ORNL
Oak Ridge
National
Laboratory
Operated by
Union Carbide Corporation for the
Department of Energy
Oak Ridge, Tennessee 37830
0 R N L ; E IS - 130
EPA
United States
Environmental Protection
Agency
Office of Research and Development
Health Effects Research Laboratory
Cincinnati, Ohio 45268
EPA-60071-79-044
REVIEWS OF THE ENVIRONMENTAL
EFFECTS OF POLLUTANTS:
X. Toxaphene
-------
OKNL/EIS-130
EPA-600/1-79-044
REVIEWS OF THE ENVIRONMENTAL EFFECTS OF POLLUTANTS: X. TOXAPHENE
Report prepared by
Patrick R. Durkin, Philip H. Howard, Jitendra Saxena, Sheldon S. Lande,
Joseph Santodonato, John R. Strange, and Deborah H. Christopher
Center for Chemical Hazard Assessment
Syracuse Research Corporation
Merrill Lane
Syracuse, New York 13210
under Subcontract No. 448 and 7663 for
Information Center Complex
Information Division
OAK RIDGE NATIONAL LABORATORY
Oak Ridge, Tennessee 37830
operated by
UNION CARBIDE CORPORATION
for the
DEPARTMENT OF ENERGY
Contract No. W-7405-eng-26
Reviewer and Assessment Chapter Author
William B. Buck
College of Veterinary Medicine
University of Illinois
Urbana, Illinois 61801
Interagency Agreement No. D5-0403
Project Officer
Jerry F. Stara
Office of Program Operations
Health Effects Research Laboratory
Cincinnati, Ohio 45268
Prepared for
HEALTH EFFECTS RESEARCH LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
CINCINNATI, OHIO 45268
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NOTICE
This report was prepared as an account of work sponsored by an agency
of the United States Government. Neither the United States Government nor
any agency thereof, nor any of their employees, contractors, subcontractors,
or their employees, makes any warranty, express or implied, nor assumes any
legal liability or responsibility for any third party's use or the results
of such use of any information, apparatus, product or process disclosed in
this report, nor represents that its use by such third party would not
infringe privately owned rights.
This report has been reviewed by the Health Effects Research Laboratory,
U.S. Environmental Protection Agency, and approved for publication. Approval
does not signify that the contents necessarily reflect the views and policies
of the U.S. Environmental Protection Agency, nor does mention of trade names
or commercial products constitute endorsement or recommendation for use.
ii
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FIGURES
Page
2.1 Schematic flow chart for toxaphene manufacture 2-7
2.2 Ionic pathways in the chlorination of camphene 2-10
2.3 Comparison of the gas chromatographs of toxaphene and Strobane. . 2-22
2.4 Gas chromatogram from 30 ng of toxaphene (A) before alkali
treatment and after alkali treatment (B) 2-46
2.5 Specific ion monitoring with gas chromatography of toxaphene
and DDT residues 2-48
2.6 LMRGC for toxaphene in New Orleans drinking water extract .... 2-49
3.1 Relative toxicity of toxaphene components as compared with
toxaphene (as 1) to freshwater blue-green algae,
Anacystis nidulans 3-17
5.1 Uptake of Cl-36 labelled toxaphene and toxaphene related
residues in mosquito fish exposed to 2 ppm Cl-36 toxaphene. . . . 5-7
5.2 The rate of uptake of whole-body levels of toxaphene by rainbow
trout in Miller Lake. Average concentration of toxaphene in
water was 1.20 ppb for 1963 and 0.84 ppb for 1964 5-19
5.3 Variation, with time, of toxaphene concentration in bluegills
from Fox Lake, mean and range indicated 5-20
5.4 Dose-response curves for susceptible and resistant strains of
mosquito fish exposed to toxaphene for 36 hours 5-50
5.5 Activity of large oysters and growth of small oysters exposed
to 0.1 ppm toxaphene 5-65
5.6 Effects of six levels of toxaphene on survival in selected
aquatic invertebrates 5-93
5.7 Graph of the relative toxicities of Sephadex LH-20-methanol
column fractions to mosquito larvae and brine shrimp 5-96
5.8 Graph of the relative toxicities of TLC separated fractions
to mosquito larvae and brine shrimp 5-97
5.9 The effects of several concentrations of toxaphene on clam
eggs and larvae 5-101
5.10 Survival curves showing the net effect of toxaphene on ring-
necked pheasant reproduction 5-125
5.11 Log dose-probit lines for toxaphene treated lady beetles based
on 72-hour observations 5-164
iii
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FIGURES (Continued)
6.1 Tissue levels in female rats fed toxaphene in the diet at
concentrations of 2.33 and 189 ppm
6.2 The effect of toxaphene concentration in the diet on toxaphene
tissue levels in male and female rats after twelve weeks
feeding 6"10
6.3 Excretion of radioactive products by rats treated orally with
[Cl-36] toxaphene, [Cl-36] toxaphene fractions, and [C-14]
toxaphene 6-17
6.4 Effect of feeding toxaphene and DDT on EPN detoxication,
£-demethylase activity, and N-demethylase activity of livers
of male rats 6-19
6.5 Log dose response curves of the acute oral toxicity of toxaphene
to rats under different dietary conditions 6-25
6.6 Log dose response curves for male and female rats to acute
oral and dermal toxaphene intoxication 6-26
7.1 Atmospheric samples collected from the RV Trident, 1973-1974. . . 7-8
7.2 Monthly average of apparent toxaphene concentration of
manufacturing plant effluent, 1970-1974 7-22
7.3 Amount of toxaphene measured in Clayton Lake (New Mexico) water
from the 3.05 meter level before and after applications of
toxaphene 7-26
7.4 Gas chromatograms of standard toxaphene and toxaphene
extracted from Comstock Lake Water 7-45
7.5 GC patterns of toxaphene from sediments of Fox Lake and
Ottman Lake compared to standard toxaphene 7-46
7.6 Degradation of toxaphene in soil unamended or amended with
alfalfa meal and incubated in a moist aerobic environment or
under moist and flooded anaerobic conditions 7-51
8.1 Materials flow diagram for toxaphene, 1972 8-5
IV
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.Ely viKCv;,.io ; ; ,-,_ ,~!iL, \L"fri LIBRARY
TABLES UNIVERSITY D£ CIN.CIN.NATl
Page
2.1 Physical properties of technical toxaphene 2-4
2.2 Toxaphene product specifications 2-5
2.3 Composition of the products from ionic chlorination of camphene. 2-12
2.4 Components in toxaphene separated by silica gel adsorption
chromatography and then gas chromatography with qualitative
analysis by chemical ionization mass spectrometry 2-14
2.5 Approximate composition of polychlorinated toxaphene constituents
based upon field ion mass spectrum measurement 2-20
2.6 Structure and properties of constituents isolated from toxaphene 2-23
2.7 Analytical methods for toxaphene 2-38
2.8 Measurements of variability, recovery, and sensitivity for
several toxaphene analytical procedures 2-39
2.9 Gas chromatographic columns used for toxaphene analysis 2-41
2.10 Relative retention times of pesticides and related compounds
and their electron capture detection response 2-42
2.11 Comparison of colors obtained by the reaction of pesticides
with diphenylamine-zinc chloride 2-51
2.12 Recommended procedure for toxaphene measurement 2-53
3.1 Effect of field application of toxaphene on soil bacteria. . . . 3-2
3.2 Effect of toxaphene on processes in soil catalyzed by bacteria . 3-3
3.3 Toxicity of toxaphene to soil fungi 3-7
3.4 Accumulation of toxaphene in fish-food organisms exposed to
multiple toxaphene dosages 3-10
3.5 Toxicity of toxaphene to unicellular algae 3-12
3.6 Effect of toxaphene on the physiological processes of uni-
cellular algae and phytoplankton 3-19
4.1 Uptake of Cl-36-labelled toxaphene by salt marsh cordgrass,
Spartina alterniflora 4-4
4.2 Toxaphene residues in aquatic vegetation in lakes treated with
toxaphene for eradication of rough fish 4-5
4.3 Ambient toxaphene levels in noncrop plants 4-7
v
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TABLES (Continued)
4.4 Residue of toxaphene detected in crop plants
4.5 Comparison of washing with and without surfactants on removal
of toxaphene from vegetables ................... 4-12
4.6 Toxaphene residues found in food and drug administration
market basket survey, 1964 to 1975. ............... 4-14
4.7 Toxaphene residues in tobacco and tobacco products ........ 4-16
4.8 Response of various crops to application of large amounts
of toxaphene ........................... 4-19
4.9 Effect of 30 ppm toxaphene, added at the time of planting,
on growth and respiration of various crop plants ......... 4-21
4.10 Effect of toxaphene on the growth and size of vegetable seedlings 4-22
4.11 Effect of application of toxaphene to tobacco on cigarette flavor 4-25
5.1 Whole-body residues of toxaphene in brook trout fry continuously
exposed to toxaphene ....................... 5-9
5.2 Whole-body residues of toxaphene in yearling brook trout con-
tinuously exposed to toxaphene .................. 5-10
5.3 Toxaphene residues found in adult brook trout fillet and
remaining tissue after 161 days of exposure ........... 5-13
5.4 Toxaphene residues in adult brook trout after transfer to
uncontaminated water ....................... 5-15
5.5 Toxaphene total body residues in fish from Davis Lake ...... 5-18
5.6 Toxaphene residues in fish from various fresh-water systems . . . 5-22
5.7 Collagen, calcium, and phosphorus concentrations in fathead
minnow backbones as affected by 150 day continuous exposures
to various levels of toxaphene .................. 5-26
5.8 Backbone composition of brook trout fry exposed to toxaphene.
Mean values expressed on basis of dried weight of backbone. . . . 5-28
5.9 Inhibition of catfish brain, kidney, and gill ATPases by
toxaphene ............................ 5-29
5.10 Acute toxicity of toxaphene to various fish during static
exposures ...... . ................... , 5-31
VI
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TABLES (Continued)
Page
5.11 Effects of temperature in the acute toxicity of toxaphene
to fish 5-36
5.12 The relative acute toxicity to fish of some common pesticides
compared to toxaphene 5-41
5.13 The effects of chronic toxaphene exposure on the growth of
fathead minnows 5-44
5.14 The effect of chronic toxaphene exposure in the growth of
brook trout fry 5-45
5.15 The effect of toxaphene on brook trout reproduction 5-48
5.16 Use of toxaphene as a fish control agent 5-53
5.17 Accumulation of toxaphene by oysters exposed to 1 ppb singly
or in combination with 1 ppb DDT and 1 ppb parathion 5-59
5.18 Effects of toxaphene exposed Daphnia on fish 5-60
5.19 Toxaphene residue in plankton from Big Bear Lake exposed to
0.03 and 0.10 ppm treatments two weeks apart 5-62
5.20 Acute toxicity of toxaphene to aquatic invertebrates 5-67
5.21 LD50 values for aquatic invertebrates exposed to toxaphene. . . . 5-73
5.22 Occurrence of bottom organisms in toxaphene treated lakes
before and after poisoning 5-78
5.23 Plankton samples taken before and after toxaphene treatment . . . 5-79
5.24 Invertebrates collected before and after treatment with 0.1 ppm
toxaphene 5-80
5.25 Bottom fauna in two Colorado lakes expressed as organisms
per square meter 5-85
5.26 Number of organisms per liter in Wolfe Butte Reservoir before
and after application of 35 ppm toxaphene 5-87
5.27 Number of plant-inhabiting organisms and bottom fauna in
Wolfe Butte Reservoir before and after application of 35 ppm
toxaphene • 5-88
5.28 Numbers of organisms per liter in Raleigh Reservoir before and
after treatment with toxaphene - 25 ppb initially followed by
90 ppb on day 53 5-90
vii
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TABLES (Continued)
Page
5.29 Numbers of plant-inhabiting organisms and bottom fauna in
Raleigh Reservoir before and after treatment with toxaphen ^ ^^
- 25 ppb initially followed by 90 ppb on day 53
5.30 Toxicities to brine shrimp and mosquito larvae of chlorination ^
products of exo-2,10-dichlorobornane
5.31 Toxaphene residues in bobwhite quail, rabbits, and white-taxied
deer collected from treated soybean fields in Alabama during
1968 - 1969
5.32 Toxaphene residues in ring-necked pheasant
tissues 5-106
5.33 Toxaphene residues in fish-eating birds found dead at Tule Lake
and Lower Klamath Refuges between 1960 and 1962 5-107
5.34 Toxaphene residues in dead birds found in treated habitats. . . . 5-108
5.35 Toxaphene residues in bird tissues 5-110
5.36 Toxaphene residues in bird eggs 5-111
5.37 The effects of toxaphene on thyroid size and 2-hour uptake of
I131 by the thyroid of Bobwhite quail 5-112
5.38 The effects of toxaphene on body weight, adrenal weight, and
liver weight of Bobwhite quail 5-113
5.39 Acute toxicity of toxaphene to birds and terrestrial wildlife . . 5-116
5.40 Toxicity data for birds exposed to toxaphene 5-117
5.41 Acute oral LD50 in mg/kg to Mallards of various ages (95%
confidence limits in parentheses) 5-119
5.42 Bird census on selected plots sprayed with toxaphene 5-120
5.43 Toxicity of toxaphene to geese used to control grass and weeds
in cotton fields 5-122
5.44 Egg production, fertility, and hatchability 5-126
5.45 The effect of toxaphene on the hatching of hen's eggs 5-127
5.46 Percent hatchability resulting from injection of toxaphene into
the yolk of fertile eggs after seven days' incubation ...... 5-128
5.47 Effect of toxaphene on percent digestion of dry matter in vitro
by Mule Deer rumen bacteria „ 5-130
viii
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TABLES (Continued)
5.48 Residues of toxaphene in fat samples of meat and poultry
products at slaughter in the United States 5-132
5.49 Toxaphene content of raw whole milk and products
manufactured subsequently 5-133
5.50 Toxaphene concentration (ppm) in milk from cows fed daily
rations containing varying amounts of added toxaphene 5-135
5.51 Toxaphene concentration (ppm) in milk from cows during feed-off
period following feeding of rations containing varying amounts
of added toxaphene 5-136
5.52 Toxaphene (ppm) in milk from cows fed different levels of
toxaphene in the diet 5-137
5.53 Insecticide (ppm) in milk from cows sprayed twice at 3-week
intervals with 0.5% sprays of toxaphene 5-139
5.54 Insecticide (ppm) excreted in milk of dairy cows sprayed twice
daily for 21 days with 1 ounce of 2.0% oil solution of toxaphene. 5-140
5.55 Parts per million of insecticides stored in the fat of cattle
and sheep that had known amounts added to their diet 5-141
5.56 Parts per million of toxaphene in fat of calves sprayed with
0.5 percent toxaphene 5-142
5.57 Parts per million of toxaphene on forage on days indicated
after application 5-143
5.58 Residues in beef cattle fed hay from fields sprayed with
toxaphene 5-144
5.59 Effects of toxaphene sprays applied to suckling calves 5-147
5.60 Effects of oral administration of toxaphene formulations 5-148
5.61 Effects of toxaphene sprays or dips applied to adult animls . . . 5-150
5.62 Penetration of Cl-36 labelled toxaphene into the American
cockroach after topical application of 75 micrograms 5-153
5.63 The distribution of Cl-36 labelled toxaphene in tissues of
Leucophaea maderae 5-154
5.64 Acute toxicity of toxaphene to various insects on topical
application 5-158
ix
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TABLES (Continued)
Page
5.65 Toxicity of toxaphene to boll weevils having various
combinations of larval habitat, age, and adult food 5-166
5.66 Acute topical LDSO's of toxaphene and toxaphene fractions
to the house fly 5-168
5.67 The topical toxicity of toxaphene, two column fractions,
and two components 5-170
5.68 Synergistic effects of toxaphene with DDT and methyl
parathion on topical application 5-176
5.69 Efficacy of toxaphene in the control of various insect pests 5-182
5.70 Efficacy of toxaphene in combination with other insecticides 5-186
5.71 Alkali bee, leafcutting bee, and honey bee 24-hour percent
mortalities from insecticide treatments on alfalfa,
Washington, 1966 5-190
6.1 Uptake of radioactivity in various rat tissues and organs
following a single-dose of Cl-36 toxaphene 6-6
6.2 Distribution of C-14 toxaphene and C-14 toxicant B in rat
tissue after single intubation (residue in ppm) 6-8
6.3 Fat storage of toxaphene in two-year feeding studies of
rats and dogs 6-12
6.4 Excretion of radioactivity in urine and feces of rats
following a 20 mg/kg dose of Cl-36 toxaphene 6-14
6.5 Elimination of labelled toxaphene, toxaphene fractions
I-VII, and toxicants A and B 14 days after single oral dose. 6-15
6.6 Influence of various dietary levels of toxaphene on the
activity of microsomal enzymes of the livers of male and
female rats 6-20
6.7 Estimated dose-response information for toxaphene in oral
and dermal applications 6-24
6.8 Acute oral toxicity of technical toxaphene to laboratory
mammals 6-27
6•9 Case studies of toxaphene poisoning in humans in which
ingestion is the primary route of entry 6-30
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TABLES (Continued)
6.10 Summary of histopathologic findings at autopsy on albino
rats which died from oral administration of toxaphene 6-33
6.11 Acute dermal toxicity of toxaphene to laboratory mammals .... 6-34
6.12 Acute toxicity of toxaphene on inhalation, intravenous, and
intraperitoneal injections 6-37
6.13 Acute intraperitoneal LDSO's of toxaphene and toxaphene
fractions to male mice 6-39
6.14 Acute intraperitoneal LDSO's of toxaphene and toxaphene
fractions to mice 6-40
6.15 Subacute oral toxicity of toxaphene 6-43
6.16 Subacute inhalation toxicity of toxaphene 6-45
6.17 Chronic toxicity of toxaphene at low dietary levels to
laboratory mammals 6-47
6.18 Tumors in mice associated with oral administration of strobane . 6-49
6.19 Toxaphene chronic feeding studies in rats 6-51
6.20 Toxaphene chronic feeding studies in mice 6-52
6.21 Analyses of the incidence of primary tumors in male rats
fed toxaphene in the diet 6-55
6.22 Analyses of the incidence of primary tumors in female rats
fed toxaphene in the diet 6-56
6.23 Analyses of the incidence of primary tumors in male mice
fed toxaphene in the diet 6-58
6.24 Analyses of the incidence of primary tumors in female mice
fed toxaphene in the diet 6-59
6.25 Expected and observed oral LDSO's of toxaphene plus other
pesticides in rats 6-63
7.1 Average monthly atmospheric levels of toxaphene (ng/cu m)
in Stoneville, Mississippi 7-3
7.2 Maximum toxaphene levels found in air samples of nine
localities 7-5
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TABLES (Continued)
Page
7.3 Toxaphene found in air samples from Stoneville, Mississippi
the first day of each sampling week • • 7~6
7.4 Concentrations of airborne toxaphene over the Western
North Atlantic 7~9
7.5 Toxaphene residues in air samples at five North American sites . 7-11
7.6 Toxaphene residues recovered from Flint Creek, Alabama and
the Hartselle Water Treatment Plant 7-14
7.7 Comparison of insecticide recovery from sediment and water,
Hartselle, Alabama Water Treatment Plant 7-15
7.8 Toxaphene concentrations in California surface waters and
their sediments 7-17
7.9 Toxaphene residues in tile effluent drainage from a plot not
previously treated with toxaphene 7-19
7.10 Toxaphene residues in sediments of the lower Mississippi
River and its tributaries in 1966 7-21
7.11 Toxaphene concentrations in three sediment cores, by 10 cm
increments, collected from Terry Creek, Brunswick, Georgia,
June 10, 1971 7-23
7.12 Toxaphene residues in Miller Lake and Davis Lake, Oregon .... 7-25
7.13 Toxaphene in Wisconsin lakes, 1965 . 7-28
7.14 Toxaphene monitoring in U.S. croplands: Application rates
and soil residues 7-29
7.15 Cropland soil monitoring for toxaphene in six states, 1967 . . . 7-32
7.16 Toxaphene in cropland soil by state, fiscal year 1970 7-33
7.17 Toxaphene residue in cropland soil by cropping region,
fiscal year 1970 7-34
7.18 Toxaphene residues in soil from eight cities, 1969 7-35
7.19 Toxaphene in a terrestrial-aquatic model ecosystem 7-44
8.1 Toxaphene production go
8.2 Toxaphene consumption in 1972 „ o_/
xii
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CONTENTS Page
1.0 Summary 1-1
1.1 Discussion of Findings 1-1
1.1.1 Chemical Properties and Analytical Techniques . . . 1-1
1.1.2 Environmental Occurrence 1-3
1.1.3 Environmental Cycling and Fate 1-4
1.1.4 Biological Aspects in Microorganisms 1-5
1.1.5 Biological Aspects in Plants 1-6
1.1.6 Biological Aspects in Wild and Domestic Animals . . 1-7
1.1.7 Biological Aspects in Humans and Test Animals . . . 1-10
1.1.8 Food Chain Interactions 1-12
1.2 Conclusions 1-13
2.0 Chemical and Physical Properties and Analysis 2-1
2.1 Summary 2-1
2.2 Physical Characteristics of Technical Toxaphene 2-3
2.3 Toxaphene Production 2-6
2.3.1 Manufacturing Process 2-6
2.3.2 Chemistry of Toxaphene Production 2-9
2.4 Composition of Toxaphene 2-13
2.4.1 Number and Molecular Formula of Constituents in
Toxaphene 2-13
2.4.2 Comparison of the Compositions of Toxaphene and
Strobane 2-19
2.4.3 Fractionation and Isolation of Toxaphene and
Strobane 2-21
2.5 Chemical Reactivity 2-25
Xlll
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CONTENTS (continued) Pa§e
2.5.1 Hydrolysis 2~25
2.5.2 Reactions Used for Analytical Clean-Up and to
Prepare Derivatives of Toxaphene 2-26
2.5.3 Oxidation-Reduction 2~27
2.5.4 Photochemistry 2"28
2.6 Analysis 2"29
2.6.1 Considerations in Analysis 2-29
2.6.2 Sample Collection and Extraction 2-29
2.6.2.1 Air 2-30
2.6.2.2 Water 2-30
2.6.2.3 Soils and Sediments 2-31
2.6.2.4 Biological Specimens 2-32
2.6.3 Clean-Up and Pretreatment 2-33
2.6.4 Toxaphene Measurement 2-37
2.6.4.1 Gas Chromatography 2-40
2.6.4.1.1 Effect of Environmental Weathering Upon the GC
Peak Pattern 2-40
2.6.4.1.2 Attempts to Simplify the Toxaphene GC Pattern .... 2-44
2.6.4.1.3 Gas Chromatography - Specific Ion Monitoring 2-45
2.6.4.2 Other Chromatographic Methods 2-47
2.6.4.3 Colorimetry 2-50
2.6.4.4 Total Chlorine Determination 2-52
2.6.5 Recommended Toxaphene Analysis 2-52
References „ 2-55
xiv
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CONTENTS (continued) Page
3.0 Biological Aspects in Microorganisms 3-1
3.1 Summary 3-1
3.2 Bacteria 3-1
3.2.1 Metabolism 3-4
3.2.2 Effects 3-4
3.3 Fungi 3-6
3.3.1 Metabolism 3-6
3.3.2 Effects 3-6
3.4 Unicellular Algae 3-8
3.4.1 Uptake and Metabolism 3-9
3.4.2 Effect on Growth 3-11
3.4.3 Effect on Physiological Processes 3-18
3.5 Effect of Toxaphene on Protozoa 3-18
References 3-21
4.0 Biological Aspects in Plants 4-1
4.1 Summary 4-1
4.2 Nonvascular Plants - Higher Fungi, Macro-Algae, and Mosses . . . 4-2
4.3 Vascular Plants 4-2
4.3.1 Noncrop Plants 4-2
4.3.1.1 Metabolism: Uptake, Adsorption, and Residue 4-2
4.3.1.2 Effects 4-8
4.3.2 Crop Plants 4-8
4.3.2.1 Metabolism: Uptake, Adsorption, and Residues .... 4-8
4.3.2.2 Translocation 4-17
4.3.2.3 Effects 4-17
References 4-26
xv
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CONTENTS (.continued)
5.0 Biological Aspects in Wild and Domestic Animals .......... 5~
5.1 Chapter Summary .......................
5.1.1 Fish and Amphibians ...............
5.1.2 Aquatic Invertebrates ......... ..... 5~2
5.1.3 Birds and Terrestrial Wildlife ......... 5-3
5.1.4 Domestic Animals ................ 5-4
5.1.5 Terrestrial Insects and Other Terrestrial
Invertebrates .................. 5-5
5.2 Fish and Amphibians ..................... 5-6
5.2.1 Metabolism ................... 5-6
5.2,1.1 Absorption/Bioconcentration ........... 5-6
5.2.1.2 Transport and Distribution ........... 5-11
5.2.1.3 Biotransformation ................ 5-12
5.2.1.4 Elimination ................... 5-14
5.2.1.5 Residues .................... 5-14
5.2.2 Effects ..................... 5-25
5.2.2.1 Physiological and Biochemical Effects ...... 5-25
5.2.2.2 Toxicity .................... 5-30
5.2.2.2.1 Acute Toxicity ................. 5-30
5.2.2.2.2 Subacute and Chronic Toxicity .......... 5-42
5.2.2.2.3 Reproductive and Teratogenic Effects ...... 5-47
5.2.2.2.4 Resistance/Tolerance .............. 5-47
5.2.2.2.5 Toxaphene Field Studies ............. ' 5-51
5.2.2.2.6 Behavioral Effects ............... 5-54
5.2.2.2.7 Drug Interactions ................ 5-55
5.2.2.2.8 Effects on Amphibians .............. 5-55
xvi
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CONTENTS (continued) Page
5.3 Aquatic Invertebrates 5-57
5.3.1 Metabolism 5-57
5.3.2 Effects 5-63
5.3.2.1 Physiological or Biochemical 5-63
5.3.2.2.1 Sublethal Toxicity 5-64
5.3.2.2.2 Acute Toxicity 5-66
5.3.2.2.3 Chronic Toxicity 5-98
5.3.2.2.4 Embryotoxic Effects 5-99
5.3.2.3 Resistance/Tolerance 5-100
5.4 Birds and Terrestrial Wildlife 5-103
5.4.1 Metabolism 5-103
5.4.1.1 Absorption 5-103
5.4.1.2 Transport and Distribution 5-103
5.4.1.3 Biotransformation 5-103
5.4.1.4 Elimination 5-103
5.4.1.5 Residues 5-103
5.4.2 Effects 5-109
5.4.2.1 Physiological or Biochemical 5-109
5.4.2.2 Toxicity 5-115
5.4.2.2.1 Acute 5-115
5.4.2.2.2 Chronic 5-123
5.4.2.2.3 Other Effects 5-124
5.5 Domestic Animals 5-131
5.5.1 Metabolism 5-131
xvii
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CONTENTS (continued) Page
5.5.1.1 Absorption ................ . • J
5.5.1.2 Transport and Distribution ......... . 5-131
5.5.1.3 Biotransf ormation .............. 5-131
5.5.1.4 Elimination ................. 5-131
5.5.1.5 Residues ................... 5-131
5.5.2 Effects ................... 5-145
5.5.2.1 Physiological or Biochemical ......... 5-145
5.5.2.2 Toxicity ..... .............. 5-145
5.5.2.2.1 Acute .................... 5-145
5.5.2.2.2 Chronic ................... 5-151
5.5.2.2.3 Other .................... 5-151
5.6 Terrestrial Insects and Other Terrestrial Invertebrates . . 5-152
5.6.1 Metabolism .................. 5-152
5.6.2 Effects ................... 5-155
5.6.2.1 Physiological and Biochemical Effects .... 5-155
5.6.2.2 Toxicity ................... 5-157
5.6.2.2.1 Acute Toxicity to Insects .......... 5-157
5.6.2.2.1.1 Acute Toxicity of Technical Toxaphene to Insocts 5-157
5.6.2.2.1.2 Acute Toxicity of Toxaphene Components to
Insects ................... 5-167/
5.6.2.2.2 Resistance in Insects ............ 5-169
5.6.2.2.3 Synergistic Effects in Insects ........ '5-174
5.6.2.2.4 Reproductive Effects in Insects ....... 5-180
5.6.2.2.5 Field Studies ................ 5-180
XVlll
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CONTENTS (continued) Page
5.6.2.2.5.1 Pest Insects 5-181
5.6.2.2.5.2 Beneficial Insects 5-188
5.6.2.2.6 Toxicity to Other Terrestrial Invertebrates . . . 5-191
References 5-193
6.0 Biological Aspects in Humans and Laboratory Animals 6-1
6.1 Chapter Summary 6-1
6.2 Metabolism 6-4
6.2.1 Absorption 6-4
6.2.2 Transport and Distribution 6-5
6.2.3 Biotransformation 6-11
6.2.4 Elimination 6-13
6.3 Effects 6-16
6.3.1 Physiological or Biochemical 6-16
6.3.1.1 Normal Physiological or Biochemical Functions . . 6-16
6.3.1.2 Effects on Normal Physiological or Biochemical
Functions 6-16
6.3.2 Toxicity 6-22
6.3.2.1 Acute Toxicity 6-22
6.3.2.1.1 Technical Toxaphene 6-23
6.3.2.1.1.1 Oral Toxicity 6-23
6.3.2.1.1.2 Acute Dermal Toxicity 6-32
6.3.2.1.1.3 Other Routes of Entry 6-36
6.3.2.1.2 Toxaphene Fractions and Components 6-38
6.3.2.2 Subacute Toxicity 6-42
6.3.2.3 Chronic Toxicity 6-46
6.3.2.4 Carcinogenicity 6-48
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CONTENTS (continued) PaSe
6.3.2.5 Mutagenicity 6~54
6.3.2.6 Teratogenicity 6~61
6.3.2.7 Drug Interactions 6~62
References 6~64
7.0 Environmental Distribution and Transformation 7-1
7.1 Summary 7-1
7.2 Monitoring 7-2
7.2.1 Residues in Air 7-2
7.2.2 Residues in Water and Sediment 7-7
7.2.3 Residues in Soil 7-27
7.3 Environmental Fate 7-36
7.3.1 Mobility and Persistence in Air 7-36
7.3.2 Mobility and Persistence in Water 7-38
7.3.3 Mobility and Persistence in Soil 7-47
References 7-54
8.0 Environmental Interactions and Their Consequences 8-1
8.1 Summary 8-1
8.2 Sources of Toxaphene 8-1
8.2.1 Toxaphene Production and Consumption 8-1
8.3 Environmental Cycling of Toxaphene 8-6
8.4 Food Chains 8-9
8.4.1 Toxaphene in Food 8-9
8.4.2 Terrestrial Ecosystems 8-9
8.4.3 Aquatic Ecosystems 8-10
References > 8-12
xx
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1.0 SUMMARY
1.1 DISCUSSION OF FINDINGS
1.1.1 Chemical Properties and Analytical Techniques
Technical toxaphene is a synthetic organic chemical which is manufactured
by chlorinating the diterpene, camphene, to a 67 to 69 percent chlorine content
(Section 2.3). The technical mixture contains more than 175 polychlorinated
diterpenes, none of which is present in more than a few percent (Section 2.4).
Technical toxaphene is an amber, waxy solid which melts in a range from 65 to
90 C and has a vapor pressure of 0.17 to 0.4 mm Hg at 25 C. Its solubility
in water is ca. 40 micrograms per liter, but it is soluble in most organic
solvents (Section 2.2).
The chemistry of technical toxaphene is typical of the chlorinated paraf-
fins. While it hydrolyzes slowly below pH less than 10, it rapidly reacts with
strong alkali. It is reduced by active metals such as sodium and reacts with
Lewis acids such as aluminum chloride. It shows little photochemical reactivity
within the sunlight spectrum (Section 2.5).
Biochemical behavior has not been studied with technical toxaphene, but
has been examined for two of the isolated toxaphene constituents: Toxicant A
(a mixture of 2,5-endo, 2,6-exo, 8,9,9,10-octachlorobornane and 2,5-endo, 2,6-
exo, 8,8,9,10-octachlorobornane) and Toxicant B (2,5-endo, 2,6-exo, 8,9,10-
heptachlorobornane). Both toxaphene constituents reacted with biological iron
protoporphyrin systems (in vitro) to yield reductively dechlorinated and de-
hydrochlorinated products (Section 2.5).
The recommended method for environmental toxaphene residue analysis is
gas chromatography (GC) with electron capture or microcoulometric detection.
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While GC with detection by mass spectrometry (specific ion monitoring) is
specific and sensitive than the conventional detectors, its cost is prohibitive
for many laboratories. Less expensive (but also less sensitive) alternatives
to GC are colorimetry (diphenylamine derivative) and chromatographic methods
such as thin-layer chromatography (TLC) and reverse phase TLC (Sections 2.6.4.2
and 2.6.4.3).
Numerous interferences, including naturally occurring materials, other
pesticides, and synthetic organic chemicals, can affect GC and other forms of
analysis, with the possible exception of GC with mass spectral detection (Sec-
tion 2.6.4.1.3). To prevent sample contamination (especially by organic plas-
ticizers), glass equipment should be used for collecting and pretreating samples.
Air samples are collected by impingers, filters, and adsorbents. Poly-
urethane foam plugs are an excellent media for collecting atmospheric toxaphene
(Section 2.6.2.1). With aqueous samples the best procedure is to extract toxa-
phene with a pure hydrocarbon solvent such as ri-hexane (Section 2.6.2.2).
Soils and sediments are extracted by a number of techniques; the two most popu-
lar are mechanical shaking and the Soxhlet extraction with solvent (hexane-
acetone azeotrope is the preferred solvent) (Section 2.6.2.3). Biological
samples (including foods) are liquified, if necessary, and extracted with
hydrocarbon solvents. They often contain large concentrations of naturally
occurring interferences, such as fats, oils, and pigments. These can be re-
moved by hexane/acetonitrile partitioning, treating with strong acid, and/or
column chromatography (Section 2.6.2.4). Several procedures are available
for removing synthetic organic chemicals which interfere with toxaphene GC.
Generally, these procedures include treatment with strong acid (e.g., fuming
sulfuric), treatment with strong base, and column chromatography (Sections 2.5.2
and 2.6.3).
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The individual chlorinated diterpenes which comprise the complex toxaphene
mixture differ in their physical, chemical, and toxicological properties. Sev-
eral laboratories have separated toxaphene into fractions based upon differences
in the polarity, adsorption behavior, solubilities, and other properties of
the individual components. Seven distinguishable constituents have been iso-
lated in fairly pure form and their compositions and properties evaluated (Sec-
tion 2.4.3). While one constituent is actually a mixture of two isomeric octa-
chlorinated bornanes, all others have been identified as single compounds. The
eight compounds which have been identified include seven polychlorinated bornanes
and one polychlorinated dihydrocamphene. Environmental weathering apparently
alters toxaphene composition due to differences in rates of chemical and bio-
chemical degradation and removal by physical processes (e.g., evaporation, dis-
solution). However, GC chromatograms of the residues are still recognizable,
although bioassay of these residues demonstrates that the toxicity has often
been considerably changed. While the relationships between the change in
composition and toxicity are important, they are largely unknown.
1.1.2 Environmental Occurrence
Toxaphene is a synthetic product, the only known source of which is commer-
cial production. Therefore, any contamination results from release during pro-
duction or commercial use of toxaphene. Approximately 50 million kg of toxaphene
are annually produced in the United States (Section 8.2).
Soil and water monitoring studies (Sections 7.2.2 and 7.2.3) suggest that
toxaphene is not as widespread an environmental contaminant as other chlorina-
ted hydrocarbon insecticides, such as DDT and dieldrin. Toxaphene is rarely
1-3
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detected in soil, water, or sediment samples that have not received direct or
nearby applications of the pesticide for crop protection (mostly cotton), rough
fish control, or loss from a manufacturing plant. When detected, toxaphene water
concentrations rarely exceed ppb levels, while sediment and soil samples usually
are in the ppm range. 'This lack of widespread detection of toxaphene may be due,
in part, to the difficulty encountered in trace analysis of such a complex mix-
ture of compounds (Section 2.6) and the high volatility of toxaphene (Section 2.2),
In contrast to the lack of water and soil contamination, toxaphene appears to be
a widespread atmospheric contaminant. Sampling of western North Atlantic Ocean
air indicates that the mean levels of toxaphene (0.7 to 0.8 ug/cu m) are equal
to or twice those of PCB's and ten times higher than those of other pesticides
such as DDT (Section 7.2.1). This high concentration probably results from
both the large scale use of toxaphene and its volatility.
1.1.3 Environmental Cycling and Fate
A quantitative understanding of the cycling of toxaphene is not available,
but some field and monitoring studies provide some insight into the processes
that take place. Much of the toxaphene which is released into the environment
will evaporate at a relatively fast rate because of its high volatility. Toxa-
phene in the atmosphere (in the vapor state or adsorbed to particulate matter)
can travel long distances. Toxaphene residues in air have been detected near
the island of Bermuda, 1200 km from the southeastern cotton belt (Section 7.2.1).
Part of the toxaphene that is sprayed on crops will land on the foliage or soil.
The more volatile toxaphene components will evaporate from the foliage, while
the less volatile components are washed onto the soil (Section 8.3). Toxaphene
residues in soil can evaporate, be carried by erosion to rivers and lakes when
1-4
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adsorbed to small particles of soil (less than 1 percent of applied toxaphene
is found in runoff - Section 7.3.3), or leach into ground water. Most studies
have found very little soil leaching, although one investigation demonstrated
that toxaphene could leach through three feet of soil (Section 7.3.3). Toxaphene
residues in water have a tendency to adsorb to suspended particles or sediment,
which explains the low (ppb) concentrations found in water (Section 7.3.2).
Degradation of toxaphene in the environment is poorly understood. Toxaphene
is not susceptible to hydrolysis or photolysis (Section 7.3.2) but does appear
to biodegrade in soil under anaerobic conditions (Section 7.3.3). Several re-
searchers have noted that longer retention time (less volatile) peaks in the
gas chromatogram of soil and water toxaphene residues decrease, while shorter
retention time (more volatile) peaks increase. Residues from water and sediment
do not appear to be products of dehydrohalogenation (Section 7.3.2). The gas
chromatograms of atmospheric toxaphene residues are only slightly changed.
The rate of environmental degradation can vary considerably. One study
determined that the concentration of toxaphene in lake sediment decreased by a
factor of 2 every 20 days. However, concentrations of toxaphene in lakes treated
3 to 9 years before monitoring contained 1 to 4 micrograms per liter in water
and 0.2 to 1 ppm in sediment (Section 7.3.2). The reported half-residence time
for toxaphene in soil varies from 100 days to 11 years (Section 7.3.3). The
latter is considered a maximum value.
1.1.4 Biological Aspects in Microorganisms
Toxaphene is not toxic to soil bacteria and fungi or to the microbiological
processes important to soil fertility at concentrations even higher than those
used for controlling soil insects (Section 3.2.3.3). Inhibition of growth and
1-5
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photosynthesis by toxaphene at concentrations lower than 1 ppm has been observed
in many species of fresh water and marine unicellular alga (Section 3.4.2). No
single toxaphene fraction is exclusively responsible for toxicity. Toxaphene
is stimulatory to growth, carbon fixation, or cell population of algae, soil
bacteria, and fungi at-very low concentrations.
The uptake and metabolism of toxaphene by microorganisms has not been
adequately studied. The available data show that microorganisms do not bioac-
cumulate significant quantities of the pesticide from the medium (Sections 3.2.1,
3.3.1, 3.4.1). No degradation of toxaphene by microorganisms has been noted.
1.1.5 Biological Aspects in Plants
The uptake and metabolism of toxaphene by plants has not received much
investigation. Aquatic plants are able to accumulate appreciable quantities
of toxaphene from the medium (Section 4.3.1.1). Salt marsh (cod grass) col-
lected from a contaminated estuary contained as much as 36.3 ppm toxaphene in
leaves. Significant toxaphene residues have also been detected in many crop
plants (Section 4.3.2.1) when toxaphene is directly applied (rarely occurs
commercially). The residues were higher in most crops immediately after harvest,
but decreased upon incubation. Whether crop residues resulting from direct
application are due to surface deposition or to translocation of toxaphene in
plants is unclear, although the former is thought to be the most important.
Toxaphene residues are reduced by commercial packing house washing and during
food processing (4.3.2.1). Exposure to toxaphene via food is not appreciable
as demonstrated by the FDA market basket survey. Though toxaphene is not
recommended for use in tobacco, residues have been detected in tobacco and
tobacco products.
1-6
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Toxaphene is not toxic to most crop plants at concentrations recommended
for control of insects (15 to 20 kg/ha, annually) (Section 4.3.2.3). Some ad-
verse affect has been noted in the case of cauliflower, cabbage, and tomatoes.
Toxaphene causes off-flavor in tobacco when applied to tobacco foliage. No in-
formation concerning the effect of toxaphene on non-crop plants is available.
1.1.6 Biological Aspects in Wild and Domestic Animals
Both aquatic and terrestrial animals are able to absorb and store toxaphene
in significant quantities. Quantitative data, however, are available only on
aquatic species. Fish and aquatic invertebrates are able to concentrate toxa-
phene from water by factors of several thousand (Sections 5.2.1.1 and 5.3.1).
Although model ecosystem studies indicate that biomagnification may occur (Sec-
tion 8.4.3), one field study has shown that predator fish accumulate less toxa-
phene than prey fish (Section 5.2.1.5). While this finding might suggest dif-
ferent rates of toxaphene biotransformation and/or elimination in these two
groups of fish, experimental data on toxaphene metabolism by aquatic species
is inconclusive. Less is known about the uptake and elimination of toxaphene
in terrestrial animals. Based on residue analyses, birds seem to be able to
absorb toxic quantities of toxaphene from contaminated food (Section 5.4.1.5).
Similar studies in domestic animals have shown that toxaphene may be absorbed
from either oral or dermal exposures with detectable residues being found in
the meat or milk (Section 5.5.1.5). In two species of cockroach, toxaphene has
been shown to penetrate into the hemolymph. While no evidence was found for
toxaphene metabolism in the cockroach, a toxaphene dehydrochlorinase has been
reported in the cotton leafworm (Section 5.6.1).
The acute toxicity of toxaphene varies within each of the animal groups
on which data are available. However, in both aquatic and terrestrial animals,
1-7
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the variability is much more prominent in the invertebrates than in the verte-
brates. In fish, acute LC50 values vary by a factor of about ten, ranging from
3 ppb to 32 ppb (Section 5.2.2.2.1). In aquatic invertebrates, LC50 values vary
by a factor of two and one-half million, ranging from 7.2 ppt to 18 ppm (Sec-
tion 5.3.2.2.2). A similar, though less pronounced pattern, is seen in terres-
trial animals. Acute oral LD50 values for birds range from 10 to 300 mg/kg
(Section 5.4.2.2.1). These estimates are quite similar to the oral LD50 values
seen in experimental mammals - i.e., 25 to 270 mg/kg (Section 6.3.2.1.1.1).
In insects, acute topical LD50 values range from 5.43 mg/kg to 1789 mg/kg (Sec-
tion 5.6.2.2). While this marked variability in invertebrate forms is probably
related to the increased phylogenetic diversity of these animals, the specific
biological mechanisms involved - e.g., different rates of absorption or detoxi-
cation - are not known.
Based on the available chronic toxicity data, aquatic organisms may be
highly susceptible to toxaphene contamination. In water, concentrations of
toxaphene as low as 0.055 ppb have been shown to cause increased backbone fragility!
in fathead minnows over exposure periods of 150 days. Toxaphene concentrations
of 0.39 ppb over 60 day exposure periods also cause decreased growth in brook
trout. Pathological changes in fish chronically exposed to toxaphene include
degenerative changes in liver, kidney, gill, digestive tract, and nerve
tissue (Section 5.2.2.2.2). While less detailed information is available on
chronic toxic effects in aquatic invertebrates, toxaphene at 1 ppb has been
shown to depress growth in oysters. However, field studies on aquatic inver-
tebrate populations suggest that toxaphene concentrations of 0.070 ppb to 0.300 ppl
have no effect on these organisms. Less extensive data are available on terres-
trial organisms. In pelicans, toxaphene given in the food at 50 ppm for three
1-8
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months was lethal. However, in ring-necked pheasants, toxaphene at 300 ppm in
the diet given over the same period had no marked toxic effect (Section 5.4.2.2.2).
In the only subacute study found on domestic animals, young steers given average
doses of toxaphene at 7.9 mg/kg for four months show evidence of transient nervous
system effects (Section 5.5.2.2.2).
Chronic exposure to toxaphene may also affect the reproductive capacity
of many different types of animals. Decreased egg viability has been noted in
spawning female brook trout exposed to 0.075 ppb toxaphene. Developmental ab-
normalities have been seen in striped mullet embryos exposed to 500 ppb toxaphene,
and high mortality was found in clam larvae exposed to 250 ppb toxaphene (Sec-
tions 5.2.2.2.3 and 5.3.2.2.4). However, some field studies on toxaphene treated
lakes indicate that toxaphene does- not inhibit fish productivity at concentra-
tions of up to 1 ppb. Dietary concentrations of 300 ppm toxaphene have resulted
in decreased reproductive success in ring-necked pheasants. However, toxaphene
has not been shown to affect egg shell thickness in quail given single oral doses
of 10 mg/kg, nor does it affect hatchability of hens' eggs injected with 1.5 mg
toxaphene/egg (Section 5.4.2.2.3). In insects, egg production failure was seen
in a population of lady beetles after a single application of toxaphene at
10 micrograms per insect, but no effect on productivity was observed in milkweed
bugs exposed to sublethal levels of toxaphene for 17 generations (Section 5.6.2.2.4)
The long term effects of toxaphene may be substantially influenced by the
development of resistance. True genetic resistance has been most extensively
documented in the terrestrial insects. Various species of lepidoptera, coleoptera,
diptera, hymenoptera, and hemiptera have been shown to possess resistance to
acute toxaphene poisoning. For the most part, resistant insect strains are able
to tolerate doses of toxaphene ten times above those causing high mortality in
1-9
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corresponding susceptible strains. Although the ability to develop resistance
to toxaphene may be inherent in most insect species, the extent to which re-
sistance does develop may be quite limited. For instance, in a study on boll
weevils, a 10 to 12-fold resistance to a 2:1 toxaphene-DDT mixture developed
during a 22 generation exposure period. However, no increase in resistance
developed over an additional 20 generation exposure period (Section 5.6.2.2.3).
Resistance has also been demonstrated in fish. In these animals, resistance
factors of up to 200 have been noted. However, in most instances, resistant
strains are about 40 to 70 times less sensitive to toxaphene than susceptible
strains, and no clear distinction is made between true genetic resistance and
toxaphene tolerance (Section 5.2.2.2.4). Although there is no reason to pre-
sume that toxaphene resistance will not develop in aquatic invertebrates, only
one report of such resistance is available. In this study, a strain of fresh
water shrimp was found to be resistant to toxaphene by a factor of ten. As in
many of the fish studies, no clear distinction was made between resistance and
tolerance (Section 5.3.2.3).
1.1.7 Biological Aspects in Humans and Test Animals
Possible sources of human exposure to toxaphene include contaminated water,
food, or air. Monitoring studies suggest that water and food rarely have de-
tectable levels of toxaphene, except when toxaphene is applied or spilled near-
by the site of contamination. During localized spills, toxaphene can biocon-
centrate in fish that may be consumed by humans. Market basket surveys by FDA,
which include fish, indicate that this type of contamination rarely occurs. In
contrast, human exposure due to contaminated air appears to occur routinely,
based upon monitoring data.
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Toxaphene can be absorbed across the alimentary tract, skin, and respira-
tory tract. Like most lipophilic chlorinated pesticides, toxaphene is stored
primarily in fatty tissue. Although the metabolism of toxaphene in mammals
has not been described in detail, dechlorination mediated by the microsomal
mixed-function oxidase system seems to be a major route of detoxication. In
rats, the biological half-life of orally administered toxaphene is between 1
and 3 days, with elimination taking place via the urine and feces (Section
6.2).
The best estimates of the minimum acute oral lethal dose for man vary
between 30 and 103 mg/kg. These estimates are close to both the acute oral
LD50 values for various laboratory mammals (25 to 220 mg/kg) and, as mentioned
in Section 1.1.6, to the acute oral LD50 values for birds (10 to 300 mg/kg).
The acute dermal toxicity of toxaphene is about an order of magnitude less
than that of oral toxicity. Regardless of the route of administration, the
signs of acute toxaphene intoxication are similar in all mammals. Central
nervous stimulation, evidenced by the development of clonic-tonic convulsions,
is consistently noted. Additional signs include apprehensive behavior, saliva-
tion, vomiting, and hyperreflexia. Exposure to toxaphene-lindane mixtures
has been associated with the development of acute aplastic anemia in man
(Section 6.3.2.1).
The subacute toxicity of toxaphene has not been extensively investigated.
In humans, prolonged occupational exposure to toxaphene vapors may cause res-
piratory insufficiency and reversible lung damage. In one study with rats,
dietary levels of toxaphene at 50 ppm for 2 to 9 months have been associated
with hydropic accumulation of the liver. Other studies, however, have failed
to confirm this effect (Section 6.3.2.2).
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By far the most important studies on toxaphene toxicity are concerned with
toxaphene carcinogenicity, teratogenicity, and other reproductive effects.
Results of a study by the National Cancer Institute indicate that toxaphene fed
to mice at levels of about 100 ppm over a one and one-half year period causes a
high rate of hepatocellular carcinoma. In rats, toxaphene was associated with
an increase in the incidence of thyroid tumors (Section 6.3.2.4).
The teratogenic and reproductive effects of toxaphene are less severe. In
rats, toxaphene doses which caused high maternal mortality were associated with
an increased incidence of encephaloceles. However, at lower doses, toxaphene
does affect implantation frequency, fetal mortality, and fetal weight gain in
rats and mice, and results in decreased numbers of ossification centers in rats
(Section 6.3.2.6). Based on the dominant lethal assay in mice, toxaphene does
not appear to be mutagenic. However, mutagenicity has been demonstrated in
several bacterial assays (Section 6.3.2.5).
1.1.8 Food Chain Interactions
Human ingestion of toxaphene in food is apparently low as demonstrated by
the annual market basket survey conducted by the Food and Drug Administration
(Sections 8.4.1 and 4.3.2.1). This is probably due to the fact that (1) toxa-
phene is mostly used on cotton crops, and (2) washing and food processing re-
move considerable amounts of toxaphene (Section 4.3.2.1). Any residues that
have been detected on crops are thought to be due to surface deposition rather
than uptake from soil, although this has not been well demonstrated experimentally'
Little information on adsorption and distribution of toxaphene in birds
and terrestrial wildlife is available, but residues have been detailed in some
species (especially fish-eating birds) (Section 8.4.2). Controlled exposure
studies have demonstrated that cows fed toxaphene in their diet would have
residues in their milk.
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In aquatic ecosystems, toxaphene bioaccumulates in plants, bacteria, fungi,
green algae, oysters, plankton, daphnia, and fish (Section 8.4.3). In fish,
toxaphene is bioconcentrated by a factor of several thousand. Field studies
show that prey fish stocked in a toxaphene-treated lake concentrate more toxa-
phene than predator fish, which suggests that biomagnification may not be an
important process although other explanations are possible (Section 8.4.3).
1.2 CONCLUSIONS
1. Toxaphene is a mixture of more than 175 polychlorinated diterpenes
which are water insoluble, stable to hydrolysis at pH less than 10 but react
rapidly with strong alkali, reduce with active metals, and are stable under
sunlight irradiation.
2. Using gas chromatography with appropriate clean-up procedures, toxa-
phene concentrations of 1 ppb can be detected.
3. The major source of environmental release of toxaphene is from its
application to cotton as an insecticide.
4. Toxaphene residence time in soil and water is considerably less than
other chlorinated hydrocarbon pesticides, such as DDT, but detectable levels
(0.7 to 0.8 ng/cu m) are found in the atmosphere at sites remote from points
of application, suggesting that the overall environmental persistence is quite
high. Little is known about the environmental fate of toxaphene, one of the
highest volume pesticides presently in use.
5. Toxaphene is not toxic to plants, soil bacteria and fungi, or to the
microbiological processes important to soil fertility. Growth and photosyn-
thesis in unicellular-fresh water and marine algae are inhibited by toxaphene.
Uptake and metabolism of toxaphene by plants and microorganisms have not been
adequately investigated.
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6. Degradation of toxaphene by pure cultures of microorganisms has not
been observed. Microorganisms are unable to concentrate toxaphene to a signi-
ficant extent from the medium.
7. Exposure of humans to toxaphene via food does not appear to be appre-
ciable. Fish can bioconcentrate toxaphene, but market basket surveys, which
include fish, indicate that contamination is not widespread.
8. Aquatic organisms are highly sensitive to technical toxaphene. At
concentrations of 1 ppb and below, laboratory studies indicate that toxaphene
may lead to diminished reproductive capacity, decreased growth, and skeletal
defects. None of these results have been confirmed in field studies.
9. Dietary levels of toxaphene above 50 ppm can be lethal or can cause
decreased reproductive capacity in some species of birds. Such exposures can
occur during the use of toxaphene as an insecticide or piscicide.
10. At dietary levels of 100 ppm, toxaphene is apparently carcinogenic to
mice and possibly rats. Whether the concentrations routinely found in air will
cause cancer, is unknown.
11. Toxaphene has been shown to be mutagenic in microbial assay systems.
12. Toxaphene bioconcentrates in many aquatic organisms. In fish, con-
centrations several thousand times the concentration in water are found in
laboratory and field studies.
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2.0 CHEMICAL AND PHYSICAL PROPERTIES AND ANALYSIS
2.1 SUMMARY
Technical toxaphene is a mixture of more than 175 polychlorinated diter-
penes, all of which are present in concentrations of a few percent or less. It
is produced by chlorinating camphene to a 67 or 69 percent chlorine content.
Toxaphene is similar to a number of other chlorinated diterpenes, the most
important of which is Strobane, a suspected carcinogen. Strobane is prepared
by chlorinating a-pihene to 66 percent chlorine content. Since toxaphene and
Strobane contain many chromatographic peaks of identical retention time and
yield similar mixtures of hydrocarbons when they are reduced, they probably
contain many of the same polychlorinated species.
Overall, the physical and chemical properties of toxaphene and its indi-
vidual constituents are poorly defined. Technical toxaphene is a waxy solid
with a broad melting range (65 to 90 C), moderate vapor pressure (0.17 to
0.4 mm Hg at 25 C), and low solubility in water (ca. 40 micrograms/liter).
Its chemistry is typical of chlorinated paraffins: it hydrolyzes very slowly
in aqueous acid or neutral water but rapidly reacts with strong aqueous alkali,
probably by dehydrohalogenating; it is reactive with Lewis acids, such as alumi-
num chloride; it is reduced by active metals, such as sodium; and it is not
photochemically reactive at sunlight wavelengths.
Properties of the individual constituents apparently differ, based upon
observations of technical toxaphene. Numerous studies on toxaphene separation
have demonstrated that the constituents differ in polarity, adsorption behavior,
and solubilities in a number of solvents. Field studies on applied toxaphene
and its residues all indicate that the constituents differ in vapor pressures,
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aqueous solubilities, chemical reactivities, and other properties. Only eight
constituents have been isolated and identified, of which seven are chlorinated
bornanes and one is a chlorinated dihydrocamphene. The melting points, spectral
properties, and toxicities of the individual constituents are different. The
only work on chemical behavior of the isolated constituents has shown that Casida's
Toxicants A Ca mixture of 2,5-endo, 2,6-exo,8,8,9,10-octachlorobornane and 2,5-
endo, 2,6-exo,8,9.9,10-octachlorobornane) and B (2,5-endo, 2,6-exo,8,9,10-hepta-
chlorobornane) react with biological iron protoporphyrin systems in vitro to yield
reductively dechlorinated and dehydrochlorinated products. However, relative
rates and yields for these reactions and products are unknown.
The differential reactivities and physical properties of toxaphene consti-
tuents make the study of technical toxaphene and its residues more difficult
than other chlorinated hydrocarbon insecticides, such as DDT or aldrin. This
is manifested both in the analysis of toxaphene residues in the environment and
in the interpretation of hazards of the residues.
The recommended method for environmental toxaphene residue analysis is
by gas chromatography (GC) with electron capture or microcoulometric detection.
Detection by mass spectrometry (specific ion monitoring) is more specific and
sensitive for toxaphene residue detection, but its cost is prohibitive for many
laboratories. Less sensitive alternatives for analysis are available, such as
colorimetry (diphenylamine method) and thin-layer chromatography or related
methods (e.g., reverse-phase thin-layer chromatography). Because toxaphene
yields a large number of GC peaks over a broad range of retention times, numer-
ous interferences can affect its analysis. Analytical methods must either re-
move or correct for possible interferences.
2-2
-------
While environmental weathering alters the composition of toxaphene and the
GC pattern of its residues, the pattern can generally be recognized. The compo-
sition change apparently results when constituents are degraded or removed
(evaporation, dissolution, etc.) at different rates. Bioassay has shown that
the weathered residues are no longer toxic. However, the relationships between
the ambient environmental toxaphene residues, environmental transformations and
losses of toxaphene constituents, and the acute and chronic toxicities of the
weathered residues have not adequately been examined.
2.2 PHYSICAL CHARACTERISTICS OF TECHNICAL TOXAPHENE
Toxaphene is defined as chlorinated camphene with a combined chlorine
content of 67 to 69 percent (Guyer et al., 1971; Hartwell et al., 1974). This
complex mixture consists of more than 175 polychlorinated diterpenes (Section 2.4)
The most numerous and important of these constituents are the polychlorinated
bornanes and dihydrocamphenes. The structures and numbering sequences for the
bornane and dihydrocamphene systems are illustrated in Table 2.1.
Table 2.1 enumerates the typical properties and Table 2.2 lists the product
specifications of commercially available toxaphene. Technical toxaphene is an
amber, waxy solid with a vapor pressure ranging from 0.17 to 0.4 mm Hg at 25 C.
The broad melting range (65 to 90 C) characterizes it as a mixture rather than
a pure compound. While it is only slightly soluble in water, it is soluble
in and can be recrystallized from a number of organic solvents (Black, 1974;
Khalifa et al., 1974; Anagnostopoulos et al., 1974).
The literature has limited information on the physical properties of the
individual constituents of toxaphene. Section 2.4.3 describes recent work
published on the isolation of several (eight) constituents. The only physical
2-3
-------
Table 2.1. Physical properties of technical toxaphene
Emperical formula: (-'io^in("'^8
Average molecular weight: 414
Percent chlorine content: 67 to 69%
Physical form: amber, waxy solid
Melting point: 65 to 90 C
Vapor pressure (mm Hg): 0.17 to 0.4 at 25 C
3 to 4 at 90 C
Specific gravity at 100 C/15.6 C: 1.63 (average)
Specific gravity change per degree C: 0.0012
Pounds per gallon at 75 C: 13.8
Viscosity (Centipoises): 89 at 110 C
57 at 120 C
39.1 at 130 C
Specific heat (cal/g): 0.258 at 41 C
0.260 at 95 C
Solubility: 37 microgram/liter in water at ambient temperature. Highly
soluble in most organic solvents; greater solubility in
aromatic solvents
Octanol/water partition coefficient:C 825
.Chemical Structures
Bornane
Dihydrocamphene
(Camphane)
— • • — __—__
Sources: Adapted from Hartwell et al., 1974; Guyer et al., 1971; Hughes, 1970.
5
From Lee et al. (1968). Many reviews list the water solubility at 0.4 mg/liter
based on work by Cohen et al. (1960).
From Sanborn et al. (1976).
2-4
-------
Table 2.2. Toxaphene product specifications
a.
Total organic chlorine, % by weight:
Acidity, % by weight as HC1:
Drop softening point, C:
Infrared absorptivity at 7.2 micron:
Specific gravity at 100 C/15.6 C :
67.0-69.0
0.5 maximum
70 minimum
0.0177 maximum
1.600 minimum
a.
Source: Hartwell et al., 1974
2-5
-------
property measured for the components was their melting points, which are sub-
stantially higher than the melting range for technical toxaphene. Differences
in other physical properties of toxaphene constituents are apparent but have
not been quantitatively measured. The evidence for these differences comes
from observations on the properties and behavior of technical toxaphene.
Numerous studies on toxaphene separation have observed differential solubili-
ties, polarities, and adsorptive characteristics of the constituents (Black,
1974; Seiber et al., 1975; Holmstead et al., 1974; Anagnostopoulos et al.,
1974). Seiber et al. (1975) has cited vapor pressure differences as the cause
of compositional changes in residues for the environmental weathering of field
applied toxaphene. While differences in other physical properties are expec-
ted to influence the changes in the composition of toxaphene residues exposed
to the environment, this logical conclusion has neither been quantified nor
tested in the laboratory or field.
2.3 TOXAPHENE PRODUCTION
2.3.1 Manufacturing Processes
Toxaphene is manufactured by chlorinating camphene to 67 to 69 percent
chlorine content. Figure 2.1 describes the process flow. Camphene is derived
from turpentine, which is either a product of sulfate turpentine from the Kraft
paper process or wood turpentine from the extraction of pine stumps (Guyer et al.,
1971; Hartwell et al., 1974; Buntin, 1951, 1974; Sittig, 1967, Lawless et al.,
1972; von Rumker et al., 1974). The turpentine, which contains primarily
a-pinene, is catalytically isomerized to camphene and subsequently purified by
fractionation (Buntin, 1974). While the chlorination grade camphene may con-
tain some tricyclene, less than 5 percent of other terpenes is present (Guyer et al
1971). The chemistry of the production process is detailed in Section 2.3.2.
2-6
-------
SULFATE
TURPENTINE
ULTRAVIOLET
LIGHT
N>
I
CAMPHENE AND
OTHER TERPENES
FRACTIONATION
REACTION
GRADE
CAMPHENE
CHLORINATOR
TOXAPHENE
SOLUTION
CARBON
TETRACHLORIOE
(SOLVENTI
TURPENTINE FROM
SOUTHERN PINE STUMPS
TOXAPHENE
Figure 2.1. Schematic flow chart for toxaphene manufacture.
von Rumker et al., 1974.
Sources: Lawless et al., 1972;
-------
The process feed consists of a carbon tetrachloride solution of camphene
containing about five parts solvent to one part camphene (Sittig, 1967;
Buntin, 1974). The reactor is a lead-, glass-, or nickel-lined vessel equipped
with a heat-exchange jacket, a reflux condenser, and a well for an ultraviolet
lamp. The reaction is carried out at atmospheric pressure and the initial
reaction temperature is maintained at 85 to 90 C, which requires some cool-
ing. During the latter chlorination stages the temperature ranges from 50 to
75 C. The reaction requires from 15 to 30 hours (Sittig, 1967; Buntin, 1974).
The product work-up consists of blowing out HC1 and excess chlorine and then
distilling carbon tetrachloride in vacuo.
Although toxaphene is a complex mixture, good quality control has appar-
ently maintained a consistent product. Each commercial toxaphene batch is
bioassayed (usually with houseflies, but sometimes with other test organisms)
and physically and chemically analyzed to insure conformation with the product
specifications (Hartwell et al., 1974). The infrared absorbance (at 7.2
microns) is used to distinguish toxaphene from other polychlorinated terpenes,
such as Strobane (Hartwell et al., 1974; Guyer et al., 1971). Hartwell et al.
(1974) report that electron capture gas chromatography (Section 2.6.4.1) is
used to check the composition of each batch. Guyer et al. (1971) report that
nine samples of toxaphene retained from production at Hercules, Inc., during
the period from 1949 to 1970 were analyzed by the above methods and compared.
Since results were consistent, they concluded that technical toxaphene has
remained consistent in its composition and properties over the many years that
it has been produced by Hercules, Inc. Whether technical products from other
manufacturers are as consistent is unknown.
2-8
-------
Other polychlorinated terpenes are manufactured by essentially identical
procedures except that the composition of the starting material and the extent
of chlorination are usually different (Melnikov, 1971; Sittig, 1967). Strobane,
which is the major polychlorinated terpene that is analogous to toxaphene, is
prepared by chlorinating turpentine (which contains mainly a-pinene) to approxi-
mately a 66 percent chlorine content.
Although the pure technical toxaphene appears to have remained consistent
in its composition and properties over the years, there have been some problems
of inconsistency in toxaphene formulation for animal spray and dipping vat
preparations (Sparr et al., 1952; Radeleff and Bushland, 1960). These problems
occurred when unequal concentrations of the insecticide developed during animal
treatment with wettable powders and emulsifiable concentrates. This resulted
in inadequate pest control in some of the treated animals and toxicosis in
others. These formulation problems were solved in the early 1950's and, except
for an occasional misuse on livestock, do not represent a hazard to the animal
industry.
2.3.2 Chemistry of Toxaphene Production
Chlorination of camphene proceeds in two stages: (1) initial ionic addi-
tion of chlorine to unsaturated bonds followed by (2) free radical chlorination.
The course of the ionic chlorination has been partially detailed by chlorinating
camphene in the dark (Black, 1974; Richey et al., 1964, 1965; Jennings and
Hershbach, 1965). Figure 2.2 describes the products of the ionic reactions.
The major, initial products include exo-2,10-dichlorobornane, _3 (60 to 65 percent);
a mixture of cis- and trans-8-chlorocamphene, _2 (19 percent); and exo-6-chloro-
camphene, _5 (18 percent), which is a degradation product of 10-chlorotricyclene,
4_ (Richey et al., 1964, 1965; Jennings and Hershbach, 1965). Black (1974) reports
that subsequent ionic chlorination of 2_, 4_, and _5 yields cis- and trans-isomers
2-9
-------
PRIMARY REACTIONS
"2
CCI,
SCI
SECONDARY REACTIONS
Cl
SCI
4
Cl
Cl
Cl,
Cl
Cl
Cl
Cl,
Cl
Cl
Cl,
Cl
Cl
HCI
XCI
4
Cl
Cl
Cl
Cl
Cl Cl
•Cl Cl
I
Cl
Cl
Cl Cl
Cl
Figure 2.2. Ionic pathways in the chlorination of camphene. Sources'
Black, 1974; Seiber et al., 1975 ; Landrum et al., 1975; Jennings and Hershba1
1965; Richey et al., 1964, 1965.
2-10
-------
of exo-6,8-dichlorocamphene, j^; exo-2, endo-6,10-trichlorobornane, 7_; and exo-
2,10,10-trichlorobornane, _§_.
Products of ionic chlorination are listed in Table 2.3. Some inconsisten-
cies in the compositions reported by Black (1974) and previous work by Richey
et al. (1964, 1965) and Jennings and Hersbach (1965) are apparent but have not
been explained.
Recent work (Seiber et al., 1975; Landrum et al., 1976) suggests that
hydrogen chloride, which is produced in the initial phase of the industrial
chlorination, adds to camphene to yield exo-3-chlorocamphene, j). The assign-
ment of this reaction and product was based upon the isolation and identifi-
cation of a dihydrocamphene derivative from technical toxaphene (Section 2.4.3).
The mono-, di-, etc. polychlorinated camphenes and bornanes formed by
ionic chlorination are subsequently chlorinated by free radical pathways
(Hughes, 1970; Black, 1974; Nelson and Matsumara, 1975 a). The free radical
chlorination apparently follows the pathway:
ci £*—
RH + Cl-
R- + C10
-> 2C1
•*• R- + HC1
-> RC1 + Cl
There has been no published information to describe the preferred site of
chlorination or skeletal rearrangement of the free-radicals analogous to the
Meerwein shifts (rearrangement of the norbonyl structure) of the corresponding
cations.
Nelson and Matsumara (1975 a) examined the products from the free-radical
chlorination of the major ionic addition product, exo-2,6-dichlorobornane.
2-11
-------
Table 2.3. Composition of the products from ionic chlorination of camphene
Compound
fa
Percent
Camphene (_1) - tri cy clene
6-Chlorobornane (_5)
cis-/trans 8-chlorobornene(_2)
cis-/trans 6 ,8-dichlorobornene (6)
Unknown dichlorocamphenes
2,10-Dichlorobornane(3)
2 ,10 ,10-Trichlorobornane (_8)
2,6,10-Trichlorobornane
2,9,10-Trichlorobornane
Total
6-8
16-18
10-11
11-12
2-3
26-27
10-12
1-2
2-3
84-96
^Source: Modified from Black, 1974.
See Figure 2.2 for structure of the compounds.
2-12
-------
The product was reportedly 1.7 times more toxic to certain insects than toxa-
phene. The gas chromatographs of the chlorination product of exo-2,6-dichloro-
bornane and toxaphene are similar, although the relative heights of some peaks
differ.
2.4 COMPOSITION OF TOXAPHENE
2.4.1 Number and Molecular Formula of Constituents in Toxaphene
Literature previous to 1974 suggested that toxaphene contained 30 to 40
principal constituents (Hartwell et al., 1974; Guyer et al., 1971). However,
more recent work discloses that it contains more than 177 individual polychlori-
nated terpenes containing from five to eleven chlorines. None of the constituents
appear in concentrations greater than a few percent.
Holmstead et al. (1974) have published the most rigorous study on toxaphene
composition. They used a combination of column chromatography (CC) and gas
chromatography (GC)-mass spectroscopy (MS) to measure the molecular formula
for individual constituents. The GC pattern of technical toxaphene is too com-
plex to allow measurement of the MS of individual constituents. However, when
technical toxaphene is fractionated by CC, the GC pattern of each fraction is
simplified so that the MS of each peak can be measured. Apparently, GC separ-
ation of the components is dependent upon different properties than the CC
separation (Section 2.4.3).
Holmstead et al. (1974) eluted recrystallized toxaphene on a silica gel
column with hexane and initially collected over 700 fractions. They combined
these initial fractions to yield a total of 18 fractions which were characterized
by relatively distinct GC properties (Dexil-300 on Varaport-30 column). The
molecular formula for each component was evaluated from the GC-MS by using the
[M-C1] positive ion peak for each component. Table 2.4 lists the apparent
2-13
-------
Table 2.4. Components in toxaphene separated by silica gel adsorption chromatography.
and then gas chromatography with qualitative and quantitative analysis
by chemical ionization mass spectrometrya
Chromatographic properties
Number
C10H12C16,
1
2
3
C10HnCl7,
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28b
29
30
*31
32
C10H10C18>
33
1 t
34
35
36
f\ —i
37
38
39
40
Silica gel fraction
[M - Cl]+ = m/e. 307
2
16
16
[M - Cl]+ = m/e. 341
17
1
1-2
12
17
6
3-4
3
5
7-9
5-7
6-8
6
12
10
17
8
6
17
8-9
14
10-12
8-9
14-16
10-14
15
12-13
13
17
[M - Cl]+ = m/e 375
4
2
4
1-2
2
1-2
1-2
4
Gc R , min
9.1
15.6
15.8
9.9
10.7
11.8
12.1
12.2
12.4
12.9
13.3
13.4
14.0
14.2
14.9
15.0
15.2
15.3
15.3
15.4
15.5
15.8
15.9
16.1
16.3
16.3
16.4
16.7
16.9
17.3
17.7
21.1
11.1
11.3
11.3
12.0
12.4
12.7
13.3
13.3
Amount in toxaphene
0.02
0.92
0.52
0.47
0.60
0.45
0.06
0.48
0.04
0,45
0.05
0.44
0.45
0.74
1.25
0.46
0.16
0.97
0.30
0.10
0.60
0.65
0.17
0.47
0.61
0.80
1.58
2.48
1.05
0.90
0.30
0.65
0.07
0.03
0.07
0.24
0.13
0.32
0.76
0.21
^
a
^Source:
Holmstead et al., 1974.
^Toxicant B - see Section 2.3.3.
Toxicant A - see Section 2.3.3.
2-14
-------
Table 2.4 (continued)
Number
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69C
70
71
72
73
74
75
76
77
78
79
80
a
/Source:
Chromatographic
Silica gel fraction
1
4
3-5
1
17-18
4-5
11
7
17-18
6-7
18
1-2
6-7
2-4
5-7
9
17-18
2-4
8-11
14
8
14-16
2-4
11-12
5-8
17-18
5
9-14
5-8
9-11
18
5
11-13
5-9
15
18
13-15
16-18
9-13
14-16
Holmstead et al., 1974
properties
Gc R , min
13.8
13.8
14.6
14.7
14.8
15.0
15.3
15.4
15.5
15.7
16.1
16.2
16.3
16.5
16.7
16.7
16.9
17.1
17.1
17.1
17.4
17.4
17.5
17.5
17.7
17.7
17.9
17.9
18.0
18.2
18.2
18.4
18.4
18.5
18.5
18.5
18.7
18.7
19.1
19.3
Amount in toxaphene
0.09
0.06
1.06
0.56
0.96
0.17
0.21
0.27
0.74
0.39
0.40
0.98
0.21
1.32
0.70
0.09
0.84
1.46
0.56
0.26
0.08
2.40
1.42
0.55
2.00
1.85
0.59
2.39
1.51
1.04
0.46
0.32
0.45
1.81
0.71
0.25
0.92
0.94
2.30
1.61
,Toxicant B - see Section 2.3.3.
"Toxicant A - see Section 2.3.3.
2-15
-------
Table 2.4 (continued)
Chromatographic properties
Number
81
82
83
84
85
86
87
88
89
90
91
92
93
94
95
96
C10H9C
97
98
99
100
101
102
103
104
105
106
107
108
109
110
111
112
113
114
115
116
117
118
119
120
Silica gel fraction
17-18
9-10
12-13
17-18
7-8
11
13-14
6-9
10-13
15
18
15-16
18
13-14
18
12
I19, [M - Cl]+ = m/e. 409
17
1
1
1-2
1-4
4
1
3-4
6-7
3
4-6
1-2
3
4-5
1
5-7
14
17
3-8
10-12
1-5
7-8
18
5
Gc R , min
19.3
19.5
19.5
19.7
19.8
19.8
19.9
20.1
20.3
20.3
20.3
20.7
20.7
21.1
21.1
21.3
13.7
16.9
17.4
17.9
18.0
18.5
18.9
19.0
19.1
19.5
19.7
20.1
20.1
20.1
20.5
20.7
20.7
20.7
20.9
20.9
21.3 .
21.3
21.3
21.5
Amount in toxaphene
0.88
0.92
0.43
0.88
0.83
0.33
0.68
1.79
1.41
0.46
0.74
1.00
0.54
0.17
0.69
0.09
0.30
0.47
0.33
0.14
1.64
0.93
0.04
0.37
0.22
0.33
0.61
0.97
0.35
0.47
0.41
0.69
0.20
0.23
1.56
0.33
0.82
0.31
0.18
0.07
a.
^Source: Holmstead et al., 1974.
,Toxicant B - see Section 2.3.3.
"Toxicant A - see Section 2.3.3.
2-16
-------
Table 2.4 (continued)
Chromatographic properties
Number
121
122
123
124
125
126
127
CioH8clio
128
129
130
131
132
133
134
C10H7C111
135
C10H10C16
136
137
138
139
140
141
142
143
144
145
146
147
148
149
150
C10H9C17,
151
152
153
154
155
Silica gel fraction
11
13-16
6-10
7-11
14-17
14
14-18
, [M - Cl]* = m/e 443
3-5
2-3
5
1-3
2-4
3
1-2
, [M - Cl]+ = m/e 477
4
, [M - Cl]+ = m/e 305
1-2
1
8
2
14
8-9
11-12
14
18
15
15-16
18
13
18
13
[M - Cl]+ = m/e 339
11
14
6
17
8-12
Gc R , min
21.5
21.5
21.7
22.1
22.1
22.4
23.3
19.3
21.9
21.9
22.4
23.3
23.7
24.0
25.4
7.9
9.0
10.1
10.5
10.9
11.3
11.8
11.9
12.7
12.8
13.3
13.3
13.5
14.0
14.9
12.7
12.8
13.1
13.1
13.5
Amount in toxaphene
(%)
0.31
0.84
0.64
0.61
0.64
0.54
1.00
1.09
0.13
0.08
0.37
0.14
0.03
0.21
0.02
0.05
0.13
0.46
0.09
0.10
0.53
0.28
0.16
0.42
0.06
0.25
0.42
0.10
0.03
0.06
0.08
0.08
0,23
0.22
1.03
/Source: Holmstead et al., 1974.
Toxicant B - see Section 2.3.3.
toxicant A - see Section 2.3.3.
2-17
-------
Table 2.4 (continued)
Chromato graphic properties
Number
156
157
158
159
160
161
162
163
C10H8C18,
164
165
166
167
168
169
170
171
172
173
174
175
ClQHyClg,
176
177
ap
/Source:
Silica gel fraction
6
14
11-12
14
16
16
14-15
12-13
[M - Cl]+ = m/e. 373
2-3
2
16-17
1-3
18
5
3-4
1
3
18
3
15
[M - Cl]+ = m/e 407
1
2
Holmstead et al., 1974.
Gc R , min
13.6
14.3
14.7
14.9
14.9
15.2
15.5
16.1
13.8
14.2
14.2
15.0
15.2
15.3
15.6
15.8
15.8
15.8
16.2
17.9
16.5
18.9
Amount in toxaphene
(%)
0.13
0.59
0.81
0.29
0.15
0.37
0.58
0.60
0.13
0.20
0.47
1.47
0.42
0.10
0.46
0.04
0.30
0.59
0.08
0.29
0.36
0.14
cToxicant B - see Section 2.3.3,
Toxicant A - see Section 2.3.3.
2-18
-------
molecular formula for each of the 177 individual constituents which they ob-
served by GC. They have accounted for the observation of a constituent in two
or more fractions, but have not accounted for the possibility that some of the
peaks might have formed in the work-up or during GC. Isomers of Cir.H1AClQ
-LU ID o
and the combined isomers of cinH11Cl7 and C,QH Cl each accounted for approxi-
mately one-third of the total technical toxaphene. Most of the remaining con-
stituents correspond to unsaturated or tricyclic isomers containing from six
to eight chlorine atoms (C, ,.Hn, Cl where n is 6 to 8).
±U ID—n n
Black (1974) used the field ion mass spectrum of toxaphene to approximate
the composition of components containing from five to eleven chlorine atoms.
Table 2.5 summarizes the approximate composition of the polychlorinated species
in toxaphene based on the chlorine isotope envelope.
Holmstead et al. (1974) reduced toxaphene with triphenyl tin hydride.
They could only successfully identify bornane in the GC-MS of the reduction
product. When Beckman and Berkenkotter (1963) used sodium-liquid anhydrous
ammonia to reduce toxaphene, they suspected that a-pinene, camphene, 3-pinene,
d-limonene, and jg-cymene were present in the reduced product by virtue of
identical retention times in the GC patterns (Section 2.4.2).
2.4.2 Comparison of the Compositions of Toxaphene and Strobane
Since Strobane is a suspected carcinogen (Section 6.0), a comparison of
its composition with the composition of toxaphene is important. Strobane,
as well as other polychlorinated terpenes, is closely related in composition
to toxaphene (Guyer et al., 1971; Melnikov, 1971). Although recent research
has partly elucidated the composition of toxaphene, there has been no infor-
mation from comparative studies with other polychlorinated terpenes. Thus,
comparison of toxaphene and Strobane must rely solely on gross data from gas
chromatographic analysis.
2-19
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Table 2.5. Approximate composition of polychlorinated toxaphene constituents
based upon field ion mass spectrum measurementa
Species Ion current (%)
Cl-5 1.3
Cl-6 7.9
Cl-7 19.2
Cl-8 20.4
Cl-9 11.8
Cl-10 5.7
Cl-11 1.8
Total 68.1
Source: Black, 1974.
2-20
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Figure 2.3 compares their gas chromatograms under similar, but not identical,
conditions. Black (1976) has confirmed that the GC profiles of Strobane and
toxaphene are similar; they primarily differ in the relative peak heights of
certain components.
Beckman and Berkenkotter (1963) examined the reduction of Strobane and
toxaphene with sodium-liquid ammonia. Both yielded similar gas chromatographs.
Products, whose identities were assigned according to matching retention times,
include a-pinene, camphene, g-pinene, d-limonene, and p_-cymene. Relative GC
peak heights of the reduction products from Strobane and toxaphene differed.
2.4.3 Fractionation and Isolation of Toxaphene Components
Several groups have used chromatographic methods to fractionate toxaphene
and then have evaluated each fraction for differences in constituent composition,
physical properties, and toxicological properties (Section 6.0). A few of the
constituents have been isolated and structures have been assigned (Table 2.6).
The constituents listed in Table 2.6 are considered the most toxic components.
The constituent separations and individual component isolations have
usually used combinations of chromatographic techniques. The procedures take
advantage of differences in the separation of constituents by the different
chromatographic techniques. For example, while gas chromatography and
partition chromatography apparently separate constituents based upon chlorine
content, column chromatography and thin-layer chromatography separate
constituents based upon polarity (Holmstead et al., 1974).
Seiber et al. (1975) have challenged structural assignments by Black
(1974) and Anagnostopoulos et al. (1974). Seiber et al. (1975) demonstrated
that "non-polar 19" (Black, 1974) and Substance I (Anagnostopoulos et al.,
1974) are identical since they have identical melting points and identical
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10 12 14 «
TkM.MIlWM
18 20
Toxaphene response curves
Column, 30 feet, 30% SE-30 gum rubber on 30-60-mesh Chromosorb W, temp. 275°C,
helium flow 70 cc/min.
10 IS 20 25
Strobane response curves
Column, 2 feet, 20% SE-30 gum rubber on 30-60-mesh Chromosorb W, helium 55 cc/min,
program rate ll°/min, initial temp. 50°C.
Figure 2.3. Comparison of the gas chromatographs of toxaphe'ne and
Strobane. Source: Beckman and Berkenkotter, 1963
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Table 2.6. Structure and properties of constituents isolated from toxaphene
Name and synonyms Molecular
formula
Molecular
structure
Melting
point:,
C
Crystal
structure
Verification
of
structure
Percentage
In
toxaphene
Sources
"Toxicant B"
2,5-endo.2,6-exo,8.9,10-
hcptachlorobornane
C10H11C17
166-167 Orthorhomblc N.M.R.,
(from 5:1 hexnne- x-ray crystal-
acetone) lography
3% " Casida et al., 1974
Khalifa et a]., 1974
Turner et al., 1975
Palmer et al., 1975
(-O
CO
"Tux leant A"
2,5-endo,2,6-exo,8,8.9,10-
octachlorobornane
2,5-endo,2,6-exo,ti,9,9,10-
octachlorobornane
C10H10C18
C10H10C18
134-136
Hexagonal
(from isopropyl
alcohol)
N.M.R.
Casida et al., 1974
Khalifa et al., 1974
6% Turner et al. , 1975
Nelson and
Matsumara, 1975 a,b
2. 5, 6-exo. 8, 8, 9.10-
heptachl'orodlhydro-
camphene
C10HUC17
131-132
Parallelepiped
0.8
Selber et al., 1975
Landrum et al,, 1976
-------
Table 2.6 (continued)
Name and synonyms Molecular Molecular Melting Crystal Verification Percentage Sources
formula structure point, structure of in
C structure toxaphene
Black, ''component 13" C H Cl a^v-^" 215-220 Colorless needles N.M.R.-
2,6-exo,3-endo,8,9,10- iu It 0 ft
hexachlorobornane °r a7~~~L "~7
(tentative) C10H11C17 <='N/-£^a
(from carbon Not certain
tetrachloride)
Black, 1974
Seiber et al. , 1975
NJ
Black - "Non-polar 19"
Anagnostopoulos -
"Substance I"
2,3,5,6-exo,5-endo, or
2,3,5-exo,5,6-endo,
9,10,10-nonachloro-
bornane
C10H9C19
N.M.R.
Stereochemistry
at C-6 is not
certain
Black, 1974
Anagnostopouloe et al.
1974
Seiber et al., 1975
Anagnostopoulos -
"Substance II"
2,3,5-exo,5,6-endo,
8,10,10-octachlorobornane
C10H10C18
N.M.R.
Anagnostopoulos et al.
1974
Anagnos topoulos -
"Substance III"
2,3,5,6-end o,5~exo,
8 »9,10,10-nonachlorobornane
N.M.R.
Anagnostopoulos et al.
1974
-------
GC properties. They could not conclusively identify the stereochemistry at
C-5. Seiber et al. (1975) also suggest that "component 13" (Black, 1974)
is a heptachlorobornane isomer similar in structure to Toxicant B, since their
retention times are identical on three different columns. However, since the
melting points differ, "component 13" and Toxicant B are not identical.
2.5 CHEMICAL REACTIVITY
Chemical reactivity of toxaphene has generally been examined for the mixture
rather than by individual components. Some recent work on toxaphene reduction by
Fe(II) examined reactivity of isolated constituents (Section 2.5.4). Except
for this study (Khalifa et al., 1976), virtually no information is available
on selective reactivity of toxaphene constituents.
Most of the information on toxaphene chemistry involves the lability of
its carbon chlorine bonds (Hughes, 1970). The reactions include hydrolysis,
substitution reactions (for example, with amines), elimination reactions, and
reduction.
2.5.1 Hydrolysis
Mitchum (1963) measured the rates of loss of 1.0 mg/liter toxaphene in
filtered, natural water at temperatures of 7, 18, and 26 C. The aqueous toxa-
phene was held in the dark, but not sealed. The extrapolated time for essen-
tially complete degradation was 645 days at 26 C and 2,680 days at 7 C.
Hughes (1970) has criticized the results since the initial toxaphene concen-
tration exceeded apparent solubility. Hughes suggested that toxaphene loss
might have resulted from evaporation.
Wolfe et al. (1976) have presented evidence which establishes that toxa-
phene is relatively stable to hydrolysis in both acidic and alkaline conditions.
In their procedure, two solutions containing 1.3 ppm toxaphene in 5% aqueous
acetonitrile were adjusted to pH 3.7 and pH 10.0 and then sealed in ampoules.
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After the tubes were heated at 65 C for two days, the solutions were extracted
with chloroform and the extracts were examined by GC. No differences were
found between the GC patterns of the extracts and standard toxaphene.
2.5.2 Reactions Used for Analytical Clean-Up and to Prepare Derivatives of
Toxaphene
The dehydrohalogenation of toxaphene by strong aqueous and alcoholic alkali
has been used to prepare derivatives for GC analysis. Typical reaction condi-
tions are presented in Section 2.6.4.1.2. The products have not been well
characterized. Archer and Crosby (1966), who used ethanolic potassium hydroxide
to simplify toxaphene's GC pattern, have partially characterized the product
by infrared spectroscopy. This product exhibits a weaker absorbance in the 12
to 15 micron region, an increased absorbance in the carbon-hydrogen region at
3.5 microns, and a new carbonyl peak at 5.8 microns. These observations indi-
cate a loss of carbon chlorine bonds along with a gain in olefin and some
carbonyl production.
Several colorimetric analytical procedures utilize derivatives prepared
by toxaphene reactions with amines. The most common derivative is prepared
with diphenylamine (Graupner and Dunn, 1960; Faucheux, 1965; Adamovic, 1968;
Klein and Link, 1970; Thielemann, 1973). The procedures use different tech-
niques, e.g., Lewis acid catalysts or heat, to accelerate the reaction. Some
reaction conditions are discussed in Section 2.6.4.3. The reaction products
have not been characterized.
Toxaphene is stable in strong Bronstead acids. The stability has been
applied to several clean-up procedures: equal volume mixtures of sulfuric
and fuming sulfuric acid (Mills, 1959); sulfuric and fuming nitric acid
(Klein and Link, 1970; Kawano et al., 1969); and acetic and perchloric acid
(Hughes, 1970) (Section 2.6.3).
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2.5.3 Oxidation-Reduction
Metals and metal ions can reductively dechlorinate toxaphene. The only
identified reduction products are chloride ion and less chlorinated terpenes
(Section 2.4.1) (Beckman and Berkenkotter, 1963; Turner et al., 1975).
Toxaphene is more easily reduced by zinc and acetic acid than chlordane,
dieldrin, heptachlor, endrin, or aldrin (Hornstein, 1957; Sweeny and Fischer,
1970). Sweeny and Fischer (1970) examined chloride released by the reaction of
1 g of chlorinated hydrocarbon with 1 g of zinc powder in 20 ml of acetone
and 10 ml of 10 percent aqueous acetic acid. While toxaphene released
61.6 percent of its theoretical chloride, chloride release from the other
chlorinated hydrocarbons ranged from 19.0 to 29.6 percent of theoretical.
Khalifa et al. (1976) have partially elucidated the chemistry by which
iron in biological systems reduces toxaphene. They used standard toxaphene
and two isolated components, Toxicants A and B (see Table 2.6 for molecular
and structural formulas). Two sources of iron complexed with protoporphyrin
were examined: from hematin and from liver microsomes. They demonstrated
that toxaphene degradation proceeds by dehydrochlorination or reductive de-
chlorination and that Fe(II) is the reducing agent. They proposed that the
reductive dechlorination proceeds by the following sequence:
RC1 + Fe(II) ' [(RCl)Fe(II)] : *' + ClFe(III)
I ' I I
I I
R. + Fe(II) + H_0 RH + Fe(III) + OH
I 2 I
Regeneration of Fe(II) required a reducing agent; they used Na^O^ with
hematin and NADPH (nicotinamide adenine dinucleotide phosphate) with micro-
somes. Neither Na^S^O, or NADPH reduced toxaphene in the absence of iron
protoporphyrin.
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Khalifa et al. (1976) partially delineated the reaction products from
Toxicants A and B. Since the reaction products exhibited relatively high
R values (TLC on silica gel), they suggested that the products are more polar
than the original toxaphene constituents. They used GC-MS to identify the
molecular formulas of the primary products from reaction with the hematin-
Na2S204 system. Toxicant A (C10H10Clg) yielded two reductive dechlorination
products (C,QH Cl ) and two products with molecular formula C10H10C6* Toxi~
cant B (C10H1,C17) formed two reductive dechlorination products (C, „!!.. 2C1,),
one dehydrochlorination product (C.0H-nCl,) and two products with the molecular
formula C-.-IL^Cl-.
2.5.4 Photochemistry
Toxaphene appears to be less reactive in response to sunlight irradiation
than in response to irradiation by higher energy light. Available studies have
examined its photochemistry in solution and as a spot on chromatographic
paper.
Mitchell (1961) irradiated 141 pesticide chemicals including toxaphene
(spotted on chromatographic paper) at 253.7 nm. His results suggest that toxa-
phene is degraded more slowly than aromatic organochlorides.
Friedman and Lombardo (1975) irradiated toxaphene (9 to 10 micrograms in
petroleum ether) at 253.7 nm for 90 minutes. Their interest was to eliminate
interferences in the analysis of chlorinated paraffins and not to evaluate the
stability of toxaphene. They observed that after the above irradiation approxi-
mately 36 percent of the toxaphene peak area (GC) remained. Since sunlight
cuts off at 280 nm (Wolfe et al., 1976), toxaphene may be more stable under
environmental conditions.
Wolfe et al. (1976) have examined the effect of simulated sunlight irradia-
tion (cut-off was at 280 nm) of a saturated solution of toxaphene in filtered,
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natural water. They reported that toxaphene degraded slowly, but included no
further information on rates or products. They also reported that toxaphene
is not readily oxidized by photochemically-generated (methylene blue sensitizer)
singlet oxygen in aqueous solution.
2.6 ANALYSIS
2.6.1 Considerations in Analysis
Toxaphene analysis follows the same general procedure as the analysis of
other organochloride residues. Several references have specifically reviewed
the important procedures for toxaphene analysis (Hughes, 1970; Guyer et al.,
1971; Hartwell et al., 1974).
Prior to 1963, gas chromatography was not used for toxaphene analysis.
The most popular methods used then were total chloride determinations and
colorimetric methods (Hartwell et al., 1974). These methods were not as sensi-
tive as GC and their specificity was low in some cases (e.g., total chloride
methods measured all chlorine present in the sample).
Gas chromatographic analysis has become the method of choice for measuring
toxaphene. Since numerous organic materials can interfere with GC analysis,
the experimental procedures reported in the literature should be carefully
examined to insure that interferences were removed before analysis.
2.6.2 Sample Collection and Extraction
Samples taken for toxaphene analysis follow the same general rules as
samples taken for other organochloride residues. The sample should be repre-
sentative of the source and the collection and storage should prevent toxa-
phene loss (e.g., vaporization or degradation) or contamination by interfering
substances (Taras et al., 1971; Thompson et al., 1974). Since containers should
not introduce interferences (especially plasticizers and plastic additives),
all-glass containers and equipment should be used. Thompson et al. (1974)
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recommends that environmental samples should either be extracted within 24
hours or frozen (-12 to -18 C).
2.6.2.1 Air — Air has been sampled for low toxaphene residue concentrations
using high volume samplers (Stanley et al., 1971; Arthur et al., 1976; Bidleman
and Olney, 1974, 1975; Bidleman et al., 1976). Stanley et al. (1971) used a
combination of glass cloth filtering, impingement (Greenburg-Smith impinger
with hexylene glycol as the trapping solvent), and adsorption on alumina. Their
extraction procedure used methods that were developed for measuring food residues.
Arthur et al. (1976) used a MISCO model 88 air sampler with ethylene glycol
as the trapping solvent. Bidleman (1974, 1975, 1976) used a combination of
a glass fiber filter and a polyurethane foam plug. Residues were desorbed
by Soxhlet extraction with petroleum ether.
2.6.2.2 Water — Water can be collected as grab samples or sampled by adsorp-
tion methods, usually with activated carbon. The advantage of adsorption is
that concentrated residues from large volumes of water are obtained; its dis-
advantages include degradation on the column and poor recovery (Hinden, 1964;
Morley, 1966; Faust and Suffet, 1966).
Perhaps typical of the liquid-liquid extraction procedure is the method
used by Schafer et al. (1969). They collected 3.5 liter water samples in
narrow-necked, gallon glass jugs. Organochlorides were extracted by adding
10 ml of hexane to the jug and agitating with magnetic stirring. The hexane
layer was removed by filling the jug with water until the hexane reached the
conical neck; the hexane layer was then removed by pipette. With 2.5 ppb
toxaphene samples, their recovery was 94.7 + 11.5 percent.
One of the more common adsorption procedures is the carbon-chloroform
extraction (CCE) (Nicholson et al., 1964). During CCE procedure, a known
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volume of water is pumped through a carbon cylinder and the toxaphene is
sequentially desorbed by Soxhlet extraction with chloroform. Nicholson et al.
(1964) reported that toxaphene recovery from spiked samples averaged less than
50 percent; at 1.0 ppb, average recovery was 48 percent (range 42 to 51 percent)
and at 0.5 ppb, it averaged 42 percent (39 to 47 percent).
2.6.2.3 Soils and Sediments — Soils, suspended solids, and bottom sediments
are collected and extracted by similar procedures. Several workers (Woolson
and Kearney, 1969; Williams, 1968; Nash and Harris, 1972; Saha et al., 1969;
Saha, 1971) have evaluated the recovery of organochloride pesticides other than
toxaphene by several different extracting methods and with varying amounts of
soil moisture. They concur that recovery by all extraction methods is better
from moist (20 percent water), rather than dry, soils. Saha (1971) considers
hexane-acetone and hexane-isopropyl alcohol azeotropes the solvents of choice.
Saha et al. (1969) reported that recoveries of selected organochlorides with
hexane-isopropyl alcohol extraction by Soxhlet extractor, mechanical shaking,
ultrasonic vibration, ball mill, and blending ranged from 90 to 97 percent for
soil containing 20 percent water.
Mechanical shaking and Soxhlet methods appear the most popular methods
for extracting soils and sediments for toxaphene. Veith and Lee (1971) used
Soxhlet extraction with a hexane-acetone azeotrope for sediments. Their re-
covery for spiked samples with 30 hour extraction time was 88.3 + 2.9 percent.
Parr and Smith (1976) used the same conditions for extracting moist soil. Toxa-
phene recovery from spiked samples extracted from 24 to 48 hours approached
100 percent. Nash and Woolson (1967, 1968) extracted toxaphene and other organo-
chloride insecticides from moist soils by mechanical shaking of the sample with
hexane-isopropyl alcohol (3:1 by volume). While no recoveries were reported for
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toxaphene, they did recover other organochlorides at 85 to 100 percent. Nash et al,
(1973) used the Soxhlet method, mechanical shaking, and column extraction to
extract toxaphene and other organochlorides from moist soils. Solvents were
as follows: for Soxhlet extraction, hexane-acetone-methanol (8:1:1); for
mechanical shaking, hexane-acetone azeotrope; and for column extraction,
hexane: acetone: methanol (4:3:3). While they reported extraction efficiencies
to be best by shaking and poorest by column extraction, they noted that differ-
ences were not statistically significant. Goerlitz and Law (1974) reported a
mechanical shaking technique for extracting toxaphene and other organochloride
pesticides. The sample (50.0 g) is first agitated with acetone (40 ml); then
hexane (80 ml) is added and agitation is resumed. The solvent is decanted and
the procedure repeated. While they did not examine recovery of spiked toxa-
phene samples, recovery of other organochlorides ranged from 75 to 99 percent
with an average of 97.9 percent.
2.6.2.4 Biological Specimens — Plant and animal tissues, food, milk, blood,
and other biological samples are collected and pretreated similarly to soils
and water samples. Tissues should be stored in screw-cap jars and blood in
screw-cap vials. Tissue samples which are to be extracted within 24 hours can
be held at refrigerator temperature (+2 to +4 C), but if they are to be held
longer before extraction, they should be kept at -18 to -12 C (freezer temper-
ature) - Blood samples should be centrifuged as soon as possible after drawing.
The serum may be held at +2 to +4 C for up to 3 days or at -18 to -12 C for
longer periods (Thompson et al., 1974)- Better recovery is reported if sulfuric
acid is added to the whole blood before centrifuging (Stretz and Stahr, 1973;
Griffith and Blanke, 1974). Tissues are homogenized in a food chopper or
Waring blender, then subsequently extracted with a hydrocarbon solvent such as
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hexane, either during the blending or by shaking. Samples with high lipid
content require a partitioning (e.g., hexane/acetonitrile partitioning) to
reduce the lipids to a low enough level for clean-up by sulfonation and chroma-
tography (Section 2.6.3). Samples with low lipid concentration can begin
clean-up at the sulfonation and chromatography steps. The hexane (or other
hydrocarbon) extract is cleaned up and pretreated similarly to extracts from
other sources (Mills, 1959; Giuffrida et al., 1966; Samuel and Hodges, 1967;
Hughes, 1970; Burke, 1971; Thompson et al., 1974).
2.6.3 Clean-Up and Pretreatment
Because toxaphene is a complex mixture and numerous organochlorides can
interfere with its analysis, work-up procedures are more important to its
analysis than for organochloride pesticides of simpler composition, such as
DDT, aldrin, heptachlor, etc. Work-up procedures must remove the naturally
occurring organic products and synthetic organochlorides which will interfere
with the analysis. The procedures use differences between toxaphene and the
interferences in terms of solubility properties, adsorption characteristics,
and chemical reactivity.
Naturally occurring waxes, fats, oils, and pigments are removed by
partitioning, by sulfonation, and/or by column chromatography. Partitioning
is generally used for samples with high fat content (tissues, foods, etc.)
before acid treatment or column chromatography. The method developed by Mills
(1959) has been used for toxaphene as well as the other organochloride pesticides.
In the technique, toxaphene and the naturally occurring lipids are partitioned
between hexane and acetonitrile; the organochlorides are dissolved in the
acetonitrile and the lipids in the hexane. Mills (1959) recommends that 15 ml
or more of hexane be used per gram of fat. If less hexane is used, the dis-
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solved fat-hexane layer becomes more soluble for toxaphene and yields a low
residue measurement.
After high lipid concentrations are reduced by the partitioning proce-
dure, residual lipid concentrations can be removed by sulfonation and chroma-
tography; the sulfonated products are held up on the column. This can be
accomplished either by treating the sample extract in a hydrocarbon solvent
(hexane, petroleum ether, etc.) with a sulfuric-fuming sulfuric acid mixture
or by passing the extract through an acid-Celite column, which is prepared
from 5 grams Celite 545, 1.5 ml of concentrated sulfuric acid, and 1.5 ml of
15 percent fuming sulfuric acid (Hartwell et al., 1974).
While sulfonation removes the naturally occurring interferences, it does
not remove other organochlorides. The stronger conditions of sulfuric acid-
fuming nitric acid treatment do remove some of the organochlorides (e.g., DDT
and methoxychlor), as well as the natural products (Erro, 1967; Guyer et al.,
1971; Hartwell et al., 1974). In procedures developed by Kawano et al. (1969)
and Klein and Link (1970), the sample (ca. 50 to 100 micrograms of toxaphene)
was dried and the residue was dissolved in 5 ml of ice cold sulfuric-fuming
nitric acid. The acid solution was diluted with 150 ml of distilled water and
ice and extracted with petroleum ether (three times). Nitrated DDT and other
organochloride derivatives can then be removed by chromatographic methods.
However, this procedure will not remove polychlorinated biphenyls (PCB's).
Kowano et al. (1969) reports that toxaphene recovery from this treatment
ranges from 79 to 96 percent with a mean of 92 percent.
Archer and Crosby (1966) used alkaline dehydrohalogenation to clean up
organochloride interferences (e.g., DDT is converted to DDE which can be
separated from toxaphene by column chromatography) and to simplify the toxa-
phene GC pattern (Section 2.6.4.1.2). They reacted the toxaphene with 5 g of
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potassium hydroxide dissolved in 20 ml of aqueous alcohol (3 ml of water and
17 ml of ethanol).
Column chromatography (CC) will remove trace residues of natural products
and will fractionate organochlorides. The fractionation can divide organo-
chlorides into smaller groups or can remove potential interferences, such as
PCB's. Mills et al. (1963) developed the standard technique for fractionating
food residues by CC on Florisil using solvents of 6 percent and 15 percent
ethyl ether in petroleum ether; toxaphene elutes with the 6 percent ethyl
ether fraction. Mills et al. (1972) have developed a three solvent system for
CC on Florisil: (A) 20 percent methylene chloride and 80 percent hexane; (B)
50 percent methylene chloride, 0.35 percent acetonitrile, and 49.65 percent
hexane; and (C) 35 percent methylene chloride, 1.5 percent acetonitrile, and
48.5 percent hexane. Solvent A elutes toxaphene. In both the above proce-
dures, PCS (as well as other interfering organochlorides) will elute with the
toxaphene.
Several CC procedures are available which separate toxaphene from PCB's.
Reynolds (1969) found that PCB, heptachlor, aldrin, and DDE are eluted with
hexane while toxaphene, along with heptachlor epoxide, dieldrin, DDT, and DDE,
will elute later with 20 percent ethyl ether in hexane. Armour and Burke
(1970) reported that a silicic acid/Celite 545 column, which was prepared from
5 g acid-washed Celite and 20 g of activated silicic acid, separated PCB from
toxaphene. When they chromatographed the 6 percent ethyl ether/petroleum
ether fraction from a Florisil column, PCB eluted with petroleum ether while
toxaphene was eluted later with acetonitrile-hexane-methylene chloride (1:19:80).
While they did not examine toxaphene recovery, they did recover nine other
organochlorides by this technique at 80 to 107 percent.
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Hughes et al. (1970) used columns packed with multiple adsorbent layers
for toxaphene clean-up. The columns which were used to clean up extracts of
water and suspended solid samples were packed in three layers; Florisil; 1:1
mixture of magnesium oxide and Celite; and then anhydrous sodium sulfate. The
chromatography column for clean-up of plankton extract also contained three
layers: 4:1 mixture of Florisil and Celite; alumina (+ 1 percent water); and
anhydrous sodium sulfate. The former column was eluted with 6 percent ethyl
ether in hexane and the latter with 10 percent ethyl ether in hexane. The MgO
converts DDT to DDE, which elutes before toxaphene, but the MgO does not alter
toxaphene.
Goerlitz and Law (1974) used a two column procedure to separate organo-
chlorides in sediment extracts. They chromatographed 0.5 ml hexane extracts
on 8 cm long (10 mm id) alumina and silica gel columns and collected two
fractions on both. The alumina column yielded two fractions: A-l (0-20 ml
hexane), which contained toxaphene, aldrin, chlordan, DDT and its residues,
heptachlor, lindane, PCS, and polychlorinated napthalenes (PCN's) and A-2
(20-35 ml hexane), which contained dieldrin, endrin, and heptachlor epoxide.
Fraction A-l was chromatographed on silica gel and yielded fraction S-l
(0-25 ml with hexane), which contained aldrin, PCB, and PCN and S-2 (25-45 ml
with benzene), which contained toxaphene and the remaining organochlorides.
Although they did not report toxaphene recoveries, they obtained 75 to 99
percent recovery with fortified samples of nine other organochloride insecti-
cides.
Aquatic sediment samples contain numerous organosulfur compounds which
might not be removed by chromatographic methods. Goerlitz and Law (1971)
report that treating the sediment extract with metallic mercury prior to
column chromatography removes these potential interferences.
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2.6.4 Toxaphene Measurement
Table 2.7 lists analytical methods for toxaphene with approximate lower
detection limits. Table 2.8 compares sensitivity, recovery, and variability
for some toxaphene analysis procedures. Since gas chromatography appears to
be the most widely used procedure, their parameters are of special interest.
Detection by microcoulometry and electron capture exhibit similar sensitivi-
ties (Burke and Holswade, 1966). Toxaphene at environmental concentrations
(ca. 1 ppm) generally exceeded 70 percent recovery.
The analysis for toxaphene in ambient, environmental samples proceeds as
follows: toxaphene must be identified; the identification confirmed; and the
quantity measured. The usual procedure for pesticide analysis in ambient
samples consists of clean-up and then screening by a chromatographic method
(Burke, 1971). Gas chromatography (GC) is the preferred method, but other
chromatographic forms, in particular thin-layer chromatography (TLC), are
useful, less expensive alternatives (Ismail and Bonner, 1974). Gas chroma-
tography identification does require confirmation. Suggested methods include
other chromatographic methods (TLC or reverse phase thin-layer chromatography)
and GC with columns of different polarity. However, an independent measure-
ment is preferred. Gas chromatography using chemical ionization-mass spectro-
metric detection is a relatively recent advancement which provides specific
analysis for toxaphene at a low detection limit. However, capital and opera-
ting costs are substantially higher than other detection methods.
While colorimetry is still sometimes used to measure toxaphene in ambient
samples, infrared spectrometry and total chloride measurements are no longer
used. Total chloride, although sensitive to 100 ng, will measure all chloride.
Unless the sample is known to contain no other source of chloride, the method
2-37
-------
Table 2.7. Analytical methods for toxaphene
a
Method
Approximate
lower limit
of detection
Specific for
Gas chromatography
Electron capture detection 5 ng
Microcoulometric detection 10 ng
Specific ion mass spectrometry 5 ng
Paper chromatography 200 ng
Thin-layer chromatography 500 ng
Colorimetric (Diphenylamine) 5 yg
Infrared spectrometry 10 yg
Total chloride analysis 100 yg
All organochlorides
and other organics
All organochlorides
Specific for toxaphene
but some interferences
might be detected
(e.g., Strobane)
Other organics
Other organics
Other organochlorides
Other organochlorides
and other organics
All sources of chloride
a.
"Source: Burke and Holswade, 1966; Griffith and Blanke, 1974; Faucheux, 1965;
^Ismail and Bonner, 1974; Widmark, 1971; Thurston, 19 7t.
25 to 30 ng for % Full Scale Deflection.
30 ng for % Full Scale Deflection.
2-38
-------
Table 2.8. Measurements of variability, recovery, and sensitivity
for several toxaphene analytical procedures
Method
Sample
Recovery (%)
Variability
Sensitivity
Reference
G.C. - M.C.D."
G.C. - E.C.D. with sulfuric
acid - fuming nitric acid
clean-up
G.C. - E.C.D. with Mills,
Only, Braither work-up
G.C. - M.C.D. - extraction
by sulfuric acid method
G.C. - E.C.D. - extraction
into hexane
G.C. - E.C.I). - column
Standard toxaphene
Standard toxaphene
50 ng
100 ng
200 ng
0. 1 ppm in mills
0. 75 ppm In corn oil
0. 75 ppm in butterfat
86-96%
92-96%
79-97%
80.5
90.0
90.0
100 to 500 ng added
to 2 ml whole blood
2.5 ppb in drinking
water
100%
94.7 + 11.5
+ 11.2%
22 ng (1/2 FSD) Burke and Holswade, 1966
25-30 ng (1/2 FSD) Burke and Holswade, 1966
Kawano et al., 1969
Gluffrida et al., 1966
Griffith and Blanke,
1974
Schafer et al. , 1969
chromatography clean-up
Spectrochemical diphenylamine
with column clean-up or
sulfuric acid methylene
chloride extraction
Combustion and total chloride
analysis
J .R.
0.3 ng in natural water 97
1.0 ng in natural water 100
2.0 ng in natural water 101
From 0 to 1000 ppm in 69-100
grain extracts
0 to 1000 in blank runs 98 + 10
1 to 50 ug
+ 0.22 ng (six determinations)
+ 0.052 ng (six determinations)
+ 0.071 ng (six determinations)
1-12%
+ 7%
+ 1 to 4p
0.5% (absolute)
Hughes et al. , 1970
Graupner and Dunn,
1960
lludy and Dunn, 1957
Clark, 1962
?C.C. - M.C.D. = Cas chromatography with micro-coulometrlc detection.
G.C. - E.C.D. - Gas chromatography with electron capture detection.
-------
cannot be used. Infrared analyses have been used for product assay but are
not useful for ambient samples (Guyer et al., 1971; Hartwell et al., 1974).
2.6.4.1 Gas Chromatography — Gas chromatography is the best developed method
for both identifying and measuring toxaphene residues (Widmark, 1971;
Hartwell et al., 1974). With mass spectral (MS), electron capture, or micro-
coulometric detectors, GC is capable of measuring residues at the ppt range.
Table 2.9 lists some of the columns on which toxaphene retention times have
been characterized.
The major problems in GC analysis result from the complexity of toxaphene1 s
pattern, the pattern changes caused by environmental weathering, and the numerous
interferences with retention times within the range of the toxaphene pattern
(Erro et al., 1967). Table 2.10 lists relative retention times and response
factors (electron capture detector) for potential interfering residues (Burke
and Holswade, 1966). The problems which result from the complex GC pattern
and numerous potential interferences are alleviated by specific clean-up
procedures and by use of key peaks for quantitative analysis. The following
sections focus on the effects of environmental weathering, the methods for
optimum analysis by simplifying the GC pattern, and specific ion detection
methods.
2.6.4.1.1 Effect of environmental weathering upon the GC peak pattern —
Environmental weathering selectively transforms and degrades toxaphene consti-
tuents by chemical, biological, and physical means. Gas chromatographic examina-
tion of weathered toxaphene has established that peaks at long retention times
become reduced in size while new peaks appear at shorter retention times
(Hughes, 1970). Different environmental conditions yield residues with differ-
ing, although recognizable, toxaphene patterns (Hughes, 1970). Bioassay of
these samples showed that the toxaphene had lost its insecticidal activity.
2-40
-------
Table 2.9. Gas chromatographic columns used for toxaphene analysis
Source
Column
Relative retention
(aldrin = 1.00) of
characteristic peaks
Revenue and Beckman (1966)
Windham (1969)
Burke and Holswade (1966)
Burke and Giuffrida (1964)
Archer and Crosby (1966)
5% QF-1 on Chrom W
8% QF-1 + 2% OV-17 on
Gas Chrom Q
(1:1) 15% QF-1 on Gas
Chrom Q and 10% DC-200
on-Gas Chrom Q
10% DC-200 on Anakrom ABS
5% Dow-710 + 5% SE-30 on
Chrom W
6.0, 7.0, 8.0, and others
4.34 and others
See Table 2.10
1.18, 1.51, 1.76, 2.63, 4.31,
4.51 and others
See Figure 2. 4
Hartwell et al. (1974)
3.8% UCW-98 on Diataport S
-------
Table 2.10. Relative retention times of pesticides and related compounds
and their electron capture detection response a>->£•
Pesticide
DDT (tech.)
Methoxychlor
Kelthane (tech.)
6-BHC
Heptachlor
Dimite
Isopropyl ester 2,4,5-T
Dichlone
Dimethoate
Ronnel
Aldrin
Telodrin
0,p'-TDE olefin
Isodrin
Perthane olefin
p,p' -Kelthane
Chlorobenzilate
Methyl parathion
C,p'-DDE
Heptachlor epoxide
Malathion
p,p'-TDE olefin
Chlorbenside
Dae thai
a-Chlordane
3-Chlordane
p,p'-DDE
Parathion
Thiodan
0,p'-TDE
Cap tan
Retention time
relative to aldrin
0.39, 1.88, 2.48, 2.70, 3.28
0.51, 3.23, 4.8
0.58, 0.74, 1.09, 1.31, 1.86, 2.21, 2.37
0.68
0.81
0.82, 1.31
0.84
0.86, 2.04
0.97
0.97
1.00
1.14
1.21
1.25
1.31
1.31
1.33, 2.93
1.42
1.46
1.47
1.48
1.50
1.51
1.52
1.57
1.73
1.88
1.88
1.89, 2.92
2.04
2.10
Response
ng for ^FSD
3.5
10
6
0.5
0.5
15-20
2
4
8
1
0.6
0.8
10
0.8
4C
5
100-200
2.5
2
1
20-30
7
1.8
1.5
2
1.2
1.5
5
2-3
1.5
4
/Source: Burke and Holswade (1966).
Major peaks underlined.
ihe following conditions were used with the gas chromatograph: column: mixture of
equal portions 15% QF-1 (10,000 cst) on 80/100 mesh Gas Chrom Q and 10% DC-200
(12,500 cst) on 80/100 Gas Chrom Q, glass, 6 ft x 4 mm i.d.; column temp. 200°C,
injection temp., 225°C, gas flow rate, 120 ml N2/min; detector: electron capture,
concentric-type, tritium source; detector voltage: DC voltage at which 1 ng
heptachlor epoxide caused \ full scale recorder deflection at 1 x 10 9 AFS
sensitivity, attenuator, 1, recorder, 5 mv.
2-42
-------
Table 2.10 (continued)
Pesticide
Dieldrin
Paraoxon
0,p'-DDT
Endrin
Sulphenone
Kepone
p,p'-TDE
Ovex
p,p'-Methoxychlor olefin
Ethion
p,p'-DDT
Trithion
Endrin alcohol
Endrin aldehyde
p, p' -Methoxychlor
Pro Ian
Dilan
Mi rex
Bulan
Delta Keto 153
(endrin product)
Imidan
Tedion
Guthion
Co-Ral
Chlordane (tech.)
Toxaphene
Retention time
relative to aldrin
2.22
2.25
2.48
2.55
2.62
2.67
2.70
2.78
3.00
3.23
3.28
3.33
3.45
3.98
4.80
4.90
4.40, 4.98, 5.70
5.15
5.70
6.05
7.54
8.75
9.3
23.5
0.72, 0.81, 1.07, 1.55, 1.72, 2.78
1.70, 2.25, 2.37, 2.55, 3.03, 3.66,
4.10, 4.70, 5.8
Response
ng for ^FSD
1-1.5
15-20
3
2.5-3
3-4
7
1.5
3-4
10-12
10
3
4
4
4.5
10
10-15
15
6
5-10
7
10
8-10
50-100
50-75
7
25-30
Strobane
1.09, 1.38, 1.53, 1.69, 2.16, 2.52,
3.01, 3.56, 4.57
40
2-43
-------
2.6.4.1.2 Attempts to simplify the toxaphene GC pattern — Three methods of
simplifying the toxaphene analysis by GC have been evaluated: peaks can be
condensed by use of short columns; the number of peaks can be reduced by chemi-
cally making derivatives and quantitation can use a few characteristic peaks
instead of the whole pattern.
Witt et al. (1962) reduced the complex toxaphene pattern into a single
peak by using a 1 1/4 foot column instead of a conventional 6 foot column.
They reported a retention time of less than 2 minutes and detection to 500 ng
(microcoulometry). The method, however, is not usually feasible when other
residues with similar retention times are present, because those peaks would
overlap (Section 2.6.4.1.3).
Several groups evaluated quantitation by the use of a few key peaks of
the complex toxaphene patterns. Gaul (1966) recommended toxaphene measurement
by planimetry of the four final peaks of a chromatogram using 10 percent DC-200.
The procedure is accurate to two significant figures and yields similar values
to calculations using the whole toxaphene pattern. Gaul (1966) pointed out,
however, that GC patterns for weathered residues will differ from standard
toxaphene and results might differ between the two methods. .DDT and methoxy-
chlor can influence baseline construction for the final four peaks, but their
effect can be corrected. Bevenue and Beckman (1966) recommended calculations
use the three peak fingerprints of chromatograms obtained in a 5 foot column of
5 percent QF-1 on Chromosorb W. The toxaphene fingerprint elutes at relative
retention times (aldrin = 1.00) of 6.0, 7.0, and 8.0, while p,p'-DDT elutes
at 4.1. Klein and Link (1967) reported using various individual peaks for
estimating toxaphene.
2-44
-------
Erro et al. (1967) have criticized exclusive reliance on a fingerprint.
They report that the last four peaks might not be present, which they attribute
to weathering of the sample and/or the condition of the GC column. They con-
cluded that removal of the interferences is the surest procedure and suggested
that all samples which are to be analyzed for toxaphene should be treated with
sulfuric-fuming nitric acid.
Archer and Crosby (1966) recommended alkaline dehydrohalogenation of samples
containing toxaphene. Figure 2.4 illustrates the effect of dehydrohalogenation
on the chromatogram of toxaphene on a 9 foot column packed with 5 percent DC-710
and 5 percent SE-30 on Chromosorb W at 200 C and nitrogen flow of 40 to 60 ml/min.
The major toxaphene peak (3.50 minutes) elutes before DDT residues (6.25 min).
2.6.4.1.3 Gas chromatography - Specific ion monitoring—The best method for
toxaphene analysis is gas chromatography with specific ion monitoring (SIM) .
The technique has been developed to yield a Limited Mass Reconstructed Gas
Chromatogram (LMRGC) virtually specific for toxaphene; it is capable of detecting
less than 5 ng (Thurston, 1976).
The method requires chemical ionization mass spectrometry (CI-MS), since
the CI mode yields better resolution for the major fragments such as [M-C1],
[M-C1-HC1], and [M-C1-2HC1] ions. Electron impact mass spectrometry is not
sufficiently sensitive (Thurston, 1976; Holmstead et al., 1974; Stalling and
Huckins, 1975). The technique has been developed for the Finnigan 1015 quad-
ruple mass spectrum with System 150 data system. The programs are written for
PDF 8 (Thurston, 1976; Neher and Hoyland, 1974; Budde, 1976; Eichelberger et al.,
1974).
2-45
-------
10 8 6
Minutes
Figure 2.4. Gas chromatogram from 30 ng of toxaphene
(A) before alkali treatment and after alkali treatment (B).
Archer and Crosby, 1966.
Source:
2-46
-------
Thurston (1976) has developed LMRGC for a 60 cm x 2 nun (I.D.) glass
column packed with 3 percent SP 2100 on 80/100 Supelcon AW. A short column
can be used since detection is specific for toxaphene. The m/e 307 and 343
ions are judged as the optimum fragments for monitoring toxaphene (Thurston,
1976) . Tetrachlorobiphenyl monitored at m/e 291 or 295 is used as internal
standard. Figure 2.5 compares the reconstructed gas chromatogram (RGC) and
LMRGC for a 10:1:1:1 mixture of toxaphene, DDE, TDE, and DDT to the LMRGC for
toxaphene. Toxaphene is obscured in the RGC (reconstructed gas chromatogram),
but well-defined by its LMRGC. LMRGC also specifically detects toxaphene with
Arochlor 1260 present. Figure 2.6 compares LMRGC for toxaphene added to New
Orleans drinking water in which 80 organic chemicals were found in concentra-
tions of 0.05 to 10 micrograms per liter.
2.6.4.2 Other Chromatographic Methods—Paper chromatography, thin-layer
chromatography (TLC), and reverse phase thin-layer chromatography (RPTLC) are
less expensive although less sensitive analytical methods than GC. They can
be used to screen samples, to confirm residues identified by another method,
or for quantitative purposes [optical measurements are recommended for quanti-
tative analysis (Getz, 1971)].
Since toxaphene chromatographs as streaks in TLC methods just as it does
in GC, potential interferences must also be eliminated (Bontoyan and Jung, 1972;
Walker and Beroza, 1963; Ismail and Bonner, 1974). Alkaline treatment to simplify
chromatographic patterns has not been evaluated for chromatography other than GC.
Hartwell et al. (1974) and Guyer et al. (1971) recommend TLC on alumina
oxide treated with silver nitrate using hexane as the solvent. The technique,
as developed by Schechter (1963) and Moats (1966), has a limit of detection to
ca. 500 ng for toxaphene, which appears as a streak. The color is developed
by exposing the plate to ultraviolet light.
2-47
-------
R.I
5 I
R-J
sj
Sj
Toxaphene + DDT's. RGC.
38 * SB 98 TO 98 30 189 110 120 138 11O ISO ISO I TO
(b)
Toxaphene + DDT's.
LMRGC at 307 m/e.
a 10 20 3D •XJ SO ISO 70 W 30 190 110 IZ9 138 11O ISO ISO ITS
\CJToxaphene Standard.
LMRGC at 307 m/e.
9 19 ZB 3) K5 SO SO 79 « 3) IOO 1IO 1ZB 13B |-*J ISO I«B 170
Figure 2.5. Specific ion monitoring with gas chromatography of
toxaphene and DDT residues. Source: Thurston, 1976,
2-48
-------
New Orleans Drinking Water
Extract. 2 ul of 1:25,000
CHCl,. solution plus 0.3 ng
internal standard TCB.
New Orleans Drinking Water
Extract. 2 ul of 1:25,000
CHCl solution plus 0.3 ng
internal standard TCB plus
5 ng toxaphene.
New Orleans Drinking Water
Extract. 2 pi of 1:25,000
CHCl solution plus 0.3 ng
internal standard TCB plus
15 ng toxaphene.
a
b
Figure 2.6. LMRGC for toxaphene in New Orleans drinking water
extract. Source: Thurston, 1976.
2-49
-------
Moats (1966) prepared TLC plates at 250 microns thickness from aluminum
oxide G or H, or silica gel G or H by adding 0.2 g silver nitrate (dissolved in
aqueous alcohol) to a slurry of 30 g adsorbent in 200 to 300 ml water. The
samples were cleaned up by Florisil chromatography prior to analysis. Toxaphene
chromatographed on aluminum oxide G with hexane development yielded a streak
from Rf 0.12 to 0.40.
Ismail and Bonner (1974) have developed a similar RPTLC. They treated 30
g of aluminum oxide with 60 ml of 0.2 percent nitric acid and 10 ml of 1
percent aqueous silver nitrate. Their plates are subsequently coated to no
more than 250 microns thickness, air dried overnight, heated to 80 C for 30
minutes, cooled, and then saturated with 8 percent liquid paraffin in petroleum
ether. Their mobile solvent is acetonitrile-acetone-methanol-water (40:18:40:2),
which is saturated with paraffin oil. Color was developed by exposing the
plate to light. Toxaphene (2.5 micrograms) appeared as a characteristic
series of spots.
Faucheux (1965) used diphenylamine-zinc chloride for detecting toxaphene
and 33 other pesticides. Toxaphene was chromatographed on aluminum oxide and
developed with n-heptane. The chromogenic reagent yielded a grayish-green
streak for toxaphene (and Strobane) which would be distinguished from chlordane
(a purple streak) and other insecticide residues. Lower limit of detection is
ca. 3 micrograms.
2.6.4.3 Colorimetry—The product obtained by reacting toxaphene with diphenylamine
distinguishes toxaphene from other pesticides and has also been used for quanti-
tative analysis (Graupner and Dunn, 1960; Faucheux, 1965; Adamovic, 1968; Klein
and Link, 1970; Thielemann, 1973). Table 2.11 compares the product color from
toxaphene with colors produced with other pesticides. Graupner and Dunn (1960)
reported that toxaphene has an absorption maximum at 640 nm.
2-50
-------
Table 2.-J-. Comparison of colors obtained by
the reaction of pesticides with
diphenylamine-zinc chloride
Pesticide Product color
Toxaphene Bluish-green
Rothane Yellow
Lindane Colorless
DDT Orange-green
Endrin Green
Heptachlor Green
Methoxychlor- Yellow-green
Aldrin Purple
Dieldrin Purple
Chlordane Purple-blue
Kelthane Purple
Source: Adapted from Graupner and Dunn, 1960, and Faucheux, 1965.
2-51
-------
Klein and Link (1970) suggest the following procedure for toxaphene analysis.
The sample is cleaned up by the sulfuric-fuming nitric acid method. To ca. 20
micrograms of toxaphene residue are added 0.2 ml each of p_-diphenylamine sulfonic
acid sodium salt in methanol, and 0.50 percent zinc chloride in acetone. The
solvent is stripped and the residue heated for 2 minutes at 210 C. The residue
is dissolved with 10 ml ethanol and the absorbance is read at 625 nm. Concen-
trations measured by this method (down to ca. 0.15 ppm) were approximately
10 percent higher than from GC (electron capture detection).
2.6.4.4 Total Chlorine Determination—Toxaphene concentrations can be determined
by measurement of liberated chloride. The method is useful only where other
sources of chloride ion will not interfere. The method is not practical for
measuring ambient samples, but is discussed because of its historical importance
before GC was introduced. Chloride determination can be separated into two
parts: chloride liberation and chloride measurement.
Chloride has been quantitatively liberated from toxaphene by combustion
(Schoniger flask) and active metal reactions (Guyer et al., 1971; Hartwell et al.,
1974). Active metal systems used for this purpose include sodium-isopropyl
alcohol, sodium-liquid ammonia, sodium biphenyl, and zinc-acetic acid (Guyer et al.
1971; Hornstein, 1957). Analytical methods for the generated chloride ion in-
clude amperometric titration using silver coulometry, spectrophotometric analysis
using mercury thiocyanate-ferric ion, and specific ion electrode (Guyer et al.,
1971; Widmark, 1971).
2.6.5 Recommended Toxaphene Analysis
Table 2.12 summarizes the recommended method of toxaphene analysis, as
adapted from Hartwell et al. (1974) and Guyer et al. (1971). Hartwell et al.
(1974) recommended the use of 5 percent SE-30 and 5 percent DC-710 on Gas Chrom Q
2-52
-------
Table 2.12. Recommended procedure for toxaphene measurement
Procedure
Function
K5
Ln
Extract the sample with n-hexane.
Chromatographic clean-up by eluting the hexane
solution through sulfuric acid/Celite 545
column with 100 ml of ri-hexane
Optional chromatographic clean-up on Florisil
(6% ethyl ether/petroleum ether).
Dehydrohalogenate by evaporating the hexane and
treating the residue with 25% KOH in aqueous
ethanol at 75 to 80 C for 15 minutes.
Extract by diluting the alkaline mixture with
distilled water. Extract with 0.5 ml n-hexane.
Gas chromatography (electron capture or micro-
coulometric detection).
Column: 9' x 1/8" i.d. glass of 1:1 mixture
5% SE-30 and 5% DC-710 on Gas Chrom Q
Operating conditions: 200-210 C
Removes natural products (e.g., waxes, fats).
Removes PCB and other chlorinated hydrocarbon
interferences.
DDT and other chlorinated organics do not
interfere with GC analysis of the derivative.
Source: Adapted from Guyer et al. (1971) and Hartwell et al. (1974)
-------
as the column and recommended it be conditioned for two days at 250 C. The
method is designed for standard GC detection systems: microcoulometer or
electron capture. If mass spectral specific ion monitoring is used, the de-
hydrohalogenation step should be eliminated.
2-54
-------
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2-56
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2-57
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*—
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/
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J. Ass. Offic. Anal. Chem. 55:39-43.
f
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2-58
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Crystal and Molecular Structure of 2,2,5-Endo, 6-Exo, 8,9,10-Heptachlorobornane,
cIOHllCl7> a Toxic Component of Toxaphene Insecticide. J. Amer. Chem. Soc. 97;
408-413.
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3.0 BIOLOGICAL ASPECTS IN MICROORGANISMS
3.1 SUMMARY
Toxaphene does not significantly affect soil microorganisms - bacteria and
fungi, or microbiological processes important to soil fertility - e.g., nitri-
fication, organic matter decomposition, nodulation of legumes - at concentra-
tions even higher than those used for controlling soil insects. At lower con-
centrations, toxaphene stimulates growth, carbon fixation, or cell population
of microorganisms.
Unicellular freshwater- and marine algae are quite suceptible to toxaphene;
growth and photosynthesis are inhibited at fairly low concentrations (^ 1
ppm). Marine algae in general are more susceptible than freshwater algae. No
single, isolated fraction of toxaphene is exclusively responsible for toxicity.
One toxic fraction has been partially characterized as an octachlorobornane.
Toxaphene is also inhibitory to freshwater protozoans at a concentration of
60 ppb.
Bacteria, fungi, and unicellular algae are unable to degrade detectable
quantities of toxaphene. The microorganisms, however, sorb significant amounts
of the pesticide rapidly from the medium. The values of the distribution
constant for various microorganisms for toxaphene were in the range of 3.4 to
17.0.
3.2 BACTERIA
Toxaphene, an agricultural insecticide, is applied predominately to soil
for the control of soil insects. Its interaction with soil bacteria has received
the most attention. Studies dealing with the effect of toxaphene on soil bac-
teria and soil bacterial processes are summarized in Table 3.1 and Table 3.2.
3-1
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Table 3.1. Effect of field application of toxaphene on soil bacteria
Reference
Type of
soil
Concn. of
toxaphene
Period of
treatment
Effect on numbers
of soil bacteria
(some studies in-
clude actinomycetes)
U)
IS5
Bollen et al.,
1954
Martin et al.,
1959
Eno et al.,
1964
Eno and Everett,
1958
Smith and Wenzel,
1948
Elfadl and Fahmy,
1958
1. Chehalis silty clay
loam
2. Lake labish peat
1. Ramona sandy
loam
2. Holtville sandy
clay loam
Leon fine sand
Arrendondo
loamy fine
sand
Branchville sandy
loam and cadorus
silt loam (with
and without
cottonseed meal)
Silt loam soil
11.2 to 22.4 kg/ha
22.4 kg/ha,
5 annual
applications
16.8 kg/ha,
12 semi-
annual
applications
12.5 to 100 ppm
250 to 1250 ppm
(simulating
112 to 560 kg/ha
acre in the field)
4.9 to 40.3 kg/ha
10 to 20 days
5 to 6 years
3 to 6 years
1 to 16 months
incubation in
greenhouse
5 to 96 days in-
cubation in
greenhouse
30 to 136 days
incubation in
greenhouse
No measurable effect
No measurable effect
No measurable effect
No measurable effect
Stimulation of growth
Lower concns: stimu-
latory;
Higher concns.: some-
what inhibitory
-------
Table 3.2. Effect of toxaphene on processes in soil catalyzed by bacteria
Reference
Type of
soil
Soil microbial
function
examined
Concn. of
toxaphene
Period of
treatment
Effect
Eno and Everett,
1958
Arredondo
loamy fine
sand
Nitrification, C02
evolution, nodu-
lation of legumes
12.5 to 100 ppm 1 to 16 months Stimulation of
in greenhouse nitrification
and CO- evolu-
tion; no signi-
ficant effect on
nodulation
U)
I
U)
Martin et al., 1. Ramona sandy Organic matter decom- 22.4 kg/ha
1959 loam position, nitri-
2. Holtville sandy fication
clay loam
5 to 6 years
No effect
Eno et al.,
1964
Elfadl and
Fahmy, 1958
Leon fine
sand
Silt loam
Nitrification
CO^-production
urea hydrolysis
Nodulation of
legumes
16.8 kg/ha,
12 semi-
annual
applications
4.9 to
40.3 kg/ha
3 to 6 years
30 to 136 days
incubation in
greenhouse
No significant
effect
Stimulation at
low doses up
to 9 Ibs/acre;
No effect at
higher con-
centrations
-------
3.2.1 Metabolism
The uptake and metabolism of toxaphene in bacteria has been reported by
Paris et al. (1975) in some detail; this is the only study on the subject.
The test organisms used included a gram negative bacterium, Flavobacterium
harrisonii, isolated from citrus plant effluent, and a gram positive bacterium,
Bacillus subtilis, isolated from Soya Creek. The bacteria were unable to
degrade detectable quantities of toxaphene. The sorption of toxaphene by
bacteria, as determined by following the decrease in pesticide concentration
in the culture medium, was rapid. The cultures reached equilibrium with
toxaphene in the medium within 30 minutes and no further change was noted over
24 hours. Extraction of toxaphene from the cells accounted for all the pesti-
cide in the parent form. Sorption of pesticide followed the Freundlich equation;
the values of the distribution coefficient (mg pesticide per mg dry wt of the
organism/mg pesticide per mg water) were calculated to be 3.4 and 5.2 for ]J.
subtilis and JF. harrisonii, respectively, which shows that the 1J. subtilis
sorbed toxaphene less effectively than _F. harrisonii.
When microorganisms with sorbed pesticides move to aqueous environments,
they release some of the compound to the environment. The sorption in B^.
subtilis and 7_. harrisonii was reversible (Paris et al., 1975). Desorption of
equilibrium was achieved in the same time with comparable k values.
3.2.2 Effects
Information concerning the interaction of toxaphene with soil bacteria
has been derived largely from field studies. Bollen et al. (1954) added
11.2 -22.4 kg of toxaphene per hectare to field soils. After 10 and 20 days,
no effect on numbers of bacteria and streptomyces was noted. In another
3-4
-------
study, five annual applications of toxaphene at 22.4 kg/ha in the field soils
produced no measurable effect on numbers of soil bacteria during the five year
period and up to 10 months after the fifth application (Martin et al., 1959).
Other researchers (e.g., see Eno et al., 1964) have also found no measurable
change in soil bacterial population following addition of toxaphene. This
inference is also supported from the studies carried out under controlled
conditions in a greenhouse (Elfadl and Fahmy, 1958; Eno and Everett, 1958). A
few studies have shown that toxaphene stimulates bacterial growth in soil at
low concentrations (Smith and Wenzel, 1948; Elfadl and Fahmy, 1958; Eno and
Everett, 1958). The stimulatory effect may be due to a catalytic effect of
small quantities of toxaphene on certain physiological processes of the cell,
or to a favorable shift in the ecological balance in the soil, e.g., destruc-
tion of certain predators (Eno and Everett, 1958). The actinomyces, on the
other hand, failed to show any increase due to the insecticide.
The effect of toxaphene on the microbiological processes important to
soil fertility, e.g., nitrification, organic matter decomposition, etc., has
been studied by many researchers. Eno and Everett (1958) reported stimulation
of nitrification (^ 10%) and C0~-evolution-rate (^ 8%) one month after applica-
tion of toxaphene to soil. The concentration of toxaphene ranged from 12.5 to
100 ppm (approximately equivalent to 28-224 kg/ha of the active ingredient of
the insecticide). Sixteen months after insecticide application, no significant
change occurred in the microbiological processes. No significant effect by
toxaphene on soil microbiological processes was demonstrated by other investi-
gators (Eno et al., 1964; Martin et al., 1959).
To evaluate the effect of toxaphene on nodulation of legumes, Eno and
Everett (1958) treated soil with toxaphene and 35 days later planted with
3-5
-------
Rhizobium-inoculated sweet clover seed. An inspection for nodules 10 weeks
later revealed no significant effect on nodulation on the clover grown in
treated soil. Similarly, Elfadl and Fahmy (1958) noted no effect of toxaphene
at the higher concentrations tested (40.3 kg/ha) on nodulation of Rhizobium-
inoculated cowpea. At lower concentrations of toxaphene (< 10 kg/ha), an
increase of 20% to 40% was observed in the number of nodules per plant.
3.3 FUNGI
3.3.1 Metabolism
The degradation and accumulation of toxaphene by a pure culture of fungi
has been briefly investigated by Paris et al. (1975). In this study, an
Aspergillus sp. was isolated from a chicken plant effluent on Rose Bengal
agar. Subsequently, it was grown on glucose in the presence of toxaphene.
The fungus was unable to degrade detectable amounts of toxaphene during a 12
week incubation period. The sorption of toxaphene by Aspergillus sp. was
relatively slow with the cultures reaching equilibrium with the medium in
nearly two hours. The fungi formed clumps while growing but the time required
for equilibration was independent of the clump size. The sorption of the
insecticide followed the Freundlich equation, and the value of distribution
coefficient was calculated to be 17.0, which is higher than for bacteria
(Section 3.2). This suggests that fungi sorb toxaphene more effectively than
bacteria. The ability of toxaphene to desorb from fungi when placed in toxaphene-
free environment was not investigated.
3.3.2 Effects
A large number of the reported studies have dealt with the effect of tox-
aphene on soil fungi. The experimental conditions and results of the studies
are summarized in Table 3.3.
3-6
-------
Table 3.3. Toxicity of toxaphene to soil fungi
Reference
Type of
soil
Concn. of
toxaphene
Period & conditions
of treatment
Effect on
growth
Eno and Everett,
1958
Smith and Wenzel,
1948
r
Eno et al., 1964
Martin et al., 1959
Bollen et al.,
1954
Arredondo
loamy fine
sand
Brandville
sandy loam and
cadorus silt
loam (with and
without cotton-
seed meal)
Leoni fine sand
Ramona sandy
loam
Holtville sandy
loam
Chehalis silty
clay loam
Lake labish peat
12.5 to 100 ppm
250 to 1250 ppm
(simulating
112 to 560 kg/ha
in the field)
1 to 16 months incubation
in greenhouse
5 to 96 days incubation
in greenhouse
16.8 kg/ha, 12 semi- 3 to 6 years, field
annual applications application
22.4 kg/ha, 5 annual
applications
11.2 to
22.4 kg/ha
5 to 6 years, field
application
10 to 20 days, field
application
0 to 22%
stimulation
Stimulation
- cell number
more than
doubled
No measurable
effect
No measurable
effect
Stimulatory at
10 Ibs/acre;
No significant
effect at
20 Ibs/acre
-------
As can be seen in Table 3.3, the toxicity of toxaphene to soil fungi has
been assessed largely from field and greenhouse studies. Field application of
toxaphene of 16.8-22.4 kg/ha has commonly not adversely affected fungi (Eno et
al., 1964; Martin et al., 1959). In fact, in a number of studies, toxaphene
has increased fungal cell numbers (Smith and Wenzel, 1948; Eno and Everett,
1958; Bollen et al., 1954). Smith and Wenzel (1948) noted nearly doubling of
the cell numbers by toxaphene in a greenhouse study at concentrations simu-
lating 112-560 kg/ha toxaphene application in the field. The normal applica-
tion rate of toxaphene as an insecticide is only 1.12-6.7 kg/ha.
A differential effect of toxaphene on three species of fungi was noted by
Bollen et al. (1954). Field application of toxaphene at 11.2 kg/ha to a peat
soil caused an overall increase of 62% in the total number of fungi. A study
of the comparative effect of toxaphene on three genera of fungi (Penicillium,
Aspergillus, and Mucor) revealed that toxaphene caused an increase in the
counts of Penicillium, whereas the numbers of Aspergillus and Mucor decreased.
Similar changes were caused by toxaphene at 20.4 kg/ha; however, the effect
was less pronounced. Conversely, Martin et al. (1959) have noted no signifi-
cant influence of toxaphene on numbers or kind of soil fungi in sandy loam or
sandy clay loam soil. The findings of Bollen et al. (1954) may be suggestive
of the potential of toxaphene for altering microbial number and affecting
microbial community structure in certain soils.
3.4 UNICELLULAR ALGAE
Unicellular algae constitute an important constituent of the food chain
and a broad base for primary productivity in our aquatic environment. Con-
sequently, it is important to evaluate the effect of environmental contamin-
ants on freshwater and marine algae.
3-8
-------
The interaction of toxaphene with freshwater and marine algae has been
the subject of numerous studies. The effect on growth and photosynthetic
processes of the cell have been studied most frequently.
3.4.1 Uptake and Metabolism
Information concerning uptake and metabolism of toxaphene in unicellular
algae is virtually nonexistent. Paris et al. (1975) briefly examined the extent
of sorption of toxaphene by the green alga, Chlorella pyrenoidosa. The culture
of algae was analyzed.at intervals by centrifuging an aliquot and determining
the pesticide concentrations in the supernatant. The algae was noted to be
in equilibrium with the medium within 10 minutes. The sorption of toxaphene
was represented by the empirical equation of Freundlich; the value of the
distribution constant (mg pesticide per mg dry wt of the organism/mg pesticide
per mg water) for toxaphene by algae was 17.0. Toxaphene sorption was also
demonstrated in field samples containing algae (Scenedesmus sp. and Chlorella
sp.) and other microorganisms. The extraction and analysis of the whole algal
cultures revealed no degradation of toxaphene by green algae after 24 hours of
incubation.
In an experiment designed to determine bioaccumulation/biomagnification
potential of toxaphene, Schoettger and Olive (1961) exposed cultures of the
planktonic alga, Scenedesmus incrasstulus, to 0.01 ppm toxaphene for 384 hours.
The accumulated toxaphene was extracted from algae with benzene, the extract
redissolved in isopropyl alcohol. The presence of toxaphene in the extract was
bioassayed using Notropis sp. of fish as the test organism. The results in-
dicated that the algae had not accumulated a sufficient quantity of toxaphene
to kill any of the test fish (Table 3.4). On the other hand, periphytons (composed
3-9
-------
Table 3.4. Accumulation of toxaphene in fish-food organisms exposed to multiple toxaphene
dosages
Daily dosage
of toxaphene
(ppm)
Toxaphene
formulation
used
Exposure
Period
(days)
No of
cells/liter
% Fish surviving
up to 120 hrs.
(Test fish:
Notropis sp.)
Planktonic algae - as fish-food organism
0.01
0.01
100% technical
grade
40% wettable
powder
16
16
1,850 x 10
3,300 x 10
100
100
Periphyton (composed of Cladophora sp., Anacystis sp., Scenedesmus sp., Novicula sp., Diatoma sp.,
and Ciliophora sp.) as fish-food organisms
0.01
0.01
100% technical
grade
40% wettable
powder
13
13
Not specified
Not specified
0
0
a
Source: Modified from Schoettger and Olive, 1961.
-------
of various algae, diatoms, and ciliates) exposed to toxaphene, extracted and
bioassayed in a similar manner, revealed concentrations of toxaphene high
enough to kill fish within 24 hours. It was suggested that periphyton accumu-
lated toxaphene by incorporating it into the mass rather than by adsorption.
3.4.2 Effect on Growth
The salient features of the studies dealing with the effect of toxaphene
on growth are given in Table 3.5. Ukeles (1962) tested the toxicity of toxa-
phene on five species of marine algae. A toxaphene solution prepared in sterile
sea water was added in appropriate concentration (final concn. 0.01-0.15 ppm)
to the sterile basal medium which had been supplemented with the test species
of algae. The algal cultures were incubated under continuous illumination
and growth rates were measured by the determination of transmittance at 530 nm.
The concentrations of toxaphene required to cause complete inhibition of growth
of the five species of marine phytoplankton are shown in Table 3.5. Of the
various species, the Monochrysis sp. was the most sensitive to toxaphene since
complete inhibition of growth was observed at a toxaphene concentration as low
as 10 ppb. Other species were also inhibited by toxaphene but at relatively
higher concentrations.
Unlike the effect of toxaphene on marine algae, the effect on freshwater
algae has been studied by a number of researchers. The earliest study was
that of Palmer and Maloney (1955) who studied the effect of toxaphene at a
concentration of 1.5 ppm on several laboratory strains of unicellular algae
representing blue-greens, greens, and diatoms. The criteria for selecting
the algal species was their ability to produce rapid and uniform growth under
laboratory conditions. The effect of toxaphene was assessed in a qualitative
3-11
-------
Table 3.5. Toxicity of toxaphene to unicellular algae
Concn. of
toxaphene
Reference Test sp. (ppm)
A. Marine algae
Ukeles, 1962 Proloccus sp. 0.15
Chlorella sp. 0.07
Dunaliella sp. 0.15
Phacodactylum 0.04
tricornutum
Monochrysis 0.01
lutheri
B. Freshwater algae
Palmer and 1. Blue-green algae 1.2
Maloney, 1955 Cylindrospermum
licheniforme
Microcystis
aeroginosa
2. Green algae "
Scenedesmus
obliquus
Chlorella
variegata
3. Diatoms "
Gomphonema
parvulum
Nitzschia palea
Period of
treatment
(days) Effect on growth
10 100% inhibition
10 100% inhibition
10 100% inhibition
10 100% inhibition
10 100% inhibition
up to 21 days
Partially toxic
Toxic
it
Partially toxic up
to 7 days, non-
toxic afterwards
Non-toxic
ii
Partially toxic
Partially toxic up
to 7 days, non-
toxic afterwards
-------
Table 3.5 (continued)
Reference
Test sp.
Concn. of
toxaphene
(ppm)
Period of
treatment
(days)
Effect on growth
U)
i
B. Freshwater algae
(continued)
Stadnyk et al., Green algae
1971 Scenedesmus
quadricaudata
O'Kelley and
Deason, 1976
21 strains from
the Harrier River
0.1
1.0
0.001
0.01
0.1
Up to 10
Up to 10
14
14
14
No significant
decrease in cell
number and bio-
mass
19% decrease in
cell number, no
significant effect
on biomass
12% strains inhibited
(growth >50%)
6% no effect
3% stimulated (growth
2% strains inhibited
(growth >50%)
13% no effect
6% stimulated (growth
1% strains inhibited
(growth <50%)
7% inhibited (growth
>50%)
8% no effect
5% stimulated (growth
-------
Table 3.5. (continued)
Reference
Test sp.
Concn. of
toxaphene
(ppm)
Period of
treatment
(days)
Effect on growth
Freshwater algae
(continued)
O'Kelley and
Deason, 1976
(continued)
21 strains from
the Warrier River
(continued)
1.0
14
8% strains inhibited
(growth <50%)
7% inhibited (growth
>50%)
6% no effect
U)
Belonging to Chlorella sp.; Nitzschia sp.; Scenedesmus
.nastrum sp., Honoraphid1urn sp., and Golinkiniopsis sp.
sp. ; Carteria sp. ; IColiella sp.
-------
manner by comparing the amount of visible growth. The authors noted no effect
on Chlorella (a green algae); however, the growth of other algal cultures was
partially to fully inhibited at 1.5 ppm toxaphene.
The fresh water algae as a group are very heterogeneous, both physiologically
and morphologically. Predictions of pesticide interaction with algae in the
field, based upon the data with common laboratory strains, may be subject to
error. Several researchers have realized this difficulty and have used strains
of algae isolated from streams, lakes, and ponds in their toxicity studies.
Stadnyk et al. (1971) used a field-collected strain of green alga, Scenedesmus
quadricaudata, for studying toxaphene toxicity. The test medium was supplemented
with micronutrients to approximate the conditions of a eutrophic lake. The
authors reported no significant effect of toxaphene at concentrations ranging
from 0.1 to 1 ppm on cell number or biomass of the plankton algae. However,
carbon fixation in Scenedesmus was markedly stimulated in the presence of tox-
aphene (Section 3.3.2). In a more elaborate study, O'Kelley and Deason (1976)
tested the influence of toxaphene on the growth of 21 strains of algae which
were isolated from the Warrior River near Tuscaloosa, Alabama. Following treat-
ment, cultures were incubated in a growth chamber without aeration. Growth was
measured turbidimetrically at the end of two weeks. Only a few algal isolates
were inhibited by 0.001 or 0.01 ppm of toxaphene; however, significant numbers
were inhibited when the concentration was increased to 0.1 to 1.0 ppm. At
toxaphene concentrations up to 0.1 ppm, a small proportion of the isolates
(^ 5%) showed stimulated growth (Table 3.5).
Matsumura and coworkers (Nelson and Matsumura, 1975a) have attempted to
isolate and identify the components of toxaphene which are toxic to the blue-green
3-15
-------
alga, Anacystis nidulans, and other aquatic organisms. The components were
separated by a Sephadex LH-20-methanol column and thin-layer chromatography.
Following the addition of the separated fractions, the algal cultures were in-
cubated in light and aerated with air enriched to 2% CC>2 (v/v). The growth
rates were calculated from short term experiments lasting 26 to 30 hours. The
relative toxicity values were obtained by comparing growth rates in standard
toxaphene with separated fractions at a concentration of 1 ppm. Toxaphene at
this concentration caused nearly 1/3 reduction in the growth rates of the algae.
Figure 3.1 shows that no single, isolated fraction was exclusively responsible
for toxaphene toxicity, and that the activity was quite evenly distributed
among several fractions. This led the authors to conclude that several com-
ponents of toxaphene are toxic to algae, and that the toxic action is relatively
non-specific. A toxic fraction which was nearly 1.35 times more toxic than
toxaphene was partially characterized as an octachlorobornane.
Nelson and Matsumura (1975b) used a simplified approach to study toxic
fractions of toxaphene. They chlorinated purified exo-2,10-dichlorobornane,
a major early chlorination product of camphene, to form a "simplified toxaphene"
which was then used for isolation of toxic components. The four chlorination
products purified were only slightly toxic to the blue-green alga, Anacystis
nidulans, at 1 ppm (growth constant for chlorination products = 0.76 to 0.93 per
day, compared to control, 0.96 per day). This was in contrast to toxaphene,
which strongly inhibited algal growth at similar concentrations (growth con-
stant = 0.32 per day), and suggested that exo-2,10-dichlorobornane was not a
precursor for the components of toxaphene which are toxic to algae.
3-16
-------
1.2 -i
£ 0.8
i 0.6-
z
uj
5 0.4-
2
2 0.2-1
JK
0.0
ALGAE
-15
I I
16 17
I
18
1
19
I
20
I
21
I
22
I I I
23 24 25
I
26
I \
27 28
I
29-
FRACTION NUMBER
z i-o-i
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cc
£ 0.6-
UJ
§ 0.4-
0.2-
O
* 0.0
ALGAE
\
G
I
M
TLC FRACTION
Figure 3.1. Relative toxicity of toxaphene components as compared with
toxaphene (as 1) to freshwater blue-green algae, Anacystis nidulans.
Source: Nelson and Matsumura, 1975 a.
k values represent growth rate; k value ratios were calculated at
toxicant concentrations of 1 ppm. Average k value for toxaphene
= 0.327 + 0.009; k value for control = 0.96 + 0.072.
A. Fractions collected from Sephadex LH-20 - methanol column.
B. Fractions separated by thin layer chromatography.
3-17
-------
3.4.3 Effect on Physiological Processes
Information is available concerning the effect of toxaphene on the photo-
synthetic process of the cell (Table 3.6). This physiological process has re-
ceived attention because of the fact that an alteration of the photosynthetic
process at the algal level is ultimately reflected in altered production at
higher trophic levels.
The effect of toxaphene on the photosynthetic process has generally been
assessed by examining the effect on C-14 labelled COj-assimilation in the cell.
In a four hour exposure of toxaphene at 1 ppm, Butler (1963) reported almost
complete inhibition of carbon fixation in estuarine phytoplankton (no details
of species given) (Table 3.6). Stadnyk et al. (1971) measured the effect of
toxaphene on C-14 labelled CO -assimilation of the plankton algae, Scenedesmus
quadricaudata, obtained from field collection. The experimental procedure in-
volved withdrawal of aliquots of toxaphene-treated culture at different inter-
vals; the incubation of cultures with C-14 labelled Na.CO for four hours; and
the determination of C-14 in the cells. Toxaphene was found to cause a marked
stimulation of carbon fixation. As much as 450% increase in carbon fixation
was observed at 1 ppm toxaphene. The degree of stimulation decreased with
continued incubation with toxaphene. The mechanisms of carbon-fixation stimu-
lation by toxaphene is unclear at the present time.
3.5 EFFECT OF TOXAPHENE ON PROTOZOA
A field study concerning the effect of toxaphene on protozoa and other
plankton in a reservoir has been described by Hoffman and Olive (1961). The
reservoir, a single basin lake with a surface area of 24.3 ha and a depth of
15 feet, was treated with toxaphene at a concentration of 0.06 ppm and the
3-18
-------
Table 3.6. Effect of toxaphene on the physiological processes of unicellular algae and
phytoplankton
I
M
MO
Reference
Butler, 1963
Test species
Estuarine phyto-
plankton (no
Concn. of
toxaphene
tested
(ppra)
1
Physiological
Period of process
treatment studied
C02-fixation
Effect
observed
91% inhi
bition
Stadnyk et al. ,
1971
other details
given)
Green algae
Scenedesmus
quadricaudata
0.1
Up to 10 days
C0?-fixation
expressed as
per unit
culture
volume, or
per cell
1.0
Initially
48% in-
crease, no
significant
effect after-
wards
Initially
450% in-
crease,
30% in-
crease after
ten days
-------
number of plankton organisms was determined for a year period, starting in
October. The authors noted that, following the application of toxaphene to
the reservoir, the turbidity decreased to the point that the bottom became
visible. This suggested a toxic action of toxaphene on the aquatic micro-
organisms. Protozoans decreased from high counts taken in October to nearly
zero in December. Furthermore, in the control reservoir the number of protozoan
increased, whereas the number in the treated reservoirs remained at nearly zero.
These results suggest that toxaphene was inhibitory to protozoans and other
microorganisms at the concentration studied.
3-20
-------
REFERENCES
Bolien, W.B., Morrison, H.E. and Crowell, H.H. 1954. Effect of Field
Treatments of Insecticides on Number of Bacteria, Streptomyces, and Molds
in Soil. J. Econ. Entomol. 47:302-306.
Butler, P.A. 1963. Commercial Fisheries Investigations. In: Pesticide -
Wildlife Studies, U.S. Dept. of Interior, Fish and Wildlife Service, Circ.
167, pp. 11-25.
Elfadl, M.A. and Fahmy, M. 1958. Effect of DDT, BHC, and Toxaphene on
Nodulation of Legumes and Soil Microorganisms. Agr. Res. Rev. 36;339-350.
Eno, C.F. and Everett, P.H. 1958. Effect of Soil Applications of 10 Chlori-
nated Hydrocarbon Insecticides on Soil Microorganisms and the Growth of
Stringless Black Valentine Beans. Soil Sci. Soc. Amer. Proc. 22:235-238.
Eno, C.F., Wilson, J.W., Chisholm, R.D. and Koblitsky, L. 1964. The Effect
of Soil Applications of Six Chlorinated Hydrocarbon Insecticides on Vegetable
Crops and Soil Microorganisms. Proc. Florida State Hort. Soc. 77:223-229.
Hoffman, D.A. and Olive, J.R. 1961. The Effects of Rotenone and Toxaphene
Upon Plankton of Two Colorado Reserviors. Limnol. Oceanogr. 6_:219-222.
Martin, J.P., Harding, R.B., Cannell, G.H. and Anderson, L.D. 1959. In-
fluence of Five Annual Applications of Organic Insecticides on Soil Biological
and Physical Properties. Soil Sci. 87:334-338.
Nelson, J.O. and Matsumura, F. 1975 a. Separation and Comparative Toxicity of
Toxaphene Components. J. Agric. Food Chem. 23(5);984-990.
Nelson, J.O. and Matsumura, F. 1975 b. A Simplified Approach to Studies
of Toxic Toxaphene Components. Bull. Environ. Cont. and Toxicol. 11(4);464-470.
O'Kelley, J.C. and Deason, T.R. 1976. Degradation of Pesticides by Algae.
U.S. Environmental Protection Agency, Athens, Georgia. Report EPA-600/3-76-002,
42 pp.
Palmer, C.M. and Maloney, T.M. 1955. Preliminary Screening for Potential
Algicides. Ohio J. Sci. 55:1-8.
Paris, D.F., Lewis, D.L., Barnett, J.T. and Baughman, G.L. 1975. Microbial
Degradation and Accumulation of Pesticides in Aquatic Systems. U.S. Environ-
mental Protection Agency, Corvallis, Oregon. Report EPA-660/3-75-007, 45 pp.
Schoettger, R.A. and Olive, J.R. 1961. Accumulation of Toxaphene by Fish-
Food Organisms. Limnol. Oceanogr. ^:216-219.
Smith, N.R. and Wenzel, M.E. 1948. Soil Microorganisms are Affected by
Some of the New Insecticides. Soil Sci. Soc. Amer. Proc. 12:227-233.
3-21
-------
Stadnyk, L. , Campbell, R.S. and Johnson, B.T. 1971. Pesticide Effect on
Growth and 14C Assimilation in a Freshwater Algae. Bull. Environ. Cont.
and Toxicol. Ji:l-8.
Ukeles, R. 1962. Growth of Pure Cultures of Marine Phytoplankton in the
Presence of Toxicants. Appl. Microbiol. 10:532-537.
3-22
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4.0 BIOLOGICAL ASPECTS IN PLANTS
4.1 SUMMARY
The information concerning the interaction of toxaphene with plants has
been confined to vascular crop and noncrop plants. Aquatic plants are able to
accumulate appreciable amounts of toxaphene from the medium. That toxaphene
can accumulate in plants is also demonstrated by the fact that toxaphene in
large quantities has been detected in the aquatic weeds collected from the
vicinity of a toxaphene discharge point. Considerable toxaphene residues have
also been detected in various crop plants - beet leaves and roots, cabbage,
collard, ladino clover seed greenings, onion greens, and snap beans - when
these crops have received a toxaphene application. The residues were higher
in most crops immediately after harvest, but decreased greatly upon incubation.
A significant reduction in toxaphene residues occurs during commercial packing
house washing and in food processing. In general, detergent washing removed
more toxaphene than washing with water. A survey by FDA shows that human expo-
sure to toxaphene via food is not appreciable.
In spite of the fact that toxaphene is not recommended for use in tobacco,
significant quantities have been detected in tobacco and tobacco products.
Toxaphene caused off-flavor in tobacco when applied to tobacco foliage; how-
ever, soil applications of toxaphene produced no significant effect on flavor.
Toxaphene is not phytotoxic to most crop plants at concentrations recom-
mended for control of insects (15 to 20 kg/ha, annual). The growth and develop-
ment of cauliflower, cabbage, and tomato has, however, been shown to be affected
to some extent at such concentrations of toxaphene. At concentrations 10 to
20 times the normally recommended dose, detrimental effects of toxaphene on
oat plants have been noted. No information concerning the effect of toxaphene
on noncrop plants is available.
4-1
-------
Toxaphene generally does not impart off-flavor to raw and canned vege-
tables. Some adverse effect was noted in cooked rutabagas, onions, and carrots.
Application of toxaphene to the soil appears to cause more instances of changes
in flavor than did applications to the foliage. It is believed that crop
residues result primarily from surface deposition, and only little if any
translocation of toxaphene occurs in plants.
4.2 NONVASCULAR PLANTS - HIGHER FUNGI, MACRO-ALGAE, AND MOSSES
No studies were encountered in the literature which dealt with the inter-
action of toxaphene with nonvascular plants. Among the miscellaneous samples
analyzed for toxaphene by Reimold and Durant (1972 a) was the macro-algae Ulva
(sea lettuce). The samples were collected from a Georgia estuary into which
toxaphene wastes (from a manufacturing plant) were discharged. Significant
concentrations of toxaphene were not detected in the algae in this study (de-
tection limit 0.15 ppm).
4.3 VASCULAR PLANTS
4.3.1 Noncrop Plants
The effect and metabolism of toxaphene in noncrop plants has not been
investigated extensively. In view of the fact that toxaphene has been sug-
gested for use in rehabilitation of fresh water fishing lakes and reservoirs,
a number of the reported studies have dealt with the distribution and analysis
of toxaphene in aquatic fauna and flora following application of toxaphene for
fish eradication. Some information is also available on the levels of toxa-
phene in vegetation in toxaphene-contaminated areas. Virtually no information
on the effect of toxaphene on noncrop plants is available.
4.3.1.1 Metabolism; Uptake, Adsorption, and Residue — Information on the
metabolism and fate of toxaphene in noncrop plants is important in the overall
assessment of the environmental hazards of toxaphene. Noncrop plants may hold
4-2
-------
toxaphene and/or toxaphene metabolites, and recycle them into water by elution
or lysis. Also, the terrestrial and aquatic animals feeding on the vegetation
may ingest toxaphene and concentrate it in their bodies. Reimold (1974), while
studying the levels of toxaphene in organisms collected from the contaminated
estuary, noted that the salt marsh cordgrass, Spartina alterniflora, contained
36.3 ppm toxaphene in leaves, 4.9 ppm in the seed heads, and 1.9 ppm in the
roots. The salt marsh sediment in the same area contained only 32.5 ppm toxa-
phene. Realizing that the high toxaphene content of S. alterniflora represents
a translocation of toxaphene from sediment into the plant tissue, the author
undertook a detailed study of the movement of toxaphene through the salt marsh
sediment into S. alterniflora using Cl-36-labelled toxaphene. The plants col-
lected from a marsh were placed in sea water medium containing toxaphene and,
following various incubation periods, Cl-36 activity and toxaphene concentra-
tion in various parts of the plant were determined. The toxaphene content in
the plant at 26 days represented nearly 20 to 25 percent of the total insecti-
cide initially present in the sea water culture medium (Table 4.1). The
insecticide content decreased on continued incubation. The data further
showed that the plant roots and rhizomes sorbed greater amounts of toxaphene
than leaves and stems. Toxaphene and Cl-36 in the plant after a 100-day
incubation were 1.8 and 12.6 percent, respectively. The presence of Cl-36 in
excess of that due to the presence of toxaphene as detected by the gas chroma-
tographic method suggests that metabolic alteration of the insecticide had
occurred.
Residues of toxaphene in aquatic vegetation following application of
toxaphene for the purpose of eradication of rough fish have been examined by a
number of researchers (Table 4.2). Kallman et al. (1962) reported accumula-
tion of high concentrations of toxaphene in the rooted aquatic weed, Potamogeton.
4-3
-------
Table 4.1. Uptake of Cl-36-labelled toxaphene by salt marsh cordgrass,
Spartina alternifloraa
Days
of
incubation Plant part
26 Leaves and stems
26 Roots and rhizomes
100 Leaves and steins
100 Roots and rhizomes
% Uptake
of i % Uptake of L
available toxaphene" available Cl-36"
0.621 + .
20. 1+5.
0.048 + .
1.82 + 0.
60^ 2.17 + 1.4C
2 22.5 + 1.6
04 0.6 + 0.36
7 12.02 + 3.6
^Modified from Reimold, 1974.
The sea water culture medium initially contained 100 ppm toxaphene +
4. 316 ]iC± Cl-36 equivalent to 83 mg toxaphene.
Mean + standard deviation.
4-4
-------
Table 4.2 Toxaphene residues in aquatic vegetation in lakes treated with toxaphene for eradication
of rough fisha
Estimated
application rate Interval
of toxaphene between treatment
Aquatic vegetation analyzed (ppb) Lake treated and analysis
Potumogeton sp. 10 Clayton Lake, Mew Mexico 48 hours
30 (in two appli- Clayton Lake, New Mexico 96 hours
cations of 10
and 20)
50 (in three Clayton Lake, New Mexico 9 days
appli ca t ions
of 10, 20, and
20)
J>-
1 Aquatic plants (species name 40 Davis Lake, Oregon 4 years
""" not given)
5 years
88 (in two appli- Miller Lake, Oregon 1 year
cations)
2 years
3 years
Residue fppm)
2.8
4.9
14.6
0.39
(0.2-0.6)a
0.21
(0.1-0.9)
4.59
(0.3-15.5)
2.78
(0.39-7.5)
5.77
(1. 87-13. l)a
Reference
Kallman e t al . ,
1962
tv;i 1 J in;in u t a 1 . ,
1962
K;il liaan e t al . ,
1962
Terriere et al . ,
1966
Terriere et al. ,
1966
Range of detected toxaphene residues.
-------
At the application rate of 50 ppb, the concentration of toxaphene in the
aquatic plant was as high as 4.4 ppm after 5 days of incubation, and increased
to 14.6 ppm after 9 days. The uptake and fate of toxaphene beyond 9 days was
not investigated by the authors. In agreement with these results, Terriere et
al. (1966) have also noted accumulation of appreciable residues of toxaphene
in aquatic plants. Plants absorbed up to 15.5 ppm toxaphene when growing in
water containing approximately 2 ppb of the insecticide (residue detected in
water). The analysis of the various parts of the plant revealed that roots
usually contained the highest concentration of toxaphene.
The ambient toxaphene levels in aquatic vegetation growing in toxaphene-
contaminated areas have been determined by Reimold and Durant (1972 b). The
salt marsh cordgrass, Spartino alterniflora, growing in the vicinity of a
toxaphene discharge point from a toxaphene manufacturing plant, accumulated
toxaphene in the leaves to the extent of 36 ppm (Table 4.3). Other parts of
the plant contained relatively low concentrations of toxaphene. Contrary to
these findings, the uptake studies of Reimold (1974) with Cl-36-labelled
toxaphene had revealed that the plant roots and rhizomes sorbed the greatest
amount of toxaphene (Table 4.1). The difference between the two observations
was believed to be due to differences in uptake from sea water medium as
opposed to marsh soil. The results, however, clearly demonstrate that toxa-
phene can be concentrated in Spartina when it occurs in the growth medium.
Reimold and Durant (1974) also monitored the toxaphene content in salt marsh
cordgrass before, during, and after dredging of toxaphene-contaminated sedi-
ment and found the concentration to be highest during dredging rather than
before or after.
4-6
-------
Table 4.3. Ambient toxaphene levels in noncrop plants
Plants analyzed
Sail mursh cordgrass
Salt marsh cordgrass
Salt marsh cordgrass
Sampling location
Salt marsh bordering
Terry Creek,
Brunswick , Georgia
(near toxaphene
discharge point)
Locations other than
Terry Creek within
the Brunswick
study at'ea
Terry Creek, Brunswick
Georgia » before,
during, and after
dredging operation
Source ol Loxapliene
Ln thu vicinity,
it ,iy
Toxaphene manuf acturl np,
plant - Brunswick
Ope rat ions.
Hercules , Inc .
Toxaphene manufacturing
plant - Brunswick
Operations ,
Hercules , Inc,
Toxaphene manufacturing
"plant - Brunswick
operations ,
Hercules , Inc.
Toxiiphent!
concent r;itiun in
the vli-inity
.if thi- s.-impling
location
Marsh sediment
-ontained 32.5 ppm
tuxaphene
Marsh sediment
contained toxaphene
ranging from
0. 81 to 63.7 ppm
<10 ppm In surface
sediment
Toxaphenu leve Ls
i it p Ian tfcj Re f e rtince
16,3 pum 1 leaves; Ketmold and Ourant,
4.9 ppm in -
-------
4.3.1.2 Effects — No information was found in the literature concerning the
effect of toxaphene on noncrop plants.
4.3.2 Crop Plants
Toxaphene, like other pesticides, can contaminate and accumulate in man's
food supplies. Consequently, its interaction with crop plants has received a
considerable amount of emphasis. The toxaphene residues remaining in man's
food and the adverse effect of toxaphene on plants have been extensively
investigated. The information available on these and other aspects of toxaphene
interaction with crop plants is summarized below.
4.3.2.1 Metabolism; Uptake, Adsorption, and Residues — Whereas the uptake and
adsorption of toxaphene in crop plants has not been studied, the residues remain-
ing in various crops after applying the insecticide have received a considerable
amount of attention. The residues have been determined following both soil and
foliar applications.
The conditions of the test and toxaphene residues as found on various crops
are shown in Table 4.4. Considerable toxaphene residues were found on beet
leaves and roots, cabbage, collards, ladino clover seed greenings, onion
greens, and snap benas. Other crops investigated contained only traces or
nondetectable levels of toxaphene. In general, lesser amounts of toxaphene
were found when the insecticide was applied in soil prior to planting. The
residues were higher in most crops immediately after harvest, but decreased
greatly upon incubation.
The removal of toxaphene residues from vegetables by washing has been
studied by Van Middelem (1966). Detergent washing was found to remove consider-
ably more toxaphene residues from the vegetables tested - tomatoes, green
beans, celery, and mustard greens - than the water wash alone (Table 4.5).
4-8
-------
Table 4.4 Residue of toxaphene detected in crop plants
Crop studied
Beets
Table beet (early wonder)
Sugar beet (U.S. 56)
Bell peppers
Cabbage
Collards
Corn
Co t ton seed
Col Con .seed oil
Cotton meal
Appli cation
rate
(kg/ha)
1.7 - 3.3
3.3
3.3
2.8 - 5.6
1.1 - 4.4
33.6 (20% toxaphene
dust)
125 - 250
123 (wettable '
powder formu-
lation)
App 11 i'.'i L i on procedure
and
number of applications
1-3 dust applications
Single preplanting treatment
Single preplanting treatment
1-3 dust applications
1-3 applications of dust or spray
Single dust application at the
time of harvest
Single application of technical
toxaphene
96.9 Ibs in a single preplanting
application, plus 13.9 Ibs in
a single foliar application
Interval
between
treatment
and harvest
7-9 days
9 days
18 days
1 hr-4 wks
1-3 wks
0-13 days
Crop planted
16 yrs after
application
of insecticide;
crop harvested
and analyzed at
maturity
Residue (ppm)
4.3-41 in tops
3.3-16 in roots
Below delectable level
Below detectable level
0.2-2.9
0-5.6
4.9-168.0
Hondetectable
0.1
0.4
0.13
Reference
Wene, 1958
Muns et al. , 1960
Muns et al. , 1960
Wene, 1958
Wene, 1958
Brett and Bowery, 1958
Nash and Harris, 1973
Randolph et al. , 1960
Ladine clover
seed screenings
1.1 - 1.7 DDT plus
2.2 - 3.3 toxaphene Two applications
21.1 in composite
s amp1e
Archer, 1970
-------
Table 4.4 (continued)
Application Application procedure
rate and
Crop studied (kg/ha) number of applications
Lettuce 2.5 - 3.7 1-3 dust applications
Oats 125 - 250 Single application of technical
grade toxaphene
Onion 1.7 - 3.3 1-3 spray applications
.p-
1 Potato (white rose) 3.3 Single preplanting treatment
O
Snap beans 33.6 (20% toxaphene Single dust application at the
dust) time of harvest
Sorghum 140 (wettable 108 kg/ha in a single pre-
powder) planting application, plus
31.6 kg/ha in two foliar
applications
Soybean 125 - 250 Single application of technical
grade toxaphene
Interval
between
treatment
and harvest Residue (ppm) Reference
3 hrs-3 wks 0-1.7 Wene, 1958
Crop planted Monde tec table Nash and Harris, 1973
16 yrs after
application
of the insec-
ticide; crop
harvested and
analyzed at
maturity
1-3 wks 5-71 in tops; Wene, 1958
0.1-4.6 in bulbs
14 days 0.3 Muns et al. , 1960
0-12 days 1.5-8.10a Brett and Bowery, 1958
— -- Nondetectable Randolf et al. , 1960
Crop planted Noiidetuctnlile Nash and Harris, 1973
16 yrs after
application
of the insec-
ticide; crop
harvested and
analyzed at
maturity
-------
Table 4.4 (continued)
Application
race
Crop studied (kg/ha)
Stringbeans 2.8 - 5.6
Tomatoes 33.5 (20% toxaphene
dust)
Wheat 125 _ 250
•P-
1
I-1
Application procedure
and
number of applications
Single dust application
Single dust application at the time
of harvest
Single application of technical
grade toxaphene
Interval
between
t rea tment
and harvest Residue (ppm) Reference
3 hrs-3 wks 0-11.7 Wene , 195«
0-12 days 0.15-4. 0£ Brett and Bowery, 1958
Crop planted Nonde tectable Nash and Harris, 1973
16 yrs after
application
of insecti-
cide; crop
harvested
and analyzed
at maturity
For comparison, FDA tolerances of toxaphene on snap beans, celery, and tomatoes are 7 ppm (Brett and Bowery, 1968; Van Middelem, 1966).
-------
Table 4.5. Comparison of washing with and without surfactants on removal of
toxaphene from vegetables^
Toxaphene, ppm
Washing treatments
None
Water
0.1% Polyether alcohol
1% Neutral soap
Tomatoes
3.69
3.31
3.19
3.27
Green beans
31.07
24.91
13.18
12.25
Celery
60.44
25.81
76.99
16.42
Mustard green
233.64
100.02
23.09
20.77
^Modified from Van Middelem, 1966.
4-12
-------
For example, the toxaphene residue on mustard greens which were harvested 12
hours after treatment was reduced 57 percent by water washing alone and 90
percent following the detergent washing. Significant reductions were also
noted in toxaphene residues in celery following commercial packinghouse wash-
ing (Van Middelem and Wilson, 1960). In food processing, 20 to 60 percent of
the residues are removed by processing, washing, peeling, and heating of
fruits and vegetables (Hartwell et al., 1974).
Estimates of toxaphene exposure from dietary intake can be made from the
U.S. Food and Drug Administration (FDA) market basket survey, the FDA survey
of unprocessed food and feed samples, and the U.S. Department of Agriculture
(USDA) survey of meat and poultry. In the FDA market basket survey, food
samples are prepared for consumption (i.e., cooked, or otherwise processed)
prior to monitoring for pesticide residues (Duggan and McFarland, 1967). The
market basket items are grouped by commodity class (e.g., dairy products,
leafy vegetables, legume vegetables) and are intended to represent a 2-week
diet for a 16- to 19-year-old male (Duggan and Corneliussen, 1972). The
results of these surveys, from their inception to the most recently published
report, are summarized in Table 4.6. From 1964 to 1972, food samples were ob-
tained from five cities: Boston, Massachusetts; Baltimore, Maryland; Los Angeles,
California; Kansas City, Missouri; and Minneapolis, Minnesota. Of the 26
positive samples encountered during this period, 19 were in Los Angeles, four
were in Baltimore, and one was in Boston. Based on the estimates of daily in-
take made by Duggan and Corneliussen (1972), and assuming an average body weight
of 70 kg, the estimated daily dose of dietary toxaphene over the period of
June 1964 to April 1970 was 0.021 ug toxaphene/kg body weight/day. This esti-
mate is based on food samples from a limited number of cities, most of which
4-13
-------
Table 4.6. Toxaphene residues found In food and drug administration market basket survey, 1964 to 1975
r
Monitoring
Period
June
June
June
June
June
June
June
June
Aug.
Aug.
Aug.
1964-April 1965
1965-April 1966
1966-April 1967
1967-April 1968
1968-Aprll 1969
1969-April 1970
1970-April 1971
1971-July 1972
1972-July 1973
1973-July 1974
1974-July 1975
No. of
Composits
216
312
360
360
360
360
360
420
360
360
240
No. of % Commodities contaminated Range of Daily
Composits Occurrence (No. of composite of levels Intake
Positive Each commodity contaminated) (mg/kg)
0 0.0 — — 0
3 1.0 Leafy vegetables (1) and 0.048-0.38 0.002
garden fruits (2)
0 0.0 — — 0
4 1.1 Meat, fish, or poultry (1) 0.064-0.375 0.002
leafy vegetables (1) ,
garden fruits (2)
13 3.6 Garden fruits (6), meat, 0.022-0.33 0.004
fish, or poultry (1) ,
legume vegetables (2) ,
root vegetables (1) ,
leafy vegetables (3)
4 1.1 Leafy vegetables (2), 0.080-0.132 0.001
garden fruits (2)
1 0.3 Root vegetables (1) trace
1 0.2 Leafy vegetables (1) 0.1
0(l)a 0.0 — (0.005)a
3 0.8 Garden fruits (3) trace-0.163
1 0.4 Leafy vegetables (1) 0.118
Reference
Duggan et al., 1966
Duggan et al., 1967
Martin and Duggan,
Corneliussen, 1969
Corneliussen, 1970
Corneliussen, 1972
Manske and Corneliu
1974
Manske and Johnson,
Johnson and Manske,
Manske and Johnson,
1968
ssen,
1975
1975
1976
Johnson and Manske, 1977
Strobane.
-------
are not located in areas of high toxaphene usage. The more recent (1972 to
1975) results of the market basket survey suggest that the current daily
dietary dose may be substantially lower; however, it is equally possible that
the dietary doses for individuals located in the Mississippi Delta (an area of
high toxaphene usage) could be substantially higher. Most toxaphene exposure
is associated with leafy vegetables and garden fruits.
Toxaphene residues in significant quantities have also been demonstrated
in tobacco and tobacco products (Table 4.7). This is in spite of the fact
that toxaphene is not registered for use on tobacco, and its use has been dis-
couraged for many years. Toxaphene is used on cotton, and Domanski and coworkers
(1975) state that the residues found in tobacco may be due, at least in part,
to drift during application of the insecticide to cotton planted in close prox-
imity to tobacco fields. Over 90 percent of the fire-cured tobacco samples
were positive for toxaphene in 1972 (Domanski et al., 1975). This shows a con-
siderable increase from 1970 when only 30 percent of the samples contained
toxaphene (Domanski et al., 1973). Significant amounts of toxaphene were found
in other types of tobacco also. For example, 50 percent of the 1972 burley
samples had toxaphene concentrations greater than 0.5 ppm (Domanski et al., 1975).
A survey for toxaphene residues on commercial tobacco products revealed that
the insecticide was present in all the products analyzed (Table 4.7). Toxaphene
residues in most tobacco products were considerably higher in 1971 than the
concentration found in the 1973 auction market samples. For example, toxaphene
in cigarettes decreased from 3.3 ppm in 1971 to 1.4 ppm in 1973 (Domanski et al.,
4-15
-------
Table 4.7. Toxaphene residues in tobacco and tobacco products
Product studied
Flue cured tobacco
Hurley tobacco
Fire cured tobacco
Dark air cured tobacco
Light air cured tobacco
Cigarettes
f
1— "
O\ Cigars
Light cigars
Chewing tobacco
Smoking tobacco
Snuff
Sampling location
Auction market (1972 crop)
Auction market (1972 crop)
Auction market (1972 crop)
Auction market (1972 crop)
Auction market (1972 crop)
Retail market
Retail market
Retail market
Retail market
Retail market
Retail market
Sample details
Composite of 50 leaves
(5 leaves from each farmer)
Composite of 50 leaves
(5 leaves from each farmer)
Composite of 50 leaves
(5 leaves from each farmer)
Composite of 50 leaves
(5 leaves from each farmer)
Composite of 50 leaves
(5 leaves from each farmer)
Individual samples of various
brands were composited
Individual samples of various
brands were composited
Individual samples of various
brands were composited
Individual samples of- various
brands were composited
Individual samples of various
brands were composited
Individual samples of various
brands were composited
Residue (ppm)
0.51-1.96
0.43-1.24
0.35-4.0
0.36-1.69
0.19
3.3 in 1971
1.4 In 1973
<0.6 in 1971
0.3 in 1973
<0.6 in 1971
0.1 in 1973
1.4 in 1971
0.5 in 1973
1.6 in 1971
1.0 in 1973
1.2 in 1971
0.6 in 1973
Reference
Domanski et al. ,
Gibson et al. ,
Domanski et al. ,
Gibson et al. ,
Domanski et al. ,
Gibson et al. ,
Domanski et al. t
Gibson et al. ,
Domanski et al. ,
Gibson et al. ,
Domanski et al. ,
Domanski et al. ,
Domanski et al. ,
Domanski et al. ,
Domanski et al. ,
Domanski et al. ,
1975;
1974
1975;
1974
1975
1974
1975;
1974
1975;
1974
1974
1974
1974
1974
1974
1974
-------
1974). This suggests that toxaphene use with or near tobacco was greater in
the past and is slowly declining.
4.3.2.2 Translocation — A search of literature has revealed no experimental
data concerning translocation of toxaphene in crop plants. It is believed that
crop residues result primarily from surface deposition, and only little, if any,
translocation of toxaphene occurs in plants (Hartwell et al., 1974). Accordingly,
root crops will be expected to contain only traces of toxaphene.
4.3.2.3 Effects— The effect of toxaphene on emergence, growth, physiological
functions, and yield of various crop plants has been reported. In assessing
the phytotoxic effect of insecticide treatment from field studies, the damage
caused by the insecticide should be compensated for by the protection afforded
by the insecticide. Such compensation, however, is not always easy to determine
and, therefore, the results of such studies should be viewed with caution.
In an attempt to determine if accumulation of toxaphene in the soil might
become detrimental to cotton, Franco et al. (1960) studied the effect of increas-
ing soil concentrations of the insecticide on the growth of cotton plants. The
studies were carried out in Mitscherlich pots in a greenhouse with two types of
soil - one sandy and the other clay. In sandy soil, toxaphene added to the pot
at a dose corresponding to the field dose of 72.3 kg/ha (calculated on area
basis), when applied in emulsion form, showed some toxicity- However, the in-
secticide applied in powdered form equivalent to a field dose of 101.5 kg/ha
did not exhibit any toxic effect. Toxaphene also failed to exert significant
toxic effects in clay soil. Joyce (1954) reported spotting of leaves in cotton
caused by toxaphene, but noted no prolonged effect on the plant and no subsequent
deleterious effect on yield. The growth and development of cotton as affected
4-17
-------
by a toxaphene-DDT mixture (2:1) has been studied by Brown and coworkers (1961,
1962). Both yield and boll production were significantly increased in field
plots upon treatment with toxaphene and DDT. No significant effect of the
insecticide mixture was observed on internode length, number of internodes,
plant height, percent of boll shed, germination, percent lint, boll components
seed index, lint index, boll weight, and seeds per boll. The results suggest
that toxaphene, as well as DDT, was relatively nontoxic to cotton.
Randolph et al. (1960) conducted field experiments to determine the
effect of large amounts of toxaphene applied to soil on the germination,
growth, and yield of several crops. The initial dosage was equivalent to the
amount of toxaphene applied normally during 5 years for the control of cotton
insects. Subsequently, normal foliar treatments were made annually to the
crops grown in the treated plot. Cotton was grown immediately after applica-
tion; sorghum 1 year after application; and alfalfa 2 years following cotton.
No significant difference in the number of plants per acre and yield of the
above crops was noted between the treatments and the control (Table 4.8). At
the dosage of toxaphene equivalent to the amount that would be applied for
control of insects in cotton over a 10 and 20-year period, adverse effects on
some crops were observed. Most severely affected were the oat plants; the
plant stand and yield of oat forage and grain were significantly less than
those from untreated plots. The data must be considered, however, keeping in
mind that the dose of toxaphene in these studies was several fold higher than
normally recommended for insect control.
The effect of toxaphene on the growth and respiration of corn, oats, peas,
and cucumber has been evaluated by Lichtenstein et al. (1962). The plants were
4-18
-------
Table 4.8. Response of various crops to application of large amounts of toxaphene
-P-
Rate and mode of application
A. 121.5 kg/ha preplanting
application plus 14-19 Ibs/acre
annual foliar application
B. 165.3 kg/lia preplanttng
application
C. 309.4 kn/ha preplanting1"
application
Crops grown
during 4-year period
following application
Cotton , immediately after application
Sorghum, first year
Alfalfa, second year
Alfalfa, third year
Cotton, immediately after application
Oats, first year
Cotton, third year
Cotton, immediately after application
Oats , first year
Cotton, third year
Plants/ha
112
88
102
124
101
75
94
95
63
115
Crop response examined
Average height Yield
% of control
102 117, seed cotton
106, forage;
103, grain
—
93, seed cotton
88 , forage ;
101, grain
97, seed cotton
86, forage;
101, grain
^Modified from Randolph eC al.» 1960
Calculated amount that would be applied in 10 years.
Calculated amount that would be applied in 20 years.
-------
grown in quartz sand to keep sorption of the insecticide to a minimum. In general,
toxaphene was toxic to these plants as revealed from stem and root length measure-
ments, respiration of excised root tips, and dry weight per unit length of root
tip (Table 4.9). A slight reduction over control was noted in peas in the ratio
of root and stem length. However, considering that the conditions of treatment
were extreme (for example, the quartz sand does not possess any insecticide-
retention qualities), it is unlikely that toxaphene will be toxic to peas. In
a greenhouse study, Hagley (1965) noted no significant effect of toxaphene at
an application rate of 1.57 kg/ha on the growth and size of seedlings of cauli-
flower and tomato, but in cabbage a reduction in the size of the seedlings was
observed (Table 4.10). At 15.7 kg/ha, the insecticide was found to reduce
growth rate and the size of seedlings, but did not significantly affect root
development. In some plants, toxaphene also caused marginal and interveinal
chlorosis and necrosis of the lower leaves. The greatest phytotoxicity was
observed in tomatoes resulting in the death of 50 and 33 percent of the seed-
lings in the second and third weeks of growth, respectively.
Probst and Everly (1957) observed no noticeable effect of toxaphene when
applied as wettable powder in the soil on emergence, growth, yield, and chemical
composition of soybeans. The chemical variables of soybean'tested included
protein, oil content, and iodine number of the oil. The dosage (44.8 kg/ha)
used in these experiments was much greater than that recommended for the con-
trol of soil insects.
Considerable effort has been expended on the investigation of toxaphene
as a possible cause of off-flavor in food products. Generally, researchers
have attempted to answer three questions: (a) is the difference real; (b) is
4-20
-------
Table 4.9. Effect of 30 ppm toxaphene, added at the time of planting, on growth
and respiration of various crop plants^-
Plant variable
examined
Stem length
fa
Root length
Root length /stem length
Crop
Corn
88
87
1.02
response in %
Peas
114
108
0.67
of control plants
Oats Cucumber
— —
— —
— —
Respiration of excised
root tipsc 99.9
Dry matter content
of 10 mm root tipc 88
83.9
87
103.6
88
116.9
104
^Modified from Lichtenstein et al., 1962.
21 days after planting.
7 days after planting.
4-21
-------
Table 4.10. Effect of toxaphene on the growth and size of vegetable seedlings
Crop response in percent of control plants
Toxaphene dose
(kg active ingredient/ha)
Plant growth
variable studied
Cauliflower Chinese cabbage
Tomato
1.5
15.6
Average weekly increase in size
Average weight of stem and foliage
Average weight of roots
Average weekly increase in size
Average weight of stem and foliage
Average weight of roots
107.0
77.6
136.0
61. 0*
77.6
109.0
71.5
77.0
69.5t
83.6
80.5
87.6
88.0
50.6fc
66.0L
74.5
Modified from Hagley, 1965.
Significantly different from untreated mean.
-------
the difference commercially important, and (c) is the difference good or bad.
Kramer and Ditman (1956) reported no significant difference in the taste of four
processed products - tomato juice, tomatoes, snap beans, and lima beans - which
had been treated with a 2 percent toxaphene aerosol. Potatoes treated in a
similar fashion showed an improvement in flavor. The information on the flavor
change in this study was obtained by evaluating the taste scores of the judges
by both the triangular (an example of an attributes method) and the variable
method. Birdsall et al. (1957) studied the effect of toxaphene on flavors of
vegetables grown in the presence of the insecticide applied by soil treatment
and by foliar treatment. The rate of application of the insecticide (6.7 kg/ha
on vegetable foliage, 26.9 kg/ha in soil) was four times the normal rate of
application. No significant difference in taste was noted in treated and
untreated raw and canned vegetables tested - beets, sauerkraut, carrots,
onions, potatoes, squash, pumpkin, radishes, and cucumbers. Undesirable changes
in flavor, however^ occurred in cooked rutabagas, onions, and carrots. Appli-
cation of toxaphene to the soil appears to cause more instances of changes in
flavor than did applications to the foliage.
While many insecticides have been reported to cause an increase in the
disaccharide content of rutabagas and red beets by stimulating sucrose synthesis,
toxaphene was found to have no significant effect (Monnberg, 1959).
The ability of toxaphene to impart off-flavor to tobacco has been inves-
tigated by Townes et al. (1958). The contamination of tobacco in several ways
was examined, including: (a) build-up of soil residues] (b) uptake of insecti-
cidal residues in tobacco grown in subsequent years; and (c) direct foliar
application of tobacco. Following curing, the tobacco was given to cigarette
4-23
-------
companies which made it into unblended cigarettes and then submitted the ciga-
rettes to taste—testing panels. No off-flavor in tobacco grown in fields
receiving soil applications of toxaphene or in fields previously planted with
cotton treated with toxaphene was noted (Table 4.11). Toxaphene, however,
caused off-flavor when 'applied to tobacco foliage.
4-24
-------
Table 4.11. Effect of application of toxaphene to tobacco on cigarette flavon
a.
Toxaphene formulation
used
20% Toxaphene dust
75% E.C.b
10% Dust
42% Strobane E.G.
20% Toxaphene dust
Mode of
application
Foliar
Foliar
Foliar
Foliar
Single application
Application rate
(kg/ha)
5.4
' 2.1
1.7-2.0
2.0
112.0
Report
of panel
Objectionable
Objectionable
Objectionable
Objectionable
Satisfactory
20% Toxaphene
75% E.G.
to soil
Eight applications
to cotton foliage
in preceeding year;
tobacco grown in
the same soil in
succeeding year
Foliar application
within 10 days of
harvest
78.4-112.0
Satisfactory
Objectionable
^Modified from Townes et al. , 1958.
E.G., emulsified concentrate.
4-25
-------
REFERENCES
Archer, T.E. 1970. Toxaphene and DDT Residues in Ladino Clover Seed Screenings.
Pest. Monitor. J. _4(2):27-30.
Birdsall, J.J., Weckel, K.G., and Chapman, R.K. 1957- Effect of Chlorinated
Hydrocarbon Insecticides on Flavors of Vegetables. J. Agr. Food Chem. _5_(7) :
523-526.
Brett, C.H. and Bowery, T.G. 1958. Insecticide Residues on Vegetables. J. Econ.
Entomol. 51:818-821.
Brown, L.C., Lincoln, C., Frans, R.E., and Waddle, B.A. 1961. Some Effect of
Toxaphene-DDT and Calcium Arsenate on Growth and Development of Cotton. J. Econ.
Entomol. 54-(2) :309-311.
Brown, L.C., Cathey, G.W., and Lincoln, C. 1962. Growth and Development of
Cotton as Affected by Toxaphene-DDT, Methyl Parathion, and Calcium Arsenate.
J. Econ. Entomol. 55:298-301.
Corneliussen, P.E. 1969. Pesticide Residues in Total Diet Samples (IV).
Pestic. Monit. J. _2(4):140-152.
Corneliussen, P.E. 1970. Pesticide Residues in Total Diet Samples (V).
Pestic. Monit. J. _4(3) =89-105.
Corneliussen, P.A. 1972. Pesticide Residues in Total Diet Samples (VI).
Pestic. Monit. J. 1(4):313-330.
Domanski, J.J., Laws, J.M., Haire, P.L., and Sheets, T.J. 1973. Insecticide
Residues on 1971 U.S. Tobacco Products. Tob. Sci. 17:80-81.
f
Domanski, J.J., Haire, P.L., and Sheets, T.J. 1974. Insecticide Residue on
1973 U.S. Tobacco Products. Tob. Sci. 18;115-116.
yDomanski, J.J., Haire, P.L., and Sheets, T.J. 1975. Insecticide Residues on
V1972 U.S. Auction-Market Tobacco. Beitr. Tabakforschung 8_, Heft 1, 39-43.
Duggan, R.E. and McFarland, F.J. 1967- Assessments Include Raw Food and Feed
Commodities, Market Basket Items Prepared for Consumption, Meat Samples Taken
at Slaughter. Pestic. Monit. J. J.(l):l-5.
i "'""
Duggan, R.E. and Corneliussen, P.E. 1972. Dietary Intake of Pesticide Chemicals
in the United States (III), June 1968 - April 1970. Pestic. Monit. J. _5(4):331-3A1,
Duggan, R.E., Barry, H.C. and Johnson, L.Y. 1966. Pesticide Residues in Total
Diet Samples. Science 151(3706);101-104.
Duggan, R.E., Barry, H.C. and Johnson, L.Y. 1967. Pesticide Residues in Total
Diet Samples (II). Pestic. Monit. J. _1(2):2-12.
4-26
-------
Franco, C.M., Fraga, C.C., Jr., and Neves, O.S. 1960. Effect of Addition of
Insecticides to Soil on the Development of Cotton Plant. Bragantia 19(2);13-25.
Gibson, J.R., Jones, G.A., Dorough, H.W., Lusk, C.I., and Thurston, R. 1974.
Chlorinated Insecticide Residues in Kentucky Burley Tobacco: Crop Year 1963-72.
Pestic. Monit. J. 7_(4) :205-213.
/
Hagley, E.A.C. 1965. Effect of Insecticides on the Growth of Vegetable Seedlings.
J. Econ. Entom. 58(4) -.777-778.
Hartwell, W.V., Datta, P.R., Markley, M., Wolff, J., and Billings, S.C. 1974.
Aspects of Pesticidal Use of Toxaphene and Terpene Polychlorinates on Man and
the Environment. Office of Pesticide Programs, EPA.
Johnson, R.D. and Manske, D.D. 1975. Pesticide Residues in Total Diet
Samples (IX). Pestic. Monit. J. 9_:157.
/Johnson, R.D. and Manske, D.D. 1977. Pesticide and Other Chemical Residues
'in Total Diet Samples (XI). Pestic. Monit. J. 11:116.
Joyce, R.J. 1954. Entomological Section. Annual Report of the Research Division,
Ministry of Agriculture. Sudan Gov't. 106-160.
/Kallman, B.J., Cope, O.B., and Navarre, R.J. 1962. Distribution and Detoxification
* of Toxaphene in Clayton Lake, New Mexico. Trans. Amer. Fisheries Soc. 91(14):66-69.
Kramer, A. and Ditman, L.P. 1956. A Simplified Variables Taste Panel Method
for Detecting Flavor Changes in Vegetables Treated with Pesticides. Food Tech.
10:155-159.
>Lichtenstein, E.P., Millington, W.F., and Cowley, G.T. 1962. Effect of Various
''Insecticides on Growth and Respiration of Plants. J. Agr. Food Chem. 10(3):251-256.
Manske, D.D. and Corneliussen, P.E. 1974. Pesticide Residues in Total Diet
Samples (VII). Pestic. Monit. J. 8^(2) : 110-124.
./lanske, D.D. and Johnson, R.D. 1975. Pesticide Residues in Total Diet Samples
' (VIII). Pestic. Monit. J. jK2):94-105.
Manske, D.D. and Johnson, R.D. 1976. Pesticide and Metallic Residues in
Total Diet Samples (X). Pestic. Monit. J. 10:134.
Martin, R.J. and Duggan, R.E. 1968. Pesticide Residues in Total Diet Samples (III).
Pestic. Monit. J. J1(4):ll-20.
Monnberg, B. 1959. Effect of Certain Chlorinated Hydrocarbons on the Sugar
Components in Plants. Finska Kemists. Medd. 68(2):66-76.
w'mins, R.P., Stone, M.W., and Foley, F. 1960. Residues in Vegetable Crops
* Following Soil Applications of Insecticides. J. Econ. Entomol. 53(5):832-834.
4-27
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^
'
Nash, R.G. and Harris, W.G. 1973. Chlorinated Hydrocarbon Insecticide Residues
in Crops and Soil. J. Environ. Quality _2(2) :269-273.
Prgfoat, A.H. and Everly, R.T. 1957. Effect of Soil Insecticides on Emergence,
Growth, Yield, and Chemical Composition of Soybeans. Agro. J. 49:385-387.
Randolph, N.M., Chisholm, R.D., Koblitsky, L., and Gaines, J.C. 1960. Insecticide
Residues in Certain Texas Soils. Texas Agri. Expt. Sta., Misc. Publication, MP 447.
Reimold, R.J. and Durant, C.J. 1972 a. Survey of Toxaphene Levels in Georgia
Estuaries. Georgia Marine Science Center, University of Georgia, Technical
Report No. 72-2.
Reimold, R.J. and Durant, C.J. 1972 b. Monitoring Toxaphene Contamination in a
Georgia Estuary. Georgia Marine Science Center, University of Georgia, Technical
Report No. 72-8.
Reimold, R.J. 1974. Toxaphene Interactions in Enteractions in Estuarine Ecosystems.
Georgia Marine Science Center, University of Georgia, Technical Report No. 74-6.
Reimold, R.J. and Durant, C.J. 1974. Toxaphene Content of Estuarine Fauna and
Flora Before, During, and After Dredging Toxaphene-Contaminated Sediments.
Pestic. Monit. J. _8(l):44-49.
Terriere, L.C., Kiigemagi, U., Gerlach, A.R., and Borovicka, R.L. 1966. The
Persistence of Toxaphene in Lake Water and Its Uptake by Aquatic Plants and
Animals. J. Agr. Food Chem. 14(1);66-69.
f
Townes, H.K., Smith, C.F., Rabb, R.L., Bowery, T.G., and Gunthrie, F.E. 1958.
Insecticide Residues as a Source of Off-Flavor in Tobacco. Tob. Sci. j^:90-94.
Middelem, C.H. 1966. Fate and Persistence of Organic Pesticides in the
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Van Middelem, C.H. and Wilson, J.W. 1960. Unpublished data. Cited in
Van Middelem, C.H., 1966.
Jwene, G.P. 1958. Toxaphene Residues on Certain Vegetables at Various Time
Intervals After Application. J. Rio Grande Valley Hort. Soc. 12:106-110.
4-28
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5.0 BIOLOGICAL ASPECTS IN WILD AND DOMESTIC ANIMALS
5.1 CHAPTER SUMMARY
Toxaphene has a broad spectrum of acute toxicity. It kills aquatic
protozoans at levels as low as 7 ppt and can kill calves at oral doses of
50 rag/kg. Organisms at all levels have demonstrated the ability to bioconcen-
trate the pesticide, thus potentially passing on to the next higher trophic
level a toxic or metabolically altered dose of toxicant.
5.1.1 Fish and Amphibians
The effects of toxaphene on fish and amphibians are important due to the
position that fish occupy as the top consumers in most aquatic environments,
and because of possible entry into other food webs including higher vertebrates
and man.
Fish are able to absorb and bioconcentrate toxaphene in significant
quantities. A large portion of this is complexed with the lipid stores. In
young fish, large quantities of toxaphene are associated with the yolk sac.
As the fish matures and loses its yolk sac, toxaphene levels decrease. Later,
as the fish grows older and develops its own lipid stores, toxaphene levels
again increase. Some species can concentrate up to 76,000 times the amount
detectable in the surrounding water (Mayer et al., 1975). There is no evidence
of toxaphene biotransformation in fish. Toxaphene residues remain in fish for
many months, or up to several years, following toxaphene application to lakes.
Prey fish appear to accumulate more than predator fish. Under identical
conditions, prey fish - bluegills and suckers - accumulated 9.4 and 10.6 ppm,
respectively, whereas predator fish - bass, northern pike, and walleyes - had
body residues of 2.2, 3.3, and 1.2 ppm, respectively (Hughes and Lee, 1973).
5-1
-------
Toxaphene causes increased calcium-to-collagen ratios in fish, resulting
in increased fragility of the bones. Similarly, decreases in bone collagen
amino acids such as alanine, valine, leucine, isoleucine, lysine, phenylalanine
and hydroxyproline are prevalent. Decreased vitamin C content and a strong
inhibition of ATPase activity occur in toxaphene-exposed fish (Mayer and
Mehrle, 1975 a, b, 1976).
Because of its use as a pesticide, acute toxicity data are abundant
(Section 5.2.2.2.1). Toxaphene is not selectively toxic to fish species
yielding LC50 values.ranging from 2 to 32 ppb. Compared to other pesticides,
however, toxaphene is an extremely potent fish toxicant.
Subacute or chronic exposure to toxaphene results in growth depression,
degenerative changes in the liver with a proliferation of rough endoplasmic
reticulum, mitochondrial swelling and condensation of nuclear chromatin. Also
noted are kidney necrosis, necrotic changes in the digestive tract and damage
to the central nervous system.
Finally, fish species exposed to toxaphene have been reported to develop
a resistance to the pesticide. Toxaphene-tolerant mosquito fish tested in
Ferguson's laboratory (1964, 1965 a, 1969) had 6 to 48 times more resistance
than fish without prior exposure to toxaphene. Later studies revealed that
many species of fish can develop increased tolerances to toxaphene, and that
certain populations had a genetic resistance to toxaphene.
5.1.2 Aquatic Invertebrates
Aquatic invertebrates from unicellular plankton species to oligocheates
have been shown to bioaccumulate toxaphene. Indeed, toxaphene residues in
many invertebrate plankton dwellers often exceed the original toxaphene appli-
cation level for periods up to 9 months post-application (Johnson, 1966).
5-2
-------
Toxaphene concentrations in excess of one ppm block egg development in
clams, while 0.25 ppm toxaphene will kill 67% of the larvae. Most inverte-
brates develop hyperactivity, loss of equilibrium and coordination, paralysis,
and death in response to toxic levels of toxaphene.
Sublethal doses of toxaphene decrease activity and slow down shell de-
composition in those aquatic invertebrates which possess them.
An abundance of acute toxicity data exists. Wide ranges in lethal doses
have been observed from the estuarine copepod, Acartia tonsa, with an LC50
(96 hr) of 7.20 ppt (Khattat and Farley, 1976) to the naiads of Pteronarcys
California with an LC50 (24 hr) of 18.0 ppm.
Pathologically, acute exposure to toxaphene causes vacuolated granular
cells in the hepatopancreas often times coupled with a contraction of the in-
testine. There is occasionally noted a sloughing of the stomach epithelium,
irregular spaces in the nervous system, and the deposition of an undefined
orange-brown pigment in the body spaces (Courtenay and Roberts, 1973).
Chronic exposure to toxaphene results in decreased weight, tissue changes
in the kidney, gills, and digestive tubules, and the invasion of the body
cavity by a mycelial fungus (Lowe et al., 1971).
Many species have developed a resistance to toxaphene. In areas of high
toxaphene use, freshwater shrimp have a toxaphene tolerance 3.5 times greater
than non-exposed shrimp (Naqvi and Ferguson, 1970). As with fish, the ability
to bioaccumulate large quantities of toxaphene has implications that extend to
higher trophic levels.
5.1.3 Birds and Terrestrial Wildlife
No information is available on absorption, transport and distribution,
biotransformation and elimination in birds and terrestrial wildlife.
5-3
-------
Residues in birds are well documented. Residue levels in birds appear
to be indicative of duration of exposure, exposure level, and amount of adi-
pose tissue.
At the biochemical level, toxaphene has been shown to increase thyroid
weight along with a possible stimulation of iodine incorporation.
Acute toxicity studies have yielded a variety of LCSO's in a wide range
of species. These values range from 50 to 316 mg/kg body weight depending on
age, sex, route of administration, etc., and are summarized in Table 5.39.
Chronic toxicity data are limited. Pathological changes in pelicans fed
toxaphene-treated sardines included a decrease in the subcutaneous and mesintery
fat deposits (Keith, 1965). Ring-necked pheasants survived three months of
300 ppm toxaphene treatment with only a transitory weight loss and an increased
vacuolation of liver tissue (Genelly and Rudd, 1956 a). However, quail exposed
to 0.1 percent toxaphene experienced 100 percent mortality in 13 days (Linduska
and Springer, 1951).
Toxaphene, at high dietary doses, has also been shown to decrease egg
laying, egg hatchability, and post-hatching survival. These factors, coupled
with the bioconcentration factor many birds experience in their diet, could
have profound effects on the survival of susceptible bird species.
5.1.4 Domestic Animals
The metabolic data which are presently available deal only with toxaphene
tissue residues. Toxaphene use to control ectoparasites has led to the appear-
ance of residues in the meat and milk of these animals.
Young calves are susceptible to acute toxaphene poisoning when applied as
a spray or dip. Death has resulted from application levels as low as one percent.
5-4
-------
Chronic exposure data are rather limited. Animals fed toxaphene-treated
hay experienced transitory central nervous system effects and no other overt
signs of toxicity. Toxaphene has also been shown to depress rumen function.
The potential for contamination of meat and milk with toxaphene should
be adequately monitored so as to protect man from excessive exposure.
5.1.5 Terrestrial Insects and Other Terrestrial Invertebrates
Metabolically, toxaphene has been shown to penetrate the hemolymph and to
bind with the nervous system tissue. Physiologically, toxaphene appears to
alter central nervous system activity and neuron ion permeability. In the
house fly, toxaphene alters the metabolism of formate by increasing uric acid
and decreasing proline levels.
Toxaphene has been widely used for the protection of cotton crops from
lepidoptern pests for over 20 years. Acute toxicity data for these pests and
other insects are presented in Table 5.59.
Resistance to toxaphene has been demonstrated in several orders of
insects. Resistant insect strains can tolerate doses of toxaphene ten times
greater than those causing death in susceptible strains. In attempts to
explain this developed or genetic resistance, Reiser et al. (1953) found that
in boll weevils the survivors had 10 percent more fat per unit body weight.
Moore et al. (1967) correlated survival with higher levels of palmitic and
oleic acids. Moore and Taft (1972) found higher levels of triglycerides in
the survivors. The development of completely resistant strains of pest organ-
isms is not a unique problem associated with toxaphene, but one which should
be continuously monitored.
5-5
-------
5.2 FISH AND AMPHIBIANS
Because of the general interest in fish as an environmental resource and
the use of toxaphene as a piscicide, extensive information is available on the
effects of toxaphene on fish. This information is presented in the appropriate
sections below. Amphibians have received much less attention, and all of the
information on these animals is presented in Section 5.2.2.2.8.
5.2.1 Metabolism
5.2.1.1 Absorption/Bioconcentration — Relatively few controlled studies have
been conducted on the absorption of toxaphene by fish. While the toxaphene
residue studies described in Section 5.2.1.5 indicate that fish are able to
absorb toxaphene directly from the water and/or from contaminated fish food
organisms, such studies do not include sufficient details on exposure conditions
and the effects of toxaphene metabolism to justify quantitative statements on
toxaphene absorption and bioconcentration. Nevertheless, given the informa-
tion described below, fish seem able to rapidly bioaccumulate toxaphene from
water and to bioconcentrate this compound by a factor of several thousand.
The short term uptake kinetics of Cl-36 labelled toxaphene in mosquito
fish have been examined by Schaper and Crowder (1976). In this study, fish were
exposed to 2 ppm toxaphene in water for eight hours, and toxaphene uptake in
whole fish was determined at one-hour intervals. As indicated in Figure 5.1,
absorption was linear under these conditions with detectable concentrations
of toxaphene appearing in whole fish after one hour. After eight hours, toxa-
phene content in whole fish averaged 0.586 micrograms/gram (ppm), thus indica-
ting that bioconcentration did not occur during this relatively brief exposure
period.
5-6
-------
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5678
Exposure Time (hours)
Figure 5.1. Uptake of Cl-36 labelled toxaphene and
toxaphene related residues in mosquito fish exposed to 2 ppm Cl-36 toxaphene
Source: Schaper and Crowder, 1976.
5-7
-------
During longer exposure periods to markedly lower levels of toxaphene,
bioconcentration of toxaphene has been demonstrated in both bluegills and brook
trout. In static 96-hour exposures to toxaphene concentrations of 17.3 ppb and
70.3 ppb, bluegills accumulated mean whole body concentrations of 9.4 ppm and
7.7 ppm, respectively (Hughes, 1970). However, the differences between these
means were not statistically significant at p less than 0.05, suggesting that
at both test concentrations the mechanisms of toxaphene absorption were satura-
ted. At toxaphene concentrations of 0.5 ppb and below, Mayer and coworkers (1975)
have shown that in brook trout both the amount of toxaphene absorbed and the
extent of bioconcentration are generally directly proportional to the concentra-
tion of toxaphene in the water. However, the time-course of toxaphene absorp-
tion and bioconcentration is somewhat complex. As indicated in Table 5.1,
brook trout fry continuously exposed to toxaphene at concentrations of 0.041 ppb
to 0.500 ppb concentrated toxaphene from the water by a factor of up to 76,000.
Maximum whole-body residue and maximum bioconcentration at all exposure levels
occurred after 15 days, declined between days 15 and 60, then increased up to
day 90. The extraordinarily high levels of toxaphene at day 15 were partially
attributed to the presence of the yolk sac which contains large quantities of
lipids, and thus might be expected to store appreciable amounts of toxaphene.
The decrease in tissue residue and bioconcentration by day 60 could correspond
to the disappearance of the yolk sac and a relatively low level of body lipids.
The increased body levels of toxaphene by day 90 are most likely attributable
to the increasing lipid content of maturing fry (Mayer et al., 1975).
Yearling brook trout seem to bioconcentrate toxaphene less extensively
than do trout fry (Table 5.2). During 161 day exposure periods, these yearlings
5-f
-------
Table 5.1. Whole-body residues of toxaphene in brook trout fry contin-
uously exposed to toxaphene
Exposure level
0.
0.
0.
0.
0.
(ppb)
b
041
(.039 ± .003)
075
(.068 ± .004)
125
(.139 ± .010)
270
(.288 ± .022)
500
(.502 ± .040)
Statistic
d
Residue
Number
BCF
Residue
Number
BCF
Residue
Number
BCF
Residue
Number
BCF
Residue
Number
BCF
Days of exposure
7
0.2
2
4,900
1.0
2
13,300
2.2
2
17,600
4.5
2
16,700
9.2
2
18,400
15
2.6
2
63,400
3.7
2
49,300
8.3
2
66,400
18
2
66,700
38
2
76,000
60
0.4
2
9,300
0.9
2
12,000
1.8
2
14,400
J
-
-
_
-
90
0.6
2
14,600
1.4
2
18,700
2.6
2
20,800
_
-
-
_
-
"
^Source: Modified from Mayer et al., 1975.
Nominal concentration of toxaphene in water (micrograms/liter).
Steasured concentration of toxaphene in water ± the standard error
.(micrograms/liter). •> f f, \
\hole-body toxaphene residue from composite sample of several try U gJ
(micrograms/gram).
^ioconcentration factor: toxaphene residue in whole-body samples - toxaphene
/concentration in water.
"All fish were dead.
5-9
-------
Table 5.2. Whole-body residues of toxaphene in yearling brook trout continuously exposed to
toxaphene
Ul
I
Exposure level
0.
0.
0.
0.
0.
(ppb)
fa
041
(.039 ± .003)
075
(.068 ± .004)
125
(.139 ± .010)
270
(.288 ± 22)
500
(.502 ± .040)
Statistic Days of exposure
d
Residue
Number
BCF
Residue
Number
BCF
Residue
Number
BCF
Residue
Number
BCF
Residue
Number
BCF
3 7 10 18 24
,
(ppm) N.D." N.D. < 0.1
444
<2,400 -
< 0.1 < 0.1
4 4
< 1,300 < 1,300
N.D. < 0.1 0.3 ± 0.11
444
<800 2,400
< 0.1 0.3 ± 0.06
4 4
< 370 1,100
< 0.1 0.5 ± 0.04 2.0 ± 0.12
444
< 200 1,000 4,000
60
<: 0.1
4
< 2,400
0.1 ± 0.02
6
1,300
0.5 ± 0.03
4
4,000
1.8 ± 0.12
6
6,700
4.5 ± 0.46
4
9,000
140
0.3 + 0.005
4
7,300
0.8 + 0.08
6
10,700
0.6 ± 0.06
4
4,800
3.5 ± 0.30
5
13,000
7.7 ± 0.10
4
15,400
0
4
0
5
0
3
2
8
8
16
161
.2 +
4
,900
.4 ±
7
,300
.4 ±
4
,200
.4 ±
8
,900
.0 ±
3
,000
0.03
0.04
0.06
0.19
0.55
/Source: Modified from Mayer et al., 1975.
Nominal concentration of toxaphene in water (micrograms/liter) .
.Measured concentration of toxaphene in water ± the standard error (micrograms/liter),
Whole-body toxaphene residue ± the standard error in micrograms/gram.
,Bioconcentration factor: body residue T toxaphene concentration in water.
None detected, detection limit O.Q5 micrograms/gram.
-------
concentrated toxaphene from the water by factors of from 3,200 to 16,000. As
with the trout fry, the extent of toxaphene bioconcentration at a given time
was generally proportional to exposure concentrations, although fish in the
0.125 ppb exposure group concentrated less toxaphene than would be expected
after 60 days. At all exposure levels below 0.500 ppb, maximum toxaphene
bioconcentration occurred on or after day 140 of exposure and declined appre-
ciably by day 161. The reasons for this decline were not examined.
The transfer of toxaphene to fish from fish food organisms has received
little study. As described in Section 5.3.1, Schoettger and Olive (1961) found
that fish died when fed on Daphnia which had been exposed previously to sublethal
levels of toxaphene. However, these investigators did not determine if lethal-
ity was attributable directly to the ingestion of toxaphene in contaminated
Daphnia or to leaching of toxaphene from the Daphnia with subsequent reabsorp-
tion by the fish. Recently, Sanborn and coworkers (1976) have followed the
fate of C-14 labelled toxaphene in a terrestrial-aquatic model ecosystem. At
a concentration of approximately 44.4 ppb toxaphene in water, mosquito fish
concentrated toxaphene by a factor of 4,247 over a three-day exposure period.
5.2.1.2 Transport and Distribution — As with the pattern seen in mammals
(Section 6.1.2), the limited available information on fish suggests that fat
tissue is a primary site of toxaphene storage. In bluegills exposed to toxa-
phene in toxaphene-treated lakes for periods of up to 350 days, a higher cor-
relation was found between toxaphene concentration and fat content in whole
fish, fish fillets, and fish liver than between toxaphene content and whole
animal or organ weight. Further, the edible portion of the fish (fillets)
contained only one-sixth the concentration of toxaphene found in whole fish
5-11
-------
and only about one-half the concentration of toxaphene found in liver (Hughes,
1970; Hughes and Lee, 1973). Similar results have been found in 161 day expo-
sures of adult brook trout to concentrations of toxaphene ranging from 0.041 ppb
to 0.500 ppb (Mayer et al., 1975). Toxaphene residues found in trout fillet
were about one-third to one-fourth of those found in the remaining tissue (Table 5.3),
5.2.1.3 Biotransformation — The ability of fish to metabolize toxaphene cannot
be determined from the available data. In 161 day exposures of brook trout to
2 ppm toxaphene, Maye.r and coworkers (1975) noted that GLC peak patterns of
toxaphene tissue residues differed from those of toxaphene standards. For the
most part, these differences indicated preferential tissue storage of the more
highly chlorinated lipophilic toxaphene isomers. Similar alterations in chro-
matographic patterns have been noted in fish taken from toxaphene-treated lakes
(Hughes, 1970; Hughes and Lee, 1973; Terriere et al. , 1966). While these re-
sults would certainly be expected if fish significantly metabolized various
toxaphene components, such alterations in peak patterns also could be explained
on the basis of preferential absorption or elimination without actual biotrans-
formation. Further, because toxaphene is known to be altered in aquatic systems
(Section 7.3.2), altered peak patterns in fish taken from toxaphene-treated
lakes do not necessarily indicate that toxaphene is metabolized by fish.
Schaper and Crowder (1976) have found no evidence that mosquito fish are
able to metabolize toxaphene. After eight hours exposure of these fish to 2 ppm
Cl-36 labelled toxaphene, virtually all of the recoverable label was found in
the nonpolar fraction (hexane) and none in the polar fraction (water). While
these results do not rule out the possibility of minor metabolic conversions,
they do suggest that substantial dechlorination or conversion of toxaphene to
water soluble metabolites did not occur.
5-12
-------
Table 5.3. Toxaphene residues found in adult brook trout fillet and
remaining tissue after 161 days of exposure
Exposure,
ppb Statistic Fillet Offal
b c.
0.041 Meant S.E. <0.10 0.42± 0.05
Number 4 4
0.075 Mean ± S.E. 0.17 ± 0.04 0.62 ± 0.07
Number 4 4
0.125 Mean ± S.E. 0.21 ± 0.3 0.66 ± 0.11
Number 4 4
0.270 Mean ± S.E. 0.87 ± 0.13 3.0 ± 0.2
Number 4 4
0.500 Mean ± S.E. 4.0 ± 0.72 11 ± 1.2
Number 3 3
^Source: Mayer et al., 1975.
Nominal concentration.
Standard error.
5-13
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5.2.1.4 Elimination — The elimination of toxaphene by fish seems to depend,
at least in part, on the total body burden of toxaphene. In a follow-up to the
absorption studies described in Section 5.2.1.1, Mayer and coworkers (1975)
placed adult brook trout, which had been previously exposed to known levels of
toxaphene, into toxaphene-free water and monitored the decrease in total body
residue over a 56 day period (Table 5.4). Only fish from the two higher expo-
sure concentrations, 0.27 ppb and 0.5 ppb, eliminated detectable quantities of
toxaphene, 32 percent and 51 percent, respectively. As indicated in the previous
section, the lower chlorinated toxaphene isomers appeared to be eliminated more
readily than the more highly chlorinated isomers based on shifts in chromato-
graphic patterns of toxaphene residues. Trout in the 0.075 ppb and 0.125 ppb
exposure groups released low, but consistent, levels of toxaphene throughout
the 56 day depuration period.
The failure to detect toxaphene in fish from the 0.041 ppb exposure group
after 28 and 56 days may be attributable to the analytical technique which was
not quantitatively valid below residue levels of 0.1 ppm and had a minimum
detection limit of 0.05 ppm (Mayer et al., 1975). Schaper and Crowder (1976)
were also unable to detect toxaphene elimination in mosquito fish during a
six-hour period after exposure to 2 ppm toxaphene for eight hours. The total
body residue in these fish, 0.6 ppm, approximated that of the 0.125 ppb exposure
group in the study by Mayer and coworkers (1975).
5.2.1.5 Residues — Studies on toxaphene residues in fish fall into two basic
groups. The first involves periodic monitoring in lakes which had been treated
with toxaphene as part of fish eradication programs. Such studies are useful
in that they often provide some indication of the duration and extent of toxaphene
5-14
-------
Table 5.4. Toxaphene residues in adult brook trout after transfer to
uncontaminated watera
(ppm)
fa
Exposure
ng/1
0.041C
0.075
0.125
0.270
0.500
Statistic
Mean ± S.E.
Number
Mean ± S.E.
Number
Mean ± S.E.
Number
Mean ± S.E.
Number
Mean ± S.E.
Number
Days
7
0.1 ± 0.03
3
0.3 ± 0.00
3
0.7
2
2.2 ± 0.17
3
7.5 ± 0.62
3
after cessation of
14
0.1
2
0.2 ± 0.13
3
0.8
2
1.6 ± 0.10
3
5.8 ± 0.59
3
toxaphene exposure
28
N.D/
2
0.4 ± 0.10 0.3
3
0.9 0.7
2
1.4 ± 0.35 1.5
3
5.8 ± 0.58
3
56
N.D.
2
± 0.
3
± 0.
3
± 0.
3
3.7
2
06
03
07
^Source: Mayer et al. , 1975.
Toxaphene concentration in water before cessation of toxaphene exposure.
^Nominal concentration.
,Standard error.
None detected, detection limit 0.05 ppm.
5-15
-------
exposure, as well as information on the kinetics of toxaphene uptake and species
specific differences in toxaphene bioconcentration. The second type of study
usually involves one-time measurements of toxaphene fish residues in aquatic
systems not specifically treated with toxaphene. These studies presumably
give some indication of ambient levels of fish contamination by toxaphene.
Toxaphene residues found in fish from toxaphene-treated lakes are generally
consistent with the results of laboratory studies described previously and indi-
cate that fish bioconcentrate toxaphene by a factor of several thousand. Kallman
and coworkers (1962) monitored the uptake of toxaphene by rainbow trout and
bullheads in Clayton Lake, New Mexico. Seventy-two hours after the initial
application of toxaphene, toxaphene water levels ranged from undetectable
(limits of detection not given) to 12 ppb. Assuming zero values for all samples
in which toxaphene was not detected, the average water concentration for this
period was 2.4 ppb. Over this period, trout concentrated toxaphene by factors
of 180 to 2250 having toxaphene residues of 0.43 ppm to 5.4 ppm. Over the
same period, bullheads concentrated toxaphene by factors of 200 to 1750 with
toxaphene residues ranging from 0.51 ppm to 4.2 ppm. Five days after initial
treatment, average toxaphene water concentration was 12 ppb (2 ppb on the
windward side and 21 ppb on the lee side). By this time, bullheads were se-
verely affected by the toxaphene and had mean body residues of about 13 ppm.
After toxaphene water levels in the lake stabilized at about 1 ppb, trout ex-
posed for seven days had toxaphene whole-body residues between 0.8 ppm and
2.5 ppm (bioconcentration factors of 800 to 2,500). In similar exposures for
17 days, trout bioconcentrated toxaphene by factors of 1,300 to 3,500. By
523 days after initial toxaphene treatment, no toxaphene was detected in the
water, but trout exposed for 108 to 170 days had body residues of 0.3 ppm.
5-16
-------
In a similar study, Terriere and coworkers (1966) monitored toxaphene
water levels and fish residues in two Oregon Lakes: Davis Lake treated in the
fall of 1961 at a calculated concentration of 88 ppb and Miller Lake treated in the
fall of 1958 at a calculated concentration of 40 ppb. Results for Davis Lake are
summarized in Table 5.5. Rainbow trout consistently accumulated greater levels of
toxaphene than Atlantic salmon. As in the experimental exposures by Mayer and co-
workers (1975), fish concentrated the toxaphene by a factor of several thousand.
For Miller Lake, residue levels were determined only in brook trout for one year,
1964. Here, toxaphene water concentration was 0.84 (0.7 to 1.1) ppb and whole body
residues in brook trout were 12.46 (8.30 to 24.80) ppm with a bioconcentration
factor of about 14,800. The rate of toxaphene uptake by brook trout in Miller
Lake over 10 to 18 day periods was determined by suspending the fish in live
boxes for varying periods of time (Figure 5.2). As in the short-term uptake
studies by Schaper and Crowder (1976), uptake appeared to be linear over the
first ten days but plateaued after this time in the 1964 test.
Hughes and Lee (1973) present long term kinetic data on toxaphene residues
in bluegills from Fox Lake (Figure 5.3). Two groups of fish were monitored in
this study: those that had been placed into the lake 11 months after initial
toxaphene treatment (stocked), and young bluegills from the successful spawn-
ing of this first group (hatched). At all times during this study, toxaphene
concentration in the water was below detectable limits (1 ppb). However, the
peak concentration at 200 days after stocking in both groups of fish is•pre-
sumed to represent the time that toxaphene water concentration became so low
that water-tissue equilibrium reversed in favor of toxaphene elimination. It
is interesting to note that after 787 days, toxaphene residues in both groups
5-17
-------
Table 5.5. Toxaphene total body residues in fish from Davis Lake
Toxaphene concentration
in water, ppb
Rainbow trout
Residue, ppm
BCF
Atlantic salmon
Residue, ppm
BCFC
1962
0.63
(0.5-0.9)
5.7
(1.2-12.0)
9,050
2.75
(2.6-2.9)
4,370
Year
1963
0.41
(0.3-0.6)
7.72
(2.75-13.7)
18,830
3.24
(1.11-5.50)
7,900
1964
<0.2
3.5
(3.2-3.8)
> 17,500
1.8
(1.5-2.1)
> 9,000
/Source: Terriere et al., 1966.
All figures give mean value and range in parentheses.
Bioconcentration factor: average tissue residue T average
concentration of toxaphene in water.
5-18
-------
1963 TESTS
46 a 10 12 14 l< II
OATS IN LAKE
Figure 5.2. The rate of uptake of whole-body levels of toxaphene by
rainbow trout in Miller Lake. Average concentration of toxaphene in water was
1.20 ppb for 1963 and 0.84 ppb for 1964. Source: Terriere et al. , 1966.
5-19
-------
100
o>
>»
o*
J-ao
1/1
2 60-
u
a.
<
x 2.0
O
\ STOCKED BLUEGILLS
HATCHED BLUEGILLS
IOO 200 3OO 400 500
DAYS AFTER STOCKING
600
700
800
Figure 5.3. Variation, with time, of toxaphene concentration
in bluegills from Fox Lake, mean and range indicated.
Source: Hughes and Lee, 1973.
5-20
-------
of fish approached 1.0 ppm which is about the concentration at which Mayer and
coworkers (1975) were able to demonstrate no significant elimination after
53 days (Section 5.2.1.4). In another phase of this study, Hughes and Lee
(1973) found that predator fish accumulated less toxaphene than prey fish.
Six months after stocking, bluegills and suckers, both prey fish, had total
body residues of 9.4 ppm and 10.6 ppm, respectively. At the same time, the
predator fish bass, northern pike, and walleyes had body residues of only 2.2
ppm, 3.3 ppm, and 1.2 ppm, respectively.
Studies giving toxaphene residues in fish from aquatic systems not speci-
fically treated with toxaphene are summarized in Table 5.6. Some of these
studies reenforce various aspects of toxaphene metabolism previously described.
This is particularly true with reference to the distribution of toxaphene de-
scribed in Section 5.2.1.2. The results of Grzenda and Nicholson (1965)
suggest that, in most species studied, the edible portion of the fish tends to
accumulate less toxaphene than offal. The presence of toxaphene at 50 ppm in
carp fat as compared to only 0.44 ppm in the gills is consistent with the
supposition that toxaphene is preferentially sequestered in fat tissue (Johnson
and Lew, 1970).
Two recent monitoring surveys failed to find toxaphene in fish samples.
Reimold and Shealy (1976) monitored pesticide residues in fish collected from
eleven estuaries representing all the Atlantic drainage basins in Georgia and
South Carolina. The surveys were conducted semi-annually from 1972 to 1974.
Although dieldrin, DDT, PBC's and mercury were found in many samples, toxaphene
was not detected. Similarly, toxaphene was not found in fish taken from a
New Jersey salt marsh in 1967 and 1973 (Klaas and Belisle, 1977).
5-21
-------
Table 5.6. Toxaphene residues in fish from various fresh-water systems
Ln
I
Fish
Rainbow trout
Bullhead
White sucker
Carp
Northern pike
Catfish
Carp
Channel catfish
Threadfin shad
Sonoran sucker
Gilia sucker
Location Tissue
New Mexico (lake) Whole body
M II II II
South Dakota (lake) Whole body minus
internal organs
n ii
M ti
Mississippi, commercial Fillet
fish ponds
Colorado River Basin Fat
Gills
" " Muscle
Red muscle
Scraped skin
Fat
11 " Whole fish
Skin and muscle
Gills
Ovary
Visera
" " Skin and muscle
Kidneys
Viscera
" " Whole fish
Skin and muscle
Viscera
Residue (ppm)
(average range)
0.06
0.41
0.083
0.176
0.074
1.98
(0.0-20.67)
50.0
0.44
6.8
0.55
1.32
9.8
(8.2-11.38)
1.05
1.73
4.75
0.70
1.71
2.19
(0.0-0.25)
4.23
32.52
(2.75-172.92)
25.00
2.81
(0.71-4.91)
34.09
(25.24-42.94)
Reference
Kallman et al. , 1962
Hannon et al. , 1970
Hawthorne et al., 1974
Johnson and Lew, 1970
-------
Table 5.6 (continued)
Fish
Chain pickerel
White sucker
White perch
Fresh water drum
Spotted sucker
01 n
\ Carp
NJ
OJ
Smallmouth buffalo
Largemouth bass
Channel catfish
Green sunfish
Largemouth bass
Redhorse suckers
Green chubs
Gizzard shad
Warmouth bass
Location
Maine
Delaware River
Lake Ontario
Lake Erie
South Carolina
Louisiana
Arizona
Arkansas
Oklahoma
Utah
Alabama Basin
it n
it n
n n
M n
n n
Tissue
Whole-body
n ii
n n
n n
n n
n n
n n
M n
n n
n ii
Fillet
Offal
Fillet
Offal
Fillet
Offal
Fillet
Offal
Fillet
Offal
Fillet
Offal
Residue (ppm)
(average range) Reference
0.36 Henderson et al.,
1969
1.60
1.25
.01, 0.24
0.03
0.03
0.01
0.01, 0.02
0.01
0.01
0.34 Grzenda and Nicholson,
0.55 1965
0.80
3.30
0.99
2.10
0.96
2.90
1.32
1.70
0.30
0.71
-------
Table 5.6 (continued)
I
M
Fish Location
Largemouth bass California
Brown bullhead " "
Carp
Channel catfish " "
Black crappie " "
Tui chub " "
Pumpkinseed " "
Rainbow trout " "
Bluegill Louisiana
Channel catfish " "
r*^r,^vA fhlA " "
Tissue
Flesh
Viscera
Flesh
Flesh
Viscera
Fat
Whole-body
Whole-body
n n
Flesh
Whole-body
n n
n n
Residue
(average
0.
1.
0.
0.
0.
0.
0.
1.
0.
0.
0.
2.
i
05
13
06
01
05
4
03
09
04
22
48
23
/. Q
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(n
(ppm)
range)
0-0.
2-2.
0-0.
0-0.
0-0.
0-8.
0-2.
0-2.
0-6.
n_/,
3)
0
19)
1)
1)
0)
57)
06)
6)
7q\
Reference
Keith and Hunt, 1966
Epps et al. , 1967
-------
5.2.2 Effects
5.2.2.1 Physiological and Biochemical Effects — Chronic toxaphene exposure
causes biochemical alterations associated with the "broken back" syndrome in
fathead minnows, brook trout, and channel catfish (Mayer et al., 1975; Mayer
and Mehrle, 1976; Mehrle and Mayer, 1975 a,b). Clinical details of this syn-
drome are discussed in Section 5.2.2.2.2. Fathead minnows exposed to toxa-
phene concentrations ranging from 0.055 ppb to 0.621 ppb for 150 days had
significantly increased levels of calcium but decreased levels of collagen in
the backbone (Table 5.7). Phosphorous levels, however, were not affected.
Because collagen is the primary constituent of the organic matrix around which
calcification and mineralization occur, Mehrle and Mayer (1975 a) proposed
that the increased calcium to collagen ratio made the backbones of these fish
more fragile and more liable to fracture. As detailed in Section 5.2.2.2.2,
electrical shock experiments support this conclusion. In addition to the
effects on collagen and calcium, all exposures caused significant (p less than
0.05) decreases in the following amino acids of bone collagen: alanine,
valine, leucine, isoleucine, lysine, phenylalanine, and hydroxyproline. In
the skin of exposed fish, only hydroxyproline content decreased. However,
because this amino acid is important in the formation of collagen, which is in
turn essential for wound healing, this effect may further limit the ability of
fish to cope with environmental stress.
Using hydroxyproline as an index of collagen synthesis, Mehrle and Mayer
(1975 b) have demonstrated that toxaphene concentrations as low as 0.068 ppb
inhibit collagen synthesis in brook trout sac fry by seven days after hatching.
Similar to the results with minnows, continuous exposure to low level toxaphene
concentrations for up to 90 days caused a pronounced decrease in collagen con-
tent and somewhat less pronounced increases in calcium content in the backbones
5-25
-------
Table 5.7. Collagen, calcium, and phosphorus concentrations in fathead minnow backbones
as affected by 150 day continuous exposures to various levels of toxaphene
t_n
1
N>
Backbone constituent 0
Collagen 32.27
Calcium 11.81
Phosphorus 4.08
,Source: Mehrle and Mayer,
F „ _ - - o A /, . F „ ~ , - 1 cr
Toxaphene
0.005
26.89C
15.10C
3.55
1975 a.
i
concentration (ppb) ,
0.132 0.288 0.621 f value LSD
•05
22.92C 19.94C 22.39C 7.64 2.14
23.26C 20.84C 24.31C 3.27 2.85
3.66 3.78 3.48 2.63 0.19
/I'U'UO >-.w-r, '(J-Ui _-.^J-.
Significantly different from controls (P <0.05),
-------
of these fish (Table 5.8). Because proline hydroxylase (the enzyme responsible
for the conversion of proline residues to hydroxyproline residues) requires
ascorbic acid as a cofactor, Mehrle and Mayer (1975 a,b) proposed that a local
vitamin C deficiency might account for these biochemical effects. Recently,
Mayer and Mehrle (1976) did show that decreased collagen content in the back-
bones of channel catfish was accompanied by decreased backbone vitamin C content
after toxaphene exposures similar to those described above. In that liver
vitamin C was not affected, these investigators suggest toxaphene induces
hepatic microsomal hydroxylases - e.g., P-450 - which utilizes vitamin C and
thus causes a shift of vitamin C from the backbone to the liver.
Like many chlorinated pesticides, toxaphene exerts a strong inhibitory
effect on fish ATPase activity. In vitro, toxaphene at 40 ppm inhibited
NaKMg-ATPase, NaK-ATPase, and MgATPase activities in trout gill microsomes by
49 percent, 36 percent, and 66 percent, respectively. At 4 ppm, only NaKMg-
ATPase (8.6 percent inhibition) and Mg-ATPase (24 percent inhibition) were
significantly affected (Davis et al., 1972). Similar inhibition has been dem-
onstrated in three types of ATPase activity in catfish brain, kidney, and gill
(Table 5.9). Oligomycin insensitive (i.e., non-mitochondrial) Mg-ATPase was
the most severely inhibited ATPase in all tissue. For the most part, brain
and gill ATPases were more severely affected than kidney ATPases (Desaiah and
Koch, 1975).
Toxaphene, along with a number of other chlorinated and organophosphate
insecticides, has also been shown to inhibit oxygen uptake by bluegill liver
mitochondria. At a concentration of 0.41 mg toxaphene per ml of reaction
medium, mitochondrial oxygen uptake was depressed by 69 percent. No signifi-
cant effect, however, was seen on phosphate uptake (Hiltibran, 1974). The
5-27
-------
Table 5.8.
Backbone composition of brook trout fry exposed to toxaphene.
expressed on basis of dried weight of backbone
Mean values
M
OQ
Days after
hatching, and
Toxaphene concn. (ppb)
backbone constituent
30 days
Collagen
(mg HyP/g)
Phosphorus (%)
Calcium (%)
60 days
Collagen
(mg HyP/g)
Phosphorus (%)
Calcium (%)
90 days
Collagen
(mg HyP/g)
Phosphorus (%)
Calcium (%)
0
51.34
5.73
1.23
20.10
12.05
2.64
19.48
10.60
10.20
0.039
34.66^
7.30
0.98
f\
13.86
10.. 81
2.85
j
16.42
15.61
15.34
0.068
26.l6d
9.89
1.33
,
14.02
9.89
2.54
i
16. 22^
20.25^
20.61°
0.139 0.288
19.76 24.99
8.80 10.90
1.05 1.43
(\ 0
10.98j
7.7ia
1.31
d
16.42*
19.87^
21.14a
.Source: Mehrle and Mayer, 1975 b.
\ = 6-8.
jCollagen estimated by determination of hydroxyproline.
Significantly different from controls (LSDg.os)-
All fish exposed to 288 ppb toxaphene died within 60 days and all those exposed to
0.502-ppb (not shown) died within 30 days.
-------
Table 5.9. Inhibition of catfish brain, kidney, and gill ATPases by toxaphene
a
Toxaphene
(micromoles)
NaK-ATPase
Brain Kidney Gill
% Inhibition
Oligomycin sensitive
Ms-ATPase
Brain Kidney Gill
Oligomycin insensitive
Mg-ATPase
Brain Kidney Gill
Ln
I
ho
1.8
3.6
7.2
14.5
29.0
8.2 -8.4
25.9 29.5
41.3* 31.6
48.0** 33.9
50.0** 33.7
-2.0
22.2
35.4*
36.1*
37.4*
19.3
24.5
46.1*
46.1
59.3*
-49.7
- 8.5
-3.1
22.9
42.3
-6.7
-2.5
9.0
40.9**
50.9***
29.7* 1.5
42.5** 16.0
49.9***28.0*
72.0***36.2**
74.8***39.7**
27.6*
33.9*
47.8**
56.6***
60.5***
a
/Source: Modified from Desaiah and Koch, 1975.
^Statistically significant when determined by Student's t test, *P <0.05, **P <0.01, ***P <0.001.
"Negative (-) values indicate per cent stimulation of enzyme activity.
-------
inhibition of oxygen uptake is consistent with the signs of respiratory dis-
tress seen in toxaphene-poisoned fish (Section 5.2.2.2.1).
5.2.2.2 Toxicity
5.2.2.2.1 Acute toxicity — Because toxaphene has been used as a piscicide in
fish management programs (Section 5.2.2.2.5), many studies have been conducted
on the acute toxicity of toxaphene to fish. In that such fish management
programs have been directed towards the elimination of rough fish in favor of
game fish, several of these studies have attempted to determine if toxaphene
is selectively toxic'to certain families or species of fish. As summarized in
Table 5.10, such selective toxicity does not appear to exist. Of the eight
families of fish for which acute toxicity estimates are available, the 96-hour
LCSO's range from 2 ppb to 32 ppb. However, no consistent differences are
apparent among the various families of fish. For example, the salmonidae, a
family consisting largely of desirable game fish, has 96-hour LC50 values
ranging from 2.5 ppb to 11 ppb. The ictalurids and cyprinids, which are less
regarded as game fish, have comparable LC50 ranges of 2.7 to 13 ppb and 4 to
32 ppb, respectively. Such a comparison, based on the combined results of
different investigators using a variety of different conditions and techniques,
may be of limited validity. As discussed in the following paragraphs, such
factors as pH, temperature, and alkalinity may affect acute toxicity estimates.
Nonetheless, even in the individual studies which tested a variety of different
species from different families, no clear pattern of species or family sensi-
tivity emerges. Although Macek and McAllister (1970) were able to demonstrate
substantial and consistent differences in the response of various families of
fish to carbamate and organophosphorous insecticides, no such differences were
observed with the organochlorine insecticides DDT, lindane, and toxaphene. Of
5-30
-------
Table 5.10. Acute toxicity of toxaphene to various fish during static exposures
FAMILY
Organism Size
~ Average (range)
Common name ° °
wgt . in grams
(Genus species) Length in mm
CEtJTKARCHIDAE
Bluegill sunfish 2.9 g
(Lepomis macrochirus) (1-2 g)
-
(0.6-1.5 g)
Green sunfish 3.17 g
(Lepomis cyanellus)
Redear sunfish
(Lepomis microlophus)
Largemouth bass
Temperature
°C
20°
25°
18°
18.3"
20°
18°
18°
Other
conditions
PH
PH
PH
PH
PH
PH
PH
7.4, hardness
7.4, hardness
7.1, alk.
7.1, alk.
35
35
7.4, hardness
7.1, alk.
7.4, alk.
35
35
LC in ppb after
specified interval
(95% confidence interval)
24 hr
24 ppm
20 ppm 7.5
ppm
ppm 6.8(6.2-7,5)
24 ppm
ppm
ppm
36 hr 48 tir 72 hr 96 hr Reference
23 Ferguson
3.8 3.5 Henderson
18(10-30) Macek and
2.6(2.2-3.0) Macek et
38 Ferguson
13(8-17) Macek and
2(1-3) Macek and
et
et
Me
al.
et
al. , 1964
al. , 1959
Allister,
, 1969
al. , 1964
McAllister,
McAllister,
1970
1970
1970
(Micropterus salmoides)
CYPRINIDAE
Carp
(Cyprinus carpio)
Goldfish
(Carassius auratus)
Golden shiner
(Notemigonus crysoleucas)
minnow, Blantnose
(Pimephales notatus)
18°
pH 7.1, alk. 35 ppm
56(48-67)
4(3-5)
(1-2 g)
(33-64 mm)
2.43 g
25° pH 7.4
18°
pH 7.1, alk. 35 ppm
17.2° pH 7.9, alk. 36 ppm
20°
pH 7.4, hardness 24 ppm
17.2° pH 7.9, alk. 36 ppm
17.2° pH 7.9, alk. 36 ppm
8.2
86
27
200
6.8
35
30
7.0
35
28
6.2
14
5.6
14(11-19)
28
< 5.0
8.8
Ludemann and Neumann, 1960
Macek and McAllister, 1970
Henderson et al., 1959
Macek and McAllister, 1970
Mahdi, 1966
Ferguson et al., 1964
Mahdi, 1966
Mahdi, 1966
-------
Table 5.10 (continued)
I
OJ
ho
FAMILY
Organism Size
,, Average (range)
Common name ^ „
wgt. in grains Temperature
(Ceruis species) Length in mm °C
CYPRINIDAE (cont'd)
minnow, Fathead (1.
(Pimephales promelas) (1
Stoneroller
(Campos toma anomalum)
CASTEROSTEIDAE
Threespine stickleback (0.
.0-1.5 g)(51-64
.0-2.0 g)(38-64
(51-64
-
-
mm)
mm)
mm)
17
.38-0.77 g) (22-44 mm)
25°
25°
24°
18°
.2°
20°
-
PH
PH
PH
pH
PH
pH
Other —
conditions
7.4,
7.4,
7.1,
7.9,
7.1,
7.1.
hardness 20 ppm
alk. 212 ppm
alk. 35 ppm
alk. 36 ppm
salinity 5%
salinity 25%
LC in ppb after
specified interval
(95% confidence interval)
24 hr 36 hr 48 hr 72 hr 96 hr
13
13 7.5 7.5
5.7(4.1-8.0)
14(9-22)
54 44 35 32
12 9.8 8.6
8.8 7.8
Reference
Cohen et al. , 1960
Henderson et al., 1959
Hooper and Crzcnda,
Macek and McAllister
Mahdi, 1966
Katz, 1961
1957
, 1970
ICTALURIDAE
Black bullhead
(Ictalurus melas)
Channel catfish
(Ictalurus punctatus)
PERCIDAE
Yellow perch
(Perca flavescens)
POECIUIDAE
Kuppy
(Lebistes reticulatus)
20° pH 7.4, hardness 24 ppm
18° pH 7.1, alk. 35 ppm
17.2° pH 7.9, alk. 36 ppm 15
18° pH 7.1, alk. 35 ppm
18° pH 7.1, alk. 35 ppm
(0.1-0.2 g)(19-25 ppm) 25° pH 7.4, hardness 20 ppm 42
3.75 Ferguson et al., 1965 a
5(4-7) Macek and McAllister, 1970
4.3 4.2 2.7 Mahdi, 1966
13(10-17) Macek and McAllister, 1970
12(9-14) Macek and McAllister, 1970
24
20 Henderson et al., 1959
-------
Table 5.10 (continued)
Ol
Co
FAMILY
Organism Size
,, Average (range)
Common name e
wgt. in grams Temperature
(Genus species) Length in mm °C
POECIL1IDAE (cont'd)
Mosquito fish
(Gambusia affinis) 0.28 g
(30-40 mm)
1.08 g
SALMON IDAE
salmon, Chinook (1.45-5 g) (51-114 nn
(Oncorhynchus tshawytscha)
salmon, Coho (2.7-4.1 g)(57-76 mn
(Oncorhynchus kisutch)
trout, Brook 16 months old
(Salvelinus fontinalis)
trout. Brown
(Salmo trutta)
trout, Rainbow 1 g
(Salmo g-airdneri) 3.2 g (51-79 mm)
21°
24°
20°
«) 20°
n) 20°
13°
-
13°
12. i
20°
13°
12°
pH 7
pH 7
pH 7
pH 7.
pH 7,
pH 1.
r
pH 7
pH 7
pH 7
Other
conditions
.8, hardness 28 ppm
.4, hardness 24 ppm
.1, alk. 51 ppm
.1, alk. 51 ppm
.1, alk. 35 ppm
-
.1, alk. 35 ppm
.1, alk. 51 ppm
. 1, alk. 35 ppm
.9, alk. 36 ppm
LC in ppb after
specified interval
(95% confidence interval)
24 hr 36 hr 48 hr 72 hr 96 hr
10
1.0
45 24 9 8
31(LC 63)
10
860
7.9 3.3 2.7 2.5
13.0 10.5 10.0 9.4
8(6-10)
10.8
3(2-5)
7.6 4.4 2.7
11.5 8.4 8.4 8.4
11(8-13)
30 14.5 11.1 8.4
Reference
Boyd and Ferguson, 1964
Burke and Ferguson, 1969
Chaiyarach et al. , 1975
Dziuk and Plapp, 1973
Ferguson et al., 1965 a
Schaper and Crowder, 1976
Katz, 1961
•Katz, 1961
Macek and McAllister, 1970
Mayer et al. , 1975
Macek and McAllister, 1970
Cope, 1965
Katz, 1961
Macek and McAllister, 1970
Mahdi, 1966
SERRANIDAE
Slriped bass
(Morone saxatilis)
2.3 g (60 mm)
17° salinity 30%
4.4(2-9) Korn and Earnest, 1974
-------
the twelve species of fish tested by Macek and MacAllister (1970), bullheads
(ictalurids), carp (cyprinids), largemouth bass (centrarchids), and brown trout
(salmonids) were significantly more susceptible to toxaphene than the remain-
ing eight species. The four susceptible species had 96-hour LC50 values rang-
ing from 2 to 5 ppb, while the remaining species had a corresponding range of
8 to 14 ppb. In all but one instance, the LC50's for the sensitive species were
significantly different (p less than 0.05) from those of the other species.
Thus, while sensitive and tolerant species of fish were found, the pattern of
sensitivity did not fall into family groups. The centrarchids contained both
the most and least sensitive species: the largemouth bass with a 96-hour LC50
of 2 ppb and the bluegill sunfish with a 96-hour LC50 of 18 ppb.
An examination of the results of other studies summarized in Table 5.10
which used more than one species of fish also reveals no clear patterns of
species or family sensitivity. Of the six species of fish tested by Mahdi
(1966), the black bullhead (ictaluridae) was more susceptible to toxaphene
than any of the four species of cyprinids at 17 C but not at 11.6 C. Further,
at the lower temperature, the rainbow trout (salmonid) was more susceptible
(LC50 8.4 ppb) than the ictalurid or any of the cyprinids (LCSO's 14 to 94
ppb). Similarly, Katz (1961) found no remarkable differences in the toxicity
of toxaphene to three species of salmonids and one gasterosteid. Given the
marked effect that preexposure, genetic resistance, and a variety of experi-
mental parameters can have on such acute toxicity estimates (following para-
graphs and Section 5.2.2.2.4), the relatively narrow range of LC50's noted in
Table 5.10 does not suggest major species or family specific differences in
the acute toxicity of toxaphene to fish.
5-34
-------
In the treatment of freshwater lakes with toxaphene as a piscicide,
various investigators have noted that certain lakes remain toxic to fish for
longer periods of time than other lakes receiving comparable doses of toxa-
phene (Section 5.2.2.2.5). While these findings could be attributable to
differing rates of toxaphene breakdown in different aquatic systems (Section
7.3.2), the possibility that different toxaphene formulations or varying water
quality characteristics might also affect the toxicity of toxaphene to fish
also has been examined. Based on the current available information, detailed
below, substantial (greater than 10 C) increases in temperature do seem to
increase the toxicity of toxaphene to most fish, but the increase is generally
less than a factor of five. Other water quality variables, such as pH, alkal-
inity, hardness, and salinity have less pronounced effects, and the formulation
of toxaphene used seems to have little, if any, effect. None of these factors
approach, in magnitude, an apparent importance of preexposure or genetic
resistance (Section 5.2.2.2.4).
Most toxicants have a positive temperature coefficient - i.e., toxicity
increases with increasing temperature. Macek and coworkers (1969) found this
relationship to hold for a variety of commercial pesticides including toxaphene.
As summarized in Table 5.11, most of the available data on the toxicity of
toxaphene to fish also indicate a positive temperature coefficient. In all
but one instance where temperature differs by more than 10 C, the toxicity of
toxaphene is increased at the higher temperature. The single exception, the
24-hour LC50 values for the goldenshiner, is of questionable significance be-
cause confidence intervals are not given and because a positive temperature
coefficient is suggested at 48 and 72 hours.
In general, the magnitude of the positive temperature coefficient seems
to decrease or stay about the same as the exposure period increases. For the
5-35
-------
Table 5.11. Effects of temperature in the acute toxicity of toxaphene to fish
FAMILY Temperature
Organism °C
CENTRARCHIDAE
Bluegill sunflsh 12.7
ID 1
lo . J
23.8
CYPRINIDAE
Fathead minnow 10
24
Stoneroller 11.6
Ui 17'2
1 22.8
LO
<^ Goldfish 11.6
17.2
22.8
Bluntnose minnow 11.6
17.2
22.8
Goldenshiner 11.6
17.2
22.8
1CTALURIDAE
Black bullhead 11.6
17.2
22.8
SAI.MONIDAE
Rainbow trout 7.2
12.7
18.3
24 hr
9. 7
(8.4-11)
6.8
(6.2-7.5)
6.6
(5.5-7.9)
20
(14-28)
5.7
(4.1-8.0)
62
54
9
270
86
54
> 4000
200
15
12.5
27
13.4
> 48
15
12
16.0
7.6
5.0
LC5() in
(95% confidence
48 hr
27
44
7.8
115
35
50
860
35
8.4
7
6.6
> 48
4.3
4.2
8.4
4.4
2.8
ppb
Interval)
72 hr
27
35
< 5
94
28
50
150
14
6.5
6.2
6.5
34
4.2
3
96 hr R.I.T. Reference
3.2
(2 8-3 7) 1-3 Macek et al., 1969
2.6
(2.2-3.0)
2.4
(2.1-2.7)
3.5 Hooper and Grzenda, 1957
14 > 2.8 Mahdl, 1966
32
< 5
94 1.9
28
50
30 4.8
8.8
6.3
b
0.93
< 5
6
25 13.9 Mahdi, 1966
2.7
1.8
5.4 3.0 Cope, 1965
2.7
1.8
£R.I.T. - Relative increase in toxicity:
• unless otherwise specified.
at highest temperature divided by LC,._ at lowest temperature. 96 hr value used
24 hr
used.
-------
bluntnose minnow, the R.I.T. (relative increase in toxicity as defined in
Table 5.11) between 11.6 C and 22.8 C is more than 267 in 24-hour exposures
but drops to 4.8 in 96-hour exposures. The only apparent exception to this
pattern is the black bullhead which has R.I.T. values of greater than 4 and
13.9 at 24 hours and 96 hours, respectively. As with the estimates of acute
toxicity discussed above, the rather wide variability found within the cyp-
rinids, and the limited information available on the other families, cautions
against general statements on the family or species sensitivity to the temper-
ature effect. However, it is interesting to note that the black bullhead,
which seems to be among the fish more sensitive to toxaphene, also has a
markedly higher R.I.T. than the other species listed in Table 5.11.
In addition to temperature, a variety of other water characteristics such
as pH, alkalinity, hardness, and salinity, could affect measurements of acute
toxicity. However, relatively little detailed information is available in
these factors in estimates of toxaphene toxicity. Hooper and Grzenda (1957)
noted a slight, but not statistically significant, increase in the toxicity of
toxaphene to fathead minnows over 24-hour exposures at 10 C with increasing
alkalinity. At an alkalinity of 6 ppm, the LC50 (95 percent confidence inter-
val) was 36 (26 to 50) ppb. When the alkalinity was raised to 212 ppm, the
corresponding toxicity estimate was 20 (14 to 28) ppb. Using the same species,
Henderson and coworkers (1959) also noted a slight increase in the toxicity of
toxaphene at alkalinities of 18 ppm and 360 ppm with corresponding LC50 values
of 7.5 ppb and 5.1 ppb.
The effect of pH has not been systematically evaluated. Workman and
Neuhold (1963) tested goldfish and mosquito fish using two toxaphene formula-
tions in waters from three different lakes with pH values of 7.0, 7.8, and 8.3.
5-37
-------
Statistically significant differences among the various waters were uncommon.
In that the lakes also differed markedly in dissolved solids (46 to 238 ppm),
the significant differences which were found could not be unequivocally attri-
buted to pH.
The only other water quality characteristic which has received any atten-
tion is salinity. As summarized in Table 5.10, Katz (1961) demonstrated a
slight increase in the toxicity of toxaphene to the threespine stickleback
(Gasterosteidae) as salinity increased from 5 parts per thousand to 25 parts
per thousand.
No information is available on the effect of dissolved oxygen on toxaphene
fish toxicity. Courtenay and Roberts (1973) attempted to obtain such informa-
tion but were unsuccessful due to problems in exposure technique. This is
particularly regrettable, given the speculations of Schaper and Crowder (1976)
described below, that respiratory distress may be a major factor in acute
toxaphene exposures.
Toxaphene is or has been available in a variety of formulations. The
studies summarized in Table 5.10 identify the toxaphene used as commercial
formulations with toxaphene concentrations of 0.48 kg/liter, 0.72 kg/liter, or
0.96 kg/liter, or as "pure" or technical grade toxaphene. Those studies which
used pure or technical toxaphene in a specified solvent (usually acetone) con-
ducted appropriate solvent controls, the details of which are not included in
this report. No systematic studies are available on the potential effect of
the various solvents or emulsifying agents used in commercial formulations on
the toxicity of toxaphene to fish. Two studies, however, have compared the
potency of different toxaphene formulations. Mayhew (1955) exposed fingerling
rainbow trout to toxaphene as an emulsifiable concentrate or wettable powder
5-38
-------
at concentrations (as toxaphene) ranging from 50 to 1000 ppb. No fish sur-
vived exposure to the lowest concentration of either formulation for over 16
hours. Because of this high mortality, standard LC50 values cannot be calcu-
lated. Nevertheless, the emulsifiable concentrate seemed slightly more toxic,
killing all fish exposed to 100 ppb after 12 hours. Identical exposure to the
wettable powder killed 70 percent of the fish. Similarly, Workman and Neuhold
(1963) compared the toxicity of sinking and floating formulations of toxaphene.
Using various combinations of temperature, water type, and species, statistically
significant differences between these two formulations were demonstrated in
only one of eight trials. Thus, there is no clear indication that the formula-
tion of toxaphene used markedly affects the toxicity estimates given in Table
5.10.
The mechanism by which toxaphene adversely affects fish is not known.
Like most chlorinated hydrocarbon insecticides, toxaphene is assumed to act
primarily in the nervous system, and such an assumption is consistent with
most of the signs of toxaphene poisoning in fish. Henderson and coworkers
(1959) found that the responses of fathead minnows, bluegills, goldfish, and
guppies to nine chlorinated hydrocarbon pesticides, including toxaphene, were
essentially the same. Initially, these fish displayed hyperactivity followed
by periods of muscular spasms. As intoxication progressed, the major sign of
poisoning was loss of equilibrium accompanied by periodic short, jerky move-
ments. Similar patterns have been seen in carp, tilapia, trout, bullheads,
mosquito fish, and goldfish (Gruber, 1959; Kallman et al., 1962; Ludemann and
Neumann, 1960; Schaper and Crowder, 1976; Workman and Neuhold, 1963). The
period of initial hyperactivity may be concentration dependent. Workman and
Neuhold (1963) note this effect only at water concentrations of more than 3
5-39
-------
ppm toxaphene. The loss of equilibrium seen in the latter stages of intoxica-
tion is characterized by partial paralysis, impaired locomotion, and a loss of
dorso-ventral orientation with the fish turning on to their sides either at
the surface of the water or the bottom of the exposure vessel (Ludemann and
Neumann, 1960; Workmann and Neuhold, 1963). In addition to these signs of
apparent nervous system damage, Schaper and Crowder (1976) have noted signs of
respiratory distress in mosquito fish. Shortly after exposure to 2 ppm toxa-
phene, fish began to display swimming behavior characteristic of low water
oxygen levels, and rapid gill ventilation was noted throughout the exposure.
As discussed in Section 5.3, suffocation rather than nerve damage has been
proposed as a mechanism of toxaphene1s effect on aquatic invertebrates.
Little detailed information is available on the total body burden of
toxaphene necessary to cause signs of intoxication in fish. Bullheads showing
advance signs of toxaphene poisoning after three to six days exposure in a
toxaphene-treated lake had total body toxaphene concentrations of 8 to 15 ppm
(Kallman et al., 1962). However, in mosquito fish showing similar signs of
intoxication after exposure to 2 ppm toxaphene for up to eight hours, total
body residues of only 0.42 to 0.75 ppm were found (Schaper and Crowder, 1976).
Compared to other common pesticides, toxaphene is an extremely potent fish
toxicant on acute exposure. Table 5.12 summarizes some of the available infor-
mation on the toxicity of toxaphene relative to some other pesticides. Infor-
mation on additional pesticides and other fish species is given in the studies
cited in this table. With the exception of endrin, toxaphene is almost invari-
ably more toxic to fish than other chlorinated hydrocarbons and consistently
more toxic than organophosphorous or carbamate pesticides.
5-40
-------
Table 5.12. The relative acute toxicity to fish of some common pesticides compared to toxaphene
a
l-n
I
Pesticides
Endrin
Dieldrin
DDT
Heptachlor
Lindane
Malathion
Parathion
Carbaryl
„ b
Carp
14.0
0.83
0.98
0.15
0.20
0.003
0.016
-
c.
Rainbow trout
8.0
0.80
0.80
0.44
0.18
0.005
-
-
Fish
Bluegills
5.8
0.44
0.22
0.18
0.045
-
-
Striped bass Bullhead"
47.0
0.22
8.3 1.0
1.5
0.60 0.078
0.31 0.00039
0.25
0.0044 0.00025
/Relative toxicity calculated as LC..Q of toxaphene -s- LC
Source: Ludemann and Neumann, 1960.
/Source: Cope, 1966.
Source: Henderson et al., 1959.
/Source: Korn and Earnest, 1974.
"Source: Macek and McAllister, 1970.
of specified pesticide.
-------
Other topics dealing with the acute toxicity of toxaphene to fish are
resistance/tolerance and drug interactions, which are discussed in
Sections 5.2.2.2.4 and 5.3.2.2.7, respectively.
The fish toxicity of individual toxaphene components has received rela-
tively little attention. A mixture of two toxaphene components, 2,2,5-endo,6-
exo,8,8,9,10-octachloroborane and 2,5,5-endo,6-exo,8,9,9,10-octachloroborane
(toxicant A), is about twenty-five times more toxic to goldfish than technical
toxaphene and has a 24-hour LC50 of 1.7 ppb to this species. Another component,
2,2,5-endo, 6-exo,8,9,10-heptachloroborane (toxicant B) is only about five
times more toxic to this species than technical toxaphene, having a 24-hour
LC50 of 8.6 ppb (Turner et al., 1975). Similar patterns have been noted in
the toxicity of these components to mice (Table 6.13, Section 6.2.2.1.2) and
houseflies (Table 5.61, Section 5.6.2.2.1.2).
5.2.2.2.2 Subacute and chronic toxicity— Relatively few controlled studies
are available on the effects of prolonged exposures of fish to toxaphene. While
studies on fish kill in toxaphene-treated lakes can be used to supplement the
available experimental information, such studies are inherently epidemiological,
involving ill-defined dosage data, uncontrolled, and often undefined exposure
conditions, and few details on toxic response other than mortality estimates.
Consequently, such case studies are discussed separately (Section 5.2.2.2.5).
As described in Section 5.2.2.1, Mayer and coworkers (Mayer and Mehrle,
1976; Mayer et al., 1975; Merhle and Mayer, 1975 a, b) have demonstrated that
prolonged toxaphene exposure causes decreased spinal collagen in fathead minnows,
brook trout, and channel catfish. Using fathead minnows, Mehrle and Mayer
(1975 a) related this biochemical effect directly to increased backbone fragil-
ity. Six fish from each toxaphene-treated group (0.055 ppb to 0.621 ppb for
5-42
-------
150 days) were exposed to sublethal electrical shocks. In each instance, four
of the six fish had multiple complete fractures of the spine. No such breaks
were seen in control fish given identical electrical shock treatments. In
addition to spinal weakness, all exposures resulted in decreased growth by 150
days (Table 5.13). However, although such growth depression was statistically
significant, the extent of depression ranged from only 4 percent to 22 percent.
As with the effects on collagen synthesis, the extent of growth retardation
was not dose-related. All of these chronic effects were associated with total
body residues of 5.9 to 52 ppm. Excessive mortality was seen only in fish
exposed to 1.23 ppb for 150 days (Mehrle and Mayer, 1975 a).
Although these investigators did not attempt to demonstrate spinal fragility
in other fish species, comparable studies on growth and mortality in brook
trout suggest that this game fish is markedly more sensitive to toxaphene than
fathead minnows. Brook trout fry exposed to toxaphene at concentrations as
low as 0.039 ppb evidenced significantly decreased growth after 60 days (Table
5.14). With these fish, the extent of growth depression was approximately 50
percent. This effect was associated with total body residues generally under
1 ppb. Thus, the increased sensitivity of these fish compared to minnows is
not attributable to more rapid or complete absorption. Further, complete
mortality was seen in all fry exposed to 0.288 ppb for 60 days and 0.502 ppb
for 30 days (Mehrle and Mayer, 1975 b). Yearling brook trout are somewhat
less susceptible than fry (Mayer et al., 1975). At exposures of 0.270 ppb for
180 days, growth of yearlings, expressed both as length and weight gain, was
depressed 24 percent. In similar exposures to 0.5 ppb, length was reduced 24
percent and weight gain reduced 46 percent. All of these differences from
control fish were significant at p less than 0.05. In addition to growth
5-43
-------
Table 5.13. The effects of chronic toxaphene exposure on the growth of fathead minnows'*
Days of
exposure
60
90
Ul
£ 150
Toxaphene concentration
Measurement
length
weight
length
weight
length
weight
(mm)
(g)
(mm)
(g)
(mm)
(g)
0
30.4
0.27
40.0
0.68
48.6
1.28
0.055
30.2
0.27
39.2
0.66
46. 2C
1.14
0.132
30.
0.
39.
0.
45.
1.
6
28
2
64
2C
oic
0.
30
0
38
0
46
1
(ppb)
288
.1
.27
.9
.63
.6C
.14C
0.621
30.
0.
38.
0.
46.
1.
0
27
8
62 -
2C,
04C
b
F value
0.34
0.42
0.76
0.52
2.44
2.52
/Source: Mehrle and Mayer, 1975 a.
CF.05 = 2.26; F01 = 3.12.
Significantly different from controls
-------
Table 5.14. The effect of chronic toxaphene exposure in the growth of brook trout fry
Ln
1
•IS
Ln
Toxaphene
concn
(ppb)
0
0.039
0.068
0.139
0.288
0.502
Fry length in mm
(standard deviation in parentheses)
0 day
16.5
(0.8)
16.8
(1.1)
16.8
(1.1)
16.8
(1.2)
16.3
(1.0)
16.4
(1.0)
30 days
23.4
(1.2)
23.3
(2.2)
23.6
(2.0)
22. 06
(2.1)
19.46
(1.6)
c.
60 days
34.3
(4.4)
31.46
(3.3)
31.3b
(3.4)
30.86
(4.3)
c
c
90 days
42.0
(9.2)
35. 86
(6.2)
38.3
(8.2)
35.16
(7.7)
c
c
Weight (g)
at
90 days
0.81
(0.60)
0.446
(0.30)
0.596
0.44
0.436
(0.37)
c
e
.Source: Mehrle and Mayer, 1975 b.
Significantly different from controls
All fish dead.
-------
depression, hyperexcitability, discoloration, and increased incidences of fin
erosion were seen in these fish. The subacute toxicity of toxaphene to year-
ling trout, expressed as 14 day LC50 (95 percent confidence interval), was 3.6
(3.2 to 4.0) ppb. Although mortality data from the chronic exposure was not
suitable for calculating a chronic LC50, Mayer and coworkers (1975) indicate
that 180 day exposures to 0.288 ppb caused about 50 percent mortality in
yearling brook trout.
Various pathological lesions have been reported in fish after subacute or
chronic exposure to toxaphene. Like many chlorinated hydrocarbon pesticides,
toxaphene seems to cause histological alteration of liver tissue. In five-
month exposures of Leiostomus xanthurus (spot) to toxaphene concentrations of
0.01 ppb and 0.1 ppb, no growth retardation was noted, but unspecified degenera-
tive changes were seen in the livers of treated fish (Lowe, 1964). Similarly,
Cope (1966) also reports liver damage in the trout exposed to toxaphene (details
of exposure not specified). In toxaphene exposures of from 1 ppb to 100 ppb,
carp displayed dose-related ultrastructural changes in hepatic cells charac-
teristic of enzyme induction. Such changes included proliferation of the
rough endoplasmic reticulum, swelling of the mitochondria, and condensation of
nuclear chromatin (Rutschke and Brozio, 1975).
In addition to liver degeneration, Lowe (1964) reports a pronounced
thickening in the gill lamellae of toxaphene-treated fish but no damage to
renal or nervous tissue. However, in the fry of largemouth bass given toxa-
phene in food at 10 percent of the 96-hour TLm for 14 days, Courtenay and
Roberts (1973) found extensive necrosis of kidney tissue with almost complete
degeneration of the kidney tubules. Necrotic changes were also seen in the
lining of the digestive tract. The only positive report of nervous system
damage is given by Kayser and coworkers (1962). In this instance, cell swelling,
5-46
-------
chromatolysis, and agglutination of the Nissal substance around the cell
nuclei were found in the brain cells of carp intoxicated with toxaphene.
5.2.2.2.3 Reproductive and teratogenic effects — Toxaphene seems to have an
adverse effect on egg development when exposure occurs via gravid females.
Boyd (1964) noted that 5 percent of the gravid mosquito fish aborted after
surviving toxaphene exposures that caused 10 to 40 percent mortality (toxaphene
concentration not specified). Similarly, egg viability was markedly reduced
when spawning female brook trout were exposed to toxaphene concentrations of
0.075 ppb and above (Table 5.15). However, eggs obtained from females not
exposed to toxaphene were incubated in solutions of 0.039 ppb to 0.502 ppb
toxaphene for twenty days prior to hatching and no decrease in egg hatchabil-
ity was seen (Mayer et al., 1975; Mehrle and Mayer, 1975 b).
Courtenay and Roberts (1973) have noted developmental abnormalities in
striped mullit (Mugil capbalus) embryos exposed to toxaphene concentrations
of up to 0.5 ppm (more detailed dosage data not available). These organisms
exhibited enlargement and proliferation of glandular structures, apparently
mucous glands, in the optic region. Because such changes might be attributable
to epithelial irritation rather than to actual maldevelopment, the teratogenic
implications of these findings are questionable (Courtenay and Roberts, 1973).
5.2.2.2.4 Resistance/tolerance — Ferguson and coworkers (Boyd and Ferguson,
1964; Burke and Ferguson, 1969; Ferguson et al., 1964, 1965 a) were the first
to note that fish of the same species taken from different locations differed
substantially in their susceptibility to toxaphene. Boyd and Ferguson (1964)
associated these differences with different levels of preexposure of fish popu-
lations to pesticide runoffs. Mosquito fish were collected from four areas of
Mississippi, one of which was reasonably free of pesticide contamination, and
5-47
-------
t_n
I
00
Table 5.15. The effect of toxaphene on brook trout reproduction
a.
Exposure,
ppb
Control
0.041
0.075
0.125
0.270
0.500
Number of
females
spawning
4
6
6
6
4
1
Number
of
spawns
9
8
9
12
7
1
Eggs
per
spawn
380
406
344
271
264
617
Eggs
per
female
855
541
516
542
462
617"
% Viability
69
60
22C
15C
6C
oc
/Source: Mayer et al., 1975.
Nominal concentration.
Significantly different from controls (P less than 0.05).
-------
allowed to breed in control ponds containing no pesticides. The fish population
from the area free of pesticides, which was designated as susceptible, had a
36-hour LC50 to toxaphene of 10 ppb. Fish populations from the other areas
had LCSO's of 60 ppb to 480 ppb. In that these LCSO's were obtained from second
or third generation offspring, genetic resistance rather than some form of pre-
exposure tolerance was proposed. In a subsequent study on mosquito fish, Burke
and Ferguson (1969) identified a fish population 200 times more resistant to
toxaphene than the susceptible population. However, dose response curves for
both populations of fish had about the same slope (Figure 5.4). Similar patterns
of resistance have been reported in mosquito fish populations in Texas (Dziuk
and Plapp, 1973). Subsequent studies also found evidence of resistance in other
species of fish. As summarized in Table 5.10, Ferguson and coworkers (1964)
determined 36-hour LCSO's for susceptible strains of goldenshiners, bluegills,
and green sunfish at 30 ppb, 38 ppb, and 62 ppb, respectively. However, cor-
responding LCSO's in the same three species of fish collected in areas of high
pesticide runoff were 1200 ppb, 1600 ppb, and 1500 ppb, respectively, giving
resistance factors of from 39 to 70. In a similar study, various populations
of black bullheads were found to differ from each other in their susceptibility
to toxaphene by a factor of up to 13. In general, all of these studies indica-
ted that resistance was developed not only to toxaphene but also to a variety
of other chlorinated pesticides.
Further supporting the conclusion of Boyd and Ferguson (1964) that fish
populations in highly contaminated areas possess genetic resistance rather than
some form of tolerance, Lowe (1964) found that preexposure of spots to toxaphene
concentrations of 0.01 ppb or 0.1 ppb for five months did not increase the
tolerance of these fish to acute toxaphene poisoning. In fact, fish chronically
5-49
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100
2- 75
o
50
25
Susceptible
I
0 0.25 0.50 0.75 1.00 1.25
Concentration in ppb
1.50
100
I" 75
S
o
50
25
Resistant
I
50 100 150 200 250
Concentration in ppb
300
Figure 5.4. Dose-response curves for susceptible and resistant strains
of mosquito fish exposed to toxaphene for 36 hours. Source: Burke and Ferguson, 1969,
5-50
-------
exposed to sublethal levels of toxaphene were more sensitive to acute doses
than fish with no history of preexposure. While it thus seems probable that
fish populations in areas contaminated with pesticides do possess genetic
resistance to toxaphene poisoning, the mechanisms of such resistance are not
known. Finley and coworkers (1970) have suggested that high levels of pesti-
cides, particularly DDT and toxaphene which appear in freshwater after heavy
rains, may exert the primary selection pressure in the development of resis-
tance. Fabacher and Chambers (1971) have noted some correlation between
resistance and increased body lipids in mosquito fish. However, patterns of
cross-resistance made it unlikely that this factor alone could account for
resistant populations. In studies on the inhibition of succinic dehydrogenase
in mitochondrial preparations from resistant and susceptible mosquito fish,
Moffett and Yarbrough (1972) noted that the patterns of inhibition between
mitochondria with intact and disrupted membranes suggested a relationship
between resistance and membrane permeability. However, no relationship was
found between acute toxicity data and the patterns of the in vitro enzyme
inhibition.
Whatever the mechanism of genetic resistance, such resistance can alter
the toxicity of toxaphene to a given species of fish by a factor of up to
several hundred. Recalling the mere 16-fold difference in the 96-hour LC50
estimates for the various species of fish covered in Table 5.10, it seems
apparent that preexposure conditioning, rather than the phylogenetic position
of a given fish species, will largely determine the ability of a fish popula-
tion in the environment to tolerate acutely toxic doses of toxaphene.
5.2.2.2.5 Toxaphene field studies — Toxaphene once enjoyed wide usage as a
piscicide in rough fish control programs. Such programs usually involved complete
5-51
-------
elimination of fish in a given lake. After a suitable period of detoxication,
the lake was then restocked with the desired species of game fish. Such programs,
which have been conducted since 1948, have been exhaustively reviewed by Hughes
(1970). In terms of assessing the potential adverse effects of toxaphene in
the environment, such studies must be interpreted with caution. Perhaps the
greatest problem is determining the precise extent of fish exposure to toxaphene.
Table 5.16, which summarizes several studies on toxaphene use as a fish poison,
expresses toxaphene exposure as a theoretical dose. This is defined as the
concentration of toxaphene which would be present in the water if the total
amount of toxaphene added to the water mixed evenly and remained stable. This
does not happen during the actual application of toxaphene. Kallman and co-
workers (1962) found radical differences in the concentration of toxaphene in
treated lake water for periods of up to 73 days after initial application.
Toxaphene concentrations were usually higher on the leeward side of the lake
and also appeared to be greater in deeper water. Further, toxaphene concentra-
tion never reached the theoretical maximum of 50 ppb, remaining below 10 ppb
for most measurements. Similar failures to reach the theoretical dose have been
noted by Hughes and Lee (1973), who partially attributed this to the water in-
solubility of toxaphene. Additional factors which might cause the rapid elimina-
tion of toxaphene from lake water are discussed in Section 7.3.2.
While these studies cannot be used to quantitatively refine estimates of
toxaphene fish toxicity, they are eminently useful in indicating what does not
happen under actual conditions of environmental exposure. As described in
Section 5.2.2.2.3, toxaphene has been shown to have a marked effect on egg
viability in fish at a concentration of 0.075 ppb. This information would cer-
tainly seem to suggest that toxaphene, at very low levels, would profoundly
5-52
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Table 5.16. Use of toxaphene as a fish control agent
Ol
I
Ln
U)
Location
North Dakota
Number
of lakes
treated
16
Characteristics
of the lakes
2.6-370 ha, maxi-
mum depth of 2.4-
Theoretical
concentration
5-35 ppb
Fish mortality
data
Incomplete to complete
fish kills
Detoxication data
5 of 7 lakes detoxified
after 7 months
Reference
llenegar,
Warnick
1966 and
, 1966
Wisconsin 4
Wisconsin 8
New Mexico 1
British Columbia 7
Oregon 2
Montana 1
8 meters
pH 3.2-9.9, hardness 51-
2,385
5.3 - 1063 ha, 100 - 420 ppb
maximum depth 5-9.8 meters
pH 8.2-8.6, alk. 104-128 ppm
24-110 ha, depth 100 ppb
3.5-8 meters
pH 8.0-9.2
32 ha, depth 5.7 meters 50 ppb
pH 8.5, alkalinity 114 ppm
4-55 ha, depth 3.3-
16 metere
pH 7.9-8.4
229 ha, 44 meters deep
1299 ha, 6 meters deep
5.2 ha, depth 4 meters
pH 7.4-8.6
10 - 100 ppb
40 ppb
88 ppb
78 ppb
Smaller fish more suscep- No data available on other
tible than larger fish lakes
Minimum dose for complete
kill, 20 ppb
No correlations of toxicity
to water chemistry
Complete kills of bullhead Successful restocking after Hughes and Lee, 1973 and
and stunted bluegill
populations
Substantial fish kill,
112 kg/ha
Complete fish kills
Complete fish kills
Complete fish kills
Smaller fish more
susceptible than
larger fish
Hughes, 1970
Johnson et al., 1966
7-10 months
Toxaphene in water at
1-4 ppb for up to
nine years after treat-
ment
Successful restocking after Kallman et al., 1962
12 months
Toxaphene in water at less
than 1 ppb by 523 days
after treatment
Lakes still toxic to fish
after 9 months
Stringer aad McMynn,
1958
Remained toxic for 6 years Terriere ec al., 1966
Remained toxic for 1 year
Remained toxic for 4 months Wollitz, 1963
-------
limit the development of stable fish populations in the environment. Based on
the studies summarized in Table 5.16, such effects are not seen in toxaphene-
treated lakes. After varying periods, most of these studies specify that the
treated lakes were detoxified and were able to support productive and healthy
fish populations. Both Hughes and Lee (1973) and Kallman and coworkers (1962)
indicated that this happened in water containing 1 ppb toxaphene. The failure
of toxaphene to affect fish productivity under such circumstances could be
attributable to many factors. The most obvious of these is that the composition of
toxaphene monitored in water after prolonged exposure periods is different from
that of the originally applied commercial toxaphene. Hughes (1970) has reported
that toxaphene extracted from treated lake water is less toxic to fish than
commercial toxaphene. Thus, based on these fish control studies and other in-
formation given in Section 7.3.2, it may be reasonable to conclude that the toxic
components in toxaphene that are responsible for the adverse effects on fish re-
production may be selectively removed from aquatic systems.
5.2.2.2.6 Behavioral effects— Subtle effects on the behavioral patterns -
e.g., feeding activity or predator avoidance - could substantially affect the
ability of an organism to survive under normal environmental circumstances.
Such affects, however, are difficult to document under controlled conditions.
As previously discussed (Sections 5.2.2.2.1 and 5.2.2.2.2), toxaphene seems to
cause hyperactivity in fish on both acute and chronic exposure. Using a highly
complex experimental design, Warner and coworkers (1966) attempted to define
less obvious behavioral changes. In this experiment, goldfish were continuously
exposed to toxaphene at concentrations of 0.44 ppb and 1.8 ppb for 96 hours.
The fish were kept in an exposure chamber designed both to detect and record
5-54
-------
the response of these fish to various external stimuli - e.g., light and shock.
Comparing the behavior of exposed fish to controls, these investigators concluded
that toxaphene causes "behavioral pathology of considerable severity." The nature
of this pathology and the possible effects that it might have on goldfish under
normal environmental stress are not described.
Kynard (1974) has demonstrated that both susceptible and resistant mosquito
fish avoid toxaphene concentrations of 250 ppb and above.
5.2.2.2.7 Drug interactions — There is no evidence that toxaphene has a
synergistic or antagonistic effect with other pesticides in its effect on fish.
Macek (1975) found that toxaphene/parathion and toxaphene/DDT combinations had
only additive combined action in their acute toxicity to bluegills. Similar
results have been reported for toxaphene/methyl parathion, toxaphene/DDT, and
toxaphene/endrin combinations using mosquito fish (Ferguson and Bingham, 1966).
5.2.2.2.8 Effects on amphibians — Very little attention is given to this group
of animals by aquatic toxicologists, and this lack of concern is evidenced in
the toxaphene literature. Nonetheless, based on the limited available data,
these animals seem markedly less susceptible than fish to toxaphene. Sanders
(1970) determined the acute toxicity of toxaphene to western chorus frog tad-
poles (Pseudacris triseriatu) and one week old Fowler's toads (Bufo woodhousii).
For the toad, 24, 48 and 96-hour LCSO's with corresponding 95 percent confi-
dence intervals were 600 (300 to 1200) ppb, 290 (200 to 420) ppb, and 140
(60 to 350) ppb, respectively. Even higher values were found with the tadpole;
1700 (500 to 3200) ppb, 700 (400 to 1200) ppb, and 500 (100 to 1100) ppb. The
symptoms of toxaphene poisoning, however, were similar to those seen in fish
and included initial excitability, loss of equilibrium, and death. Also similar
5-55
-------
to fish, toxaphene was among the more toxic pesticides to the toad, being exceeded
only by endrin and trifluralin. Mulla (1962, 1963) recommended the use of toxa-
phene for frog and toad control. Although giving no LC50 estimate, Mulla (1963)
indicated that toxaphene causes complete eradication of three species of frogs
when applied at levels of about 1.2 kg/ha.
As with fish, Ferguson and Gilbert (1967) noted resistant or tolerant
populations of frogs and toads, with the pattern of resistance/tolerance apparently
related to preexposur'e to pesticide runoffs. In these animals, 36-hour LC50
values were extraordinarily high, ranging from 50 ppm to 500 ppm. No attempt
was made to demonstrate a genetic basis to this lack of susceptibility. However,
potential genetic resistance to DDT has been described in frogs (Boyd et al.,
1963) and, given the pattern seen in fish, could also apply to toxaphene.
5-56
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5.3 AQUATIC INVERTEBRATES
5.3.1 Metabolism
Information on the absorption, transport, distribution, biotransfortnation,
and elimination of toxaphene in marine and freshwater invertebrates is not as
detailed as in vertebrate species (Section 6.1). Butler et al. (1962) have
demonstrated a failure of molluscs, oysters in particular, to continue the
normal deposition of their shell when exposed continuously to 0.1 ppm, with dif-
ferences detectable as early as 24 hours.
Metcalf (1955) suggests that, unlike cyclodiene insecticides, toxaphene
is distributed throughout the invertebrate animal's body by the hemo-
lymph and has no effect on the nervous system. Wagner (1974) states that in
those aquatic organisms which are especially sensitive to this class of compounds,
the uptake of oxygen through the gills is disrupted, and death is associated with
suffocation rather than nervous disorders. In the marine environment the dehydro-
chlorination of this chemical under alkaline conditions may enhance its toxicity.
To date no studies in aquatic invertebrates have been found that utilized
Cl-36 or C-14 labelled toxaphene. Any conclusions about sites of absorption,
transport, and distribution within the organism and biotransformation of the
compound are hard to predict without such studies. Similarities in many metabolic
pathways and fate of the toxaphene may be common in many organisms and similar
to those described in Section 6.1. However, unique species-specific metabolic
differences are likely to exist.
Oysters exposed to 1 ppb toxaphene for 36 weeks bioaccumulated the compound
throughout the exposure period until spawning, at which time there was a noticeable
drop in tissue levels of toxaphene. Likewise, when oysters were exposed to a
5-57
-------
combination of DDT, toxaphene, and parathion, at 1 ppb each, there was bioac-
cumulation throughout the exposure period accompanied by the drop during spawning.
Between four and twelve weeks after the removal of these oysters from the toxicants,
they had eliminated the toxaphene from their tissues (Lowe et al., 1971). Table 5.17
characterizes these data on tissue accumulation. The drop in toxaphene levels
following spawning indirectly suggests that the oyster can rid itself of a sig-
nificant portion of its body load of toxicants in this manner.
Schoettger and Olive (1961) have reported that Daphnia magna and I), pulex
accumulate sub-lethal doses of toxaphene. When these cladocerans serve as the
food source for the kokanee salmon, Orcorhynchus nerka kennerlyi, and shiners,
Notropis sp., mortality is high in the fish (Table 5.18). This study suggests
that toxaphene is stored unchanged or bound in the Daphnia in a form which remains
toxic to other members of the food chain. It was not reported by Schoettger and
Olive (1961) whether these Daphnia irreversibly bound the toxaphene, or if it might
be lost when the organisms were returned to a toxaphene-free environment. The
levels that are stated by Schoettger and Olive (1961) as sublethal to Daphnia
and the accumulated dosages imply that the Daphnia and water in which they are
living have equivalent levels of toxaphene. If they are in equilibrium with
the toxaphene as suggested, then their introduction into the fish aquaria would
be expected to leech out the toxaphene stored by the Daphnia. The mortality
in the fish could then be the result of an interaction of toxaphene levels
present in their food and in the aquaria water. Results of these experi-
ments indicate that single sublethal dosages of toxaphene are inadequate to
produce accumulations in fish-food organisms which cause mortalities among the
test fish (Table 5.18). Multiple sublethal doses permitted sufficient amounts
5-58
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Table 5.17. Accumulation of toxaphene by oysters exposed to 1 ppb
singly or in combination with 1 ppb DDT and 1 ppb parathiona
Week
12
24
36b
40
44
48
Pesticide residue (ppm of wet tissue weight)
Single exposure Combination exposure
20.0 1.0
23.0 30.0
8.0 9.0
3.0
NDC
ND
/Source: Modified from Lowe et al., 1971.
End of pesticide exposure.
Not detectable, less than 0.010 ppm.
5-59
-------
Table 5.18. Effects of toxaphene exposed Daphnia on fish
Ul
o
Trial
no.
1
2
3
4
5
6
7
8
9
10
11
12
Dosage
(ppm)
0.03
0.03
0.03
0.03
0.03
0.03
• 0.03
0.03
0.03
0 01 /
', 0D
0.13
0. 01
0.1
0.02
0.1
Exposure
period
(hr)
36
72
96
192
216
288
312
408
624
312
240
120
Species
of
fish
0 . ne rka
kennerlyi
0. nerka
kennerlyi
0. nerka
kennerlyi
0. nerka
kennerlyi
0. nerka
kennerlyi
0. nerka
kennerlyi
0. nerka
kennerlyi
0. nerka
kennerlyi
0. nerka
kennerlyi
0. nerka
kennerlyi
NoCropis
sp.
Notropis
sp.
No.
of
fish
4
4
4
4
8
4
8
4
4
8
I?
12
Test fish
no. of fish
surviving
after (hr)
24 48 72 96 120 144
4444^
44444
4444-
44444 4
77766 5
44444 4
8888-
44444 4
44444 4
00000
94322
10 i 5 4 3
Control fish
no. of fish
surviving
after (hr)
96 120 144
44-
44-
4 - -
444
887
444
8 - -
444
444
88-
12 12
12 11
.Source: Modified from Schoettger and Olive, 1961.
Total dosage of multiple, sublethal dosages.
-------
of toxaphene to accumulate in Daphnia to cause a significant increase in mortality
in kokanee salmon and in shiners. It might be concluded that sublethal, acute
dosages are detoxified before they accumulate. However, sublethal, chronic dosages
may exceed the detoxification rate and permit toxaphene to accumulate in the
Daphnia.
Toxaphene residues were obtained from plankton in Big Bear Lake, California,
following two toxaphene treatments of 0.03 and 0.10 ppm two weeks apart (Table 5.19).
Bioaccumulation and biomagnification of toxaphene occurred for 114 days post-
application. Values present in the plankton at this time were sometimes greater
than the original application. At 265 days the values in the plankton were
still greater than 1.5 times the original application (Johnson, 1966). The
presence of this level of toxaphene in the plankton was toxic to the fish,
and produced toxaphene levels in the fish that were considered unsafe for ten
months post-application.
Naqvi (1973) states that tubificid worms (EL sowerbyi) are capable of
accumulating toxaphene in large quantities in nature, and that worms collected in
the Mississippi River Delta contained large amounts which he was unable to quanti-
tate.
Bioconcentration in oysters (Lowe et al., 1971) and in plankton (Johnson,
1966) are documented. Bioconcentration is indicated in Daphnia (Schoettger and
Olive, 1961). After the pesticide has entered the organism, it may be stored as
residues, either unchanged for long periods or decomposition may take place in
the animal body, with storage or excretion of the breakdown products. The fate
of the pesticide and the capacity of aquatic animals to remove, store, or metabolize
it may be related to the previous pesticide burden (Section 5.3.2.3), and the po-
tential for food chain amplification at higher trophic levels exists.
5-61
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Table 5.19. Toxaphene residue in plankton from Big Bear Lake exposed
to 0.03 and 0.10 ppm treatments two weeks aparta
Date
Oct. 4,
Oct. 19,
Jan. 20,
Jan. 20,
Mar. 7,
Apr. 10,
Apr. 10,
Apr. 20,
May 11,
June 13,
June 20,
June 20,
June 26,
June 26,
June 26,
June 26,
June 27,
June 27,
June 2 7 ,
Toxaphene residue
(ppm)
1960
1960
1961
1961
1961
1961
1961
1961
1961
1961
1961
1961
1961
1961
1961
1961
1961
1961
1961
1.
50.
73.
97.
50.
20.
30.
16.
17.
19.
10.
12.
12.
0.
19.
4
0
0
0
0
0
0
0
0
0
0
0
0
0
0
trace
2.
2.
2.
0
0
0
Days since last
treatment
5
21
114
114
160
194
194
204
225
251
258
258
264
264
264
264
265
265
265
Source: Modified from Johnson, 1966.
5-62
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5.3.2 Effects
5.3.2.1 Physiological or Biochemical — With expanding use of chemical pesticides
it has become vital to understand their activity and eventual fate in the natural
environment. By and large, we are aware of the initial animals or plants that
these chemicals control or kill. However, our knowledge of their secondary
« effects is limited. It is probable that many of these substances and, possibly,
their more toxic oxidation products, find their way into our watersheds, estuaries,
and coastal waters. The coastal waters are important to us commercially as a home
for many of our economically important marine species. Fairly extensive research
/
is now available to evaluate the effects of chemicals on non-target freshwater
and marine organisms.
Toxaphene at very high concentrations (greater than 1 ppm) is required to block
the development of clam eggs, but levels of less than 0.25 ppm kill 67% of the
larvae (Section 5.3.2.2.4). It is well-established that early embryogenesis is under
the control of m-RNA manufactured in the ovary prior to the completion of oogenesis.
These m-RNA compounds (or masked messengers, as they are called) direct the early
development of the organism. Only following gastrulation are the maternal and
paternal chromosomes of the new individual required to manufacture their own m-RNA.
Failure of the larvae to continue development could be the result of the lack
of DNA-directed RNA production and hence a loss in RNA-directed enzyme and protein
synthesis. This in no way answers the question of where toxaphene is bound
in the cell or what its specific activity within the cell is. However, one
could suggest that the above effects might be the result of (1) an interaction
with enzymes or proteins that causes a change in normal activity; (2) an
interaction at the genetic level preventing genetic transcription, altering
genetic transcription, or even causing a mutation; (3) an interaction of toxaphene
with cellular or subcellular membranes that might mimic the above two alternatives.
5-63
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Albaugh (1972) has described the symptoms of the acute toxicity response
in the crayfish as hyperactivity, loss of equilibrium and coordination, paral-
ysis, and death. Death occurred 1 to 24 hours after the symptoms were first
observed. Those animals surviving at 48 hours completely recovered when
transferred to clean water, even though many had displayed advanced symptoms
of poisoning. These physiological responses of the organism to toxaphene seem
to indicate a response that is similar to that described in higher animals
(Section 6.2.2.1.1.1). These responses are thought to be of neural origin.
This would seem to conflict with the information of both Metcalf (1955) and
Wagner (1974); however, no specifics were given in the last two instances,
only generalities without supporting experimental data. It seems cogent to
surmise that toxaphene may have a neurotoxic effect in aquatic invertebrates.
5.3.2.2.1 Sublethal toxicity— Oysters, Crassostera virginica, were exposed
for four weeks to a 0.1 ppm concentration of toxaphene (Butler et al., 1962).
Initially, they measured both shell movements employing a kymograph and growth
of new shell in yearling oysters. It became evident to Butler et al. that
measurement of shell growth was their most critical bioassay. The analysis of
the activity records of the large oysters and growth of the small oysters is
summarized in Figure 5.5. No growth was evident in the experimental group
after the second week. The harmful effects of toxaphene were also clearly
demonstrated by the 70 per cent decrease in activity of experimental oysters,
as compared to controls, at the end of the fourth week of the experiment.
Sublethal concentrations were calculated in a study by Schoettger and
Olive (1961) for the purpose of feeding these exposed animals to another com-
ponent of the food chain. Sublethal concentrations were as follows: for
Daphnia pulex, 0.03 ppm after 168 hours; for Daphnia magna, 0.03 ppm after 120
hours; and for damselfly nymphs, 0.004 ppm after 96 hours.
5-64
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100
80-
g 40H
o
20-
0
a
Control (C)
Experimental (E)
4 5
WEEKS
Figure 5.5. Activity of large oysters and growth of small oysters exposed
to 0.1 ppm toxaphene. Source: Butler et al., 1962.
5-65
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Needham (1966) determined the levels of toxaphene which numerous freshwater
aquatic invertebrates would tolerate for 24 hours without any mortality. The
highest concentration was 1 ppm for leeches (Hirudinea), while the least resistant
organism was a Coleopteran (Haliplus) which would tolerate only 10 ppb. Table 5.20
gives the data for the 14 genera tested. The nine orders of invertebrates tested
are listed commencing with the most tolerant order and concluding with the least
tolerant: Hirudinea, Hydracarina, Gastropoda, Trichoptera, Ononata, Hemiptera,
Ephemeroptera, Amphipoda, and Coleoptera.
5.3.2.2.2 Acute toxicity— Khattat and Farley (1976) exposed the estuarine copepod
Acartia tonsa Dana to toxaphene at concentrations ranging from 0.03 to 300 ppt
(nanograms/liter). They calculated an LC50 of 7.20 ppt (Table 5.21).
In an extensive study by Courtenay and Roberts (1973), the toxic effects of
toxaphene on Sesarma cinereum (drift line crab), Penaeus duorarum (pink shrimp),
and Rhithropanepeus harrisii (mud crab) were examined. The 96-hour LC50 for
stage I zoea in ^. cinereum was 0.054 ppb. The LC50 increased about tenfold to
0.076 ppb for stage II zoea and to 0.74 ppb for stage III zoea. The LC50 again
increased about tenfold to 6.8 ppb for stage IV zoea and to 8.4 ppb for the megalopa.
Courtenay and Roberts (1973) also studied the synergistic effects of temperature,
salinity, and oxygen saturation on the expression of the toxaphene toxicity. In
general, higher temperatures, higher salinities, and lower oxygen saturation in-
creased the toxicity of toxaphene to one or more of the zoeal or megalopa stages
evaluated (Table 5.20).
The 96 hour LC50 for P_. durorarum decreased from 2.2 ppb for nauplii to
1.8 ppb for protozoeae to 1.4 ppb for mysis. In this study there was a signifi-
cant interaction at every stage in the temperature-toxaphene tests. A significant
interaction of salinity-toxaphene was observed only in the mysis stage. The nauplius
stage showed significant mortality only when toxaphene was present. Decreased
oxygen levels had no effect on the toxaphene toxicity (Table 5.20).
5-66
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Table 5.20. Acute toxicity of toxaphene to aquatic invertebrates
Ol
I
cr>
Reference Organism Concentration
Khattat and Acartia tonsa 300.0 ppt
Farley, 1976 Dana 30.0
3.0
0.3
0.03
Courtenay and Sesarma cinereum 0.05 ppb (25 C)
Roberts, 1973 Stage I - zoeal (35 C)
(25°/oo salinity)
(34 1 oo salinity)
(75 /o oxygen
saturation)
(100 /o oxygen
saturation)
Stage II - zoeal 0.5 ppb (25 C)
(35 C)
(30Q/oo salinity)
(40 /oo salinity)
(75 /o oxygen
saturation)
(100 /oo oxygen
saturation)
Stage III - zoeal 0.5 ppb (25 C)
(35 C)
(30°/oo salinity)
(40 /oo salinity)
(75 /o pxygen
saturation)
(100/Q oxygen
saturation)
Percent
mortality
77.5 (96 hr)
67.5
35.0
27.5
30.0
0.0
62.5
17.5
100.0
5.0
0.0
15.0
70.0
20.0
40.0
87.5
22.5
2.5
67.8
22.5
45.0
32.5
10.0
Control
mortality
5
5
5
5
5
0.0
10.0
10.0
27.5
0.0
0.0
2.5
22.5
7.5
0.0
20.0
2.5
2.5
40.0
22.5
32.5
27.5
22.5
-------
Table 5.20. (continued)
CTv
OO
Reference Organism Concentration
Courtenay and Sesarma cinereum 5.0 ppb (25 C)
Roberts, 1973 Stage IV - zoeal (35 C)
(30°/0o salinity)
(45°/0o salinity)
(75 /o oxygen
saturation)
(100 /o oxygen
saturation)
Megalopa 5..0 ppb (25 C)
(35oC)
(30 /oo salinity)
(50°/oo salinity)
(75 /o oxygen
saturation)
(100 /o oxygen
saturation)
Penaeus duorarum 1.0 ppb (25 C)
nauplius (35 C)
(35°/oo salinity)
(45°/oo salinity)
(75. /o oxygen
saturation)
(100 /o oxygen
saturation)
Percent
mortality
27.5
80.0
40.0
85.0
100.0
32.5
2.5
10.0
17.5
77.5
5.0
15.0
20.0
100.0
17.5
15.0
2.5
5.0
Control
mortality
35.0
47.5
27.5
37.5
32.5
22.5
17.5
17.5
10.0
40.0
7.5
25.0
10.0
32.5
0.0
0.0
0.0
0.0
-------
Table 5.20. (continued)
Ln
I
vo
Reference
Courtenay and
Roberts, 1973
Courtenay and
Roberts, 1973
Naqvi and
Ferguson, 1968
Organism
Penaeus dourarum
protozoea
mysis
Rhithropanopeus
harisii
Stage I - zoeal
Six species of
freshwater
cyclopoids
Percent Control
Concentration mortality mortality
1.0 ppb
1.0 ppb
10 ppb
20
30
40
50
15 ppb
25
35
45
(25 C) 27.5
(35 C) 97.5
(35°/oo salinity) 22.5
(45°/oo salinity) 40.0
(75 /o oxygen 87.5
saturation)
(100°/o oxygen 15.0
saturation)
(25 C) 30.0
(35 C) 90.0
(35°/oo salinity) 15.0
(50°/oo salinity) 67.5
(75 /o oxygen 12.5
saturation)
(100°/o oxygen 15.0
saturation)
10.0
10.0
7.0
32.0
75.0
0.0")
8.5
30.1
28.3.,
5.0
37.5
22.5
25.0
32.5
10.0
15.0
25.0
17.5
32.5
12.5
15.0
history 58.0 ^
, of pesti- 56.0 L
cide 64.7 (
contamination 84. oj
little
prior
pesticide
contaminatior
-------
Table 5.20 (continued)
Reference Organism
Concentration
Percent
mortality
Control
mortality
Naqvi and Eupera singleyi
Ferguson, 1968
Physa gyrina
Ol
I
Harris and
Jones, 1962
Needham, 1966
Culicoides
cariipennis
larvae
Hirudinea
Gammarus sp.
Hydracarina
Callibaetis sp.
100 ppb
200
300
400
500
600
700
350 ppb
400
450
500
550
88 ppb
412
1 ppm
100 ppb
200
300
500
1 ppm
150 ppb
300
400
500
0.0
0.0
15.0
50.0
50.0
60.0
40.0,
20.0'
60.0(
35.0
55.0
100.0^
50.0
90.0
0.0
0.0
61.0
79.0
100.0
0.0
0.0
29.0
87.0
100.0
history
of pesti-
cide
contamination
history
of pesti-
cide
contamination
80.0
75.0
100.0
100.0
100.0
9.0
9.0
9.0
9.0
0.0
0.0
0.0
0.0
0.0
little
prior
pesticide
contamination
little
prior
pesticide
contamination
-------
Table 5.20 (continued)
Ul
I
Reference Organism Concentration
Needham, 1966 Aeschna sp. 200 ppb
275
350
450
Lestes sp. 450 ppb
500
600
850
Notonecta sp. 275 ppb
300
400
600
Sigara sp. 50 ppb
75
100
150
Limephilus sp. 500 ppb
550
600
650
Haliplus sp. 10 ppb
40
50
75
Percent
mortality
0.0
16.0
60.0
100.0
0.0
19.0
67.0
100.0
0.0
29.0
61.0
100.0
0.0
40.0
75.0
100.0
0.0
51.0
80.0
100.0
0.0
55.0
89.0
100.0
Control
mortality
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
-------
Table 5.20 (continued)
Un
I
N>
Reference Organism Concentration
Needham, 1966 Hydroporus sp. 60 ppb
100
300
450
Dytiscus sp. 15 ppb
50
60
.75
Gyrinus sp. 65 ppb
100
150
185
Percent
mortality
0.0
27.0
65.0
100.0
0.0
24.0
42.0
100.0
0.0
22.0
40.0
100.0
Control
mortality
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
Lymnaea sp.
700
0.0
0.0
-------
Table 5.21. LC50 values for aquatic invertebrates exposed to toxaphene
Reference Organism LC50
Khattat and Acartia tonsa Dana 7.2 ppt (96 hr)
Farley, 1976
Courtenay and Sesarma cinereum
Roberts, 1973 Stage I - zoeal 0.054 ppb (96 hr)
Stage II - zoeal 0.760
Stage III - zoeal 0.740
Stage IV - zoeal 6.8
Megalopa 8.4
Penaeus duorarum
nauplius 2.2 ppb (96 hr)
protozoea 1.8
mysis 1.4
Rhithropanopeus harrisii
Stage I - zoeal 43.75 ppb (96 hr)
Stage II - zoeal 288-345
Stage III - zoeal 290-430
Chaiyarach et al., Palaemonetes kadiakensis 0.091 ppm (24 hr)
1975 0.068 (48 hr)
0.046 (72 hr)
0.036 (96 hr)
Procambarus simulans 0.45 (24 hr)
0.29 (48 hr)
0.24 (72 hr)
0.21 (96 hr)
Rangia cuneata 940 (24 hr)
699 (48 hr)
488 (72 hr)
460 (96 hr)
Hooper and Ephemera simulans 9.5 ppm (24 hr)
Grzenda, 1957 nymphs
Daphnia magna 1.5
Gammarus fasciatus 0.06
Asellus intermedius 0.10
Naqvi and Palaemonetes kadiakensis
Ferguson, 1970 Bluff Lake - no history 20.9 ppb (24 hr)
of exposure
Hollandale'l prior history 229.0
Belzoni f of insecticide 89.0
Sky Lake J exposure 44.0
5-73
-------
Table 5.21 (continued)
Reference
McDonald, 1962
Albaugh, 1972
Ferguson et al., 1965b
Organism LC50
Gammarus lacustris lacustris 1.0 ppm
0.5
0.1
0.05
Procambarus acutus 60.7 ppb
90.2
Palaemonetes kadiakensis 57.5
170.0
(72 + 10 min)
(96 + 11 min)
(360 + 40 min
(460 + 80 min
(48 hr-clean
(48 hr-cotton
area)
(36 hr-clean
(36 hr-cotton
Sanders and Cope, 1966
Naqvi, 1973
Sanders and Cope, 1968
Pigatti and Mello, 1961
Harris and Jones, 1962
Guyer et al., 1971
Simocephalus serrulatus
Daphnia pulex
Branchiura sowerbyi
Pteronarcys California
naiads
Pteronarcello badia naiads
Claassenia sabulosa- naiads
Culex pipens fatigans
larvae
Culicoides variipennis
larvae
Crassostera Virginia
Peneus aztecus
Peneus duorarum
Palemonetes kadiakensis
Palemonetes
pregio
Palemonetes maciodactylus
Daphnia pulex
Daphnia serrulatus
Gammarus fasciatus
Gammarus lacustris
area)
19.0 ppb (48 hr-60 F)
10.0 (48 hr-7C F)
15.0
(48 hr)
1.0 ppm (72 hr LC100)
18.0 ppm (24 hr)
7.0 (48 hr)
2.3 (96 hr)
9.2 ppm (24 hr)
5.6 (48 hr)
3.0 (96 hr)
6.0
3.2
1.3
14 ppm
(24 hr)
(48 hr)
(96 hr)
0.09 ppm (24 hr)
0.02 ppm (96 hr)
0.0027 (48 hr)
0.0042
0.006
0.0052
0.037
0.015
0.01
0.022
0.07
5-74
-------
Table 5.21 (continued)
Reference Organism LC50
Guyer et al., 1971 Pteronareys California 0.007 ppm (48 hr)
Clossonia sabulosa 0.0032
Pteronarcella bodia 0.0056
Ischmura verticalis 0.086
5-75
-------
The LC50 for stage I of R.. harrisii was determined to be 43.75 ppb. Values
for stages II and III ranged from 288 to 430 ppb (Table 5.21). Additionally,
Courtenay and Roberts (1973) histologically evaluated JL cinereum, R.. harrisii,
and Callinectes sapidus (blue crab). In stage I of S_. cinereum, the hepatopancreas
had vacuolated granulated cells coupled with contraction of the intestine in one
of the four specimens examined. In stage II, 20% of the animals exhibited no
j-
effects, 50% had the vacuolated granulated hepatopancreas cells, and 30% had
necrotic spaces (undefined by the authors). Forty percent had intestinal con-
traction, 10% had a slight dilation, and 50% had no other apparent intestinal
anomaly. In stage III all animals examined showed extensive hepatopancreas
degeneration and 80% exhibited the contraction of the intestine.
Stage I C_. sapidus larvae exhibited no histological changes at toxaphene
concentrations of 0.0005 or 0.00001 ppb. However, 50% of the stage III larvae
exposed to 0.01 ppb showed hepatopancreas changes similiar to those found in
j^. cinereum.
In stage I R.. harrisii, one of the three specimens examined, had necrosis
of the intestine and sloughing of the stomach epithelium. Stages II and IV
showed damage to the nervous tissue in the form of irregular spaces between
the fibers.
The authors stated that in all three species deposition of an orange-brown
pigment has been found in the body spaces. The pigment deposition was especially
heavy along the intestine and attempts to dissolve the pigment in water, alcohol,
or xylene failed.
In a study by Chaiyarach et al. (1975), three species of aquatic inverte-
brates were exposed to acute levels of toxaphene. The organisms were Palaemonetes
kadiakensis (grass shrimp), whose LC50 was 0.091 at 24 hours and 0.036 ppm at
5-76
-------
96 hours; Procambarus simulans (crayfish), whose LC50 was 0.45 ppm at 24 hours
and 0.21 ppm at 96 hours; and Rangia cuneata (mactrid clam), whose LC50 was
940 ppm at 24 hours and 460 ppm at 96 hours (Table 5.21). It is interesting
to note that Rangia is the most tolerant species tested, and that it is also of
commercial value as the meat is eaten by man in part of its distribution range.
In 1954 eight lakes in British Columbia were treated with "Fish Tox" -
a rotenone based chemical containing 10% toxaphene (Stringer and McMynn, 1958).
Two lakes each were treated with 0.01, 0.03, 0.07, and 0.10 ppm. An evaluation
of the effects of toxaphene on the bottom fauna was carried out with 30 pre-
poisoning and 70 post-poisoning dredgings taken from each lake. Qualitative
results are summarized in Table 5.22. Shrimp (Gammarus or Hyallela) were found
in six of the eight lakes prior to treatment, but were absent in subsequent
sampling. Midge larvae (chironomidae) were present in all lakes initially and
at one month post-poisoning. However, at nine months post-poisoning, the midge
larvae were absent in the two lakes treated at the 0.10 ppm level. Aquatic earth-
worms (Qligochaeta) were present in all lakes, both before and after treatment.
Leeches (Hirundinea), although not present in all lakes at nine months post-
treatment, were present in the two 1.0 ppm lakes, indicating that these organisms
were not affected by the levels of toxaphene employed in this experiment. Mayfly
nymphs (Ephemeroptera) were present in six of the lakes prior to treatment. The
authors stated that subsequent samples contained no nymphs except for Taylor
Lake (0.01 ppm). However, their data indicated (Table 5.23) that mayfly nymphs
were present in Spectacle Lake (0.07 ppm) at all three sampling periods. Dragonfly
and damselfly nymphs (Odonata) were found in most lakes initially and appear to
5-77
-------
Table 5.22. Occurrence of bottom organisms in toxaphene treated lakes
before and after poisoning
Lake
„ , Alleyne Taylor Gladstone
Organism /A , .
b
A BCABCABC
Shrimp 0 --C4--0--
Midge Larvae 4 44444444
Aquatic Earth- 4 44444444
worms
Leeches - _____ + + _
u> Mayfly Numphs - --44-4--
oo Dragonfly and 4 --444444
Damself ly
Nymphs
Freshwater 4 44444444
Snails
Round
A B
4 0
4 0
4 0
0
4 0
4 0
4 0
Gallagher Spectacle Summit
m) (0.07 ppm) (0.07 ppm) (0.10 ppm)
CABCABCABC
_ + __ + __ + --
444444444-
4444444444
----4-4444
-4--4444--
_ + __ + + + + _-
-44444444-
Lady King
(0.10 ppm)
A
4
4
4
-
-
-
4
B C
-
4
4 4
4
-
_
4 4
a
/Source: Modified from Stringer and McMynn, 1958.
A - before treatment; B - one month after treatment; C - nine months after treatment.
4 indicates organism is present; - indicates organism is not present; 0 indicates that lake was not
sampled.
-------
Table 5.23. Plankton samples taken before and after toxaphene
treatment
Lake
0 . Alleyne Taylor Gladstone Round Gallagher Spectacle Summit Lady King
m (0.01 ppm) (0.01 ppm) (0.03 ppm) (0.03 ppm) (0.07 ppm) (0.07 ppm) (0.10 ppm) (0.10 ppm)
A BCABCABCABCABCABCABCABC
Cladocera:
Daphnia - + + + + + + - + + + - + + - + + + + - - + + +
Bosmina + + + + + + + + + + + + + + + + - + + + + + + +
Copepoda:
Cyclops + + + + + - + + - + + + + + - + - + + + - + + +
Diaptomus + 4-_______-j-__-(-4._ + _____4._4-
^ Nauplius larvae+ + + + + - + + + -- + + + + + - + + -- + + +
i
^ Rotifera + + + + + + + + + + + + + + + + + + + + + + + +
Flagellates + + + + + + + + + + + + + + + + + + + + + + + +
Diatoms + + + + + + + + + + + + + + + + + - + + + + + +
/Source: Modified from Stringer and McMynn, 1958.
A - before treatment; B - one month after treatment; C - nine months after treatment.
+ indicates organism is present; - indicates organism is not present.
-------
Table 5.24. Invertebrates collected before and after treatment with 0.1 ppm toxaphene
Location and
date of sampling
Horseshoe Lake
Sept. 20
(1 day before
treatment)
Oct. 15
April 29
^ July 29
i
o McCarty Lake
Aug. 3
(1 day before
treatment)
Aug. 24
May 25
June 23
Average number of animals per square meter
Diptera Ondonata
989.9 68.9
167.9
4.3
36.6 116.2
4605.3
6.5
-
9856.2
Ephemeroptera Oligochaeta Mollusca Others Total
516.5 5.4 19.4 32.3 1678.6
43.0 51.6 51.6 4.3 314.1
12.9 8.6 - 43.0
116.2 68.9 - 335.7
12.9 - - 4.3 464.8
6.5 6.5 - 20.4
47.3 - - 47.3
172.2 10,028.3
a.
Source: Modified from Hooper and Grzenda, 1957.
-------
be eliminated at about the 0.07 ppm level as they were absent in one lake at
this level and in both 1.0 ppm lakes. Freshwater snails (Gastropoda) were taken
in most dredgings and appear to be unaffected by these dosages of toxaphene.
In order to determine the effect of the various toxaphene concentrations
on the planktonic groups, Stringer and McMynn (1968) qualitatively sampled the
eight lakes, of which two each were poisoned with 0.01, 0.03, 0.07, and 0.10 ppm.
The results are summarized in Table 5.23. Although there is some fluctuation
in the planktonic groups evaluated, it appears that toxaphene had no effect
at the levels employed in this study.
When Hooper and Grzenda (1957) studied two lakes in Michigan (Horseshoe and
McCarty Lakes) that were treated with 0.1 ppm toxaphene, they found a large portion
of the benthos was killed in a month (Table 5.24). Horseshoe Lake invertebrates
seemed more affected than those from McCarty Lake. In Horseshoe Lake, the mollusks
and certain oligochetes were the only groups not harmed. A few midges and mayflies
survived for more than three weeks, but these species were not collected the fol-
lowing spring. Repopulation in Horseshoe Lake had begun by late July of the
following year, with most groups of invertebrates represented. However, several
aquatic diptera, e.g. Chaborus, were not present. Eleven months after treatment,
McCarty Lake had a greater standing crop than existed prior to treatment. The
bottom fauna was almost entirely a single species of midge (Tendipes plumasus).
In a study with the freshwater shrimp (Palaemonetes kadiakensis), Naqvi
and Ferguson (1970) collected shrimp from three areas in Mississippi with a
history of insecticide exposure. For comparison, they used shrimp from a wildlife
refuge which had no history of agricultural pesticides. The LC50 for shrimp from
the wildlife refuge was 21 ppb, and for those shrimp obtained from areas of prior
pesticide exposure, the LC50 ranged from 44 to 229 ppb (Table 5.21). The Naqvi
5-81
-------
and Ferguson study (1970) and the data reported by Chaiyarach et al. (1975)
agree very well on the LC50 value for JP. kadiakensis (Table 5.21). Both studies
collected the test animals from ditches with a history of past pesticide exposure,
and the Chaiyarach study LC50 falls in the midrange of the three pre-exposed
sampling areas of the Naqvi and Ferguson study, 91 and 44, 89 and 229 ppb, re-
spectively.
McDonald (1962) studied the freshwater shrimp (Gammarus lacustris lacustris),
and calculated LCSO's for four concentrations of toxaphene (Table 5.21). Pro-
jection of McDonald's data out to 24 hours would give an LC50 of less than 20 ppb,
which agrees with the results of Naqvi and Ferguson (1970).
Naqvi and Ferguson (1968) took six species of cyclopoid copepods, a clam
(Eupera singleyi), a snail (Physa gyrina), and a tubificid worm (Tubifex tubifex)
from a pesticide-contaminated drainage ditch and from a minimal pesticide-con-
taminated area and compared 48-hour tolerance levels to toxaphene (Table 5.20).
Representative comparisons reveal that at 45 ppb the lethality to the six cyclo-
poids is three times greater in the non-exposed organisms than it is in the pre-
exposed animals; at 700 ppb in the clam, the mortality in the pre-exposed animals
is 40% as compared to 100% in the non-exposed animals; and finally at 240 ppb
in the snail, the mortality in the pre-exposed animals is 35% and in the non-
exposed animals is 100%.
Another incidence of acute toxicity differences due to the origin and prior
exposure of the organism to pesticides is demonstrated by Albaugh (1972). This
investigation involved two crayfish populations (Procambrus acutus), one popula-
tion from a floodplain near cotton fields and the other in an area free from
pesticides. A 1.5 increase in the LC50 was noted (Table 5.21). In comparing
the LC50 of 60.7 ppb in this study for P_. acutus to the LC50 of 29 ppb in
Chaiyarach et al. (1975) for P_. simulans, the LD50's differ by a factor of two.
This could be due to the different species involved or the particular level of
resistance present.
5-82
-------
Freshwater shrimp (Palaemontes kadiokensis) obtained from a bayou fed by
run off from cotton fields were exposed for 36 hours to determine the LC50 and
were compared to shrimp from untreated areas (Ferguson et al., 1965 b). Shrimp
in the bayou had an LC50 that was three times-higher than in the shrimp obtained
from the non-pesticide-exposed area (Table 5.21).
Sanders and Cope (1966) exposed two species of daphnids (Simocephalus
serrulatus and Daphnia pulex) to toxaphene to determine the 48-hour LC50. The
values for the two species are very close, varying some with temperature, but
ranging from 7 to 29 ppm (Table 5.21).
Naqvi (1973)' exposed the tubificid worm (Brachiwa sowerbyi) to 1 ppm
toxaphene at three temperatures (4.4, 21, 32.2 C) and obtained 100% mortality
in all groups. In the same paper it is reported that these worms were exposed
to a concentration of 3.7 ppm for 9,210 mins (153 1/2 hours) and to 3.5 ppm for
7,020 mins (115 1/3 hours) with the former killing crayfish in an average of
187 mins and the latter having no effect on crayfish. It is interesting that
an LC50 at 72 hours is 1 ppm, but these worms were exposed to 3.5 times this
concentration of toxaphene for twice as long and lived to be fed to crayfish.
Personal communication with Naqvi (1976) failed to resolve this conflict.
Sanders and Cope (1968) tested the naiads of three species of stoneflies
to determine the LC50 of toxaphene at 24, 48, and 96 hours (Table 5.21). The
stoneflies tested were: Pteronarcys California, body length of 30 to 35 mm;
Pteronarcella badia, body length of 15 to 20 mm; and Classenia sabulosa, body
length of 20 to 25 mm. In all of the other nine pesticides tested, the LC50
decreased with decreasing body size; however, with toxaphene the smaller naiads
(C. sabulosa) had the intermediate LC50 (Table 5.21).
5-83
-------
Pigatti and Mello (1961), using a technical grade of toxaphene, obtained
an LC50 for mosquito larvae (Culex pipens fatigans) of 14 ppm (Table 5.21).
In a study by Harris and Jones (1962) with 15 insecticides, toxaphene was
the least toxic chlorinated hydrocarbon to Culicoides variipennis larvae, having
an LC50 of 0.09 ppm compared to an LC50 for heptachlor of 0.004 ppm.
The effects of toxaphene on the macroscopic bottom fauna in a Colorado
lake have been investigated by Gushing and Olive (1956). The experimental lake,
Reservoir No. 4, is a single basin lake with a surface area of approximately
24 ha, is 4.3 m deep, and was treated with 0.1 ppm toxaphene. The control
lake, Reservoir No. 2, is a similiar lake with a depth of 2.1 m. Sampling
was done every two weeks when possible. The results are summarized in Table 5.25.
Application of toxaphene completely eradicated the tendipedid population. In the
sample taken the day following the application, only two living organisms were
found. Four days after application, large masses of dead bloodworms washed up
on the shore indicating an immediate effect on the Chironomus larvae population.
No living tendipedidae larvae were seen in the lake again until repopulation
started some eight months later.
Chaoborus larvae showed no immediate effects in the experimental lake.
Numbers actually increased during the winter with the larvae disappearing com-
pletely after six months. The authors speculate that the absence of an Entomostraca
population, upon which the Chaoborus larvae are predaceous, led to the starvation
of the Chaoborus larvae. In the control lake the population remained relatively
stable during the winter and began to pupate in the spring. No specimens were
found in August during the height of emergence, but were present again in September.
The experimental lake failed to repopulate in September.
5-84
-------
Table 5.25. Bottom fauna in two Colorado lakes expressed as
organisms per square meter*1
Date
Oct. 21,
Oct. 26,
Poisoned
Oct. 29,
Oct. 31,
Nov. 4,
Nov. 11,
Nov. 21,
Dec. 22,
Jan. 8,
Jan. 22,
Feb. 5,
Feb. 25,
Mar. 4,
Mar. 19,
Apr. 2,
Apr. 16,
Apr. 30,
May 15,
May 29,
June 22,
July 6,
July 20,
Aug. 3,
Aug. 24,
Sept. 14
Sept. 30
1954
1954
Oct.
1954
1954
1954
1954
1954
1954
1955
1955
1955
1955
1955
1955
1955
1955
1955
1955
1955
1955
1955
1955
1955
1955
, 1955
, 1955
Tendipedidae
exp. lake control
237
377
28
226
97
161
118
32
0
0
0
0
0
0
0
0
0
0
0
11
0
151
226
484
538
151
215
398
1055
(2 alive) -
(dead.)
(dead)
(dead) 1549
(dead) 1291
-
1463
1517
1302
1775
1711
1582
1291
1517
1270
710
409
65
463
1377
861
118
129
420
Chaoborus
exp.
882
527
968
258
1291
1173
1119
958
764
527
334
118
248
32
11
0
0
0
0
0
0
0
0
0
0
0
control
484
237
_
-
-
463
398
-
495 '
409
377
355
721
59.2
323
581
312
204
280
32
32
11
0
0
151
194
Palpomyia
exp. control
0
0
0
22
0
11
0
65
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
.
-
-
0
0
-
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
22
PI
exp.
0
0
0
0
0
0
0
0
0
0
0
0
0
/">
0
0
0
0
0
0
0
43
22
97
161
75
rysa
control
0 '
0
.
-
-
0
0
-
0
o
0
0
0
0
0
0
0
0
0
0
0
0
11
0
0
0
Qlig'
exp.
0
22
75
32
22
97
54
108
452
129
-
398
570
882
732
678
1506
1162
2184
1367
1205
1797
2001
1356
• 2981
3551
nchaeta
control
22
0
.
-
-
22
11
-
11
11
43
22
32
11
0
0
11
1_1
11
11
11
43
0
0
32
75
Source: Modified from Gushing and Olive, 1956.
5-85
-------
The ollgochaeta population in the experimental lake was not adversely
affected by the application of toxaphene. In fact, numbers of oligochaetae in-
creased throughout the study, probably due to the increased organic matter that
became available following the poisoning.
Needham (1966) made an extensive study on the plankton and aquatic inverte-
brates in four North Dakota lakes. The data from Wolf Butte Reservoir, a 9.7 ha
lake with a maximum depth of 2.7 m and treated with 35 ppb, are summarized in
Table 5.26. Comparison of numbers per liter was made between pre- and post-
treatment collections. Rotifers (Keratella and Asplanchna) were the most numerous.
Keratella decreased from 91 before treatment to 15 at one week, two at one month,
and only one at one year post-treatment. Asplanchna increased from 73 to 106 at
one week, but none were present at one month or at one year post-treatment.
Cladocerans (Daphnia and Bosmina) were both less frequent one year after treat-
ment. Copepoda (Deaptomus, Cyclops, and undetermined nauplii) were also less
frequent one year after treatment. Total numbers and number of genera remained
depressed in this group one year post-treatment.
Plant-inhabiting organisms were collected simultaneously and expressed as
organisms per kilogram of vegetation (Table 5.27). Gammarus (an amphipoda) varied
throughout the study, but remained abundant. The dominant Ephemeroptera (Callibaetis
and Caenis) and Odonata (Ischnura) decreased at one week and one month after
treatment, but were abundant at one year. The Dipteran, Tendipes, decreased at
one week but rebounded at one month and was about at one-half its original frequency
at one year. Bottom fauna was sampled at the same location as the plant-inhabiting
organisms and is expressed as organisms per square meter of bottom (Table 5.27).
Only two genera were abundant, Gammarus and Tendipes. Tendipes rebounded after
a year and the Gammarus population seemed depressed. A comparison of these
last two groups (plant-inhabiting and bottom fauna) does not reveal any shift in
5-86
-------
Table 5.26. Number of organisms per liter in Wolfe Butte Reservoir before
and after application of 35 ppm toxaphene
Organism
Brachionus
Keratella
Lecane
Trichocerce
Chromogaster
Asplanchna
Polyarthra
Filinia
Hexarthra
Daphnia
Simocephalus
Ceriodaphnia
Bosmina
Chydorus
Diaptomus
Cyclops
Nauplii
Pretreatment
1
91
-
1
1
73
7
1
3
244
-
4
98
1
6
46
106
1 week
posttreatment
4
15
-
-
2
106
13
1
21
18
-
9
130
-
-
3
26
1 month
posttreatment
—
2
-
-
1
-
1
-
3
129
1
18
18
-
-
11
10
1 year
posttreatment
3
1
' 1
-
-
-
2
1
-
28
-
-
25
-
2
13
73
a.
Source: Modified from Needham, 1966.
5-87
-------
Table 5.27.
Number of plant-inhabiting organisms and bottom fauna in Wolfe Butte Reservoir before
and after application of 35 ppm toxaphene
Pretreatment
Organism
Oligochaeta
Amphipoda:
Gammarus
Hydracarina
Ephemeroptera :
Callibaetis
Caenis
Odonata :
Symetrum
01 Ischnura
oo Hemiptere:
00 Notonecta
Sigara
Coleoptera :
Haliplus
Copelatus
Hydroptila
Diptera:
Tendipes
Probezzia
Chrysops
Gastropoda :
Physa
Gyraulus
Pelecypoda:
Pisidium
fa , C
plant bottom
_.
139
11
11
13
< 2.2
88
2.2
7
4
< 2.2
-
97
< 2.2
-
271
1426
< 11
65
-
11
32
-
54
-
-
-
-
11
301
32
11
22
< 11
32
1 week Posttreatment
plant bottom
< 11
378 < 11
< 2.2
< 2.2
2.2 < 11
< 2.2 < 11
22 11
-
-
< 2.2
-
< 11
20 129
22
11
715 11
1720 11
11
1 month Posttreatment 1 year Posttreatment
plant bottom plant
< .11
161 474 583
4 - 51
273
< 11 35
- - 7
43 123
7 - 22
< 2.2 4
2.2
< 2.2 4
_
106 97 55
_
-
343 32 1949
2662 97 1478
86
bottom
32
< 11
-
< 11
< 11
-
< 11
-
-
-
-
-
269
_
-
65
11
43
a
.Source: Modified from Needham, 1966.
^Organisms per kilogram of plant.
"Organisms per square meter of bottom.
-------
populations of these organisms one year after treatment, with the exception of
Physa which increased more than sevenfold.
The second lake studied by Needham (1966) was Raleigh Reservoir, with a
surface area of 6 ha and a maximum depth of 5.5 m, and was treated with
250 ppb followed 53 days later with 90 ppb treatment. Collections were made
one day before and 11, 33, and 371 days after the first treatment; a second treat-
ment was made 53 days after the first, and the fourth collection was 318 days
after this treatment. The results of this sampling are summarized in Table 5.28.
Rotifers are represented by 15 genera, but only Brachionus and Asplanchna were
abundant. Brachinus decreased from 114 before to 108 at 11 days, 15 at 33 days,
and to only three at 371 days. Asplanchna varied from 24 before to 194 at 11 days,
16 at 33 days, and only one at 371 days. At 371 days all rotifers were scarce,
and six of the original genera were not present at all. Daphnia, Ceriodaphnia,
and Bosmina were the most abundant zooplankters prior to treatment, but were
almost absent one year later. The Copepods were also very depressed one year
post-treatment. The author points out that the lake level fell to 1.8 m the
next summer and this, coupled with the double application of toxaphene, may have
contributed to the depression in zooplankters.
The plant-inhabiting and bottom fauna data are presented in Table 5.29.
Several changes are notable. Reductions of Callibaetis, Caenis, Ischnura, and
Tendipes may be significant; all but Caenis were abundant by 37 days post-treatment.
Stringer and McMynn (1958) reported that Ephemeroptera were killed by 30 ppb.
The disappearance of Sigara after treatment appears also to be related to the
toxaphene level. However, the reduction of Gyraulus at 371 days after treatment
may be due to the lowered water level, since Hooper and Grzenda (1957) and Stringer
and McMynn (1958) found Gastropoda to be unaffected by 100 ppb toxaphene.
5-89
-------
Table 5.28. Numbers of organism/3 per liter in Raleigh Reservoir before
and after treatment with toxaphene - 25 ppb- initially followed by 90 ppb on day 53°'
Organism
Brachionus
Keratella
Platyias
Lecane
Monostyla
Trichocerca
Chromogaster
Asplanchna
Polyarthra
Synchaeta
Felinia
Testudinella
Trochosphaera
Hexarthra
Conochiloides
Daphnia
Ceriodaphnia
Bosmina
Chydorus
Diaptomus
Cyclops
Nauplii
Before
treatment
114
13
<1
<1
1
5
1
24
3
1
< 1
1
< 1
2
3
65
44
314
4
10
120
190
Day 11
108
11
17
2
1
-
3
194
15
11
1
-
-
-
11
173
156
283
17
1
9
85
Day 33
15
9
1
-
-
-
-
16
7
13
-
< 1
1
-
-
57
100
50
1
1
41
19
Day 371
3
7
-
-
5
< 1
1
1
1
-
<1
-
-
-
-
9
1
-
-
-
8
7
Source: Modified from Needham, 1966.
5-90
-------
Table 5.29. Numbers of plant-inhabiting organisms and bottom fauna in Raleigh
Reservoir before and after treatment with toxaphene - 25 ppb initially
followed by 90 ppb on day 53
Organism
Oligochaeta
A
Hirudinea
Amphipoda:
Gammarus
Hydracarina
Ephemeroptera:
Callibaetis
Caenis
Odonata:
Symptrum
Ischnura
Hemiptera:
Notonecta
Sigara
Coleoptera:
Hydroporus
Diptera:
Tendipes
Gastropoda:
Physa
Gyraulus
Pelecypoda:
Pisidium
Before/ t
plant
-
<2.2
68 '
99
145
407
.
240
4
86
4
26
11
2,559
:reatment Day 11
bottom plant bottom
43 - 86
<2.2
43 689
-------
The bottom fauna was scarce in most samplings. Oligochaeta did increase,
though not as dramatically as they did in the Gushing and Olive (1956) study.
Likewise, the reduction in Tendipedidae may be significant as Gushing and
Olive (1956) found them to be eliminated by 100 ppb.
The third lake in the study by Needham was South Lake Metigoshe, a glacial
lake of 370 ha, an average depth of 2.7 m, and treated with 10 ppb followed by
a 5 ppb treatment two days later. The last lake in this series was Odland
Reservoir, a 40.5 ha lake with a maximum depth of 4.9 m, which was treated
with 5 ppb toxaphene; The results of the last two lakes are similar. Compari-
sons of zooplankters before and after treatment revealed no marked changes.
Likewise, comparisons of plant-inhabiting organisms and bottom fauna failed to
display any numerical changes that could be attributable to the toxaphene
treatment.
In a controlled laboratory experiment, Needham (1966) exposed 6 Rotifera,
2 cladocera, and 2 Copepoda to six toxaphene concentrations ranging from 50 to
1,000 ppb for 24 hours (Figure 5.6). Marked reductions in rotifers were first
observed at 500 ppb, cladocerans at 250 ppb, and copepods at 100 ppb.
The large invertebrates were exposed to concentrations at which 100 percent
survived for 24 hours and 100 percent died within 24 hours (Table 5.20). The
Coleopteran Haliplus was the most sensitive of the species tested and the
Hirudinean was the most resistant.
In an attempt to isolate the toxic components of toxaphene, Nelson and
Matsumura (1975 a) have chlorinated exo-2,10-dichlorobornane to form what they
term a simplified toxaphene. They concluded, in their bioassay system which
included LC50 determinations in both brine shirmp and mosquito larvae, that
the degree of chlorination which was optimum for toxicity was approximated by
their chlorination product 3 (Table 5.30). Chlorination product 3 has approximately
5-92
-------
Ul
I
U)
200-j
130-
120-
110-
g 100-
'E 90-
i 80-
e
I 7°-
•Q 60-
£
8 50-
30-
20-
10-
ppm
( Organism
r
-
-
"
-
•m
atcdef abcdef
Polyarlhra Hexarthra
-
-
i
~
_
-
-i
Tl
•~
-
-
n
1 ._.
-
... hll K
Filinia Keratella Asplanchna Brach
n
-
1-1
-,
TH
r-
_.
._.
-i
TH,
-
i
del abcdef abcdef abcdef abcdef abcdef
ionus Daphnia Bosmina Diaptomus Cyclops Nauplii
- Rotifers -
-I Cladocerans 1 I-
- Copepods -
ppm - a = 50, b = 100, c = 250, d = 500, e = 750, f = 1000.
Figure 5.6. Effects of six levels of toxaphene on survival
Source: Needham, 1966.
selected aquatic invertebrates.
-------
Table 5.30. Toxicities to brine shrimp and mosquito larvae of
chlorination products of exo-2,10-dichlorobornane
Brine shrimp Mosquito larvae
Artend a salina Aedes aegypti
Chemical
exo-2 , 10-Dichlorobornane
Chlorination product 1
Chlorination product 2
Chlorination product 3
Chlorination product 4
Toxaphene
LC50 (ppm)
> 4.00
0.90
0.34
0.27
0.26
0.24
LC50 (ppm)
> 10.00
3.50
0.65
0.32
0.63
0.11
Source: Modified from Nelson and Matsumura, 1975 a.
5-94
-------
the same GC pattern as toxaphene. In an effort to clean up chlorination product 3,
a TLC preparation followed by eluting with hexane on a Florisil column was done.
Sixty-five fractions were then collected from the Florisil column. Florisil
fractions 13, 14, and 15 gave a major peak with a retention time that corresponds
to a toxic compound isolated from toxaphene earlier by Nelson. A preparative
GC of the fractions isolated a peak which was checked for purity in their bio-
assay system. The LC50 of this preparation was 0.057 ppm for the mosquito
larvae and 0.32 ppm for the brine shrimp. These values compare favorably to
toxicity values for the toxic component, isolated from toxaphene, which were
0.056 ppm for the mosquito larve and 0.32 ppm for brine shrimp.
Using a detailed combination GC, TLC, and reverse phase thin-layer chroma-
tography (RPTLC), Nelson and Matsumura (1975 b) isolated a toxic fraction from
toxaphene which is 1.87 times more toxic to mosquito larvae and 1.75 times
more toxic to brine shrimp (Figures 5.7 and 5.8). This fraction behaves as a
single component in various chromatographic systems, but was found to consist
of two components on the basis of NMR. These components have been character-
ized as octochlorobornanes.
Finally, in a status report to the Environmental Protection Agency, Guyer
et al. (1971) listed LCSO's for a variety of aquatic invertebrates. These
values are presented in Table 5.21. The lowest LC50 reported was for the
shrimp (Peneus aztecus) at 2.7 ppb, whereas the highest LC50 reported was
0.086 ppm for the damselfly (Ischmura verticalis).
In summary, acute toxicities vary greatly among aquatic organisms.
Acartia tonsa (Khattat and Farley, 1976) has an LC50 of 7.2 ppt, whereas
Tubifex tubifex (Naqvi and Ferguson, 1968) has an LC50 of greater than 6 ppm.
The LC50 varies with stages of development (Section 5.3.2.2.4), salinity, pH,
and - perhaps most importantly - with prior exposure to toxaphene and/or other
pesticides (Section 5.3.2.3).
5-95
-------
x
o
z 1-6-'
8 MH
c_;
r 1.2-
1.0-
0.8-
S 2
> LU
< I °'6H
S o °-4^
S 0.2-1
q
0.0
11
^ BRINE SHRIMP
MOSQUITO LARVAE
I I I I I I 1 1 I T I I I '
-1516 17 18 19 20 21 22 23 24 25 26 27 2829-37
FRACTION NUMBER
Figure 5.7. Graph of the relative toxicities of Sephadex LH-20-methanol
column fractions to mosquito larvae and brine shrimp. Source: Nelson and
Matsumura, 1975 b.
5-96
-------
\ / Nv
'i / s BRINE SHRIMP
MOSQUITO LARVAE
TLC FRACTION
Figure 5.8. Graph of the relative toxicities of TLC separated fractions
to mosquito larvae and brine shrimp. Source: Nelson and Matsumura, 1975 b.
5-97
-------
5.3.2.2.3 Chronic toxicity — Oysters (Crassostera virginica) grown in a mix-
ture of 1.0 ppb each of DDT, toxaphene, and parathion were studied by Lowe et
al. (1971). A significant depression in weight following the nine-month exposure
period was recorded. Weights of oysters exposed to only one of these chemi-
cals at 1.0 ppb for nine months were not significantly different from the con-
* trols. Bioaccumulation of toxaphene was noted in this study (Section 5.3.1).
During the three-month depuration period which followed, the oysters were able
to eliminate the pesticides to a non-detectable level (O.010 ppm) - Histologi-
cally, oysters raised in the pesticide mixture had tissue changes associated
with the kidney, visceral ganglion, gills, digestive tubules, and tissues
beneath the gut. A mycelial fungus invaded and caused lysis of the mantle,
gut, gonads, gills, visceral ganglion, and kidney tubules. Some of the tissues
showed an intense inflammatory reaction with leucocytic infiltration. Oysters
observed after the recovery period showed very little pathology. Oysters ex-
posed to toxaphene alone showed lysed Leydig cells, increased incidence of
pigmented cells, and pathology of the viseral ganglion and adductor muscle.
A study on the bottom fauna and zooplankton in a drainage basin in
northern Alabama, contaminated with toxaphene and benzene hexachloride, was
conducted by Grzenda et al. (1964). The amount of toxaphene present in the
water was measured by a carbon absorption method which was about 45% efficient
for its recovery of the compound. The mean seasonal recoveries for toxaphene
from the Summer of 1959 through the Fall of 1960 ranged from 29 to 140 ppt.
These authors concluded that there was no convincing evidence that continuous
toxaphene contamination at these levels influenced the zooplankton or bottom
fauna populations. In their study they evaluated eight sampling sites at
5-98
-------
different sampling times. At only one station did the variety of organism
change during the course of the year, and this was at a time following the
application of insecticide in the area. This site was 2 to 6 km downstream
from an area that was poisoned by fish poachers. A drop in the number of
genera present from 38 to 24 followed. The chronic presence of toxaphene in
the Flint Creek drainage basin in northern Alabama appears to have little
effect on the number of organisms present at levels of approximately 70 to 300 ppt
5.3.2.2.4. Embryotoxic effects — In a study by Davis (1961), in which he
exposed clam eggs and larvae (Venus mercenaria) to toxaphene concentrations
of 0.25, 0.50, 1.0, 2.5, 5.0, and 10.0 ppm, the larvae were more susceptible to
the toxic effects than the eggs. Some eggs developed normally in concentrations
up to 5.0 ppm. Since the solubility of toxaphene is presumably 1.5 ppm, the
increasing toxicity of clam eggs at 2.5, 5.0, and 10.0 ppm possibly indicates
that some of the other components of this mixture may reach toxic levels above
the 1.5 ppm range. However, clam larvae exposed to toxaphene at levels of
0.25 ppm had their growth reduced drastically and caused 50% mortality. Some
larvae survived for 12 days in the 0.50 ppm concentration, but growth of these
larvae was negligible (Figure 5.9). Davis and Hidu (1969), in what appears
to be a repeat of the Davis (1961) work, obtained results that are essentially
identical in the percentage of eggs developing through normal embryonic stages
into straight-hinge larvae. The larvacidal effect of toxaphene in the second
study was greater, as seen by 0.25 ppm toxaphene causing 50% mortality in the
first study and 67% mortality in the second. At 0.50 ppm, mortality values
of 80% and 89%, respectively, were obtained. Concentrations of 1.00 ppm and
greater prevented development in both studies. Davis and Hidu (1969) took
5-99
-------
their data and calculated a 48-hour TLm (the concentration in ppm that would
cause an approximate 50% reduction in the number of eggs developing into normal,
straight-hinge larvae). They calculated this to be 1.2 ppm. A similar calcu-
lation for the earlier report gives the same TLm. They also listed a 12-day
TLm for the larvae to be <0.25 ppm. The first report would give a 12-day TLm
of 0.25 ppm. From these data it is apparent that toxaphene permitted develop-
ment of clam eggs at concentrations considerably higher than those at which
the larvae could survive and grow. The possible biochemical significance of
this is discussed in Section 5.3.2.1.
5.3.2.3 Resistance/Tolerance — The mechanism of tolerance to insecticides
may take one or more forms. Rapid epoxidation of organochlorine compounds and
the presence and action of dehydrochlorinases are possible mechanisms, as well
as mechanical exclusion. Naqvi and Ferguson (1970), using the freshwater
shrimp, demonstrated an acquired tolerance when determining the LC50 (Section 5.3.2.2),
Shrimp collected from the Noxubee Refuge in Mississippi had a tolerance to
toxaphene which was more than 10 times less than shrimp obtained from a drainage
ditch bisecting an extensive area of cotton fields; approximately 3.5 times
less than shrimp obtained from a 2024 sq. m pond receiving drainage from several
cotton fields; and about 1.8 times less than shrimp obtained from a 2.6 sq. km.
lake also receiving runoff from numerous cotton fields.
Naqvi and Ferguson (1968) demonstrated that freshwater cyclopoids, clams,
and snails that have been exposed to pesticides previously are more tolerant
than are the same species without prior exposure. Albaugh (1972) reports the
same increase in tolerance in crayfish (Procambarus acutus) collected from
a floodplain in Texas carrying a heavy pesticide load.
5-100
-------
eggs
larvae
eggs
larvae
0.25 1.00 5.00
0.50 2.50 10.00
0.25 1.00 5.00
0.50 2.50 10.00
0.25 1.00 5.00
0.50 2.50 10.00
0.25 1.00 5.00
0.50 2.50 10.00
Toxaphene Concentration (ppm )
Figure 5. 9. The effects of several concentrations of toxaphene on clam
eggs and larvae. Source: A. Modified from Davis, 1961. B. Modified from
Davis and Hidu, 1969.
5-101
-------
The ability of these animals to tolerate and store larger body burdens is
of ecological significance. Many of these organisms tested contribute sig-
nificantly to the biomass of the trophic levels upon which vertebrates are
dependent. The ability of these lower invertebrates to develop resistance/
tolerance, carry heavier body burdens, and survive in heavily pesticide-
treated areas is a situation to be further evaluated.
Naqvi and Ferguson (1968, 1970) suggest a genetic mechanism is involved,
and that this tolerance is indeed resistance. These assumptions are based in
part on other work of Ferguson (Boyd and Ferguson, 1964; Burke and Ferguson,
1969; Ferguson et al., 1964, 1965 a) on mosquito fish (Section 5.2.2.2.4).
Although this possibility has not been experimentally tested, the suggestion
that invertebrates can develop resistance to toxaphene is strong. The possi-
bility of biomagnification at higher trophic levels and the subsequent physio-
logical and biochemical alterations that could occur are areas to be closely
monitored.
5-102
-------
5.4 BIRDS AND TERRESTRIAL WILDLIFE
5.4.1 Metabolism
5.4.1.1 Absorption — No data are available.
5.4.1.2 Transport and Distribution — No data are available.
5.4.1.3 Biotransformation — No data are available.
5.4.1.4 Elimination — No data are available.
5.4.1.5 Residues — The detection of chlorinated hydrocarbon insecticide
residues in tissues of numerous wildlife species has been a well-documented
observation for many years. In particular, studies to determine the extent
of toxaphene residue accumulation in wildlife have been undertaken by several
investigators. Causey and coworkers (1972) recently collected and analyzed
samples of edible meat from bobwhite quail (Colinus virginianus), swamp and
cottontail rabbits (Sylvilagus aquaticus and S. floridanus), and white-tailed
deer (Odocoileus virginianus). Animals were obtained either in, or adjacent
to, insecticide-treated soybean fields in Alabama, and also from areas with
little or no history of organochlorine insecticide treatment. Results of
tissue analyses in animals collected from treated soybean fields are pre-
sented in Table 5.31. These results indicate that toxaphene was detectable
in 5 of 20 quail in amounts ranging from 10.30 to 88.90 ppm (average residue
level = 32.24 ppm). Two of 31 test rabbits and 3 of 22 test deer also con-
tained toxaphene residues. Among 21 deer, 17 rabbits, and 17 quail collected
from untreated areas, toxaphene residues were detected (1.26 ppm) only in a
composite sample from three deer.
Chemical analyses for toxaphene in selected tissues of ring-necked
pheasants were conducted by Genelly and Rudd (1956 a). Groups of six-month
5-103
-------
Table 5.31. Toxaphene residues in bobwhite quail, rabbits, and white-tailed deer collected from
treated soybean fields in Alabama during 1968 - 1969
Ln
I
Animal No.
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
Quail
T
10.30
88.90
26.50
ft
17.40
18.75
ft
ft
ft
*
ft
*
ft
ft
ft
T
T
ft
Residue (ppm lipid basis)
Rabbit
* (composite of three)
* (composite of four)
ft
* (composite of six)
*
ft
12.35
ft
ft
ft
ft
ft
ft
ft
ft
1.20
ft
ft
*
*
ft
A
ft
Deer
ft
ft
ft
ft
ft
ft
ft
ft
ft
ft
ft
ft
ft
ft
1.70
ft
ft
ft
6.04
8.70
ft
ft
a
Source: Causey et al., 1972.
= <0.01 ppm; * = undetected at approximately 0.001 ppm level of sensitivity.
-------
old pheasants were exposed to various levels of toxaphene in the diet and
sacrificed after 74 days. Tissue samples were taken, extracted with benzene,
analyzed for organic chloride content, and calculations for toxaphene concen-
trations were made by comparison with normal organic chloride content in con-
trol birds. Their results are summarized in Table 5.32.
During the years from 1960 to 1962 an unusually high mortality among
fish-eating birds was noted in the Tule Lake Basin of California and Oregon
(Keith, 1965). It was suggested that runoff from surrounding agricultural
areas containing organochlorine insecticides, including toxaphene, may have
been responsible for the high incidence of death. Toxaphene was used in the
region between 1959 and 1960 at application rates of two pounds per acre.
Analyses of dead birds were made for tissue residues of toxaphene and these
results are summarized in Table 5.33. Although he pointed out that inter-
pretation of the above results is difficult, the author suggested that in-
gestion by birds of 1 to 5 ppm organochlorines in the diet may result in
tissue accumulation of 20 to 50 ppm after several months.
Keith (1965) extended his study of pesticide effects on fish-eating birds
to attempt a correlation between mortality and tissue residues of toxaphene.
Data were obtained on toxaphene residues in dead birds where death might be
attributed to the pesticide exposure due to the close association in space and
time between toxaphene applications and bird mortality. Toxaphene residue
levels found in dead birds are presented in Table 5.34.
Additional data on toxaphene residues in birds have been summarized by
Markley (1974). In his report, a study by Finley (1960) was cited involving
the application of toxaphene at 1.7 kg/ha for grasshopper control. Birds
5-105
-------
Table 5.32. Toxaphene residues in ring-necked pheasant tissues
Concentration
in total diet
(ppm) Sex
89 Female
89 Male
243 Female
243 Male
243 Female
Control Female
Experimental
period
(days)
74
74
74
74
74
74
Tissue
Liver
Fat
Liver
Testis
Liver
Liver
Testis
Liver
Fat
Liver
Fat
Concentration of
chemical in
tissue (ppm)
0.0
292.0
0.0
1,510.0
6.1
0.0
0.0
4.6
752.0
0.0
0.0
a.
Source: Modified from Genelly and Rudd, 1956 a.
5-106
-------
Table 5.33. Toxaphene residues in fish-eating birds found dead at Tula Lake
and Lower Klamath Refuges between 1960 and 1962a
Year
Species collected
White Pelican
*
Western Grebe
American Egret
Great Blue Heron
Black-crowned Night Heron
Double-crested Cormorant
Californian Gull
Ring-billed Gull
1960
1961
1962
1960
1962
1960
1962
1960
1960
1961
1962
1961
1961
1961
1962
No. of
samples
1
3
3
12
2
1
5
6
2
2
1
3
1
1
1
1
1
1
3
1
1
Tissue i Average residues (ppm)
sampled toxaphene
Carcass
Liver
Kidney
HLKM
HLKM
Brain
HLKMB
Carcass
Adipose
HLKM
Brain
Carcass
HLKMB
Carcass
Carcass
Carcass
HLKMB
Carcass
HLKM
HLKMB
HLKMB
4.0
8.0
10.3
7.6
N.D.C
N.D.
N.D.
0.3
31.5
N.D.
N.D.
9.2
N.D.
10.0
N.D.
15.0
N.D.
9.5
N.D.
N.D.
N.D.
^Source: Keith, 1965.
Carcass samples were aliquots from whole, skinned birds passed through a meat
grinder. HLKMB indicates samples consisted of a composite of 5 g each of heart,
liver, kidney, breast muscle and brain, respectively.
N.D. indicates no residues detected.
5-107
-------
Table 5.34. Toxaphene residues in dead birds found in treated habitats
Level of
Habitat exposure
Nebraska Lake 0.05 ppm
Nebraska Lake 0.40 ppm
Montana Upland 1.6 - 2.2
kg/ha
fa
Species
Blue-winged Teal (3)
Shoveller (1)
Sandpiper (1)
Black-crowned Night Heron (1)
Coot (1)
Mallard (1)
Meadowlark (2)
Meadowlark (3 young)
Wilson's Phalarope (4)
Western Kingbird (1 young)
Killdeer (2)
House Wren (2)
Brewer's Blackbird (1)
Residues
(ppm)
7
12
10
64
17
10
13
3
41
4
6
41
5
a.
^Source: Keith, 1965.
Analyses were of aliquots of whole, skinned birds; numbers of individuals
are shown in parentheses.
5-108
-------
were analyzed for toxaphene residues in their tissues with the following
results: meadowlark, 12 to 67 ppm; meadowlark nestling, 8 ppm; Wilson's
phalarope, 70 to 265 ppm; killdeer, 6.44 ppm; adult house wrens, 165 ppm;
and one Brewer's blackbird, 19 ppm. Table 5.35 presents further data from
Markley (1974) on toxaphene residues in birds.
The passage of toxaphene into the eggs of wild birds has been documented
by Keith and Hunt (1966). The results of egg sample analyses for toxaphene
residues as summarized by Markley (1974) are presented in Table 5.36.
5.4.2 Effects
5.4.2.1 Physiological or Biochemical — A recent report by Hurst et al. (1974)
has presented the first data on the effects of toxaphene on thyroid physiology.
These investigators exposed bobwhite quail to toxaphene at 5, 50, or 500 ppm
in their feed for four months. At monthly intervals, treated birds were selec-
ted (male and female) and administered a tracer dose of 1-131 by intraperitoneal
injection. The birds were sacrificed after two hours and measurements were
made of thyroid, adrenal, and liver weight, and thyroid radioactivity. Results
are presented in Tables 5.37 and 5.38. Data from toxaphene-treated birds were
compared with results obtained using untreated controls by the students' "T"
test.
The data presented above suggest that thyroid weight expressed as percent
of body weight (mg percent, Table 5.37) is elevated by toxaphene treatment
after the fourth month. However, significant increases in 1-131 uptake into
the thyroid were noted only in the group treated at the highest dosage. The
authors postulated that toxaphene may increase the release of thyroid stimu-
lating hormone, which would subsequently enhance the growth of the thyroid
5-109
-------
Table 5.35. Toxaphene residues in bird tissues
Ln
I
Species
Cowbird, Brown-Headed
Molothrus ater
Dove, Mourning
Zenaidura tnacroura
Egret, Common
Casmerodius albus
Grebe, Western
Gull, Ring-Billed
Larus delawarensis
Heron, Black-Crowned Night
Nycticorax nicticorax
Heron, Great Blue
Ardea herodias
Killdeer
Charadrius vociferus
Lark, Horned
Eremophilia alpestris
Meadowlark, Western
Pelican, White
Pelecanus erythrorhyncos
Shrike, Loggerhead
Lanius ludovicianus
Tissues •
analyzed
WB-found dead
WB-found dead
WB
Fat
Fat
WB
WB
WB-found dead
WB-sacrificed
WB-found dead
WB-found dead
Liver; Kidney
Liver ; Kidney
Heart; Liver;
Kidney; Breast
WB-sacrificed
No. of
specimens
1/1
1/1
1/1
5 samples
1 sample
3 samples
1/1
1/1
4/4
3/3
3/3
1/1
1/1
49 samples
1/1
Range or avg.
of residues (ppm)
0.98
Trace
17.0
0.0 - 39.0
Avg. = 12.66
4.8
0.0 - 15.0
Avg. =5.0
10.0
9.6
0.41 - 0.96
Avg. =0.7
Trace, 2.5, 3.3
Trace, Trace, 0.6
8.0, 13.0
9.0; 14.0
0.0 - 82.0
Avg. = 3.6
Trace
Literature
citation
Hillen, 1967
Hillen, 1967
DeWitt et al . ,
Keith and Hunt
Keith and Hunt
Keith and Hunt
DeWitt et al. ,
Hillen, 1967
Hillen, 1967
Hillen, 1967
Hillen, 1967
DeWitt et al. ,
DeWitt et al . ,
Keith and Hunt
Hillen, 1967
1962
, 1966
, 1966
, 1966
1962
1962
1962
, 1966
^Source: Markley, 1974.
-------
Table 5.36. Toxaphene residues in bird eggs
Ul
I
Tissues No. of Range or avg.
Species analyzed analyses of residues (ppm)
Cormorant, Double-Crested Yolk 2 10.0
Phalacrocorax auritus
Duck, Gadwall Yolk 5 Avg. 0.04
Anas strepera
Gull, Ring-Billed Yolk 1 0.2
Larus delawarensis
Pelican, White Egg 22 0.0 - 6.7
Pelecanus erythrorhynchos Avg. 0.39
Tern, Forster's Yolk 1 15.5
Sterna forsteri
Literature
citation
Keith and Hunt, 1966
Keith and Hunt, 1966
Keith and Hunt, 1966
Keith and Hunt, 1966
Keith and Hunt, 1966
Source: Markley, 1974.
-------
Table 5.37. The effects of toxaphene on thyroid size and 2-hour uptake of I
of Bobwhite quail
131
by the thyroid
Ln
I
Toxaphene Time of
treatment treatment No. of
Cppm) (months) animals
Control 1
2
3
4
5 I
2
3
it
50 I
2
3
4
500 1
2
3
4
6
9
10
10
6
10
10
10
6
8
10
10
6
9
9
4
Mean
thyroid wu.
(mg)
10. ,1
10.6
9.4
6.7
12.6
8.7
9.4
9.1
14.0
10.4
10.2
8.6
7.1
7.2
8.5
9.3
(1
(1
(1
(0,
(1
(0.
(0.
(1
(2
(1.
(1.
.I)6
.8)
.9)
.5)
.4)
.9)
.5)
.1)
.4)
.1)
.6)
(0.5)
(1,
(0.
(1,
(1,
,2)
.4)
.3)
.1)
Mean
thyroid we .
(mg%)
4.6
5.7
4.6
3.6
6.2
4.4
5.0
5.9
6.7
5.5
5.8
5.2
5.4
5.2
5.8
6.2
(0.6)
(0.7)
(1.0)
(0.2)
(0.9)
(0.5)
(0.3)
(0.7)**
(1.0)
(0.5)
(0.7)
(0.5)*
(0.7)
(0.3)
(0.6)
(0.3)***
Mean 2-hr, upt.iki'
of (njiTli
by total thyroid
7.
8.
6.
7.
7.
7.
5.
6.
8.
7.
6.
8.
6.
10,
9.
9.
24
60
00
40
.18
40
.70
.90
,59
80
80
40
.47
.80
.70
.80
(0.72)
(1.2)
(0.8)
(0.7)
(0.96)
(0.8)
(0.8)
(0.7)
(1.18)
(1.0)
(0.9)
(1.4)
(1.35)
(0.9)
(0.8)**
(0.6)*
ol I1 '1
L-d dost-
|KT mi
0.75
0 . 90
0.81
1 .18
0.60
0.91
0.62
0.83
0.66
0.75
0.73
1.02
0.97
1.5]
1.29
1.10
, pcrci'nt
•. "f thyroid
(0.09)
(0.15)
(0.10)
(0.17)
(0.10)
(0.09)
(0.09)
(0.10)
(0.10)
(0.09)
(0.11)
(0.20)
(0.23)
(0.12)**
(0.18)*
(0.13)
^Source: Hurst et al., 1974.
*Signii" icantly different from the controls at the 0.05 level.
**Significantly different from the controls at the 0.01 level.
***Significantly different from the controls at the 0.001 level.
-------
Table 5.38.
The effects of toxaphene on body weight, adrenal weight, and liver weight of
Bobwhite quaila
I
M
M
OJ
Toxaphene
(ppm)
Control
5
50
500
i Source :
Time of
(months)
1
2
3
4
1
2
3
4
1
2
3
4
1
2
3
4
Hurst et al., 1974.
In { \ = CJl-anHarH frrnr
Nrt n f
no . 01
animals
6
10
10
10
6
10
10
10
6
10
10
10
6
10
9
4
Mean
body wt .
(gin)
226
175
208
190
210
199
187
157
211
190
175
171
130
138
147
149
(16)b
(12)
(5)
(6)
(11)
(7)
(3)**
(9)*
(11)
(4)
(12)*
(9)
(9)***
(4)**
(8)***
(ID*
Mean
adrenal
(mg%)
9.2
9.8
8.5
8.2
7.7
8.4
10.1
11.9
8.6
10.7
11.2
10.7
9.4
11.2
12.2
9.9
(1
(0,
(0.
(0.
(0
(0.
(0.
(0.
(0.
(1,
Wt .
.6)
.7)
.8)
.9)
.6)
.9)
,6)
,7)**
.6)
.4)
(2.3)
(1.
(1
(0.
(1.
(1
,4)
.3)
.5)
.6)
.2)
Mean liver
gm
3.82
2.67
3.60
3.28
3.04
3.50
3.60
2.57
3.60
3.46
3.26
3.35
3.47
2.26
2.72
2.89
(0.60)
(0.29)
(0.30)
(0.32)
(0.43)
(0.28)
(0.35)
(0.18)
(0.53)
(0.19)*
(0.35)
(0.28)
(0.37)
(0.09)
(0.25)*
(0.57)
weight
gm%
1.75
1.53
1.72
1.73
1.43
1.75
1.92
1.64
1.67
1.83
1.89
1.96
2.81
1.64
1.84
1.90
(0.32)
(0.11)
(0.13)
(0.16)
(0.15)
(0.12)
(0.17)
(0.09)
(0.17)
(0.09)*
(0.19)
(0.14)
(0.47)
(0.07)
(0.09)
(0.23)
*Significantly different from the controls at the 0.05 level.
"Significantly different from the controls at the 0.01 level.
***Significantly different from the controls at the 0.001 level.
-------
and uptake of iodide. Moreover, as the thyroid enlarged and increased its
production of thyroid hormone, thyroid stimulating hormone secretion would
decline accompanied by a reduction in iodide uptake. As Table 5.37 points
out, each mg of thyroid tissue after four months of treatment is taking up
only as much iodide as the controls. Alternatively, it was suggested that
toxaphene may decrease the circulating half-life of thyroid hormone and there-
by cause a compensatory hypertrophy of the thyroid.
Toxaphene treatment at the highest dosage level significantly depressed
the body weight gain of birds at all time intervals. A similar, but not
statistically significant, trend was noted at the other two dosages. Dif-
ferences in adrenal and liver weights among treated and control birds were
not generally significant. It appeared, however, that some adrenal hyper-
trophy may have resulted from the toxaphene treatment after three months.
The authors did not attribute specific deleterious effects to altered
thyroid function in bobwhite quail. It was noted, however, that aberrant
thyroid function may be associated with increased reproductive failure and
abnormalities. Substantial scientific evidence exists to indicate that
chlorinated hydrocarbons in general can disrupt reproductive function in many
wildlife species.
An unusual report concerning the pharmacologic actions of toxaphene on
a Bengal tiger has recently been published (Peavy, 1975). In this case, a
female Bengal tiger, age 12 years, became poisoned after ingesting portions
of a llama calf that had died after being dipped in a toxaphene solution two
days prior to its death and subsequent feeding to the tiger. The tiger pre-
sented clinical symptoms identical to those produced by acute chlorinated
5-114
-------
hydrocarbon poisoning. These included periodic convulsions and hyperreflexia
to sudden auditory and visual, but not tactile, stimuli. Effective control of
the symptoms was obtained by administration of diazepam and methocarbamol.
5.4.2.2 Toxicity
5.4.2.2.1 Acute — The effects of acute exposure to toxaphene have been
determined in a variety of wildlife species. These investigations have gener-
ally taken the form of follow-up studies to assess the impact on wildlife from
insecticide application programs. Summarizations of acute toxicity data from
studies conducted under controlled experimental conditions have been prepared
by Pimentel (1971) and Markley (1974). These results are presented in Table
5.39. Most acute toxicity studies with wildlife are difficult to interpret
due to inadequate presentation of data and/or deficiencies in experimental
design (e.g., number of animals, age, sex, statistical analysis). A careful
examination of the experimental results as reported, however, should allow for
a relatively accurate assessment of potential hazards to wildlife which occur
by accidental exposure to toxaphene.
Dahlen and Haugen (1954) exposed bobwhite quail (8 weeks old) and mourn-
ing doves to toxaphene by oral administration in gelatin capsules. Where
death occurred from the treatment, it was usually preceded by a lack of motor
coordination and depression. Table 5.40 presents a summary of their experi-
mental data, including an approximate LD50 for each species. The method of
data tabulation used in this study is of questionable reliability and the
authors did not provide statistical limits of significance for the calculations
presented.
It was shown in a study conducted by Hudson and coworkers (1972) that the
susceptibility of mallard ducks to toxaphene poisoning is an age-dependent
5-115
-------
Table 5.39. Acute toxicity of toxaphene to birds and terrestrial wildlife
Species
Mallard duck (3-5 mo., female)
Pheasant (3-mo., female)
Bobwhlte quail (3-mo., male)
Sharp-tailed grouse (1-4 yr., male)
Fulvous tree duck (3-6 mo., male)
Lesser sandhill crane (female)
Pheasant (2-wk. old)
Coturnix (2-wk. old)
Bobwhite quail (chick)
Mallard duck (duckling)
Ln
I Mule deer (16-17 mo., male)
1 — i
^— i White pelican (young)
cr.
Chuckar partridge
Pheasant
Sage grouse
White pelican (young)
White pelican
Toxaphene^
exposure
70.7
40.0
85.4
10-20
99.0
100-316
500-550 ppm
600-650 ppm
834 ppm
563 ppm
139-240
100-400
50
200
90
100
50
Route of
administration
oral
oral
oral
oral
oral
oral
diet
diet
diet
diet
oral
oral
oral
oral
(capsule)
(capsule)
(capsule)
(capsule)
(capsule)
(capsule)
- 5 days
- 5 days
- 5 days
- 5 days
(capsule)
?
?
?
Effects
LD , no other symptoms reported
LD , no other symptoms reported
LD n, no other symptoms reported
LD , no other symptoms reported
LD,.,., no other symptoms reported
LD , no other symptoms reported
LC (5 day treatment followed by clean food for 3 days)
LC (5 day treatment followed by clean food for 3 days)
LC,... (5 day treatment followed by clean food for 3 days)
LC,... (5 day treatment followed by clean food for 3 days)
LD , no other symptoms reported
one bird died at 100 mg/kg, another was unaffected at
200 mg/kg, and a third was intoxicated at 400 mg/kg
but survived
LD . , no other symptoms reported
LD no other symptoms reported
LD,... , no other symptoms reported
two of three birds died; survivor recovered after
48 hours
two of three birds died; survivor recovered after
48 hours
Reference
Tucker and Crabtree,
1970
Heath et al. , 1970
Heath and Stickel,
, 1965
Tucker and Crabtree,
1970
Keith 1964
Post, 1949
1
Keith, 1965
Keith, 1965
a
Units of exposure are mg/kg body weight except where indicated as ppni in the diet.
-------
Table 5.40. Toxicity data for birds exposed to toxaphene
No. birds No. birds LDsg Avg. wt. Avg. no. of
Species dying (mg/kg) loss(%) days lived
Bobwhite Quail 60 22 80 - 100 25 3
Mourning Dove 8 5 200 - 250 22 3
Source: Modified from Dahlen and Haugen, 1954.
5-117
-------
phenomenon. Birds of both sexes were treated orally with a single dose of
toxaphene and resulting LDSO's were calculated (Table 5.41). Previous tests
conducted by the authors had demonstrated that nonbreeding birds do not dis-
play significant sex-dependent differences in sensitivity to pesticide effects.
The investigators obtained a biphasic age-sensitivity curve to toxaphene, as
* well as to three other central nervous system stimulants. It was suggested
that the relative insensitivity of the youngest birds was due to the functional
immaturity of the CNS; young animals were either unable to undergo full clonic
convulsions or possibly they could tolerate higher brain concentrations of the
chemical. Increasing sensitivity as the CNS matures is evident by the decreased
LD50 in birds at age 7 days. However, the trend of increasing sensitivity with
age is reversed by 3 to 5 months, and may be due to increased hepatic micro-
somal metabolism of the chemical or decreased chemical permeability of the
brain. The implication of this study is clearly that the youngest of the
species may not necessarily be the most susceptible to toxaphene poisoning.
Among several field studies to assess the impact on wildlife from in-
secticide spraying programs, the most recent was conducted by McEwen and co-
workers (1972). Toxaphene was applied to a 71,685 ha tract in New Mexico
for control of range caterpillar at the rate of 1.1 kg toxaphene in .87 liter
of fuel oil per hectare. As the data in Table 5.42 indicate, toxaphene treat-
ment resulted in a marked decrease in bird numbers in comparison to untreated
rangeland in the same area. A number of dead birds were found in the spray area
including three horned larks, two meadowlarks, one killdeer, one cowbird, and
one mourning dove. Observations made during the first week after toxaphene
spraying revealed no significant change in bird numbers or mortality.
5-118
-------
Table 5.41. Acute oral LD^ in mg/kg to Mallards of various ages (95% confidence
limits in parentheses)"1
Age
Chemical 36 + 3 Hr. 7+1 Days 3 - 5 Mo.
Toxaphene 130 30.8 70.7
(80.4 - 210) (23.3 - 40.6) (37.6 - 133)
^Source: Hudson et al., 1972.
Female.
5-119
-------
Table 5.42. Bird census on selected plots sprayed with toxaphene
Spray
rate
(kg/ha)
1.1
No. of
No. of days
counts post-spray
18 8-14
Avg . no .
Pre-spray
33.6
of birds
Post-spray
6
15.0
a.
/Source: Modified from McEwen et al., 1972.
Differs from pre-spray, a. - 0.01 and from unsprayed area, a. = 0.05.
5-120
-------
In an earlier study conducted by Eyer and coworkers (1953), controlled
field conditions were employed to evaluate the effect of toxaphene on geese in
cotton fields. Geese of the White Emden, Toulouse, and White China breeds,
age 14 to 16 weeks old, were used in the study. Toxaphene was applied to ex-
perimental cotton field plots at rates of two to six pounds per acre, either
by dust or spray. Some geese remained in the fields both during and after
toxaphene application while others were introduced for three days after the
spraying or dusting.
Table 5.43 summarizes the results obtained when geese were exposed to
toxaphene either by direct contact from spraying or by eating toxaphene-
treated vegetation. Toxaphene was apparently safe in either dust or spray
when applied at 2.2 kg per ha. At 4.4 kg per ha, birds of the Emden and
Toulouse breeds were killed within three hours of direct contact with toxa-
phene spray. At 5.5 kg/ha, direct contact with toxaphene spray produced death
in the Emden breed only within three hours. Sickness followed by recovery was
observed in the White China breed. At 6.6 kg per ha none of the geese were
affected. This apparent inconsistency in the results was not explained.
Symptoms of poisoning in the geese were generally characteristic of chlorin-
ated hydrocarbon intoxication, involving considerable central nervous system
effects. Upon post mortem examination, several distinct and consistent lesions
were encountered. First, a generalized cyanosis was observed in all birds.
In addition, congestion and edema accompanied by hemorrhages were noted in
both the heart and spleen. Several birds also displayed swollen and congested
livers.
5-121
-------
Table 5.43. Toxicity of toxaphene to geese used to control grass and weeds in cotton fields
a
Treatment
Toxaphene,
Dust
Spray
Toxaphene,
Spray
Toxaphene,
Dust
Spray
Toxaphene,
Spray
Toxaphene,
Spray
Geese Geese Geese Geese
introduced introduced introduced introduced
immediately 24 hours 48 hours 72 hours
Direct after after after after
contact treatment treatment treatment treatment
2.2 kg/ha fa
No No No ' No No
No No No No No
3.3 kg/ha
Sickness No No No No
4.4 kg/ha
No No No No No
Sickness and death No No No No
5.5 kg/ha Sickness and death No No No No
6.6 kg/ha
No No No No No
^Source: Modified from Eyer et al., 1953.
No apparent effects.
Birds showing signs of sickness later recovered.
-------
5.4.2.2.2 Chronic— Data concerning the adverse effects of long-term toxa-
phene exposures to wildlife are quite limited. Keith (1965) reported the
results from a three month toxaphene feeding study using groups of White
Pelicans. Groups of five birds each were fed with sardines which had been
injected with toxaphene in corn oil to achieve concentrations of either 10 or
50 ppm. Symptoms of poisoning occurred only in the group treated at 50 ppm;
bird deaths were recorded at days 29, 32, 34, 43, and 48 of treatment. Upon
autopsy, dead birds were noted to be practically devoid of subcutaneous and
mesentery fat deposits. Deaths also occurred among control birds. However,
these animals generally lived longer and contained greater fat deposits than
toxaphene-treated pelicans. A comparison of toxaphene and DDT with respect to
toxicity to pelicans revealed that the former insecticide is a more potent poison.
Genelly and Rudd (1956 a) conducted a detailed study on the chronic
toxicity of toxaphene to ring-necked pheasants. Four groups of ten female
birds were fed toxaphene in their diet for up to three months at levels of 25,
100, 200, and 300 ppm. No mortality resulted in any of the treatment groups.
A transitory depression of body weight was noted during the first month of the
study, but was attributed to decreased feed consumption. Histological analysis
of livers from treated birds demonstrated varying degrees of vacuolation. No
other chronic effects resulting from toxaphene treatment were encountered.
Linduska and Springer (1951) found bobwhite quail to be adversely affected
by toxaphene at dietary levels of 500 ppm and greater. When incorporated in
the diet at 1000 ppm, toxaphene produced 100 percent mortality in 13 days. At
a level of 500 ppm in the diet, a 75 percent mortality was achieved in 25
days.
5-123
-------
5.4.2.2.3 Other effects — Several parameters relevant to bird reproduction
have been measured in response to toxaphene exposure. Genelly and Rudd (1956 b)
determined the effect of toxaphene exposure on egg production, fertility, hatcha-
bility, and survival of young in ring-necked pheasants. Their results, as sum-
marized in Table 5.44, show that significant changes were noted in egg laying
rate and egg hatchability when hens were fed toxaphene in the diet at 300 ppm.
In addition, survival of young birds was significantly depressed by about five
percent during the first two weeks following hatching. An evaluation of
overall reproductive success obtained by considering the net effect of several
parameters revealed the adverse effect of toxaphene treatment (Figure 5.10).
A recent report (Bush et al., 1977) indicated that dietary toxaphene levels of
0.5, 5.0, 50, and 100 ppm given to chicks from one day of age through maturity
did not significantly alter egg production, hatchability, or fertility.
Dunachie and Fletcher (1969) investigated the effect of toxaphene on the
hatching of eggs from White Leghorn hens. Each experiment consisted of four
batches of 25 eggs each with one batch serving as a control group. Each egg
was injected with toxaphene dissolved either in corn oil or acetone. Controls
were injected with solvent only. Their results, depicted in Table 5.45, show
some unexplained inconsistencies when using acetone solutions and do not
establish a definite dose-response relationship.
Smith and coworkers (1970) examined the effects of toxaphene on egg
hatchability. Hens' eggs were injected with the chemical in corn oil directly
into the yolk after a seven day incubation period. As shown in Table 5.46,
doses of toxaphene up to 1.5 mg per egg did not significantly reduce hatchability.
Eggshell thinning caused by pesticide exposure has been partly responsi-
ble for observed reproductive failures in several species of fish-eating and
5-124
-------
Egg Production
MO Females-30 Days) Fertile Hatch
220
200 -I
13 Day Young
Control
Relative
Reproductive
Success
70%
"• Toxaphene (100 ppm ) 62 %
Toxaphene ( 300 ppm ) 46%
Figure 5.10. Survival curves showing the net effect of toxaphene on
ring-necked pheasant reproduction. Source: Modified from Genelly and
Rudd, 1956.
5-125
-------
Table 5.44. Egg production, fertility, and hatchability
Laying
rate
ppm Hen Eggs (eggs/ Eggs Eggs Fertility
in mash days laid o/day) incubated fertile (per cent)
Toxaphene
100 459 333 .725, 251 211 84.1
300 918 436 .475 337 297 88.1
Control
0 1470 1047 .712 785 717 91.3
Hatch-
Young ability
hatched (per cent)
170 80.6,
165 55.6
570 79.5
£ , Source: Modified from Genelly and Rudd, 1956 b.
Differs significantly from control at 5 per cent level.
-------
Table 5.45. The effect of toxaphene on the hatching of hen's eggs '
Toxaphene In acetone In corn oil
ppm solvent solvent
500 23/135 (17)
400 29/121 (24) 5/36 (14)
300 18/ 75 (24) 5/32 (15.5)
200 27/104 (26) 13/72 (18)
100 49/100 (49)
10 12/ 75 (16)
^Source: Modified from Dunachie and Fletcher, 1969.
Results expressed as ratio of number of eggs hatching to number
treated. Figures in parentheses indicate actual percentage of
treated eggs hatching.
5-127
-------
Table 5.46. Percent hatchability resulting from injection of toxaphene into
the yolk of fertile eggs after seven days' incubation
Toxaphene
treatment L
(mg) % Hatchability
c.
Control 92
Od 70.7
0.07 84.0
0.15 77.3
0.30 69.3
0.60 74.7
0.90 70.7
1.50 76.0
^Source: Modified from Smith et al., 1970.
Seventy-five eggs per treatment.
Eggs were drilled, sealed, and incubated with no
/injection whatever.
Eggs were injected with corn oil only.
5-128
-------
raptorial birds. Haegele and Tucker (1974) recently investigated the eggshell-
thinning effects of numerous pesticides on coturnix quail. They found that a
single oral dose of toxaphene at 10 mg/kg of body weight did not significantly
reduce eggshell thickness. No other dose levels were employed in this study,
nor were additional bird species investigated.
Several investigators have recently become aware of the adverse effects
of pesticides on rumen microbial function in wildlife animal species. Schwartz
and Nagy (1974) examined the effect of toxaphene on the in vitro fermentation
of dry matter (alfalfa hay) inoculated with rumen fluid from yearling mule deer.
The results obtained by using rumen bacteria from both wild and penned deer
are presented in Table 5.47. Inhibition of dry matter digestion was signifi-
cant only at toxaphene levels of 1000 ppm.
5-129
-------
Table 5.47- Effect of toxaphene on percent digestion of dry matter in vitr£ by
Mule Deer rumen bacteria
Rumen bacteria
source 0
Wild deer 42. lb
Penned deer 52.5
Toxaphene
10
41.3
52.3
levels , ppm
100
40.7
51.4
1000
28. 5G
c
34.5
-Source: Modified from Schwartz and Nagy, 1974.
Cumbers are percent digestion of dry matter.
Significantly different from controls at P <0.05.
5-130
-------
5.5 DOMESTIC ANIMALS
5.5.1 Metabolism
5.5.1.1 Absorption — No data are available.
5.5.1.2 Transport and Distribution — No data are available.
5.5.1.3 Biotransformation — No data are available.
5.5.1.4 Elimination — No data are available.
5.5.1.5 Residues — The use of toxaphene for the control of ectoparasites on
domestic animals has led to the appearance of toxaphene residues in both meat
and milk. Moreover, the ingestion of pesticide-contaminated forage also pro-
vides a route of entry for these chemicals into domestic animal tissues.
Data on toxaphene residues in meat and poultry products have been compiled
by the U.S. Department of Agriculture (1978) and are summarized in Table 5.48.
Toxaphene is found consistently from year to year in the fat of cattle, although
the incidence of contamination is extremely low. During the survey period
(1973 to 1978) only six samples were in excess of the tolerance limit (7.0 ppm).
Of these six violations, five were in fat samples from cattle, one of which
occurred in the first quarter of 1978.
Li and coworkers (1970) conducted extensive residue analyses on milk
products obtained from dairy cattle after daily feeding of toxaphene at 15
mg/kg of body weight. Samples were taken in duplicate from a group of two
cows and toxaphene content analyzed in the fat extracted from their milk.
Toxaphene residues in raw milk and in dairy products derived therefrom are
summarized in Table 5.49.
In an earlier study conducted by Zweig and coworkers (1963) 16 dairy cows
were fed toxaphene at 0 to 20 ppm in the feed for up to 77 days. Samples of
milk were analyzed once or twice weekly to determine the presence of toxaphene
(measured as total chloride; limit of sensitivity = 0.02 ppm). At all treat-
5-131
-------
Table 5.48. Residues of toxaphene in fat samples of meat and poultry products
at slaughter in the United States3
Number of Positive Samples/Total Number of Samples (%)
Animal
Cattle
Calves
Sheep & Goats
Swine
Chicken
Turkeys
Ducks & Geese
Rabbits
Horses
TOTAL
1973
9/710
1/84
2/289
4/232
3/530
3/517
0/95
0/19
0/44
22/2520
1974
(1.27)
(1.19)
(0.69)°
(1.72)
(0.57)
(0.58)
(0.0)
(0.0)
(0.0)
(0.87)
2/1117
0/284
1/371
2/329
1/1138
0/735
0/148
3/266
9/4388
(0.18)
(0.0)
(0.27)
(0.61)
(0.09)
(0.0)
(0.0)
(1.13)
(0.21)
1975
3/1733
0/269
0/356
0/324
0/777
0/554
0/246
0/11
0/261
3/3971
(0.17)
(0.0)
(0.0)
(0.0)
(0.0)
(0.0)
(0.0)
(0.0)
(0.0)
(0.08)
1976
3/1785
0/327
0/250
1/442
0/927
0/456
0/267
0/65
0/217
4/4736
(0.17)
(0.0)
(0.0)
(0.23)
(0.0)
(0.0)
(0.0)
(0.0)
(0.0)
(0.08)
1977
4/880
0/124
0/100
0/215
1/375
0/303
0/186
0/21
0/112
5/3216
(0.45)
(0.0)
(0.0)
(0.0)
(0.27)
(0.0)
(0.0)
(0.0)
(0.0)
(0.22)
1978b
1/432
0/62
0/36
0/179
0/191
0/64
0/39
0/14
0/20
1/1037
(0.23)
(0.0)
(0.0)
(0.0)
(0.0)
(0.0)
(0.0)
(0.0)
(0.0)
(0.10)
U.S. Department of Agriculture, 1978.
first two quarters only
listed as lamb
-------
Table 5.49. Toxaphene content of raw whole milk and products
manufactured subsequently13
Raw whole
milk, mg/kg
fat
21.9
20.8
21.9
20.8
20.8
21.9
21.9
28.3
28.7
30.1
• Source :
Product
Past, whole milk
Past, whole milk
Past. 30% cream
Skim milk
Past. 30% cream
Skim milk
Ster. cream
Butter
Buttermilk
Ster. cream
Butter
Buttermilk
Curd
Whey
Cheese
Curd
Whey
Cheese
Cond. milk
Ster. milk
Dry whole milk
Cond. milk
Ster. milk
Dry whole milk
1,1 et al. , 1970.
Toxaphene,
mg/kg fat
21.6
20.4
27.2
36.0
23.2
31.5
30.0
27.1
29.2
28.7
31.0
24.5
28.4
26.7
27.0
30.3
29.5
25.4
24.9
24.7
25.6
25.0
Storage
container
Glass
Paper
Glass
Paper
Paper
Paper
b
Parchment
Parchment
Waxed
Waxed
Paper
Can0 fc
Poly, bag
Paper
Can6 6
Poly, bag
Toxaphene In stored
mg/kg fat
7 Days
25.0
22.5
22.8
20.5
29.2
25.4
26.5
samples,
14 Days
28.3
22.8
24.8
22.3
31.7
27.8
28.3
-------
ment levels, toxaphene concentrations in milk reached a plateau after the 28th
day (Table 5.50). In addition, a rapid decline in toxaphene residues in milk
was observed about four days after the pesticide treatment ceased (Table
5.51).
In a similar study by Claborn and coworkers (1963) toxaphene was incor-
porated in the diet of 14 Jersey cows at levels of 20, 60, 100, and 140 ppm.
Two animals were used as controls and three cows were treated at each of the
four dosage levels. Total chloride analysis of milk samples (corrected by sub-
traction of chloride-content in untreated samples) demonstrated the presence
of toxaphene residues in the milk at all dosages (Table 5.52). Residue levels
reached a plateau after about four weeks of toxaphene treatment, and decreased
rapidly upon withdrawal from the contaminated feed. Omental fat samples were
collected by Claborn et al. (1963) after eight weeks of toxaphene treatment
and analyzed for residue content. Residue levels in animals on the 60, 100,
and 140 ppm dietary dosages were 8.4, 14.3, and 24.3 ppm, respectively.
Pesticide residues in the milk of dairy cows fed toxaphene-treated
alfalfa hay were demonstrated more than two decades ago (Bateman et al.,
1953). At an application rate of 1.1 kg per ha of hay, toxaphene was detected
(total chloride method) in the milk of two cows at an average of 2.5 and 2.3
ppm over a 112 day period. Hay treated at 2.2 and 4.4 kg of toxaphene per ha
produced residues of 4.3 and 3.9, and 18.2 and 8.3 ppm, respectively, in the
milk.
Toxaphene sprayed on domestic animals has also resulted in toxaphene
residues in various tissues. Claborn and coworkers (1963) applied toxaphene
by spray to three groups of two Jersey cows each. Two groups were sprayed
twice at three week intervals with toxaphene in the form of either a 0.5
5-134
-------
Table 5. 50. Toxaphene concentration (ppm) in milk from cows fed daily rations containing varying
amounts of added toxaphene
Days
2
5
7
9
11
14
18
21
25
28
32
35
42
49
56
63
70
74
77
/Source:
2.5
0.04
0
0.05
0.06
0.03
0.03
0.02
0.07
b
0.07
Zweig et al. , 1963.
ppm
5
0.08
0.10
0.05
0.10
0.07
0.20
0.09
0.05
0.03
0.01
0.06
0.07
0.08
Toxaphene added
10
0.13
0.14
0.06
0.19
0.08
0.10
0.05
0.15
0.11
0.05
0.12
0.10
0.16
to ration
15
___
0.26
0.18
0.11
0.14
0.14
0.15
0.13
0.34
0.31
0.08
0.16
0.09
0.16
20
___
0.16
0.08
0.18
0.24
0.20
0.22
0.14
0.26
0.18
0.11
0.19
0.18
0.18
The 2.5 ppm level was discontinued after the 32nd day.
-------
Ln
I
GJ
ON
Table 5. 51. Toxaphene concentration (ppm) in milk from cows during feed-off period following feeding
of rations containing varying amounts of added toxaphene
Days after
toxaphene
feeding was
discontinued
1
4
7
8
11
14
18
21
25
27
30
34
41
ppm Toxaphene added to
2.5
0.07
0.03
0
0
5
0.01
0.01
0.01
0
0
0
0
0
— — —
10
0.07
0.01
0.03
0
0
0
0
0
ration
15
0.09
0.05
6
0.08
0.02
0
0
0
0
0
0
20
0.09
0.02
0.06
0.02
0.02
0.02
0.01
0
0.01
0
0
^Source: Zweig et al., 1963.
These results for the llth day and all subsequent dates exclude results of one cow of this
group which had become virtually dry.
-------
Table 5.52. Toxaphene (ppm) in milk from cows fed different levels of toxaphene in the diet
(Each sample consisted of 2 liters from morning milk and 1 liter from afternoon milk)
Oi
i
Dosage ,
ppm
20
60
100
140
Control
Cow -
no.
3
4
5
Av.
6
7
8
Av.
9
10
11
Av.
12
13
14
Av.
1
2
Av.
Weeks fed
1
0.17
0.26
0.16
0.20
0.61
0.61
0.47
0.56
0.90
0.87
0.85
0.87
1.46
1.13
1.74
1.44
0.00
0.00
0.00
2
0.24
0.31
0.24
0.26
0.65
0.69
0.50
0.61
0.99
1.00
1.05
1.01
1.56
1.09
2.36
1.67
0.02
0.00
0.01
3
0.24
0.31
0.24
0.26
0.74
0.87
0.65
0.75
0.92
1.08
1.04
1.01
1.68
1.40
2.32
1.80
0.00
0.00
0.00
4
0.31
0.41
0.35
0.36
0.70
0.66
0.67
0.68
1.06
1.19
1.19
1.15
1.75
1.45
2.47
1.89
0.07
0.04
n.06
5
0.29
0.24
0.35
0.33
0.67
0.69
0.53
0.63
0.87
1.13
0.92
0.97
1.31
1.23
1.96
1.50
0.20
0.11
0.16
6
0.33
0.42
0.35
0.37
0.68
0.77
0.69
0.71
0.96
1.04
0.89
0.96
1.39
1.23
2.31
1.64
0.17
0.14
0.16
7
0.25
0.31
0.26
0.27
0.47
0.53
0.48
0.49
0.93
0.97
0.68
0.86
1.36
1.53
2.24
1.71
0.00
0.00
0.00
8
0.21
0.25
0.24
0.23
0.44
0.52
0.48
0.48
0.90
0.96
0.88
0.91
1.52
1.44
2.51
1.82
0.00
0.00
0.00
Weeks after shift to
untreated feed
1
0.10
0.06
0.06
0.07
0.08
0.14
0.16
0.13
0.11
0.18
0.17
0.15
0.19
0.30
0.46
0.32
0.00
0.00
0.00
2
0.01
0.04
0.02
0.02
0.05
0.11
0.13
0.10
0.05
0.16
0.18
0.13
0.17
0.22
0.80
0.40
0.00
0.00
0.00
3
....
0.04
0.09
0.09
0.07
0.06
0.15
0.12
0.12
0.21
0.26
0.20
0.00
0.00
0.00
"Source: Claborn et al.f 1963.
-------
percent emulsion, or a 0.5 percent suspension of the wettable powder. The
third group was treated twice daily for 21 days with one ounce of a 2.0 percent
toxaphene spray in kerosene. Milk samples obtained from all sprayed cows
contained detectable toxaphene residues (measured as total organic chlorine
and corrected for chlorine content in control milk samples). These results
are presented in Tables 5.53 and 5.54. In all cases, residue levels rapidly
decreased after cessation of toxaphene treatment.
Roberts and Radeleff (1960) demonstrated in hogs that toxaphene was
probably not substantially accumulated in fat. Four weeks after the spray
application of a 0.5 percent toxaphene emulsion, samples of omental and renal
fat from eleven hogs contained no chemical residues (determined by a modifi-
cation of the total chlorine combustion method). Unfortunately, samples were
not taken either during or shortly after the toxaphene treatment period.
Dietary levels of up to 100 ppm fed to beef cattle and sheep for 16 weeks
resulted in toxaphene residues in omental fat of 20 and 28 ppm, respectively.
After discontinuing exposure, fat residues decreased to 12 to 14 ppm at four
weeks and 0.5 and 3.0 ppm at eight weeks (Claborn, 1956; Table 5.55). Sheep
and cattle fed 25 ppm toxaphene had fat residues of 8 and 12 ppm, respectively,
after 16 weeks of feeding. Claborn (1956) also reported on studies in which
12 beef cattle were sprayed with 0.5 percent toxaphene 12 times at 2-week
intervals. Omental fat samples were taken surgically prior to each spray
application. Table 5.56 shows that after the 12th spraying fat residues were
11 to 16 ppm and had decreased to 4 to 5 ppm four weeks after the last spraying.
No toxaphene was detected in the fat of any animals after the first two sprayings.
Claborn (1956) compared fat storage of several organochlorine insecticides
in cattle and sheep. He also reported milk residues after spraying 0.5 percent
5-138
-------
Table 5.53. Insecticide (ppm) in milk from cows sprayed twice at 3-week
intervals with 0.5% sprays of toxaphene
5
Emulsion spray Suspension spray
Sampling date
Before spraying
Days after first
spraying:
1
2
3
5
7
14
21
Days after second
spraying:
1
2
3
5
7
14
21
/Source: Claborn et al. ,
Toxaphene
0
0.60
0.61
0.44
0.23
0.16
0.06
0.08
0.52
0.55
0.41
0.24
0.16
0.10
0.06
1963.
Toxaphene
0
0.64
0.74
0.58
0.26
0.20
0.12
0.12
0.71
0.81
0.57
0.31
0.18
0.08
0.02
The insecticides were calculated in ppm of milk containing 4% butterfat
The values given are averages from the analysis of two milk samples.
Each sample consisted of 10 grams of butterfat. The titration of the
control sample from each cow was used as the blank and was substracted
from subsequent samples.
5-139
-------
Table 5.54. Insecticide (ppm) excreted in milk of dairy cows sprayed twice
daily for 21 days with 1 ounce of 2.0% oil solution of toxaphenea
Sampling date Toxaphene
Before spraying 0
Days after spraying
started:
1
3
7
14
21
Days after spraying
ceased:
7
14
21
0.12
0.41
0.33
0.26
0.30
0.09
0.10
0.06
/Source: Claborn et al. , 1963.
Calculated from organic chlorine determinations for milk containing
4% butterfat. The values given are averages from the analyses of
two milk samples of 3 liters each.
5-140
-------
Table 5.55. Parts per million of insecticides stored in the fat of cattle and sheep
that had known amounts added to their diet
Ln
I
Insecticide Dosage Animal — 7
(ppm) i
weeks
Toxaphene 100 Ewe 29
21
19
Wether 20
25
16
Av. 22
Steer 25
—
30
Heifer 23
26
Av. 26
25 Ewe 1
1
0
Wether 1
2
4
Av. 2
Heifer 2
3
1
Steer 4
1
Av. 2
After
8
weeks
30
19
18
17
19
24
21
27
45
34
27
35
34
1
1
1
0
4
4
2
4
4
9
4
1
4
feeding
12
weeks
—
—
25
—
—
0
25
36
43
29
25
33
33
3
3
1
9
1
1
3
11
7
9
11
12
10
16
weeks
30
21
19
17
19
17
20
37
52
29
33
39
38
11
7
11
—
4
6
8
16
12
16
8
9
12
4
weeks
19
9
11
10
15
10
12
10
29
24
10
15
14
After feeding ceased
8 20 32 36
weeks weeks weeks weeks
0
0
0
0
0
3
0.5
—
9
0
3
0
3
(From Claborn, 1956)
-------
Table 5.56. Parts per million of toxaphene In fat of calves sprayed
with 0.5 percent toxaphenea
Animal
3rd
Steer 0
2
7
—
Av. 3
2 weeks after indicated spraying
4th
5th
6th 7th
8th
9th 10th
Emulsion prepared in laboratory
7 — 7 — — 10
11 — o
3 0 10
17 — — 8
2
5
Emulsion
—
12
from
Humble Oil
Steer 2
—
—
0
0
—
Av. 1
—
3
11
—
—
—
7
9
—
—
13
7
—
10
4
1
—
7
—
9
7
7 10
llth
10
12
—
11
12th
13
9
—
11
After 12th spraying
4 weeks
—
5
5
6 weeks
0
5
—
2
concentrate furnished by
and
—
4
5
—
—
8
6
Refining
—
—
—
7
—
—
7
Company
—
—
9
—
7
—
8
9
12
—
—
—
—
12 9
—
—
6
—
8
—
7
13
20
—
—
—
—
16
—
—
—
—
—
4
4
—
—
2
—
6
—
4
aAll samples taken from the control animals were negative or within the experimental error of the method,
, which is + 4 ppm.
None found in any sample after the first two sprayings.
(From Claborn, 1956)
-------
emulsions and suspensions of toxaphene and strobane. Of the insecticides
studied the order of their storage in fat from greatest to least was as
follows: aldrin > dieldrin > BHC > DDT > chlordane > lindane > endrin >
heptachlor > toxaphene > methoxychlor. Methoxychlor was the only insecticide
studied that did not cause some storage in the body fat.
Another study (Ent. Res. Service, ARS, USDA, 1959) involved the grazing
of range forage treated with 24 oz. toxaphene/A in 1 gal. diesel oil. The
forage was analyzed for toxaphene residues and the following results were ob-
tained (Table 5.57).
Table 5.57- ppm Toxaphene on Forage on Days Indicated
After Application
Days I 1 714 4l84~
ppm
Toxaphene 8.59 4.62 3.50 3.61 1.66 0.99
*
Each value represents a composite of 8 sampling sites.
Five yearling cattle were placed on the pasture at the time of spraying and
four were placed seven days later. They were kept on the pasture 103 and 96
days, respectively. Omental fat samples were taken: 1) before being placed
on the pasture, 2) when removed from the pasture (103rd and 96th day), and 3)
at the time of slaughter (120th day). Table 5.58 shows the fat residues of
toxaphene. The important finding in this study was that toxaphene
residues did not increase above the tolerance of 7 ppm established for meat.
Schanzel and Chemtai (1974) recently noted in a report on problems re-
lated to tick control in Uganda, that high levels of toxaphene were being dis-
covered in milk and milk products in Uganda. The authors cited data which
revealed levels of 3 ppm in milk, 15 ppm in cream, 21 ppm in cottage cheese,
and 87 ppm in milk fat.
5-143
-------
Table 5.58. Residues in beef cattle fed hay from fields
sprayed with toxaphene
Dosage
oz/A
24
No. of
Animals
5
4
Days
Exposed
103
96
Pre-
exposure
0.1
0.2
Toxaphene in
End of
Exposure
3.5
3.2
Fat (ppm)
At
Slaughter
0.5
0.5
(From Ent. Res. Div., ARS, USDA, 1959)
5-144
-------
5.5.2 Effects
5.5.2.1 Physiological or Biochemical — No data are available.
5.5.2.2 Toxicity
5.5.2.2.1 Acute — Because toxaphene is registered for use in the control of
ectoparasites on livestock, several acute toxicity investigations have been
conducted employing domestic animals. For the most part, these studies focused
on defining "minimum toxic" and "maximum non-toxic" doses.
Radeleff (1949) described in detail the clinical and pathologic signs
associated with 27 cases of experimental toxaphene poisoning in goats, sheep
and cattle. Each case was detailed individually. The onset of clinical signs
occurred within a few minutes to a few hours following exposure. Although not
all clinical signs observed were seen in a single poisoned animal, there was
sufficient similarity between animals that one can summarize a consistent syn-
drome associated with toxaphene poisoning.
An affected animal became at first apprehensive and hypersensitive and,
in some cases, belligerent. Soon blepharospasms and fasciculations of the
facial and cervical muscles appeared, followed by clonic spasms of the cervical
muscles, then those of the forequarters and, finally, those of the hindquarters.
The spasms were either continuous leading to rapid death or, more frequently,
appeared intermittently at regular or irregular intervals concurrent with the
appearance of muscular spasms. There was an increased flow of saliva and accom-
panying chewing movements producing froth. As the seizure activity increased,
animals often became frenzied, lost coordination, walked aimlessly about, moved
in circles or jumped imaginary objects. Abnormal posturing such as resting the
sternum on the ground while the hindlegs remained in the standing position, or
persisting in keeping the head between the forelegs while standing were frequently
observed. The clinical signs progressed to clonic-tonic seizures accompanied by
5-145
-------
periods of paddling movements, nystagmus, grinding of the teeth and groaning
or grunting in a comatose attitude.
Marsh and coworkers (1951) reported sublethal doses of 35 to 110 rag/kg
for toxaphene fed to 17 heifers and young cows weighing from 500 to 800 pounds.
The minimum lethal dose was 144 mg/kg. In earlier work, Radeleff (1949) treated
goats and sheep by oral drenching with toxaphene. At 100 mg/kg one of two sheep
was killed; at 170 mg/kg one of two goats was killed, whereas two sheep sur-
vived the same exposure.
Reports from field observations have indicated that young dairy and beef
calves are particularly susceptible to poisoning by toxaphene when applied as
a spray or dip. Radeleff and Bushland (1950) sprayed calves, two to six weeks
old, with three different formulations of toxaphene at various concentrations.
Their results, as summarized in Table 5.59, show that death was produced by
treating calves at concentrations as low as one percent. The effects of oral
toxaphene administration to three month old calves, adult goats, and adult
sheep were also investigated. These results are presented in Table 5.60.
In addition, the results obtained by treating adult cattle, goats, and sheep
with toxaphene sprays or dips are summarized in Table 5.61. A strict adherence
to expected dose-related increases in mortality was not observed with any of
the animal species or treatments. The nature of the apparent increased sus-
ceptibility of young calves to toxaphene poisoning cannot be explained by
these results.
Choudbury and Robinson (1950) reported that successive daily oral doses
of toxaphene at 25, 37.5, 50, and 75 mg/kg caused no adverse effects in one
goat. However, death occurred after feeding 100 mg/kg on days 9 and 10 and
150 mg/kg on day 11. In another animal, a dose of 100 mg/kg on days 1 and 2
followed by 150 mg/kg on day 3 caused death on day 4.
5-146
-------
Table 5.59. Effects of toxaphene sprays applied to suckling calves
a
L/i
Concentration of Total Number . Formulation
toxaphene animals treatments administered
8.0% 3 1 XE-65
KE-65
WP-40
4.0% 3 1 XE-65
KE-65 ,
WP-40
1.5% 12 2 XE-65
KE-65
WP-40
1.0% 11 1 XE-65
WP-40
0.75% 12 8 XE-65
KE-65
WP-40
Number of
animals
1
1
1
1
1
1
4
4
4
8
3
4
4
4
Results
Died
Died
Affected
Affected
Unaffected
Affected
2 Affected
2 Unaffected
1 Died
2 Affected
1 Unaffected
3 Affected
1 Unaffected
1 Died
7 Unaffected
3 Unaffected
1 Affected
3 Unaffected
4 Unaffected
4 Unaffected
Source: Radeleff and Bushland, 1950.
When animals were treated more than once, the interval between sprayings was 4 days.
XE-65 = An emulsion concentrate containing toxaphene 65 per cent, xylene 25 per cent,
Triton X-100 10 per cent; KE-65 = A commercially formulated emulsion concentrate con-
taining 65 per cent toxaphene dissolved in kerosene; WP-40 = A commercially formulated
/wettable powder containing 40 per cent toxaphene.
Two consecutive sprayings of 4% concentration.
-------
Table 5.60. Effects of oral administration of toxaphene formulations
Ul
I
Co
Animals
treated
Calves
(3-month-
old)
Goats
(Adult)
Sheep
(Adult)
Dose
toxaphene
95 mg/kg
50 mg/kg
50 mg/kg
100 mg/kg
170 mg/kg
250 mg/kg
50 mg/kg
100 mg/kg
Total Formulation i
animals administered
3 XE-65
KE-65
WP-40
3 XE-65
KE-65
WP-40
3 XE-65
KE-65
WP-40
3 XE-65
KE-65
WP-40
3 XE-65
KE-65
WP-40
3 XE-65
KE-65
WP-40
3 XE-65
KE-65
WP-40
3 XE-65
KE-65
WP-40
Number of
animals
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
Results
Unaffected
Unaffected
Unaffected
Died
Affected
Affected
Affected
Affected
Affected
Affected
Affected
Unaffected
Died
Affected
Affected
Died
Died
Died
Unaffected
Unaffected
Unaffected
Affected
Affected
Died
-------
Table 5. 60.(continued)
Ln
I
Animals
treated
Sheep
(Adult)
(cont'd)
^Source:
Dose
toxaphene
170 mg/kg
250 mg/kg
Radeleff and
Total
animals
3
3
Bushland, 1950.
Formulation /
administered
XE-65
KE-65
WP-40
XE-65
KE-65
WP-40
Number of
animals
1
1
1
1
1
1
Results
Affected
Affected
Affected
Died
Affected
Died
XE-65 = An emulsion concentrate containing toxaphene 65 per cent, xylene 25 per cent,
Triton X-100 10 percent; KE-65 = A commercially formulated emulsion concentrate con-
taining 65 per cent toxaphene dissolved in kerosene; WP-40 = A commercially formulated
wettable powder containing 40 per cent toxaphene.
-------
Table 5.61. Effects of toxaphene sprays or dips applied to adult animals
Ul
I
Animals
treated
Steers
Goats
Sheep
Concentration of Total Number
toxaphene animals treatments
8.0% 1 1
4.0% 20 1
8.0% 3 1
4.0% 3 1
8.0% 3 1
4.0% 3 1
Formulation /
administered
XE-65
XE-65
KE-65
WP-40
XE-65
KE-65
WP-40
XE-65
KE-65
WP-40
XE-65
KE-65
WP-40
XE-65
KE-65
WP-40
Number of
animals
1
7
6
7
1
1
1
1
1
1
1
1
1
1
1
1
Results
Affected
1 Affected
6 Unaffected
6 Unaffected
7 Unaffected
Died
Died
Affected
Unaffected
Unaffected
Unaffected
Affected
Died
Affected
Affected
Affected
Unaffected
/Source: Radeleff and Bushland, 1950.
XE-65 = An emulsion concentrate containing toxaphene 65 per cent, xylene 25 percent,
Triton X-100 10 per cent; KE-65 = A commercially formulated emulsion concentrate con-
taining 65 per cent toxaphene dissolved in kerosene; WP-40 = A commercially formulated
wettable powder containing 40 per cent toxaphene.
-------
5.5.2.2.2 Chronic— Little information is available concerning the effects
of long-term toxaphene exposures to domestic animals. In early studies, Marsh
(1949) fed toxaphene-treated hay (8.8 kg/ha) to 14 yearling steers. Treatment
continued for more than four months, with daily toxaphene intake averaging
7.9 mg/kg. Two animals developed transient central nervous system effects.
Lambs fed toxaphene-treated hay at 1.1, 2.2, 4.4, and 8.8 kg per ha showed no
toxic symptoms.
5.5.2.2.3 Other — In addition to the data which have been accumulated con-
cerning the mortality produced in domestic animals by toxaphene poisoning,
recent work has shown that the pesticide may also affect rumen microbial ac-
tivity. Kutches and coworkers (1969, 1970) investigated the action of toxa-
phene on in vitro fermentation using rumen liquor obtained from a sheep. They
measured pesticide effects on dry matter disappearance, volatile fatty acid
production, and changes in protozoal numbers. Toxaphene caused a statistic-
ally significant depression of dry matter disappearance of 20 and 28 percent
when added to the rumen liquor at levels of 750 and 1,000 ug/ml. No depression
was produced at lower concentrations. The authors noted that among the insec-
ticides tested (DDT, sevin, malathion, toxaphene), toxaphene was the most in-
hibitory. None of the pesticides tested caused a significant decrease in
volatile fatty acid production. However, toxaphene produced the greatest
depression of protozoal numbers among the pesticides tested.
5-151
-------
5.6 TERRESTRIAL INSECTS AND OTHER TERRESTRIAL INVERTEBRATES
5.6.1 Metabolism
Although toxaphene has been used as an insecticide for over twenty years,
little is known about the metabolism of this compound in insects. With the
exception of a recent study by Crowder (1976), no information is available on
the absorption, transport, distribution, and elimination of toxaphene. The
rate of toxaphene penetration after topical application has been determined
by Crowder (1976) using the American cockroach, Periplaneta americana. After
application of 75 micrograms of toxaphene per cockroach, penetration into the
hemolymph and other body tissue was rapid over the first 8 hours, appeared to
stabilize between 8 and 24 hours, and increased markedly between 24 and 48 hours
(Table 5.62). The marked increase in absorption between 24 and 48 hours cor-
responded with the onset of mortality on these animals. The distribution of
toxaphene was determined in another cockroach species, Leucophaea maderae
(Table 5.63).
Insects showing signs of toxaphene intoxication tended to have much higher
levels of toxaphene in the hemolymph and the nerve cord at 120 hours after
exposure than did insects showing no sign of intoxication. About five percent
of the toxaphene associated with the nerve cord seemed to be tightly bound to
nervous tissue and was not removed after 20 minutes of rinsing in insect saline.
At the subcellular level, toxaphene binding to nerve tissue was greatest in
the nuclei, nerve sheath, and microsomal components (Crowder, 1976).
Based on gas chromatographic analysis of toxaphene residues in nervous
tissue, Crowder (1976) found no evidence for biotransformation of toxaphene
by the cockroach. However, Abd El-Aziz and coworkers (1966) reported the
5-152
-------
Table 5. 62. Penetration of Cl-36 labelled toxaphene Into the American cockroach after
topical application of 75 micrograms
Ln
LO
Time,
hours
1/2
1
2
4
8
24
48
Hemo lymph
Percent of
Micrograms applied dose
1.11
2.79
2.15
5.08
4.57
6.04
11.09
1.48
3.72
2.87
6.77
6.09
8.05
14.79
Carcass
Micrograms
8.84
8.00
5.47
7.82
11.75
12.97
51.56
Percent of
applied dose
11.78
10.67
7.29
10.42
15.66
17.29
68.75
Total
Micrograms
8.37
10.13
6.35
15.23
23.67
25.63
62.66
Percent of
applied dose
+ SE
11.16 + 4.0
13.51 + 5.8
8.46 + 3.4
20.31 + 7.7
31.56 + 6.5
34.17 + 4.3
83.54 + 11.7
/Source: Crowder, 1976.
Carcass rinsed in acetone to remove unabsorbed toxaphene.
-------
Table 5.63. The distribution of CI-36 labelled toxaphene in tissues of
Leucophaea maderaea
Tissue
Memo lymph
Nerve cord
Fat body
Alimentary canal
Number
of
insects
75
21
12
126
27
12
75
21
12
75
21
6
b
Disposition
A
A
S
A
A
S
A
A
S
A
A
S
Hour
time
<120
120
120
<120
120
120
<120
120
120
<120
120
120
Average
(Pg/g)
193.88
67.02
563.50
654.00
184.59
268.71
263.89
309.62
299.03
272.97
260.72
305.02
^Source: Crowder, 1976.
A = Asymptomatic, animals show no signs of intoxication.
S = Symptomatic, animals show signs of nervous system stimulation.
5-154
-------
presence of a toxaphene dehydrochlorinase in the dry acetone powder extracts
of a toxaphene-resistant strain of the cotton leafworm, Prodenia litura. Using
colorimetric methods for the analysis of toxaphene, these investigators esti-
mated 5 to 50 percent breakdown of toxaphene under a variety of in vitro conditions.
Gluathione and diethyl ether were necessary cofactors for the reported break-
down of toxaphene.
5.6.2 Effects
5.6.2.1 Physiological and Biochemical Effects — The mechanism by which toxaphene
adversely affects insects is not known. Most investigators feel that toxaphene,
like other cyclodiene insecticides, is a neuroactive agent which acts by alter-
ing ion permeability in the neurons, rather than by inhibiting cholinesterase
(Matsumura, 1975). Using central nerve cord preparations from the German
cockroach, Blatella germanica, incubated in 100 micromolar toxaphene solutions,
Wang and Matsumura (1970) distinguished three stages of response. The first
stage, which began after twelve minutes, was characterized by frequent
(5 to 7/second), short (0.03 to 0.05/second) bursts of spontaneous activity with
maximum spike intensity of 70 to 80 microvolts. In the second stage, which began
after 17 minutes, spike intensity decreased to 50 to 70 microvolts. Bursts became
irregular, less frequent (1 to 4/second), and of longer duration (0.1 to 0.4 seconds)
After 25 minutes of incubation, spontaneous bursts became weaker, less frequent,
and shorter. The threshold of response for toxaphene was 0.6 micromolar, with
an average latent period of 14 minutes. Similar bursts of spontaneous nerve
discharge were caused by a variety of the cyclodiene insecticides (Wang and
Matsumura, 1970). Spontaneous high amplitude, high frequency nervous discharge
has also been demonstrated in nerve cord preparations from the American cockroach,
5-155
-------
Periplaneta americana, at toxaphene concentrations of 1 to 100 millimolar.
At lower concentrations, 0.1 to 0.01 millimolar toxaphene had little effect
on nerve cord activity. Preliminary observations on the rate of chloride ion
uptake by the nerve cord indicated that changes in chloride ion permeability
may be related to changes in spontaneous neural activity (Crowder, 1976).
Although toxaphene is not generally considered to act by acetylcholinesterase
inhibition, Litzbarski and Litzbarski (1974) have demonstrated in vitro acetyl-
cholinesterase inhibition in both the housefly and Galleria mellonella, the
greater wax moth. During ten-minute incubation periods, the toxaphene con-
centrations causing 50 percent inhibition were about 500 ppm for the moth and
3400 ppm for the fly.
Toxaphene has also been shown to alter formate metabolism in houseflies
(Cline and Pearce, 1963). Flies injected with C-14 labelled formate and then
treated topically with 3 micrograms toxaphene per fly demonstrated a 5-fold
increase in labelled uric acid, but a 2.5-fold decrease in labelled proline
after three hours. This effect was also seen with other chlorinated insecti-
cides but not with organophosphate or carbamate insecticides. Because pro-
line is metabolized to gamma-aminobutyric acid which functions in the trans-
mission of nerve impulses, Cline and Pearce (1963) suggested that proline
depletion accompanied by a presumed increase in gamma-aminobutyric acid may
be involved in the neurotoxic effects of the chlorinated hydrocarbons.
Moore and Taft (1964) were not able to demonstrate in vitro succinic
dehydrogenase inhibition by toxaphene in boll weevil homogenates.
5-156
-------
5.6.2.2 Toxicity
5.6.2.2.1 Acute Toxicity to Insects
5.6.2.2.1.1 Acute Toxicity of Technical Toxaphene to Insects — Information on
the acute toxicity of toxaphene to various insects in topical application is
summarized in Table 5.64. Based on the rather limited number of LD50 estimates
given in units of microgram toxaphene per gram insect, the coleopteran, dipteran,
and hemipteran pest species, along with the largely beneficial hymenopteran
species seem to be susceptible to toxaphene over similar dose ranges, about
2-80 micrograms/gram. In contrast, the lepidopteran species, most of which are
economically significant cotton pests, and the coleopteran lady beetles, seem
to be much less susceptible to toxaphene with LC50 values commonly over 1000
micrograms/gram. However, the significance of this generalization is difficult
to assess. Besides the normal problems in comparing toxicity estimates by
different investigators, the development of toxaphene-resistant insect strains
may have had a pronounced effect on these toxicity estimates. In an attempt
to minimize this effect so that more valid interspecies comparisons can be
made, only the lowest LD50 values from studies measuring toxicity in more than
one population are presented in Table 5.64. Nevertheless, as discussed more
fully in Sections 5.6.2.2.2 and 5.6.2.2.4, toxaphene has enjoyed wide use in
the protection of cotton crops from lepidopteran pests for over 20 years.
During this time, many strains or populations of these insect pests have be-
come resistant to toxaphene. Thus, the higher LD50 values for the lepidopterans
may reflect the high selection pressure for toxaphene resistance on this order
of insects as a group and/or the use of highly resistant lepidopteran strains
collected from areas of intense toxaphene exposure.
5-157
-------
Table 5.64. Acute toxicity of toxaphene to various insects on topical application
Ul
1
M
Ln
CO
ORIJER
Genus species
(Common name)
COLEOI'TERA
Anthonomus grandls
(Boil weevil)
Coleomegilla maculaca
(Lady beetle)
llippodainia convergens
(Lady beetle)
DIPTERA
Musca domestics^
(House fly)
11EMIPTERA
Lygus hesperus
(Alfalfa bug)
Slope of
A, „, LD50 (95;.: confidence interval) lt>8 ^°se~
Observation probit
Stage hours micrograms/gram micrograms/insect line
Adult 48 5.43
48 240
72 12.5
72 81.1
48 1.296 (0.780-1.812)
48 20.388 (17.815-23.453)
Adult 72 1789 22
Adult 48 1069
Adult 48 70.0
Adult — 33 (30-35)
Adult — 2.1
Adult 48 0.463 2.58
Reference
Fye et al. , 1957
Roussel et al. , 1959
Burkhalter and Arant, 1960
Bass and Rawson, 1960
Brazzel and Ship, 1962
Hopkins et al. , 1975
Atallah and Newsom, 1966
Burke, 1959
Ohsawa et al. , 1975
I.andrura et al. , 1976
Anagnos topoulos et al.,
1974
Leigh and Jackson, 1968
Dose estimation based on average insect weight. See text for details.
-------
Table 5. 64 (continued)
ORDER
Genus species
(Common name)
HYMENOPTERA
Apis roellifera
(Honey bee)
Slope of
log dose-
Observation LD50 (95% confidence interval) probit
period in response
Stage hours micrograms/gram micrograms/insect line Reference
Adult 48 19.08 ' Graves and Mackensen, 1965
Adult 48 0.144 Torchio, 1973
Bracon mellitor
(a parasitic wasp on
the boll weevil)
Adult
48
14.2
0.035
Johansen and Davis, 1972
Adams and Cross, 1967
Ln
I
Ln
Brucliophayus roddi
(Alfalfa seed chalcid)
Campole Lis perdis tivc tus
(a parasitic wasp on
tlie boll weevil)
Megachile rotundata
(Alfalfa leaf-cutting bee)
Nomia melanden
(Alkali bee)
Vuspula pensylvanica
(Western yellowjacke t)
Adult
Adult, female
Adult
24
Adult, male 24
Adult, female 24
Adult, female 72
Adult, female 72
48
48
2.12
0.676 (0.0512-0.0892) 2.45 Bacon and Riley, 1963
0.1769 (0.1635-0.1913)
0.2633 (0.2505-0.2767)
0.216
0.0023
3.36
5.67
67.0
Lingren et al., 1972
Johansen, 1972
Torchio, 1973
Torchio, 1973
Johansen and Davis, 1972
-------
Table 5.64 (continued)
Ul
1
M
(^
O
ORDER
Genus species
(Common name)
I.El'IDOPTERA
lleiiotliis drmigera
Heliothis pusrctigera
Heliochis virescens
(Tobacco budworm)
Heliothis zea
(Bollworm)
Peccinophora gossypiella
(Pink bollworm)
Prodenia litura
(Egyptian cotton leafworm)
Stage
Larva
Larva
Larva
Larva
Larva
Larva
Larva
Adult
Larva,
4th instar
Larva,
4th instar
Observation
period in
hours
48
48
48
48
48
48
48
48
—
24
Slope of
LD50 (95% confidence interval) lo& ^ose
probi t
microg rams/gram
[380.0]
[106.0]
[3,280]
[2,500]
5,020
320 (180-460)
[794]
1371 (890-2130)
1430
mlcrograms/insect line
13.61
3.70
82.0
100.0
0.71
32.0
0.534
1 2.0
Reference
Wilson, 1974
Wilson, 1974
Guzman-Varon et al
Adkisson, 1964
Wolfenbarger, 1973
Brazzel, 1964
Adkisson, 1964
Lowry and Berger,
. , 1974
1964
Salama et al. , 1966
Eldefrawi et al.,
1964a
Pseudaletia unipuncta
(Army wo mi)
Larva
56.2
Weinman and Decker, 1951
-------
An additional problem in making valid comparisons among the various species
is that most of the acute toxicity data is presented in units of micrograms/insect.
While such information may be of some use in intraspecies comparisons, the rela-
tively large variation in size among insect species severely limits the utility
of this data in interspecies comparisons. This limitation is most apparent in
the hymenoptera data on bees compared to the parasitic wasps. In studies giving
the average weight of the insects used, microgram/gram units are estimated in
Table 5.64. However, because insect weights in a given species vary widely,
no attempt has been made to approximate insect weights and convert LD50 units
in the other studies.
For the most part, the methodology used in these studies is similar.
Acetone is used as the toxaphene vehicle. Appropriate dilutions are applied
in 1 to 3 microliter quantities to the dorsal thoracic region. In all cases,
appropriate vehicle controls are used. Control mortality usually ranged from
0 to 3 percent. When excessive mortality was noted in the control group
(e.g., Johansen and Davis, 1972), the solvent volume was lowered until an
acceptable control mortality was obtained. In most studies, the LD50 esti-
mates were corrected for control mortality using Abbott's (1925) formula.
Although 48 hour LD50 estimates are the most common, some studies give
mortality estimates based on 24 or 72 hour exposures. The effect of such
variation on the LD50 estimates cannot be defined, given the limited informa-
tion on the speed of action of toxaphene. Using the residue-film method in
toxicity studies on the housefly, Sun (1971) estimated LC50's for toxaphene
after 3 and 22 hours at 188 micrograms/jar and 22 micrograms/jar, respectively.
Taking a ratio of these values as an index of speed of action, Sun (1971)
5-161
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classified toxaphene as relatively slow acting when compared to a number of
other organochlorine insecticides, organophosphorus insecticides, or carbamates.
In agreement with the observation, Torchio (1973) noted substantial differences
in mortality between 24- and 48-hour exposure periods in three species of
bees. However, for the honey bee and the alkali bee, no deaths occurred after
48 hours. With the leafcutting bee, 20% of the deaths occurred between 48 and
72 hours. All bees surviving 72 hours also survived up to 96 hours. If this
pattern is common to other insects, 48- and 72-hour LC50's should be relatively
similar to each other, but substantially lower than 24-hour LCSO's. This is
consistent with Crowder's (1976) observation of the rate of toxaphene absorp-
tion in cockroaches (Section 5.6.1)-
Various factors, such as weight, age, developmental state, sex, and diet,
have been shown to affect the toxicity of toxaphene to insects. The effects
of weight and age on toxaphene toxicity to the larvae of two lepidopterans have
been examined by Eldefrawi and coworkers (1964 a) and Weinman and Decker (1951).
In both studies, increasing larval weight was associated with decreasing toxa-
phene toxicity. In the Egyptian cotton leafworm (Prodenia litura) topical
LDSO's to 3rd, 4th, and 5th instar larvae were 2.64, 1.43, and more than 7.30
milligrams/gram insect, respectively. The corresponding weight ranges for
these instars were 10 to 15 mg, 40 to 60 mg, and 180 to 200 mg (Eldefrawi et
al., 1964 a). In the armyworm (Pseudaletia unipuncta), Weinman and Decker
(1951) found a well-defined trend between topical toxicity and larval weight.
In four groups of larvae weighing less than 150 mg/larva, 151 to 250 mg/larva,
251 to 350 mg/larva, and more than 351 mg/larva, the 48-hour LD50's were 33.7,
49.8, 58.2, and 67.0 micrograms/gram, respectively. These investigators did
5-162
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not indicate which larval instars of the armyworm were tested. While both
studies suggest that increasing larval size is accompanied by decreasing toxi-
city of toxaphene, the lack of confidence intervals for the LD50's prevents
an evaluation of the significance of these findings. In both studies, the
maximum differences in toxicity among the larval groups are only about 2-fold.
The effect of diapause on toxaphene toxicity has been examined in the
adult lady beetle, Coleomegilla moculata (Atallah and Newsom, 1966). The
beetles were collected and maintained in diapause until the time of testing,
with mating used as the criterion for return to activity. The resulting LD50
for the diapausing beetle (44.2 micrograms/beetle or 3619 micrograms/gram)
was about twice that of the active beetle (Table 5.64). These differences were
significant at the 0.05 level. As illustrated in Figure 5.11, the slopes of the
log dose-probit response lines for both stages are parallel, indicating similar
heterogenicity and possibly similar modes of action in the two stages. Atallah
and Newsom (1966) conclude that the relatively gradual slopes - e.g., high degree
of heterogenicity - were indicative of a population currently developing resis-
tance to toxaphene.
Possible differences in the response of males and females to toxaphene
has received little attention. With Bracan mellitor, Adams and Cross (1967)
noted lower LD50 values (as micrograms/insect) in males than in females. This
difference was attributed to the smaller size of the male wasps. Insufficient
information is given in their report to determine if both size and sex could
have been involved. For another wasp species, Campoletis perdistinctus, males
were also found to be more susceptible than females in terms of dose per wasp
(Table 5.64). Although no reference was made to size differences (Lingren et al.,
1972), male ichneumonid wasps are generally much smaller than females. In a
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100
90
80
70
f 60
o
5 50
•M
c
oj
30
20
10
0
345 10 20 30 4050 100 200300400
( Micrograms/Beetle)
Figure 5.11. Log dose-probit lines for toxaphene treated lady
beetles based on 72-hour observations. Source: Atallah and Newsom, 1966.
5-164
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modified contact toxicity test in which male and female houseflies were exposed
to various concentrations of toxaphene in different types of soil samples, male
flies were consistently about twice as sensitive to toxaphene based on measure-
ments of LT90's (Qadri, 1972). (Different soil types had no apparent effect on
toxicity.)
In the boll weevil, Anthomonus grandis, larval habitat, adult food source,
as well as weevil age, have been shown to have significant influence on the
topical toxicity of toxaphene (Bass and Rawson, 1960). In their study, larvae
were collected either from cotton squares (square-reared) or bolls (boll-reared)
and emerging adult weevils were tested at either two or nine days of age. During
the test period, adult weevils were fed from only one source, either blooms,
bolls, or squares. The results, summarized in Table 5.65, indicated that boll-
reared, boll-fed, nine-day old weevils were 61 times more tolerant to toxaphene
than square-reared, bloom-fed, two-day old weevils. The single factor con-
tributing most to susceptibility was adult food source. Adults fed on cotton
bolls were much less susceptible than those fed on cotton squares, regardless
of larval habitat or age. In a field study, Gaines and Mistric (1952) also
found that boll-reared larvae were four times more tolerant of toxaphene appli-
cations than square-reared larvae.
The oral toxicity of toxaphene has been determined in the 4th instar
larva of two lepidopteran species. In both cases, toxaphene was more toxic
orally than topically. In the armyworm, Psudaletia unipuncta, the typical
LD50 was 56.2 micrograms per gram, while the oral LD50 was 34.1 micrograms
per gram (Weinman and Decker, 1951). As in topical applications, smaller
larvae seemed more susceptible to toxaphene than larger larvae. In the
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Table 5.65. Toxiclty of toxaphene to boll weevils having various combinations of larval habitat, age,
and adult
Ln
I
o\
. . LD50 and fiducial limits in yg/p
Age of weevils 6 °
Adult food
(days)
Toxaphene
Square-reared
Blooms
Squares
Squares
Bolls
Bolls
2
2
9
2
9
81.1 +
140.9 +
512.1 +
2,099.0 +
4,385.0 +
31.2
58.4
194.9
707.0
1,544.5
Boll-reared
Squares
Squares
Bolls
Bolls
Source: Modified
2
9
2
9
from Bass and Rawson, 1960.
330.4 +
1,067.0 +
4,982.0 +
4,943.0 +
102.8
849.8
153.0
1,537.5
-------
Egyptian cotton leafworm, Prodenia litura, Salama and coworkers (1966) found a
greater proportioned difference between the topical LD50 (1.371 micrograms/gram)
and oral LD50 (0.250 micrograms/gram).
Other types of toxicity tests have been conducted on toxaphene. The
utility of two of the most frequently used, the immersion method and the dry
film method, has been recently evaluated by Hopkins and coworkers (1975).
Although such tests are useful in screening for efficiency (Section 5.6.2.2.5),
actual dosage data in terms of quantity of toxicant per animal cannot be
obtained from such methods. Consequently, such studies are not detailed in
this section. They are included, when appropriate, in discussions of syner-
gistic effects (Section 5.6.2.2.3) and efficacy (Section 5.6.2.2.5).
Only one study using injection as the exposure route has been encountered.
Crowder (1976) estimated the 48-hour LD50 of toxaphene by injection to the
cockroach, Leucophaea moderae, at 2.37 micrograms/gram.
5.6.2.2.1.2 Acute toxicity of toxaphene components to insects — As discussed
previously (Section 2.0), toxaphene is a complex mixture of various components.
Three of these components, two in toxicant A and one in toxicant B, have been
identified (footnote in Table 5.66). In addition, a few investigators have
separated toxaphene into various fractions. The toxicity of some toxaphene
fractions and the identified components to houseflies are summarized in Table
5.66. In the seven toxaphene fractions tested by Ohsawa and coworkers (1975),
there is no apparent relationship of toxicity to either chlorine content or
the presence of toxicants A and B. The least toxic fraction, fraction I, has
the same chlorine content as the most toxic fraction, IV. While the two
highly toxic fractions (III and IV) both contain toxicant B, fraction V, which
contains both toxicant A and B, is somewhat less toxic than technical grade
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Table 5.06. Acute topical LD50's of toxaphene and toxaphene fractions to the
house fly*1
Technical toxaphene
Expected values of
Fractions I - VII
Fraction I
Fraction II
Fraction III2'
0
Fraction IV
d e.
Fraction V '
Fraction VI
Fraction VII
Toxicant A"
Toxicant B^
LD50
(micrograms/g)
70
75
316
139
42
33
76
101
171
16
32
Toxicity
relative to
toxaphene'-'
1.0
0.93
0.22
0.50
1.67
2.12
0.92
0.69
0.41
4.38
2.18
Percent Cl
68.6
68.1
68.3
68.8
68.1
66.8
64.1
21.2
^Source: Modified from Khalifa et al., 1974 and Ohsawa et al., 1975.
LD50 of toxaphene-j-LD50 of component.
Expected value calculated as harmonic mean, i.e., (X) = N/E (1/Xi).
Includes Toxicant B.
/Includes Toxicant A.
Toxicant A: Identified as mixture of 2,2,5-endo,6-exo,8,8,9,10-octa-
chloroborane and 2,5,5-endo,6-exo,8,9,9,10-octachloroborane (Turner et al.,
1975)..
Toxicant B: Identified as 2,2,5-endo,6-exo,8,9,10-heptachloroborane.
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toxaphene. An identical pattern is seen in the toxicity of these fractions
and components to mice (Table 6.13, Section 6.2.2.1.2). The expected value of
the LD50 for fractions I through VII does not indicate any synergistic effect
among the toxaphene components.
Landrum and coworkers (1976) also present data on the toxicity of toxaphene
fractions and two toxaphene components to houseflies (Table 5.67). Component 1
in this study is the same heptachloroborane designated as toxicant A by Ohsawa
and coworkers (1975). Landrum and coworkers (1976), however, found this compo-
nent to be less toxic relative to toxaphene than did the previous study. None-
theless, both studies indicate that this heptachloroborane is more toxic than
technical toxaphene. The other component identified by Landrum and coworkers
(1976), a heptachlorodihydrocamphene, is significantly less toxic than technical
toxaphene.
Three additional polychloroborane components of toxaphene, which have
been isolated but not yet absolutely identified, were found to be 1.75 to 3.5
times more toxic than toxaphene on topical application to houseflies (Anagnostopoulos
et al., 1974).
5.6.2.2.2 Resistance in insects — Resistance to toxaphene has been demonstrated
in several orders of insects including: lepidoptera, coleoptera, diptera,
hymenoptera, and hemiptera. Although quantitative estimates of such resistance
vary markedly and are sometimes difficult to compare, resistant insect strains
are often able to tolerate doses of toxaphene ten times above those causing
pronounced mortality in corresponding susceptible strains. For target organisms,
such as the lepidopteran, some coleopteran, and hemipteran species, the develop-
ment of resistance can severely affect the efficacy of toxaphene as a control
agent.
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Table 5.6 7. The topical toxicity of toxaphene, two column fractions, and two
components'1
House fly
Compound LD50 relative
or mixture LD50 (mg/kg) to toxaphene
Toxaphene
Column Fraction 3
Column Fraction 7
Component 1
Component 3
b
33(30-35)
40(37-44)
50(44-55)
31(27-36)
40(36-43)
1.0
c
0.83
0.66°
1.10
0.83C
/Source: Landrum et al. , 1976.
The values in parentheses represent 95% confidence limits.
/Relative LDSO's which differ significantly (p = 0.05) from toxaphene.
Same as Toxicant B, Table 5.61.
2,4,6-exo,8,8,9,10-heptachlorodihydrocamphene.
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A number of lepidopteran larvae of the noctuidae family have been shown to
be resistant to toxaphene or toxaphene:DDT combinations. In Texas populations
of Heliothis virescens (the tobacco budworm), the development of toxaphene resis-
tance followed the development of DDT resistance. In 1963, this species was re-
sistant to DDT but still highly susceptible to DDT:toxaphene combinations (Brazzel,
1963 and 1964). However, between 1963 and 1965, the LD50 value of a 2:1 toxaphene-DDT
mixture increased by a factor of six, going from 0.57 mg/g larva to 3.52 mg/g
larva. A similar pattern of resistance was also seen in the closely related
pesticide, Strobane (Adkisson, 1967). Over about the same period, a four-fold
increase in LD50's was seen for the same species in Louisiana, with the resistant
strain having a LC50 of 4.3 to 8.6 micrograms/larva (Graves et al., 1963 and 1967).
In a 1970 Mississippi study, which compared the toxicity of a 2:1 toxaphene-DDT
mixture to Heliothis virescens, resistant strains were found in two of the five
sample locations and had values of 450 micrograms/larva and over 1000 micrograms/larva.
Comparable values in the strains designated as susceptible ranged from 34 to
52 micrograms/larva. These results indicate that the resistant strains tolerated
9 to 29 times more toxaphene than the susceptible strain (Harris, 1972). However,
this study differs in an important respect from the Texas and Louisiana studies
in that the susceptibility of the populations was not measured over a period of
time. Thus, while the variability in the Heliothis virescens population found
by Harris (1972) clearly suggests the potential for resistance to develop, the
actual development of resistance was not demonstrated. In the same study, no
such variability was seen in five populations of Heliothis yea (the bollworm).
Similarly, in an Australian study, Wilson (1974) has demonstrated a five-fold
resistance to toxaphene-DDT in some populations of Heliothis armigera, but no
resistance in Heliothis punctigera.
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Resistant populations of the Egyptian cotton leafworm (Prodenia litura),
also a noctuid, were found during the early 1960's by Ali and coworkers (1962).
The extent of resistance ranged from factors of about 8.5 to 24. However, in
a follow-up study, both laboratory-reared progeny of the resistant strains and
field populations showed an apparent reversal of toxaphene resistance. This
reversal occurred after toxaphene use had been discontinued, with carbaryl and
trichlorfon being used as substitutes (Eldefrawi et al;, 1964 b).
Two species of coleoptera, Anthonomus grandis (the boll weevil) and
Tribolium castaneum (a flour beetle), also have demonstrated resistance to
toxaphene. As in studies on the tobacco budworm, an early study in five boll
weevil populations in Alabama showed no evidence of developing toxaphene re-
sistance (Burkhalter and Arant, 1960). However, after a couple of years, re-
sistant populations were reported in Georgia and Texas (Brazzel and Ship, 1962;
Graves and Roussel, 1962; Tippins and Beckham, 1962). In the Texas study the
magnitude of resistance in the field populations varied from negligible to a
factor of 142, when compared to a laboratory-reared susceptible strain with
a LD50 of 7.6 micrograms/weevil (Brazzel and Ship, 1962). In a Louisiana
study, no evidence was found for the development of resistance in field popu-
lations of boll weevils. However, Graves and Roussel (1962) did demonstrate
a 10- to 12-fold increase in the LD50 of a 2:1 toxaphene-DDT mixture in an
experimental population of boll weevils exposed to this mixture for 22 genera-
tions. Although continued exposure for 20 additional generations did not result
in further resistance, the toxaphene-DDT resistant strain was found to be cross-
resistant to three organophosphates and two carbamates.
Some attempt has been made to determine the physiological basis of toxaphene
resistance in the boll weevil. In comparing the lipid levels of boll weevils
5-172
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surviving and succumbing to a given toxaphene exposure, Reiser and coworkers (1953)
found that surviving animals consistently had about 10 percent more fat per unit
body weight. In a study on both laboratory-reared and field collected boll
weevils, Moore and coworkers (1970) associated boll weevil survival after toxa-
phene exposure to higher levels of palmitic and oleic acids and lower levels of
stearic, linoleic, and linolenic acids than were found in weevils succumbing to
exposure. In a subsequent study, Moore and Taft (1972) have associated sur-
vival with significantly higher levels of triglycerides. Further, these in-
vestigators found significantly higher levels of triglycerides in untreated
controls than in surviving animals, suggesting increased triglyceride use by
the animal during toxaphene exposure. No relationship was found between sur-
vival and phospholipid levels (Moore and Taft, 1972).
Toxaphene resistance in Tribolium castaneum, another coleoptera, has been
demonstrated only in a laboratory-reared, lindane-resistant strain. This lindane-
resistant strain demonstrated more than a 9-fold cross-resistance to toxaphene,
compared to the lindane-susceptible strain (Bhatia and Pradhan, 1972). No such
cross-resistance toxaphene was seen in a DDT-resistant strain of the same species
(Bhatia and Pradhan, 1970).
Information on toxaphene resistance in other pest orders is somewhat sketchy.
Grayson (1953) reports the development of marginal resistance to toxaphene in
the hemipteran Oncopeltus fasciatus (the milkweed bug) after laboratory rearing
of 17 generations exposed to sublethal levels of toxaphene. Based on 10-second
emersions of the insects in suspensions of toxaphene, the selected strain had
a LC50 of 0.345 g/liter and the control strain a LC50 of 0.190 g/liter. In
studies of pesticide resistance in the body louse, Pediculus humanus, both
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Eddy (1952) and Busvine (1953) indicate that DDT-resistance does not confer
cross-resistance to toxaphene. However, DDT-resistant houseflies, Musca domestica...
have been shown to be cross-resistant to toxaphene and Strobane (Bodenstein and
Fales, 1962; Pratt and Babers, 1950). Quantitative estimates of resistance
cannot be made based on the data presented in either of these studies.
Toxaphene resistance in beneficial insects also has received relatively
little study. Atkins and Anderson (1962) demonstrated that DDT-resistant honey
bees, Apis snellifera, are remarkably cross-resistant to toxaphene. Toxaphene
exposures killing fifty percent of the DDT-susceptible bees in 140 to 163 hours
caused no mortality above control levels in DDT-resistant bees over periods
greater than 560 hours. In Bracon mellitor, a hymenopteran parasite of the
boll weevil, no resistance to toxaphene could be demonstrated over a five-
generation selection exposure study. However, under the same conditions, re-
sistance did develop to a 1:1 toxaphene-DDT mixture and to DDT (Adams and Cross,
1967). Thus, those studies cited above which indicate a resistance to toxaphene-
DDT combinations are not definitive proof of the developed resistance to toxa-
phene alone.
5.6.2.2.3 Synergistic effects in insects — In the control of agricultural
pests, toxaphene has been frequently used in combination with other organo-
chlorine pesticides such as DDT or with organophosphorous insecticides such as
methyl parathion. While some field studies indicate that such combinations
are more efficacious than toxaphene alone (Section 5.6.2.2.5), relatively few
attempts have been made to experimentally demonstrate synergism between toxa-
phene and these other pesticides. Only about half of these studies provide
sufficient data for quantitative estimates of synergistic effects.
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A variety of methods are available for the analysis of joint toxic action.
Although some entomologists (e.g., Atallah and Newsom, 1966; Wolfenbarger, 1973;
Wilson, 1974) seem to prefer the calculation of co-toxicity coefficients pro-
posed by Sun and Johnson (1960) using houseflies, this reviewer will use Finney's
(1952) model for the calculation of an expected LC50 based on the joint harmonic
mean of the LD50 for mixtures (Table 5.68). This method has enjoyed wide use by
toxicologists in the analysis of many pesticide and nonpesticide formulations
(e.g., Ohsawa et al., 1975; Smyth et al., 1969). The ratio of expected LD50
to observed LD50 is used as the quantitative estimate of synergism, with ratios
of less than 1 indicating a less than additive effect and values greater than
1 indicating a more than additive effect. Using this method to analyze syner-
gistic effects in all possible pairs of 27 chemicals, Smyth and coworkers (1969)
noted, based on an analysis of ratio variance, that ratios below 0.40 suggest
antagonistic effects, while ratios above 2.70 suggest actual synergisms. Inter-
mediate values (0.40 to 2.7) were attributed to random variation. Although such
significance estimates are not necessarily valid for all groups of chemicals,
they may serve as useful guides in the analysis of toxaphene synergisms.
Studies on toxaphene synergism from which ratios of expected LD50 values
may be calculated are given in Table 5.68. In several studies, an apparent
synergism exists between toxaphene and DDT. All four Heliothis species,
important cotton pests, show this effect which is also seen in the honey bee
and some strains of boll weevil.
In the boll weevil study, some inferences can be drawn concerning the
relationship of resistance to synergism. Two strains were used, one sensitive
to DDT and toxaphene and the other sensitive to DDT but resistant to toxaphene.
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Table 5.68. Synergistic effects of toxaphene with DDT and methyl patrathlon on topical application
Ui
I
Toxa-
phene ;sy nurgis t
Synergist ratio Organism
DDT 2 : 1
2:1
1:1
1:1
2:1
1:1
1:2
2:1
2:1
2:1
Anthonmis grandi a
(Boll weevil)
Suscept ible
Resistant
Apis melli fera
(Honey bee)
Heliothis armipera
Ord strain
Durham strain
Heliothis puac tizea
Ord strain
Heliothis virescens
(Tobacco budworm)
Heliothis virescens
(Tobacco budworm)
Heliothis virescens
(Tobacco budworm)
Hello this virescens
(Tobacco budworm)
Heliothis virescens
(Tobacco budworm)
Heliothis virescens
(Tobacco budworm)
48 hourfl
I.D50
toxaphene
(0
(6
19
22
13
3.
(5
(5
(5
.240)
.80)
.08
.53
.61
70
.02)
.02)
.02)
346.0
78.67
82
.86
48 hour
LD50
synergist
(3
(2
7.
80
12
0.
(4
(4
(4
1.
1.
17
.00)
.40)
91
.71
.24
88
.41)
.41)
.41)
02
26
.92
Observed
L050 of
combination
(0.360)
(0.750)
4.4
10.56
4.05
0.30
(0.21)
(0.49)
(1.54)
0.54
1.40
3.34
6
Expected
LD50 of
combination
0.
4.
13.
35.
12.
1.
4.
4.
4.
2.
2.
29.
265
263
10
23
89
42
85
70
61
03
48
47
Ratio of
expected
to observed
joint
toxi ci ty
(E/0)
0.
5.
2.
3.
3.
4.
23.
9.
2.
3.
1.
8.
74
68
98
33
18
73
09
59
99
76
77
82
Re f erence
Roussel et al. , 1959
Graves and Mackensen, 1965
Wilson, 1974
Wilson, 1974
Wolfenbarger, 1973
Wolfenbarger, 1973
Wolfenbarger, 1973
Guzman-Varon et al. , 1974
Guzman-varon et al. , 1974
Guzman-Varon et al. , 1974
Expected value calculated as harmonic mean, i.e., I/expected LC50 = Pt/LD50 of toxaphene + Ps/LD50 of synergist where Pt and Ps are proportions of
(jtoxaphene and synergist, respectively. v * "uno ui
Numbers in parenthesis indicate units of mg/g. All other doses are in units of micrograms/lnsects.
-------
Table 5.68. (continued)
Ln
I
Toxa-
phene : synergic t
Synergist ratio Organism
DDT
fiethyl
p a r a t h i on
2:1 Heliothis zea
(Bollworm)
c l.ygus hesperus
1960
1961
1962
2:1 Pectinophora gossypiella
(Pink bollworm)
DDT-resistant
2:1 Anthonomus grandis
(Boll weevil)
1973
1974
18 hour"
LD50
toxaphene
(0.32)
0.463
1.038
0.737
0.534
24.5
20.3
48 hour
LD50
synergist
(2
0.
0.
0.
1.
0.
0.
.71)
894
258
877
164
061
056
Observed
UJ50 of
combination
(0
0.
0.
0.
0.
0.
0.
.12)
390
252
597
615
194
123
Expected
LD50 of
combination
0.
0.
0.
0.
0.
0.
0.
45
610
413
801
658
,184
.169
Ratio of
expected
to observed
joint
toxici ty
(E/0)
3.83
1.56
1.64
1.34
1.06
0.95
1.37
Reference
BraifEel, 1964
Leigh and Jackson, 1968
Lowry and Berger, 1964
Hopkins et al. , 1975
a
Expected value calculated as harmonic mean, i.e., I/expected LC50 = PC/LD50 of toxaphene + Ps/LD50 of synergist where Pt and Ps are proportions of
, toxnphene and syaergis t, respectively.
Cumbers in parenthesis indicate units of mg/g. All other doses are in units of micrograms/insects.
Not specified, 1:1 assumed in calculation of predicted LC50.
-------
Marked synergism is noted only in the toxaphene-resistant strain. The increased
synergistic effect in this strain was associated with relatively flat and slightly
diverging log dose-probit response lines for all applications. In the toxaphene
sensitive strain, however, the dose-response line was flat only for the DDT
application. The corresponding lines for both toxaphene and toxaphene-DDT appli-
cations were much steeper and almost identical to each other (Roussel et al.,
1959). This seems to suggest that toxaphene and DDT interacted very weakly,
if at all, in the toxaphene-sensitive strain, with most of the lethality
attributable directly to the toxaphene. In the study on Heliothis armipera,
the Ord strain was more resistant than the Dunham strain to both toxaphene and
DDT. However, the levels of synergism in the two strains were about the same
(Wilson, 1974). In the tobacco budworm, the most marked synergism is seen in
the strain resistant to DDT (Guzman-Varon et al., 1974).
The effect of toxaphene:DDT ratio on synergism has been examined only by
Wolfenbarger (1973). Using a strain of Heliothis virescens about equally sensi-
tive to both toxaphene and DDT, this investigator demonstrated that both toxicity
and the degree of synergism were directly related to the proportion of toxaphene
in the toxaphene-DDT mixture. However, with the boll worm, Heliothis zoa, the
highest toxicity was achieved with a 1.1:toxaphene:DDT mixture. Because a
LC50 for toxaphene to this species was not given, details of this study are not
summarized. However, using a modification of the Sun and Johnson (1960) method
and assuming negligible toxaphene toxicity, Wolfenbarger (1973) noted increasing
synergistic behavior as the proportion of toxaphene decreased.
The remaining information summarized in Table 5.68 suggests no synergistic
effect between toxaphene and DDT in Lygus hesperus or in a DDT-resistant strain
of Pectinophora gossypiella. Further, Hopkins and coworkers (1975) were not able
5-178
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to demonstrate a synergistic effect between toxaphene and methyl parathion in
the boll weevil. These results are consistent with field studies which indica-
ted that toxaphene does not usually enhance the protective effects of methyl
parathion on boll weevil infestations of cotton (Hopkins et al., 1970).
The synergism of dimethyl sulfoxide and toxaphene in boll weevils has
been examined by Moore and coworkers (1970). In this study, toxaphene, DDT,
and a 2:1 mixture of toxaphene:DDT were applied to a 90 mm disc of filter
paper. Boll weevils were left in contact with the treated paper for 48 hours,
at which time mortality was recorded. In a parallel series of exposures,
40 percent dimethyl sulfoxide was added to each of the three test preparations.
The 48-hour LDSO's without dimethyl sulfoxide were 178, 3000, and 1.7 mg
insecticide/90 mm filter paper for toxaphene, DDT, and (2:1) toxaphene-DDT,
respectively. Using the harmonic mean, these results indicate a synergistic
ratio of 154. While supporting the evidence for toxaphene-DDT synergism
outlined above, this high synergistic ratio may not be comparable to those
described in Table 5.68 because of the differing route of administration. The
addition of dimethyl sulfoxide resulted in the following LDSO's for toxaphene,
DDT, and (2:1) toxaphene-DDT: 141, 1600, and 14.7. While a slight increase
in toxicity is apparent for both toxaphene and DDT, the synergistic ratio of
the toxaphene-DDT mixture is lowered to about 14.
Although other studies are available in the joint toxicity of toxaphene
with some other pesticide, these studies do not provide sufficient data for
an analysis of synergism. Many such studies described the development of re-
sistance to toxaphene-DDT combinations (Section 5.6.2.2.2). Others (e.g.,
Lentz et al., 1974; Rajak and Perti, 1967) are generally screening studies
concerned with the development of more efficient pesticide combinations. This
aspect of toxaphene synergism is discussed in Section 5.6.2.2.5.
5-179
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5.6.2.2.4 Reproductive effects in insects — Remarkably little is known about
the long term effects of toxaphene exposure to insects. As reflected in the
preceding sections, most of the research generated on toxaphene has been con-
cerned with assessing the usefulness of toxaphene as an insecticide and, secon-
darily, in determining the immediate effects that such use might have on bene-
ficial insects - e.g., bees and certain parasitic wasps. Consequently, the
ability to assess the effects of background levels of toxaphene contamination
on insects is limited.
Toxaphene may severely affect the reproductive potential of lady beetles,
Coleomegilla maculata (Atallah and Newsom, 1966). After application of single
topical doses of toxaphene as low as 10 micrograms per beetle - about 820 micro-
grams/gram - twenty mated pairs of beetles placed in oviposition cages failed
to produce eggs during the remaining life span of the female. Under the same
conditions, untreated pairs showed normal egg laying. However, egg production
in beetles surviving single doses of a 2:1 toxaphene-DDT mixture at 7.5 micro-
grams per beetle was not significantly decreased, although the survival potential
of the F-l progeny was reduced by 40%. These results would seem to suggest that
DDT blocks the adverse reproductive effects of toxaphene or that the reproduc-
tive failure in the toxaphene-only treated group was not due to toxaphene itself.
In a similar study in which milkweed bugs, Oncopeltus fasciatus, were ex-
posed to sublethal levels of toxaphene for 17 bug generations, no significant
reduction was seen on the average number of eggs per female (Grayson, 1953).
5.6.2.2.5 Field studies — Toxaphene, either alone or in combination with
other insecticides, has been used as a pest control agent on a number of crops
including: alfalfa, cotton, corn, sorghum, soybeans, peanuts, citrus fruits,
5-180
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and various vegetables. An extensive review of these applications giving
details on target organisms and recommended application procedures is avail-
able from Hercules Incorporated (1972). The efficacy of such application is
related to three factors: the suppression of pest species; the effect on
beneficial species, such as pollinators or pest predators; and the phototoxic
effects of the pesticide application. The phytotoxicity of toxaphene has been
discussed in Section 4.0. This section will focus on the effects of toxaphene
field applications on pest insects and beneficial insect species.
5.6.2.2.5.1 Pest insects — Toxaphene is recommended for the control of numerous
insect pests (Hercules Incorporated, 1972). Published reports on the effective-
ness of toxaphene applications are summarized in Table 5.69. Similar to the
estimates of acute toxicity given in Table 5.66, the effects of toxaphene field
applications vary considerably from species to species. For instance, toxaphene
significantly reduced the Adelphocoris superbus infections in alfalfa fields
after toxaphene spray applications of .45 - 1.1 kg/ha (Lilly and Hobbs, 1962),
while much higher spray applications of up to 22 kg/ha gave inferior control
of Amphimallon majalis on meadow and pasture sod (Shorey et al., 1958). However,
such field studies are particularly difficult to compare to each other. A
number of meteorological factors, such as humidity, sunshine, dew, wind, and
rain, affect the results of a given field study (Gaines and Mistric, 1952).
In most of the studies summarized in this section, sufficient details are not
included on these factors to determine their significance in the effectiveness
of control. A number of physiological and behavioral characteristics of the
insects may also influence the effectiveness of insecticide applications. For
example, Gaines and Mistric (1952) have found that toxaphene is more effective
5-181
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Table 5.69. Efficacy of toxaphene in the control of various insect pests
L/l
1
M
CO
N3
Pest
Aconita dacia
(Brown cotton leafwonn)
Adelphocoris superbus
(Superb plant bug)
Agrotis orthogonia
(Pale western cutworm)
Agrotis ypsilon
(Black cutworm)
Alabama arglllacea
(Cotton leafworm)
Amphlmallon majalis
(European chafer)
.
Dose
Crop (kfl/lia) Comments
Cotton 2.8 Effective control for up to 6 days
Alfalfa 0.45 - 1.1 Significantly reduced infestations
Winter wheat 1.1 Tailed to give practical control
within 7 days
Cotton 1. 7 Very effective for up to two weeks
Cotton 2.2 Satisfactory control, 92.8% mortality
after 48 hours
Meadow and pasture 22.0 Inferior control during year of appli-
sod cation
Ref e rence
Lloyd and Martin, 1956
Lilly and Hobbs , 1962
DePew and Harvey, 1957
Kamel and Shoeb, 1958
Adkisson, 1958
Shorey et al. , 1958
Blissus leucopterus
(Chinch bug)
Bothynoderes punctiventris
(Sugar beet: weevil)
Chaetoenema pulicaria
(Corn flea beetle)
Coleophora alcyonipennella
(Clover case bearers)
Sorghum
Sugar beets
Sorghum
White clover
3.3
1.0
2.2
17.0
Good control for over one week
Up to 100% control
75% control for 11 days in 1958 less
effective in 1960
Effective control
Randolph and Newton,
1959
Hanolache et al., 1965
Henderson et al., 1962
Hoy, 1960
-------
Table 5.69 (continued)
Ln
|-j
OO
U)
Pest Crop
Collnis nit-ida
(June beetle) Clover
Eutrombicula alfreddugesi
(a chigger) Woodlands
Frank li niella sp.
(Tlirips) Cotton
Frankiiniella fusca
(Tlirips) Peanuts
Frankliniella fusca
(Tlirips) Peanuts
lleliothls zea
(Boll worm) Lima beans
lleliothls zea
(Boll worm) Cotton
Kotochalla junodi
(Wattle bagworm) Acacia
Lygus hesperus
(Lygus bugs) Lima beans
Dose
(kg/ha)
2.0
2.2
0.84
1.7
1.7
2. 46
1.7
2,52
3.3
Comments
Effective for first inatar larva,
erratic control of second Instar
Very effective for 4 weeks
Effective control
90.7% reduction in pest
Significant thrip mortality but no
increase in crop yield
Adequate control
58% kill in 48 hours
Nearly complete mortality 10 to 14 days
Effective control for up to one month
Reference
Davich et al. , 1957
Keller and Couck , 1957
Davis et al. , 1958
Dogger, 1956
Morgan et al. , 1970
Stone et al. , 1960
Nemec, 1972
Ossowski, 1958
Stone et al. , 1960
-------
Table 5.6 9.(continued)
Ln
1
fast: Crop
Psallus seriatus
(Cotton flej liopper) Cotton
Pyrausta nubilalis
(European corn borer) Peppers
Rhapalosiphum raaidis
(Corn leaf aphid) Sorghum
Dose
(kg/ha) Comments Reference
0.84 Effective control Davis et al. , 1958
2.2 Infestation reduced by 50% Hof master et al. , 1960
4.9 78% control by 5 days after application Henderson et al. , 1964
CO
-------
in the control of boll weevils when applied early in the growing season. As
discussed in Section 5.6.2.2.1, the dietary history of the insect may be
important in this effect. The type of toxaphene formulation may also influ-
ence the efficacy of toxaphene in a particular species. Dogger (1956) found
that 173 kg/ha of toxaphene spray produced a 90.7 percent reduction of a
Frankliniella species on peanuts. Granulated formations of 10 percent toxa-
phene, at 224 kg/ha, however, produced only a 64.8 percent reduction in these
things. Lastly a variety of different methods are used to assess toxaphene
efficacy. While some studies report efficacy in terms of increased crop
yield, others use estimates of mortality based on field surveys over periods
of a few days to several months. Thus, these field studies, while providing
practical information on toxaphene utility, are not readily comparable to each
other or to the toxicity estimates given in Section 5.6.2.2.1.
Similar field studies on the efficacy of toxaphene in combination with
other insecticides are summarized in Table 5.70. Toxaphene-methyl parathion
mixtures are frequently used, and toxaphene-DDT combinations have been commonly
used in previous years. As with field studies using only toxaphene, consider-
able variation is apparent among the different species. The tobacco budworm,
Heliothis virescens, is only 30 percent controlled by a 1.7 and 2.2 kg/ha
application of toxaphene and methyl parathion (Nemec, 1972), while a .56 and
1.1 kg/ha toxaphene-methyl parathion application was effective in the control
of the boll worm, Heliothis zoa (Turnipseed et al., 1974). A formulation con-
taining a greater amount of toxaphene (1.7 and .56 kg/ha toxaphene-methyl
parathion) also produced high (97 percent) mortality in Heliothis zoa after
48 hours (Nemec, 1972).
5-185
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Table 5.70. Efficacy of toxaphene in combination with other insecticides
a
Pest
An tlionomus grandis
Anthonomus grandis
Anticarsia gemmatalis
Franklinie lla sp.
Heliothis sp.
Heliothis virescens
Ln
1
£- lleliothis zea
Heliothis zea
Crop
Cotton
Cotton
Soybeans
Cotton
Cotton
Cotton
Soybeans
Cotton
Application
(kg/ha) Comments
2.2 T + 1.1 MeP Superior control
2.2 T + 1.1 DDT 100% kill in 48 hours
0.56 T +1.1 MeP Minimum effective rate
0.84 T + 0. 28 MeP Effective but no improvement over
toxaphene alone; MeP alone had
no effect
2. 2 I + 1.1 MeP Significant reduction in boll damage,
not much more effective than MeP
1.7 T + 2.2 MeP 30% kill in 48 hours, significantly
more effective than MeP alone but
no more effective than toxaphene
alone
0.56 T + 1.1 MeP Minimum effective rate
2.2 T + O.S4 MeP Satisfactory control, better than
Reference
Harding, 1970
Nemec, 1972
Turnipseed et
Davis et al. ,
Harding, 1972
Neinec, 1972
Turnipseed et
Hopkins et al
al. , 1974
1958
al., 1974
. , 1970
Heliothis zea
Cotton
1. 7 T + 0.56 MeP
MeP alone
Almost complete kill within 48 hours,
not much better than 0.84 MeP
Nemec, 1972
Heliothis zea
Heliothis zea
Cotton
Cotton
1.7 T + 0.
1. 7 T + 0.
,56
,56
Ph
G
Almost complete kill.
or Ph alone
90% kill in 48 hours,
or G alone
better
better
than T
than T
Nemec, 1972
Hemec, 1972
flKEY: T - toxaphene; MeP = methyl parathion; G = galuron; Ph = phosvel; M = malathion.
-------
Table 5.70 (continued)
Pest
lleliothis zea
lleliothis zea
Lygus hesperus
Lygus sp.
Lygus sp.
Plathypenascaba
1 i Psallus seriatus
CO
SpodopLera littoralis
Crop
Cotton
Cotton
Cotton
Lima beans
Lima beans
Soybeans
Cotton
Cotton
Application
(kg/ha)
1. 7 T + 0.56 EfN
2. 2 T + 1.1 DDT
3. 3 T + 1. 1 M
3. 3 T + 1.1 M
3. 3 T + 1. 7 DDT
0.56 T + 1.1 MeP
0. 84 T + 0. 28 MeP
0. 84 T + 0. 36 M
3. 9 T + 1. 0 EPN
Comments
87% kill in 48 hours; better than
T alone but not as good us 1.1 EPN
alone
30% kill in 48 hours; not as good as
toxaphene alone
Effective control
Good control for 3 to 4 weeks, better
than toxaphene alone
Less yield than with M combination
Minimum effective rate
Both M and MeP combinations increased
cotton yield by 67 kg/ha
86% control after 3 days
Nemec,
Nemec ,
Falcon
Shorey
Shorey
Reference
1972
1972
et al. , 1968
et al. , 1965
et al. , 1965
Turnipseed et al. , 1974
Davis
Kamel
et al. , 1958
and Mitri, 1970
KJiV: T = toxaphene; MeP = methyl parathion; G - galuron; Ph = phosvel; M = malathion.
-------
Toxaphene combinations with DDT have given mixed results. In Spodoptera
littoralis, the Egyptian cotton leafworm, a 3.8, 1.9, and .66 kg/ha combi-
nation of toxaphene, DDT, and methyl parathion was almost twice as effective
initially as a 5.5 and 1.5 kg/ha combination of toxaphene and methyl parathion
(Kamel and Mitri, 1970). Shorey and coworkers (1965), however, found that a
toxaphene-DDT mixture was much less effective in the control of several Lygus
species than a similar mixture of toxaphene malathion. Similarly, in the con-
trol of Heliothis zoa, a 1.7 and .56 kg/ha mixture of toxaphene and methyl
parathion provided a 97 percent kill on cotton in 48 hours, while a 2.2 and
1.1 kg/ha application of toxaphene-DDT caused only 30 percent mortality over
the same period (Nemec, 1972). As indicated in Table 5.70, toxaphene combined
with organophosphorous insecticides is often effective in the control of many
agricultural insect pests. However, as discussed in Section 5.6.2.2.3, quanti-
tative estimates of synergism for these combinations are not available.
5.6.2.2.5.2 Beneficial insects — The limited toxicological data - i.e., LD50
estimates - available on the beneficial insects have been summarized in
Table 5.64 and discussed in Section 5.6.2.2.1. These beneficial insects are
mostly hymenopterans including two major groups, the bees and the parasitic
wasps. In addition, some information was presented on the lady beetle, a
coleopteran. However, most of the information on the effects of toxaphene on
beneficial insects comes from field studies in which toxaphene was applied and
observations made on the effects of treatment on nontarget organisms.
Several insecticide combinations which include toxaphene have been used
in the control of pests on alfalfa. Since bees are essential to the pollina-
tion of this crop, the control measures must be relatively harmless to bees.
5-188
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Johansen and Eves (1967) determined the sensitivity of three species of bees
to mixtures of toxaphene and three other pesticides (Table 5.71). Both toxa-
phene-DDT mixtures and toxaphene-oxydemetonmethyl (Meta-Systox-R) caused sig-
nificant mortality. Toxaphene-systox caused only slight mortality in the alkali
bee (Johansen and Eves, 1967). Toxaphene alone was also found to be moderately
toxic to the leafcutting bee. Exposure to foliage which had been treated at
3.3 kg/ha toxaphene three hours prior resulted in 35 to 76 percent mortality
after 24 hours. When exposure was delayed until 24 and 48 hours after treat-
ment, resulting 24-hour mortalities dropped to 41 to 53 percent and 29 percent,
respectively (Johansen et al., 1963). In a similar study, conducted in Egypt,
honeybees were exposed to cotton plants which had been sprayed with a 60 percent
emulsion of toxaphene at 8.6 liters/ha. Plants held for two or more days
before exposure to bees caused less than 10 percent mortality (Wafa et al., 1963).
Ibrahim and coworkers (1967), in a related study, assessed the contact toxicity
of toxaphene sprays to the honeybee. Toxaphene concentrations of 0.24 to
0.396 percent caused no deaths during six-hour exposures at application rates
of 7.4 to 12 liters/ha.
Field studies also indicate that toxaphene is not extremely toxic to
bees. Palmer-Jones and coworkers (1958) found no evidence of bee mortality in
clover fields treated with toxaphene at 5.5 kg/ha. Only very low bee mortali-
ties (0.77 percent) were seen in fields treated weekly with 4.4 to 17.7 kg/ha
toxaphene for eight weeks (Weaver, 1950). However, toxaphene may have a re-
pellant effect on bees. Atkins and coworkers (1970) noticed a decrease in the
frequency of bee foraging in fields that had been treated with 4.4 kg/ha
toxaphene. Similarly, a 50 percent drop in foraging by honeybees was seen in
an alfalfa field sprayed with 3.3 kg/ha toxaphene (Todd and Reed, 1969).
5-189
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Table 5.71. Alkali bee, leafcutting bee, and honey bee 24-hour percent mortalities from insecticide
treatments on alfalfa, Pullman, Washington, 1966a
Caged with treated foliage, age of residues
Alkali bee Leafcutting bee Honey bee
Material kg/ha 3hr 1 day 3 days 5 dayu 3 hr 1 day 3 day:; 5 days 7 days 3 hr 1 day 3 days
Toxapliene 3.6 kg E 3.3+
+ Demeton (Systox)
0.9 kg E 0.28 10 0
Toxaphene 3.6 kg E 3.3+
+ Oxydemetonmethyl
I (Meta-Systox-R) 0.9 lcE E 0.28 26 5 — — 42 18 — — — 10 2
MD
O
Toxaphene - DDT
0.9 - 0.6 kg E 3.3-1.7 — 40 12 0 -- 96C 76 24 8 — 22
/^Source: Johansen and Eves, 1967.
Cooler temperature at the time of application caused greater residual action than in Toxaphene-Demeton test.
Daily maximum temperatures were 62 to 75 F during this alfalfa leafcutting bee test.
-------
The reports on the effects of toxaphene on entomophagus insects - i.e.,
beneficial insects which prey on pest insects - generally indicate that toxaphene
adversely affects these animals. Fenton (1959) noted that field treatments with
toxaphene (3.3 kg/ha) and several other pesticides caused marked mortality in
both beneficial and pest species. However, because of the resulting decrease
in entomophagus insects, populations of pest species on treated plots subse-
quently increased above those in control plots. In cotton and alfalfa fields
treated with toxaphene at 2.8 to 3.2 kg/ha, Orius, Geocoris, Nabis, and
Hippodamia predator species were substantially depressed. Treatments with a
2:1 toxaphene-DDT mixture at 4.9 kg/ha had a somewhat greater effect (van den
Bosch et al., 1956). Similar reductions in these insects and in Sinea diadema
and syrphids were noted in cotton fields treated with 1.5 and 2.9 kg/ha toxaphene-
DDT mixtures (Stern et al., 1959). Multiple treatments of cotton fields with
toxaphene at 11.1 to 17 kg/ha drastically reduced populations of syrphids,
assassin bugs, lady beetles, flower bugs, lace wings, and Geocoris species
(Gaines, 1955). More recently, Walker and coworkers (1970) have demonstrated
that toxaphene treatments of cotton fields initially caused an 80 percent
decline in Hypodamia convergens, Orius insidiosus, and a Seyminus species. By
three weeks after treatment, populations of these insects were still 60 percent
below those in control plots. Shepard and Sterling (1972) have also shown that
toxaphene applied to cotton at 1.1 kg/ha reduced by 50 percent the population of
various insect predators, including Orius, Seyminus, Hippodamia, Ceratomeyilla,
Chrysopa, Nabis, Podisus, Collops, Zelus, and Geocoris species.
5.6.2.2.6 Toxicity to other terrestrial invertebrates — A limited amount of
information is available on the toxicity of toxaphene to millipedes, spiders,
5-191
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worms, and snails. Of the three species of millipedes tested, only Spinotarsus
fiedleri seems susceptible to toxaphene. This species experienced 60 percent
mortality by seven days after being fed meal containing 0.2 percent toxaphene.
Two other millipede species, Poratophilus pretorianus and Foratophilus robustus^
experienced no mortality after identical treatments (Fiedler, 1965). Redmond
and Brazzel (1968) tested the toxicity of a 2:1 toxaphene-DDT mixture to the
striped lynx spider, Qxyopea salticus. On topical application of 1 microliter
drops, the 72-hour LCSO's for two spider populations were 0.069 and 0.079 micro-
grams /milliliter. Sufficient information is not given in this report to convert
the LC50 values to LD50's in micrograms toxaphene/gram spider. In a field study,
spider populations were initially depressed by 70 percent and remained 50 percent
below control levels one month after treatment of a cotton field with toxaphene
at 1.1 kg/ha (Walker et al., 1970). As discussed in Section 5.6.2.2.5.2, similar
effects were noted on beneficial insect species. Decreases in worm populations
have also been seen in worms treated with toxaphene at 6.8 kg/ha (Hopkins
and Kirk, 1957) and 1.7 kg/ha (Legg, 1968)- Toxaphene has also been shown
to cause some mortality in European brown snails at concentrations of 80,000 ppm
in bran. No effects, however, were seen at concentrations of 20,000 ppm or
5,000 ppm (Pappas and Carman, 1955).
5-192
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/
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5-209
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6.0 BIOLOGICAL ASPECTS IN HUMANS AND LABORATORY ANIMALS
6.1 CHAPTER SUMMARY
In evaluating the potential hazard of toxaphene to man and other mammals,
carcinogenicity should be a major concern. Strobane, a pesticide closely re-
lated to toxaphene, has been shown to cause a high rate of liver tumors in
mice (Innes et al., 1969). A study by the National Cancer Institute (1979)
found that toxpahene, at dietary levels of around 100 ppm given over a one and
a half year period, is associated with a high incidence of hepatocellular
carcinoma in mice. Rats also evidenced a substantial increase in the incidence
of some malignant tumors.
Teratogenicity and other reproductive effects are additional areas of
concern. In rats, toxaphene given on days 7 to 16 of gestation resulted in an
increased incidence of encephaloceles and resulted in high maternal mortality.
Thus, this effect would be of most concern in cases of accidental poisoning of
pregnant females. However, even at lower doses to rats and mice - 15 and
25 mg/kg/day - dose related changes were seen in implantation frequency, fetal
mortality, and fetal weight gain. In rats only, decreases were seen in sternal
and caudal ossification centers (Chernoff and Carver, 1976). Toxaphene has
not been shown to be mutagenic using the dominant lethal assay in mice
(Epstein et al., 1972).
In other respects, toxaphene is similar to many other chlorinated pesti-
cides in its biological behavior. Toxic amounts of toxaphene may be absorbed
across the alimentary tract, skin, and respiratory tract. Toxaphene seems to
be distributed throughout body tissues but is stored primarily in the fat
(Crowder and Dindal, 1974; Clapp et al., 1971). Toxaphene undergoes dechlorina-
tion, which is probably mediated by the microsomal mixed-function oxidase system.
6-1
-------
The half-life of toxaphene in rats is about 1 to 3 days and elimination takes
place via the urine and feces (Crowder and Dindal, 1974; Ohsawa et al., 1975).
The physiological and biochemical effects of toxaphene resemble those of
other chlorinated pesticides. Toxaphene induces the microsomal mixed-function
oxidase system and may thus cause a transient decrease in circulating steroid
levels (Peakall, 1976). Toxaphene also inhibits ATPases, alters lactate dehy-
drogenase isozyme patterns, and may affect vitamin A storage (Kuzminskaya and
Alekhina, 1976;- Phillips and Hatina, 1972).
The acute oral LD50 estimates of toxaphene to various laboratory mammals
vary between 25 mg/kg and 270 mg/kg (Table 6.8). As would be expected, the
vehicle used has a marked effect on such estimates (Hercules Inc., undated;
Lackey, 1949 a). Protein deficiency also increases the sensitivity of rats
to acute oral toxaphene intoxication by about a factor of three (Boyd and
Taylor, 1971). Although limited documentation on human poisoning is availa-
ble, the minimum acute oral lethal dose for man has been estimated to be be-
tween 30 and 103 mg/kg (Conley, 1952; Pollock, 1958).
The signs of acute oral toxaphene poisoning are similar in all mammals
tested. Clonic-tonic convulsions, indicative of central nervous system stimu-
lation, are the signs noted most often. The sequence of events following
administration are: a latent period of a few hours, apprehensive behavior,
salivation, vomiting, convulsions, and hyperreflexia. Death is sometimes
attributed to respiratory failure (Boyd and Taylor, 1971; Haun and Cueto, 1967;
Lackey, 1949 a; McGee et al., 1952). Renal tubular damage and fatty degenera-
tion with necrosis of the liver are two predominant pathological signs of acute
poisoning (Boyd and Taylor, 1971).
6-2
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Less detailed data are available on acute poisoning from other routes of
entry. The dermal toxicity of toxaphene to laboratory mammals seems to be an
order of magnitude below that of oral toxicity (Gaines, 1960 and 1969). While
the symptomalogy of acute dermal poisoning is similar to oral intoxication in
laboratory animals, dermal exposure to mixtures of toxaphene and lindane may
cause acute aplastic anemia in man (Pesticide Episode Response Branch, 1976).
The acute toxicity of some toxaphene components has been estimated
(Khalifa et al., 1974; Landrum et al., 1976; Ohsawa et al., 1975; Seiber et al.,
1975). Such studies are of considerable interest and may be significant in a
future understanding of the mechanism of action of toxaphene poisoning.
The subacute toxicity of toxaphene has not been studied in great detail.
Dietary doses of 189 ppm for 12 weeks and 100 to 800 ppm for six months are
reported to have no effect in rats (Clapp et al., 1971; Shelanski and Gellhorn,
undated). However, 50 ppm for 2 to 9 months has been shown to cause hydropic
accumulation in the liver (Ortega et al., 1957). Inhalation exposures to toxa-
phene dust at 0.012 mg/1 for three months causes 50 percent mortality in dogs,
but none in rats or guinea pigs (Hercules Inc., undated). In humans, subacute
exposure to toxaphene vapor may cause reversible lung damage (Warraki, 1963).
As summarized in the first paragraph of this section, chronic oral ex-
posure to toxaphene is associated with malignant tumors in mice and rats.
Toxaphene in the diet at 40 ppm may cause slight liver degeneration in dogs
after two years. In monkeys, 10 to 15 ppm in the diet over a two year period
is reported to have no effect (Hercules Inc., undated).
6-3
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6.2 METABOLISM
6.2.1 Absorption
No direct qualitative information is available on the absorption of
toxaphene. Crowder and Dindal (1974) have measured the uptake, distribution,
and elimination of Cl-36 in rat tissue at various intervals after single oral
^
intubations of 20 mg/kg Cl-36 labelled toxaphene in 0.5 ml peanut oil-gum
acacia. However, inconsistencies in toxaphene recovery during the first
24 hours after dosing preclude the use of this data in determining rates of
toxaphene absorption (Crowder, 1976). A follow-up study is currently being
conducted with mice.
Inferences on the absorption of toxaphene can be made from some of the
available toxicity data. Absorption across the alimentary tract, skin, and
respiratory tract is indicated by the adverse effects elicited by toxaphene
on oral, dermal, and inhalation exposures. Based on toxicity studies detailed
in Section 6.2.2, the vehicle used in the administration of toxaphene has a
marked influence on lethality which is probably attributable to differences
in the extent and/or rate of absorption. In oral exposures, toxaphene has a
much lower LD50 when administered in a readily absorbed vehicle - e.g. corn
oil or peanut oil - than when given in an indigestible vehicle such as kerosene.
Similarly, dermal applications of toxaphene in solution with mineral oil,
dimethyl phthalate, or water are much more toxic than similar applications of
toxaphene in powder preparations (Lackey, 1949 a and b; Conley, 1952).
Documented cases of human poisoning by toxaphene indicate that man may absorb
toxic levels on oral, dermal, or inhalation exposures (McGee et al. , 1952;
Pollock, 1958; Warraki, 1963). When administered or applied in comparable
6-4
-------
lipophilic solvents, the ratio of oral LD50 to dermal LD50 is about 0.1
(Tables 6.8 and 6.11). This suggests that toxaphene is absorbed more completely
and/or more rapidly from the alimentary tract than from the skin. The pro-
nounced variability in time to death after toxaphene ingestion (Section
6.3.2.1.1) indicates marked individual differences in the rate of toxaphene
*
absorption and/or differences in susceptibility to toxaphene intoxication.
6.2.2 Transport and Distribution
Toxaphene seems to distribute readily throughout body tissues soon after
dosing. Based upon toxaphene tissue residues in mammals after acute, subacute,
and chronic oral exposure, fat tissue seems to be the primary sight of toxa-
phene storage.
The distribution of Cl-36 in rat tissue at various times after intubation
of Cl-36 labelled toxaphene is given in Table 6.1 (Crowder and Dindal, 1974).
The values given in this table are average tissue residues for three rats at
each period after dosing. The values at day 20 are for three rats redosed with
20 mg/kg on day 9. The quantitative significance of these residue levels at
3 and 6 hours is questionable because of the low recovery of administered dose.
Further, the eratic variations of residue levels over the 20 day period are
probably attributable to marked variability in toxaphene metabolism among the
various experimental animals (Crowder, 1976). Nevertheless, these results do
indicate that measurable levels of toxaphene are transported to most body tis-
sues within 3 hours of dosing and are stored in body tissues for at least
9 days. The overall drop in tissue levels between 12 hours and day 1 is
consistent with the recovery of large percentages of the administered dose in
the feces and urine one day after dosing (Section 6.2.4). Residue levels at
6-5
-------
Table 6.1. Uptake of radioactivity in various rat tissues
and organs following a single-dose of Cl-36 toxaphene
(20 mg/kg)a
% Administered
Day
Tissue
Kidney
Muscle
Fat
Testes
Brain
Slood cells
Blood supernatant
Liver
First 2 cm ,
small intestine
T.ast 2 cm ,
-mall intestine
Large intestine
Esophagus
So Leen
Stomach
r,,;a,
3/24
0.05
0.93
0.14
0.02
0.03
3.1
0.64
0.33
0.06
0.10
0.19
0.04
0.04
3.70
9.37
6/24
0.13
1.6
0.15
.0.08
0.06
0
1.20
1.10
0.34
0.34
0.60
0
0.06
18.6
24.26
12/24
0.43
5.3
0.86
0.28
0.23
0
2.35
2.33
0.34
0.28
1.20
0.04
0.08
77.20
90.90
1
0.10
1.3
0.57
0.06
0.05
0.06
1.30
0.50
0.05
0.13
0.19
0.03
0.05
2.00
6.39
^Source: Modified from Crowder and Dindal,
^Toxaphene in 0.5 ml peanut oil acacia.
^Rats redosed with 20 mg/kg on day 9.
0
0
0
0
0
0
0
0
0
0
0
0
0
0
2
1974
dose
2
.03
.31
.04
.04
.90
.60
.31
.01
.01
.08
.01
.02
.63
.99
3 5
0.03 0.01
0.65 2.4
0.18 0.18
0.03 0.03
0 0
2.6 0
0.36 0
0 0.01
0 0
0.15 0
0.02 0.03
0.01 0.02
0 0.03
0.61 0.39
4.64 3.10
7
0
0.40
0.02
0.02
0
0
0.18
0
0
0
0.04
0
0.24
0.16
1.06
0
0
3
0
0
1
0
0
0
0
0
0
0
0
6
9
.03
.14
.65
.06
.01
.10
.09
.48
.84
.03
.06
.12
.57
20C
0
0.81
0.03
0
0
1.17
0.06
0
0.09
0
0
0
0
0
2.16
6-6
-------
nine days for single-dosed rats and 20 days for redosed rats suggest that a
relatively high amount of the retained dose is associated with blood cells.
High fat storage, however, is seen only in the single-dosed animals. As dis-
cussed above, the quantitative significance of these readings is dubious.
The distribution of C-14 labelled toxaphene and toxicant B (2,2,5-endo,
6-exo,8,9,IQ-heptachlorobornane) in rats 9 to 14 days after single oral doses
is summarized in Table 6.2 (Ohsawa et al., 1975). Fat tissue seems to be the
primary site of storage. Elevated levels in the blood, kidney, and liver are
suggestive of redistribution from fat via the circulatory system to kidney and
liver prior to urinary and fecal elimination (Section 6.2.4). No pronounced
differences are apparent between the distribution of technical grade toxaphene
and toxicant B.
In twelve week feeding studies of toxaphene to rats at levels of 2.33 to
189 ppm, toxaphene residues - determined by TLC analysis (Section 2.0) - in
omental fat, liver, and body composite samples (excluding liver and omental
fat) remained below dietary concentrations (Clapp et al., 1971). As illu-
strated in Figure 6.1 for female rats on a diet of 2.33 ppm and 189 ppm toxa-
phene, toxaphene residues in all tissue samples tended to increase during the
twelve week exposure. In male rats, however, liver and omental fat residues
peaked after 4 and 8 weeks, respectively. The relation between dietary levels
and tissue residues for both male and female rats is illustrated in Figure 6.2.
At dietary levels below 21 ppm, all three tissue components contain about the
same residue levels. However, as the dietary concentration increases above
21 ppm, tissue levels with respect to composite body samples increase in
omental fat but decrease in liver. As in the single dose studies, the pattern
of distribution illustrated in Figure 6.2 is indicative of toxaphene storage
6-7
-------
Table 6.2. Distribution of C-14 toxaphene and C-14
toxicant B in rat tissue after single intubation
(residue in ppm)
C-14 Toxaphene, C-14 Toxaphene,
14 days after dosing 14 days after dosing
(8.5 mg/kg) (19 mg/kg)
Fat
Liver
Kidney
Blood
Bone
Brain
Heart
Lung
Muscle
Spleen
Testes
0.52
0.12
0.17
0.14
0.02-0.09
0.02-0.09
0.02-0.09
0.02-0.09
0.02-0.09
0.02-0.09
0.02-0.09
/Source: Ohsawa et
,,2,2, 5-endo , 6-exo , 8
0.78
0.30
N.M.d
N.M.
N.M.
N.M.
N.M.
N.M.
N.M.
N.M.
N.M.
al. , 1975.
,9,10-Heptachlorobornane (Casida
C-14 Toxicant B,fa
9 days after dosingC
(2.6 mg/kg)
—
0.12
0.09
0.07
<0.03
<0.03
<0.03
<0.03
<0.03
<0.03
<0.03
et al. , 1974).
jAdministered in corn oil.
Not measured.
6-3
-------
100
10
UJ
cc
OMENTAL FAT
BODY COMPOSITE SAMPLE
LIVER
> 189ppm
) 2.33ppm
8
WEEKS ON DIET
12
Figure 6.1. Tissue levels in female rats fed toxaphene in
the diet at concentrations of 2.33 and 189 ppm.
Source: Modified from Clapp et al., 1971.
6-9
-------
I
I—'
o
3
Q
40
30
20
20
10 -
n 1—i—i—r
~r—i—i—i—nr
OMENTAL FAT
BODY COMPOSITE SAMPLE
LIVER
FEMALE
10 100
12 WEEK CONCENTRATION IN DIET(ppm)
1000
Figure 6.2. The effect of toxaphene concentration in the
diet on toxaphene tissue levels in male and female rats after
twelve weeks feeding. Source: Clapp et al., 1971.
-------
in fat tissue. The relatively constant liver levels but increasing fat levels
suggest that the rate of elimination via liver metabolism may be relatively
constant over dietary concentrations of 21 to 189 ppm.
Toxaphene fat storage has been measured in two year feeding studies to
,rats and dogs. These results are summarized in Table 6.3. The relation of
dietary concentration to fat storage in dogs is similar to that seen in the
12 week rat feeding studies illustrated in Figure 6.2. As in the 12 week
studies, female rats fed toxaphene for two years store greater amounts of
toxaphene in fat than do male rats.
6.2.3 B io t rans fo rma t ion
Dechlorination appears to be the primary metabolic process for toxaphene.
In the study by Crowder and Dindal (1974) detailed in Section 6.2.2, about
68 percent of the recovered activity is in the form of ionic chlorine. Of seven
Cl-36 labelled toxaphene fractions administered by intubation to rats at
doses of 8.4 to 17.2 mg/kg, all were dechlorinated by about 50 percent. Based
on the recovery of both C-14 and Cl-36 labelled toxaphene, Ohsawa and coworkers
(1975) have concluded that only 3 percent of the original dose of toxaphene
(12.7 to 14.2 mg/kg) is excreted unchanged. Only about 2 percent of C-14 la-
belled toxaphene is eliminated as C-14 labelled carbon dioxide (Ohsawa et al.,
1975).
Recently, Khalifa et al. (1976) have demonstrated that toxaphene undergoes
reductive dechlorination as well as dehydrochlorination in reduced bovine blood
hematin solutions. Similarly, toxaphene underwent about 50 percent dechlori-
nation when incubated with rat liver microsomes and reduced nicotinamide
adenine dinucleotide phosphate (NADPH) under anaerobic conditions. Both of
these reactions indicate that the hepatic microsomal mixed-function oxidase
6-11
-------
Table 6.3. Fat storage of toxaphene in two-year
feeding studies of rats and dogs
Animal
Rat , male
Rat, female
Dog
Duration
(years)
2
2
0.5
1
2
2
2
Concn. in
diet
(ppm)
400
1600
400
16QO
20
20
20
10
5
Concn. in
Fat
(ppm)
140
140
180
270
4
4.2
5.5
2.3
1.7
Concentration
Factor Reference
0.35 Lehman, 1952 a
0.088
0.45 Lehman, 1952 a
0.169
0.2 Hercules Inc.,
0.21 undated
0.275
0.23
0.34
a.
Concentration in fat/concentration in diet.
6-12
-------
system (MFOS) may be involved in toxaphene dechlorination. This enzyme system,
noted for its broad substrate specificity, utilizes cytochrome P-450 (a proto-
porphyrin similar to hematin) and requires NADPH. The cytochrome P-450, which
serves as the terminal oxidase of this enzyme system, has been shown to bind
with a variety of xenobiotics yielding two characteristic spectral patterns,
designated type I and type II binding spectra. Toxaphene interacts with the
hepatic cytochrome P-450 of rats, mice, and rabbits to yield a type I binding
spectra. As detailed by Kulkarni et al. (1975), such spectra usually indicate
that the binding compound can serve as a substrate for the cytochrome P-450
enzyme system. Thus, the hepatic microsomal mixed-function oxidase system
may play an important role in toxaphene metabolism. Evidence indicating that
toxaphene is able to induce this enzyme system is detailed in Section 6.3.1.2.
6.2.4 Elimination
As would be expected from the rapid metabolism and low volatility of
toxaphene, urinary and fecal elimination greatly predominate over respiratory
elimination. Using Cl-36 labelled toxaphene, Crowder and Dindal (1974) found
about twice as much Cl-36 in the feces as in the urine of singly dosed rats.
In redosed rats, however, the ratio was approximately equal (Table 6.4).
Ohsawa and coworkers (1975) have noted comparable patterns for Cl-36
and C-14 labelled toxaphene as well as for Cl-36 labelled toxaphene fractions
(Table 6.5). In their study, however, the amount of Cl-36 label is consistently
twice as great in the urine as in the feces. The C-14 label is distributed
more evenly, with a somewhat greater amount consistently found in the feces.
Based upon the information summarized in Table 6.5, no significant differences
are apparent between elimination patterns of the toxaphene mixture and the
fractions or components examined.
6-13
-------
Table 6.4. Excretion of radioactivity in urine and feces
of rats following a 20 rag/kg dose of Cl-36 toxaphene
(% administered dose; N=6)
Day
1
2
3
4
5
6
7
8
TotalC
Total
Single
Urine
1.46
3.20
2.89
2.35
1.82
1.19
1.15
0.54
15.30
29.10%
dose
Feces Total
23.95 25.4
7.45 10.6
1.25 4.1
1.10 3.5
1.06 2.9
1.23 2.4
0.69 1.8
0.27 0.8
37.30 52.6
70.90% 100.0%
h
Redose
Day
10
11
12
13
14
15
16
17
18
19
20
Urine
1.81
3.55
2.26
3.08
3.09
1.77
1.07
0.97
1.31
0.73
0.42
20.10
46.70%
Feces
6.00
11.60
1.40
1.10
1.20
0.60
0.40
0.40
0.20
0.00
0.00
22.90
53.30%
Total
7.8
15.2
3.7
4.2
4.3
2.4
1.5
1.4
1.5
0.7
0.4
43.0
100.0%
/Source: Crowder and Dindal, 1974.
The single-dosed animals were redosed with 20 mg/kg on the ninth day.
,% Administered dose.
% Recovered dose expressed as 100%.
6-14
-------
Table 6.5. Elimination of labelled toxaphene, toxaphene fractions I-VII,
and toxicants A and B 14 days after single oral dose
Compound
Toxaphene
Toxaphene
Toxaphene
Fraction I
Fraction II
Fraction III
Fraction IV
Fraction V
Fraction VI
Fraction VII
Toxicant A
b
Toxicant B
% Administered dose eliminated in
Dose
(mg/kg) Label Urine Feces
14.2
12.7
8.4
14.3
15.2
10.3
13.5
14.9
17.1
0.84
2.6
C-14
Cl-36
Cl-36
Cl-36
Cl-36
Cl-36
Cl-36
Cl-36
Cl-36
C-14
C-14
25.2
49.1
61.1
59.6
47.8
62.6
51.4
58.3
54.4
28.3
26.7
31.7
26.9
29.0
25.1
21.5
31.4
23.7
20.8
18.8
38.4
47.8
Expired CO.
1.2
1.8
0.7
/Source: Modified from Ohsawa et al., 1975.
Values of toxicant B for nine days after dosing.
6-15
-------
The kinetics of toxaphene elimination appear to be first order (Figure 6.3),
The half time for toxaphene elimination based on Figure 6.3 varies between 2
and 3 days (Ohsawa et al., 1975). This estimate agrees well with the apparent
half-life of toxaphene in Table 6.4 for the redosed animals (t 1/2 approxi-
mately 2 days) and the single dosed animals (t 1/2 approximately 1 day).
6.3 EFFECTS
6.3.1 Physiological or Biochemical
6.3.1.1 Normal Physiological or Biochemical Functions — Toxaphene is an
economic poison and is xenobiotic to all known life forms. As such, there
can be little reasonable doubt that toxaphene has no normal physiological or
biochemical functions.
6.3.1.2 Effects on Normal Physiological or Biochemical Functions — Like
many chlorinated hydrocarbon insecticides such as DDT, toxaphene seems to
induce the hepatic microsomal mixed-function oxidase system. However, the
inductive potency of toxaphene is generally much less than that of DDT metab-
olites. A shortening of pentobarbital sleeping time is generally considered a
good index of mixed-function oxidase induction. On single oral doses to rats,
toxaphene shortens sleeping time only at toxic levels, 50 mg/kg. DDT has a
comparable effect at one-tenth this dose (Schwabe and Wendling, 1967)- Ghazal
(1965) also found that toxaphene, given orally to rats at 100 mg/kg, decreased
N-methyl cyclohexenylmethyl barbituate sleeping time by about 20 percent for
up to 20 days. Similarly, whole liver homogenates from rats pretreated with
toxaphene or DDT in the diet for 1 to 13 weeks showed initial increased acti-
vity in three reactions characteristic of microsomal cytochrome P-450: oxida-
tive detoxication of EPN (0-ethyl-0-(4-nitrophenyl)phenylphosphonothioate),
0-demethylation of £-nitroanisole, and N-demethylation of aminopyrine
6-16
-------
<
cr
GO
O
Q
UL
O
35
\-
>
O
g
Q
100
80
60
40
20
(36
r36
Cl ) TOXAPHENEAND
Cl ] TOXAPHENE FRACTIONS
TOTAL
100
80
60
40
20
10 12 14 0
DAYS AFTER TREATMENT
*C]TOXAPHENE
TOTAL
FECES
A—A—A—6-
URINE
10
12
14
Figure 6.3. Excretion of radioactive products by rats
treated orally with [Cl-36] toxaphene, [Cl-36] toxaphene
fractions, and [C-14] toxaphene. Source: Ohsawa et al., 1975.
6-17
-------
(Kinoshita et al. , 1966). In each reaction, DDT had a markedly more pronounced
effect than toxaphene (Figure 6.4) and both chemicals generally had a more pro-
nounced effect in male rats (Table 6.6). Toxaphene also stimulates liver micro-
somal metabolism of estrone and testosterone. Liver microsomal preparations
from female rats pretreated with daily intraperitoneal injections of 25 mg/kg
toxaphene for seven days showed a 155 percent increase in estrone metabolism.
DDT, under the same conditions, resulted in a 213 percent increase (Welch et
al., 1971). Similarly, liver microsomal preparations from male rats given a
single oral dose of toxaphene at 120 mg/kg evidenced a 484 percent increase
in testosterone metabolism by five days after dosing. This increased in vitro
metabolism was accompanied by a 19 percent decrease (significant at p less than
0.02) in plasma testosterone. However, by fifteen days after dosing, testosterone
levels returned to normal even though microsomal enzyme activity was elevated
by 532 percent. Chronic oral exposure to toxaphene - 1.2 mg/kg/day x 6 months -
caused comparable increases in microsomal testosterone metabolism but did not
lower plasma testosterone levels. Thus, it appears that the initial decrease
in circulating testosterone caused by increased liver metabolism is rapidly
offset by increased testosterone production (Peakall, 1976). Because toxaphene
is known to bind with mammalian cytochrome P-450, giving a typical type I
difference spectra (Kulkarni et al., 1975), all of the above metabolic effects
may be reasonably attributed to hepatic cytochrome P-450 induction.
Some additional enzyme systems are also affected by toxaphene. At a
concentration of 330 micromolar, toxaphene inhibits the in vitro activity of
the mitochondrial succinoxidase system of beef heart by 75.9 percent and the
6-18
-------
TOXAPHEME
DDT
300-i
WEEKS ON DIET
400-,
13 0 1
EPN DETOXICATION
WEEKS ON DIET
iOO-i
200 -
100 -
0 1
WEFKSON DIET
400-1
300-
o
e
£ 200-
o
o
100 -
J_
13 0 1
O-DEMETHYLASE
WEEKSOIM DIET
13
WEEKSON DIET
I
600-
500-
350-
300-
200-
100 -
13 0 1
N-DEMETHYLASE
WEEKSON DIET
13
Figure 6.4. Effect of feeding toxaphene and DDT
on EPN detoxication, .a-demethylase activity, and N-
demethylase activity of livers of male rats.
Source: Kinoshita et al., 1966.
6-19
-------
Table 6.6. Influence of various dietary levels of toxaphene on the activity
of microsomal enzymes of the livers of male and female rats
* Length of feeding
(weeks)
Activity
Male Female
(5 ppm)
fa
Dietary level
Male Female
(25 ppm)
Male Female
(50 ppm)
EPN Detoxification
1 90%
3 115%
6 120%
13 125%
114%
97%
103%
90%
145%
145%
140%
140%
146%
132%
103%
110%
150%
140%
160%
150%
135%
135%
110%
130%
0-Demethylase
1
3
6
13
140%
110%
120%
130%
106%
82%
113%
98%
180%
140%
150%
110%
134%
101%
112%
116%
200%
115%
155%
120%
140%
109%
95%
120%
N-Demethylase
1
3
6
13
75%
140%
120%
125%
158%
127%
114%
118%
125%
155%
125%
130%
184%
161%
184%
162%
325%
160%
145%
135%
260%
224%
200%
138%
/Source: Modified from Kinoshita et al., 1966.
Activity expressed as % of untreated control.
6-20
-------
NADH-oxidase system by 96.1 percent (Pardini et al., 1971). At 100 micromolar
concentration, toxaphene inhibits by 67 percent the active transport of glucose
through isolated mice intestine. However, the mode of action of this effect -
inhibition of sugar carrier protein, sodium ion-potassium ion-activated ATPase,
or ATP synthesis - has not been determined (Guthrie et al., 1974).
Altered lactate dehydrogenase (LDH) spectra have been demonstrated in
the liver and blood of rats given daily oral doses of toxaphene at 1.2 mg/kg
for six months or a single oral dose of 120 mg/kg. In the liver, single doses
caused a 50 percent decrease in total LDH activity over the 15 day post-exposure
observation period. This decreased activity was associated with increases in
LDH-1, LDH-2, and LDH-3, a transient decrease in LDH-4, and no significant
change in LDH-5. In the blood, single doses caused no significant change in
total LDH activity but did cause a transient decrease in LDH-1. Chronic toxa-
phene exposure decreased total LDH activity in both the blood (30 percent de-
crease) and liver (45 percent decrease). The only consistent spectral changes
in the liver were an increase in LDH-3 and a decrease in LDH-5 after three
and six months. In the blood, LDH-1 and LDH-2 activities were depressed and
LDH-5 activity elevated after six months. These changes in blood isozyme
spectra were thought to be indicative of hypoxia (Kuzminskaya and Alekhina,
1976). Gertig and Nowaczyk (1975) also noted depressed total LDH activity in
the liver, kidneys, and serum - 17, 17.5, 20 percent decreases, respectively -
of rats given daily oral doses of toxaphene at 35 mg/kg for six months. In
kidney LDH isozymes, the only constant change was a moderate (17 percent) but
statistically significant drop in LDH-1. These investigators also noted a
decrease of about 50 percent in both serum alkaline phosphatase and liver
6-21
-------
glutamate dehydrogenase after the six month exposure period. Transient changes
were seen in serum acid phosphatase (70 percent decrease after one month only),
and two liver transaminases (more than 150 percent increase after one and
three months). All of the above changes were significant at least by p less
than 0.01.
Toxaphene also may have some effect on vitamin A storage. Phillips and
Hatina (1972) exposed 28 day old female rats to toxaphene at 100 ppm in the
diet for 72 days. After exposure, these rats were mated with untreated
males, and liver vitamin A was determined in both dams and progeny at 20 days
postpartum. While no effect was seen in the dams, liver vitamin A was de-
pressed by 11.1 percent (statistically significant at p less than 0.01) in
the newborn rats. Similar results were also obtained with chlordane, lindane,
methoxychlor, and DDT.
The physiological or biochemical effects of toxaphene in man have not
received extensive study. In an epidemiclogical survey of workers exposed
to toxaphene and a variety of other pesticides over a period of months, no
effects were noted on heme synthesis as determined by delta-aminolevulinic
acid and porphobilinogen urinary excretion (Embry et al., 1972).
6.3.2 Toxicity
6.3.2.1 Acute Toxicity — As discussed in Section 2.0, technical grade
toxaphene is a complex mixture of chlorinated alicyclic hydrocarbons. Most
of the available acute toxicity information on toxaphene involves the use of
this complex mixture. Recently, however, toxaphene has been separated into
various fractions and a few of its toxic components have been identified.
Information on the toxicity of these components is discussed in a separate
section below (Section 6.3.2.1.2).
6-22
-------
6.3.2.1.1 Technical toxaphene — Like many of the chlorinated hydrocarbon
pesticides, acute intoxication with toxaphene is characterized by profound
central nervous system stimulation. Because of the patterns of toxaphene use
and application, oral and dermal exposures are likely to present the greatest
hazard to man, and cases of human intoxication by both of these routes have
been documented. Consequently, the oral and dermal toxicity of toxaphene to
experimental mammals has been studied in some detail.
However, much of the available information consists of only LD50 approxi-
mations rather than detailed dose-response data. In that the latter type of
information is highly desirable in hazard assessment, all available dose-
response data for toxaphene by both oral and dermal administration are pre-
sented separately in Table 6.7 and illustrated in Figures 6.5 and 6.6. Details
of these studies are discussed in the appropriate sections on oral and dermal
toxicity.
6.3.2.1.1.1 Oral toxicity — The acute oral toxicity of technical grade
toxaphene to a variety of laboratory mammals is summarized in Table 6.8. This
table includes only toxaphene exposures in which a readily absorbed vehicle
is used. A variety of investigators have noted that substitution of such a
vehicle with kerosene significantly raises the LD50 estimate of toxaphene.
Estimates of this change, as a ratio of LD50 with kerosene to LD50 with vege-
table oil, are 1.3 for guinea pigs, 2.0 for rats, and 5.0 for dogs (Hercules
Inc., undated). With kerosene as a vehicle, Lackey (1949 a) also noted only
sporadic mortality in dogs at toxaphene levels up to eight times above the
LD50 with peanut oil. This effect is probably attributable to decreased
absorption of toxaphene due to toxaphene affinity for kerosene.
6-23
-------
Table 6.7. Estimated dose-response information for toxaphene in oral and dermal applications
I.D1
LD50
LD99
Minimum
lethal dose
tested
Slope of
log dose
M'obi t I i ne
Time to death
Reference
Oral
K.its, male, luw protein diet 1.1
Hats, male, protelu test
diet, norma1
Kats, male, standard
191
80 + 19'
293 + 31
a
159
395
39 + 6 hours
25 + 13 hours
Boyd and Taylor,
1971b
laboratory chow
Kats, male
1
^ Rats, female
Dermal
Ra t s , ma 1 e
Ra t s , female
139 220 + 33 300
15 90 50
(67-122)
44 88 63
(70-9.1)
72 1075 800
(717-1613)
230 780 400
(600-1014)
•^15 24 + 9 hours
3.0 3 hours-5
10.0 4 hours-1
2 3 hours-4
4.3 4 hours-5
days Gaines, 1960 and
1969"
day
days
days
.Mean + SE.
^See Figure 6.6. for graphical presentation of data.
"JMean (957 confidence interval).
See Figure 6.7. for graphical presentation of data.
-------
99
95
90
80
70
60
50
40
30
20
10
5
LOW PROTEIN
DIET/
PROTEIN TEST
DIET NORMAL
0.1
1.0 2.0
LOG DOSE ( mg/kg )
3.0
4.0
Figure 6.5. Log dose response curves of the acute
oral toxicity of toxaphene to rats under different dietary
conditions. Source: Boyd et al. , 1971.
6-25
-------
7.0
> 6.0
CC
o
I 5.0
m
§ 4.0
3.0
1.0 2.0 3.0
LOG DOSE (mg/kg )
4.0
Figure 5.6. Log dose response curves for male and female
rats to acute oral and dermal toxaphene intoxication.
Source: Gaines, 1960 and 1969.
6-26
-------
Table 6.8. Acute oral toxicity of technical toxaphene to laboratory mammals
Organism
Rats
Unspecified strain
Wistar, male,
3-4 weeks, 50-60 g,
fasted
It
Sherman, male,
>90 days, >175 g,
fasted
Sherman, female,
>90 days, >175 g,
fasted
Mice
Cats
Dogs
Rabbits
Guinea Pigs
Vehicle
Unspecified
Cottonseed oil
Peanut oil
Peanut oil
Peanut oil
Peanut oil
Corn oil
Corn oil
Corn oil
Unspecified oil
Peanut oil
Unspecified oil
Peanut oil
Corn oil
Unspecified oil
Peanut oil
Corn oil
Unspecified oil
LD50
(mg/kg)
69
220 + 33a
90(67-122)
80(70-91)
40
90
120-125
60
112
80
25-40
100
•\-25
49
100
75-100
270
80
Reference
Lehman, 1951
Boyd and Taylor, 1971
Gaines , 1960
Gaines, 1960
Shelanski and Gellhorn,
undated
Hercules Inc. , undated
Shelanski and Gellhorn ,
undated
Hercules Inc. , undated
Hercules Inc. , undated
Rico, 1961
Hercules Inc. , undated
Rico, 1961
Lackey, 1949 a
Hercules Inc. , undated
Rico, 1961
Hercules Inc. , undated
Hercules Inc. , undated
Rico, 1961
(
Standard error.
95% Confidence interval.
6-27
-------
Even when comparable readily absorbed vehicles are used, considerable
variation is still evident both within and among the various species cited in
Table 6.8. Such variability could be attributable to a number of factors in-
cluding differences in strain, sex, age, handling, care, and initial health
of the experimental animals. Based on the results of Gaines (1960, 1969),
sex does not appear to be a significant variable in the LD50 estimate for
Sherman rats. However, on both oral and dermal administrations, the dose
response curve for female rats has markedly higher slopes than that for male
rats (Table 6.7, Figure 6.6). Because confidence limits are not given for
these slopes, the significance of this difference cannot be determined. Diet,
however, may have a pronounced effect on estimates of toxaphene toxicity. Boyd
and Taylor (1971) have shown that rats on a protein deficient diet (3.5 percent
casein) for 28 days prior to dosing have LD50 values about three times below
those of rats fed on a diet containing normal protein levels (24 to 26 percent
casein). Details of these results are summarized in Table 6.6 and Figure 6.5.
Similar increases in susceptibility of protein-deficient rats to chemical in-
sult have been demonstrated not only for toxaphene but also for a variety of
chlorinated hydrocarbon and organophosphorous insecticides (Boyd and Taylor,
1971). The reasons for the increased sensitivity of protein-deficient animals
is not completely understood. However, as indicated in Section 6.2.3, the
hepatic, microsomal cytochrome P-450 system may contribute to toxaphene detoxi-
cation. In that protein deficiency generally depresses cytochrome P-450 con-
centration and activity (Campbell and Hayes, 1974) , one of the factors involved
in the increased susceptibility of protein-deficient animals to toxaphene may
be a decreased detoxication ability. As illustrated in Figure 6.5, the in-
creased susceptibility of the protein-deficient rats is not uniform throughout
6-28
-------
the dose-response curve. The disproportionately low LD1 estimate for protein-
deficient rats suggests that a subpopulation of this group has an extraordinarily
low tolerance to toxaphene poisoning.
Because of the many experimental variables that could influence the LD50
estimates given in Table 6.8 and because confidence limits are not given for
*
most of these estimates, no significant differences in species susceptibility
can be identified. The acute oral LD50 estimates for laboratory mammals
range from 25 mg/kg to 270 mg/kg. The minimum acute lethal dose for man has
been estimated to lie between 30 and 103 mg/kg (Conley, 1952; Pollock, 1958).
Based on the estimates of mammalian toxicity discussed above and on cases of
accidental ingestion by humans detailed below, this approximation of human
lethal dose seems reasonable.
Like most of the chlorinated hydrocarbon insecticides, toxaphene acts as
a central nervous system stimulant in acute intoxication. The precise mechanism
of this effect, however, is not known. The most consistent clinical sign of
acute toxaphene poisoning fs clonic-tonic convulsions. In fatal exposures,
death is usually attributable to respiratory failure. For the most part,
patterns of intoxication are similar in dogs, rats, and humans. Published
reports of human cases of toxaphene poisoning by ingestion are summarized
in Table 6.9. Initial signs of intoxication usually do not appear until a
few hours after ingestion. Rats responded to single oral doses of toxaphene
with hypersensitivity, tremors, and convulsions. Symptoms appeared after
1 1/2 hours and lasted for up to 24 hours. Although some delayed deaths were
noted up to the sixth day after dosing, most deaths occurred during the first
24 hours (Lehman, 1951). In dogs receiving lethal doses of toxaphene, the
6-29
-------
Table 6.9. Case studies of toxaphene poisoning in humans in which ingestion is the primary
route of entry
1
u>
0
Case No. 1 2 3 4
Subject(s) Male Male, 4 yrs Male, Male, 2 yrs
2yr8mo lyrSmo
Nature of
toxaphene Wax Emulsion in 60% in 20% in solution
water solvents
Dose Unknown Unknown ^-100 mg/kg Unknown
Time to react
to onset of
symptoms '^7 hours 2 hours N.S. N.S.
5
Female , 20 yrs
Female, 16 yrs
Female, 12 yrs
Residue of spray
in food
9.5-47 mg/kg
1.5-4 hours
6
Male, adult
Male , young
Female , adult
Residue of spray
in food
Unknown
4 hours
7
Female, 9 mo
Powder, 13.8%
toxaphene ,
7.04% DDT
Unknown
A few hours
Synip toms
Convulsions Convulsions at Convulsions, Convulsions, intermittent; Nausea; vomiting;
2-5 minute intermittent mild cerebral excitement; convulsions
intervals aimless jerking motion and
excessive muscular tension
of extremities; marked
pharyngeal and laryngeal
spasms; labored respira-
tion; cyanosis
No nausea; spon- Vomiting;
taneous vomiting; diarrhea; convul-
convulsions; jerk- sions; hyperre-
ing and transitory flexia; tachycar-
movements; muscular dia; b.p.140/110;
rigidity; periods labored respira-
of unconsciousness; tion; respiratory
aranesia(?) failure
Outcome Death Death
Time to death
or recovery 9.5 hours 6 hours
Death Recovery
11 hours 12 hours
Recovery
1-12 hours
Recovery
<1 day(?)
Death
'>-9 hours
.Sources: Cases 1-6, McCee et al., 1952; Case 7, Haun and Cueto, 1967.
See text for dose estimation.
-------
first signs of intoxication include apprehensive behavior, salivation, and
vomiting which appear 1 1/2 to 2 hours after dosing (Lackey, 1949 a). In
some cases of human poisoning, however, early signs of intoxication may be
severe - e.g., Case 6 in Table 6.9. Along with convulsions, hyperreflexia
has been noted in dogs (Lackey, 1949 a), rats (Boyd and Taylor, 1971), and
humans (Haun and Cueto, 1967). In Case 7 in Table 6.9, hyperreflexia was an
outstanding feature in the latter stages of intoxication. While Doroshchuk
(1974) indicates that both nembutal an^d urethane can be used to control spasms
and normalize respiration in severely intoxicated rats and cats, Haun and
Cueto (1967) found that phenobarbital did not diminish the severe "violent
spasm" induced by "very minor stimulation" in Case 7 in Table 6.9. However,
except for the consistent presence of convulsions, the symptoms seen in cases
of acute human intoxication are somewhat variable. For instance, Case 6
in Table 6.9 is somewhat peculiar in that neither nausea nor spontaneous
vomiting occurred. Given the lack of dosage information, the significance
of such differences is questionable. As with many toxins, the expected sur-
vival time is inversely proportional to the dose. This has been specifically
noted by Boyd and Taylor (.1971) for rats and may have partially contributed
to the increased survival time of the protein-deficient rats noted above.
In four fatal human poisonings, death occurred between 6 and 11 hours after
ingestion. This relatively short period prior to death may indicate that
relatively massive doses were consumed. Specific lethal dose information,
however, is available on only one child. In this case, the seventeen month
old child is said to have consumed one-half a tablespoon of 60 percent toxa-
phene (McGee et al., 1952). Assuming a body weight of 14 kg, this would equal
a dose of about 100 mg/kg.
6-31
-------
Additional reports of oral poisoning in humans have not been encountered
in the published literature. A draft of a report on poisoning incidents in-
volving toxaphene compiled by the Pesticide Episode Response Branch, E.P.A.
(1976) has recently become available. This report contains information vol-
untarily provided to the E.P.A. by various state, federal, and private organi-
zations. Most of these individual reports, which vary widely in quality and
detail, involve exposure to toxaphene along with a variety of other active
agents. In no cases are estimates of toxaphene doses available. Of the 43
cases of human poisoning described in this report, four cases deal with acci-
dental poisoning of children. Symptoms of poisoning are consistent with
those described in the published reports: vomiting, convulsions, cyanosis,
and coma. Two of the four cases were terminal. In a fifth case, an adult
apparently attempted suicide by toxaphene ingestion. High levels of toxaphene
were found in the blood, but the only symptom given is jaundice (Pesticide
Episode Response Branch, 1976).
Pathology data on acute oral toxaphene poisoning are available only on
rats (Table 6.10). The renal tubular damage is similar to that seen on sub-
acute exposures. However, the fatty degeneration noted in Table 6.10 for
acute intoxication does not correspond to the hydropic degeneration noted
in Section 6.3.2.2 for subacute intoxication.
6.3.2.1.1.2 Acute dermal toxicity — Based on the available information sum-
marized in Table 6.11, toxaphene seems much less toxic on dermal exposures
compared to oral exposures. However, the solvent used in dermal exposures
may markedly affect toxicity estimates. Lehman (1952 b) noted only moderate
skin irritation and hyperexcitability in rabbits after a single 24 hour exposure
6-32
-------
Table 6.10. Summary of histopathologic findings at autopsy on albino rats
which died from oral administration of toxaphene'1*'3
Organ
His topathology
Adrenal glands
Brain
Gastrointestinal tract
Cardiac stomach
Pyloric stomach
Small bowel
Cecum
Colon
Heart
Kidneys
Liver
Lungs
Muscle
(ventral abdominal wall)
Salivary (submaxillary) glands
Skin
Spleen
Testes
Thymus gland
Cortical sinusoids frequently congested
Capillary-venous congestion and hemorrhage in
the meninges; capillary congestion of the brain
Granulocyte-infiltrated ulcers in late deaths
Congestion of capillaries in the lamina propria
near the mouths of the gastric glands
Capillary congestion of the villi
Occasionally mild capillary congestion of the
lamina propria
Normal appearance
Occasionally capillary congestion and capillary
hemmorrhage
Cloudy swelling of the proximal and distal
convoluted tubules and congestion of the loop
of Henle
Extensive diffuse fatty degeneration and early
necrosis
Occasionally congestion and hemorrhage of the
parenchyma
Normal appearance
Serous glands shrunken and ducts empty
Normal appearance
Contracted red pulp
Varying degrees of inhibition of spermatogenesis
Varying degrees of loss of thymocytes and
capillary-venous congestion
a.
/Source: Boyd and Taylor, 1971.
Animals fed a low protein diet exhibited, in addition, signs of
inhibited development of most organs and tissues.
6-33
-------
Table 6.11. Acute dermal toxicity of toxaphene to laboratory mammals
a
Organism Vehicle
Rats
Sherman, male,
>90 days, >175 g,
unfasted Xylene
as Rats
oj Sherman, female,
*" >90 days, >175 g,
unfasted Xylene
Rats Xylene
Rabbits Dust
Rabbits Peanut
oil
Dose
(mg/kg)
1075
(717-1613)
780
(600-1014)
930
>4000
<250
Dermal :0ral
Response LD50 Ratio^
LD50 (95% VL2
Confidence
Interval)
LD50 (95% 9.75
Confidence
Interval)
LD50 ^10
LD50
LD50 2.5-3.3
Reference
Gaines , 1960 and 1969
Gaines, 1960 and 1969
Hercules Inc. , undated
Hercules Inc., undated
Hercules Inc. , undated
/See Table 6.7 for dose-response data.
Ratio of dermal toxicity to comparable oral toxicity from Table 6.7.
-------
to toxaphene (dry, waxy form) at 4000 mg/kg. In contrast, toxaphene in 20 per-
cent dimethyl phthalate caused tremors leading to convulsions and death at doses
of less than 780 mg/kg. A similar solvent dependence is apparent in the rabbit
studies summarized in Table 6.11. Gaines (1960, 1969) specifically designed
^his experimental protocol so that the oral and dermal toxicity of a variety
of pesticides could be reliably compared (Table 6.7, Figure 6.2). In these
studies, Gaines (1960, 1969) noted that compounds with oral and dermal toxici-
ties on the same order of magnitude are most likely to present an occupational
hazard to man. The relatively high ratio of dermal to oral toxicity for toxa-
phene suggests that this pesticide is not likely to present such a hazard.
This tentative assumption may be true for toxaphene, at least in cases
of acute dermal exposure. Only one case of human poisoning by cutaneous ab-
sorption has been published and, based on the available details, the symptoms
that developed cannot be unequivocally attributed to toxaphene. In this in-
stance, a 70 year old male had his hands in contact with a toxaphene—lindane
solution for two hours. After 10 hours, the following symptoms developed:
headache, poor coordination, lassitude, severe nausea, and vomiting. Over
the next week, this individual exhibited mild hyperthermia, flaccid muscula-
ture, and decreased response to stimuli. Only after 9 days did the individual
become semicomatose. At no time were convulsions or hyperreflexia noted
(Pollock, 1958). These signs and symptoms are not characteristic of toxaphene
or lindane poisoning (Matsumura, 1975) and differ markedly from the previously
described cases of acute oral toxaphene poisoning in humans. While clinical
signs of intoxication may be expected to show some variation with different
routes of entry, such profound variation is uncommon with the chlorinated
6-35
-------
insecticides. Gaines (1960, 1969) noted no difference between signs of intoxi-
cation in rats orally and dermally exposed to a variety of pesticides. Lackey
(1949 a, b) similarly notes no remarkable differences in the response of dogs
to subacute oral and dermal doses of toxaphene (Section 6.3.2.2). However,
two cases of acute aplastic anemia associated with dermal exposure to toxa-
phene/lindane mixtures have been reported (Pesticide Episode Response Branch,
1976). One of these cases resulted in death due to acute myelomonocytic leu-
kemia which was presumed to be secondary to the development of aplastic anemia.
A recent review by the National Academy of Sciences (1977) links over 50 cases
of aplastic anemia with exposure to lindane or benzene hexachloride, although
no firm causal relationship could be demonstrated. Consequently, the role of
toxaphene in the development of acute aplastic anemia is uncertain.
Hayes (1963) has estimated that a single dermal application of 46 g or
daily applications of 2.4 g "over a period of days is very dangerous" to
humans. For a 70 kg man, the single dose estimate is approximately 660 mg/kg.
Documentation of this estimate is not provided by Hayes (1963).
6.3.2.1.1.3 Other routes of entry —• Very little information is available on
the toxicity of technical grade toxaphene by routes other than oral and dermal
exposure (Table 6.12). The acute intraperitoneal LD50 for mice approaches the
lower level of the oral LD50 estimates for mammals (Table 6.8), suggesting
that incomplete absorption may be a major factor in the variability of the
oral LD50 estimates. Unlike oral exposures, intravenous injections of toxa-
phene to curarized animals cause a pronounced increase in blood pressure
indicative of vasomotor stimulation (Conley, 1952). Thus, the lower LDSO's
with intravenous injections may be associated with a different mode of action
caused by high toxaphene blood levels not attained with other routes of admin-
istration.
6-36
-------
Table 6.12. Acute toxicity of toxaphene on inhalation, intravenous, and intraperitoneal injections
Organism
Route
Vehicle
Dose
Response
Reference
I
LJ
Mice
Rats
Mice, male
18-20 g
Rats
Mammals
Inhalation Oil-based 0.2 mg/100 cc air/ LC50 estimate Conley, 1952
mists min x 2 hrs
Inhalation 40% dust 3.4 g/1 air x 1 hour 50% mortality Hercules Inc., undated
Intraperi-
toneal
injection
Intravenous
injection
Intraperi-
toneal
injection
Dimethyl 42 mg/kg
sulfoxide,
100 yI/mouse
Peanut oil 13 mg/kg
and acacia,
1% toxaphene
LD50
LD50
Khalifa et al. , 1974
Shelanski and Gellhorn,
undated
N.S.
10-15 mg/kg
LD50 range Conley, 1952
-------
The inhalation LD50 value cited by Conley (1952) is based on exposures of
mice to 0.2 mg/100 cc air/minute for varying periods of time. During exposure
periods of 112, 116, and 158 minutes, mice mortality was 40, 100, and 60
percent, respectively. Because no significant lung damage was noted in fatally
exposed mice, death was attributed to toxaphene absorption rather than to the
oil-based mist used as the vehicle. No cases of acute inhalation poisoning in
humans have been reported.
Toxaphene, as a 20 percent solution in kerosene, applied to eyes of
rabbits and guinea pigs at doses of 2 mg/kg and 10 mg/kg, respectively, for 14
days caused only transient eye irritation with no permanent eye injury (Hercules
Inc., undated).
6.3.2.1.2 Toxaphene fractions and components — As discussed previously
(Section 2.0), toxaphene may be separated into various fractions with each
fraction containing a complex mixture of several toxaphene components. The
chemical identity of these components has been determined in only a few in-
stances. Toxicity data on toxaphene fractions and a few of the identified
components are summarized in Tables 6.13 and 6.14. To emphasize that the
fractions separated by Ohsawa and coworkers (1975) are chemically different
from those of Seiber and coworkers (1975), fractions isolated by the former
group are identified by Roman numerals (I to VII), and those of the latter by
Arabic numeral (1 to 8). As previously defined (Section 2.0), the specific
compounds isolated by Casida et al. (1974) and Turner et al. (1975) are
referred to as toxicants A and B.
In the seven toxaphene fractions tested by Ohsawa and coworkers (1975),
there is no apparent relationship of toxicity to either chlorine content or
6-38
-------
Table 6.13. Acute intraperitoneal LD50's of toxaphene and toxaphene fractions
to male
Technical Toxaphene
Expected Values of
Fractions I- VII"
Fraction I
Fraction II
Fraction III2"
Fraction IV
Fraction V ' J
Fraction VI"
Fraction VII
Toxicant A
Toxicant B
LD50 b
(mg/kg)
42
45
252
110
16
25
52
63
194
3.1
6.6
Toxicity relative
to toxapheneC-
1.
0.
0.
0.
2.
1.
0.
0.
0.
13.
6.
0
93
16
38
63
68
81
67
22
55
36
% Cl
68.
60.
68.
68.
68.
68.
66.
64.
21.
6
8
1
3
8
1
8
1
2
^Source: Modified from Khalifa et al., 1974 and Ohsawa et al., 1975.
In 100 microliters of dimethyl sulfoxide per dose.
jLD50 of toxaphene * LD50 of component.
Expected value calculated as harmonic mean, i.e. (x)ii = N/£(l/JXi).
/Includes Toxicant A.
^Includes Toxicant B.
^Toxicant A: Identified as mixture of 2,2,5-endo, 6-exo, 8,8,9,10-octa-
chloroborane and 2,5,5-endo, 6-exo, 8,9,9,10-octachloroborane
, (Turner et al., 1975).
Toxicant B: Identified as 2,2,5-endo, 6-exo, 8,9,10-heptachlorobor.ane
(Casida et al., 1974).
6-39
-------
Table 6.14. Acute intraperitoneal LDSO's of toxaphene and toxaphene fractions
to micea
Technical Toxaphene
Expected Values of
Fractions 2-8^
Fraction 1
Fraction 2
Fraction 3£
Fraction 4
Fraction 5
Fraction 6
Fraction 7
Fraction 81$
LD50
(mg/kg)b
32
41.4
29
23
65
95
79
53
177
Toxicity relative
to toxapheneC-
1.0
0.77
1.10
1.39
0.49
0.34
0.41
0.60
0.18
% (w/w)
technical
toxaphene
2.1
38.1
33.9
12.0
8.0
3.1
1.5
2.2
-Source: Seiber et al., 1975.
In dimethyl sulfoxide (amount unspecified).
TLD50 of toxaphene T LD50 of component.
Expected value calculated as Z(LD50i • % of toxaphene i).
/Contains Toxicant B.
°Major component of fraction identified as 2,5,6-_exo-8,8,9 ,10-hepta-
chlorodihydrocamphene.
6-40
-------
the presence of toxicants A or B. The least toxic fraction, I, has about the
same chlorine content as the most toxic fractions, III and IV. While both of
these highly toxic fractions contain toxicant B, fraction V, which contains
both toxicants A and B, is less toxic than technical grade toxaphene. The
average LD50 computed as the harmonic mean of the seven fractions suggests an
additive rather than synergistic relation among these fractions (Ohsawa at al.,
1975). Similar patterns have been noted in the toxicity of these fractions and
components to house flies (Section 5.6). As discussed in Section 6.2.3, dif-
ferences in the toxicity of these components cannot be directly attributed to
differences in detoxication.
The toxaphene fractions separated by Seiber and coworkers (1975) have a
range of toxicity (Table 6.14) comparable to those noted in Table 6.13. While
no definite pattern is apparent in the toxicity of the various fractions, high
toxicity was associated with those fractions eluted by relatively apolar sol-
vents - i.e., fractions 2 and 3 by 40 percent and 50 percent benzene in hexane.
These fractions also comprised significantly greater percentages of the original
technical toxaphene. As with the highly toxic fractions, III and IV, separated
by Ohsawa and coworkers (1975), toxicant B was found by Seiber and coworkers
(1975) in the fraction with the highest relative toxicity, fraction 3. Con-
sistent with the results of Ohsawa et al. (1975), the expected value for the
LD50 of fractions 2 to 8 does not indicate any synergistic effect among the
toxaphene fractions.
Landrum and coworkers (1976) have recently determined the toxicity of two
toxaphene column fractions to mice on intraperitoneal injection. Column frac-
tion 3 had an LD50 of 38 (32 to 45) mg/kg which was not significantly different
6-41
-------
from that found for technical toxaphene, 33 (27 to 41) mg/kg. However, the
other column fraction, designated column fraction 7, was only about one-half
as toxic as toxaphene, having an LD50 of 67 (58 to 78) mg/kg. About one-half
of this latter fraction consisted of 2,5,6-exo,8,8,9,10-heptachlorodihydro-
camphene, which has been shown to be significantly less toxic than technical
toxaphene to houseflies (Section 5.6.2.2.1.2).
6.3.2.2 Subacute Toxicity — Table 6.15 summarizes the effects of subacute
oral administration of toxaphene to laboratory mammals. Except for convulsions
observed in dogs given 5 mg/kg/day, none of the exposures detailed in Table 6.15
resulted in clinical signs of toxaphene poisoning. The ability of dogs to tol-
erate large cumulative doses (176 to 424 mg/kg) when given at 4 mg/kg/day sug-
gests a rather sharp threshold level for central nervous system stimulation.
This is consistent with information discussed in Section 6.2.4, showing that
toxaphene is eliminated relatively rapidly. A similar pattern is seen in rats
on intraperitoneal injection. Ohsawa and coworkers (1975) have found that
male rats injected with 50 mg toxaphene (approximately 300 mg/kg) every 48 hours
tolerated cumulative doses of 700 to 2000 mg/kg (over ten times the single
oral LD50 dose) before marked lethality occurred.
In subacute exposures which do not cause apparent central nervous system
stimulation, no increases in mortality are noted. However, pathological changes
of the kidneys and liver, as well as changes in blood chemistry, seem to be
common features of subclinical toxaphene intoxication.
Ortega and coworkers (1957) (using rats) and Lackey (1949 a) (using dogs)
have noted similar changes in liver histology. Morphologically, these changes
appear as vacuoles of plasma with occasional red blood cells found within
6-42
-------
Table 6.15. Subacute oral toxicity of toxaphene
Organism
Mice, both albino and
wild s t rai ns
R;its
RJLS
Rats, Sherman, male
and female, ^100 g
Rats and guinea pigs
Dugs
Vehicle Duration
Diet Several weeks
or months
Diet 12 weeks
N.S. 7 months
Diet 2-9 months
Diet 6 months
Corn oil "A few days1'
Corn oil 44 days
Corn oil 106 days
Estimated
Dose cumulative
(mg/kg/day or dose
ppm in diet) (mg/kg)
50 mg/kg/day >300
(250-480 ppm)
189 ppm
1.2-4.8 mg/kg/day -v.250-1000
50 and 200 ppm
100-800 ppm
5 mg/kg/day ^15-35
4 mg/kg/day 176
4 rag/kg/day 424
Response
Changes in blood chemistry and
urine protein
No apparent adverse effects
Temporary change in blood chemistry
Questionable liver pathology
No significant effect
Convulsion
Questionable liver pathology;
renal tubular degeneration
Questionable liver pathology;
renal tubular degeneration
Reference
Baeumler, 1975
Clapp et al. , 1971
Grebenyuk, 1970
Ortega et al. , 1951
Shelanski and
Gellhorn,
undated
Lackey , 1949 a
Lackey, 1949 a
Lackey, 1949 a
I See text for details.
N.S. - not specified.
-------
hepatic cells. This condition, referred to as hydropic accumulation, is dis-
tinct from fatty degeneration. In neither rats nor dogs was hydropic accumu-
lation associated with the destruction of hepatic cells. However, Ortega and
coworkers (1957) also noted occasional masses of red blood cells invading the
cytoplasm of liver cells in areas of hypertrophy and margination. In addition
to liver damage, Lackey (1949 a) also noted wide-spread degeneration of the
tubular epithelium, occasionally accompanied by inflammation of the pelvis of
the kidney. Identical pathological changes were seen in dogs surviving pro-
longed dermal exposures to toxaphene (Lackey, 1949 b).
As noted in Table 6.15, alterations in clinical chemistry have also been
seen in subacute oral toxaphene exposures. Mice with no clinical signs of
intoxication evidenced consistent increases in serum acid phosphatase, glutamic-
pyruvic transaminase, and gamma-glutyamyl transpeptidase activities, along
with increased neutrophil counts and changes in urine protein (Baeumler, 1975).
At a much lower daily dose, rats had only a transient increase in serum alka-
line phosphatase during the fifth month of intoxication and showed no variation
in urine hippuric acid (Grebenyuk, 1970). Increases in all of the above en-
zyme activities are consistent with the mild liver pathology associated with
subacute toxaphene exposure.
Lehman (1952 b) states that the 90 day dermal LD50 of toxaphene (as a
dry wax) is 40 mg/kg in rabbits. No details of symptoms or pathology are
provided.
The subacute inhalation toxicity of toxaphene is summarized in Table 6.16.
Details of these studies were not available for this review. Hercules Inc.
(undated) indicates that, unlike subacute oral exposures, these inhalation
exposures did not result in abnormal blood chemistry or hematologic measurements.
6-44
-------
a.fa
Table 6.16. Subacute inhalation toxicity of toxaphene '
Concentration
Organism (ing/liter air) Form
Rat
Rats and rabbits
Rats and rabbits
Rats
Rats, dogs, and
guinea pigs
Rats
Dogs
Guinea pigs
Rats
Dogs
Guinea pigs
0.25
0.04
0.20
0.50
0.004
0.012
0.012
0.012
0.04
0.04
0.04
Dust
Mist
Mist
Mist
Dust
Dust
Dust
Dust
Dust
Dust
Dust
Duration
1 week
3 weeks
3 weeks
3 weeks
3 months
3 months
3 months
3 months
3 months
3 months
3 months
Response
100% mortality
No observed effects
No observed effects
No observed effects
No observed effects
No mortality
50% mortality
No mortality
27% mortality
67% mortality
20% mortality
a.
/Source: Hercules Inc., undated.
All exposures far 6 hours./day, 5 days/week.
6-45
-------
The most pronounced effect prior to death was severe weight loss. As in
subacute oral exposures, female rats had slight focal cellular necrosis of
the liver.
Hercules Inc. (undated) has conducted controlled dermal and inhalation
exposures of humans to toxaphene. Toxaphene doses of 300 mg/day applied to
the skin of 50 volunteers for 30 days produced no gross toxic effects. Simi-
larly, cotton patches treated with toxaphene produced neither sensitization
nor primary skin irritation when applied to the skin of 200 subjects. In
daily 30 minute inhalation exposures to toxaphene mists of 500 mg/cu m air,
humans exposed for ten consecutive days followed by three daily exposures
three weeks later showed no adverse effects based on physical examinations as
well as blood and urine tests (Shelanski, 1974).
However, Warraki (1963) has attributed two cases of acute bronchitis with
military lung shadows to inhalation of toxaphene during applications of toxa-
phene spray. Both individuals, male adults, had been exposed to toxaphene
sprays from 1 1/2 to 2 months before the onset of pulmonary insufficiency.
Maximum breathing capacity was between 19 and 22 percent of normal. All ad-
verse effects, including pulmonary lesions, were reversible within three
months after toxaphene exposure was discontinued. No central nervous system
effects were noted. One case of allergic rhinitis in a worker exposed to
toxaphene by inhalation has been reported. However, details on the duration
of exposures were not given (Pesticide Episode Response Branch, 1976).
6.3.2.3 Chronic Toxicity — Long term exposures to low dietary levels of
toxaphene are summarized in Table 6.17- All studies note some form of liver
pathology in rats at dietary levels of 100 ppm or above. At 100 ppm, cytoplasmic
6-46
-------
Table 6.17. Chronic toxicity of toxaphene at low dietary levels to laboratory
mammals
Organism
Rats,
Sprague Dawley
Racs
Rats
Rats
Duration
of feeding
3 generations
Lifetime
Lifetime
2 years
2 years
Toxaphene
concentration
in diet
25 ppm
100 ppm
25 ppm
100 ppm
25 ppra
25 ppm
100 ppm
1000-1600 ppm
Dogs
Dogs
Dogs
Monkeys
2 years
2 years
1360 days
(V3.7
years)
2 years
5-20 ppm
40 ppm
200 ppm
5 mg/kg/day
10-15 ppm
(•x-0.64-
0.78 mg/kg/day)
a.
Response
No effect
Liver pathology
No effect
Liver pathology
Liver pathology
No effect
Slight liver
damage
CNS stimulation
No effect
Slight liver
degeneration
Moderate liver
degeneration
Liver necrosis
No clinical or
histological
effects
Reference
Kennedy et al. , 1973
Lehman, 1952 a
Fitzhugh and Nelson,
1951
Hercules Inc. , undated
Hercules Inc. , undated
Hercules Inc. , undated
Hercules Inc. , undated
Administered in capsules containing toxaphene dose in corn oil; 5 mg/kg/day
equivalent to 200 ppm in diet.
6-47
-------
vacuolization similar to that seen on subacute oral exposure was noted by
Kennedy and coworkers (1973). Lehman (1952 a) noted both cytoplasmic vacuoli-
zation and fatty degeneration of the liver in rats fed 100 ppm. At 25 ppm,
Fitzhugh and Nelson (1951) did observe increased liver weight with minimal
liver cell enlargement. Unpublished studies on rats, dogs, and monkeys by
Hercules Inc. (undated) are in general agreement with the above published re-
ports. The lowest dietary level of toxaphene producing unequivocal liver
damage over a two year feeding period is 20 ppm. Only at relatively high
concentrations - i.e., 1000 ppm - does chronic toxaphene exposure elicit
central nervous system effects characteristic of acute intoxication. The
relevance of these studies to other aspects of toxaphene toxicology is dis-
cussed in subsequent sections.
No cases of chronic human intoxication have been encountered in the
literature. In an epidemiclogical survey of workers exposed to toxaphene and
a variety of other pesticides, no evidence of abnormal heme synthesis or ab-
normal sympathoadrenal activity - based on urinary analysis of unconjugated
catecholamines - has been noted (Embry et al., 1972).
6.3.2.4 Carcinogenicity — None of the chronic feeding studies detailed in
Section 6.3.2.3 noted any increase of tumor incidence in the experimental
animals. However, a closely related pesticide, Strobane (chlorinated ter-
penes), has been shown to cause an increase in tumor production in male but
not female mice (Table 6.18, Innes et al., 1969). In this study, two strains
of mice were used and designated strain X (C57BL/6xC3H/Anf,F1) and strain Y
(C57BL/6xAKR,Fl) . Seven days after birth, the mice received 4.64 mg/kg/day
of Strobane in 0.5 percent gelatin by intubation until the mice were four
6-48
-------
Table 6.18. Tumors in mice associated with oral administration of strobane
a
Strain
Number of
tumors
Relative Risk
compared to
positive control
1
vo Relative Risk
compared to
positive control
X
Y
X
Y
X
Y
Number of mice
necroscopied Hepatomas
(M) (F) (M) (F)
15 18 2
18 1.8 LI
1.56
i.
23.846
0.136
2.04
0
0
0
0
0
0
Pulmonary tumors Lymphomas Total tumors
(M) (F) (M) (F) (M) (F)
11 52 8
00 0 0 11
b
1.40 2.07 7.10 2.81 2.90
A,
00 00 6.92
0.550
1.110
3
0
2.11
0
0.099
0
Sum
c
2.56
„
4.73
.265
.704
.Source: Modified from Innes et al., 1969.
increased tumor yield significant at 0.01 level.
Increased tumor yield significant at 0.05 level.
-------
weeks old. The dose was calculated by the weight of the seven day old mice
and not adjusted for weight gain. At age four weeks, Strobane was administered
in the food at a concentration of 11 ppm. Based on the weight and food con-
sumption at four weeks, this concentration resulted in daily doses of about
4.6 mg/kg. This schedule was maintained for eighty weeks, using thirty-six
mice of each strain evenly divided by sex. Male mice of strain X developed a
significantly (p less than 0.01) higher incidence of lymphomas. Males of
strain Y, however, showed significant (p less than 0.01) increases only in
hepatomas.
In this study, hepatomas are classified as liver tumors which have not
metastasized. Such a classification does not imply "that these tumors are
benign. Indeed, it seems more reasonable to conclude that the great majority
had malignant potential" (Innes et al., 1969). The positive control consisted
of combined data from mice exposed to seven known carcinogens: urethane,
ethylene imide, amitrol, aramite, dihydrosafrole, isosafrole, and safrole.
Although the increased incidence of hepatomas in male mice of strain Y admini-
stered Strobane compared to the positive control has no absolute definitive
value, it is indicative of highly positive tumorigenic potential.
Under contract to the National Cancer Institute, Gulf Research Institute
has recently completed a carcinogenicity bioassay of toxaphene (National Cancer
Institute, 1979). It should be noted that this study, which was conducted from
1971 to 1973, did not follow current NCI protocols (NCI, 1977). Specifically,
only ten organisms were used in matched control groups, and matched-fed control
groups were not utilized. In this study, groups of Osborne-Mendel rats and
B6C3F1 hybrid mice were exposed to technical-grade toxaphene in the diet for
80 weeks. Details of the dose schedule and number of organisms used are pro-
vided in Tables 6.19 and 6.20.
6-50
-------
Table 6.19. Toxaphene chronic feeding studies in rats
Sex and Initial
Test No. of ,
Group Animals
MALE
Matched-control 10
Low-dose 50
High-dose 50
ui FEMALE
t— •
Matched-control 10
Low-dose 50
High-dose 50
Toxaphene
in diet13
(mg/kg)
0
1,280
640
320
0
2,560
1,280
640
0
0
640
320
0
1,280
640
0
Time
(weeks)
2
53
25
2
53
25
55
25
55
25
on Study Time-weighted Average
Observed doseJ
(weeks) (mg/kg)
108-109
556
28
1,112
28
108-109
540
30
1,080
30
^National Cancer Institute, 1979.
All animals were 5 weeks of age when placed on study.
^Initial doses shown were toxic; therefore, doses were lowered after 2 weeks and again at 53 or 55 weeks,
,as shown.
All animals were started on study on the same day.
eWhen diets containing toxaphene were discontinued, dosed rats and their matched controls were fed control
,,diets without corn oil for 20 weeks, then control diets (2% corn oil added) for an additional 8 weeks.
•'Time-weighted average dose _ £(dose in ppm x no. of weeks at that dose)
Z(No. of weeks receiving each dose)
-------
Table 6.20. Toxaphene chronic feeding studies in mice'
Sex and
Test
Group
MALE
Matched-control
Low-dose
High-dose
FEMALE
^ Matched-control
ro
Low-dose
High-dose
Initial
No. of ,
Animals
10
50
50
10
50
50
Toxaphene
in diet0
(mg/kg)
0
160
80
0
320
160
0
0
160
80
0
320
160
0
Time
Dosed^
(weeks)
19
61
19
61
19
61
19
61
on Study
Observed
(weeks)
90-91
11
10
90-91
11
10
Time-weighted Average
dose-'
(mg/kg)
99
198
99
198
^National Cancer Institute, 1979.
All animals were 5 weeks of age when placed on study.
^Initial doses shown were toxic; therefore, doses were lowered at 19 weeks, as shown.
All animals were started on study on the same day.
When diets containing toxaphene were discontinued, dosed mice and their matched controls were fed control
,4iets without corn oil for 7 weeks, then control diets (2% corn oil added) for an additional 3 to 4 weeks.
^Time-weighted average dose _ E(dose in ppm x no. of weeks at that dose)
E(No. of weeks receiving each dose)
-------
Toxaphene was added to the feed in acetone. In addition, 2 percent corn
oil was added to the diet as a dust suppressant. Actual dietary toxaphene
concentrations, which were confirmed by gas-liquid chromatography, did not
deviate from the nominal concentration by more than 6.9 percent. In addition
to the matched control groups indicated in these tables, pooled control groups
were used in the statistical analyses. For rats, pooled controls consisted of
matched controls from similar bioassays on captan, chloraben, lindane, mala-
thion, and picloram, as well as the matched controls from the toxaphene bio-
assay. For mice, pooled controls consisted of matched controls from similar
bioassays on lindane, malathion, phosphamidon, and tetrachlorvinphos, as well
as the matched controls from the toxaphene study. Organisms used in all pooled
control groups were of the same strains, from the same suppliers, and examined
by the same pathologists.
During the course of this study, both rats and mice evidenced signs of
general toxic effects. Both male and female rats in the high-dose group de-
veloped body tremors at week 53. From week 52 to week 80, other clinical signs,
which occurred primarily in toxaphene-dosed rats, included diarrhea, dyspnea,
pale mucous membranes, alopecia, rough hair coats, dermatitis, ataxia, leg
paralysis, epistaxis, hematuria, abdominal distention, and vaginal bleeding.
Female rats in both dose groups had lower mean body weights than the matched
controls. No dose-related effect on mortality was noted in any of the rat
test groups. In mice, males and females in each dose group displayed a signi-
ficant increase in mortality when compared to the matched controls. In high-
dose male mice, mean body weights were generally lower than those in the
matched control group. Clinical signs of toxicity in mice included abdominal
distention, diarrhea, alopecia, rough hair coats, and dyspnea.
6-53
-------
The effects of dietary toxaphene on tumor incidence in male rats, female
rats, male mice, and female mice are summarized in Tables 6.21, 6.22, 6.23, and
6.24, respectively.
In male rats in the high dose group, a significant increase was noted in
the incidence of follicular-cell carcinomas or adenomas of the thyroid. Of the
nine thyroid tumors which were found in this group, two were carcinomas. A
significant increase of follicular-cell adenomas of the thyroid was also noted
in the high-dose group of female rats. No carcinomas of the thyroid were found
in this group. In both of these groups, the development of thyroid tumors was
dose-related. A significant increase was also noted in the incidence of chromo-
phobes, adenomas, chromophobe carcinomas, and adenomas of the pituitary in the
high-dose group female rats. However, an examination of historical control
data on the incidence of pituitary tumors in female rats suggested that an
association between the administration of toxaphene and the development of
pituitary tumors could not be maintained based on the results of this study.
In both male and female mice, significant increases were noted in the
incidence of hepatocellular carcinomas and in the incidence of hepatocellular
carcinomas combined with neoplastic nodules of the liver.
Based on the results of this study, the National Cancer Institute has con-
cluded that "Toxaphene was carcinogenic in male and female B6C3F1 mice, causing
increased incidences of hepatocellular carcinomas. The test results also sug-
gest carcinogenicity of toxaphene for the thyroid of male and female Osborne-
Mendel rats" (National Cancer Institute, 1979).
6.3.2.5 Mutagenicity — Epstein et al. (1972) have used a modified dominant
lethal assay in mice to evaluate the mutagenic potential of a variety of chemical
agents including toxaphene. In this study, four groups of male ICR/Ha Swiss mice
6-54
-------
Table 6.21. Analyses of the Incidence of primary tumors in male rats fed toxaphene in the diet
a.,£>
I
Ui
Ui
Topography; Morphology
Liver: Neoplastic Nodule
P Values
Weeks to First Observed Tumor
Pituitary: Chromophobe Adenoma,
Carcinoma, NOS, or Adenoma, NOS
P Values
Weeks to First Observed Tumor
Adrenal: Adenoma, NOS, Cortical
Adenoma, or Carcinoma
P Values^6
Weeks to First Observed Tumor
Spleen: Hemangioma
P Value sd
Weeks to First Observed Tumor
f
Thyroid: Follicular-cellJ
Carcinoma or Adenomac
P Values
Weeks to First Observed Tumor
Matched
Control
1/9 (11)
N.S.
109
3/7 (43)
N.S.
102
4/9 (44)
P = 0.019
0/9 (0)
N.S.
—
1/7 (14)
N.S.
109
Pooled
Control
1/52 (2)
N.S.
- —
8/46 (17)
N.S.
—
5/52 (10)
(N) N.S.
0/49 (0)
N.S.
—
2/44 (5)
P - 0.007
—
Low
Dose
6/44 (14)
P = 0.034**
108
13/42 (31)
N.S.
85
5/41 (12)
P - 0.043 (N)*
85
3/45 (7)
N.S.
83
7/41 (17)
N.S. P
104
High
Dose
4/45 (9)
N.S.
94
5/31 (16)
N.S.
95
3/37 (8)
P - 0.020 (N)*
85
3/42 (7)
N.S.
85
9/35 (26)
- 0.008**
56
^National Cancer Institute, 1979.
Dosed groups received time-weighted average doses of 556 or 1,112 ppm.
Number of tumor-bearing animals/number of animals examined at site (percent).
Beneath the incidence of tumors in a control group is the probability level for the Cochran-Armitage test when
P less than 0.05; otherwise, not significant (N.S.) is indicated. Beneath the incidence of tumors in a dosed
group is the probability level for the Fisher exact test for the comparisons of that dosed group with the
matched-control group (*) or with the pooled-control group (**) when P less than 0.05 for either control group;
otherwise, not significant (N.S.) is indicated.
,A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
•'Data being reevaluated. See Test.
-------
Table 6.22. Analyses of the incidence of primary tumors in female rats fed toxaphene in the diet
Topography: Morphology
Integumentary System: Malignant
Fibrous Histiocytoma of the Sub-
cutaneous Tissue0
P Values
Weeks to First Observed Tumor
Mammary Gland: Fibroadenoma
P Values
Weeks to First Observed Tumor
Liver: Hepatocellular Carcinoma
or Neoplastic Nodule*3
P Values
Weeks to First Observed Tumor
Pituitary: Chromophobe Adenoma,
Carcinoma, or Adenoma, NOSC
P Values
Weeks to First Observed Tumor
Q
Thyroid: Follicular-celle Adenoma
P Values
Weeks to First Observed Tumor
Matched
Control
0/10 (0)
N.S.
—
0/10 (10)
N.S.
87
1/10 (10)
N.S.
109
3/8 (38)
P = 0.046
85
0/6 (0)
P = 0.022
—
Pooled
Control
0/55 (0)
N.S.
6/55 (11)
N.S.
—
1/55 (2)
N.S.
—
17/51 (33)
P = 0.012
—
1/46 (2)
P = 0.008
—
Low
Dose
1/50 (2)
N.S.
105
10/50 (20)
N.S.
19
5/42 (12)
N.S.
108
15/41 (37)
N.S.
75
1/43 (2)
N.S.
102
High
Dose
3/49
N.S.
83
10/49
N.S.
67
4/40
N.S.
109
23/39
P = 0.
79
7/42
P = 0.
105
(6)
(20)
(10)
(59)
013**
(17)
021**
-------
Table 6.22. Analyses of the incidence of primary tumors in female rats fed toxaphene in the diet
(Cont'd)
ON
Oi
Topography; Morphology
Adrenal: Cortical Adenoma or
Carcinoma13
P Values
Weeks to First Observed Tumor
Uterus: Endometrial Stromal
Polyp^
P Values0
Weeks to First Observed Tumor
Matched
Control
0/8 (0)
N.S.
—
0/9 (0)
N.S.
—
Pooled
Control
3/50 (6)
N.S.
—
5/53 (9)
N.S.
—
Low
Dose
3/44 (7)
N.S.
104
9/41 (22)
N.S.
87
High
Dose
6/43
N.S.
87
5/45
N.S.
109
(14)
(11)
^National Cancer Institute, 1979.
Dosed groups received time-weighted average doses of 540 or 1,080 mg/kg.
^Number of tumor-bearing animals/number of animals examined at site (percent).
Beneath the incidence of tumors in a control group is the probability level for the Cochran-Armitage test
when P less than 0.05; otherwise not significant (N.S.) is indicated. Beneath the incidence of tumors in
a dosed group is the probability level for the Fisher exact test for the comparison of that dosed group
with the matched-control group (*) or with the pooled-control group (**) when P less than 0.05 for either
control group; otherwise, not significant (N.S.) is indicated.
Data being reevaluated. See text.
-------
Table 6.23. Analyses of the incidence of primary tumors in male mice fed toxaphene in the diet
Ln
00
Topography: Morphology
Liver: Hepatocellular Carcinoma
P Values
Weeks to First Observed Tumor
Liver: Hepatocellular Carcinoma
or Neoplastic Nodulec
P Values
Weeks to First Observed Tumor
Matched
Control
0/10 (0)
P less than
0.001
—
2/10 (20)
P less than
0.001
90
Pooled
Control
4/48 (8)
P less than
0.001
—
7/48 (15)
P less than
0.001
—
Low
Dose
34/49 (69)
P less than
0.001*
P less than
0.001**
73
40/49 (82)
P less than
0.001*
P less than
0.001**
73
High
Dose
45/46 (98)
P less than
0.001*
P less than
0.001**
59
45/46 (98)
P less than
0.001*
P less than
0.001**
59
^National Cancer Institute, 1979.
Dosed groups received time-weighted average doses of 99 or 198 mg/kg.
^Number of tumor-bearing animals/number of animals examined at site (percent).
Beneath the incidence of tumors in a control group is the probability level for the Cochran-Armitage test
when P less than 0.05; otherwise not significant (N.S.) is indicated. Beneath the incidence of tumors in
a dosed group is the probability level for the Fisher exact test for the comparison of that dosed group
with the matched-control group (*) or with the pooled-control group (**) when P less than 0.05 for either
control group; otherwise, not significant (N.S.) is indicated.
-------
Table 6.24. Analyses of the incidence of primary tumors in female mice fed toxaphene in the diet
I
^o
Topography: Morphology
Liver: Hepatocellular Carcinoma
P Values^
Weeks to First Observed Tumor
Liver: Hepatocellular Carcinoma
or Neoplastic Nodulec
P Values
Weeks to First Observed Tumor
Matched
Control
0/9 (0)
P less than
0.001
—
0/9 (0)
P less than
0.001
—
Pooled
Control
0/48 (0)
P less than
0.001
—
0/48 (0)
P less than
0.001
—
Low
Dose
5/49 (10)
P = 0.030**
89
18/49 (37)
P = 0.026*
P less than
0.001**
89
High
Dose
34/49 (69)
P less than
0.001
P less than
0.001**
72
40/49 (82)
P less than
0.001*
P less than
0.001**
72
^National Cancer Institute, 1979.
Dosed groups received time-weighted average doses of 99 or 198 mg/kg.
^Number of tumor-bearing animals/number of animals examined at site (percent).
Beneath the incidence of tumors in a control group is the probability level for the Cochran-Armitage test
when P less than 0.05; otherwise not significant (N.S.) is indicated. Beneath the incidence of tumors in
a dosed group is the probability level for the Fisher exact test for the comparison of that dosed group
with the matched-control group (*) or with the pooled-control group (**) when P less than 0.05 for either
control group; otherwise, not significant (N.S.) is indicated.
-------
were given toxaphene intraperitoneally - single doses of 36 mg/kg and
180 mg/kg - and orally - five doses of 8 mg/kg/dose and 16 mg/kg/dose.
After dosing, the treated males were mated to groups of untreated females
over an 8-week period. Based on measurements of early fetal deaths per
pregnancy and the percent of females with early fetal deaths, the toxaphene-
treated groups did not differ significantly from controls. Thus, in this
strain of mice, toxaphene apparently does not produce chromosomal abnormali-
ties that preclude zygote development.
Hill (1977) has summarized information on the mutagenicity testing of
toxaphene in bacterial systems. Ames tests have been conducted on Salmonella
typhimurium strains TA-1535, TA-1537, TA-1538, TA-98, and TA-100 with and
without metabolic activation by non-induced mammalian liver fractions. Posi-
tive results were obtained for strains TA-98 (frameshift mutation) and TA-100
(base pair substitution) only in tests without metabolic activation. All other
tests were negative. A "high temperature" toxaphene has elicited positive dose
response increases in strains TA-98 and TA-100 only with metabolic activation.
All the above tests were, conducted by Litton Bionetics Inc. for Hercules Inc.
In addition to these studies, work has been conducted on the mutagenicity
of toxaphene in the Salmonella system by Dr. Kim Hooper of Bruce Ames' group
in Berkeley, California (Hill, 1977). His results indicate that toxaphene
and toxaphene subfractions are mutagenic to strain TA-100 with and without
activation by Aroclor-induced rat microsomes. Mutagenic activity was decreased
in those tests using microsomal activation.
A recently completed study by U.S. EPA (1978) found no significant differ-
ences in the rates of chromosomal aberrations in leukocytes between groups of
individuals occupationally exposed to toxaphene and groups with no occupational
exposures to toxaphene.
6-60
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6.3.2.6 Teratogenicity — In a study by Kennedy et al. (1973), male and
female rats were fed dietary levels of 25 mg/kg diet and 100 mg/kg toxaphene.
Gross and microscopic pathology of F3 weanlings revealed no indication of
teratogenic effects. Further, no statistically significant variations from
controls were noted in either dose group for any of the following parameters:
mating index, fertility index, pregnancy index, parturition index, mean viable
liter size, live birth index, 5-day survival index, lactation index, or wean-
ing body weights of offspring. One of 16 females from each dose group resorbed
an entire litter. This was not seen in any of the 32 control females but did
occur in tests with another pesticide, delnav.
In multigeneration studies of mice given toxaphene at 25 mg/kg diet, no
effects on fertility, gestation, viability, lactation, or survival indices were
observed (Keplinger et al., 1970).
In addition to these long-term dietary studies, one study (Chernoff and
Carver, 1976) has been conducted in which toxaphene in corn oil was adminis-
tered to pregnant female rats and mice from days 7 to 16 of gestation at doses
of 15, 25, and 35 mg/kg/day. All doses produced signs of maternal and fetal
toxicity but no teratogenic effects.
DiPasquale (1977) has examined the effects of toxaphene on fetal guinea
pig development. In this study, toxaphene was administered to pregnant females
at a dose of 15 mg/kg body weight orally from day 21 to day 35 of gestation.
No effects were noted on anatomical development of the fetus. The only sign
of fetotoxicity was a decrease in collagen-containing structures. This was
attributed to a functional deficiency of vitamin C related to mixed-function
oxidase induction. Maternal guinea pigs showed a slight loss of body weight
but no effects were seen on maternal liver weight or mortality.
6-61
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6.3.2.7 Drug Interactions— As indicated in Table 6.25, toxaphene offers a
slight protective effect to rats when administered by intubation at equitoxic
doses with three of seven tested pesticides (Keplinger and Deichmann, 1967).
Because both pesticides were given at about the same time, the protective
effect is probably not attributable to liver microsomal mixed-function oxidase
induction. However, hepatic microsomal mixed-function oxidase induction may
account for the protective effects of aldrin and dieldrin on toxaphene toxi-
city to rats noted by Deichmann and Keplinger (1970). In this study, the 96
hour LD50 of toxaphene to rats receiving no pretreatment was 78 mg/kg. In
rats given single oral doses of aldrin or dieldrin at 30 mg/kg four days prior
to toxaphene dosing, the 96 hour LD50's of toxaphene were 180 mg/kg and 165
mg/kg, respectively. This decrease in toxaphene toxicity corresponded to
increased 0-dealkylase and 0-demethylase activities in the liver homogenates
of pretreated animals. A decrease in toxaphene toxicity was also seen in rats
after feeding on diets containing 400 ppm DDT for two weeks. In these animals,
the 96 hour LD50 of toxaphene was 215 mg/kg, about three times that seen in
untreated controls.
Toxaphene has been found to have no ability to supress pentylene tetar-
zole-induced convulsions in rats (Ghazal, 1965).
As discussed in Section 6.3.2.1.1.2, dermal exposure to toxaphene/lindane
mixtures has been associated with the development of acute aplastic anemia in
man. This observation, however, is based solely on rather limited epidemio-
logical data.
6-62
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Table 6.25. Expected and observed oral LD50's of toxaphene plus other
pesticides in ratsa
Other
pesticide
Malathion
Carbaryl
V-C 13
Delnav
Parathion
Diazinon
Trithion
Expected LD50
(rag/kg)
1000
445
120
90
67
150
65
Observed LD50
(mg/kg)
820
524
146
112
100
227
121
Ratio
E/0
1.22
0.85
0.82
0.80
b
0.67
b
0.66
b
0.54
-Source: Keplinger and Deichmann, 1967.
Ratio considered indicative of protective effects by toxaphene.
6-63
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/
i Innes, J.R.M. , Ulland, B.M. , Valerio, M.G. , Petrucelli, L. , Fishbein, L. ,
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/Kennedy, G.L., Jr., Frawley, J.P., and Calandra, J.C. 1973. Multigeneration
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Technical Toxaphene. J. Agr. Food Chem. 22(4);653-657.
Khalifa, S., Holmstead, R.L., and Casida, J.E. 1976. Toxaphene Degradation
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/ Kinoshita, F. , Frawley, J.P., and DuBois, K.P. 1966. Quantitative Measurement
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Juiz'minskaya, U.A. and Alekhina, S.M. 1976. Effect of Chlorocamphene on the
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in the Rabbit and Dog. J. Ind. Hyg. Toxicol. 31:155-157.
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I
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6-67
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6-68
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7.0 ENVIRONMENTAL DISTRIBUTION AND TRANSFORMATION
7.1 SUMMARY
Soil and water monitoring studies suggest that toxaphene is not as wide
spread an environmental contaminant as other chlorinated hydrocarbon insecti-
cides, such as DDT and dieldrin. Toxaphene is rarely detected in soil, water,
or sediment samples that have not received direct applications of the pesti-
cide for crop protection or rough fish control. This may be due, in part, to
the difficulty encountered in trace analysis of such a complex mixture of com-
pounds and the high volatility of toxaphene. Sampling of western North Atlantic
Ocean air indicates that mean levels of toxaphene are equal to or twice those
of PCB's and ten times higher than those of other pesticides such as DDT.
This probably results from both the large scale use of toxaphene and its vola-
tility. These results also suggest that toxaphene, which is detected near
the island of Bermuda, has travelled 1200 km.
Degradation of toxaphene in the environment is poorly understood. Several
researchers have noted that longer retention time (less volatile) peaks in
the gas chromatogram of toxaphene residue decrease while shorter retention time
(more volatile) peaks increase. Residues from water and sediment do not appear
to be products of dehydrohalogenation. The rate of degradation varies considerably.
One study determined that the concentration of toxaphene in sediment decreased
by a factor of 2 every 20 days. However, concentrations in lakes treated
3 to 9 years before monitoring contained 1 to 4 micrograms per liter iij water
and 0.2 to 1 ppm in sediment. Adsorption to sediment and suspended solids
appears to be an important process for removing toxaphene from water. The
calculated half-life for evaporation of toxaphene from water is 10.4 hours.
7-1
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The reported half-residence time for toxaphene in soil varies from
100 days to 11 years. The latter is considered to be a maximum value. Most
studies suggest that little leaching of toxaphene through soil occurs, al-
though one investigation found toxaphene in ground water below a treated plot.
7.2 MONITORING
7.2.1 Residues in Air
The highest toxaphene residues in air have been found in areas where it
is applied for agricultural purposes (especially cotton production) (Arthur et al.,
1976; Stanley et al., 1971; Tabor, 1965, 1966). Studies in cotton growing
areas demonstrate that airborne residues are highest during the cotton growing
season and decrease to low levels after harvesting, but spring tilling releases
soil residues to the air. The recent identification of ng/cu m levels over the
Atlantic Ocean, where toxaphene has not been applied, establishes that toxaphene
residues move with air currents analagous to DDT (Bidleman et al., 1976; Bidleman
and Olney, 1975).
Arthur et al. (1976) reported a three year (January 1972 to December 1974)
study of toxaphene residues at Stoneville, Mississippi, which is located in the
southern cotton belt. They sampled on a weekly schedule in which they collected at
intervals of 4.29 minutes per hour (12 hours per week). They used a MISCO
Model 88 air sampler containing ethylene glycol as the trapping solvent.
Table 7.1 summarizes the average monthly toxaphene concentration for the
three years. The highest average monthly concentrations were detected during
July and August. The highest average weekly concentration was 1746.5 ng/cu m.
Although they do not explain the sharp decrease in average residues for 1973
compared to 1972, they attribute the increase in 1974 over 1973, in part,
to a 22 percent increase in toxaphene use.
7-2
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Table 7.1. Average monthly atmospheric levels of toxaphene (ng/cu m)
in Stoneville, Mississippi^
January
February
March
April
May
June
July
August
September
October
November
December
Average
1972
0.0
13.0
68.0
67.4
32.4
44.2
400.7
1540.0
827.9
97.9
9.3
0.0
258.4
1973
0.0
0.0
16.8
10.8
46.8
109.9
41.1
268.8
322.6
161.1
0.0
9.9
82.3
1974
10.9
9.7
19.1
27.7
44.3
38.6
175.0
903.6
524.6
114.8
32.9
12.6
159.5
Source: Arthur et al., 1976.
-------
Stanley et al. (1971) examined toxaphene and 18 other pesticides and
pesticide metabolites for six months at nine locations: Baltimore, Maryland
(urban); Buffalo, New York (rural); Dothan, Alabama (rural); Fresno, California
(urban); Iowa City, Iowa (rural); Orlando, Florida (rural); Riverside, California
(urban); Salt Lake City, Utah (urban); and Stoneville, Mississippi (rural).
The sites were sampled during 1967 and 1968; sites were examined at different
times during this period. They sampled twice a month for seven day intervals,
with air being collected for either 12 or 24 hours per day. Toxaphene was
only detected at three rural locations in southern states. Table 7.2 lists
the maximum levels and the number of samples containing toxaphene. Toxaphene
residues on the first day of each sampling week at Stoneville, Mississippi,
are summarized in Table 7.3.
Tabor (1965, 1966) monitored air in seven communities located in southern
agricultural areas and three eastern urban areas. Toxaphene was detected only
in communities located in the two cotton growing areas (Leland, Washington
County, Mississippi, and Newellton, Texas). Toxaphene residues were not detec-
ted in the urban communities or the five rural communities located in fruit
or vegetable growing areas. At Leland, toxaphene was detected in 6 of 15
samples collected from July to September 1963; concentrations ranged from
1.2 to 7.5 ng/cu m. Toxaphene was found in 6 of 10 samples taken during the
same period at Newellton, Texas (3.1 to 15 ng/cu m).
Bidleman and Olney (1975) measured atmospheric toxaphene residues over
the east coast of the U.S., near Bermuda, and over the open ocean (Western
North Atlantic Ocean) during 1973 to 1974. They collected residues by draw-
ing 200 to 3000 cu m of air through a device consisting of a Gelman A glass
7-4
-------
Table 7.2. Maximum toxaphene levels found in air samples of nine localities
Site
Baltimore
Buffalo
Do than
Fresno
Iowa City
Orlando
Riverside
Salt Lake City
Stoneville
Number of
total samples
123
57
90
120
94
99
94
100
98
Toxaphene range Samples containing
(ng/cu m) toxaphene
0
0
27.3 - 79.0 11
0
0
20.0 - 2520 9
0
0
16.0 - 111.0 57
a
Source: Adapted from Stanley, 1968, and Stanley et al., 1971.
-------
Table 7.3. Toxaphene found in air samples from Stoneville,
Mississippi the first day of each sampling weeka
Date
August 14-16
August 21-22
September 11-12
October 2-3
July 1-2
July 15-16
July 29-30
August 12-13
Toxaphene level, ng/cu m
283
373
701
161
68
116
62
135
Source: Stanley et al., 1971.
7-6
-------
fiber filter and a polyurethane foam plug (Section 2.6.2). Toxaphene was
analyzed by standard gc procedures, both with and without alkaline dehydro-
chlorination (Section 2.5.2). Although toxaphene had weathered, the collected
residues were clearly recognized. Figure 7.1 and Table 7.4 describe the
-sampling sites and measured residues. At Bermuda, 36 samples were collected:
26 when winds blew from inside a 90 E to 240 SW selected sector (ISS) (air
passed over hundreds of kilometers of open ocean and not over the island of
Bermuda); 7 when winds blew from outside this selected sector (OSS) (over
portions of the island); and 3 at periods of variable winds. Bidleman and
Olney (1975) concluded that toxaphene residues did not originate from Bermuda
for the following reasons: (1) toxaphene had not been used on Bermuda for
the prior 3 to 4 years; (2) mean concentrations for measurements OSS
(0.81 + 0.45 ng/cu m) and ISS (0.72 + 0.09 ng/cu m) were similar; and (3) com-
parable residue levels were measured over the open ocean and over Bermuda.
The authors concluded that the logical source of toxaphene was the U.S.
cotton belt, 1200 km away.
Bidleman et al. (1976) monitored toxaphene residues at five North
American sites for benchmark data. Sample collection followed the same pro-
cedure as that employed in the sampling of air over the Western North Atlantic.
Table 7.5 describes the residues detected at the five sites. Toxaphene and
other chlorinated hydrocarbons were never applied at the sites sampled. The
lowest residue levels were measured at the northern sites (Rhode Island and
Canada) and the highest levels at the southern sites (in Georgia and Arizona).
7.2.2 Residues in Water and Sediment
Although toxaphene has not been detected in any routine monitoring studies
of U.S. surface waters (Weaver et al., 1965; Brown and Nishioka, 1967;
7-7
-------
- 2O
60U
Figure 7.1. Atmospheric samples collected from the RV Trident, 1973 - 1974
—» (see Table 7-4 for cruise dates)
§a, TR-137; O, TR-145; 3) , TR-152; A, TR-153.
Source: Bidleman and Olney, 1975.
7-J
-------
Table 7.4. Concentrations of airborne toxaphene over the Western North Atlantic
a
Location
Dates
Toxaphene (ng/cu m)
Bermuda
32°15'N 64°50'W
Bermuda
32°15'N 64°50'W
Sapelo Island, Georgia
31°15'N 81°05'W
Cruise TR-137
33°20'N, 65°15'W
34°40'N, 66°15'W
37°40'N, 68°10'W
38°50'N, 69°15'W
40°30'N, 70°20'W
Cruise TR-145
32°00'N, 65°00'W
32°40'N, 67°20'W
36°40'N, 75°20'W
37°QO'N, 76°00'W
37°20'N, 76°10'W
40°30'N, 72°40'W
Mar. - Dec., 1973
Mar. - Oct., 1974
May 20-26, 1975
June 4-9, 1973
Oct. 20 - 31, 1973
< 0.02 - 3.3, 1.0 (11)
0.10 - 1.9, 0.57 (25)
1.7 - 5.2, 2.8 (6)
0.70
1.2
1.1
0.52
1.6
0.87
0.57
0.39
0.77
0.77
0.65
/Source: Bidleman and Olney, 1975.
Midpoint of the collection track.
/The mean blank corresponded to 0.03 ng/cu m for a 1,000 cu m sample.
Range, mean and (number of samples).
-------
Table 7.4 (continued)
Location
Dates
Toxaphene (ng/cu m)
I
I—1
o
Cruise TR-152
31°20'N, 64°50'W
31°30'N, 65°20'W
32°00'N, 65°30'W
40°40'N, 66°30'W
41°30'N, 68°30'W
42°20'N, 66°40'W
Cruise TR-153
33°40'N, 57°00'W
34°30'N, 58°40'W
40°30'NJ 70°00'W
May 9-19, 1974
May 25 - 31, 1974
0.16
< 0.1
0.39
< 0.06
< 0.04
0.05
0.11
0.58
0.04
.Source: Bidleman and Olney, 1975.
Midpoint of the collection track.
/The mean blank corresponded to 0.03 ng/cu m for a 1,000 cu m sample.
Range, mean and (number of samples).
-------
Table 7.5. Toxaphene residues in air samples at five North American sites
Location and date
Kingston, RI , 1975
Sapelo Island, GA, 1975
Organ Pipe Cactus National Park,
AZ, 1974
Hays, KS, 1974
Number
of
samples
6
6
6
3
Range
(ng/cu m)
0.04 - 0.4
1.7 - 5.2
2.7 - 7.0
0.083 - 2.6
Northwest Territories,
Canada, 1974 3 0.04 - 0.13
Source: Adapted from Bidleman et al., 1976.
7-11
-------
Lichtenberg et al., 1970; Manigold and Schulze, 1969; Schulze et al., 1973;
Schafer et al., 1969; Mattraw, 1975), 1,2,3,4,5,7,7-heptachloronorbornene, which
is claimed as a possible toxaphene component (Sanborn et al., 1976), has been found
in New Orleans drinking water at 0.06 ppb (U.S. EPA, 1974). Since the compound is
neither a terpene nor a likely metabolite, the claim that toxaphene is its source
is in serious doubt. The routine monitoring studies have screened for residues
by procedures which will detect the simple organochloride residues (e.g., DDT and
its residues, dieldrin, heptachlor epoxide) at significantly lower concentrations
than toxaphene (Section 2.6.1). Lichtenberg (1971) and Schulze et al. (1973) place
the toxaphene lower detection limit at 0.5 to 1.0 ppb, whereas other organochlor-
ides can be detected at concentrations about two orders of magnitude lower.
While Lichtenberg (1971) points out that a lower toxaphene detection limit
could be achieved if toxaphene were specifically sought, the routine monitor-
ing studies are either vague about toxaphene's lower detection limit or con-
firm that it is significantly higher than the other organochloride residues.
Toxaphene has been detected and monitored in waters and sediments where
its origin was known. These include surface and ground waters in agricultural
areas where toxaphene was applied to crops; lakes where toxaphene was added
for rough fish control; and surface waters surrounding a toxaphene manufac-
turing plant at Brunswick, Georgia.
The Flint Creek, Alabama watershed was monitored during the years 1959
to 1965 (Nicholson et al., 1964; Nicholson et al., 1966; Nicholson, 1969;
Grzenda and Nicholson, 1965; Grzenda et al., 1964). Although the watershed
drains a diversified agricultural area, the study identified small cotton
farms as the major pesticide users (Nicholson et al., 1964).
7-12
-------
Natural water from Flint Creek and potable water from the Hartselle
Water Treatment Plant (which is drawn from Flint Creek) were sampled in pairs
by carbon-chloroform extraction (Section 2.6.2). Table 7.6 compiles data on
toxaphene application in the watershed drainage area and the residues in the
raw and finished waters. Actual residue concentrations in the waters are
probably higher than reported by a factor of two,, since the average toxaphene
recovery from spiked samples was less than 50 percent (Section 2.6.6). Toxa-
phene concentration remained below 1 ppb in finished and raw waters through-
out the study period. Water treatment by the Hartselle plant apparently did
not significantly reduce the toxaphene concentration (Cohen et al., 1961).
Toxaphene residues varied seasonally. The maximum level, which was ob-
served in summer, corresponded to the time that maximum application was ex-
pected. After toxaphene usage in the area had decreased by approximately an
order of magnitude (in 1963 to 1964), the residues in water rapidly dropped.
Nicholson et al. (1964) could not explain why toxaphene residues increased
during 1960 to 1961, although its application decreased by approximately
one-third from 1959 to 1960.
The Flint Creek study also measured residues in biota (Section 4.0 and
5.0), soil (Section 7.2.3), and sediment. Grzenda and Nicholson (1965) re-
ported that no toxaphene was detected in 58 samples of mud from eight sites
in Flint Creek. Sediment samples were collected between June and December
1963. Table 7.7 summarizes the number of positive detections of toxaphene,
DDT, and BHC residues in suspended sediments of raw water, in filtered, raw
water, and in treatment plant sediments at the Hartselle Water Treatment
Plant. Since they detected toxaphene less often on suspended solids than DDT,
7-13
-------
Table 7.6. Toxaphene residues recovered from Flint Creek, Alabama and
the Hartselle Water Treatment Planta
year
1959-60
1960-61
1961-62
1962-63
1963-64
1964-65
1965
Te
to
[Hi
kilo£,ra
63.
42.
72.
60.
S.
9.
9.
3
5
4
7
4
0
5
Toxaphene
clinical Summer
usage Untreated water
ms (pounds) ] Mean Range
(56.5) 0.105 0.010-0.260
(37.9) 0.140 0.010-0.270
(64.6) 0.145 0.110-0.164
(72.0) 0.054 0.025-0.125
(7.5)
(b.O)
(8.5)
Treated
water
Mean
0.
0.
0.
0.
0.
0.
0.
105
210
074
095
16|sic]
05
01
0.
0.
Range
040-0.280
,010-0.410
0.037-0.111
0.
0.
0.
0.
045-0.154
04-0.08
,00-0.11
,00-0.08
concentration (ppb)
Fall
Untreated Treated
water water
Mean Mean
0.029 0.079
0.067 0.063
0.040 0.030
0.049 0.066
0.03
0.01
0.00
Winter
Untreated Treated
water water
Mean Mean
0.030 0.
0.027 0,
0.051 0.
0.050 0,
0.
0,
046
,018
,052
,043
,001
.00
Spring
Untreated Treated
water water
Mean Mean
0.049 0.052
No sample
No sample
0.01
0.01
0.00
Source: Adapted from Nicholson et al., 1964, 1966.
-------
Table 7.7. Comparison of insecticide recovery from sediment and water,
Hartselle, Alabama Water Treatment Planta
Sample source
No. sample
Percent positive for
DDT
DDE Toxaphene BHC
I
I—'
Ln
Sediment from treatment
plant settling basis
Suspended sediment extracted
from raw water by filtra-
tion prior to carbon
filtration'3
Carbon adsorption samples
collected from water after
removal by above filtration
45
77
77
71
69
13
64
62
12
18
10
31
22
17
74
a
/Source; Nicholson et al., 1966.
A Cuno Micro-Klean filter that removed sedimentary particles larger than 25 microns was used,
Smaller particles pass through to the carbon adsorption units.
-------
they concluded that aquatic toxaphene residues are primarily dissolved rather
than adsorbed onto particulates.
Bailey and Hannum (1967) monitored toxaphene in California surface waters
and their sediments. Table 7.8 summarizes their measurements. Toxaphene con-
centrations were higher in sediments than in the water. They also noted that
the pesticide concentrations generally were greater on the smaller size parti-
cles.
Johnston et al. (1967) discovered that toxaphene and other insecticide
residues had contaminated an aquifer in the San Joaquin Valley, California.
The contamination was detected in an experimental plot in which the movement
of other insecticides was being examined. A plot, to which pesticides had
not previously been applied, was subdivided into three blocks. The blocks
were flooded and subsequent drainage from each block was analyzed. Although
toxaphene had never been applied, it was detected in the drainage (Table 7.9).
They concluded that its source was contaminated ground water. Johnston et al.
(1967) also detected toxaphene residues in 13 of 66 analyses of other San
Joaquin Valley tile drainage effluents (average 0.528 ppb and range 0.130 to
0.950 ppb) and in 60 of 61 analyses of surface effluent in Panoche Drain Water
(average 2.009 ppb and range 0.100 to 7.900 ppb).
In their annual reports, the San Joaquin District, California Department
of Water Resources (1968 to 1969), which monitored the area's water, detected
toxaphene in 51 of 422 (12 percent) tile drainage effluents (0.02 to 0.50 ppb),
in 216 of 447 (48 percent) of surface drains in Central Valley (0.04 to 71.00 ppb)
in 88 of 712 (12 percent) of Central Valley surface waters (0.02 to 0.93 ppb),
and in 8 of 200 (4 percent) of California bays and surface waters.
7-16
-------
Table 7.8. Toxaphene concentrations in California surface waters and their sediments
a
Sampling station
Streams and bays
Concentration (micrograms per liter)
Surface Water
Sediment
Maximum Minimum Average Maximum Minimum Average
I
M
^J
Feather River at Nicolaus Bridge
American River at Sacramento
Sacramento River at Walnut Grove
Mokelumne River at Highway 99
Little Connection Slough at
Atherton Road
Middle River at Victoria Canal
Delta Mendota Canal at Head
San Joaquin River at Antioch
Suisun Bay at Martinez
Napa River at Duttens Landing
San Pablo Bay at Point San Pablo
San Francisco Bay at Berkley Pier
San Francisco Bay at Treasure Island
San Francisco Bay at San Mateo Bridge
0.4
0.03 0.10
0.04
0.160
0.120
0.320
0.090
0.03
0.050
0.050
0.08
0.145
0.063
0.08
0.23 0.03 0.13
0.26
130
57
170
140
110
110
Source: Bailey and Hannum, 1967.
-------
Table 7.8 (continued)
I
M
00
Concentration (micrograms per liter)
Sampling station
Streams and bays
Golden Gate Bridge at Fort Point
San Joaquin River at Vernailes
San Joaquin River at Fremont Ford
Salton Sea near North Shore
Alamo River
All American Canal at Alamo River
Surface Water Sediment
Maximum
0.93
0.46
0.40
0.65
0.08
Minimum
0.02
0.04
0.05
0.30
0.04
Average Maximum Minimum Average
0.26
0.13
0.14
0.47
0.06
Agricultural Drainage
Reclamation District No. 108 Drain
Colusa Basin Drain
Staten Island Drain
Roberts Island Drain at Whiskey Slough
Panoche Drain
Salt Slough
0.230
5.50
0.440
0.100
0.040
1.467
0.169
210
110
380
-------
Table 7.9. Toxaphene residues in tile effluent drainage from a plot not
previously treated with toxaphene^
Toxaphene concentration (ppt)
Block A Block B Block C
First flooding
Second flooding
Third flooding
50
175
550
500
50
0
100
0
0
Source: Johnston et al., 1967-
7-19
-------
Barthel et al. (1969) examined organochloride residues in sediments of
the lower Mississippi River and its tributaries during 1966. Toxaphene was
detected at few of the sampling sites; Table 7.10 summarizes sites and con-
centrations .
The University of Georgia Marine Institute (Reimold, 1974; Reimold and
Durant, 1972 a, b, 1974; Durant and Reimold, 1972) has monitored toxaphene
contamination in surface waters, sediment, and biota of waters receiving the
effluent of the Hercules, Inc. plant located on Terry Creek, Brunswick,
Georgia. This plant is the largest producer of toxaphene in the U.S.
Figure 7.2 describes the average monthly toxaphene concentration in the plant's
effluent from 1970 to 1974. It has decreased from a high of 2332 ppb in
August 1970 to a low of 6.4 ppb in June 1974. Dye experiments have shown
that the effluent is diluted by a factor of 10 after it reaches Terry Creek
(Reimold, 1974). The Institute (Reimold and Durant, 1972 a, b; Durant and
Reimold, 1972) analyzed sediment at three locations downstream of the plant
outfall. Samples were collected prior to a dredging operation in June 1971.
at sites downstream: 0.2 miles from the outfall at a location 50 yards from
its intersection with another creek; 0.8 miles from the plant outfall; and
1.4 miles from the plant outfall and 50 yards from the end of Terry Creek
(junction with Back River). Table 7.11 summarizes toxaphene concentration
in sediment cores to 80 cm deep. Reimold and Durant (1972 b) measured
32.56 ppm as the average toxaphene concentration within Terry Creek Marsh.
The highest residue concentration measured in the surrounding water was
0.015 ppm.
Mattraw (1975) screened for organochloride residues in soil, water, and
its sediment in southern Florida from 1968 to 1972. The area includes undeveloped
7-20
-------
Table 7.10.
Toxaphene residues in sediments of the lower Mississippi River
and its tributaries in 1966a
River
Site
Concentration of^
toxaphene/Strobane in
oven-dried sample
(ppm)
Mississippi River
Sunflower River
West Memphis, Arkansas
River mile 727.7
Site UL-2
Site UL-7
Coahoma County, Mississippi
Site M
Site M
Site D
Site D
0.4
0.1
11.19
11.93
8.85
10.87
Sunflower River
Sunflower County,
Mississippi
Site U
Site D
2.45
6.57
Ditch 24,
Sunflower River
Jones Bayou
Sunflower County,
Mississippi
Bolivar County, Mississippi
Site U
Site D
13.18
4.52
3.91
a.
Source: Barthel et al. , 1969.
Strobane is a mixture of chlorinated diterpenes which is produced by
chlorinating a-pinene. It closely resembles toxaphene (see Section 2.4.2).
7-21
-------
I
K>
T
0
X
A
P
H
E
N
E
P
P
B
I.OOO—-
100—=
10—
ASONDJFMAMJJASONOjFMAMJJASONOjFMAMJJASONDJFMAMJ
1970
1971
1972
1973
1974
Figure 7.2. Monthly average of apparent toxaphene concentration of manufacturing plant effluent,
1970 - 1974. Source: Reimold, 1974.
-------
Table 7.11. Toxaphene concentrations in three sediment cores, by 10 cm increments, collected from
Terry Creek, Brunswick, Georgia, June 10, 1971a
Residues (ppm) and content of sediment
I
N3
U)
Sampling
location
Site C
Site 11 C
d
Site E
a.
^Source
, Horth
/South
Surf, to
10 cm
1,858.3
mud and
some chips
111.85
mud and
few chips
35.5
mud
: Durant and Reimold,
shore of Terry Creek,
shore of Terry Creek,
10 - 20 cm 20
1,340.5
mud and
few chips
615.64
mud and
many chips
35.47
mud
1972 b.
50 yd from Junction with Dupree
.8 mile from the plant outfall
- 30 cm
1,324.0
mud
16.04
mud
21.9
mud
Creek and .2
and one-half
30 - 40
1,367.2
mud
17.46
mud
70.65
mud
mile from
cm 40 - 50 cm 50 - 60 cm 60 - 70 cm 70 - 80 cm
1,236.7 433.6 68.5 83.2
mud mud mud mud
5.42 3.4 2.88 No sample
mud mud mud
79.8 21.0 18.5 5.27
mud mud mud mud
toxaphene plant outfall.
the distance from the outfall to Back River.
QQh f\f H^f\l V ! l.at-
-------
lands, agricultural lands, and coastal urban areas. Toxaphene residues were
not detected in 146 samples of surface water. Although Mattraw (1975) defines
the limit of detection for water samples at 0.005 ppb for all organochlorides,
the limit is expected to be higher for toxaphene (Sections 2.6.1 and 7.2.3).
Toxaphene was detected (claimed lower detection limit of 0.05 ppb) in 3.2 percent
of sediment samples and 4.2 percent of cropland soil.
Keith (1966) examined toxaphene and other chlorinated hydrocarbon insec-
ticides in the sediments of Tule Lake and Lower Klamath Lake National Wildlife
Refuge in northeastern California. Both lakes are impoundments which are used
for agricultural purposes. While the lakes were not directly treated with
toxaphene, their waters were used for irrigation and subsequent agricultural
drainage was returned. Keith (1966) estimated that waters of the refuge were
used from five to seven times for irrigation. Sediment toxaphene concentra-
tions in these lakes ranged from 0.0 to 0.2 ppm for the years 1960 and 1961.
Terriere et al. (1966) monitored residues in two Oregon lakes to which
toxaphene was applied as a piscicide. Table 7.12 describes the residue con-
centrations in the water and bottom sediment. Miller Lake is deep and bio-
logically sparse, while Davis Lake is shallow and rich in aquatic life.
Kallman et al. (1962) applied 0.05 ppm of toxaphene to Clayton Lake,
New Mexico in three stages. It was added as 0.7 kg per liter (emulsifi-
able concentrate) for rough fish control. Figure 7.3 describes the additions
and subsequent monitoring of aqueous toxaphene at the 3.05 meter level. The
solid columns denote amounts in water from the windward shore and the barred
columns from the lee shore. Vertical arrows and their numbers denote the
times at which applications were made and the amounts applied. They found
7-24
-------
Table 7.12. Toxaphene residues in Miller Lake and Davis Lake, Oregon
(Year) 1958
Toxaphene concentration (ppb)
Miller Lake
Davis Lake
1962
1963
1964
1961
1962
1963
1964
Toxaphene
application rate
40
88
i
NJ
Ln
Aqueous residue
2.10
(0.7-3.1)
1.2
(0.7-1.6)
0.8
(0.7-1.1)
0.63
(0.5-0.9)
0.41
(0.3-0.6)
< 0.2
Bottom mud
750 391
(390-1430) (330-13800)
650
(100-1000)
800
(100-3100)
Terriere et al., 1966.
-------
.03 —
-
E
Q.
.5 .02 —
UJ
Z
UJ
0.
X
0
o
I-
I .01-
.02ppr
1
.02 pom
1
5 .01 ppm
^
>
f
n
. 1
n
/ i
'
t
71
' 0
n
ND- NOT DETECTED
r
1
il ii n
v — »i — • \ — »i — m\ — -,-> — H — n — K — n — _ —
4 5 6 14
DAYS AFTER FIRST APPLICATION
20
31
38
523
Figure 7.3. Amount of toxaphene measured in Clayton Lake (New Mexico) water from the 3.05 metcl: ievel
before and after applications of toxaphene. Source: Kallman et al., 1962.
-------
a maximum toxaphene concentration in sediment of 0.15 ppm, which was measured
six days after the first addition.
Johnson et al. (1966) examined toxaphene residues in Wisconsin lakes to
which toxaphene was applied for fish control. Table 7.13 describes the toxa-
phene application and residues found in 1965.
7.2.3 Residues in Soil
Stevens et al. (1970) examined insecticide application and soil residues
at 51 sites from 1965 to 1971. They found that 17 sites regularly applied
pesticide, 16 sites applied pesticide at least once but that application was
not regular, and 18 sites never had applied a pesticide. No toxaphene resi-
dues were detected at the sites of limited or no pesticide use. Table 7.14
summarizes the known application rates and residue levels. Although only
5 sites reported that toxaphene had been applied, toxaphene residues were
detected at 7 sites. The 2 sites with residues but no history of application
were Tule Lake, California, and Wenatchee, Washington. The regular pesticide
use croplands were classified into three groups: vegetable and/or cotton;
tree fruit; and small grain and root crops. Toxaphene was detected at
60 percent of the fields in the vegetable and/or cotton croplands (range
0.66 to 9.38 mg/kg), at one orchard (7.72 mg/kg), and at 12 percent of the
small grain and root croplands (range 0.11 to 2.01 mg/kg).
EPA's National Soil Monitoring Program (NSMP) has monitored soils of
croplands, non-croplands, and urban areas since 1967. Wiersma et al.
(1972 b) examined cropland in 43 states and non-cropland in 11 states during
fiscal year 1969. They detected toxaphene in only 1 of 199 non-cropland
soils (concentration 0.52 ppm). Toxaphene was detected in 4.2 percent (73
7-27
-------
Table 7.13. Toxaphene In Wisconsin lakes, 1965
Concentration (ppb)
1
N>
00
Lake
Little Green
Emily
Kusel
Marl
Big Twin
Wilson
Round
Corns tock
(surface)
6.5 meters
Year of
treatment
1956
1959
1960
1960
1963
1964
Hay 1965
1964 £. 1965
June, 1965C
Trea tmen t
rate
100
100
100
100
100 & 50
2.5 - 3.5
5 Epilimnlon only
5 + 5
100
Water
1
4
3
3
2
4
2
20
4
Suspended
matter
40
20
200
9
20
80
200
100
500
Aquatic
plants
400
70
80
40
50
80
50
-~
Sediment
20
200
400
1000
800
500
600
1000
.Source; Johnson et al., 1966,
Remaining in water after filtration through Whatman GF/A glass filters.
Comstock Lake was treated 14 days prior to sampling.
-------
Table 7.14. Toxaphene monitoring in U.S. croplands: application rates and
soil residues'"1
Location
Toxaphene
applied
kg/ha (Ibs/acre) Year
Toxaphene residue
mg/kg
Year
Kern County, California
Lower Rio Grande Valley,
California
Field
Field
Field
Field
Field
Monmouth
1
2
3
4
5
County, New Jersey
18.
J.
2.
53.
7.
1.
38.
10.
1.
33.
44.
3
4
3
0
9
4
4
2
1
0
2
(16.2)
(3.0)
(2.0)
(47.0)
(7.0)
(1.25)
5
(34.0)
(9.0)
(1.0)
(29.25)
(39.16)
1956-1964
1965
1966
1958-1964
1965
1966
1958-1964
1965
1966
1955-1964
1956-1964
2.
1.
1.
2.
2.
~
90
98
77
01
43
-
Oct.
Oct.
Oct.
Oct.
Oct.
1966
1966
1966
1966
1966
Dade County, Florida
Field
Field
Field
Field
Field
Eastern
Field
Field
Field
Field
Field
1
2
3
4
5
South Carolina
1
Z
3
4
5
4.
9.
4.
2.
4.
2.
10.
35.
22.
3.
16.
J.
53.
10.
20.
42.
5
0
5
3
5
5
2
6
4
4
9
4
0
2
3
9
(4.0)
(8.0)
(4.0)
(2.0)
(4.0)
(2.2)
(9.0)
(31.59)
(19.90)
(3.0)
(15.0)
(3.0)
(47. n)
(9.0)
(18.0)
(38.0)
1965
1958-1964
1955
1966
1965
1962-1964
1965
1962-1964
1965
1952-1964
1956-1964
1966
1957-1964
1965
1966
1956-1964
1.
2.
0.
4.
4.
9.
7.
2.
1.
5.
0.
2.
21
64
66
42
14
38
00
99
05
64
99
04
Mar.
Mar.
Mar.
tlov.
Mar.
Nov.
Mar.
Aug.
Aug.
Aug.
Aug.
Aug.
1968
1968
1968
1966
1968
1966
1968
1966
1966
1966
1966
1966
faStevens et al., 1970.
1.9 Ug/ha (1.70 Ib/acre) Strobane was also applied (see Table 7.10, footnote fa).
7-29
-------
Table 7.14. (continued)
Location
Toxaphene applied
kg/ha (Ib/acre)
Year
Toxaphene residue
mg/kg
Year
Wenatchee, Washington
Orchards 1, 2, 3, 4
Orchard 5
Adams County, Pennsylvania
Bererien County, Michigan
Western North Carolina
Yuma, Arizona
Central, Georgia
Quincy and Moses Lake,
Washington
Fields 1 and 5
Field 2
Field 3
Field 4
Tulelake, California
Field 1
Fields 2 5
Weld County, Colorado
Fields 1, 2, 4
Field 3
Field 5
Urbana, Illinois
Western Iowa
Eastern Virginia
1959-1967
7.72
3.4 (3.0)
1.7 (1.5)
30.3 (27.0)
1963-1964
1965
1956-1964
0.95
0.58
Unknown
1966
0.78
2.01
0.11
Oct. 1967
Oct. 1967
Sept. 1967
Oct. 1966
Sept. 1967
Sept. 1967
^Stevens et al., 1970.
1.9 ke/ha (1.70 Ib/acre) Strobane was also applied (see Table 7.10, footnote b).
7-30
-------
of 1729) of cropland sites with a concentration of 0.10 to 11.72 ppm. The
mean toxaphene concentration for all cropland sites was 0.07 ppm. Wiersma et al.
(1971) sampled 242 cropland and 117 non-cropland soils in six states in 1967-
Toxaphene residues were not detected in non-cropland soil. Results for the
cropland soils are presented in Table 7.15. Residues were only found in two
states; Georgia (10 sites) and Idaho (1 site).
Crockett et al. (1974) published data for toxaphene application and soil
residues at 1506 cropland sites in 35 states, which were monitored during
fiscal year 1970. Toxaphene was reportedly used at 33 (2.45 percent of sites)
sites and applied at an arithmetic mean rate of 0.023 kg/ha. Table 7.16
summarizes the results by state. Toxaphene was detected at 27 sites: 13 in
California; 5 in Mississippi; 3 in Georgia; 2 in Louisiana and South Carolina;
and one in Florida, Oklahoma, and Kentucky. Toxaphene residues at detected
sites ranged from 0.79 to 8.75 ppm. Residues are listed by crop in Table 7.17.
Toxaphene was most often detected at sites classified as cotton or irrigated
lands and the residues found at those sites had the highest concentration:
arithmetic mean of 0.68 ppm for irrigated land and 0.32 ppm for cotton farming.
Wiersma et al. (1972 a) measured toxaphene and other pesticide residues
in soil from eight cities in 1969. Table 7.18 summarizes the toxaphene data.
In each city, fifty random sites were sampled. Toxaphene residue was detec-
ted at only four sites: two sites in Miami and one site each in Houston and
Manhattan (Kansas). While toxaphene residues never exceeded 4 percent of the
sites in any city, DDT residues were detected at 40 percent (Houston) to
100 percent (Miami) of sampled urban sites.
7-31
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Table 7.15. Cropland soil monitoring for toxaphene in six states, 1967
a.
*
State
Georgia
Idaho
Maine
Nebraska
Virginia
Washington
Overall
Sites
sampled
30
33
8
107
19
45
242
Sample sites
treated with
toxaphene
(percent)
13.3
5.3
Sites with
toxaphene Toxaphene
residues residues
(percent) (ppm)
33.3 0.55
3.1 0.44
0
0
0
0
4.6 0.13 ,
a
'Source: Wiersma et al. , 1971.
7-32
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Table 7.16. Toxaphene in cropland soil by state, fiscal year 1970'
a
Toxaphene concentration
(ppm)
Times
State detected
All sites
Alabama
Arkansas
California
Florida
Georgia
Illinois
Indiana
Iowa
Kentucky
Louisiana
Michigan ,
Mid-Atlantic states
Minnesota
Mississippi
Missouri
Nebraska
New England States
New York
North Carolina
Ohio
Oklahoma
Pennsylvania
South Carolina
South Dakota
Tennessee
Virginia, West Virginia
Wisconsin
27
C
C
13
1
3
c
c
c
1
1
c
c
c
5
c
c.
c
c
c
c
1
c
2
c
c.
c
c
Sites
sampled
1506
21
47
65
17
28
140
78
150
30
26
54
19
120
29
81
106
20
38
30
69
65
32
17
106
25
26
67
Range of
Arithmetic detected
mean residues
0.06 0.79-8.75
0.61 0.79-7.63
0.34 5.71
0.17 1.21-2.27
0.03 0.89
0.20 5.32
0.73 2.79-8.75
0.02 1.60
0.34 2.44-3.34
/Source: Adapted from Crockett et al., 1974.
For all sites samples.
/None reported.
Mid-Atlantic states include Maryland, Delaware, and New Jersey.
eNew England states include Maine, New Hampshire, Massachusetts,
Rhode Island, and Connecticut.
7-33
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Table 7.17. Toxaphene residue in cropland soil by cropping region,
fiscal year 1970a
Crop
Corn
Cotton
Cotton and general farming
General farming
Hay and general farming
Irrigated land
Small grains
Vegetables
Vegetables and fruit
Times
detected
(percent)
N.D.C
8.91
3.42
3.40
N.D.
20.51
N.D.
N.D.
2.38
Sites
analyzed
713
101
117
147
184
39
105
72
42
Arithmetic
mean ,
concentration
(ppm)
N.D.
0.32
0.09
0.07
N.D.
0.68
N.D.
N.D.
0.14
.Source: Adapted from Crockett et. al. , 1974.
For all sites analyzed.
Not detected.
7-34
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Table 7.18. Toxaphene residues in soil from eight cities, 1969
City
Bakersfield, CA
Camden , N J
Houston, TX
Manhattan, KS
i
£ Miami, FL
Milwaukee , WI
Salt Lake City, UT
Waterbury, CT
c.
, Percent
Range Average positive sites
N.D.d
N.D.
O.lie <0.01 2.0
12.07e 0.24 2.0
14.79 - 52.73 1.34 4.0
N.D.
N.D.
N.D.
a
Source: Wiersma et al., 1972 a.
uBased on total number of samples (49).
^Percent based on number of sites with residues greater than or equal to the sensitivity limits.
Not detected.
One value.
-------
The National Soil Monitoring Program examined cropland soils of the corn
belt states (Carey et al., 1973), 10 onion growing states (Wiersma et al.,
1972 c), and 9 sweet potato farming states (Sand et al., 1972). No toxaphene
residues were detected from sweet potato or corn growing soils, but residues
were found in 12.7 percent of the 76 onion growing sites. While toxaphene
residues averaged 0.55 ppm at the onion growing sites, at positive detection
sites they ranged from 2.19 to 7.77 ppm.
Grzenda and Nicholson (1965) examined toxaphene residues in soil samples
from fields surrounding Flint Creek, Alabama. Their work was part of an in-
tensive study of toxaphene, DDT, and BHC use and residues (Section 7.2..2).
They detected toxaphene in 58 percent of the soils collected from 33 cotton
fields. Mean concentration (33 fields) was 0.41 ng/kg. Toxaphene concentra-
tions (positive sites) ranged from 0.16 to 1.60 ng/kg.
Mullins and his coworkers (1971) examined pesticide applications and
residues in major agricultural areas of Colorado. Of the 50 sites evaluated,
9 reported toxaphene applications with an average amount applied of 3.14 ppm.
Toxaphene (at 1.00 ppm) was detected at only one site.
7.3 ENVIRONMENTAL FATE
7.3.1 Mobility and Persistence in Air
The relatively high vapor pressure of toxaphene (0.17 to 0.4 mm Hg) com-
pared to other chlorinated hydrocarbon pesticides (0.000001 to 0.000000001 mm Hg)
suggests that atmospheric processes will have considerable impact upon the
environmental fate of toxaphene. Toxaphene has been detected in air in con-
centrations as high as 2520 ng/cu m, the concentration varying depending upon
location (near or remote from crop spraying) and time of year (high during
summer months when pesticides are being used most (Section 7.2.1).
7-36
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The several monitoring studies that have detected toxaphene in air all
illustrate the ability of toxaphene to be transported by atmospheric currents.
This is best demonstrated by the work of Bidleman and coworkers (Bidleman and
Olney, 1975; Bidleman et al., 1976) who detected toxaphene 1200 km out to sea.
When the trajectories of the samples were examined, it was found that samples
with North American air sources differed by no more than a factor of two from
the samples whose trajectories did not pass over the continent. The authors
indicated that these results suggest that toxaphene has a sufficiently long
lifetime in the atmosphere to become well-mixed in the North Atlantic atmos-
phere. However, the higher concentrations detected when the air passed over
North America suggests that a likely source of airborne toxaphene is from its
use in the southern U.S. cotton growing areas. The mean levels of toxaphene
detected in the North Atlantic Ocean air are equal to or twice those of PCB's
and ten times higher than those of other pesticides, such as DDT.
The form in which toxaphene is present in the atmosphere (particulate or
gaseous) is unknown. Bidleman and Olney (1975) found that less than 5 percent
of the total toxaphene was found on glass fiber filters (collects 98 percent
of particles >_ 0.015 micrometer) which suggests that toxaphene may be mostly
gaseous. However, their air flow rates (0.48 to 0.76 cu m/min) in the sampler
might allow evaporation from the particulate matter.
The stability of toxaphene in the air has not been studied. For example,
no vapor phase photolysis studies are available. However, comparison of gas
chromatographs of standard toxaphene and an air sample obtained in Bermuda
(Bidleman and Olney, 1975) show some slight changes in peak heights and re-
tention times. These differences might be explained by selective volatiliza-
tion of the more than 175 compounds included in toxaphene. Seiber et al. (1975)
7-37
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found that a toxaphene air sample gathered above a cotton field and analyzed
by gas chromatography showed a substantial enrichment of the more volatile com-
ponents. Enrichment of the less volatile components was found on leaf surfaces.
7.3.2 Mobility and Persistence in Water
Although toxaphene is a widely used insecticide, few laboratory studies
have been conducted on its environmental fate in aqueous systems. Most in-
vestigations relevant to the environmental behavior of toxaphene in water
have been field studies, mostly monitoring studies of water systems near cotton
fields or lakes that have been treated with toxaphene for rough fish control.
Toxaphene has not been detected in routine ambient water monitoring in-
vestigations, which might suggest a lack of persistence of toxaphene in water
systems. An alternative explanation is that the analytical methods used for
screening chlorinated hydrocarbon pesticides are not as sensitive for toxaphene
as the methods are for less chemically complex compounds.
Analysis of water samples which contain toxaphene (Section 7.2.2) provides
some insight into the mode of transport. Nicholson et al. (1966) detected
toxaphene less often than DDT in suspended solids, even though the percentage
of samples containing DDT in the water by carbon chloroform extraction was
lower than the percentage containing toxaphene. These authors concluded that
aquatic toxaphene residues are primarily dissolved rather than adsorbed onto
particulates and are not removed by conventional water treatment plants
(Cohen et al. , 1961). This suggestion is supported by the work of Grzen-da and
Nicholson (1965) who found amounts of toxaphene in river water varying from
30 to 140 ppt in Flint Creek, Alabama, but no toxaphene in river bottom sediment
7-38
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On the other hand, Bailey and Hannum (1967) found higher concentrations of
toxaphene in sediment than in water and the concentration was greater on
smaller sized particles.
Toxaphene residues in water may be removed by a variety of mechanisms
including biodegradation, hydrolysis, photodecomposition, evaporation, or
codistillation, and adsorption to sediment or suspended particulates (followed
by settling to the sediment). Studies conducted on lakes where toxaphene was
used for rough fish control provide considerable information on the importance of
the above mechanisms (Johnson et al., 1966; Kallman et al., 1962; Terriere et al.,
1966; Veith and Lee, 1971; and Hughes, 1970). Toxaphene was used for rough
fish control (piscicide) because it has the greatest fish toxicity of any
chlorinated hydrocarbon, except endrin, and has a lower acute human toxicity
than endrin, dieldrin, aldrin, or rotenone (Johnson et al., 1966). In addition,
toxaphene's lower cost facilitated its use as a piscicide.
Prior to the use of modern analytical techniques, a number of studies
which were based upon bioassays (fish kills) determined that a number of fac-
tors affected the rate of detoxification of toxaphene treated lakes. A variety
of parameters have been suggested including sunlight, pH, temperature, dis-
solved oxygen concentration, alkalinity, hardness, concentration applied,
turbidity, and presence of bacteria (Johnson et al., 1966; Veith and Lee, 1971).
Studies using gas chromatographic methods could find little correlation be-
tween these factors. Johnson et al. (1966) studied eight Wisconsin lakes,
almost all of which were classified as hard water, alkaline, shallow, eutrophic
lakes with large amounts of plankton and higher plants in near shore areas.
The lakes were treated with 0.1 ppm of toxaphene 3 to 9 years prior to sampling
7-39
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and contained 1 to 4 micrograms/liter of toxaphene in water, 0.2 to 1 ppm in
sediments, and 0.05 to 0.4 ppm in plants. Although these residues were de-
tected, the authors concluded that shallow eutrophic lakes detoxify rapidly.
This finding is consistent with the results of Terriere et al. (1966) who
studied two toxaphene treated Oregon lakes, one a deep and biologically sparse
lake and the other a shallow and biologically rich lake. With the shallow
lake, detoxification occurred in one year (concentration in water 0.6 micro-
gram/liter), similar to the rapid detoxification observed by Johnson et al.,
(1966). However, with the deep lake the water concentration remained at
1.2 micrograms/liter after five years.
Veith and Lee (1971) concluded that toxaphene adsorption to sediment
is a major detoxification mechanism which is consistent with the above con-
clusion that treated lakes which are stratified (deep lakes) remain toxic to
fish longer than well-mixed, shallow lakes. They analyzed three lakes in
Wisconsin that had been treated with toxaphene and also conducted a desorp-
tion study with toxaphene on sediment. The analysis of lake samples by gas
chromatography with electron capture detection demonstrated that the con-
centrations in water rapidly decreased while the concentrations in sediment
increased to a maximum and then decreased by a factor of 2 every 20 days.
The authors noted that this rate of loss from sediment greatly differs from
the 11 year half-life in soil reported by Nash and Woolson (1967). Analysis
of various sediment sections (0 to 5 cm, 5 to 10 cm, 10 to 15 cm) showed that
the concentration of toxaphene was much higher in the top layer. By measuring
the time between treatment and the first detection of toxaphene in sediment,
Veigh and Lee (1971) calculated the rate of vertical transport. For the
three lakes, the values varied from 0.4 to 1.1 cm/day.
7-40
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The desorption study of Veith and Lee (1971) consisted of placing various
amounts of Ottman Lake sediments which contained 2.0 micrograms of toxaphene
per gram (dry weight) in flasks containing filtered Ottman Lake water. The
flasks were maintained at 23 to 25 C for 10 days with mixing four times daily.
At 10 days the samples were filtered (glass fiber) and both water and sediment
were analyzed. The toxaphene concentration remained below the detectable limit
(1 microgram/liter) in water and was essentially the same as the starting con-
centration in sediment, thus suggesting strong adsorption between toxaphene
and the sediment.
For each of the three lakes, Veith and Lee (1971) calculated the per-
centage of the applied toxaphene that was in the sediments at the time of
maximum concentration. These percentages were 47.3 percent for Fox Lake at
51 days after treatment; 46.3 percent for Ottman Lake at 191 days; and
13.5 percent for Silver Lake at 275 days. Taking into account the low con-
centration in water (<1 microgram/liter), the authors concluded that "greater
than 50 percent of the toxaphene applied may have been lost during applica-
tion, lost through evaporation and/or codistillation with water vapor, and
degraded by microbial activity." As is the case now and was noted then by
the authors, the significance of these additional processes has not been
evaluated. However, some information on these processes is available.
Monitoring data of the air above the surface of toxaphene treated lakes
or laboratory studies of the rate of evaporation of toxaphene from water are
not available. However, Mackay and Leinonen (1975) have modified some work
of Liss and Slater (1974) in order to provide a method for calculating the
half-life of evaporation of low-solubility compounds from water bodies to
7-41
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the atmosphere using the vapor pressure, water solubility, and molecular weight
of a compound. Using the available data for toxaphene (water solubility =
.037 mg/liter; vapor pressure = 0.17 mm Hg; average molecular weight = 414),
the calculated half-life for toxaphene evaporation from water is 10.4 hours.
The rate of evaporation for each component of the toxaphene mixture will be
different, and it is likely that changes in the gas chromatographic finger-
print could be attributed to selective evaporation (Seiber et al., 1975).
Few well controlled studies of the degradability of toxaphene in water
systems have been conducted. Hughes (1970) reviewed a study by Mitchum (1963)
where 1.0 mg/liter toxaphene was placed in some filtered, natural waters of
Wyoming and held in the dark at temperatures of 26 C, 18 C, and 7 C for 521 days.
Analysis of the water by paper chromatograph was undertaken every three months.
Extrapolation of the toxaphene concentration in water vs. time curve provided
estimated times for complete breakdown (645 days for 26 C; 2,680 days for
7 C). Interpretation of the data is difficult because no particulate matter
is present (adsorption to particulates may affect rate of degradation). The
presence of microorganisms in filtered water is unclear, and the initial con-
centration of toxaphene exceeds the water solubility of toxaphene (some loss
might be due to precipitation [Hughes, 1970]).
Wolfe et al. (1976) have conducted some screening studies of hydrolysis
and photolysis of toxaphene in aqueous media. They placed toxaphene in air-
saturated dilute acidic and alkaline medium at 65 C. After two days they
detected no change in the gas chromatographic fingerprint. In another study, they
concluded that the photolysis of toxaphene under sunlight is a very slow pro-
cess in pure water and that photochemically-generated singlet oxygen does not
readily oxidize toxaphene. Sanborn et al. (1976) studied the fate of radiolabelled
7-42
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(C-14 at the 8 carbon) and unlabelled toxaphene in a terrestrial-aquatic model
ecosystem. Gas chromatography with electron capture detection was used with
the unlabelled material for comparison. The results (Table 7.19) show that
toxaphene is stable enough to be bioaccumulated in substantial quantities in a
variety of aquatic organisms. On the other hand, the results also indicate
some breakdown to new compounds or unextractables. However, in terms of the
degradability of toxaphene in natural water systems, the results are difficult
to interpret. The normal operation of the terrestrial-aquatic model ecosystem
uses sand, rather than sediment, which most likely will have considerable
impact on the rate of breakdown as well as the degree of bioaccumulation.
Investigators monitoring water and sediment samples have long noticed a
difference between the gas chromatography fingerprint of residue toxaphene and
standard toxaphene. Johnson et al. (1966) detected 1 to 4 micrograms/liter of
toxaphene in eight treated Wisconsin lakes. This concentration of standard
toxaphene is used to kill small fish, yet the residue concentration was not
toxic to fish. The authors found differences in the gas chromatographic
fingerprint and suggested that components of toxaphene, especially the ones
responsible for fish toxicity, degrade at different rates. Similar results
were found by Terriere et al. (1966) and Hughes (1970). Hughes et al. (1970)
concluded that selective degradation of components does occur to a limited
extent at a slow rate in the aquatic environment.
Hughes (1970) is the only researcher who has attempted to characterize
the alterations that occur. Typical fingerprint alterations for toxaphene
residues from lake water or sediment are illustrated in Figures 7.4 and 7.5.
In general, the gas chromatographic changes consist of the diminution of
7-43
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Table 7.19. Toxaphene in a terrestrial-aquatic model ecosystem^
-P-
«£c
Total C
Toxaphene 0. 70
I 0.57
IV 0.34
Origin 0.00
Unext rac table
Toxaphene equivalents (ppm)
Oedogonium Phyea Gulex
Water (alga) (snail) (mosquito)
0.04441 13.2941 17.6198 2.2570
0.00159 10.9743 15.2639 1.4147
0.00106 1.7535 1.8360 0.2359
0.00164 0. 1130 0. 0585 trace
0.02002 0.0944 0.0211 0.2022
0.01328 2.2156 1.1153 1.1245
Gambusia
(fish)
10.3977
6.7523
2.4923
0.1487
0.1674
4.2264
^Source: Sanborn et al. , 1976. ,
C Jl.nCl (67 - 69% chlorinated camphene) 8 - C
10 10 o
.Silica Gel OF 254, Skellysolve B (bp 68°C) : diethyl ether: acetone 80:20:10, by volume.
-
. ,
Roman numerals - cliemical structure unknown.
-------
a - SURFACE WATER 7/23/66
b - 2.5 NANOGRAM ( ng) TOXAPHENE STANDARD
01
oo
O
Q.
00
UJ
a.
cc.
01
a
cc
o
CJ
ai
CC
TIME (MINUTES)
Figure 7.4. Gas chromatograms of standard toxaphene and toxaphene extracted
from Cornstock Lake Water. Source: Hughes, 1970.
7-45
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a - FOX LAKE SEDIMENT (190 DAYS)
b - OTTMAN LAKE SEDIMENT (191 DAYS )
c - STANDARD TOXAPHENE ( 2 ng )
01
00
z
O
a.
V)
LLJ
CC
GC
LU
Q
cc
O
u
12 10 8 64
TIME (MINUTES)
Figure 7.5. GC patterns of toxaphene from sediments of Fox Lake and Ottman
Lake compared to standard toxaphene. Source: Hughes, 1970.
7-46
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peaks near the end of the pattern with increasing strength of the early eluting
peaks (Hughes, 1970). This same effect occurs when standard toxaphene is de-
hydrohalogenated. Hughes (1970) ruled out a dehydrohalogenation process for
the formation of residue toxaphene by demonstrating that residue toxaphene
is unaffected by nitration, whereas dehydrohalogenated standard toxaphene is
altered by nitration.
In summary, the available information suggests the label "hard," as used
with dieldrin and DDT, does not apply to the persistence of toxaphene residue
toxicity in water (Hughes, 1970). Also, the persistence of toxaphene in the
chemical sense also appears to be somewhat less than that of other organochlor-
ine pesticides. For example, the concentration in sediment decreases by a
factor of 2 every 20 days. Nevertheless, detectable quantities of toxaphene
are found in sediment and water 3 to 9 years after treatment of a lake.
Concentrations of toxaphene in water rapidly decrease because the chemical is
absorbed on suspended particulates and sediment.
7.3.3 Mobility and Persistence in Soil
Toxaphene has infrequently been detected in soils which have not been
exposed to the pesticide during crop protection (Section 7.2.3). For example,
Wiersma et al. (1972 b) detected toxaphene in only one soil sample out of 199
non-crop samples. A study of soils in 8 cities detected toxaphene in 4 out
of 400 sites and only in southern cities. This suggests that the major source
of soil toxaphene residues is from its direct use as a pesticide, mostly for
the protection of cotton.
Estimates of the half residence time for toxaphene in soil vary from
100 days (LaFleur et al., 1973) to 11 years (Nash and Woolson, 1967). Because
7-47
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of the experimental conditions used, Nash and Woolson (1967) expected that
their results provided an upper limit of persistence. In 1951 they applied
toxaphene to Congarec sandy loam soil in approximate concentrations of 55.8
and 137.0 ppm (Nash et al., 1973). The plots were surrounded by concrete
walls and the pesticide was thoroughly mixed before being placed to a depth
of 38 cm and provided with gravel and tile drainage. After 14 years, 45 percent
of the toxaphene remained in the soil. The authors felt that the treatment
and maintenance of the soil samples would allow minimum leaching, volatiliza-
tion, photodecomposition and mechanical losses because of the thorough mixing
and minimum tillage and probably little biological decomposition because of
the high rate of application which may have been toxic to the soil micro-
organisms. Twenty years after treatment Nash et al. (1973) used the same
plot to compare the efficiencies of three methods (shake, Soxhlet, and column)
for extracting toxaphene from soil. By taking the mean value for the Soxhlet
and shake extraction recoveries at both treatment rates, they determined that
45.1 percent of the total amount of toxaphene residue remained after 20 years.
LaFleur et al. (1973) used a 10 x 10 meter plot of Dunbar soil which
overlay a water table that was usually less than 3 meters from the surface.
Toxaphene was sprayed at a rate of 100 kg/ha and then hand raked into the soil.
They plotted the change in concentration with time and determined that the
loss was rapid at first but slower after 80 days. The calculated half resi-
dence time was 100 days (32 ppm to 4 ppm in 1 year).
The residence time of toxaphene in soil has been reported by a number
of other investigators. Westlake and SanAntonio (1960) found that the levels
of toxaphene in air-dried soil decreased from 140 to 85 ppm over a six year
7-48
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period. Adams (1967), using the data of Foster et al. (1956), has calculated
the approximate half-life in years of various pesticides in three soils. The
values for toxaphene were 2.0, 0.8, and 0.8 for Beltsville, Mississippi, and
New Jersey soils, respectively. Hermanson et al. (1971) followed for 9 years
the concentration of toxaphene in a Holtville sandy clay that received a treat-
ment of 22.4 kg/ha for 4 years followed by no treatment for 5 years. They
rated toxaphene as 0.18 on a persistency index where 1.00 was equal to no
degradation during the first year (DDT = 0.26). They also calculated a
persistence half-life of 4 years which was the same as the value for DDT.
A slight correlation between persistence and vapor pressure was noted.
Seiber et al. (1975) analyzed soil samples from a California cotton field
6 and 12 months after spraying. They found a concentration of approximately
5 ppm at 6 months and approximately 3.5 ppm at 12 months. Bradley et al.
(1972) found that the toxaphene residue in acidic loam declined during the
year sampling period while the DDT residue stayed basically the same.
This range of residence times may be explained by climate and soil
factors. Chemicals can be degraded or lost from soil by volatilization
and codistillation, leaching, oxidation, hydrolysis, and microbial degrada-
tion (Edwards, 1966). Some photodegradation is possible but only on the
top surface. These processes are affected by climate and soil factors,
such as soil type, pH, organic matter content, mineral (especially clay)
content, drainage and soil moisture, and cropping (Edwards, 1966- Adams,' 1967).
Although the effect of these factors cannot be quantitatively correlated,
some general qualitative statements are possible. Heavy clay soils appear
to increase the persistence of chlorinated hydrocarbons (Harris, 1966; Edwards,
1966) and temperature increases result in increased rates of loss (Edwards,
1966). Little evaporation occurs from dry soil (which may explain the results
7-49
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of Westlake and SanAntonio, 1960) while considerable evaporation occurs from
wet soil (Adams, 1967). Biodegradation may increase on addition of carbon
sources such as manure because of the increase in metabolic activity of the
microorganisms (Adams, 1967). Edwards (1966) has listed the following fac-
tors affecting soil insecticide persistence in decreasing order of importance:
(a) the chemical structure of the insecticide;
(b) the type of soil to which the insecticide is applied, especially
depending on the organic matter content; amount of rainfall reaching
treated soil and the moisture in the soil;
(c) the microbial population of the treated soil; depth of cultivation
of the insecticide; mean temperatures in the treated soil;
(d) the mineral content and acidity of the treated soil; amount of
plant cover over treated soil; formulation and concentration of
the insecticide.
The effect of these environmental factors on the fate of toxaphene in
soil has been studied by only a few investigators. In a recent study, Parr
and Smith (1976) studied the degradation of toxaphene in Crowley silt loam
(pH 6.0) under a variety of conditions. They used both aerobic and anaerobic
conditions as well as a sterilized sample that was later inoculated with
microorganisms. Both the loss of toxaphene and the cumulative release of
carbon dioxide were measured. In order to see the effect of organic matter,
some samples were amended with alfalfa meal. The percent toxaphene remaining
(starting concentration 10 ppm) after various times is depicted in Figure 7.6.
The results show that no degradation occurs under aerobic conditions, while
substantial breakdown occurs under anaerobic conditions. Addition of extra
7-50
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100
ENVIRONMENTS:
• CO;-FREE AIR. MOIST (0.3 BAR)
A NITROGEN, MOIST (0.3 BAR)
*• NITf'OGEN FLOODED, UNSTIRRED
NTFOGEN, FLOODED, STIRRED
MOIST (0.3 BAR),
AUTOCLAVED, REINOCULATED
REINOCULA7ED
CLDSED SYMBOLS:
ALFALFA MEAL (1%)
SYMBOLS:
UNAMENDED
TIME-WEEKS
Figure 7.6. Degradation of toxaphene in soil unamended or amended with alfalfa
meal and incubated in a moist aerobic environment or under moist and flooded
anaerobic conditions. Source: Parr and Smith, 1976.
7-51
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organic matter has a positive effect on degradation under anaerobic conditions.
It appears that microorganisms are degrading the toxaphene since no loss occurs
in a sterilized, anaerobic sample until it is inoculated. Plimmer (1974)
reported that anaerobic conditions, such as flooded soils or sediment, appear
to accelerate reductive metabolism. Parr and Smith (1976) also measured the
redox potential and concluded that degradation is very rapid between 0 to -100 mV.
Leaching of toxaphene through soil has been examined by several researchers.
Swoboda and coworkers (1971) analyzed three plots of Houston black clay soil
(50 to 60 percent montmorillonitic clay) which received a total of 11.2, 20.16,
and 24.64 kg/ha. The percentage of the applied toxaphene recovered in the
soil varied from 10 to 20 percent, depending upon the rate of application. By
measuring the concentration of toxaphene at various depths, it was determined
that 88 to 95 percent of the toxaphene was found in the top 12 inches (measured
by foot increments down to 60 inches). The authors concluded that very little
leaching occurred. Nash and Woolson (1968) reported similar results with
toxaphene residues measured in Congarec sandy loam soil 15 years after appli-
cation. They found 85 percent of the toxaphene in the upper 23 cm, which
probably corresponds to the cultivated layer. A study of toxaphene desorption
from soil using various solvents indicated that little toxaphene would be
desorbed from soil by water (LaFleur, 1974).
Although all the above information suggests that toxaphene does not leach
through soil, LaFleur and coworkers (1973) found that toxaphene was leached
through Dunbar soil overlying a water table less than 3 meters from the sur-
face. This may have been assisted by the high total rainfall encountered (13
percent higher than mean for the past 72 years).
7-52
-------
The ability of toxaphene to migrate from the surface to ground water was
suggested by the work of Johnston et al. (1967). They detected toxaphene in
flooded plots where they were studying the movement of other pesticides.
Since they had not used toxaphene, they concluded that the toxaphene was from
contaminated ground water.
Bradley et al. (1972) examined the pesticide runoff from two locations
each of which were either treated with toxaphene (26.8 kg/ha) or a combination
of toxaphene (268 kg/ha) and DDT (13.4 kg/ha). Only 0.3 to 0.6 percent of the
applied toxaphene was detected in the runoff. Of that recovered, 75 percent
was associated with sediment (96 percent with DDT). The amount in the runoff
was dependent upon climatic factors such as frequency, intensity, and duration
of rain and soil physical properties. The toxaphene runoff into a farm pond
was enough to exceed the 96 hour medium tolerance limit for blue gill (3.5 ppb)
Only two studies have discussed the alteration of the gas chromatographic
fingerprint resulting from the weathering of toxaphene in soil (Parr and
Smith, 1976; Seiber et al., 1975). Seiber et al. (1975) found that the less
volatile components (longer retention time components) tended to be enriched.
This was similar to what was found with residues on foliage which was attribu-
ted to selective evaporation. The effect in soil may be attributed to drain-
age from leaves since the residue concentration in soil increased at first.
In contrast, Parr and Smith (1976) found a diminution of the later peaks
when toxaphene was incubated with soil under anaerobic conditions. This re-
sult is similar to that which Hughes (1970) found with toxaphene residues in
water and sediment.
7-53
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Hughes, R. 1970. Studies on the Persistence of Toxaphene in Treated Lakes.
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/
>/oi
.ughes, R.A., Veith, G.D. and Lee, G.F. 1970. Gas-Chromatographic Analysis
of Toxaphene in Natural Waters, Fish, and Lake Sediments. Water Res. 4_(8):
547-58.
Johnson, D.W., Lee, G.F. and Spyridakis, D. 1966. Persistence of Toxaphene
in Treated Lakes. J. Air Wat. Pollut. Intern. 10:555-560.
Johnston, W.R., Ittihadish, F.T., Craig, K.R. and Pillsbury, A.F. 1967. In-
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Kallman, B.J., Cope, O.B. and Navarre, R.J. 1962. Distribution and Detoxi-
cation of Toxaphene in Clayton Lake, New Mexico. Trans. Am. Fish Soc. 91:
14-22.
Keith, J.O. 1966. Insecticide Contaminations in Wetland Habitats and Their
Effects on Fish-Eating Birds. J. Appl. Ecol. ^(suppl.):71-85.
LaFleur, K.S. 1974. Toxaphene-Soil-Solvent Interactions. Soil Sci. 117(4);
205-10.
LaFleur, K.S., Wojeck, G.A. and McCaskill, W.R. 1973. Movement of Toxaphene
and Fluometuron Through Dunbar Soil to Underlying Ground Water. J. Environ.
Qual. _2(4):515-518.
Lichtenberg, J.J. 1971. Analytical Quality Control Laboratory, EPA, Cincinnati,
Ohio. Personal communication in Hartwell et al., 1974.
Lichtenberg, J.J., Eichelberger, J.W., Dressman, R.C. and Longbottom, J.E. 1970.
Pesticides in Surface Waters of the United States - A 5-Year Summary, 1964-68.
Pest. Monit. J. 4_:71-86.
rLiss, P.S. and Slater, P.G. 1974. Flux of Gases Across the Air-Sea Interface.
Nature 247:181-184.
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Mackay, D. and Leinonen, P. 1975. Rate of Evaporation of Low-Solubility
Contaminants from Water Bodies to Atmosphere. Environ. Sci. Technol. 9^(13):
1178-1180.
Manigold, D.B. and Schulze, J.A. 1969. Pesticides in Selected Western Streams
A Progress Report. Pestic. Monit. J. _3:124-135.
Mattraw, H.C. 1975. Occurrence of Chlorinated Hydrocarbon Insecticides -
Southern Florida - 1968-72. Pestic. Monit. J. J9:106-115.
Mitchum, D.L. 1963. Study of Fish Toxicants. Job Completion Report, Proj.
No. FW-3-R-10, Game and Fish Lab. Research, State of Wyoming, pp. 28-32. Re-
viewed in Hughes, 1970.
/Mullins, D.E., Johnson, R.E. and Starr, R.I. 1971. Persistence of Organo-
/ chlorine Insecticide Residues in Agricultural Soils of Colorado. Pestic.
V Monit. J. 1(3):268-275.
Nash, R.G. and Woolson, E.A. 1967- Persistence of Chlorinated Hydrocarbon
Insecticides in Soils. Science 157:924-927.
Nash, R.G. and Woolson, E.A. 1968. Distribution of Chlorinated Insecticides
in Cultivated Soil. Soil Sci. Soc. Amer. Proc. 32:525-527.
Nash, R.G., Harris, W.G., Ensor, P.D. and Woolson, E.A. 1973. Comparative
Extraction of Chlorinated Hydrocarbon Insecticides from Soils 20 Years After
Treatment. J. Assoc. Offic. Anal. Chem. 56:728-732.
Nicholson, H.P. 1969. Occurrence and Significance of Pesticide Residue in
Water. J. Washington Acad. Sci. 59(4-5);77-85.
Nicholson, H.P., Grzenda, A.R., Lauer, G.J., Cox, W.S. and Teasley, J.I. 1964.
Water Pollution by Insecticides in an Agricultural River Basin. I. Occurrence
of Insecticides in River and Treated Water. Limnol. Oceanog. £:310-318.
Nicholson, H.P., Grzenda, A.R. and Teasley, J.I. 1966. Water Pollution by
Insecticides: A Six and One-Half Year Study of a Watershed. Proc. Symp.
Agric. Waste Waters Report #10 of Water Resources Center. University of
California, pp. 132-141.
Parr, J.F. and Smith. S. 1976. Degradation of Toxaphene in Selected Anaerobic
Soil Environments. Soil Science 121:52-57.
Plimmer, J.R. 1974. Symposium on Toxaphene Composition and Environmental Fate.
168th National Meeting of the American Chemical Society, Atlantic City, N.J.,
Sept. 9-13.
Reimold, R.J. 1974. Toxaphene Interactions in Estuarine Ecosystems. National
Technical Information Service COM-75-10104/8GA. Springfield, Virginia.
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Reimold, R.j. and Duranti, C.J. 1972a. Survey of Toxaphene Levels in Georgia
Estuaries. Technical Report Series Number 72-2. Georgia Marine Science Center,
Skidaway Island, Georgia. 51 pp.
Reimold, R.J. and Duranti, C.J. 1972b. Monitoring Toxaphene Contamination in
a Georgia Estuary. National Technical Information Service COM 73-1072/.
Springfield, Virginia.
Reimold, R.J. and Durant, C.J. 1974. Toxaphene Content of Estuarine Fauna
and Flora Before, During, and After Dredging Toxaphene-Contaminated Sediments.
Pestic. Monit. J. 8^(1):44-49.
Sanborn, J.R., Metcalf, R.L., Bruce, W.N. and Lu, P.-Y. 1976. The Fate of
Chlordane and Toxaphene in a Terrestrial-Aquatic Model Ecosystem. Environ.
Entomol. _5(3) : 533-538.
Sand, P.F., Wiersma, G.B. and Landry, J.L. 1972. Pesticide Residues in Sweet
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Annual Summaries of Water-borne Chlorinated Hydrocarbon Pesticide Program.
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Hazardous Materials Advisory Committee, U.S. EPA.
Schafer, M.L., Peeler, J.T., Gardner, W.S. and Campbell, J.E. 1969. Pesticides
in Drinking Water: Waters from the Mississippi and Missouri Rivers. Environ.
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Schulze, J.A., Manigold, D.B. and Andrews, F.L. 1973. Pesticides in Selected
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/Seiber, J.N., Landrum, P.F., McChesney, M.M. and Madden, S.C. 1975. Toxaphene
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'Stevens, L.J., Collier, C.W. and Woodham, D.W. 1970. Monitoring Pesticides
in Soils from Areas of Regular, Limited, and No Pesticide Use. Pestic. Monit.
J. j4(3): 145-164.
Swoboda, A.R., Thomas, G.W., Cady, F.B., Baird, R.W. and Knisel, W.G. 1971.
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7-57
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Trans. N.Y. Acad. Sci. Ser. 2., 28;569-578.
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Program - Six States, 1967. Pestic. Monit. J. ,5:223-227.
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JFY 1969 - National Soils Monitoring Program. Pestic. Monit. J. 6^:194-201.
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in Onions and Soil - 1969. Pestic. Monit. J. 5_: 345-347.
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Chemical and Photochemical Transformation of Selected Pesticides in Aquatic
Systems. U.S. EPA. Office of Research and Development. Athens, Georgia.
7-58
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8.0 ENVIRONMENTAL INTERACTIONS AND THEIR CONSEQUENCES
8.1 SUMMARY
Toxaphene is released into the environment primarily from its application
as an insecticide for the protection of cotton, mostly in the southern states.
Present production of toxaphene is estimated at 49 million kg per year.
Toxaphene sprayed on cotton either evaporates or is deposited upon soil or
foliage. Toxaphene in air can travel for long distances (1200 km). Residues
in soil will evaporate, degrade, or, depending upon the soil and climate
conditions, sometimes will leach into the ground water. Evaporation from
water or adsorption to sediment or suspended solids appear to be the primary
mechanisms for removal of toxaphene from water. Human exposure to toxaphene
through food or terrestrial sources appears to be very limited, although some
residues have been detected in birds, wildlife, and food products. Bioconcen-
tration of toxaphene in aquatic organisms is very sizable (in fish up to
75,000 times the concentration found in water) and may result in human expo-
sure and residues in fish-eating birds.
8.2 SOURCES OF TOXAPHENE
Toxaphene is a complex mixture of polychlorinated diterpenes which is
produced by the chlorination of the diterpene, camphene. It is now the most
commonly used chlorinated hydrocarbon insecticide. While release to the en-
vironment primarily occurs when it is applied for pest control, some is re-
leased during manufacture from formulating plants and from accidental spills
(Gerakis and Sficas, 1974). There are no known natural sources or inadvertant,
man-made sources of toxaphene.
8.2.1 Toxaphene Production and Consumption
Since the agricultural chemical industry will not release precise figures
on production or consumption for proprietary reasons (VonRumker et al., 1974),
8-1
-------
the available data are only estimates. Table 8.1 describes production by manu-
facturer and plant location for the years 1972 and 1975. The largest production
plant is the Hercules Inc. plant located at Brunswick, Georgia. Other plants
which have large manufacturing capacity are the Tenneco Chemicals (Fords, NJ)
and Sonford Chemical Co. (Houston, TX). As a result of the environmental
regulations limiting the use of persistent, organochloride insecticides,
toxaphene production and consumption has been rapidly increasing. Ifeadi
(1975) projects that production will reach 105.9 million kilograms of active
ingredient in 1980 (compared to 49.1 million kilograms in 1975) based on the
present market conditions.
Toxaphene mainly enters the environment through application as an insecti-
cide. It is consumed in distinct regional and seasonal patterns. It is
primarily applied in the southern states and in California for the control of
pests infesting cotton farms (Guyer et al., 1971; Hartwell et al., 1974;
VonRumker et al., 1974). Although the amount used for other farming applica-
tions is small, significant amounts of toxaphene are still used as a livestock
insecticide. In the past, toxaphene has been used for rough fish control in
lakes. Table 8.2 and Figure 8.1 describe the consumption by geographical
region for 1972 (VonRumker et al., 1974). Hartwell et al. (1974) have sug-
gested that the consumption in California might be understated. While VonRumker
and co-workers have suggested that about 8 percent of the toxaphene is consumed
in California, Bailey and Hannum (1967) have stated that about 20 percent of the
total pesticides for the U.S. are consumed in California.
Seasonal variation in toxaphene consumption follows the need for its
application in its major use, cotton farming. Several investigators (Nicholson
et al., 1966; Tabor, 1966; Stanley et al., 1971; Willis et al., 1976) have
concluded that the usual time of application for toxaphene ranges from late
8-2
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Table 8.1 Toxaphene production
Manufacturer
Location
Capacity
(millions of
kilograms)
Production
(millions of
kilograms)
1972
1975
Hercules, Inc. Brunswick, GA
Synthetics Dept.
Tenneco Chemicals Fords , NJ
Intermediates Div.
Sonford Chemical Co. Houston, TX
Vicksburg Chemical Vicksburg, MS
Co.
TOTAL
22.7-34.1
56.8
18.2
2.3
25.4
9.1
0
0
34.5
29.5
9.1
9.1
1.4
49.1
Source: Adapted from VonRumker et al. , 1974 and Ifeadi, 1975.
3-3
-------
Table 8.2 Toxaphene consumption in 1972
a.
Distribution of Consumption
Toxaphene (millions of kilograms)
U.S. Production 34.5
Imports 0
Exports 8.2
U.S. Consumption 26.3
Industrial
Region Agricultural Commerical
Northeast Negl. 0.1
Southeast 9.1 0.1
North Central 0-5 0.1
South Central 14.3 0.1
West 2.0 0.1
TOTAL 25.9 0.5
Total
0.1
9.2
0.5
14.4
2.1
26.3
Source: VonRumker et al. ,1974.
3-4
-------
CO
I
Ul
^Missouri I East North Central
Toxaphene Millions
1972 Estimated: kg Active In9red.ent
. U. S. Production 34-4
. Imports None
. Exports 8 2
.U.S. Supply 263
^"Production Plant
Figure 8.1 Materials flow diagram for toxaphene, 1972. Source: von Rumker et al., 1974.
-------
July into early August. This period corresponds to the incursion by boll
weevils and bollworms. Toxaphene is sometimes applied in the spring to
control cutworm infestation.
8.3 ENVIRONMENTAL CYCLING OF TOXAPHENE
A variety of degradative and transport processes may effect the environ-
mental fate of toxaphene (Section 7.3). Intermedia processes which affect the
environmental cycling of toxaphene include evaporation, water leaching from
soil, adsorption to soil followed by erosion, washout by rain, and adsorption
to particulate matter followed by precipitation (effective in both water and
air). The high vapor pressure of commercial toxaphene, compared to other
organochlorine pesticides, assures that atmospheric processes are extremely
important. Because commercial toxaphene consists of numerous components,
these intermedia processes have considerable potential for altering the
chemical composition of the toxaphene residue.
Quantitative information on the cycling of toxaphene is not available.
Most studies have been closely tied to a particular release source; (1) con-
ventional crop (cotton) spraying, (2) treatment of lakes for rough fish control,
or (3) loss in water from production plants.
Toxaphene sprayed on crops will be either deposited on the foliage or soil
or will evaporate. Air monitoring information (Arthur et al., 1976; Stanley
et al., 1971; Tabor, 1965, 1966) indicates that atmospheric residues of toxaphene
are highest in the southern cotton belt and that the residue concentrations are
highest during the months of July and August when the greatest rates of appli-
cation occur. The evaporated toxaphene, probably as a vapor, although it might
adsorb to particulate matter, can travel long distances (1200 km) and become
well mixed in the atmosphere (Bidleman and Olney, 1975). Monitoring of toxaphene
8-6
-------
residues in air at locations remote from application sites indicate that the
chemical makeup of toxaphene is only slightly altered. In contrast, Seiber
et al. (1975) found that toxaphene detected above a cotton field showed a sub-
stantial enrichment of the more volatile components. The remote monitoring
results suggest that significant amounts of all the toxaphene components may
be lost to the atmosphere during spraying. Swoboda et al. (1971) in their
field study, could only recover 10 to 20 percent of the applied toxaphene in
the soil.
Of the sprayed toxaphene that reaches the foliage and soil, substantial
amounts appear to evaporate. Seiber et al. (1975) concluded that the more
volatile components which are deposited on foliage evaporate, while the less
volatile components are washed off and drop into the soil. In contrast, in
soil, the percentage of the more volatile components increases, suggesting
some type of degradation of the less volatile components. Carlin et al. (1974)
found no evidence of metabolism of 36Cl-toxaphene applied to cotton plants
maintained in a closed all-glass system for 7 days. Thus, the mechanism of
loss was volatilization.
Toxaphene residues in soil may be transported by erosion, leaching, or
evaporation. The rate of toxaphene evaporation from soil has not been studied,
but is likely to be dependent upon a variety of soil factors, especially soil
moisture (Edwards, 1966). Bradley et al. (1972) found that 0.3 to 6.0 percent
of the applied toxaphene was detected in the runoff (75 percent associated with
sediment). Monitoring of a watershed which drains an agricultural area con-
taining small cotton farms (Nicholson et al., 1964) also indicates that water
contamination may result from runoff. Whether toxaphene residues in soil can
leach into ground water is probably dependent upon the particular soil and
climate conditions. Swoboda and coworkers (1971) determined that 88 to 95
percent of the toxaphene residue was found in the top 30.5 cm (measured by
8-7
-------
foot increments down to 152.4 cm). Similar results were found by Nash and
Woolson (1968). However, LeFleur et al. (1973) found that toxaphene was
leached through soil overlying a water table less than 3 meters from the
surface. Johnston et al. (1967) also found suggestive evidence that toxaphene
from contaminated ground water could migrate into flooded plots.
Considerable study of lakes treated with toxaphene for rough fish control
suggest that toxaphene is removed from water by evaporation or codistillation
and by adsorption to sediment or to suspended particulate matter, followed by
settling. Shallow, biologically rich lakes become detoxified (in respect to
fish) faster than deep, biologically sparse lakes (Terriere et al., 1966),
which is consistent with a toxaphene adsorption mechanism (Veith and Lee,
1971). However, monitoring data from river systems suggests that adsorption
may not reduce the concentration of toxaphene in river water. For example,
Grzenda and Nicholson (1965) found concentrations of toxaphene in river water
varying from 30 to 140 ppt, but no toxaphene in river bottom sediment. In
contrast, Bailey and Hannum (1967) found higher concentrations of toxaphene in
sediment than in water. The calculated half-life for toxaphene evaporation
from water is 10.4 hours, and monitoring studies by Veith and Lee (1971) have
suggested that greater than 50 percent of the toxaphene applied to lakes is
lost through evaporation or microbial degradation. As with soil residues,
toxaphene residues detected in water systems have a decreased percentage of
less volatile material, but it has been shown that this is not due to dehydro-
chlorination (Hughes, 1970).
Reimold and Durant (1972) have studied toxaphene residues in water systems
which receive effluents from the Hercules production plant in Terry Creek,
8-8
-------
Brunswick, Georgia. They concluded that toxaphene was confined to suspended
material in the water and the concentration of residues in the sediment de-
creased with an increase in distance away from Terry Creek. These results
again suggest that adsorption to sediment may be an important process for
removing toxaphene from water.
8.4 FOOD CHAINS
8.4.1 Toxaphene in Food
Human ingestion of toxaphene in food is apparently low. The annual market
basket survey conducted by the Food and Drug Administration during a four year
period (Table 4.6) showed that most products contained no detectable concen-
trations of toxaphene. The only exceptions were leafy vegetables and garden
fruits, both of which had very low levels (calculated intake of approximately
0.001 mg per day). The low concentrations in food are probably due to the fact
that toxaphene is mostly used on cotton crops, and washing and food processing
remove considerable amounts of toxaphene (Van Middlelem, 1966; Van Middelem
and Wilson, 1960; Hartwell et al., 1974). Toxaphene residues have been detec-
ted in tobacco products (Domanski et al., 1975), probably as a result of drift
from cotton spraying in nearby fields.
8.4.2 Terrestrial Ecosystems
Bioaccumulation of toxaphene residues in terrestrial ecosystems is not well
understood. Uptake and adsorption of toxaphene in plants has not been studied,
although residues have been detected in some instances. These residues are
apparently due mostly to surface deposition (drift or intentional spraying),
rather than uptake from soil (Hartwell et al., 1974). Little information on
adsorption and distribution in birds and terrestrial wildlife is available,
8-9
-------
but residues have been detected in some species. For example, in a study in
Alabama, Causey et al. (1972) detected toxaphene in 5 of 20 (average concen-
tration 30 ppm) quail, and detectable concentrations were found in 2 of 31
rabbits and 3 of 22 deer. Toxaphene residues have been detected in fish-
eating birds (Keith, 1965) and in non-fish-eating birds (Markley, 1974), and
the passage of toxaphene into the eggs of wild birds has been documented.
Toxaphene residues have not been detected in domestic animals except in
controlled exposure studies. Several studies have demonstrated that cows fed
toxaphene in their diet (Zweig et al., 1963; Claborn et al., 1963) or in
treated hay (Bateman et al., 1953) have residues in their milk. However,
toxaphene has not been detected in milk in the annual market basket survey
(Section 8.4.1).
8.4.3 Aquatic Ecosystems
Toxaphene appears to have considerable potential for bioaccumulating in
aquatic organisms, resulting in possible exposure to humans. Aquatic plants
are able to bioaccumulate toxaphene from the water (Section 4.0). For example,
Terriere et al. (1966) found 15.5 ppm toxaphene in aquatic plants which were
exposed to water containing 2 ppb. Other levels of the food chain, such as
bacteria, fungi, and green algae (Paris et al., 1975), are also capable of
bioaccumulating toxaphene. Bioconcentration of toxaphene in aquatic inverte-
brates, such as oysters (Lowe et al., 1971), plankton (Johnson, 1966), and
daphnia (Schoettger and Olive, 1961), has also been documented.
Several studies have indicated that toxaphene is bioconcentrated in fish
by a factor of several thousand. This conclusion has been supported by
several monitoring studies (Kallman et al., 1962; Terriere et al., 1966;
8-10
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Hughes and Lee, 1973) and laboratory investigations (Hughes, 1970; Sanborn
et al., 1976; Mayer et al., 1975). Sanborn et al. (1976) exposed mosquito
fish for three days to 44.4 ppb toxaphene (C-14 labelled) in a model ecosystem
and found a concentration factor of 4247. Mayer et al. (1975) have demon-
strated that toxaphene can be concentrated up to 75,000 times in fish, with
the rate appearing to be dependent upon the lipid concentration in the fish.
Hughes and Lee (1973) found that the concentration in prey fish stocked in a
toxaphene treated lake was more than the concentration in predator fish, which
suggests that biomagnification may not be an important process. The bioconcen-
tration of toxaphene in fish is further supported by numerous field monitoring
studies (Table 5.6) which have detected toxaphene in fish samples. However,
the levels that have commonly been detected are relatively low and in some
instances no toxaphene is detected. For example, Reimold and Shealy (1976)
monitored chlorinated pesticide residues in fish collected from 11 estuaries
representing all the Atlantic drainage basins in Georgia and South Carolina.
Although they detected dieldrin, DDf, and PCB's, no measurable residues of
toxaphene were found even though a toxaphene manufacturing plant was located
in New Brunswick, Georgia.
3-11
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Hartwell, W.V., Datta, P.R., Markley, M., Wolff, J., Aspelin, A. and Billings,
S.C. 1974. Aspects of Pesticidal Use of Toxaphene and Terpene Polychlorinates
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Hughes, R. 1970. Studies on the Persistence of Toxaphene in Treated Lakes.
University of Wisconsin, Ph.D. Thesis, Published by University Microfilms,
Ann Arbor, Michigan.
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Ifeadi, C.W. 1975. Screening Study to Development Background Information
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Johnson, W.C. 1966. Toxaphene Treatment of Big Bear Lake, California. Calif.
Fish and Game 52(3);173-179.
Johnston, W.R., Ittihadish, F.T., Craig, K.R. and Pillsbury, A.F. 1967. In-
secticides in Tile Drainage Effluent. Water Resources Research 3^:525.
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cation of Toxaphene in Clayton Lake, New Mexico. Trans. Amer. Fisheries Soc.
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Nicholson, H.P., Grzenda, A.R. and Teasley, J.I. 1966. Water Pollution by
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8-15
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TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)
\. REPORT NO.
EPA-600/1-79-044
2.
3. RECIPIENT'S ACCESSION«NO.
4. TITLE AND SUBTITLE
Reviews of the Environmental Effects of Pollutants:
X. Toxaphene
5. REPORT DATE
November 1979
6. PERFORMING ORGANIZATION CODE
7. AUTHOR(S)
P.R. Durkin, P.H. Howard, J. Saxena, S.S. Lande,
J. Santodonato, J.R. Strange, and D.H. Christopher
8. PERFORMING ORGANIZATION REPORT NO
ORNL/EIS-130
9. PERFORMING ORGANIZATION NAME AND ADDRESS
Center for Chemical Hazard Assessment
Life and Material Sciences Division
Syracuse Research Corporation
Merrill Lane
10. PROGRAM ELEMENT NO.
susa
11. CONTRACT/GRANT NO.
Contract No. W-7405-eng-26
Subcontract No. 448 and 7663'
12.
Health Effects Research Laboratory
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, Ohio 45268
13. TYPE OF REPORT AND PERIOD COVERED
Final Report •
14. SPONSORING AGENCY CODE
15. SUPPLEMENTARY NOTES
16. ABSTRACT
The environmental effects of toxaphene are extensively reviewed. Information
is presented on chemical properties and analytical techniques, environmental occurrence,
cycling, and fate, as well as on food chain interactions. Biological aspects of
toxaphene in microorganisms, plants, wild and domestic animals, and humans and test
animals are reviewed, including metabolism, toxicity, carcinogenicity, mutagenicity,
and teratogenicity.
17.
KEY WORDS AND DOCUMENT ANALYSIS
DESCRIPTORS
b.lDENTIFIERS/OPEN ENDED TERMS
c. COSATl Field/Group
Pollutants
Toxaphene
Toxicology
Health Effects
8. DISTRIBUTION STATEMENT
Release to public
19. SECURITY CLASS (This Report)
Unclassified
21. NO. OF PAGbS
495
20. SECURITY CLASS (This page)
Unclassif-ipH
22. PRICE
EPA Form «20-1 (9-73)
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