EPA-600/1-76-010a
January 1976
Environmental Health Effects Research Series
ASSESSMENT OF TOXICITY OF
AUTOMOTIVE METALLIC EMISSIONS
Volume I
Health Effects Research Laboratory
Office of Research and Development
U.S. Environmental Protection Agency
Research Triangle Park, North Carolina 27711
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RESEARCH REPORTING SERIES
Research reports of the Office of Research and Development,
U.S. Environmental Protection Agency, have been grouped into
five series. These five broad categories were established to
facilitate further development and application of environmental
technology. Elimination of traditional grouping was consciously
planned to foster technology transfer and a maximum interface in
related fields. The five series are:
1. Environmental Health Effects Research
2. Environmental Protection Technology
3. Ecological Research
4. Environmental Monitoring
5. Socioeconomic Environmental Studies
This report has been assigned to the ENVIRONMENTAL HEALTH EFFECTS
RESEARCH series. This series describes projects and studies relating
to the tolerances of man for unhealthful substances or conditions.
This work is generally assessed from a medical viewpoint, including
physiological or psychological studies. In addition to toxicology
and other medical specialities, study areas include biomedical in-
strumentation and health research techniques utilizing animals -
but always with intended application to human health measures.
This document is available to the public through the National
Technical Information Service, Springfield, Virginia 22161.
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EPA-600/l-76-010a
January 1976
ASSESSMENT OF TOXICITY OF AUTOMOTIVE METALLIC EMISSIONS. VOLUME I;
Assessment of Fuel Additives Emission Toxicity via Selected
Assays of Nucleic Acid and Protein Synthesis
By
David J. Holbrook, Jr., Ph.D.
Department of Biochemistry
School of Medicine
University of North. Carolina
Chapel Hill, North Carolina 27514
Contract No. 68-02-1205
Project Officer
Ms. Frances P. Duffield
Catalyst Research. Program Office
Health Effects Research Laboratory
Research Triangle Park, North Carolina 27711
U. S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF RESEARCH AND DEVELOPMENT
HEALTH EFFECTS RESEARCH LABORATORY
RESEARCH TRIANGLE PARK, NORTH CAROLINA 27711
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DISCLAIMER
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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ABSTRACT
Various parameters of toxicity have been studied for salts of manga-
nese, lead, palladium and platinum. Following intraperitoneal injection,
the acute toxicities (LD-50 doses, 14-day observation period, doses ex-
pressed in molar quantities) in decreasing order, were: PtCl, > Pt(SO, )'
4H20 (from B. F. Goldsmith) > PdCl^l^O >Pt(S04>2 '4H20 (from K and K Lab-
oratories), MnCl2'4H20>PdSO, > PtCl2> PbCl-. Following oral administration,
the acute toxicities (LD-50 doses), in decreasing order, were: PtCl, >
Pt(S04)-4H20> PdCl2'2H20, RuCl3> MnCl2.4H20 > PdS04> PtCl2>Pt02> PbO,
PdO, Mn02.
Following dietary (via drinking fluid) administration of soluble salts
of Pb2+ (PbCl2) or Pt + (PtCl4 or Pt(SC>4)2'4H20) , the highest concentra-
tions of metallic cations occurred in the kidney, intermediate levels in
the liver, and generally lower levels in the spleen, heart, testes, brain
and blood. In rats which survived for 14-days following the administration
of approximately the oral LD-50 and intraperitoneal LD-50, the kidneys con-
tained approximately 16 and 37 ugPt/g wet tissue, respectively, and the
livers contained approximately 2 and 34 ug Pt/g wet tissue, respectively.
Weights of five organs (liver, kidney, spleen, heart, testes) were
measured in rats which had been treated with various metallic salts in the
diet (either drinking fluid or solid feed). The organ weights were ex-
MnCl2'4H20 at a level of
pressed as a percentage of body weight. Rats which received.1.6 or 3.7
g/liter (8.3 or 18.6 mmoles/liter) for 90 days did not show statistically
significant changes in the weights of any organs. PbCl2> at a level of
1.0 g/liter (3,7 mmole/liter) for 30 or 90 days, consistently increased the
kidney weights of treated rats (17% and 23% above control in the 90-day
experiments). The use of saturated solutions of PdCl2'2H20 (8 days) or of
PdSO, (8 or 30 days) as the drinking fluid did not cause a consistent
ill
-------
change in any of the organ weights of the treated rats.
If PtCl, was added to the drinking fluid at 183 mg/liter (0.5 mmoles/
liter) for 30 or 90 days, or at 550 rag/liter (1.6 nmoles/liter) for 8 days,
no consistent changes were observed in the organ weights. If the concen-
tration-duration of PtCl, was increased to 550 mg/liter (1.6 mmoles/liter)
for 30 days or 825 mg/liter (2.4 ranoles/liter) for 9 days, the kidneys were
increased in weight by approximately 6% in each of four experiments and the
testes were increased by approximately 11% in each of four experiments; how-
ever, in each tissue in each individual experiments, the differences between
control and metal-treated animals showed statistically significant differences
(p<0.05) or trends (p<0.10) in only about one-half of the experiments.
After experimental rats were maintained on metal-containing diets for
approximately 8, 30 or 90 days, hepatic microsomes were isolated and the fol-
lowing parameters related to in vitro drug metabolism were measured: yield
of microsomal protein/g liver; in vitro activities of aniline hydroxylase
and aminopyrine demethylase; content of cytochromes P-450 and b5/mg micro-
somal protein. Treatment with MnCl2'4H20, 1.6 g/liter (8.3 mmoles/liter)
or 3.7 g/liter (18.6 mmoles/liter) for 90 days did not alter any of the
studied parameters of drug metabolism. The administeration of low levels
2+
of Pd (saturated solution of PdCl2'2H20 for 8 days or saturated solution
of PdSO, for 8 or 30 days) in the drinking fluid resulted in somewhat de-
creased activities of aniline hydroxylase and aminopyrine demethylase.
PtCl2 (saturated solution as drinking water) did not produce consistently
statistically different levels of aniline hydroxylase or aminopyrine
demethylase.
4+
A wide range of dose levels-duration of soluble salts of Pt did not
cause consistent changes in the levels of aniline hydroxylase or aminopyrine
iv
-------
demethylase in liver tissue; dosages and durations used included 0.5 mmoles/
liter (183 mg PtCl,/liter) for 30 and 90 days; 1.6 mmoles (550 mg PtCl,/
liter or 750 mg Pt(S04>2'4H20/liter for 8 days or for 30 days (PtCl^ only).
Work has been completed on the development of a rapid and convient
method for the analysis of ribosomal RNA in studies of RNA synthesis.
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CONTENTS
Page
Abstract ill
I. Introduction 1
II. Materials and Methods 2
III. .Lethal Dose Studies 3
IV. Metal Content of Various Tissues 11
V. Organ Weights 15
VI. Drug Metabolism In Vitro 19
VII. Studies on RNA Synthesis In Vivo 28
VIII.References 29
IX. Appendix: Data on Lethal Dose Levels 30
vi
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I. INTRODUCTION
A. Contract. These studies were conducted pursuant to contract num-
ber 68-02-1205 and project number DU-73-B439 with the Enviornmental Pro-
tection Agency.
B. Importance £f compounds. Lead salts are an emission product from
mobile (or automotive) emission Bourses due to the addition of tetraethyl
lead to gasoline. Because of known toxic properties of lead salts, it has
been proposed that alkyl manganese compounds be substituted as a fuel addi-
tive for tetraethyl lead. With the introduction of platinum and palladium
in the catalytic converters of 1975-model year vehicles, it is of concern
to determine the quantities of platinum and palladium metal and salts which
will be in emission products and the biological effects of these compounds
on mammalian tissues.
C. Studies undertaken. Consequently, experiments were undertaken
in this laboratory to study the effects of various metal salts on the fol-
lowing: acute lethal dose following oral and intraperitoneal administra-
tion, growth of animals receiving the salts in feed or drinking fluid, the
tissue concentration of some of the metals in various organs, the size of
selected organs, the in vitro activity of two representative microsomal
drug-metabolizing enzymes and the cytochrome P-450 and b5 concentrations
in liver, the activity in vitro of eucaryotic DNA polymerases, and RNA syn-
thesis in vivo.
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II. MATERIALS AND METHODS
All experimental studies were conducted With male Sprague-Dawley
rats. The animals were received at 3-3.5 weeks of age and were main-
tained for 1-1.5 weeks before use. The mean body weights were usually
100-110 g when the rats were used for the lethal-dose experiments or
started on the diets.
In the lethal dose experiments, the salts were administered orally
(via stomach tube) or intraperitoneally. The rats were observed through
a 14-day observation period. In the completed experiments, the LD-50
values were calculated by the method of Litchfield and Wilcoxon (1).
In the diet experiments, four rats were maintained per cage.
The metallic salt under study was dissolved in the drinking fluid.
Animals consumed feed and drinking fluid ad libitum. Analyses for
metals were performed on samples from three lots of feed (Purina
Laboratory Chow). The feed contained (mean + std. dev.): 56 + 5 rag
Mn/kg feed and 0.99 + 0.07 rag Pb/kg feed; the analyses of the three
lots for platinum were 0.09, < 0.02, and ^0.02 rag Pt/kg feed. Measure-
ments were made of the body weights of individual rats and feed and fluid
consumption per cage of four rats at 7-day intervals during the course of
each diet experiment.
At the termination of the dietary experiments, samples of liver
were used for the isolation of microsomes. Aniline hydroxylase was
measured by the method of Imai et al. (2), modified by the addition of
HgCl2 (3). Aminopyrine demethylase was measured by the formation of
formaldehyde (Nash reaction) (4).
-------
The analyses of the rat tissues for platinum, lead and manganese
were carried out by Yoakum, Stewart and Sterrett (5) of Stewart
Laboratories, Inc. by an emission spectrochemical method.
III. LETHAL DOSE STUDIES
The rats used in these studies usually had a mean body weight of
100-110 grams. The LD-50 values and the 95% confidence limits were
determined by the method of Litchfield and Wilcoxin (1).
-------
A. Oral administration. The summary of the lethel dose studies fol-
lowing oral administration are given in Table 1.
In terms of the LD-50 (dose lethal to 50% of the animals within the
14-day . observation period), the acute toxicities (expressed in molar quan-
tities), in decreasing order, were: PtCl >
, PtCl2>Pt02> PbO, PbCl2>Mh02, PdO.
4+
Thus, the two soluble Pt salts were found to be the most toxic salts
following oral administration. As anticipated, the poorly absorbed, insol-
uble salts, namely PbCl7, PtCl_, PbO, PtO,,, PdO and MnO», were the least
the latter Z Z
toxic. In ^ named cases, doses could not be increased sufficiently to
attain 50% lethality in the experimental rats and still maintain the volume
administered to 2% or less of the body weight.
In each of these cases, the LD-50 dose is greater than the 5,000 mg/kg
body weight which the National Institute for Occupational Safety and Health
uses as a criterion for inclusion as a toxic substance in the Toxic Substances
List. 1972 edition;^ ' thus, in terms of the acute LD-50, the salts PbCl9,
J ~ ~ £.
PbO, PdO, Pt02, and Mn02 would be considered "non-toxic" by this standard.
Also included in Table 1 are the LD-10 and LD-90, i.e., the doses
which cause the death of 10% and 90%, respectively, of the rats in the 14-
days after oral administration. In those cases where data are available,
the order of toxicities for the oral LD-10, "the minimal lethal dose", is
the same as that given above for the order of the LD-50 values.
The slopes of the toxicity curves can be compared by the ratio of the
LD-90 to LD-10 (Table 1). For example, MnCl2*4H20 is relative non-toxic
and the oral LD-10 dose is 6.3 mmoles/kg body weight. However, if the dose
was increased 1.4-fold to 9.0 mmoles/kg, 90% of the treated rats died. In
contrast, PtCl, was orally the most toxic of the salts tested; the ratio
of the LD-90 to LD-10 was approximately 5.0. The other soluble or partially-
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Table 1.
Lethal Doses Following Oral Administration
Unit
Compound
ptci4
Pt(S04)2'4H20
PdCl2-2H20
RuCl3
MnCV4H20
PdS04
(per kg body
weight)
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
LD-50
(957= confidence limits)
0.70 (0.51-0.96)
240 (171-320)
136 (99-188)
2.2 (1.57-3.1)
1010 (720-1400)
430 (310-600)
2.7 (2.2-3.4)
590 (470-730)
290 (240-360)
3.2 (2.4-4.0)
650 (500-830)
310 (240-400)
7.5 (7.0-8.1)
1490 (1380-1610)
410 (380-450)
>7.5
>1500
>790
LD-10
0.31
104
60
1.37
630
270
1.56
330
166
1.78
370
180
6.3
1260
350
--
--
--
LD-90
1.57
530
310
3.5
1620
690
4.8
1030
520
5.4
1130
550
9.0
1780
490
--
--
--
LD-90
LD-10
5.1
2.6
3.1
3.0
1.4
--
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Table 1 (continued)
Unit
(per kg body
weight)
LD-50
(95% confidence limits)
LD-10
LD-90
LD-90
LD-10
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
>8
>2,000
>1,400
>35
>8,000
>6,900
>45
>10,000
>9,300
»35
»9,600
>>7,200
» 82
>> 10,000
>> 8,700
(est.) 135
(est.) 12,000
(est.) 7,400
--
--
--
--
--
*30
<6,700
<6,200
21-35
5,800-9,600
4,300-7,200
--
--
--
(est.) 60
(est.) 5,200
(est.) 3,300
-------
soluble salts tested had intermediate LD-90 to LD-10 ratios of 2.5-3.1.
acute
If the^toxicities following oral administration are expressed as mg
of cation/kg of body weight, the LD-50 values were in the following de-
creasing order: PtCl, > PdCl2'2H20, RuCl3>MnCl2'4H20, Pt(S04)2'4H20 >
PdS04>PtCl2> Mn02, Pt02, PbCl2> PbO, PdO.
Likewise, if the LD-10 values are expressed in terms of mg of cation/
kg, the acute toxicities are in the following decreasing order: PtCl,>
PdCl2'2H20, RuCl3>Pt(S04)2-4H20>MnCl2'4H20»Mn02, PbCl2> PbO.
B. Intraperitoneal inlection. The summary of the lethal dose
studies following intraperitoneal injection are given in Table 2. Values
are given for the LD-50, LD-10 and LD-90 and each parameter is expressed
in terms of mmoles/kg body weight, mg salt/kg body weight, and mg cation/
kg of body weight.
-------
In terms of the LD-50, the acute toxicities (expressed in molar
quantities) in decreasing order, were: PtCl, >Pt(SO,) '4H-0 (Goldsmith) >
PdCl -2H20>Pt(S04)2-4H20 (K and K Laboratories), MnCl2'4H20> PdSO, >
PtCl2> PbCl2.
On a molar basis, the LD-10 are in the following decreasing order:
PtCl4> Pt(S04)2«4H20 (Goldsmith )>PdCl2*2H20>MnCl2-4H20, Pt(S04)2'4H20
(K and K)> PdS04> PbCl-. This was exactly the same order as the molar
LD-50 values.
The limiting dosages differentiating toxic and nontoxic substances
used for inclusion of a substance in the Toxic Substances List (7) is
2,000 mg/kg following an intraperitoneal injection in rats. According
to this standard, all of the tested compounds are "toxic" after in-
traperitoneal injection.
If the LD-50 following intraperitoneal injection is expressed in
terms of mg cation/kg body weight, the acute toxicities of the compounds
decrease in the following order: PtCl4> MnCl2'4H20> PdCl2'2H20, Pt(S04)2«
4H20 (Goldsmith)> Pt(S04)2-4H20 (K and K Lab), PdS04> PtCl2> PbCl2.
The intraperitoneal LD-10 values, expressed in mg cation/kg body
weight, are in essentially the same sequence as the eight most toxic
compounds listed for the intraperitoneal LD-50 values.
C. Duration £f survival. The rapidity of death in non-surviving
rats following oral administration varied widely. For example, rats
receiving approximately the oral LD-50 survived for ^1.0 day, PtCl,,
Pt(S04)2'4H20; 1-2.5 days, RuCl3; 3.5-4 days, MnCl^I^O; or 5 days,
PdCl7*2H70. For rats which received approximately the intraperitoneal
LD-50, the non-surviving rats lived for^l.O day, MnCl2'4H20; 3.0-3.5
days, Pt(S04)2'4H20, PdCl2'2H20; and 4.0-4.5 days,
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Table 2.
Lethal Doses Following Intraperitoneal Injection
Compound
PtCl,
4H20
(K and K Lab)
(Goldsmith)
PdCl2'2H20
PdSO,
PbCl,
PtCl,
Unit
(per kg body
weight)
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
mmoles
mg salt
mg cation
LD-50
(95% confidence limits)
0.11 (0.09-0.15)
38 (29-50)
22 (17-29)
0.68 (0.60-0.76)
310 (280-350)
132 (117-149)
0.70 (0.61-0.80)
138 (120-159)
38 (33-44)
0.3-0.4
138-184
59-78
0.57 (0.45-0.72)
121 (95-154)
60 (48-77)
1.42 (1.11-1.81)
290 (220-370)
151 (118-193)
8.5 (5.0-14.4)
240 (1400-4000)
1760 (1050-3000)
2.5 (1.58-4.0)
670 (420-1060)
490 (310-770)
LD-10 LD-90
LD-90
LD-10
0.56
260
110
0.56
111
31
0.2-0.3
92-138
39-59
0.39
84
42
(est.)
(est.)
(est.)
1.6
440
330
0.82
380
160
0.87
172
48
0.4-0.6
184-280
78-117
0.82
175
87
0.77 1.8
156 370
82 195
16.8
4700
3500
1.5
1.6
2.1
2.4
10
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In rats which survived for 7 to 14 days after administration of doses which
were two-thirds or less of the oral or intraperitoneal LD-50, significant lack
of weight gain was noted during days 0-7 in rats receiving most of the compounds.
However, weight gain, (expressed in grams) was approximately the same in the
the
treated and control rats during days 7-14 of the observation period.
Two samples of Pt(SO,K'AH-O were tested in the intraperitoneal lethal
dose studies; one was purchased from ICN-K and K Laboratories and a second
sample from D. F. Goldsmith Chemical and Metal Corp. The two samples differed
in their acute toxicities by approximately 2-fold. It has not been possible
to identify the cause of the differences.
Detailed data on the acute toxicities are given in the attached Appendix.
The data include the duration of survival by all animals and non-surviving
animals and the weight gain during the two weeks following metal administration
to the surviving animals. Also included are the plots of the probits versus
log dose which were used in the method of Litchfield and Wilcoxon.(l) to
evaluate the LD-10 and LD-90 values.
10
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IV. METAL CONTENT OF VARIOUS TISSUES
Analyses for lead, manganese and platinum were conducted by
Yoakum, Stewart and Sterrett (5). In a series of rats treated for
90-91 days, the control rats ingested approximately 0.15 g of
manganese (from the solid feed). The tissue concentration of Mn
was 1.4 and 1.0 pg Mn/g wet tissue in the liver and kidney, respect-
ively. In Mn-treated rats, which received 8.3 mM MnCl .4H 0 as the
drinking fluid and ingested approximately 2.3 g of Mn per rat
during the 90-91 day interval, the concentration of Mn was somewhat
increased, namely 2.8 and 1.6 /ig Mn/g of wet tissue in the liver and
kidney, respectively. The Mn concentration in spleen, heart, testes
and blood was not increased in the tissues of Mn-treated rats.
A second group of rats received 3.6 mM PbCl in the drinking
water for 90-91 days and ingested approximately 3 g of lead per rat
during the interval; control rats ingested < 0.01 g of Pb in the
solid feed during the same interval. Kidney showed a marked accumula-
tion of Pb (to 11.1 pg Pb/g of wet tissue) in the lead-treated rats;
in the same rats the concentration in liver was 1.2 jig Pb/g of wet
tissue. The corresponding levels in the control rats were approximately
11
-------
0.3 pg Pb/g of wet tissue in both kidney and liver. The other
tissues -- spleen, heart, testes and blood -- did not exhibit
appreciably higher levels of Pb in the Pb-treated rats.
Soluble Pt salts were included in the drinking fluid of rats
for 8-9 day intervals. The approximate total Pt intake (rag Pt per
rat) and data on the tissue concentration of Pt in various tissues
are presented in Table 3 . Although the Pt concentrations in tissues
of untreated control rats often attain levels measurable by the
technique used by Stewart Laboratories, Inc., the levels are low
and are generally less than 0.1 jag Pt/g of wet tissue. For the higher
4+
levels of Pt intake in the Pt-treated rats, the highest tissue
concentrations of Pt occurred in the kidney and ranged from 4.5-5
;ug Pt/g of wet tissue. High levels, ranging from 0.7-2.5 ;jg Pt/g,
also occurred in the liver. In contrast, brain showed only a very
low level of Pt which may reflect a contribution from the blood.
Separate experiments were conducted on the tissue concentrations of
Pt in rats which received a saturated solution of PtCl as the
2
drinking fluid for 30-31 days. In the PtCl«-treated rats, the mean
Pt concentration for liver, kidney and spleen were <0.08/jg Pt/g
of wet tissue.
In Table 4 are presented the Pt concentration of tissues
removed from rats which had survived for the 14-day observation
period in lethal-dose experiments. The doses of Pt(SO ) .4H 0
42 2
administered by both the oral and intraperitoneal routes were
approximately 90% of the LD-50 values by the respective routes.
12
-------
Table 3. Pt content of tissues of rats maintained on drinking
fluid containing Pt salts.
Diet (drinking
fluid)
Pt salt concn.
(mg Pt/lit)
Duration of diet
(days)
Total Pt intake < 0.01
Control Pt(S04)2.4H20
106
319
26
80
PtCl,
319
60
Tissue Tissue concentration of Pt
(ug Pt/g wet tissue)
Liver 4. 0.02
+ 0.02
Kidney < 0.23
+ 0.45
Spleen 4 0.08
+ 0.08
Heart 4. 0.02
+ 0.01
Testes < 0.014
+ 0.010
Brain
Blood 0.10
+ 0.13
0.07
(0.04-
0.09)
0.26
± °-05
0.02
(0.01-
0.01)
0.02
0.04
+ 0.05
-
0.05
0.85
(0.73-
0.97)
4.6
(4.5-
4.7)
0.13
0.25
-
0.015
+ 0.002
0.22
(0.09-
0.36)
2.2
(2.0-
2.5)
4.8
+ 0.5
0.24
-
-
-
0.23
(0.19-
0.27)
Control rats are those from diet experiments after approximately
8 or 30 days; 5-7 values for blood, spleen and heart, 13-16 values for
liver, kidney and testes.. +, standard deviation is given for means
with 4 values; ranges are indicated in parentheses for means of 2 values,
13
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Table 4. Pt concentration in rat tissues following the
administration of single high doses of Pt(SO, )2.4H20
Treatment
Route
Dose of Pt
(mg Pt/kg)
Tissue
Liver
Kidney
Spleen
Controls
Pt(S04)2.4H20
Oral Oral I. p.
382 113
Tissue Concentration of Pt
(ug Pt/g wet weight of tissue)
< 0.01
(0.004-0.006)
< 0.008
(0.004-0.004)
-£ 0.013
(0.007-0.011)
2.3 34
(1.2-3.5) (30-38)
16 37
(13-19) (28-46)
3.3 16
(2.3-4.2) (12-20)
Heart
Testes
Brain
Blood
0.02
0.011
(0.009-0.013)
0.01
0.008
0.8
0.5
3.0
1.2
(0.4-0.6) (0.9-1.5)
0.10 0.6
(0.07-0.14)(0.07-1.1)
3.3
1.0
Range of 4 values for control liver, kidney and spleen,
and range of 2-3 values of all other tissues are given
in parentheses. Control values are the mean values of
Pt concentration in 2 rats which received orally NaCl.
14
-------
During the two-week observation period, the rats gained weight at a
rate from one-third to three-fourths the rate of the control rats.
In the orally treated rats, the highest concentration of Ft occurred
in the kidney (approximately 16 ug Pt/g) and appreciable levels of
Pt also occurred in liver and spleen (range, 1-4 ug Pt/g of wet
tissue). In the intraperitoneably treated rats, the kidney, liver
and spleen showed very high levels of Pt in the range of 10-40 ug
of Pt/g of wet tissue.
In a comparable lethal Hose experiment, rats were treated
orally with a dose of MnCl .4H 0 equivalent to 100% of the oral
LD-50 value and the tissues were analyzed in surviving rats at the
end of the 14-day observation period. In contrast to the finding
with the Pt salt, the oral administration of a single, large but
nonlethal dose of MnCl. .4H 0 to rats did not result in the retention
2 2
after 14 days of excess concentrations of Mn in any of the tissues
analyzed (Table 5). Due to low levels of absorption and/or a
high capacity for excretion of the Mn, the tissue Mn levels of the
experimental rats were approximately equal to the levels found in
control rats.
4+
These studies show that in rats treated with soluble Pt
salts, appreciable levels of the metal can be found in the kidney,
liver and spleen. Further studies will be necessary to determine the
effects of the Pt and other metals on various biochemical reactions.
V. ORGAN WEIGHTS
Weights of five organs (liver, kidney, spleen, heart and testes)
were measured in rats which had been treated with various metallic
15
-------
Table 5. Mn concentrations in rat tissues following
the oral administration of a single large dose of
Treatment Controlsa
Dose of Mn 416
(rag Mn/kg)
Tissue Tissue Concentration of Mn
(ug Mn/g wet weight of tissue)
Liver
Kidney
Spleen
Heart
Testes
1.60
+ 0.87
0.75
+ 0.50
1.46
+ 1.99
0.55
+ 0.35
0.44
+ 0.35
1.9
(1.3-2.5)
1.3
(1.0-1.5)
1.3
(1.1-1.5)
0.7
0.5
(0.4-0.5)
Brain 0.3 0.03
Blood 0.86 0.4
-I- 0.44 (0.2-0.6)
Control values are from rats treated orally.with NaCl, and rats on
diet experiments for approximately 8 or 30 days. Means + standard
deviations are given for 6-7 samples of spleen, heart and blood and for
13-18 samples of liver, kidney and testes from control rats; ranges are
given in parentheses where two values are available from Mn-treated rats.
16
-------
salts in the diet (either drinking fluid or solid feed). The organ
weights were expressed as the percentage of the body weight. In the
discussions which follow, no consistent changes in organ weights occurred
unless specifically noted. In general, following treatment with most
metallic salts (at the doses used) weight changes were not observed in
liver, spleen and heart; several metals changed the kidney weight and a
few salts changed the testes weight.
MnCl2'4H20. MnCl2'4H20, in the drinking fluid at 8.3 or 18.6
mmoles/liter (1640 or 3690 mg/liter) for 90 days did not bring about
major changes in organ weights, although there was some enlargement (12,
13, and 8% above control; not statistically significant) in the spleen
of all three experimental groups of animals. None of the other organs
of rats receiving MnClj^H^O in the drinking fluid showed consistent
changes.
PbCl2. In rats which received 3.7 mmoles/liter (1022 mg/liter)
for 30 days or for 90 days, the size of the liver was increased (6-12%
above control; only one statistically significant) in three of the four
experiments. In contrast, the kidney size was increased in both the
30-day dietary experiments (7% and 6% above control; not statistically
significant) and the 90-day experiments (17% and 23% above control;
each statistically significant.) Consistent changes in organ weights
were not noted in the cases of spleen, heart and testes although spleen
showed increased size in two of four experiments. In a single experi-
ment, PbCl2 (8.3 mmoles/liter or 2300 mg/liter) for 30 days caused 18%
and 25% (neither statistically significant) increases in kidney and
spleen size, respectively. The increase in kidney size due to treat-
2+
ment with Pb is consistent with the data by Hirsch (6) and by others.
PdCl2 (anhydrous) and PdCL^^O. Although the addition of Pdd2
(anhydrous) to the feed (13.2 mmoles/kg or 2345 mg/kg; 30-days) caused
17
-------
changes in several organ weights, the pattern was not consistent to that
found in a second experiment. The use of a saturated solution of PdCl.
2H«0 as the drinking fluid (8-days) also did not cause a consistent change
in any of the organ weights.
PdSO,. The use of a saturated solution of PdSO, as the drinking fluid
for 8 or 30 days did not cause a consistent pattern of changes in the organ
weights of the experimental rats. In a single experiment in which solid
PdSO, was added for 30-days to the feed at a level of 5.9 mmoles/kg feed
(1.19 g salt/kg feed), no changes were observed in organ weights; each rat
received a mean of 3.8 mmoles of Pd salt (0.40 g of Pd) during the total
diet period.
PtCl,. When this salt was added to the drinking fluid at 0.5 mmoles/
liter (183 mg salt/liter) for 30 days or 90 days, or at 1.6 mmoles/liter
(550 mg salt/liter) for 8 days, no consistent changes were detected in the
weights of the five organs. If the concentration was increased to 1.6 mmoles/
liter (550 mg salt/liter) for 30 days or to 2.4 mmoles/liter (825 mg salt/
liter) for 9 days, the weight of the kidneys were increased by approximately
6% in each of 4 experiments but only two experiments gave p<0.1.
In addition, in rats treated at the higher levels (1.6 mmoles/liter
for 30 days or 2.4 mmoles/liter for 9 days), the testes were increased in
weight by approximately 11% in each of the 4 experiments but the difference
was statistically significant (p<0.05) in only one experiment.
Pt(SO, )2'4H_0. At a concentration of 1.6 mmoles/liter (750 mg salt/
liter) in the drinking fluid in 8-9 day experiments, increased kidney
weight was observed in only one of 2 experiments and increased testes
weight (approximately 8%) was found in each of the two experiments but none
of the differences were statistically significant.
18
-------
VI. DRUG METABOLISM IN VITRO
Selection o_f substrates. A wide variety of substrates are metabo-
lized by the NADPH-dependent mixed function oxidase system of hepatic
microsomes. The interaction of substrates with the cytochrome P-450
may produce one of several types of spectral changes. Various substrates
produce a type I spectrum which is characterized by a peak at 385-390 nm
and a trough at 419-425 nm; alternatively, other substrates produce a
type II spectrum (trough at: 390-405 nm; peak at 426-435 nm) upon inter-
action with cytochrome P-450 (8-10). In the current study, it was con-
sidered desirable to select: one representative substrate for each type
of interaction with cytochrome P-450. Consequently, in vitro drug
metabolism studies by isolated hepatic microsomes were conducted with
two substrates: the N-demethylase of aminopyrine, a type I substrate,
and the p-hydroxylase of aniline, a type II substrate\ (11, 12). How-
ever, the removal of one N-methyl group from aminopyrine yields a
metabolite which has a type II spectrum and which is N-demethylated (11).
The yield of microsomal protein (mg/g liver) found in this study is
appreciably greater than the yield of approximately 20 mg/g liver reported
in various publications for liver of fed rats. The difference can be at-
tributed predominantly to the fasting period of approximately 14 hours
(range 13-15 hours) used before the removal of rat tissues in these
studies. For example, McLean and Day (13), using male Wistar rats
weighing approximately 150-250 g, the levels of microsomal protein
(mg/g liver) were 25 ± s.d. 2 and 38± s.d. 10 in fed and 18-hour-fasted
rats, respectively; the latter value is in close agreement with the
yield of 40 ± s.d. 5 (15 groups) found in the livers of 14-hour-fasted,
Sprague-Dawley rats in this study. The control values obtained in these
experiments for the activities of aniline hydroxylase and aminopyrine
demethylase and for the content of cytochrome P-450 and cytochrome b5
are within the range of control values reported for male rats in various studies.
IP
-------
After experimental rats were maintained on metal-containing diets
for approximately 8, 30 or 90 days, hepatic microsomes were isolated
and parameters related to drug metabolism were measured. Data on pre-
sented in Table 6. Microsomal protein was expressed as mg microsomal
protein/g liver; aniline hydroxylase activity, nmoles p-aminophenol pro-
duced/mg microsomal protein/20 min; aminopyrine demethylase, nmoles form-
aldehyde produced/mg microsomal protein/ 10 min; cytochrome P-450 and
cytochrome b5, nmoles/mg microsomal protein.
Treatment with MnCl?-4H70 for approximately 90 days at either 8.3
mmoles (1.6 g salt)/liter or 18.6 mmoles (3.7 g salt)/liter did not ap-
(Table 6A),
pear to consistently change any of the parameters^ Treatment with PbCl?
at 3.7 mmoles (1.0 g salt)/liter for 90 days or 8.3 mmoles (2.3 g salt)/
liter for approximately 30 days did not affect recovery of microsomal
protein or the activity of aniline hydroxylase; for the other three para-
meters, it was not possible to detect a trend since data were obtained for
only one experiment each.
PdCl2'2H-0, a slightly soluble salt, was administered to rats as a
saturated solution as the drinking fluid for 8 days and a marked decrease
was noted in the activities of aniline hydroxylase and aminopyrine demethy-
(Table 6B),
However, caution must be exercised in the interpretation of this
trend. PdSO, , which is also a slightly soluble salt, was administered
as a saturated solution as the drinking fluid for 8 days or for approxi-
mately 30 days. In each of these experiments no trend of changes was ob-
(Table 6C),
served in the activities of the two enzymes » At this time, it is not pos-
sible to explain the apparent differences as a result of the administration
2+
the two slightly soluble salts of Pd
20
-------
Table 6A.
Dietary
concn. ,
nxnoles
/liter
or kg
(mg salt
Metallic
Salt
MnCl7-
4H20^
MnCl7-
4H2(T
Dietary
Group
58
Control ,
paired
59
Un-
treated
62
Control,
Paired
63
Mn-
treated
/liter
or ka)
8.29
(1640)
/liter
18.65
(3690)
/liter
Mean
metal
consumption
/rat/
Days on diet
diet period
93 2.6
mmoles
93 40.3
mmoles
(2.2g Mn)
88 2.8
mmoles
88 71.8
mmoles
(3.9g Mn)
Microsomal
protein
(rag/8
liver)
45.9
±2.8
50.3
±3.4
m
109%
36.4
±0.8
36.2
±2.1
ns
Aniline
hydroxylase
14.5
±1.4
13.9
±0.4
ns
22.5
±1.1
24.0
±4.0
ns
Aminopyrine
d erne thy lase
62.2
±5.0
56.5
±6.9
ns
73.7
±6.0
75.0
±8.1
ns
Cytochrome
P-450
0.870
±0.046
0.812
±0.088
ns
0.750
±0.098
0.710
±0.060
ns
Cytochrome
b5
0.369
±0.026
0.326
±0.028
*
887.
0.389
±0.039
0.395
±0.037
ns
-------
Table 6B.
Metallic
Salt
PdCl2'2H20
Dietary
concn. ,
mmoles Mean
/liter metal
or kg consumption
(mg salt /rat/
Dietary /liter Days on diet
Group or kg) diet period
36 8
Control ,
paired
37 (satd. 8
Pd- soln.)
treated
31 8
Control ,
paired
30 . (satd. 8
Pd- soln.)
treated
Microsomal
protein
(mg/g
liver
45.4
±2.6
51.6
±4.1
m
113%
43.6
±3.6
41.8
±1.4
ns
Aniline
hydroxylase
14.2
±2.4
11.0
±1.8
m
77%
14.5
±0.5
9.7
±2.6
*
67%
Aminopyrine Cytochrome
demethylase P-450
57 .8 run
±1.0
52.0 nm
±3.4
*
90%
67 .7 nm
±4.9
44.9 nm
±8.9
**
66%
Cytochrome
b5
nm
nm
nm
nm
-------
Table 6C.
Metallic
Salt
PdS04
*T
Dietary
Group
68
Control,
paired
69
Pd-
treated
Dietary
concn. ,
mmoles
/liter
or kg
(mg salt
/liter
or kg)
(satd.
soln.)
Mean
metal
consumption
/rat/
Days on diet
diet period
8
8
Microsomal
protein
(mg/g
liver
42.1
±1.9
42.5
±3.5
ns
Aniline
hydroxylase
15.0
±1.5
16,4
±2.1
ns
Aminopyrine Cytochrome
demethylase P-450
56 .3 nm
±3.6
49.4 nm
±6.9
ns
88%
Cytochrome
b5
nm
nm
PdSO,
ho
U)
60
Control,
paired
61
Pd-
treated
(satd.
soln.)
31
31
48.4
±2.3
19.1
±1.0
76.8
±8.4
0.587
±0.083
0.282
±0.017
47.2
±3.3
ns
17.3
±4.0
ns
90%
73.6
±15.0
ns
0.632
±0.049
ns
0.280
±0.023
ns
-------
2+
PtCl-, an "insoluble" salt of Pt , was administered to rats as a
saturated solution as the drinking fluid for approximately 30 days. No
consistent changes were observed in the activities of aniline hydroxyl-
ase and aminopyrine demethylase although the former enzyme showed de-
creased activity in two of the three experiments (Table 6D).
4+
The Pt salts, PtCl, and Pt(SO,)2'4H20, are very soluble and were
included in the drinking water. A wide range of dosages and durations
were used in the dietary experiments: 0.54 mmoles (183 mg PtCl,)/liter
for 30 and 90 days, and 1.6 mmoles (550 mg PtCl, or 750 mg Pt(SO,)'4H20/liter)
for 8 days and for 30 days (PtCl, only). Under none of the conditions
were consistent changes observed in the level of the drug metabolizing
enzymes (aniline hydroxylase or aminopyrine demethylase), or in the re-
covery of microsomal protein in the liver (Tables 6E, 6F).
It is apparent that the activities of the two representative drug
metabolizing enzymes and amounts of cytochromes P-450 and b5 are not
4+2+
extremely sensitive to lower levels of dietary Pt and Pd . In order
4+ 2+
to administer in the diet sufficient Pt and Pd to affect these para-
meters of drug metabolism, it will be essential or preferable to administer
the metallic salts in the solid feed rather than in the drinking fluid.
Obviously, the use of higher doses of the insoluble or slightly soluble
salts will require administration in the solid feed. Such studies are
currently in progress.
24
-------
Table 6D.
N>
Ui
Dietary
concn. ,
mmoles Mean
/liter metal
or kg consumption
(mg salt /rat/
Metallic Dietary /liter Days on diet
Salt Group or kg) diet period
PtCl2 7 31
Control ,
paired
8 (satd. 31
Pt- soln.)
treated
9 30
Control,
paired
11,12 (satd. 30
Pt- soln.)
treated
29 29
Control,
paired
28 (satd. 29
Pt- soln.)
treated
Microsomal
protein
(mg/g
liver
46.0
±3.2
43.9
±3.0
ns
55.0
±11.3
43.1
±3.0
m
787.
46.5
±3.1
44.6
±1.4
ns
Aniline
hydroxylase
16.3
±2.2
13.5
±1.4
m
83%
11.4
±1.3
8.3
±1.4
*
737,
16 .-0
±3.1
15.9
±2.3
ns
Aminopyrine Cytochrome
demethylase P-450
64 . 3 run
±20.7
63.9 nm
±17.6
ns
55.9
±14.3
81.6 nm
±13.7
*
1467.
80.1 nm
±16.9
73. 1 nm
±7.9
ns
Cytochronw
b5
nm
nm
nm
nm
nm
-------
Table 6E.
Metallic
Salt
ptci4
Dietary
Group
22
Control,
paired
23
Pt-
treated
Dietary
concn. ,
mmoles
/liter
or kg
(mg salt
/liter
or kg)
0.544
(183)
/liter
Mean
Metal
consumption
/rat/
Days on diet
diet period
30 0.00
mmoles
30 0.54
mmoles
(105 mg Pt)
Microsomal
protein
(mg/g
liver
41.1
±4.4
40.2
±3.1
ns
Aniline
hydroxylase
16.5
±3.5
15.5
±0.5
ns
Aminopyrine
demethylase
74.0
±16.1
76.3
±11.2
ns
Cytochrome Cytochrome
P-450 b5
nm
run
nm
20
Control ,
paired
21
Pt-
treated
0.544
(183)
/liter
29
29
0.00
mmoles
0.47
mmoles
(91 mg Pt)
41.7
±3.7
40.8
±3.2
ns
12.8
±3.0
13.6
±3.7
ns
64.8
±10.4
70.6
±22.0
ns
nm
nm
nm
nm
PtCl,
52
Control,
paired
53
Pt-
treated
0.544
(183)
/liter
91
91
0.00
mmoles
1.97
mmoles
(384 mg Pt)
49.1
±4.9
46.7
±3.9
ns
11.6
±1.2
14.3
±2.1
m
123%
55.7
±10.6
63.4
±3.4
ns
114%
0.790
±0.140
0.763
±0.128
ns
0.324
±0.044
0.348
±0.050
ns
-------
Table 6F.
Metallic
Salt
ptci4
Dietary
Group
46
Control,
paired
47
Pt-
treated
Dietary
concn. ,
tnmoles
/liter
or kg
(mg salt
/liter
or kg)
1.63
(550)
/liter
Days on
diet
29
29
Mean
metal
consumption
/rat/
diet
period
0.00
mmoles
1.63
mmoles
(317 mg Pt)
Microsomal
protein
(mg/g
liver
39.5
±3.6
42.1
±2.7
ns
Aniline
hydroxylase
27.3
±2.0
25.3
±1.6
ns
Aminopyrine
demethylase
105.7
±9.5
104.3
±9.4
ns
Cytochrome
P-450
0.478
±0.031
0.575
±0.107
ns
120%
Cytochrome
b5
0.326
±0.027
0.324
±0.011
ns
N>
38
Control,
paired
39
Pt-
treated
1.63
(550)
/liter
29
29
0.00
mmoles
1.34
mmoles
(261 mg Pt)
45.6
±1.2
42.6
±8.4
ns
25.4
±2.5
23.4
±2.6
ns
85.8
±6.6
86.2
±12.9
ns
nm
nm
nm
nm
27
Control,
paired
26
Pt-
treated
1.63
(550)
/liter
29
29
0.00
mmoles
1.27
tnmoles
(248 mg Pt)
47.9
±3.4
45.6
±2.6
ns
14.0
±1.3
15.0
±1.4
ns
76.6
±5.9
77.7
£7.7
ns
nm
nm
nm
nm
-------
VII. STUDIES ON RNA SYNTHESIS IN VIVO
Two major rRNA-containing peaks resulted from centrifugation of
resuspended hepatic polysomes on a sucrose gradient containing 0.1 M
NaCl and 0.001 M EDTA. The peaks had sedimentation coefficients of
approximately 27S and 43S. ^260^280 ratios °^ tne isoiated peaks in-
dicated a greater content of protein than in corresponding peaks pre-
pared from phenol-SDS or SDS-extracted rRNA. When compared with profiles
of phenol-SDS or SDS-extracted rRNA, material treated with only EDTA
and NaCl exhibited greater homogeniety and/or tighter conformation.
Increases in gradient NaCl concentration (0.25 M or 0.5 M) resulted in
additional peaks, all having S values greater than 27S. As indicated
by labeling studies for 1 h and 24 h, the EDTA-NaCl procedure was com-
parable to the SDS extraction procedure for observation of rRNA. Ribo-
somal RNA from RNase-treated polysomes (5 mg or 11 mg RNA/0.1 ug RNase)
showed greater structural integrity following EDTA-NaCl treatment than
following SDS extraction. When compared with diethylpyrocarbonate as
a means of improving resolution of rRNA following RNase degradation of
polysomes, the EDTA-NaCl procedure gave equally satisfactory results
with significantly greater convenience. A manuscript describing this
study has been prepared.
28
-------
VIII. REFERENCES
1. Litchfield, J. T., and Wilcoxon, F. A simplified method of evaluating
dose-effect experiments. J. Pharmacol. Exp. Ther. 96: 99 (1949).
2. Imai, Y., Ito, A., and Sato, R. Evidence for biochemically different
types of vesicles in the hepatic microsomal fraction. J. Biochem.
(Tokyo) 6£: 417 (1966).
3. Chhabra, R. S., Gram, T. E., and Fouts, J. R. A comparative study of
two procedures used in the determination of hepatic microsomal aniline
hydroxylation. Toxicol. Appl. Pharmacol. 22; 50 (1972).
4. Schenkman, J. B., Remmer, H., and Estabrook, R. W. Spectral studies
of drug interaction with hepatic microsomal cytochrome. Mol. Pharmacol.
3: 113 (1967).
5. Yoakum, A. M., Stewart, P. L., and Sterrett, J. E. Method development
and subsequent survey analysis of biological tissues for platinum, lead
and manganese content. Environ. Health Perspect. (1975)
6. Hirsch, G. H. Effect of chronic lead treatment on renal function.
Toxicol. Appl. Pharmacol. .25: 84 (1973).
7. Christensen, H. E. (ed.), National Institute for Occupational Safety and
Health Toxic Substances List. 1972 Edition (1972).
8. Schenkman, J. B., Remmer, H. and Estabrook, R. W. Mol. Pharmacol., 3_,
113-123 (1967).
9. Mannering, G. J., Sladek, N. E., Parli, C. J. and Shoeman, D. W. In
Microsomes and Drug Oxidations (J. R. Gillette e_t ajL., ed.). Academic
Press (1969).
10. Hayes, J. R. and Campbell, T. C. Biochem. Pharmacol., 2.3, 1721-1731 (1974).
11. Correia, M. A. and Mannering, G. J. Mol. Pharmacol., £, 470-485 (1973).
12. Flynn, E. J., Lynch, M. and Zannoni, V. G. Biochem. Pharmacol., 21. 2577-
2590 (1972).
13. McLean, A. E. M. and Day, P. A. Biochem. Pharmacol., 23, 1173-1180 (1974).
29
-------
X. APPENDIX: DATA ON LETHAL DOSE LEVELS
Values are the means (or weighted means) ± (upper) standard devia-
tions ± (lower) standard errors; the number of values is given in paren-
theses. NA, not applicable or not applied. In the column of "percentage
survival", the underlined values were used for the determination of the
LD-50, LD-10 and LD-90 doses by the method of Litchfield and Wilcoxon.
The standard deviations and standard error in the percentage survival
column are calculated from the products of the percentage survival and
the number of rats in each experiment in which that dose was tested.
Statistical analyses (student's t-test) were applied only to weekly weight
gains: ***, p<0.001; **, p<0.01; *, p<0.05; m, 0.05* p< 0.10; no mark-
ing used were p>0.10. Percentage changes of the weight gains are indi-
cated only where p<0.10 or where the percentage was less than 90% or
more than 110% of the control values.
a
"Corrected value from Table 1 of Litchfield and Wilconon for meas-
ured 0% or measured 100% survival.
"Expected value" was <0.01% or >99.99% by method of Litchfield and
Wilcoxon and not used in determination of lethal doses.
£
Value not used in determination of lethal doses.
Value approximated by assumption of 0.25 survivors/number tested
where 0% survival was measured, and 0.25 non-survivors/number tested
where 100% survival was measured.
30
-------
MnCl '4H20 / Intraperitoneal injection
Dose
(mg salt
/kg
body
weight) Expt.
Controls 21,25,
32,34
400 2,21,
25
200 2,21,
25
141.4 32,34
100 2,21,
25
50 21,25
Number
survivors
/number Percentage
tested survival
10/10 NA
100 .0
±0.0
±0.0
0/11 NA.b
o.'o
±0.0
±0.0
0/10 2.5d
0.0
±0.0
±0.0
4/12 33.3
33.3
±0.0
±0.0
11/11 97. 7d
100.0
±0.0
±0.0
11/11 NAb
lOO.'O
±0.0
+0.0
Duration of
survival (hr)
all
animals
336
±0
±0
(10)
2
±2
±1
(ID
8
±4
±1
(10)
120
±159
±46
(12)
336
±0
±0
(ID
336
±0
±0
(ID
non-
survivors
NA
2
±2
±1
(11)
8
±4
±1
(10)
12
±3
±1
(8)
NA
NA
Body
weight
(g) on
day 0
117
±8
±3
(10)
112
±12
±4
(11)
112
±9
±3
(10)
114
±13
±4
(12)
112
±8
±2
(11)
112
±8
±2
(11)
WL-l«h
days
0-7
63
±11
±3
(10)
NA
NA
18
±34
±17
(4)
*
29%
33
±19
±6
(9)
***
52%
45
±11
±3
(11)
**
71%
t K'-'in
days
0-14
110
±13
±4
(10)
NA
NA .
56
±57
±28
(4)
m
51%
81
±21
±7
(9)
**
74%
96
±15
±4
(ID
*
87%
(K)
days
7-14
48
±9
±3
(10)
NA
NA
38
±25
±12
(4)
ns
79%
48
±12
±4
(9)
ns
51
±12
±4
(11)
ns
31
-------
MnCl_.4H70 / Intraperitoneal injection
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2.2
2.3
.10
Dose (log)(mg salt/kg body weight)
32
-------
/ Oral administration
Dose
(mg salt
/kg
body
weight) Expt.
Controls 5,8,
14,23
2500 1,5,
14,23
2000 5,8,
14,23
1750 8,14,
23
1500 5,8,
14,23
1250 1,8,
14,23
1000 5,8,
14,23
Number
survivors
/number Percentage
tested survival
14/14 NA
1/12 NA°
8.3
±15.1
±4.3
0/12 l.O3
0.0
±0.0
±0.0
2/12 16.7
16.7
±14.7
±4.2
6/12 50.0
50.0
±28.9
±8.3
10/11 90.9
90.9
±12.6
±3.8
12/12 99. 9a
100.0
±0.0
±0.0
Duration of
survival (hr)
all non-
animals survivors
336
±0
±0
(14)
40
±94
±27
(12)
22
±23
±7
(12)
106
±114
±33
(12)
213
±133
±38
(12)
307
±97
±29
(11)
336
±0
±0
(12)
NA
13
±11
±3
(11)
22
±23
±7
(12)
60
±43
±14
(10)
90
±48
±19
(6)
NA
NA
Body
weight
(g) on
day 0
107
±10
±3
(14)
109
±9
±2
(12)
103
±7
±2
(12)
109
±8
±2
(12)
103
±14
±4
(12)
111
±10
±3
(11)
105
±7
±2
(12)
Weight gain (g)
days days days
0-7 0-14 7-14
51
±10
±3
(14)
NA
NA
NA
22
±22
±9
(6)
**
43%
39
±25
±9
(7)
ns
76%
40
±19
±5
(12)
m
78%
100
±31
±8
(14)
NA
NA
NA
84
±26
±10
(6)
ns
84%
103
±26
±10
(7)
ns
99
±20
±6
(12)
ns
49
±23
±6
(14)
NA
NA
NA
62
±9
±3
(6)
m
127%
64
±10
±4
(7)
m
131%
59
±14
±4
(12)
ns
120%
33
-------
MnCl_.4H20 / Oral administration
> "
3
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01
00
rt
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-------
MnO? / Oral administration
Dose
(mg salt
/kg
body
weight) Expt.
Controls 33,38
10,000 33,38
6,666 38
Number
survivors
/number Percentage
tested survival
6/6 MA
100.0
±0.0
±0.0
6/10 60.0
60.0
±0.0
±0.0
9/11 81.8
81.8
Duration of .
survival (hr) ^
all
animals
336
±0
±0
206
±168
±53
(10)
285
±113
±34
(11)
non- (g) on
survivors day 0
NA 99
±9
±4
(6)
11 103
±8 ±10
±4 ±3
(4) (10)
NA 99
±10
±3
(11)
Weight gain (g)
days
0-7
68
±11
±4
(6)
58
±15
±6
(6)
ns
85%
51
±18
±6
(9)
*
75%
days
0-14
128
±12
±5
(6)
120
±16
±6
(6)
ns
102
±44
±15
(9)
ns
80%
days
7-14
60
±7
±3
(6)
62
±6
±2
(6)
ns
50
±28
±9
(9)
ns
83%
35
-------
MnCL / Oral administration
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Dose (log)(mg salt/kg body weight)
36
-------
PbCl, / Intraperitoneal injection
Dose
(mg salt
/kg
body
weight)
Controls
2083
1250
750
450
270
Expt.
12,21,
28,41,
48
28,41,
48
21,28,
41,48
12,21,
28,41,
48
12,21,
28,41
21,28
Number
survivors
/number Percentage
tested survival
13/13 NA
100.0
±0.0
±0.0
9/16 56.2
56.2
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±3.6
11/16 68.8
68.8
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±6.8
12/16 75.0
75.0
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±7.5
11/12 91.7
91.7
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±3.6
7/7 NA
100.0
±0.0
±0.0
Duration of
survival (hr)
all
animals
336
±0
±0
(13)
227
±130 .
±33
(16)
257
±126
±31
(16)
304
±74
±18
(16)
325
±39
±11
(12)
336
±0
±0
(7)
non-
survivors
NA
87
±39
±15
(7)
82
±63
±28
(5)
206
±101
±50
(4)
NA
NA
Body
weight
(g) on
day 0
111
±9
±3
(13)
101
±6
±2
(16)
110
±9
±2
(16)
106
±9
±2
(16)
111
±6
±2
(12)
116
±4
±2
(7)
Weight gain
days
0-7
55
±19
±5
(13)
30
±24
±8
(10)
*
55%
31
±18
±5
(12)
**
56%
27
±18
±5
(15)
fCyCK
49%
29
±25
±7
(12)
**
53%
29
±16
±6
(7)
**
53%
days
0-14
125
±17
±5
(10)
76
±36
±12
(9)
**
61%
63
±37
±11
(11)
***
50%
57
±37
±11
(11)
"K'ffff
46%
54
±44
±16
(8)
***
43%
74
±30
±11
(7)
**
59%
(g)
days
7-14
62
±11
±4
(10)
41
±21
±7
(9)
*
66%
31
±30
±9
(11)
**
50%
32
±16
±5
(11)
***
52%
28
±24
±9
(8)
**
45%
44
±16
±6
(7)
*
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37
-------
FbCl. / Intraperitoneal Injection
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CHAMPION LINE NO. 674 - PROBABILITY X QO
Dose (log) (ing salt/kg body weight)
38
-------
PbCl, / Oral administration
Dose
(mg salt
/kg
body
weight) Expt.
Controls 51
9645 51
5787 51
Number
survivors
/number
tested
2/2
9/10
9/10
Duration of .
survival (hr) ^
v ' weight
Percentage
survival
NA
100.0
90.0
90.0
1
90.0
90.0
all
animals
NA
305
±97
±31
(10)
325
±35
±11
(10)
non- (g) on
survivors day 0
NA NA
NA 118
±12
±4
(10)
NA 116
±10
±3
(10)
Weight gain (g)
days
0-7
NA
44
±13
±4
(9)
54
±27
±9
(10)
days
0-14
NA
107
±12
±4
(9)
115
±31
±10
(9)
days
7-14
NA
63
±5
±2
(9)
55
±14
±5
(9)
39
-------
PbO / Oral administration
Dose
(mg salt
/kg
body
weight) Expt.
Controls 33,39
10,000 33,39
6,666 39
Number
survivors
/number Percentage
tested survival
6/6 NA
100.0
±0.0
±0.0
7/11 63.6
63.6
±15.7
±4.7
6/9 66.7
66.7
Duration of
survival (hr)
all
animals
336
±0
±0
(6)
217
±165
±50
(11)
228
±161
±54
(9)
non-
survivors
NA
9
±4
±2
(4)
13
±4
±2
(3)
Body
weight
(g) on
day 0
105
±6
±2
(6)
103
±10
±3
(11)
97
±4
±1
(9)
Weight gain (g)
days
0-7
64
±15
±6
(6)
32
±19
±7
(7)
**
50%
44
±12
±5
(6)
*
697»
days
0-14
106
±33
±13
(6)
96
±15
±6
(7)
ns
105
±14
±6
(6)
ns
days
7-14
43
±18
±7
(6)
64
±8
±3
(7)
*
149%
61
±12
±5
(6)
m
142%
40
-------
PbO / Oral administration
0)
o
.. h_.. o
CHAMPION LINE
3.8
NO. 674 - PROBABILITY X 9O
4.0
Dose (log)(mg salt/kg body weight)
41
-------
PdCl2'2H20 / Intraperitoneal Injection
Dose
(mg salt
/kg
body
weight) Expt.
Controls 17,24,
37
279 17,24
186 17,24,
37
124 17,24,
37
82.7 24,37
55.1 24,37
Number
survivors
/number Percentage
tested survival
8/8 NA
100.0
±0.0
±0.0
0/5 NA
0.0
±0.0
±0.0
1/11 9.1
9.1
±8.7
±2.6
4/11 36.4
36.4
±19.5
±5.9
11/11 92. 6a
100.0
±0.0
±0.0
11/11 NA
100.0
±0.0
±0.0
Duration of
survival (hr)
all
animals
336
±0
±0
(8)
17
±6
±3
(5)
59
±94
±28
(11)
172
±138
±42
(11)
336
±0
.±0
(11)
336
±0
±0
(11)
non-
survivors
NA
17
±6
±3
(5)
32
±22
±7
(10)
78
±58
±22
(7)
336
±0
±0
(11)
336
±0
±0
(11)
Body
weight
(g) on
day 0
97
±7
±2
(8)
95
±6
±2
(5)
96
±5
±2
(ID
101
±4
±1
(11)
98
±8
±2
(11)
97
±5
±2
(11)
Weight gain (g)
days
0-7
60
±8
±3
(8)
NA
NA
*
33
±19
±9
(5)
*
55%
59
±8
±2
(11)
ns
57
±11
±3
(11)
ns
days
0-14
118
±15
±5
(8)
NA
NA
86
±8
±4
(4)
***
73%
119
±11
±3
(11)
ns
116
±21
±6
(ID
ns
days
7-14
57
±9
±3
(8)
NA
NA
46
±6
±3
(4)
*
81%
60
±8
±2
(11)
ns
59
±13
±4
(ID
ns
42
-------
PdCl2.2H-,0 / Intraperitoneal injection
OJ
c
0)
P.. :::.
1.9 2.0
CHAMPION UNIi NO. 074 - PROS AIM I ITY X 9O
2.1
2.2
2.3
2.4
Dose (log)(mg salt/kg body weight)
43
-------
PdCl2-2H20 / Oral administration
Dose
(mg salt
/kg
body
weight) Expt.
Controls 15,16,
18,25
1410 16,18,
25
940 15,16,
18,25
627 15,16,
18,25
418 15,16,
18,25
279 15,16
18,25
186 15
Number
survivors
/number Percentage
tested survival
9/9 NA
100.0
±0.0
±0.0
0/11 2.2a
0.0
±0.0
±0.0
2/11 18.2
18.2
±31.2
±9.4
5/11 45.5
45.5
±32.6
±9.8
7/10 70.0
70.0
±29.2
±9.2
11/11 96. 2S
100.0
±0.0
±0.0
3/3 NA
100.0
Duration of
survival (hr)
all
animals
336
±0
±0
(9)
58
±57
±17
(ID
125
±108
±32
(11)
213
±130
±39
(ID
276
±100
±32
(10)
336
±0
±0
(11)
336
±0
±0
(3)
non-
survivors
NA
58
±57
±17
(ID
79
±31
±10
(9)
110
±76
±31
(6)
136
±52
±30
(3)
NA
NA
Body
weight
(g) on
day 0
108
±13
±4
(9)
109
±11
±3
(11)
102
±11
±3
(11)
106
±8
' ±3
(11)
103
±8
±3
(10)
107
±11
±3
(11)
98
±5
±3
(3)
Weight gain
days
0-7
57
±8
±3
(9)
NA
NA
14
±23
±9
(6)
***
257,
11
±20
±7
(8)
***
357o
37
±28
±8
(11)
*
65%
49
±10
±6
(3)
ns
86%
days
0-14
115
±13
±4
(9)
NA
NA
69
±31
±14
(5)
**
60%
57
±40
±15
(7)
**
50%
89
±30
±9
(11)
*
77%
106
±14
±8
(3)
ns
(8)
days
7-14
59
±9
±3
(9)
NA
NA
51
±13
±6
(5)
ns
86%
42
±23
±9
(7)
m
71%
52
±9
±3
(11)
ns
887,
57
±4
±3
(3)
ns
44
-------
PdCl2.2H20 / Oral adminiatration
&
;>f
-fl
iB
±it
5 s
H
B o
3 §
ca
-------
PdO / Oral administration
Dose
(mg salt
/kg
body
weight) Expt.
Controls 33,54
10,000 33,54
Number
survivors
/number
tested
5/5
6/6
Percentage
survival
NA
100.0
±0.0
±0.0
100.0
100.0
±0.0
±0.0
Duration
survival
- all
of
(hr)
non-
animals survivors
336
±0
±0
<5>
336
±0
±0
(5)
NA
NA
Body
weight
(8) on
day 0
108
±7
±3
(5)
103
±13
±5
(6)
ns
Weight gain
days
0-7
69
±12
±5
(5)
50
±18
±7
(6)
m
72%
days
0-14
130
±10
±4
(5)
98
±35
±14
(6)
m
75%
(8)
days
7-14
62
±7
±3
(5)
48
±17
±7
(6)
ns
77%
46
-------
PdSO, / Intraperitoneal injection
Dose
(mg salt Number
/kg survivors
body /number Percentage
weight) Expt. tested survival
Controls 22,40, 9/9 NA
47 100.0
±0.0
±0.0
450 40 0/4 NA
0.0
300 40,47 5/10 50.0
50.0
±21.5
±6.8
245 47 5/9 55.6
55.6
200 22,40, 8/10 80.0
47 80.0
±17.2
±5.4
133.3 22,40 4/4 NA
100.0
±0.0
±0.0
Duration of
survival (hr)
all
animals
336
±0
±0
(9)
52
±52
±26
(4)
219
±127
±40
(10)
245
±114
±38
(9)
294
±88
±28
(10)
336
±0
±0
(4)
non-
survivors
NA
52
±52
±26
(4)
101
±40
±18
(5)
132
±60
±30
(4)
NA
NA
Body
weight
(g) on
day 0
107
±4
±2
(9)
101
±6
;±3
(4)
103
±7
±2
(10)
110
±10
±3
(9)
105
±6
±2
(10)
104
±13
±6
(4)
Weight gain (g)
days
0-7
62
±10
±3
(9)
NA
44
±10
±4
(5)
**
717.
38
±16
±7
(6)
**
61%
31
±16
±6
(8)
***
50%
34
±21
±10
(4)
*
55%
days
0-14
126
±19
±6
(9)
NA
89
±39
±17
(5)
m
71%
86
±33
±15
(5)
*
68%
75
±34
±12
(8)
**
60%
90
±33
±17
(4)
m
71%
days
7-14
64
±12
±4
(9)
NA
45
±30
±13
(5)
ns
70%
46
±18
±8
(5)
m
72%
44
±25
±9
(8)
m
69%
56
±14
±7
(4)
ns
88%
47
-------
/ Intraperitoneal injection
3
OJ
-------
Ptd2 / Intraperitoneal injection
Dose
(mg salt
/kg
body
weight) Expt.
Controls 18,52
53
1111 52,53
741 52,53
578 52,53
450 18,53
250 18,53
Number
survivors
/number Percentage
tested survival
6/6 NA
100.0
±0.0
±0.0
1/4 NA
25.0
±16.6
±8.3
3/9 33.3
33.3
±18.9
±6.3
3/6 50.0
50.0
±38.7
±15.8
9/10 90.0
90.0
±5.3
±1.7
8/10 80.0
80.0
±10.5
±3.3
Duration of
survival (hr)
all
animals
336
±0
±0
(6)
89
±165
±82
(4)
151
±167
±56
(9)
175
±177
±72
(6)
307
±91
±29
(10)
329
±18
±6
(10)
non-
survivors
NA
7
±6
±3
(3)
58
±118
±48
(6)
13
±21
±12
(3)
NA
NA
Body
weight
(g) on
day 0
104
±24
±10
(6)
108
±21
±11
(4)
112
±12
±4
(9)
107
±8
±3
(6)
117
±12
±4
(10)
117
±16
±5
(10)
Weight gain (g)
days
0-7
66
±18
±7
(6)
NA
6
±23
±12
(4)
**
97,
16
±12
±7
(3)
**
24%
39
±16
±5
(9)
**
59%
53
±11
±3
(10)
ns
80%
days
0-14
124
±17
±7
(6)
NA
53
±52
±30
(3)
m
43%
58
±27
±15
(3)
**
47%
100
±25
±8
(9)
*
81%
107
±21
±7
(8)
ns
86%
days
7-14
57
±4
±2
(6)
NA
46
±24
±14
(3)
ns
81%
43
±15
±9
(3)
ns
75%
61
±12
±4
(9)
ns
56
±12
±4
(8)
ns
49
-------
PtCl, / Intraperltoneal Injection
Dose
(mg salt
/kg
body
weight) Expt.
Controls 42,45,
50
108 42,45
64.8 42,45,
50
38.9 45,50
23.3 45,50
14.0 45
Number
survivors
/number Percentage
tested survival
6/6 NA
100.0
±0.0
±0.0
0/6 NA
0.0
±0.0
±0.0
1/10 10.00
10.0
±12.2
±4.1
2/10 20.00
20.0
±0.0
±0.0
10/10 97. 5d
100.0
±0.0
±0.0
5/5 NA
100.0
±0.0
±0.0
Duration of
survival (hr)
all
animals
336
±0
±0
(6)
0
±0
±0
(6)
34
±106
±34
(10)
107
±123
±39
(10)
336
±0
±0
(10)
336
±0
±0
(10)
non-
survivors
NA
0
±0
±0
(6)
1
±0
±0
(9)
50
±29
±10
(8)
NA
NA
Body
weight
(g) on
day 0
95
±12
±5
(6)
105
±7
±3
(6)
101
±9
±3
(10)
100
±6
±2
(10)
100
±9
±3
(10)
102
±8
±3
(5)
Weight gain
days
0-7
61
±6
±3
(6)
NA
NA
NA
22
±22
±7
(10)
***
57
±11
±5
(5)
ns
days
0-14
120
±11
±5
(6)
NA
NA
NA
87
±26
±8
(10)
**
73%
96
±6
±3
(5)
**
80%
(8)
days
7-14
59
±7
±3
(6)
NA
NA
NA
62
±14
±4
(10)
ns
38
±11
±5
(5)
**
647,
50
-------
PtCl, / Oral administration
Dose
(mg salt
/kg
body
weight) Expt.
Controls 31,43,
46
660 29
440 29,46
293,3 31,43,
46
195.6 31,43,
130.4 31,43,
46
36.9 31,43
Number
survivors
/number Percentage
tested survival
7/8 NA
87.5
±23.1
±8.2
0/6 NA
0.0
1/9 11.11
11.1
±10.5
±3.5
4/10 40.0
40.0
±22.0
±6.9
8/10 80.0
80.0
±18.3
±5.8
7/10 70.0
70.0
±31.6
±10.0
2/3 NA
66.7
Duration of
survival (hr)
all
animals
310
±72
±26
(8)
0
±0
±0
(6)
38
±112
±37
(9)
140
±170
±53
(10)
270
±138
±44
(10)
236
±162
±51
(10)
240
±166
±96
(3)
non-
survivors
NA
0
±0
±0
(6)
1
±3
±1
(8)
9
±10
±4
(6)
NA
1
±0
±0
(3)
NA
Body
weight
(g) on
day 0
100
±15
±5
(8)
98
±9
±4
(6)
100
±6
±2
(9)
109
±10
±3
(10)
103
±12
±4
(10)
102
±9
±3
(10)
106
±15
±9
(3)
Weight gain (g)
days
0-7
65
±18
±7
(7)
NA
NA
16
±38
±19
(4)
*
25%
NA
57
±13
±5
(7)
NA
days
0-14
108
±35
±13
(7)
NA
NA
81
±34
±17
(4)
ns
75%
NA
115
±22
±8
(7)
NA
days
7-14
44
±29
±11
(7)
NA
NA
65
±14
±7
(4)
ns
148%
NA
58
±22
±8
(7).
NA
51
-------
PtCl, / Oral administration
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CHAMPION LINE NO 674 - PROBABILITY
Dose
X 90
(log)(mg salt/kg body weight)
52
-------
PtO« / Oral administration
Dose
(mg salt
/kg
body
weight) Expt.
Controls 9,20,
33,51
8000 9,33,
51,52
4444 20,52
Number
survivors
/number Percentage
tested survival
9/9 NA
100.0
±0.0
±0.0
5/7 71.4
71.4
±48.8
±18.4
5/6 83.3
83.3
±12.9
±5.3
Duration of ,
survival (hr) . ^
v weight
all
animals
336
±0
±0
(9)
253
±143
±54
(7)
326
±25
±10
(6)
non- (g) on
survivors day 0
NA 106
±16
±5
(9)
NA 118
±11
±4
(7)
NA 115
±21
±9
Weight gain
days
0-7
66
±20
±7
(9)
68
±15
±7
(5)
ns
62
±18
±7
(6)
ns
days
0-14
117
±35
±12
(9)
114
±17
±8
(5)
ns
106
±36
±16
(5)
ns
90%
(g)
days
7-14
ril
±18
±6
(9)
47
±10
±4
(5)
ns
37
±28
±13
(5)
ns
7370
53
-------
Pt(SO,)2'4H20 / Intraperitoneal Injection
Dose
(mg salt
/kg
body
weight)
Controls
600
400
326.6
267
178
Expt.
14,22,
24,32,
14,22,
24
14,22,
24
32, 3'4
14,22,
24
14,22,
24
Number
survivors
/number Percentage
tested survival
16/16 NA
100.0
±0.0
±0.0
0/10 NAb
0.0
±0.0
±0.0
1/11 9.1
9.1
±20.2
±6.1
2/12 16.7
16.7
±0.0
±0.0
10/11 90.9"
90.9
±8.7
±2.6
11/11 NAb
100.0
±0.0
±0.0
Duration of
survival (hr)
all
animals
336
±0
±0
(16)
28
±16
±5
(10)
67
±92
±28
(11)
117
±106
±31
(12)
308
±93
±28
(11)
336
±0
±0
(11)
non-
survivors
NA
28
±16
±5.
(10)
41
±23
±7
(10)
73
±31
±10
(10)
NA
NA
Body
weight
(g) on
day 0
106
±10
±3
(16)
100
±4
±1
(10)
102
±11
±3
(11)
108
±11
±3
(12)
101
±8
±2
(11)
104
±11
±3
(11)
Weight gain (g)
days
0-7
59
±7
±2
(16)
NA
NA
NA
39
±17
±5
(10)
**
66%
36
±14
±4
(11)
***
61%
days
0-14
114
±14
±4
(16)
NA
NA
NA
96
±26
±8
(10)
m
84%
94
±32
±10
(11)
m
82%
days
7-14
55
±13
±3
(16)
NA
NA
NA
56
±11
±3
(10)
ns
58
±21
±6
(11)
ns
4+
Ft salt obtained from ICN/K and K Laboratories
54
-------
Pt(SO,)2.4H-0 / InfcrapiBsritoneal injaction
T
3
0)
0)
60
CO
4-1
V
a
1 M
2.4 2.5
CHAMPION LINE NO. 674 - PROBABILITY X 9O
2.6
Dose (log)(mg salt/kg body weight)
-------
pt(so4).
4H20 (Goldsmith) / Intraperitoneal injection
Dose
(mg salt
/kg
body
weight) Expt.
Controls 56,57
400 55
267 55,56
57
119 56,57
52.7 56
Number
survivors
/number Percentage
tested survival
4/4 NA
100.0
±0.0
±0.0
1/3 NA
33.3
0/7 3.6d
0.0
±0.0
±0.0
6/7 85.7
85.7
±17.8
±6.7
4/4 NA
100.0
Duration of
survival (hr)
all
animals
336
±0
±0
114
±167
±96
(3)
76
±13
±5
(7)
307
±78
±29
(7)
336
±0
±0
(4)
non-
survivors
NA
NA
76
±13
±5
(7)
NA
NA
Body
weight
(g) on
day 0
105
±18
±9
(4)
121
±16
±9
(3)
112
±13
±5
(7)
114
±7
±3
(6)
120
±18
±9
(4)
Weight gain (g)
days
0-7
52
±12
±6
(4)
NA
NA
8
±31
±13
(6)
*
16%
56
±16
±8
(4)
ns
days
0-14
106
±15
±8
(4)
NA
NA
56
±34
±14
(6)
*
52%
117
±24
±12
(4)
ns
days
7-14
54
±5
±2
(4)
NA
NA
47
±19
±8
(6)
ns
88%
61
±9
±4
(4)
ns
113%
4+
Ft salt obtained from D. F. Goldsmith Chemical and Metal Corp.
56
-------
/ Oral administration
Dose
(mg salt
/kg
body
we igh t )
Controls
2025
1350
900
600
400
Expt.
14,16,
23,25
14,16,
23,25
14,16,
23,25
14,16,
23,25
14,16,
23,25
14,16,
23
Number
survivors
/number Percentage
tested survival
11/11 NA
100.0
±0.0
±0.0
1/10 10.0
10.0
±31.6
±10.0
1/10 10.0
10.0
±31.6
±10.0
5/10 50.0
50.0
±45.1
±14.3
9/10 90.0
90.0
±16.1
±5.1
7/7 99. 83
100.0
±0.0
±0.0
Duration of
survival (hr)
all
animals
336
±0
±0
(ID
46
±104
±33
(10)
43
±103
±33
(10)
172
±173
±55
(10)
307
±92
±29
(10)
336
±0
±0
(7)
non-
survivors
NA
13
±24
±8
(9)
11
±10
±3
(9)
8
±4
±2
(5)
NA
NA
Body
weight
(g) on
day 0
106
±12
±3
(ID
109
±13
±4
(10)
109
±11
±3
(10)
109
±10
±3
(10)
108
±11
±3
(10)
109
±7
±3
(7)
Weight gain
days
0-7
50
±8
±2
(11)
NA
NA
27
±33
±15
(5)
ns
54%
37
±25
±8
(9)
ns
74%
52
±15
±6
(7)
ns
days
0-14
108
±16
±5
(11)
NA
NA
78
±48
±21
(5)
ns
72%
106
±17
±6
(9)
ns
116
±20
±8
(7)
ns
(8)
days
7-14
58
±11
±3
(ID
NA
NA
51
±19
±8
(5)
ns
88%
70
±29
±10
(9)
ns
64
±6
±2
(7)
ns
110%
57
-------
Pt(SO^)2.4H20 / Oral administration
t-
ft
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ii:
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00
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uit
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rr
tit
i\
iR
s 2.6 2.7 2.8
CHAMPION LINE NO. 674 - PROBABILITY X 90
2.9
3.0
3.1
3.2
3.3
Dose (log)(mg salt/kg body weight)
58
-------
RuCl, / Oral administration
Dose
(mg salt
/kg
body
weight) Expt.
Controls 4,7,
35,36
1350 35,36
900 35,36
600 7,35,
36
400 4,7,
35,36
200 4
Number
survivors
/number Percentage
tested survival
8/9 NA
88.9
±22.0
±7.3
0/11 4.8a
0.0
±0.0
±0.0
3/11 27.3
27.3
±12.2
±3.7
5/12 41.7
41.7
±28.9
±8.3
12/12 90. la
100.0
±0.0
±0.0
2/2 NA
100.0
Duration of
survival (hr)
all
animals
335
±4
±1
(9)
36
±16
±5
(11)
139
±132
±40
(11)
150
±164
±47
(12)
336
±0
±0
(12)
336
non-
survivors
NA
36
±16
±5
(11)
65
±45
±16
(8)
18
±17
±7
(7)
NA
NA
Body
weight
(8) on
day 0
114
±16
±5
(9)
110
±11
±3
(ID
109
±7
±2
(11)
102
±10
±3
(12)
114
±13
±4
(12)
132
Weight gain (g)
days
0-7
57
±11
±4
(8)
NA
NA
44
±7
±4
(3)
*
77%
45
±12
±5
(6)
m
79%
NA
days
0-14
113
±16
±6
(8)
NA
10
±31
±18
(3)
***
9%
102
±19
±8
(5)
ns
90%
101
±26
±7
(12)
ns
89%
NA
days
7-14
52
±17
dbS
(7)
NA
NA
71
±4
±2
(3)
*
137%
56
±11
±5
(6)
ns
NA
59
-------
RuCl, / Oral administration
$
8
...I.
-ifi
m
t
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itt-ty
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2.6
2.7
2.8
2.9 3.0
3.1
CHAMPION' I.INT NO. 074 - PROBABILITY X 90
Dose (log)(mg salt/kg body weight)
60
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TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)
1. REPORT NO.
EPA-600/l-76-010a
2.
4. TITLE AND SUBTITLE
ASSESSMENT Of TOXfCITY OF AUTOMOTIVE METALl
Volume Ij Assessment of Fuel Additives Emis
yia Selected Assays of Nucleic Actd and Prc
7. AUTHOR(S)
David J, Hoi brook, Jr,
3. RECIPIENT'S ACCESSION-NO.
5. REPORT DATE
.1C EMISSIONS January 1976
>sion Toxicit'P'PERFORMINGORQANIZATION CODE
)tein Syrithes s
8. PERFORMING ORGANIZATION REPORT NO.
9. PERFORMING ORGANIZATION NAME AND ADDRESS
Department of Biochemistry
School of Medicine
University of North. Carolina
Chapel Hill, N.C. Z7514
12. SPONSORING AGENCY NAME AND ADDRESS
Heal th Effects Research Laboratory
Office of Research and Development
U.S. Environmental Protection Agency
Research Trianqle Park, N.C. 27711
10. PROGRAM ELEMENT NO.
1AA601
11. CONTRACT/G'RANT NO.
68-02^1 Z05
13. TYPE OF REPORT AND PERIOD COVERED
Final
14. SPONSORING AGENCY CODE
EPA-ORD
15. SUPPLEMENTARY NOTES
'\
16. ABSTRACT
Various parameters of toxic
palladium, and platinum. A
intraperitoneal injection a
PtCl4, Pt (5041.2, PdCl2., MnC
Concentrations of metallic
as are effects on weights o
Also following dietary admi
following parameters relate
microsomal protein/g livery
aminopyrtne deroethylase; co
protein.
Development of a rapid and
in studies of RNA synthesis
17.
ity have been studied for salts of manganese, lead,
cute toxicities (LD^50. doses I are reported for both
n.d oral administration for the, following salts j
lz., PdS04, PtCl2., RuCla, PtOz» PH), PdO, and MnOz-
ions following dietary^ administration are reported,
f five organs Giver, kidney, spleen, heart, testes).
nistration, hepatic microsomes were isolated and the
d to^in^vltro drug metabolism were measured: yield of
^tn VHrb" activities of aniline hydroxylase and
nteht of ' cytochromes Pr45Q and 55/mg microsomal
convenient method for the analysis of ribosomal RNA
is reported..
KEY WORDS AND DOCUMENT ANALYSIS
a. DESCRIPTORS
Toxlcity Metabolism
Manganese
Lead (metal I
palladium
platinum
Exhaust emissionsRihonucleic acids
Ribonucleic acids
18. DISTRIBUTION STATEMENT
RELEASE TO PUBLIC
b.lDENTIFIERS/OPEN ENDED TERMS
19. SECURITY CLASS (This Report)'
UNCLASSIFIED
20. SECURITY CLASS (This page)
UNCLASSIFIED
c. COS AT I Field/Group
06 F, T
21 D
21. NO. OF PAGES
67
22. PRtCE
EPA Form 2220-1 (9-73)
61
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