O,O-bis(methylphenyl) hydrogen dithiophosphate

Endpoint summary

Description of key information

Repeated dose toxicity test was done according to OECD Guideline 422 under GLP compliance. Crj strain rats were used. No test item-related influence was noted on the reproduction parameters in any treatment group. The qualitative sperm staging revealed no test item-related spermatogenic changes. NOAEL (no-observed-adverse-effect level): above 137.1/182.8 mg/kg b.w./day, p.o. Effects on the development of the F1 offspring (pups) NOAEL (no-observed-adverse-effect level): 45.7 mg/kg b.w./day, p.o.

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

Remarks:

combined repeated dose and reproduction / developmental screening

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Crj: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories Research, Models and Services Germany GmbH, Sandhofer Weg 7, 97633 Sulzfeld
- Age at study initiation: 59 days
- Weight at study initiation: Males: 319.0 g to 353.1 g, Females: 209.8 g to 248.4 g
- Housing: singly (With the exception of the mating period) in MAKROLON cages (type III plus) with a basal surface of approx. 39 cm x 23 cm and a height of approx. 18 cm
- Diet (e.g. ad libitum): ssniff R-Z V1324 (ssniff Spezialdiäten GmbH, 59494 Soest, Germany) . The food was offered ad libitum with the exception of the night before the day of blood withdrawal for laboratory examination
- Water (e.g. ad libitum): Tap water was offered ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C ± 3°C
- Humidity (%): of 55% ± 15%
- Photoperiod (hrs dark / hrs light): 12 h d / 12 h l

IN-LIFE DATES: From: Males and females : September 5th 2012 To: Males:October 17, 2012 Females:November 3, 2012

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:

The test item formulations were freshly prepared on each administration day.

VEHICLE
- Concentration in vehicle: Variable, the dose of the test item was adjusted to the animal's actual body weight daily.
- Amount of vehicle: 10 mL/kg b.w./day
- Lot/batch no.: Batch no.: 11078709, Caesar & Loretz GmbH, 40721 Hilden, Germany.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

For the analysis of the test item-vehicle mixtures samples of approx. 2 x 1 mL were taken at the following time points and stored at ≤ minus 20°C until analysis at LPT:

Start of treatment period:
Concentration and stability:
Immediately after preparation of the test item vehicle mixtures as well as 8 and 24 hours after storage of the test item preparations at roomtemperature:
3 samples/dose level group (groups 2 to 4)
Number of samples: 3 x 4 = 12
Homogeneity:
At start of administration, during (middle) administration and before administration to the last animal of each dose level group:
3 samples/dose level group (groups 2 to 4).
Number of samples: 3 x 4 = 12

End of treatment period:
Concentration:
During treatment with the test item always before administration to the last animal/dose level group:
1 sample/dose level group (groups 2 to 4).
Number of samples: TD 28 (1 x 3) = 3
Number of samples: TD 41 (1 x 3) = 3
Sum of all samples: 30
Sum of all aliquots: 60

The samples were labelled with the study number, test item, test species, type of sample, aliquot number, concentration, test day, sampling time and date.

Duration of treatment / exposure:

Males
The daily administration of the test item started two weeks before mating and lasted until the day before sacrifice (considering a minimum total dosing period of at least 28 days).

Females
The daily administration of the test item started two weeks before mating and lasted up to at least day 3 of lactation.

Frequency of treatment:

once daily

Dose / conc.:

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

Dose / conc.:

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

Dose / conc.:

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

Dose / conc.:

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

Dose / conc.:

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

Remarks:

The high dose level was increased from 137.1 mg/kg to 182.8 mg/kg on test day 18, because the effects observed with 137.1 mg/kg after 2 weeks appeared to be too slight.

No. of animals per sex per dose:

80 animals (40 male and 40 female rats),10 animals/sex/group.
A sufficient number to grant at least 8 pregnant females per group for evaluation of the F0 generation.

Control animals:

yes, concurrent vehicle

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least daily, The frequency was increased when signs of toxicity were observed.
Cageside observations included skin/fur, eyes, mucous membranes, respiratory and circulatory systems, somatomotor activity and behaviour patterns. The onset, intensity and duration of any signs observed were recorded.
Individual animals were observed before and after dosing at each time of dosing for any signs of behavioural changes, reaction to treatment or illness.
In addition, animals were checked regularly throughout the working day from 7:00 a.m. to 3:45 p.m. On Saturdays and Sundays animals were checked regularly from 7:00 a.m. to 11:00 a.m. with a final check performed at approximately 3:30 p.m.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: at least daily, The frequency was increased when signs of toxicity were observed.
Additionally, once before the first exposure (to allow within-subject comparisons) and once a week thereafter, detailed clinical observations were made in all animals outside the home cage in a standard arena and at the same time, each time preferably by observers unaware of the treatment. Signs noted included changes in skin, fur, eyes, mucous membranes, occurrence of secretions and excretions and autonomic activity (e.g. lacrimation, piloerection, pupil size, and unusual respiratory pattern). Changes in gait, posture and response to handling as well as the presence of clonic or tonic movements, stereotypies (e.g. excessive grooming, repetitive circling), difficult or prolonged parturition or bizarre behaviour (e.g. selfmutilation, walking backwards) were also recorded.
Dated and signed records of appearance, change, and disappearance of clinical signs were maintained on clinical history sheets for individual animals.

BODY WEIGHT: Yes
- Time schedule for examinations: Males and females were weighed on the first day of dosing, weekly thereafter and at termination. During gestation, females were weighed on days 0, 7, 14 and 20 and within 24 hours of parturition (day 1 post-partum) and day 4 post-partum. Bodyweights were recorded individually for each adult animal.
The pups were weighed within 24 hours of parturition (day 1 post-partum) and on day 4 post-partum.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: At the end of the pre-mating period
- Under anaesthesia: Yes
- Animals fasted: Yes
- How many animals: 5 male and 5 female animals randomly selected from each group

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At the end of the pre-mating period
- Animals fasted: Yes
- How many animals: 5 male and 5 female animals randomly selected from each group

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Shortly before scheduled sacrifice
- Dose groups that were examined: 5 males and 5 females per group
- Battery of functions tested: Screening of sensory reactiviity to simuli of different types (eg. auditory, visual and proprioceptive sitimuli), grip strength and motor activity.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Statistics:

Toxicology and pathology data were captured, whenever possible, using the depart-mental computerized systems (Provantis® Integrated preclinical software, version 8.2.0, Instem LSS Ltd.). Raw data not fully compatible with the computerized systems were maintained on paper according to appropriate SOPs.

For all numerical values homogeneity of variances was tested by using the BARTLETT chi-square test. If the variances were homogeneous, the DUNNETT test (p ≤ 0.01) was used to compare the experimental groups with the control group.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

FO generation:
Male animals:
At the high dose level (137.1/182.8 mg Cresyl-P1/kg b.w./day) slight or moderate salivation was noted in 5 of 10 animals between test days 5 and 8 and at the end of the study between test days 41 and 43. Piloerection was noted for 2 of 10 animals between test days 19 and 24.
Female animals:
At the high dose level (137.1/182.8 mg Cresyl- P1/kg b.w./day) slight or moderate salivation was noted in 2 of 10 animals during the second test week with a duration of 1 or 2 days. Animal no. 77 showed a slight salivation on gestation day 21 and animal no. 71 showed a moderate salivation on lactation day 3.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

No test item-related mortality was noted.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Male animals:
In the high dose group (137.1/182.8 mg Cresyl-P1/kg b.w./day) a decreased body weight was noted from test day 8 until test day 42, reaching statistical significance (p≤0.05) on test days 29, 36 and on test day 42 (p≤0.01), with a maximum reduction of 10.9% in comparison to the control rats.
Body weight gain was reduced in the high dose group (137.1/182.8 mg Cresyl-P1/kg b.w./day) during the whole study, reaching statistical significance at the end of the first test week on test day 8 (p≤0.01) and test days 29, 36 (p≤0.05) and test day 42 (p≤0.01).
Female animals:
At the high dose level (137.1/182.8 mg Cresyl-P1/kg b.w./day) a slightly decreased body weight was noted from the end of the gestation period (gestation day 20) until the end of the lactation period, reaching statistical significance (p≤0.05) on lactation day 4, with a maximum reduction of 8.1% in comparison to the control rats.
This slight decrease in body weight was not reflected by a reduction of body weight gain.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

A statistically significant reduction in food consumption compared to the control (at p ≤ 0.01) was noted in the high dose group (137.1/182.8 mg Cresyl-P1/kg b.w./day during the first test week.

Haematological findings:

no effects observed

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Male animals:
In the high dose group (137.1/182.8 mg Cresyl-P1/kg b.w./day) a statistically significant (p ≤ 0.05) decrease was noted in the albumin and in the total protein concentration.
Female animals:
No test item-related influence was noted.

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Male and female animals:
The histomorphological examination revealed a mild to moderate squamous cell hyperplasia with mild to moderate hyperkeratosis in the forestomach of the male and female rats of the high dose groups (137.1/182.8 mg Cresyl-P1/kg b.w./day). These lesions were considered to be test item-related.
An additional examination of the forestomach of the male and female rats of the low and intermediate dose groups (13.7 and 45.7 mg Cresyl-P1/kg b.w./day) revealed no changes.
Spermatogenesis was not affected in the high dose group (137.1/182.8 mg Cresyl-P1/kg b.w./day).

Key result

Dose descriptor:

NOAEL

Effect level:

ca. 45.7 mg/kg bw/day (actual dose received)

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: Effects on the F0-generation

Key result

Critical effects observed:

not specified

Conclusions:

NOAEL (no-observed-adverse-effect level): 45.7 mg/kg b.w./day, p.o.

Effects on reproductive toxicity:
No test item-related influence was noted on the reproduction parameters in any treatment group.
The qualitative sperm staging revealed no test item-related spermatogenic changes
NOAEL (no-observed-adverse-effect level): above 137.1/182.8 mg/kg b.w./day, p.o.
Effects on the development of the F1 offspring (pups)
NOAEL (no-observed-adverse-effect level): 45.7 mg/kg b.w./day, p.o.

Executive summary:

Repeated dose toxicity test was done according to OECD Guideline 422 under GLP compliance. Crj strain rats were used. No test item-related influence was noted on the reproduction parameters in any treatment group. The qualitative sperm staging revealed no test item-related spermatogenic changes. NOAEL (no-observed-adverse-effect level): above 137.1/182.8 mg/kg b.w./day, p.o. Effects on the development of the F1 offspring (pups) NOAEL (no-observed-adverse-effect level): 45.7 mg/kg b.w./day, p.o.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

45.7 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Good quality

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

No classification is warranted based on the findings in the OECD 422 study according to Regulation (EC) No 1272/2008.