Phosphoric acid, mixed esters with biphenyl-4,4'-diol and phenol

Endpoint summary

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Version / remarks:

of 1995

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: References detailed in Robust study summary section "Any other information on materials and methods incl. tables"

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Sprague Dawley rats, strain: Crl:CD(SD)(SPF) with appropriate range of bodyweight at study start.
- Source: Charles River Japan Hino Breeding Center.
- Age at study initiation (treatment start): Ca. 5 weeks.
- Assignment to dose groups: By bodyweight stratified randomization prior to treatment start.
- Weight at study initiation (treatment start): Males: minimum 145 g, maximum 167 g,
Females: minimum 117 g, maximum 137 g.
- Housing: Inside a barriered rodent facility in hanging stainless steel cages with wire mesh floor:
for quarantine and acclimation: In groups of 3 or 5/cage by sex
after assignment to dose groups: At 1 animal/cage
During urine collection individual housing in metabolism cages (15-17 h, treatment day 28 & recovery day 14).
- Diet (ad libitum, except during urine collection): MF pelleted diet (Lot No. 0803, Oriental Yeast)
- Water (ad libitum, also during urine collection): Chlorinated water from Hita City supply.
- Acclimation period (including 6 days of quarantine): 9 days before treatment start.

Diet and housing materials were autoclaved at 121°C prior to use. Routine analysis of the batch of diet used and drinking water produced results within acceptable ranges.

ENVIRONMENTAL CONDITIONS
The animal room was maintained at (target ranges for temperature and relative humidity):
- Temperature (°C): 22 to 25°C
- Relative Humidity (%): 56 ± 8 %
- Photoperiod (artificial lighting): 12 hrs day / 12 hrs night
- Rate of air exchange: 10 to 15 changes/h

Route of administration:

oral: gavage

Vehicle:

olive oil

Details on oral exposure:

- Concentration in vehicle: The concentration of the test material in vehicle varied between dose groups thus allowing constant dosage volume in terms of mL/kg bw/day.

- Amount (dose volume by gavage): 10 mL/kg bw/day.

- For concentrations of test material in vehicle at different dose levels, see Table 1 in "Any other information on materials and methods incl. tables"

- Dosing formulations were prepared fresh at about weekly intervals before (for analytical confirmation of concentration) and during the treatment period.

- Justification for choice of vehicle:
In a vehicle trial, the test material was not dissolved in purified water or olive oil, but was satisfactorily suspended in olive oil, when prepared at 10% w/v. Therefore, olive oil was chosen as a vehicle. In addition, in the present main study, satisfactory concentrations, homogeneity and stability of the test material in the chosen vehicle were confirmed.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

- Test material stability during the dosing period was confirmed by IR spectrophotometry.
- Chemical analysis of test material formulations by high performance liquid chromatography with UV-VIS detection (HPLC/UV-VIS).
- Chemical analysis confirmed that the prepared formulations were within 99.5 to 102% of the corresponding nominal concentration (conducted
prior to treatment start, at all dose levels) .
- Homogeneity of the test material formulations and their stability over 8 days were confirmed (by HPLC/UV-VIS) at the high and low dose levels.

Duration of treatment / exposure:

28 days

Frequency of treatment:

Once daily, 7 days/week on 28 consecutive days

Dose / conc.:

25 mg/kg bw/day (actual dose received)

Dose / conc.:

150 mg/kg bw/day (actual dose received)

Dose / conc.:

1 000 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

5 (Main study animals; all dose groups)
5 (Recovery animals; only vehicle control and high dose group)

Control animals:

yes, concurrent vehicle

Details on study design:

Dose selection rationale:
Dose selection in the present 28-day oral toxicity study was based on a 7-day repeated dose range-finding oral toxicity study with doses of 0 (vehicle control), 25, 250, 500 and 1000 mg/kg bw/day. In the 7-day study, adverse effects attributable to treatment with the test material were not evident. Salivation seen after administration was not considered to represent an adverse effect.

Study design of the present 28-day study:
5 m + 5 f / main study group: Vehicle control, low, mid and high dose groups, termination after 28 treatment days
5 m + 5 f / recovery group: Vehicle control and high dose groups, termination after 28 treatment days followed by 14 treatment-free recovery days

Positive control:

Not applicable.

Observations and examinations performed and frequency:

The following observations and examinations were performed on all animals:
- Clinical signs and mortality
during treatment period: Four times a day (before, during and just after dosing during the morning, and in the afternoon)
during recovery period: Twice daily
- Detailed clinical observations
and arena observations: Before treatment start and once a week during the treatment and recovery periods **.
- Functional Observation Battery:* During treatment week 4 on all animals.
- Body weights: At dose group allocation (during acclimation) and twice a week during the treatment and recovery periods.
In addition, immediately prior to necropsy for determination of organ to bodyweight ratios.
- Food consumption: At dose group allocation, once or twice a week during the treatment period and twice a week during recovery.

* FOB including sensory reactivity tests (approach, contact/touch, tail pinching and auditory startle, pupillary & righting reflexes)**, grip strength**
and locomotor activity.

** "Blind" recording, i.e. the treatment or dose level were not disclosed during recording.

- Haematology/Clinical chemistry (after overnight fasting for 16 to 20 hours with water available ad libitum):
main study animals: At completion of the treatment period.
recovery animals: At completion of the treatment-free recovery period

- Urinalysis (urine accumulated for 15-17 hours).
main study animals: At the end of the treatment period
recovery animals: At the end of the recovery period
Animals were deprived of food but had access to water during urine collection in metabolism cages.

Hematology parameters examined:
Red blood cell count, white blood cell count, platelet count, reticulocyte count, hemoglobin concentration, hematocrit value, differential leukocyte counts,
protrombin time, activated partial thromboplastin time, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration.

Blood chemistry parameters examined:
Total protein, albumin, A/G ratio, blood urea nitrogen, creatinine, glucose, total cholesterol, total bilirubin, triglyceride, sodium, potassium,
chloride, calcium, inorganic phosphorus, alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, gamma-glutamyl transpeptidase,
cholinesterase.

Urinary parameters examined:
Volume, colour, turbidity, osmotic pressure, pH, protein, glucose, blood, sediment (red blood cells, white blood cells, epithelial cells, casts crystals).

Sacrifice and pathology:

- Sacrifice and gross pathology (full macroscopic external and internal examination with tissue collection):
Main study animals: End of treatment period (Day 29)
Recovery animals: End of treatment free recovery period (Day 43)

- Organs Weights: The following organs of all main study and recovery animals were weighed at necropsy:
Brain, heart, liver, spleen, thymus, adrenals, kidneys, testes, epididymides, ovaries.

- Histopathology of main study animals for the vehicle control and 1000 mg/kg bw/day groups (organs microscopically examined):
Brain (cerebrum, cerebellum, pons), eye balls, pituitary gland, thyroid, parathyroids, heart, thymus, liver, spleen, adrenals, kidneys, testes,
epididymides, prostate, seminal vesicles, ovaries, uterus, vagina, lungs, trachea, forestomach, glandular stomach, duodenum, jejunum, ileum, caecum, colon, rectum, lymph node (axillary, mesenteric), urinary bladder, spinal cord, sciatic nerve, bone marrow (femur).
All dose groups: Gross lesions

Other examinations:

No

Statistics:

Data regarding body weights (excluding those at the time of necropsy), food consumption, hematological examinations, blood chemical examinations, urine volume, urine osmotic pressure, organ weights and FOB metrical data (grip strength and locomotor activity counts) were analyzed by using the Bartlett's test for homogeneity of variance. If the variances were homogeneous at a significance level of 5%, one way analysis of variance was performed. If there was a significant difference in this analysis, the difference between the vehicle control group and each of the treatment group was analyzed by the Dunnett's test. If the variances were not homogeneous, the Kruskal-Wallis's test was used. If there was a significant difference in this analysis, the difference between the vehicle control group and each of the treatment group was analyzed by the nonparametric Dunnett's test.

FOB numerical data (defecation and urination) was analyzed by using the Kruskal-Wallis's test. If there was a significant difference in this analysis, the difference between the vehicle control group and each of the treatment group was analyzed by the nonparametric Dunnett's test.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

but not adverse

Mortality:

mortality observed, treatment-related

Description (incidence):

salivation directly after gavage dosing was observed in few animals, not considered as adverse effect

Body weight and weight changes:

no effects observed

Description (incidence and severity):

see Figures attached in "Attached background material"

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

see Figures attached in "Attached background material"

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Description (incidence and severity):

statistically significant changes not considered to be toxicologically relevant

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

statistically significant changes not considered to be toxicologically relevant

Urinalysis findings:

no effects observed

Description (incidence and severity):

statistically significant changes not considered to be toxicologically relevant

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

In view of a lack of dose relationship, a statistically significant reduction in forelimb grip strength in males at 150 mg/kg/day was not considered to be toxicologically relevant.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

In view of a lack of dose relationship, a statistically significant reduction in epididymis weight in males at 150 mg/kg/day was not considered to be toxicologically relevant.

Gross pathological findings:

no effects observed

Description (incidence and severity):

incidental findings, toxicologically not relevant

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

incidental spontaneous background lesions. Toxicologically, these did not distinguish treated animals from concurrent controls.

Histopathological findings: neoplastic:

not examined

Details on results:

CLINICAL SIGNS AND MORTALITY
There were no deaths. Toxicologically relevant clinical signs or effects on sensory reactivity, grip strength or motor activity were not evident. Therefore, sensorimotor function was not tested on recovery animals. Salivation occurring just after administration in one female at 25 mg/kg/day and in both sexes at 150 and 1000 mg/kg/day increasing in number of affected animals with increasing dose was not considered to be an indication of neurotoxicity or to represent an adverse effect. The number of urinations in males of the 1000 mg/kg/day dose group statistically significantly higher than in concurrent controls was restricted to treatment week 4. In the absence of histopathological changes in urinary organs and of overt nervous effects in the present study, this statistically significant difference was not attributed to treatment with the test material. Statistically significantly lower forelimb grip strength in males at 150 mg/kg/day than in concurrent controls was not considered to be toxicologically relevant, because there were a lack of dose relationship in males and no confirmatory findings in females.

BODY WEIGHT AND FOOD CONSUMPTION
There were no adverse effects of treatment with the test material on bodyweight or food consumption.

HAEMATOLOGY
Mean corpuscular volumes (MCV) statistically significantly lower in main study males at 25 and 150 mg/kg/day than in concurrent controls were without toxicological significance, because there was a lack of dose-relationship in males and females were unaffected at all dose levels.

CLINICAL CHEMISTRY
Serum total cholesterol statistically significantly lower in recovery females at 1000 mg/kg/day than in concurrent controls at the end of the recovery period was considered to be fortitious and of no toxicological importance, because of a lack of confirmatory findings in treated main study animals at the end of the treatment period.

URINALYSIS
Urine volume statistically significantly higher and urine osmotic pressure statistically significantly lower in recovery females at 1000 mg/kg/day than in concurrent controls at the end of the recovery period were considered to be fortitious and of no toxicological importance, because of a lack of confirmatory findings in treated main study animals at the end of the treatment period.

NEUROBEHAVIOUR
In view of a lack of dose relationship, forelimb grip strength statistically significantly lower in males at 150 mg/kg/day than in concurrent controls at the end of the treatment period was not considered to be toxicologically relevant.

ORGAN WEIGHTS
There were no changes in organ weights or organ to terminal bodyweight ratios attributable to treatment with the test material. At the end of the treatment period, epididymis weight to terminal bodyweight ratios were statistically significantly higher at 150 mg/kg/day than in concurrent controls. In view of a lack of dose relationship, this difference was not considered to be toxicologically relevant. At the end of the recovery period, male kidney weights statistically significantly lower than in concurrent controls were also not attributable to treatment.

GROSS PATHOLOGY AND NON-NEOPLASTIC HISTOPATHOLOGY
Macroscopic or microscopic pathology findings, toxicologically, did not distinguish treated animals from concurrent controls and therefore were not attributable to treatment with the test material. In view of the absence of relevant findings in main study animals, recovery animals were not histologically examined.

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Key result

Critical effects observed:

no

Conclusions:

There were no adverse effects in the present study. Therefore, the "No Observerd Adverse Effect Level" (NOAEL) was the highest dose level tested, i.e. 1000 mg/kg/day in both sexes.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Study duration:

subacute

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

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

The NOAEL for systemic toxicity in adult male and female rats after 4 weeks of oral treatment with ADK STAB FP-800 was the highest tested dose of 1000 mg/kg/day. This does not necessitate any classification regarding repeated exposure according to European classification rules [REGULATION (EC) 1272/2008].