Triphenyl phosphate

Endpoint summary

Description of key information

The NOAEL for repeated dose toxicity of triphenyl phosphate in male and female rats can be determined with 105 mg/kg bw/d,  the lowest NOAEL obtained in the recently performed Guideline study with subchronic exposure.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

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

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Qualifier:

according to guideline

Guideline:

EU Method B.26 (Sub-Chronic Oral Toxicity Test: Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system 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

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

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.

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.

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.

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

Critical effects observed:

not specified

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)

 

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.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

105 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: dermal

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: purity of test substance not stated, individual quantitative results not reported. Also assessed by OECD.

Qualifier:

equivalent or similar to guideline

Guideline:

EPA OPPTS 870.3200 (Repeated Dose Dermal Toxicity -21/28 Days)

GLP compliance:

not specified

Specific details on test material used for the study:

description: coarse white solid

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Details on test animals or test system and environmental conditions:

supplier: Marland Breeding FArms, inc. Hewitt, New Jersey 07421
weight at initiation of treatment: males mean 2.41 kg, females mean 2.42 kg
acclimation period: 24 days
food and water: ad libitum
environmental conditions: 12h light/dark cycle

Type of coverage:

other: no dressing used after application of test substance but collars were used to prevent ingestion of the material

Vehicle:

ethanol

Details on exposure:

Route of Administration: dermal exposure for 6h/day, 5 days/week over 21 to 23 days

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

10 g samples of the test material were taken one week pretest and on the last day of the study and sent to the sponsor for analytical confirmation of the test material

Duration of treatment / exposure:

21-23 days

Frequency of treatment:

5 days/week

Remarks:

Doses / Concentrations:
100 and 1000 mg/kg bw/day
Basis:
nominal per unit body weight

No. of animals per sex per dose:

10 males and 10 females per dose; the skin of the first 5 rabbits of each sex in each group was abraded twice weekly

Control animals:

yes, concurrent vehicle

Details on study design:

Post-exposure period: no
Two days prior to application of the test material an area equal to approximately 25% of the total body surface area was carefully and closely clipped on the dorsal surface. All rabbits were reclipped twice weekly.
The skin of the first 5 rabbits of each sex in each group was abraded twice weekly by producing shallow incisions, not sufficiently deep to cause bleeding but penetrating the horny layer of the epidermis.

Statistics:

yes: body weight, hematology and clinical chemistry parameters, organ weights and oran/body weight ratios were analysed.

Clinical signs:

no effects observed

Dermal irritation:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

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):

no effects observed

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

effects observed, treatment-related

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Details on results:

The only treatment related effect was a depression of acetyl cholinesterase in plasma, erythrocytes and brain.

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day

Sex:

male/female

Basis for effect level:

other: highest dose tested

Critical effects observed:

not specified

CLIN.CHEMISTRY: decrease of acetylcholinesterase activity in plasma,  erythrocytes and brain in males and females
ALL OTHER PARAMETERS: comparable to control/ no effects
2 rabbits of the control group (one male, one female) died spontaneously during the course of the study.

Conclusions:

The findings detected at the site of treatment as well as other parameters (mortality, clinical symptoms, body weight, heamatology, clinical chemistry, necropsy, organ weights, histopathology of >30 tissues (including reproductive organs and nervous system) were not different to controls. The only treatment related effect seen was a depression of acetyl cholinesterase in plasma, erthrocytes and brain of TPP treated rabbits. No clinical or histological correlations was found. No quantitative data was reported for this endpoint. This effect was not considered as of toxicological relevance.

Executive summary:

'The toxicity of TPP after repeated dermal exposure was determined in rabbits. Ten male and 10 female animals per group were treated on clipped, intact (half of the animals) and abraded skin (half of the animals), 6 hours/day, five times/week for three weeks with doses of 0, 100 and 1000 mg/kg bw/day under open conditions. Ingestion was prevented by means of a collar. TPP was applied as a 50% (w/v) solution in ethanol. The application volume was 0.2 mL or 2 mL/kg bw/day. Control animals were treated with 2 mL/kg bw/day ethanol alone. The findings detected at the site of treatment as well as other parameters (mortality, clinical symptoms, body weight, heamatology, clinical chemistry, necropsy, organ weights, histopathology of >30 tissues (including reproductive organs and nervous system)) were not different to controls. The only treatment related effect seen was a depression of acetyl cholinesterase in plasma, erthrocytes and brain of TPP treated rabbits. No clinical or histological correlations was found. No quantitative data was reported for this endpoint. This effect was not considered as of toxicological relevance.' (OECD SIDS, 2002)

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rabbit

Additional information

In a short-term study on triphenyl phosphate (TPHP) published recently by NTP (NTP 2018) male Sprague Dawley rats were treated once daily for 4 days by oral gavage with doses of 55, 110, 220, 441, and 881 mg/kg bw. On Day 5, animals were sacrificed, standard toxicological measures assessed (clinical signs, body weight, some clinical biochemistry parameters, liver and brain weight), and in a transcriptomic approach the liver was taken for gene expression studies using microarrays. Dose-dependent and statistically significant increases in absolute (at ≥ 220 mg/kg bw) and relative (at ≥ 441 mg/kg bw) liver weight and serum cholesterol levels (at ≥ 110 mg/kg bw) became obvious. A dose-dependent decrease in free thyroxine was observed at ≥ 220 mg/kg bw. Additional explorative endpoints were investigated as biomarkers of exposure, e.g. a decrease in serum cholinesterase levels at all doses was recorded, and transcriptomic analyses revealed polysaccharide biosynthetic process and oligodendrocyte development as the most sensitive gene sets for which a reliable estimate of the BMD could be made.

 

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 in 2015. 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. 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. 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. 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.

 

The results of the 90 day feeding study are well in line with those of a 28 day feeding study performed in 2007. In this study triphenyl phosphate was given in dietary concentrations of 0, 250, 1000 or 4000 ppm to 5 male and 5 female Wistar rats per dose group for a period of 4 weeks following OECD TG 407. This corresponded to an averaged (over the whole study) test substance intake of 23, 104 or 508 mg/kg bw/day in males and 39, 161 or 701 mg/kg bw/day in females.

 

The subacute and subchronic study are compared in the table below. Slight histopathological and functional changes in the liver in males at 104 mg/kg bw/d that were considered as adverse in the 28 day study could also be seen in the 90 day study at >= 105 mg/kg bw. However, in this study at the recorded severities and since degenerative changes in the liver were absent they were not considered to be adverse.

 

	Rat 28d oral toxicity study (OECD 407), supporting study of 2007	Rat 90d oral toxicity study (OECD 408), key study of 2015
Exposure route	oral via diet	oral via diet
Test substance uptake males	0, 23, 104, 508 mg/kg bw/d	0, 20, 105, 583 mg/kg bw/d
Test substance uptake females	0, 39, 161, 701 mg/kg bw/d	0, 22, 117, 632 mg/kg bw/d
mortality / clinical signs	none	none
effects considered as adverse	absolute and relative liver weight increased (absolute by 22 and 13% for males and females, respectively) at the highest dose; minimal to slight histopathological changes, ASAT activity decreased, cholesterol concentration increased at 104 mg/kg bw/d and above in males and at 701 mg/kg bw/d in females	Absolute liver weight increased at all doses in males; absolute and relative liver weight increased in males and females at the highest dose; minimal to slight centrilobular hepatocellular hypertrophy at 105 mg/kg bw and above in males and at 632 mg/kg bw in females is not considered as adverse at the recorded severities and degenerative changes were absent
NOAEL males	23 mg/kg bw/d	105 mg/kg bw/d
NOAEL females	161 mg/kg bw/d	117 mg/kg bw/d

 

Overall, the NOAEL for repeated dose toxicity of triphenyl phosphate in male and female rats can be determined with 105 mg/kg bw/d, the lowest NOAEL obtained in the recently performed Guideline study with subchronic exposure.

 

The toxicity of TPP after repeated dermal exposure was determined in rabbits (Monsanto, 1979). Ten male and 10 female animals per group were treated on clipped, intact (half of the animals) and abraded skin (half of the animals), five times/week for three weeks with doses of 0, 100 and 1000 mg/kg bw/day under open conditions. Ingestion was prevented by means of a collar. TPP was applied as a 50% solution in ethanol. The application volume was 0.2 mL or 2mL/kg bw/day. Control animals were treated with 2mL/kg bw of ethanol. 'The findings detected at the site of treatment as well as other parameters (mortality, clinical symptoms, body weight, heamatology, clinical chemistry, necropsy, organ weights, histopathology of >30 tissues (including reproductive organs and nervous system)) were not different to controls. The only treatment related effect seen was a depression of acetyl cholinesterase in plasma, erthrocytes and brain of TPP treated rabbits. No clinical or histological correlations were found. No quantitative data was reported for this endpoint. This effect was not considered as of toxicological relevance.' (OECD SIDS, 2002)

 

Triphenyl phosphate (11042-J) is currently (status June 2021) within the testing procedure of the National Toxicology Program (NTP) with the following tests for human health related toxicity:

-       2 week toxicity study on 5 B6C3F1/N mice per sex with doses of 0, 1875, 3750, 7500, or 30000 ppm triphenyl phosphate in food (C20720; status: data in review)

-       ‘Modified One-Generation Reproduction Study’ on Harlan Sprague Dawley rats, including neurotoxicology assessment (MOG11042; status for the respective dose-range finding study with administration via food: data in review)

Justification for classification or non-classification

Triphenyl phosphate was tested for repeated dose toxicity after oral (28d and 90d rat feeding studies) and dermal exposure (21d rabbit dermal study). The NOAEL for repeated oral toxicity in rats was determined with 105 mg/kg bw/d, obtained a subchronic study. In the dermal study no systemic toxicity became obvious up to the highest tested dose of 1000 mg/kg bw. Thus, according to EU Regulation 1272/2008 no classification for repeated dose toxicity is warranted.