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Toxicological information

Repeated dose toxicity: oral

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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:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report Date:
2015

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Qualifier:
according to
Guideline:
EU Method B.26 (Sub-Chronic Oral Toxicity Test: Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Qualifier:
according to
Guideline:
EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: pellets
Details on test material:
- Name of test material (as cited in study report): Triphenyl phosphate
- Physical state: White pellets
- Analytical purity: 99.5%
- Lot/batch No.: CH13/085
- Stability under test conditions: stable
- Storage condition of test material: Room temperature

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: approximately 6 weeks
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days

Administration / exposure

Route of administration:
oral: feed
Details on oral exposure:
The required amount of test substance was ground to a fine powder and mixed directly with some powder feed without the use of a vehicle (premix) and subsequently mixed with the bulk of the diet. Elix water (approximately 15% in total) was added to aid pelleting. The pellets were dried for approximately 24 hours at 35ºC before storage. The control animals received similarly prepared pellets but without the test substance. Diets were prepared once weekly.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of diet preparations were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations, in Weeks 1, 6 and 13). Stability over 18 days at room temperature under normal laboratory light conditions was also determined (highest and lowest concentration, in Week 1).
Duration of treatment / exposure:
90 days
Frequency of treatment:
continuous via diet
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0, 300, 1500, and 7500 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
0, 20, 105, and 583 mg/kg bw for males
Basis:
actual ingested
Remarks:
Doses / Concentrations:
0, 22, 117, 632 mg/kg bw for females
Basis:
actual ingested
No. of animals per sex per dose:
10
Control animals:
yes, plain diet
Details on study design:
Based on the results of a 28-day range finding study (Project 505939; see Appendix 5), the dose levels for this 90-day oral gavage study were selected to be 0, 300, 1500 and 7500 ppm.

Examinations

Observations and examinations performed and frequency:
in line with Guideline requirements
Sacrifice and pathology:
in line with Guideline requirements
Statistics:
The following statistical methods were used to analyze the data:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (Ref. 1; many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex.
- The Steel-test (Ref. 2; many-to-one rank test) was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test (Ref. 3) was applied to frequency data.
- The Kruskal-Wallis nonparametric ANOVA test (Ref. 4) was applied to motor activity data to determine intergroup differences.

All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance. Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables. Test statistics were calculated on the basis of exact values for means and pooled variances. Individual values, means and standard deviations may have been rounded off before printing. Therefore, two groups may display the same printed means for a given parameter, yet display different test statistics values.

Results and discussion

Results of examinations

Details on results:
Treatment with the test substance up to 7500 ppm was well tolerated by the animals. No treatment-related mortality occurred, and no toxicologically relevant clinical signs were noted. The only in-life findings of note were a lower body weight (gain) in both sexes at 7500 ppm (approximately 21% and 12% lower for males and females at 7500 ppm, respectively), along with a slightly higher food intake (approximately 16% and 12% higher for males and females at 7500 ppm, respectively). Body weights of males showed an apparent slight dose-related decrease over the dose groups. Since these changes in body weight and food intake were considered slight in nature, these were not considered adverse in nature.

An apparent higher motor activity was recorded for males at 7500 ppm, which was not considered to be adverse in nature. Based on individual values, no clear dose-related group-response was noted, both in total mean counts and counts per interval. Also, all groups showed a similar motor activity habituation profile with a decreasing trend in activity over the duration of the test period. Also, these results were not supported by clinical observations or other functional observation tests and had no supportive morphological correlates in examined neuronal tissues.

There were treatment-related morphologic alterations in the liver, thyroid gland (males only), adrenal glands (both sexes) and stomach (females only), starting at 1500 ppm.

Histopathological findings in the liver consisted of centrilobular hepatocellular hypertrophy of the liver in males at 1500 and 7500 ppm and in females at 7500 ppm (up to slight degree). This was not considered to be adverse at the recorded severities and since degenerative changes in the liver were absent. This finding was accompanied by enlargement and red brown discolouration of the liver and higher liver weight at necropsy at 7500 ppm. The magnitude of liver weight increase at 7500 ppm (approximately 30 and 21% for males and females, respectively) was considered adverse in nature, although no supportive adverse histopathological changes were noted in the liver.

Morphological findings in the thyroid consisted of an increased incidence and/or severity of follicular cell hypertrophy of the thyroid gland in males at 1500 and 7500 ppm (up to slight degree), which might be secondary to the hepatocellular hypertrophy and is not considered to be adverse (Ref. 5). Necropsy of males at 7500 ppm showed enlargement and higher weight of the thyroid gland.

Histopathological findings in the stomach and adrenal glands were not considered to be adverse as these can also be found in control animals at similar severities and incidences. These lesions consisted of vacuolation of the limiting ridge of the stomach in females at 1500 and 7500 ppm (up to slight degree), hyperplasia and hyperkeratosis of the limiting ridge in a single female at 7500 ppm (at minimal degree), and increased vacuolation of the zona fasciculata or zona glomerulosa of the adrenal glands in males and females respectively at 7500 ppm (up to slight degree).

Changes in clinical biochemistry parameters consisted of higher total protein and calcium in males at 7500 ppm, and higher cholesterol in males and females at 7500 ppm, and in males also at 1500 ppm. These changes occurred in the absence of any correlating adverse histopathological changes, and were therefore not considered adverse in nature.

Effect levels

open allclose all
Dose descriptor:
NOAEL
Effect level:
105 mg/kg bw/day (actual dose received)
Based on:
test mat.
Remarks:
(test material intake of 1500 ppm treated males)
Sex:
male
Basis for effect level:
other: liver weight increase in males of the 7500 ppm group
Dose descriptor:
NOAEL
Effect level:
117 mg/kg bw/day (actual dose received)
Based on:
test mat.
Remarks:
(test material intake of 1500 ppm treated females)
Sex:
female
Basis for effect level:
other: liver weight in crease in females of the 7500 pm group

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Mean test article intake over the study period was as follows:

 

group

Nominal dietary inclusion level [ppm]

Average intake [mg test substance/kg body weight]

(range indicated within brackets)

males

females

2

300

20

(16-29)

22

(18-28)

3

1500

105

(81-151)

117

(97-147)

4

7500

583

(448-853)

632

(520-798)

 

Applicant's summary and conclusion

Executive summary:

Wistar rats were treated with Triphenyl phosphate for 90 consecutive days by dietary administration at dose levels of 0, 300, 1500 and 7500 ppm according to OECD 408. The mean test article intake over the study period was 0, 20, 105, and 583 mg/kg bw/d for males and 0, 22, 117, and 632 mg/kg bw/d for females. All groups consisted of 10 male and 10 female rats.

Treatment with the test substance up to 7500 ppm was well tolerated by the animals. No treatment-related mortality occurred, and no toxicologically relevant clinical signs were noted. Slight changes in body weight and food intake were not considered adverse in nature. A treatment-related increase in liver weight at 7500 ppm (approximately 30 and 21% for males and females, respectively) was considered adverse in nature, although no supportive adverse histopathological changes were noted in the liver. Histopathological findings in the stomach and adrenal glands were not considered to be adverse as these can also be found in control animals at similar severities and incidences.

Based on the liver weight increase at 7500 ppm, a No Observed Adverse Effect Level (NOAEL) for Triphenyl phosphate of 1500 ppm (corresponding to an actual test article intake of 105 and 117 mg/kg for males and females, respectively) was established.