1n 95% of the 47
females and 1n 15% of the 46 males with a significant sex difference. In 81
controls only one hepatocellular neoplasm occurred. The appearance of
tumors In the liver and stomach that do not occur spontaneously In control
rats was also reported after treatment with Aroclor 1254 (25-100 ppm 1n the
diet) suggesting that PCBs Initiate lesions de novo rather than promote the
development of naturally-occurring lesions (Ward, 1985).
7.7.2. Human. Amano et al. (1984) recently completed a 16-year cohort
mortality study of 1086 Yusho victims 1n Japan. In 1968, these victims suf-
fered an acute toxicosis from consuming rice oil contaminated with an
Industrial oil consisting of a mixture of PCBs, PCDFs and PCQs. The 581
males and 505 females sustained a total of 70 deaths (42 males vs. 45.81
expected and 28 females vs. 31.3 expected). These data are based upon
Japanese national death rates over age 40 through October 31, 1983. In this
population, for persons over 40 years of age overall cancer mortality was
greater than expected 1n men but no different In women. In male Yusho
victims 19 cancer deaths occurred vs. 11.50 expected and 1n female Yusho
victims 7 cancer deaths occurred vs. 7.20 expected. However, by organ site,
the risk of liver cancer was consistently found to be higher than expected
In both men and women during the entire 16-year observation period. Even
after a 9-16 year latent period, the risk of liver cancer 1n males was
1931A 7-88 06/30/86
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significantly higher (observed, 5; expected, 0.75; p<.01). This 1s a short
latency period when compared with most known carcinogens. In females the
results were 2 observed vs. 0.45 expected.
As a result of an 1n-depth review of this study and consideration of the
overall PCDF toxldty relationships, 1t has been hypothesized by CA6 that
the PCDFs component of the 'PCBs could have been a factor In the statisti-
cally significant liver cancers seen 1n these victims. Particular Isomers
of the Ingested PCDFs were found 1n liver tissue 1n nearly the same propor-
tions as PCBs several years later (Kuratsune et al., 1975).
7.8. EPIDEMIOLOGY -- SYSTEMIC TOXIC EFFECTS
Two mass food poisonings, the first 1n Japan 1n 1968 and the second In
Taiwan 1n 1979, were found to be caused by the Ingestlon of rice oil highly
contaminated with PCB, PCDF and PCQ, but not with PCDD. The Japanese epi-
sode Involved 1788 people at the end of 1982 and the Taiwan episode totaled
2062 patients at the beginning of 1983 (Masuda et al., 1985). It has been
roughly estimated that the average Intake during the whole Intoxication per-
iod was 973, 3.8 and 586 mg In Taiwan (Chen et al., 1985a) and 633, 3.3 and
596 mg In Japanese patients (Hayabuchl et al., 1979) for PCB, PCDF and PCQ,
respectively. The syndrome known as Yusho 1n Japan and Yu-Cheng 1n Taiwan,
has been attributed mainly to PCDF, although the presence of PCB and PCQ
should not be overlooked (Chen et al., 1981; Kashlmoto et al., 1981). The
animal data (Kashlmoto et al., 1985; Kunlte et al., 1985) Indicate that PCB
and PCQ components alone cannot cause Yusho symptoms.
Speculations on the levels of the three contaminant congeners detected
1n blood and tissues of Yusho and Yu-Cheng patients also support the hypo-
thesis that PCDFs are the main agents responsible for the poisonings (Masuda
et al., 1985; Mlyata et al., 1977b, 1985; Chen et al., 1985a).
1931A 7-89 06/30/86
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7.8.1. The Japanese Incident (Yusho disease). Yusho disease has been
described In several papers and comprehensive reviews (Kuratsune, 1972,
1975, 1980; Klmbrough, 1974; Hsu et al., 1985; Urabe and Asahl, 1985; Yoshl-
mura and Hayabuchl, 1985). Therefore, only an overview of clinical effects
will be given here.
The early symptoms presented by 89 males and 100 females 1n the Japanese
Yusho Incident are listed In Table 7-30. The available Information allows
only a qualitative description of the clinical effects; case control studies
were aimed only at Identifying the etlologlcal role of rice oil. The asso-
ciated estimates of the amount of oil consumed (Ingested quantity of PCB and
PCDF) and severity of clinical acute-subacute findings could be described 1n
a few cases (see Table 7-14) (Kuratsune, 1972, 1975; Nagayama et al., 1976;
Strlk, 1979). Other available Information will be briefly discussed.
7.8.1.2. VARIOUS CLINICAL AND BIOCHEMICAL FINDINGS — Besides those
reported for the acute and subacute phase after exposure (see Table 7-30),
other abnormal nonsystematlc findings are discussed In the following sec-
tions.
7.8.1.2.1. Liver — Slight rises In serum transamlnases and alkaline
phosphatase, occasionally low blllrubln levels; marked proliferation of
smooth endoplasmlc retlculum In a liver biopsy sample from one exposed
Individual. Unspecified liver enlargement and disturbances were also
reported.
7.8.1.2.2. L1p1d Metabolism — Serum trlglycerldes Increased to four
times the normal value, but plasma cholesterol and phosphollpld levels were
always normal. This hypertrlglycerldemla persisted for several years.
Follow-up of 24 patients showed lowered levels 5-7 years after the accident.
1931A 7-90 06/30/86
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TABLE 7-30
Signs and Symptoms of Yusho Disease 1n Adult Japanese*
SIGNS
Blackening of nails
Black spots In all pores
Excessive sweating In palms
Acnellke skin eruption
Red spots on limbs
Change 1n skin color
Swelling of hands and feet
Hardening of backs of hands
Pigmentation of mucous membranes
Sebum (gum secretion In eyes)
Hyperemla of mucous membrane of eyes
Temporary falling of eyesight
Jaundice
Swelling of upper eyelids
Fever
Vomiting
Diarrhea
SYMPTOMS
Itching
Sense of weakness
Numbness of hands and feet
Hearing difficulty
Spasms of hands and feet
Headaches
Males (n=89)
1%)
83.1
64.0
50.6
87.6
20.2
75.3
20.2
24.7
56.2
88.8
70.8
56.2
11.2
71.9
16.9
23.6
19.1
42.7
58.4
32.6
18.0
7.9
30.3
Females (n=100)
(X)
75.0
56.0
55.0
82.0
16.0
72.0
41.0
29.0
47.0
83.0
71.0
55.0
11.0
74.0
19.0
28.0
17.0
52.0
52.0
39.0
19.0
8.0
39.0
*Source: Kuratsune, 1972
1931A 7-91 06/24/86
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7.8.1.2.3. Dermal Signs — The dermal lesions described 1n Table 7-30
were the most notable manifestations of Yusho. All skin symptoms diminished
gradually but persisted with a tendency to cyst formation 1n severe cases.
This was the Initial criterion used to designate Yusho patients. The acne-
form eruptions decreased markedly after 3-4 years leaving a small number of
cysts. Black comedones disappeared between 5 and 6 years: folUcular dots
disappeared within 4 years; pigmentation disappeared after 10 years, last
from the nails. Ingrown nails are still apparent 1n 1985 (Urabe and Asahl,
1985).
Some women who consumed contaminated rice oil gave birth to "coco"
(heavily black plgmented) babies (Klkuchl et al.. 1969, 1977).
7.8.1.2.4. Porphyrln Metabolism — Strlk (1979) found no alteration
of this factor 1n Yusho patients and no other manifestations of porphyMa
cutanea tarda such as light sensitivity or hyperkeratosls. Increased excre-
tion of a-ketosterold was an occasional finding and lower than normal
1mmunoglobul1n levels were reported 1n 38 patients 2 years after the acci-
dent.
7.8.1.2.5. Ocular Signs — Although some signs subsided, 84% of 75
patients examined still showed abnormal changes of the melbomlan glands 10
years after the accident; 64% of the patients complained of eye discharge.
7.8.1.2.6. Neurological Signs — Reduced sensory nerve conduction
velocity (9 of 23 cases) was observed more frequently than reduction of
motor nerve conduction (2 cases only). No follow-up studies have been made.
Dullness, headache, heavy-headedness, indefinite stomach ache, numbness,
pains In the extremities, and swelling and pain of joints were still being
reported in 1985 (Urabe and Asahi, 1985).
1931A 7-92 06/30/86
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7.8.1.2.7. Respiratory ~ Chronic bronchitis with mucus production
was present 1n 40X of the cases. There was a very slow Improvement 1n 79
patients that were followed for 5 years. Coughing and bronchHIs-Uke symp-
toms were still being reported 1n 1985 (Urabe and Asahl, 1985). An Investi-
gation of 401 Yusho patients showed that one-half of the subjects were com-
plaining of respiratory distress. Follow-up showed a gradual Improvement
over the 10 years following onset of the disease; however, from 10-15 years
after onset Uttle or no further Improvement was observed In most cases
(Nakan1sh1 et al., 1985a,b) and some pulmonary function test Impairment per-
sisted long after the Initial Yusho Incident.
In conclusion, although the clinical profile of Yusho disease has been
described In several papers, there Is Uttle precise and quantitative Infor-
mation on clinical manifestations, Including the most severe.
7.8.2. The Taiwan Incident ("Yu-Cheng" Disease). On May, 1979, the first
report of a Yusho-I1ke disease 1n Talchung County, Taiwan was recorded with
1843 cases over a wide area up to November 1980 (Hsu et al., 1985). The
Initial diagnosis for 1670 victims was based on dermal phenomena set out In
Table 7-23. By February 1983 there were 2062 recognized cases. In nine
particularly afflicted townships, the morbidity rates were 0.095, 0.338,
0.392, 1.35, 0.248, 0.688, 0.056, 0.421 and 0.131X. The Kanechlor 400/Kane-
chlor 500 contamination In rice oil varied from 31-300 ppm, the period of
exposure from 3-9 months and the estimated total Intake per person from
0.77-1.84 g. The average latent period to the onset of visible signs was
3-4 months (range 1.5 to >6 months). In the first year, blood PCB In 613
patients ranged from 3-1156 ppb with 82.5X being 11-150 ppb; 27.6X had
levels >100 ppb. Hyperplgmentatlon and growth retardation In newborn of
pregnant mothers exposed to PCBs was noted along with enhanced mortality
1931A 7-93 06/30/86
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(weakness and lung problems). In adults, 24 deaths were reported mostly
from hepatoma, liver cirrhosis or liver diseases with hepatomegaly.
PCB-exposed patients In the first year suffered from Infections at the
following sites: respiratory tract, skin, and Immunosuppresslon (Lu and Wu,
1985). Ocular symptoms Included Increased discharge from eyes, swelling of
eyelids, weakness of eyesight, soreness, easy Irritation and easy fatigue of
eyes (Lu and Wu, 1985). Other major complaints were hyperplgmentatlon
(especially In nails) (Wong et al., 1985), headache, dizziness, general
malaise, reduced appetite, soreness and weakness of limbs, swelling or pain
of the joints and feet, Ingrown nails (especially the big toe) and numbness
of the limbs (Lu and Wu, 1985). Both sensory (In 43.6% of the patients) and
motor (1n 21.8% of the patients) nerve conduction velocity (NCV) were lower
than control (Chen et al., 1985b). The major dermatologlcal signs, as noted
above, were acne-comedones, nail pigmentation and prurltls. Abnormal men-
struation and hyperhydrosls of the palms and soles were noted 1n some
females -10%). Placental tissues obtained from women who had been exposed
3-4 years before conception, showed large Increases 1n monoxygenase enzymes,
particularly AHH (Wong et al., 1985). Males did not complain of Impotence.
Because of Immunosuppresslon (humoral and cellular), secondary mlcroblal
Infection often arose. Initially, total T, active T and helper T cells were
decreased but not B or suppressor T cells. Three years later, only the
helper 1 cells were still low giving an Immuno-regulatlng Index of 63% of
unexposed controls. Studies 3 years later showed Improvement of general
conditions, of subjective symptoms and of cutaneous changes (Lu and Wu,
1985).
1931A 7-94 06/30/86
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No association could be established between severity of the disease and
PCB level In blood. However, the Individual variations In the levels are so
wide that no conclusion could be drawn from the reported results (Lu and Wu,
1985). Correlations between Yusho and Yu-Cheng symptomatology and PCOF
levels In tissues have been discussed 1n previous sections.
7.8.3. The Blnghamton, N.Y. Transformer Accident. An Incident with
explosion and fire In a tr1- and tetrachlorlnated benzene-containing elec-
trical transformer took place In Blnghamton, New York, on February 5, 1981.
The consequent overheating led to the release of -750 8, of fluid contain-
ing 65% PCBs and 35% trl- and tetrachlorlnated benzenes. Pyrolytlc conver-
sion of these compounds led to the formation of several PCOFs and traces of
PCDDs (Schecter et al., 1984). A total of -500 persons were believed to be
exposed, but data are available only on 50 patients who voluntarily sought
medical surveillance. Among them, one case of chloracne, several transient
skin rashes, three cases of skin cancer, three subjects with liver altera-
tions with no other known cause were observed. Psychological symptoms
(nervousness, Irritability, fatigue, etc.) were recorded.
1931A 7-95 06/30/86
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8. EFFECTS OF MAJOR CONCERN AND HEALTH HAZARD ASSESSMENT
8.1. EXISTING GUIDELINES AND STANDARDS
No guidelines, standards or recommendations are available for PCDFs.
8.2. SOURCES OF EXPOSURE
PCDFs enter the environment as unwanted trace Impurities 1n commercial
mixtures of polychlorlnated blphenyls (PCBs), products derived from poly-
chlorinated phenols (PCPs), chlorinated naphthalenes, 2,4,5-trlchloro-
phenoxyacetlc add (2,4,5-T) formulations and through pyrolysls of PCBs,
2,4,5-T and polychlorlnated dlphenyl ethers. Incinerators (municipal,
Industrial and others), combusting organic materials and chlorinated com-
pounds appear to a major source of PCDFs 1n the environment. The presence
of PCDFs In the emissions from Incinerators can be explained by the pyroly-
sls of chlorinated aromatic compounds. The chlorinated benzenes, PCBs and
PCPs, which are present 1n high concentrations In emissions from waste
Incinerators, have also been suspected to act, either Individually or 1n
combination, as precursors or Intermediates for PCDF formation. PCDFs
emitted from Incinerators Include tetra-, penta-, hexa-, hepta- and octa-
chlorodlbenzofurans.
PCDFs are very widespread, appear to be highly stable 1n the environment
like PCDDs, and can bloaccumulate 1n aquatic and terrestrial organisms.
PCDFs have been detected 1n fish, turtles, seals, and fish-eating birds, In
animal and human adipose tissue, and 1n animal and human milk. Exposure to
the human population can occur by Ingesting fish from contaminated water
bodies. Human adipose tissue and breast milk from selected populations have
been found to bloaccumulate PCDFs. Considerable concern has arisen over the
public health Impact of the emission of PCDFs from waste Incinerators and
1932A 8-1 06/30/86
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presence of PCOFs 1n human adipose tissue and breast milk. Public health
Implications of nursing Infants by mothers whose milk contains residues of
PCOFs are many.
Accidental fires 1n electrical equipment, especially PCB-fllled trans-
formers and capacitors have been a source of exposure to PCDFs for a great
number of people.
Wood preserved by chlorophenols Is another source of PCDFs 1n the
environment. High levels of PCDFs have been detected In dust from sawmills,
sludge from wood dipping tanks, and other sources.
8.3. HIGH RISK POPULATIONS
Populations with high risk of exposure to PCDFs Include the following:
workers In the wood or tanning Industry who produce or use chlorophenol
preserved products; firefighters working on electric or transformer fires;
workers at the Incinerators; factory workers making or repairing trans-
formers or capacitors, using casting waxes or heat exchange systems; office
workers using carbonless copy papers containing PCBs; auto mechanics working
on cars that use leaded gasolines; people who have a high Intake of fish
from PCDF contaminated water bodies; and Infants who are nursed by mothers
whose milk contains PCDF residues.
PCDFs are extremely toxic to animals and humans as demonstrated by
short-term exposures. Long-term exposure and health experiences are notably
lacking. Signs and symptoms of toxlclty are very similar to those caused by
2,3,7,8-TCDD. Of the total 135 possible Isomer congeners so far determined
for PCDFs, 2,3,7,8-TCDF, 1,2,3,7,8-PeCDF and 2,3,4,7,8-PeCDF seem to be most
toxic. However, the relative tox1cH1es of all the congeners have not yet
been fully studied.
1932A 8-2 06/30/86
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Information on adverse health effects In humans was gathered 1n 1968 and
1979 from observations In selected Japanese and Taiwanese populations, who
accidentally consumed PCB-/PCDF-contaminated rice oil. Severe acute toxic
effects were observed 1n the affected population. Retarded growth, abnormal
I1p1d metabolism, liver dysfunction, acneform eruptions, skin pigmentation
and cutaneomucosal lesions were some of the general symptoms observed 1n the
affected Individuals. Nine babies born of mothers from these affected popu-
lations had grayish/dark pigmentation of the skin ("coco" babies), glnglval
and nails, and the majority of these babies had an unusual discharge from
the eye.
Liver and adipose tissue from affected populations examined several
years after exposure showed retention of PCDFs at ppb levels; 2,3,4,7,8-
PeCDF was found to be retained by the liver at higher concentrations than
other Isomers.
8.4. BASIS AND DETERMINATION OF RISK
There Is no experimental evidence from human, animal or mutagenesls
assays that pure PCDFs are carcinogenic or mutagenlc 1n mammals. PCDFs do
not give a mutagenlc response In mlcroblal assay systems. Furthermore, no
long-term chronic human epidemlologic or animal bloassay toxlclty data are
available that can be used for determining human health risk from chronic
exposure to pure PCDFs.
The only animal data available for hazard assessment result from
short-term exposure. Blrnbaum et al. (1986) and Blrnbaum (1986) found 3
yg/kg/day to be the lowest observed effect level (LOEL) for hydronephrosls
In the mouse fetus because of in utero exposure to 2,3,4,7,8-PeCDF during
days 10-13 of the gestation period, while for cleft palate H was 5
yg/kg/day. The LOAEL for hydronephrosls for 2,3,7,8-TCDF, 1n a mouse
teratogenlclty study by Weber et al. (1984) was found to be 10 yg/kg/day.
1932A 8-3 06/30/86
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Because of the short duration (4 days) for the j_n utero exposure to
PCDFs resulting 1n hydronephrosls and cleft palate 1n the mouse fetus, the
available teratogenlclty data are not adequate to determine human health
risk from chronic exposure. Consequently, the lowest-observed-adverse-
effect level (LOAEL) 1s determined using the TCDO-equ1valent extrapolation
approach
8.4.1. Estimation of LOAELs. Structurally and tox1colog1cally, PCDFs
resemble their PCOO congeners. Their structural-activity relationships to
AHH activity have been studied 1n a rat hepatoma cell line (Bandlera et al.,
1984b; Mason et al., 1985). Like 2,3,7,8-TCDD, PCDFs need to bind to the
cytosollc Ah receptor site for manifestation of their AHH Inducing activity.
The chlorine atoms have to occupy at least three of the four lateral
positions (2, 3, 7 and 8) and at least one vicinal carbon position must be
unsubstHuted. A linear relationship for a series of PCDFs, a PCDF mixture
typical of exposure to Yusho patients and 2,3,7,8-TCDD was obtained when
-log EC values for AHH Induction responses were plotted against -log
EDrri values for body weight loss or thymlc atrophy (Mason et al., 1985).
bU
In all cases 2,3,7,8-TCDD was found to be the most active substitute for AHH
Induction in vitro and for Iji vivo short-term toxic effects.
Taking Into consideration the AHH activity and other acute toxic
effects, 1t has been estimated that 2,3,4,7,8-PeCDF and 2,3,7,8-TCDF are
3- and 20-fold, respectively, less toxic than 2,3,7,8-TCDD (Bandlera et al.,
1984b; Mason et al., 1985) (see Table 7-29 and Figure 7-1).
Reevaluatlon of the Murray et al. (1979) primary data from a 3-genera-
tlon Sprague-Dawley rat reproductive effect study with 2,3,7,8-TCDD by
Nlsbet and Paxton (1982) and subsequently by U.S. EPA (1985) revealed that
the LOAEL for 2,3,7,8-TCDD was 0.001 vg/kg/day.
1932A 8-4 06/30/86
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Toth et al. (1978, 1979) exposed male Swiss mice to 0.007 yg/kg/week
of 2,3,7,8-TCDD for 1 year, which resulted In amyloldosis of the kidney,
spleen and liver, and dermatitis at the time of death. The duration of this
study was 649 days. In this study 0.001 yg/yg/day was also found to be
the LOAEL.
The LOAELs for 2,3,4,7,8-PeCDF and 2,3,7,8-TCDF are estlmted to be 0.003
jig/kg/day and 0.02 yg/kg/day, respectively, from the above animal long-
term chronic studies using the TCOD equivalent approach.
For comparative purposes, an attempt will now be made (In consultation
with Dr. J.J. Ryan) to determine the LOAEL for 2,3,4,7,8-PeCDF In the Yusho
poisoning Incident.
The minimum quantity of rice oil that led to an observed effect (grade 3
Yusho response) 1n a 42-year-old person after a latent period of 30 days was
121 ma,. The mean latent period 1n the Yusho episode was estimated to be
71 days (range: 20-190 days) (Hayabuchl et al., 1979). The average amount
of total PCDFs 1n the rice oil has been estimated (Ryan, 1986) from several
sources (Buser et al., 1978; Mlyata et al., 1978 and Maguda, 1982) (see
Chapter 4) to be 3.85 yg/g (ppm).
Using the peak heights on the chromatogram In the Buser et al. (1978)
study, from peak 68 It has been estimated that 2.3.4.7.8-PeCDF was -8.4% of
the total PCDFs of the Yusho oil (Ryan, 1986). The content of 2,3,7,8-TCDF
1n the oil 1s assumed to be negligible since both Masuda et al. (1982) and
Chen and Kites (1983) have found that the peak elutlng near 2,3,7,8-TCDF Is
1n fact 2,3,4,8-TCDF. Moreover, the contribution of 1,2,3,4,7,8- and
1,2,3,6,7,8-HxCDF 1s not considered to be significant since: a) HxCDFs were
present 1n lower concentrations than PeCDFs In the oil, b) HxCDFs accumu-
lated less In tissues of Yusho patients than PeCDFs (Kurokl and Masuda,
1978), and c) the HxCDFss are less toxic than PeCDFs 1n animals.
1932A 8-5 06/30/86
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Using the estimations on the total oil consumed, the average quantity of
total PCDFs 1n the oil, and the percentage of 2,3,4,7,8-PeCDF In the total
PCDFs In the oil, the LOAEL 1n a 70 kg man for 2,3,4,7,8-PeCOF 1s determined
to be:
121 mft x 3.85 ug x 0.084
71 days x 70 kg
= 0.007 yg/kg/day
For 2,3,4,7,8-PeCDF, this estimated LOAEL of 0.007 yg/kg/day from the
Yusho Incident Is supportive of the LOAEL of 0.003 yg/kg/day derived from
animal studies using the TCDO equivalent approach.
8.4.2. Derivation of Reference Dose (RfD). In the absence of long-term
chronic animal bloassay data on 2,3,4,7,8-PeCDF It seems reasonable to
determine a Reference Dose (RfD) based on the LOAEL estimated following the
TCDD equivalent approach. A composite uncertainty factor of 1000 1s used to
estimate the RfD. This uncertainty factor represents 10 because adverse
effects observed In animals are extrapolated to humans, and another 10 Is
used to account for the expected Interhuman variability In the toxldty
response to this chemical, an additional uncertainty factor of 10 1s used
because the RfD Is based on a LOAEL and not a NOAEL; thus, this latter
factor adjusts the LOAEL Into the range of the expected NOAEL. An uncer-
tainty factor of 10 to account for the short duration of the study Is not
used because the critical toxic effect, teratogenldty, 1s encompassed 1n
the duration of the study. Consequently, an uncertainty factor of
100x10=1000 Is used.
8.4.2.1. THE RfD FOR 2,3,4,7,8-PeCDF — The RfD for 2,3,4,7,8-PeCDF
1s estimated to be as follows:
, 3 x ,0-« M,/kg/day
1932A 8-6 07/01/86
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The following 1s a series of calculations that estimate the criteria for
2,3,4,7,8-PeCDF 1n various media.
1. For a 70 kg man Inhaling 2,3,4,7,8-PeCDF In 20 m3 of air/day
the criteria would be:
- 3 x 10"* yg/kg/day x 70 kg
20 mVday
= 1.1 x 10~5 yg/m3
2. For a 50 kg woman Inhaling 2,3,4,7,8-PeCDF 1n 20 m3 of air/day
3 x 10~6 yg/kg/day x 50 kg
20 mVday
- 7.5 x 10"6 yg/m3
[The above calculations assume that exposure occurs only by
Inhalation.]
3. Since 2,3,4,7,8-PeCDF residues have been detected 1n mother's
milk samples from Isolated populations, a virtually safe level
for Ingestlon of 2,3,4,7,8-PCDF 1n mothers' milk Is also estim-
ated. In this calculation, the total body weight of an Infant
1s considered to be 10 kg (ICRP, 1975). The total quantity of
fat 1n 100 ml of mother's milk 1s 4.5 g and the average dally
Intake of nursing mother's milk by the baby 1s considered to be
850 mt (ICRP, 1975). Therefore, 850 mil of mother's milk
will contain 38.25 g of fat.
Therefore, for virtual safety for an Infant, the concentration
of 2,3,4,7,8-PeCDF In fat from the nursing mother's milk should
not exceed the following:
3 x 10~6 yg/kg/day x 10 kg
38.25 g/day
= 7.8 x 10~7 vg 2,3,4,7,8-PeCDF/g of fat.
4. Since fish has been found to bloaccumulate PCDFs easily, a cri-
terion for a 70 kg man from Ingestlon of contaminated fish 1s
also determined. The average dally consumption of fish 1n the
United States 1s considered to be 6.5 g (Federal Register,
1980).
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Therefore, the criterion 1s as follows:
3 x 10"* mg/kg/day x 70 kg
6.5 g/day
= 3.2 x 10~5 mg 2,3,4,7,8-PeCDF/g of fish.
This calculation Is based on fish as the sole source of
2,3,4,7,8-PeCDF.
8.4.2.2. THE RfD FOR 2,3,7,8-TCDF — Following TCOD equivalent
approach from animal data, the LOAEL for 2,3,7,8-TCDF has been estimated
(Section 8.4.1.) to be 0.02 vg/kg/day. Based on this LOAEL, In the
absence of any chronic animal bloassay data on 2,3,7,8-TCDF H seems reason-
able to determine a RfD for this compound.
A composite uncertainty factor of 1000 1s used to estimate the RfD.
This uncertainty factor represents 10 because adverse effects observed In
animals are extrapolated to humans, and 10 to account for the expected
Interhuman variability 1n the toxlclty response to this chemical, an addi-
tional uncertainty factor of 10 because the RfD Is based on a LOAEL and not
a NOAEL; thus, this latter factor adjusts the LOAEL Into the range of
expected NOAEL. An uncertainty factor of 10 to account for the short dura-
tion of the study Is not used because the critical toxic effect, terato-
genlclty, 1s encompassed In the duration of the study. Consequently, an
uncertainty factor of 100x10=1000 1s used.
The RfD 1s estimated to be:
0.02 ug/kg/day _ ,.__ .. ..
- —**—a J- = 2 x 10 5 yg/kg/day
1000
The following Is a series of calculations that estimate criteria
for 2,3,7,8-TCDF 1n various media.
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1. For a 70 kg man Inhaling 2,3,7,8-TCDF 1n 20 m3 of air/day the
criteria would be
2 x 10"5 yg/kg/day x 70 kg
zu m3/day
= 1 x 10~* mg/m3
2. For a 50 kg woman Inhaling 2,3,7,8-TCDF 1n 20 m3 of air/day
_ 2 x 10~5 mg/kg/day x 50 kg
;>u m3/day
= 1 x 10~4 iig/m3 air.
[The previous calculations assume that exposure occurs only by
Inhalation.]
3. Since 2,3,7,8-TCDF residues have been detected 1n mother's milk
samples from specific populations, a virtually safe level for
Ingestlon of 2,3,7,8-TCDF 1n mothers' milk 1s also estimated.
In this calculation, the total body weight of an Infant Is con-
sidered to be 10 kg (ICRP, 1975). The total quantity of fat In
100 ma of mother's milk 1s 4.5 g and the average dally Intake
of nursing mother's milk by the baby Is considered to be 850
ml (ICRP, 1975). Therefore, 850 ml of mother's milk will
contain 38.25 g of fat.
Therefore, for virtual safety for an Infant, the concentration
of 2,3,7,8-TCDF 1n fat of the nursing mother's milk should not
exceed
2 x 10~5 yg/kg/day x 10 kg
»fl ,,- /Ha = 5.2 x 10-* yg 2,3,7,8-TCDF/g of fat.
oo.£i> g/oay
4. Since fish has been found to bloconcentrate PCDFs easily, a
criterion for a 70 kg man from Ingestlon of contaminated fish
1s also determined. The average dally consumption of fish In
the United States Is considered to be 6.5 g (Federal Register,
1980).
Therefore, the criterion 1s as follows:
2 x 10~s yg/kg/day x 70 kg
6.5 g/day
= 2 x 1Q-* Mg 2,3,7,8-TCDF/g of fish.
1932A 8-9 06/23/86
-------
These estimations provide general guidance only. To determine a more
accurate criteria 1t will be necessary to do a comprehensive exposure
assessment taking Into consideration the exposure through various compart-
ments of the environment and valid chronic bloassay data on 2,3,4,7,8-PeCDF
and 2,3,7,8-TCOF.
1932A 8-10 06/24/86
-------
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