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Diss Factsheets

Toxicological information

Repeated dose toxicity: oral

Currently viewing:

Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2002
Report date:
2002
Reference Type:
secondary source
Title:
Unnamed
Year:
2004
Report date:
2004
Reference Type:
secondary source
Title:
Memorandum: Science Assessment for ATBC and TEC
Author:
Boyle K
Year:
2004
Bibliographic source:
Letter, August 24, 2004 from Kathryn Boyle, Inerts Team to Dan Rosenblatt, Chief, Minor Use, Inerts and Emergency Response Branch. U.S. EPA, OPPTS.
Report date:
2004

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Qualifier:
according to guideline
Guideline:
other: EPA OPP 86-5
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Tributyl O-acetylcitrate
EC Number:
201-067-0
EC Name:
Tributyl O-acetylcitrate
Cas Number:
77-90-7
Molecular formula:
C20H34O8
IUPAC Name:
tributyl 2-acetoxypropane-1,2,3-tricarboxylate
Test material form:
liquid
Remarks:
clear, oily
Details on test material:
Acetyltributyl citrate, batch # 0000016315, Expiry date: January 2002

Test animals

Species:
rat
Strain:
Wistar
Remarks:
Han Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
Age of animals at initation of the toxicity phase of the study: 4 weeks

Administration / exposure

Route of administration:
oral: feed
Details on route of administration:
Dietary
Details on oral exposure:
Groups of 20 male and 20 female Han Wistar rats received Citroflex A-4 via the diet at target
dosages of 100, 300 or 1000 mg/kg/day for 13 weeks. A similarly constituted Control group received
untreated diet. A further 10 male and 10 female rats were assigned to the Control and high dosage
groups for a four-week recovery period following the 13-week treatment period.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Group mean achieved dosages over the 13 week treatment period at 100, 300 or 1000 mg/kg/day were
103, 306 and 1013 mg/kg/day for males and 102, 306 and 1024 mg/kg/day for females.
Duration of treatment / exposure:
13 wks for repeated dose toxicity. A 4-week recovery period was a design feature, during which high dose animals received no exposure of test material.
Frequency of treatment:
daily in feed
Doses / concentrationsopen allclose all
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
Actual: 103 mg/kg bw/d in males, 102 mg/kg bw/d in females.
Dose / conc.:
300 mg/kg bw/day (nominal)
Remarks:
Actual: 306 mg/kg bw/d in males, 306 mg/kg bw/d in females,
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Remarks:
Actual: 1013 mg/kg bw/d in males, 1034 mg/kg bw/d in females
No. of animals per sex per dose:
20. 10 additional male and female rats were assigned to the Control and high dose groups for a 4-week recovery period following the 13 week treatment period.
Control animals:
yes, plain diet

Examinations

Observations and examinations performed and frequency:
During the Toxicity phase of the study the animals were observed daily for general clinical signs and
a detailed physical examination was performed weekly. Bodyweights and food consumption were
recorded weekly. Sexual maturation was assessed in Weeks 4 and 5 and oestrous cycles were
assessed in Week 9. Functional observational battery tests were performed weekly throughout the
treatment period. Ophthalmoscopy examinations were performed during Week 1 and Week 13.
treatment and during Week 4 of recovery.
On completion of the treatment or recovery period, as appropriate, animals were killed and examined
macroscopically; selected organs were weighed and tissues were processed for microscopic
evaluation. Sperm samples were analysed for motility and morphology. Liver samples were removed
from five males and five females in each group and evaluated for peroxisome proliferation.
Statistics:
For organ weights and body weight changes, homogeneity of variance was tested using Bartlett’s test followed by Behrens-fisher test or Dunnett’s test as appropriate. Macroscopic pathology and histopathology data were assessed using Fisher’s Exact test. Estrus cycles were analyzed using the Cochran-Armitage trend test. Other statistical tests used as appropriate were: Williams’ test for a dose-related response; Student’s t-test; Shirley’s non-parametric test for a dose-related response; Steel’s test; and Wilcoxon rank sum test. Significance level was p<0.05..

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Yellow staining of the perigenital and perianal areas of high dose females
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Slighlty low body weight gain for males and females of high dose group.
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
no effects observed
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Slightly high haematocrit, haemoglobin concentrations and erythrocyte counts in males receiving 100, 300 or 1000 mg/kg/day, and slightly
high erythrocyte counts in females receiving 300 or 1000 mg/kg/day. Not toxicologically significant.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Elevations in urea, glucose, sodium potassium, chloride; slightly high albumin and albumin:gobulin ratios; low calcium and phosphorus and total protein concentrations, at various doses in males and females. These biochemical values were within historical control, data ranges, except for the sodium values for males receiving 300 or 1000 mg/kg/day.
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
Low urinary pH and high urinary protein concentrations at 13 weeks in males receiving 1000 mg/kg/day. These changes were within historical control data ranges and were not apparent at the end of the Recovery period.
Behaviour (functional findings):
no effects observed
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
After 13 weeks of treatment, high bodyweight-relative liver weights were recorded for males and females that received 1000 mg/kg/day. This resolved on completion of the Recovery period.
Gross pathological findings:
no effects observed
Neuropathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Liver peroxisome proliferation after 13 weeks with a statistically significant increases in mean palmitoyl CoA oxidase activity in mid dose males and in high dose males and females, with a statistically significant increase in the mean supernatant protein concentration. Hepatocyte hypertrophy was recorded in the livers of high dose male and female rats.
Other effects:
no effects observed
Description (incidence and severity):
No statistically significant effects on sperm motility, sperm counts or morphology.
Details on results:
Haematological investigations in Week 13 indicated slightly high haematocrit, haemoglobin
concentrations and erythrocyte counts in males receiving 100, 300 or 1000 mg/kg/day, and slightly
high erythrocyte counts in females receiving 300 or 1000 mg/kg/day. These inter-group differences
are not considered to be toxicologically significant, were within historical control data ranges for
these parameters at this laboratory and were not apparent at the end of the Recovery period.

Biochemical examination of the blood plasma in Week 13 indicated high urea concentrations in males
receiving 300 mg/kg/day and in males and females receiving 1000 mg/kg/day, high glucose
concentrations in females receiving 1000 mg/kg/day, high sodium concentrations in males receiving
100, 300 or 1000 mg/kg/day, high potassium concentrations in males receiving 1000 mg/kg/day, high
chloride concentrations in males and females receiving 300 or 1000 mg/kg/day and low calcium and
phosphorus concentrations in females receiving 1000 mg/kg/day. In addition, slightly high albumin
concentrations, with corresponding increases in the albumin to globulin ratio, were recorded in males
receiving 300 or 1000 mg/kg/day, and slightly low total protein concentrations, also resulting in a
corresponding increase in the albumin to globulin ratio, were recorded for females receiving
1000 mg/kg/day. High urea concentrations were still apparent at the end of the recovery period in
females that had previously received 1000 mg/kg/day whilst all other inter-group differences showed
full recovery. The biochemical values of the blood plasma noted above were within historical control
data ranges, except for the sodium values for males receiving 300 or 1000 mg/kg/day.

Urinalysis investigations revealed low urinary pH and high urinary protein concentrations in Week 13
in males receiving 1000 mg/kg/day. These changes were within historical control data ranges and
were not apparent at the end of the Recovery period.

Liver peroxisome evaluation after 13 weeks of treatment indicated statistically significant increases in
mean palmitoyl CoA oxidase activity in males given 300 mg/kg/day and in males and females given
1000 mg/kg/day. Statistically significant increases in the mean supernatant protein concentrations
were also reported in these same dose groups.

Hepatocyte hypertrophy was recorded in the liver of rats receiving 1000 mg/kg/day. In males this
finding tended to occur in the centrilobular region whilst in females it tended to be more generalised.
Hepatocytic hypertrophy was not evident at the end of the Recovery period.

Effect levels

Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects

Target system / organ toxicity

Key result
Critical effects observed:
no

Applicant's summary and conclusion

Conclusions:
A 13-week dietary toxicity study was undertaken in weanling F1-generation Han Wistar male and female rats which had been exposed to the test material during gestation and weaning (for approximately 6 weeks). The parental generation of these rats (F0) had been exposed to this substance for 4 weeks prior to mating and 1 week during cohabitation. A 4 week recovery period was included after the 13 weeks of exposure of adult animals (F1). Doses were 100, 300 and 1000 mg/kg bw/d. No significant clinical signs or gross pathology was observed in males or females. Non-specific toxicity was obsrved in high dose animals characteristic of adaptive changes of liver and kidney. These included weak peroxisome proliferation (not relevant to humans), hepatocyte hypertrophy, and alterations in kidney function as seen in clinical chemistry and urinalysis parameters. The NOEL for these potentially non-adverse findings was 100 mg/kg bw/d for males and 300 mg/kg bw/d for females. The overall NOAEL was 1000 mg/kg bw/d for both sexes.