3,3'-thiodi(propionic acid)

Endpoint summary

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Effects on fertility

Description of key information

The NOELs for reproductive and developmental toxicity test are considered to be 100 mg/kg/day for males (based on testicular lesions in males receiving 300 mg/kg or more), and 1000 mg/kg/day for females and offspring.

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions. Only translated summary available, actual guideline not stated.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

Deviations:

yes

Remarks:

translated summary, actual guideline not stated

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories Japan, Inc.
- Age at study initiation: (P) 10 weeks
- Weight at study initiation: (P) 335 to 399 g for males and 208 to 241 g for females
- Fasting period before study:
- Housing: individually in mesh cages with aluminum sides and a stainless steel bottom in a barrier-system room, from 18 days after pregnancy until
after lactation for 4 days, the female animals were kept in mesh cages with aluminum sides and a stainless steel bottom with a nursing tray and materials for nest building (Sunflake made by Charles River Laboratories Japan, Inc.).
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 7 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24±3
- Humidity (%): 55±20
- Air changes (per hr): 15/h
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
The test material was suspended in 0.5 w/v% carboxymethylcellulose sodium (CMC-Na) aqueous solution, preparation was performed at least once every 7 days and this liquid was stored in a cool, dark location until administration.

VEHICLE
- Concentration in vehicle: 10, 30 and 100mg/kg bw
- Amount of vehicle (if gavage): 1mL/100g bw

Details on mating procedure:

- M/F ratio per cage: 1/1
- Length of cohabitation: during the nighttime for a maximum of 5 days
- Proof of pregnancy: sperm in the vaginal plug or vaginal smear referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged (how): From 18 days after pregnancy until after lactation for 4 days, the female animals were kept in mesh cages with aluminum sides and a stainless steel bottom with a nursing tray and materials for nest building (Sunflake made by Charles River Laboratories Japan, Inc.).

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The administration liquid concentration analysis was performed on the administration liquid of all test material administration groups prepared during the first preparation and the final preparation. The results were within the standard range (within ±10%).

Duration of treatment / exposure:

The administration period for males was for 52 continuous days being the 14 days prior to mating, 14 days during the mating period and 24 days after the mating period. For females it was for 41 to 45 days being the 14 days prior to mating, the mating period (max. 5 days), and for females that copulated successfully, the gestation period through 3 days of postpartum lactation. In addition, it was 40 to 44 days through to the day prior to necropsy on the 25th day of gestation for females that did not give birth after copulation.

Frequency of treatment:

daily

Details on study schedule:

- Age at mating of the mated animals in the study: 10+2 weeks

Remarks:

Doses / Concentrations:
100, 300, and 1,000 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

12

Control animals:

yes, concurrent vehicle

Positive control:

no

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes: mean bodyweight changes in grams between the observation timepoints
- Time schedule for examinations:The body weight of males was measured at administration day 1 (first administration day), 8, 15, 22, 29, 36, 43, 50, and 53 (necropsy day) and the increase in body weight from administrationday 1 to 53 was calculated. The body weight of females was measured at administration day 1 (first administration day), 8, and 15 and the increase in body weight from administration day 1 to15 days was calculated. Further, for females confirmed to have copulated, the body weight was measured on gestation day 0, 7, 14, and 20, and for postpartum females the body weight was measured on lactation day 0 and 4 (necropsy day) and the increase in body weight was calculated from gestation day 0 to 20 and lactation day 0 to 4, respectively. The body weight of dead animals was also measured when they were discovered.

FOOD CONSUMPTION
- Food consumption for each animal determined as mean food consumption in grams/day between the observation timepoints to determine the average daily amount of feed


OTHER: The copulation rate for each group [(number of animals with successful copulation/number of animals mated) x 100] was calculated from the mating results.

Oestrous cyclicity (parental animals):

Estrous cycle observation was continued until the day copulation was confirmed, and the number of days from estrus to the next estrus was designated as the number of days in the estrous cycle and then the average estrous cycle was calculated. In addition, the incidence [(number of females showing irregular estrous cycle/number of observed females) x 100] of irregular estrous cycles (an estrous cycle of other than 4 or 5 days) during the
estrous cycle observation period was calculated.

Sperm parameters (parental animals):

Parameters examined in [all/P/F1/F2] male parental generations: for the testis, after PAS hematoxylin staining and hematoxylin eosin staining, an examination of general pathological changes in the hematoxylin eosin stained specimens was conducted, and then the sperm formation cycle (Stage VII and VIII) was examined using the PAS hematoxylin stained specimens.

Litter observations:

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring: newborns, stillbirths, genders, sex ratio, check for appearance abnormalities, average body weight by male and female per uterus, 4-day viability of newborns [(Number of newborns on lactation day 4/number of live births)x100]

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities; possible cause of death was determined for pups born or found dead.

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals on the day after the 52 day of administration
- Maternal animals: All surviving animals on lactation day 4

GROSS NECROPSY
- Males: visual observation was made of the organs and tissues: testis, epididymisseminal vesicle, prostate and other organs and tissues found with abnormalities
- Females that birthed naturally: visual observation was made of the organs and tissues: ovaries, uterus, vagina and other organs and tissues found with abnormalities, corpora lutea and implantation evidence counts
- Females determined to have not birthed naturally: visual observation was made of the organs and tissues: ovaries, uterus, and vagina
- Dead animals: prostate, seminal vesicle, ovaries, uterus, vagina, testis, epididymis and other organs and tissues found with abnormalities during the visual observation

HISTOPATHOLOGY / ORGAN WEIGHTS
These tissues were prepared for microscopic examination and weighed: prostate, seminal vesicle, ovaries, uterus, vagina, testis, epididymis and other organs and tissues found with abnormalities during the visual observation

Postmortem examinations (offspring):

SACRIFICE
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows: visual observation made of the organs and tissues

Statistics:

For body weight, body weight increase, feed amount, cumulative feed amount, average estrous cycle, corpora lutea count, implantation evidence count, gestation, number of babies born, number of stillborns, sex ratio, implantation rate, live birth rate, delivery rate, appearance abnormality occurrence rate, newborn 4-day viability, organ weight, and relative weight were first examined using Bartlett’s test for equality of variance in accordance with the automated discrimination formula. For equal variance, a significant difference between the control group and the various administration groups was verified using Dunnet’s test of multiple comparison. For unequal variance found using Bartlett’s test for equality of variance, a significant difference between the control group and the various administration groups was verified using Steel’s test. χ2 test was performed for the birth rate, copulation rate, and fertility rate. The irregular estrous cycle incidence, necropsy case-finding rate, and histopathological case-finding rate were verified using Fisher’s exact test. For the observations that found some intensification of the pathological histological observations, - was assigned as “1”, +1 as “2”, +2 as “3”, and +3 as “4” and a Mann-Whitney U test was performed. For the significant level, 5% was used for Bartlett’s test for equality of variance, and for the other tests, tests of both 5 and 1% were made. However, when the number of specimen animals was 2 or less per group, a significant difference test was not performed. The performance of born babies during the lactation period was calculated as an average of 1 specimen per mother.

Reproductive indices:

birth rate [(number of females giving birth to newborns/number of pregnant females) x 100], implantation rate [(implantation evidence count/gestation corpora lutea count) x 100], delivery rate [(total number of babies born/implantation evidence count) x 100]

Offspring viability indices:

live birth rate [(live birth count/total number of babies born) x 100]

Clinical signs:

effects observed, treatment-related

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Other effects:

not examined

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
Only in the 1,000 mg/kg bw/day group 1 male (day 49) and 2 females (day 15 and day 25) died. In the dead animals, a nasal discharge, drop in body temperature, bradypnea, piloerection, soiled fur, drooping eyelids, and dyspnea were observed several days before death. 1 case of salivation, drooping eyelids, abnormal breathing sound, decline in autonomous movement, drop in body temperature, soiled fur, and abdominal position and extended abdomen was observedin the 1,000 mg/kg bw/day group.

BODY WEIGHT (PARENTAL ANIMALS)
For males no significant differences were found. For females, although a statistical analysis did not find a significant difference between the control group and the 1,000 mg/kg group in body weight increase from administration day 1 to 15, body weight during later gestation (gestation day 14 and later), and body weight increase from gestation day 0 to 20, a low value tendency was found in the latter. In the 1,000 mg/kg group, a low value tendency for body weight compared with the control group was also observed to continue from gestation into the lactation period, but no difference in body weight increase was found during the lactation period.

FOOD CONSUMPTION (PARENTAL ANIMALS)
For males no significant differences were found. For females, although a statistical analysis did not find a significant difference between the control group and the 1,000 mg/kg group for cumulative feed amount from administration day 1 to 15, from gestation day 0 to 20, or in the average daily feed amount for postpartum day 0 to 4, a low value tendency was found.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
no significant differences

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
no significant differences

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
no significant differences

ORGAN WEIGHTS (PARENTAL ANIMALS)
no significant differences

GROSS PATHOLOGY (PARENTAL ANIMALS)
The findings observed during the necropsies were all sporadic and were deemed not to be related to the test material administration. Of the animals that died from the 1,000 mg/kg bw/day group, both the male and female exhibited stomach and large intestine lumen dilatation, the male exhibited small intestine lumen dilatation, and the female exhibited lung reddening, stomach black mottling/zones and small intestine black mottling/zones were sporadically observed. Of the 2 and 1 cases where males did not induce pregnancy in the control group and 300 mg/kg bw/day group, respectively, atrophy of the testes and epididymis and lung white mottling/zones were observed in the 300 mg/kg bw/day group animal.

HISTOPATHOLOGY (PARENTAL ANIMALS)
There was an increasing trend in the incidence of testes seminiferous tubule atrophy and prostate gland dilatation for male animals in the test material administration groups. For the testes, seminiferous epithelia vacuolation was found in the 300 mg/kg group and multinucleated giant cell formation was found in the 300 mg/kg and higher groups. For the epididymis, epididymis lumen cell debris was found in the 1,000 mg/kg group. In addition, testes seminiferous tubule atrophy, epididymis lumen cell debris, and prostate gland dilatation were found in the cases of death in the 1,000 mg/kg group, and testes seminiferous tubule atrophy, seminiferous epithelia vacuolation, and epididymis lumen cell debris were found in 1 male from the 300 mg/kg group that did not induce pregnancy. These findings indicate that test material administration has an effect on fertility. No effect on the females from test material administration was observed.

Key result

Dose descriptor:

NOEL

Remarks:

Systemic

Effect level:

100 mg/kg bw/day

Based on:

test mat.

Sex:

male

Basis for effect level:

other: Testicular and epididymal lesions at 300 and 1000 mg/kg bw/day, respectively, were considered to be treatment-related.

Key result

Dose descriptor:

NOEL

Remarks:

Systemic

Effect level:

300 mg/kg bw/day

Based on:

test mat.

Sex:

female

Basis for effect level:

other: Body weight gain and food consumption were tended to be lower at 1000 mg/kg bw/day.

Key result

Dose descriptor:

NOAEL

Remarks:

Reproduction

Effect level:

100 mg/kg bw/day

Based on:

test mat.

Sex:

male

Basis for effect level:

other: Treatment-related histopathological findings were noted in males at 300 mg/kg bw/day and higher.

Key result

Dose descriptor:

NOAEL

Remarks:

Reproduction

Effect level:

1 000 mg/kg bw/day

Based on:

test mat.

Sex:

female

Basis for effect level:

other: No adverse effects observed in females.

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings:

not examined

VIABILITY (OFFSPRING)
no effects

CLINICAL SIGNS (OFFSPRING)
no effect from test material administration during the gestation period or in the gestation corpora lutea count was found nor was an effect on sex ratio, birth rate, implantation rate, or live birth rate found.

BODY WEIGHT (OFFSPRING)
no effects

GROSS PATHOLOGY (OFFSPRING)
The post-birth day 4 necropsy found red mottling/zones on the bottom of the feet of 6 males and 4 females in the 100 mg/kg group, and for males the incidence increased significantly compared to the control group. Also, there were observed occasional cases of renal pelvis dilation, ureteral dilation, small thymus, liver brown mottling/zones, liver nodules, liver diaphragm nodules, liver reddening, brain black mottling/zones, and brown fat black mottling/zones.

Key result

Dose descriptor:

NOAEL

Remarks:

Developmental

Generation:

F1

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No treatment-related and adverse effects were noted on external observation, clinical signs, viability, growth, or necropsy of the offspring.

Reproductive effects observed:

yes

Lowest effective dose / conc.:

300 mg/kg bw/day (nominal)

Treatment related:

yes

Relation to other toxic effects:

reproductive effects in the absence of other toxic effects

Dose response relationship:

yes

Relevant for humans:

not specified

For males a 300 mg/kg bw/day dosage is suspected of have an effect on fertility, and so the no observed adverse effect level NOAEL was determined to be 100 mg/kg bw/day. No effect on female reproductive ability or infant animal development were found, and so the no observed adverse effect level NOAEL was determined to be 1,000 mg/kg/day.

Executive summary:

3,3'-Thiobispropionic acid was studied for oral toxicity in rats according to the OECD Test Guideline 421 at doses of 0, 100, 300, and 1000 mg/kg/day.

In the repeated dose oral toxicity test, 1 male and 2 female rats receiving 1000 mg/kg were found dead. Body weight gain and food consumption in females receiving 1000 mg/kg also tended to be lower than in the control group. Histological evaluation demonstrated testicular lesions in males receiving 300 mg/kg or more.

The NOELs for repeated dose oral toxicity are considered to be 100 mg/kg/day for males, and 300 mg/kg/day for females.

With regard to reproductive/developmental toxicity, infertility was observed in 2 pairs and 1 pair in the control and 300 mg/kg groups, respectively. With 300 mg/kg, a treatment-related effect on male fertility was suggested, because lesions in some reproductive organs were observed on histological evaluation of males. The test substance did not demonstrate any adverse effects on other relevant parameters.

The NOELs for reproductive and developmental toxicity test are considered to be 100 mg/kg/day for males, and 1000 mg/kg/day for females and offspring.

Effect on fertility: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

100 mg/kg bw/day

Study duration:

subacute

Species:

rat

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

3,3'-Thiobispropionic acid was studied for oral toxicity in rats according to the OECD Test Guideline 421 at doses of 0, 100, 300, and 1000 mg/kg/day.

In the repeated dose oral toxicity test, 1 male and 2 female rats receiving 1000 mg/kg were found dead. Body weight gain and food consumption in females receiving 1000 mg/kg also tended to be lower than in the control group. Histological evaluation demonstrated testicular lesions in males receiving 300 mg/kg or more.

The NOELs for repeated dose oral toxicity are considered to be 100 mg/kg/day for males, and 300 mg/kg/day for females.

With regard to reproductive/developmental toxicity, infertility was observed in 2 pairs and 1 pair in the control and 300 mg/kg groups, respectively. With 300 mg/kg, a treatment-related effect on male fertility was suggested, because lesions in some reproductive organs were observed on histological evaluation of males. The test substance did not demonstrate any adverse effects on other relevant parameters.

The NOELs for reproductive and developmental toxicity test are considered to be 100 mg/kg/day for males, and 1000 mg/kg/day for females and offspring.

However, since no clear dose-response relationship was established, classification is not warranted.

Effects on developmental toxicity

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on the available data, 3,3'-Thiobispropanoic acid does not require classification for toxicity to reproduction according to regulation (EC) 1272/2008.