5-amino-2-methoxybenzenesulphonic acid

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from publication

Data source

Reference

Reference Type:

other: Autharized database

Title:

Reproduction Toxicity Screening Preliminary Test of 2-Amino-5-Methylbenzenesulfonic Acid by Oral Administration in Rats

Author:

J-CHECK

Year:

2016

Bibliographic source:

Ministry of Health, Labour and Welfare", "Ministry of the Environment" and "National Institute of Technology and Evaluation, J-CHECK - (2016)

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

Principles of method if other than guideline:

Reproduction Toxicity Screening Test of 2-Amino-5-Methylbenzenesulfonic Acid in Rats

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

4-aminotoluene-3-sulphonic acid

EC Number:

201-831-3

EC Name:

4-aminotoluene-3-sulphonic acid

Cas Number:

88-44-8

Molecular formula:

C7H9NO3S

IUPAC Name:

2-amino-5-methylbenzene sulfonic acid

Constituent 2

Reference substance name:

2- Amino-5-Methylbenzenesulfonic Acid

IUPAC Name:

2- Amino-5-Methylbenzenesulfonic Acid

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): 2- Amino-5-Methylbenzenesulfonic Acid
- Molecular formula (if other than submission substance): C7H9NO3S
- Molecular weight (if other than submission substance): 187.2181g/mole
- Substance type: Organic
- Physical state: fine yellowish white powder
- Impurities (identity and concentrations): 0.7 %

Specific details on test material used for the study:

- Name of test material (as cited in study report): 2- Amino-5-Methylbenzenesulfonic Acid
- Molecular formula (if other than submission substance): C7H9NO3S
- Molecular weight (if other than submission substance): 187.2181g/mole
- Substance type: Organic
- Physical state: fine yellowish white powder
- Impurities (identity and concentrations): 0.7 %

Test animals

Species:

rat

Strain:

Crj: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Co., Ltd. (Kanagawa)
- Age at study initiation: 10 weeks
- Weight at study initiation: 375 to 414 g in males and 239 to 266 g in females
- Fasting period before study: No data available
- Housing: Individual housed in an aluminum front and floor stainless steel mesh breeding cage in a breeding room. Maternal animals after gestation day 18 were kept on an aluminum front and floor stainless steel mesh breeding cage with nursery until nursing 4th.
- Diet (e.g. ad libitum): NMF solid feed (radiation sterilized feed) manufactured by Oriental Yeast Co., Ltd., and was taken free during the breeding period.
- Water (e.g. ad libitum): Tap water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24 ± 2 ° C
- Humidity (%): 55 ± 10%
- Air changes (per hr): Ventilation frequency of 15 times / hour,
- Photoperiod (hrs dark / hrs light): 12 hours (7 am lights, 7 pm off)

Administration / exposure

Route of administration:

oral: gavage

Type of inhalation exposure (if applicable):

not specified

Vehicle:

silicone oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: Administration solution, which was prepared by suspending the pharmacopoeial sesame oil (Miyazawa Pharmaceutical), was sealed stored under the cold shielding until use. Test substance drug substance and administration solution of the test substance was confirmed to be stable.

VEHICLE
- Justification for use and choice of vehicle (if other than water):
- Concentration in vehicle: 0,100 ,300 and 1000 mg/kg/bw/day
- Amount of vehicle (if gavage): 0.5 ml per body weight 100 g.
- Lot/batch no. (if required): No data available.
- Purity:

Details on mating procedure:

- M/F ratio per cage: 1;1
- Length of cohabitation: 14 days
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy Sperm confirmation in vaginal plaque, and that day was taken as the 0 day of pregnancy.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

For the analysis of the concentration and homogeneity of the administration solution, samples were randomly extracted from batches of each group prepared at the start of preparation. As a result, the error with respect to the display density was in the range of 12.5 to 0.4% and within the reference range (within ± 15%). Therefore, it was confirmed that a prescribed amount of 2amino5methylbenzenesulfonic acid was contained in the administration liquid used.

Duration of treatment / exposure:

Male: 48 days
Female: 69 days

Frequency of treatment:

Daily (Once)

Details on study schedule:

- Dose selection rationale: A 14 days repeated oral toxicity Study was observed in 1 group 4 males and 4 females, and the 0,100,250,500,1000 and 2000 mg / kg / day dose was administered for 14 day daily by oral gavage. General state, body weight, food consumption, urinalysis, hematology testing, in the blood biochemical examination and organ weight, suggest change the toxicity of the test substance was observed. In the autopsy, the cecum of expansion was observed in male and female in all cases of 2000 mg / kg group. Therefore the final dose selected for this study was 100, 300 and 1000 mg/kg/bw/day.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day

Doses / concentrations 2

Dose / conc.:

100 mg/kg bw/day

Doses / concentrations 3

Dose / conc.:

300 mg/kg bw/day

Doses / concentrations 4

Dose / conc.:

1 000 mg/kg bw/day

No. of animals per sex per dose:

Total no of animals 96
0 mg/kg/bw/day- 12 male, 12 female
100 mg/kg/bw/day - 12 male, 12 female
300 mg/kg/bw/day - 12 male, 12 female
1000 mg/kg/bw/day – 12 male, 12 female

Control animals:

yes, concurrent vehicle

Details on study design:

No data available

Positive control:

No data available

Examinations

Parental animals: Observations and examinations:

Survival, clinical sign, Body weight and Food intake was observed.

Oestrous cyclicity (parental animals):

Copulation formation, average season cycle, number of days between the estrus period and the next estrus period as the sex cycle days and mating rate were examined.

Corpus luteums, number of implantation traces were examined.

Sperm parameters (parental animals):

No data available

Litter observations:

Number of births, sex, body weight of 0 and 4 days were examined.

Postmortem examinations (parental animals):

Organ weight, gross pathology and histopathology were examined.

Postmortem examinations (offspring):

Gross abnormalities of organs and tissues were observed.

Statistics:

Weight, food intake, number of corpus luteums, number of implantation traces, number of births, number of stillbirths, sex ratio, average sex, pregnancy period, implantation rate, delivery rate, birth rate, abnormal incidence of abnormal outer table, newborn 4th Multiple comparison test 24)
was performed on the survival rate, organ weight and relative weight of the rats.
For the birth rate, mating rate and conception rate, χ ^ 2 tests 5, 6) were used. The incidence of findings of pathological examination was tested using Fisher's direct probability test method 6). In addition, the average for one newborn baby during the nursing period was taken as one sample. The level of significance was set at two levels of *: P <0.05 and **: P <0.01.

Reproductive indices:

Pregnancy period, Implantation rate, delivery rate, live birth rate, mating rate were examined.

Offspring viability indices:

Viability on day 4 were examined.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Description (incidence and severity):

No clinical signs were observed in treated male and female rats due to treatment as compared to control.

Dermal irritation (if dermal study):

not specified

Mortality:

no mortality observed

Description (incidence):

No effect on survival of treated male and female rats was observed as compared to control.
In male rats, ocular secretion in 1 rat at 100 mg / kg, hair loss in 1 rat at 300 mg / kg and Crust and hair loss were observed in 1 in case at 1000 mg / kg.
In female rats, hair removal was observed in 100 mg / kg group through pregnancy and nursing periods, and in the control group, all the dead animals were found in 2 cases.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

No significant effect on body weight and body weight gain was observed in treated male rats as compared to control.

In female rats, at 100 and 1000 mg / kg, statistically significant decrease in body weight only at 4th day of nursing was observed as compared to the control.

However, since there was no obvious difference on the other measurement days and it was a change only for 1 day, and since there was no obvious change in body weight gain during the nursing period, the influence of administration of the test substance It was not thought to be an accidental change.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

In male rats, statistically significant increase in food consumption from 8 to 15 days cumulative food intake from 1 to 15 days were observed at 1000 mg / kg bw.

In female rats, no significant effect was observed as compared to control.

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

In male rats, one case skin erosion and squamous epithelium hyperplasia and cell infiltration of epididymis at 1000 mg / kg, seminiferous tubular, testicular and epididymal atrophy of testes, stromal cell proliferation, spermatozoa of epididymis, lung inflammation and liver necrosis at 300 mg / kg were observed.
However, since it is low frequency expression, it was not considered to be influence of the test substance.

In female rats, 1 case of the lung inflammation, 1 case of the stomach ulcer, 1 part of the adrenal cortex and atrophy of the thymus at 1000 mg/kg bw, 1 case of atrophy of the thymus at 300 mg/kg bw and Young yolk sac cysts in one case, inflammatory infiltration of the skin and squamous epithelium proliferation were observed in one at 100 mg/kg bw, spontaneously delivered, so it was considered that there was no effect on the ovaries of the test substance.

Atrophy of the thymus and purulent inflammation of the uterus were observed in one case in all the dead animals. No abnormal findings were observed in the other case.

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

no effects observed

Description (incidence and severity):

No effect on estrous cycle was observed in treated rats as compared to control.

Reproductive function: sperm measures:

not specified

Reproductive performance:

no effects observed

Description (incidence and severity):

No effect on number of corpus luteums, number of implantation traces, number of births and number of stillborn were observed in treated rats as compared to control.

Gestation index, Implantation rate, delivery rate, live birth rate and mating rate of treated rats as compared to control.

Effect levels (P0)

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No effect on Survival, clinical sign, Body weight and Food intake, reproductive performance, organ weight, gross pathology and histopathology

Target system / organ toxicity (P0)

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Results: P1 (second parental generation)

General toxicity (P1)

Clinical signs:

not specified

Dermal irritation (if dermal study):

not specified

Mortality:

not specified

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

not specified

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Reproductive function / performance (P1)

Reproductive function: oestrous cycle:

not specified

Reproductive function: sperm measures:

not specified

Reproductive performance:

not specified

Effect levels (P1)

Dose descriptor:

other: not specified

Based on:

not specified

Sex:

not specified

Basis for effect level:

other: not specified

Remarks on result:

other: not specified

Target system / organ toxicity (P1)

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Results: F1 generation

General toxicity (F1)

Clinical signs:

not specified

Dermal irritation (if dermal study):

not specified

Mortality / viability:

no mortality observed

Description (incidence and severity):

No effect on number of live pups was observed as compared to control.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

No effect on body weight of pups was observed as compared to control.

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Sexual maturation:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

no effects observed

Description (incidence and severity):

No gross pathological changes were observed in pups as compared to control.

Histopathological findings:

no effects observed

Description (incidence and severity):

Renal pelvic enlargement in 1 case, renal pelvic enlargement in the kidney It was recognized in 1 and 3 cases in control and 300 mg / kg respectively. In both cases, the expression was expressed in a few cases, which was not related to the administration of the test substance.

Other effects:

not specified

Developmental neurotoxicity (F1)

Behaviour (functional findings):

not specified

Developmental immunotoxicity (F1)

Developmental immunotoxicity:

not specified

Effect levels (F1)

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

1 000 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

mortality

body weight and weight gain

gross pathology

histopathology: non-neoplastic

other: No effect observed

Target system / organ toxicity (F1)

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Results: F2 generation

General toxicity (F2)

Clinical signs:

not specified

Dermal irritation (if dermal study):

not specified

Mortality / viability:

not specified

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Sexual maturation:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

not specified

Histopathological findings:

not specified

Other effects:

not specified

Developmental neurotoxicity (F2)

Behaviour (functional findings):

not specified

Developmental immunotoxicity (F2)

Developmental immunotoxicity:

not specified

Effect levels (F2)

Dose descriptor:

other: not specified

Generation:

other: not specified

Based on:

not specified

Sex:

not specified

Basis for effect level:

other: not specified

Remarks on result:

other: not specified

Target system / organ toxicity (F2)

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Overall reproductive toxicity

Reproductive effects observed:

not specified

Treatment related:

not specified

Relation to other toxic effects:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Any other information on results incl. tables

Absolute and relative organ weight of rats treated orally with 2-Amino-5-methylbenzenesulfonic acid in the preliminary reproduction toxicity screening test.

Dose levelmg/kg/day	0	100	300	1000
Male No. of animals examined	12	12	12	12
Body weight(g)	542±33	554±45	534±36	546±3
Absolute organ weight
Testes (g)	3.62±0.31	3.65±0.44	3.28±0.59	3.70±0.23
Epididymides(mg)	1313±98	1295±138	1168±121*	1316±115
Relative organ weight
Testes (g %)	0.671±0.074	0.664±0.094	0.616±0.119	0.680±0.666
Epididymides  (mg %)	243.378±27.682	235.898±34.456	219.293±27.989	242.189±27.983

Values are expressed as Mean±SD

Significant difference from control group*P<0.05

Summary of reproductive performance in rats treated orally with2-Amino-5-methylbenzenesulfonic acid in the preliminary reproduction toxicity screening test

Dose levelmg/kg/day	0	100	300	1000
No. of pairs mated	12	12	12	12
No. of pairs copulated	12	12	12	12
No. of pregnant female	11	10	12	12
Copulation index%^a	100.0	91.7	100.0	100.0
Fertility index %^b	91.7	90.9	100.0	100.0
Estrus cycle( days ,Mean±SD)	4.5±0.7	4.2±0.5	4.2±0.4	4.5±0.5

a-No. of animals with successful copulation/No. of animals mated) X100

b-No. Of pregnant animals/ No. of animals with successful copulation X 100

Values in parentheses are expressed no. of animals observed

Finding of delivery in dams treated orally with -Amino-5-methylbenzenesulfonic acid and observation on their pups(F1) in the preliminary reproduction toxicity screening test

Dose level mg /kg/day	0	100	300	1000
No. of dams observed	11	10	12	12
No. of dams delivered live pups	11	10	12	12
Duration of gestation (Mean±SD)	22.7±0.6	22.4±0.5	22.3±0.5	22.7±0.4
No. of Corpora lutea(Mean±SD)	216	170	218	222
No. of total implants(Mean±SD)	188	161	186	175
No of total implants(Mean±SD)	172	150	178	160
No. of total pups born(Mean±SD)	168	150	178	160
Male	81	69	91		79
Female	87	81	87		81
Sex ratio (male and female) (Mean±SD)	1.00	0.93	1.13		1.21s
No of total live pups on days 4(Mean±SD)
Male	66	66	85		78
Female	66	77	80		79
No. of total dead pups born(Mean±SD)	4	0	0		0
Still birth	0	0	0		0
Cannnitelism	4	0	0		0
Gestation index(Mean±SD)^a	100	100	100		100
Implantation index(Mean±SD)^b	89.5±12.3	94.7±2.0	86.6±15.8		80.2±18.2
Delivery index(Mean±SD)^c	91.6±5.6	93.5±7.7	96.2±4.8		90.9±8.6
Live birth index(Mean±SD)^d	97.4±8.6	100±0	100±0		100±0
Viability index on day 4(Mean±SD)^e
Male	77.8±39.1	96.08.4	93.1±9.7		99.1±3.2
Female	77±39.1	95.7±5.6	94.4±15.8		98±4.8

a-(No of females with live pups/ No. of pregnant females) x100

b-( No of implants/No. of corpora lutea)x100

c-(( No .of pups born/No. of implants)X100

d.(No. of live pups born / No. of pups born)x100

e.-No. of live pups on day4 after birth/No. of live pups born)X100

(Includes live pups died before observation)

 

 

 

 

 

Applicant's summary and conclusion

Conclusions:

NOAEL was considered to be 1000 mg/kg bw for P and F1 generation when Crj: CD (SD) male and female rats were treated with 2- Amino-5-Methylbenzenesulfonic Acid orally by gavage in Sesame oil for 69 days.

Executive summary:

In a reproductive toxicity study,Crj: CD (SD) male and female rats were treated with2- Amino-5-Methylbenzenesulfonic Acid in the concentration of0, 100, 300 and 1000 mg/kg/bw/day orally by gavage in Sesame oil. No effect on survival of treated male and female rats were observed. In amle rats, ocular secretion in 1 rat at 100 mg /kg, hair loss in 1 rat at 300 mg /kg and Crust and hair loss were observed in 1 in case at 1000 mg/kg. In female rats, hair removal was observed in 100 mg/kg group through pregnancy and nursing periods, and in the control group, all the dead animals were found in 2 cases. No clinical signs were observed in treated male and female rats due to treatment as compared to control.In female rats, at 100 and 1000 mg / kg, statistically significant decrease in body weight only at 4th day of nursing was observed as compared to the control. However, since there was no obvious difference on the other measurement days and it was a change only for 1 day, and since there was no obvious change in body weight gain during the nursing period, the influence of administration of the test substance It was not thought to be an accidental change.In male rats, statistically significant increase in food consumption from 8 to 15 days cumulative food intake from 1 to 15 days were observed at 1000 mg / kg bwandin female rats, no significant effect was observed as compared to control. Similarly, no reproductive toxic effect were observed onestrous cycle, corpus luteums, number of implantation traces, number of births and number of stillborn, gestation index, implantation rate, delivery rate, livebirth rate and mating rate of treated rats as compared to control. In addition,statistically significant decrease in absolute weights of epididymiswere observedin male rats but no effect on relative weight of epididymis were observed at 300 and 1000 mg/kg bw as compared to contorl.No significant changes were observed intesticular weigth of treated male rats as compared to control. In male rats, black lesions on lung in 1 animal, red spots of the liver, testis, epididymis and thinning of the coat in 1 case of the same individual gropue were observed at 300 mg/kg bw and thinning of the coat 1 case of the same individual gropue were observed at 1000 mg/kg bw. In female rats, black spots in lungstomach ulcers and white spots of the adrenal glands in 1 case each at 1000 mg/kg bw and ovary cyst and coat thinning were observed in 1 case each at 100 mg/kg bw. No findings that could be attributed to administration of the test substance were observed in any of the animals. In male rats, one case skin erosion and squamous epithelium hyperplasia and cell infiltration of epididymis at 1000 mg / kg, seminiferous tubular, testicular and epididymal atrophy of testes, stromal cell proliferation, spermatozoa of epididymis, lung inflammation and liver necrosis at 300 mg / kg were observed. However, since it is low frequency expression, it was not considered to be influence of the test substance. In female rats, 1 case of the lung inflammation, 1 case of the stomach ulcer, 1 part of the adrenal cortex and atrophy of the thymus at 1000 mg/kg bw, 1 case of atrophy of the thymus at 300 mg/kg bw and Young yolk sac cysts in one case,inflammatory infiltration of the skin and squamous epithelium proliferation were observed in one at 100 mg/kg bw, spontaneously delivered, so it was considered that there was no effect on the ovaries of the test substance. Atrophy of the thymus and purulent inflammation of the uterus were observed in one case in all the dead animals. No abnormal findings were observed in the other case. Therefore, NOAEL was considered to be 1000 mg/kg bw for P and F1 generation when Crj: CD (SD) male and female rats were treated with2- Amino-5-Methylbenzenesulfonic Acid orally by gavage inSesame oil for 28 days.