2'-acetonaphthone

Endpoint summary

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Effects on fertility

Description of key information

The study is ongoing and the template will be updated later after the completeion of the study.

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Data waiving:

other justification

Justification for data waiving:

other:

Justification for type of information:

The study is ongoing and this information will be submitted later based on ECHA communication/decision number CCH-D-2114501340-71-01/F

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

160 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

The data is K2 level and provides robbust summary

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Various reproductive/developmental toxicity studies have been investigated to observe the adverse reproductive effects occurring as a result of repeated daily oral dosing with, or exposure, to a substance for a specified period up to the expected lifespan of the test species. Often are the studies based on experiments of 60% and above structurally and functionally similar read across analogues. The studies are summarized as below: 

In the present study, one generation reproductive toxicity test of test chemical was carried out and the toxicity of reproductive to male and female animals of test chemical was examined. In the study, animals were orally administered from 10 weeks of age including mated up to 3 weeks. The males were necropsied after one week passed after the mating period. Females were allowed to spontaneously deliver after mating and necropsied with their babies on nursing 21th. Both males and females continued to be administered until the day before necropsy, during which the general condition of the parent animal, weight gain and changes in food intake were observed, and at the same time the reproductive ability, including delivery and lactation of the parent animal, and weaning of the infant was observed. The results of the parenteral examination revealed that, no mortality in treated animals at 10, 40 and 160mg/kg/day for 10 weeks,including mating period, the clinical signs examination of animlas shows, nasal discharge, eyelid ptosis or closed eyes, in group administered with 40 mg / kg or more but not in 160mg/kg group , lacrimation in 160 mg / kg administration group was transiently observed. Salivation was observed in each administration group of the test substance.No test chemical related chnages on food consumption and body weight changes were observed. Spontaneous locomotor reduction and salivation were observed transiently after administration and no abnormality was observed in both treated males and femlaes.  There was a decrease in locomotor activity, eyelid ptosis or closed eyes, and nasal discharge in group administered with 40 mg / kg or more, lacrimation in 160 mg / kg administration group. There was no change depending on the dose of the test substance in reproductive organsof male and females. The histopathological study relealed no abnormality in pituitary gland. In males, in each case in the 40 mg / kg and 160 mg / kg administration groups, the spermatogonia of the spermatogenic cells in the 14 th stage of the spermatogenesis cycle mild degeneration was observed, and cell debris was observed in the lumen of the epididymis in the example of 40 mg / kg administration group.  Unilateral seminal vesicle atrophy was observed in each group including the control group. No abnormality were seen on seminal vesicle and Coagulated glands In females, no abnormality was observed in the pituitary gland, stomach, ovary, uterus, cervix and vagina. The fetal examination results revealed, deaths in 4 offspings of control group and 2 offsping of 10 mg / kg administration group, respectively. In addition, 3 out of 6 cases in the 40 mg / kg administration group and 22 cases out of 27 cases in the 160 mg / kg administration group were judged as stillbirth, but no clinical signs of toxicity on survived pups. There was no significant difference in body weight in male and female  between the control group and each test group administered group at any time. No gross pathological changes were observed, At necropsy of born pups, morphological changes including malformations and mutations were observed in  born pups 1-2 in each group including the control group. From the observation and results, the NOAEL for P0 generation and F1 generation was considered to be 160mg/kg/day and 40mg/kg/day respectively.

In another One-Generation Reproduction Toxicity Test of test chemical in Rats was performed to estimate the reproductive toxicity profile. In the present study dose levels of 0, 12.5 ,50 and 200mg/kg used for the 10 weeks of exposure (71 dose administration) including mating period of 3 weeks. The treated animals observed for general condition, body weight measurement and food intake. Red blood cell count and white blood cell count was also measured. Estrus cycle during exposure was estimated. The offsprings were examined for number of births, general condition, number of deceased offspings, litter count, body weight, sex ratio and survival rate were measured.The postmortem of parenteral animals was carried out to examine major thoracic abdominal organs including the pituitary gland, stomach, adrenal glands, testis, epididymis, coagulated glands, seminal vesicles, prostate,ovaries, uterus and vagina. All surviving infants were necropsied by sacrificing all cases by ether inhalation on the 21th birth. At that time, the organ in which abnormality was observed. The dead child was autopsy examined for the presence or absence of abnormality on the external surface. The results of the parenteral observations revealed, transient salivation after test chemical administration in the 200 mg / kg group in both males and female. One case in the control group died due to poor feeding due to irregular occlusion. In addition, each one of 12.5 and 50 mg / kg administration group was moribund slaughtered or died by myeloid leukemia, but neither was caused by administration of the test substance. No death or moribund was observed in other treated animals. In test chemical treated males, suppression of body weight gain was observed at the end of treatment  and in females, suppression of body weight gain was observed between the early stage of administration of 200 mg / kg administration group and the middle stage of pregnancy to postpartum nursing, but the effects of administration was not observed in food intake. No change in food consumption were seen at any dose levels at 12.5, 50 and 200mg/kg/day compared to control animals.  The increase in the number of red blood cells(RBC) in the administration group of 50 mg / kg or more, no change related to the administration of the test substance was observed. At necropsy, hyperplasia of the forestomach mucosa was observed in the 200 mg / kg dose group.. In the 200 mg / kg administration group, attenuation of the degree of periportal fatty liver in the liver was also observed. For the liver, swelling was also observed at autopsy, but no distinct tissue change associated with swelling was observed. Histopathological examination of tissues revealed, no change due to administration of the test substance. In addition, changes in the pituitary, testis, epididymis, coagulated glands, seminal vesicles and prostate did not occur in any of the administration groups. In females, hyperplasia of squamous epithelium of forestomachial mucosa, but adrenal gland, pituitary gland, ovary , Uterus, uterine cervix and vagina, no change due to administration of the test substance was observed. The observations of neonates revealed, no mortality at all dose levels at 12.5,50 and 200mg/kg/day. No clinical signs of toxicity were seen at all dose levels at 12.5, 50 and 200mg/kg/day. The body weight of the 200 mg / kg administration group decreased from birth date to 21 days after birth, and growth inhibition ranging from fetal stage to nursing stage was observed. The frequency of things with external malformations such as the tail, short tail, short bending tail or small eye slightly increased in the 200 mg / kg administration group. In the 200 mg / kg administration group, abnormalities in the testis and hypoplasia of the spleen and visceral malformation of the diaphragmatic hernia were also observed. From the observations and results, the NOAEL for parents and neonates was considered to be 200mg/kg/day and 50mg/kg/day respectively.

Above study is further supported by one generation reproductive toxicity study of test material was performed according to OECD Guideline 415. The test material mixed with diet in dose concentration 0, 100, 500 and 2500 ppm and adminstered to groups of 25 rats per sex per group. Males were treated for 10 weeks and during the mated period and females for at least two weeks before mating until sacrifice. Body weights were recorded weekly during gestation period and at days 1, 4, 7, 14 and 21 during lactation period. Food consumption was measured weekly during experimental period. Food intake was recorded on presumed gestation days 7, 14 and 20 and on lactation days 4, 7, 14, 18 and 21. At birth, the number of pups born, sex and body weight of individual pups on days 1 and 4 were recorded. After standardization of litter size to 8 pups, pups were weighed individually on days 7, 14 and 21 of lactation. On day of weaning sacrifice (day 21), only one randomly selected pup per sex per litter was submitted to macroscopic examination. Fertility index for dams, sires and pup survival index were calculated. On completion of the gross pathology examination, the following tissues and organs were preserved in 10% neutral buffered formalin and submitted to microscopic examination: grossly abnormal tissues, ovaries, uterus, vagina, testes, epididymides, seminal vesicles, prostate, coagulating glands, pituitary gland and target organs of all P animals. The number of corpora lutea and implantation sites were recorded for all the dams. Histopathological examination of the parents was initially restricted to preserved organs from control and high dose group animals. There were significant lower body weights from control at high dose in males from weeks 2 to 6 and at week 8 associated with a significant lower food intake at weeks 2 and 8 at high dose in males. There was also a significant decrease in bodyweights at day 20 of the gestation period in females of the high dose group, also observed at days 4, 7 and 14 of lactation period. A statistically significant increase in relative kidney weights at mid dose in males and at high dose for males and females was observed. The increased kidney weights observed in males and females were minimal in nature (<12%) and were not associated with microscopic changes, hence considered as toxicologically insignificant changes. A statistically significant increase in relative epididymides was also recorded at high dose in males but with no related microscopic changes in left epididymides. Further, there were no corresponding change in the weight of right cauda epididymides and epididymal sperm count therefore the increased epididymal weights were considered as incidental changes.There were no test item related changes in sperm motility, cauda epididymal sperm counts and sperm morphology parameters. However, a statistically significant increase in percentage of abnormal sperms was observed in high dose males but this observed change was within the historical control data. Moreover, no fertility parameters were affected and there were no changes in the testes or epididymis grossly or histopathologically therefore, considered incidental and not related to the treatment. There were no test item related gross findings in males and females. Only single incidences of several gross findings observed in different groups were considered as incidental without any relation to test item administration. There were no test item related microscopic changes in males and females. All single or few incidences of microscopic findings observed in males and females were considered as incidental findings. The mean number and weight of male, female and total pups per litter at all the doses tested were unaffected by treatment. No treatment-related changes were observed in the data of pups up to lactation day 21 at all the doses tested. HenceNo Observed Adverse Effect Level (NOAEL) for maternal and developmental toxicity was considered to be 2500 ppm which is equivalent to 153.8 mg/kg Bwt/day for males and 393.6 mg/kg Bwt/day for females,.When male and femalerats were treated withtest chemical orally over one generation.

In an OECD 422 combined repeated dose toxicity and reproduction/developmental screening study, groups of 10 Sprague Dawley rats/sex/dose were administered acetophenone at doses of 0, 75, 225, or 750 mg/kg/day daily via gavage for a minimum of 14 days before mating and throughout the mating and gestation periods up to lactation day 3. There was a significant increase in the number of stillborn offspring among the high-dose group as compared to controls. There was a significant increase in the number of offspring dying, missing and/or cannibalized, along with an increase in the number of litters with total litter loss among the high-dose group during lactation days 1–4. There was a significant decrease in the total number of live born, viability index, and mean number of live pups per litter on lactation days 1–4. The number of mean live pups per litter was significantly lower on lactation days 1–4, and the live birth index was also reported to be out of the historical control range. Clinical signs among the high-dose group offspring included increased incidences of desquamation, cool to the touch, skin with shiny appearance, skin appearing tight with restricted movement, and a slightly increased incidence of gasping and pale skin color. There was a significant decrease in the pup weight per litter among the high-dose group on lactation days 1 and 4; this was reported to be out of the historical control ranges.

During gross pathological examination of offspring, high-dose group pups were reported with incidences of cleft palate and edema, atelectasis, dermal hypoplasia, scabbing, desquamation, and 22 dead pups with observed autolysis. Thus, the NOAEL for the developmental toxicity endpoint was considered to be 225 mg/kg/day, based on effects of treatment on viability of the offspring, alterations in clinical signs, body weight, and gross pathological alterations among the high-dose group offspring.

Effects on developmental toxicity

Description of key information

The developmental no observed adverse effect level (NOAEL) is considered to be 160 mg/kg/day for P0 generation and 40mg/kg/day F1 generation respectively.

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Justification for type of information:

Data is from Ministry of Health, Labor and Welfare report

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Principles of method if other than guideline:

Developmental toxicity test of test chemical was carried out in Sprague-Dawley rats.

GLP compliance:

not specified

Limit test:

no

Species:

other: Study 2 and 3: rat; Study 4: rabbit

Strain:

other: Study 2 and 3: Sprague-Dawley strain (Crj: CD) rats; Study 4: New Zealand White

Details on test animals or test system and environmental conditions:

Study 2: TEST ANIMALS
- Source: Tsukuba breeding center, Charles River Japan Co., Ltd.
- Age at study initiation: No data available
- Weight at study initiation: No data available
- Fasting period before study: No data available
- Housing: housed in a metallic wire mesh floor cage in a breeding room
- Use of restrainers for preventing ingestion (if dermal): yes/no: No data available
- Diet (e.g. ad libitum): No data available
- Water (e.g. ad libitum): tap water, provided ad libitum
- Acclimation period: No data available
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24 ± 1 ° C
- Humidity (%): 50 to 65%
- Air changes (per hr): about 15 times / hour
- Photoperiod (hrs dark / hrs light): lighting for 12 hours

Study 3: TEST ANIMALS
- Source: Tsukuba breeding center, Charles River Japan Co., Ltd.
- Age at study initiation: No data available
- Weight at study initiation: No data available
- Fasting period before study: No data available
- Housing: Animals were housed in a metallic wire mesh floor cage in a breeding room
- Use of restrainers for preventing ingestion (if dermal): yes/no
- Diet (e.g. ad libitum): No data available
- Water (e.g. ad libitum): No data available
- Acclimation period: No data available
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24 ± 1 ° C
- Humidity (%): 50 to 65%,
- Air changes (per hr): About 15 times / hour
- Photoperiod (hrs dark / hrs light): Lighting for 12 hours

Route of administration:

oral: gavage

Type of inhalation exposure (if applicable):

not specified

Vehicle:

CMC (carboxymethyl cellulose)

Remarks on MMAD:

No data available

Details on exposure:

Study 2: PREPARATION OF DOSING SOLUTIONS: The dosing solution was prepared by mixing the test chemical in vehicle with a magnetic stirrer
DIET PREPARATION
- Rate of preparation of diet (frequency): No data available
- Mixing appropriate amounts with (Type of food): No data available
- Storage temperature of food: No data available
VEHICLE
- Justification for use and choice of vehicle (if other than water):No data available
- Concentration in vehicle: No data available
- Amount of vehicle (if gavage): 5ml/kg
- Lot/batch no. (if required): No data available
- Purity:No data available

Study 4: Details on exposure
PREPARATION OF DOSING SOLUTIONS:
Test material dissolved in 0.5% Carboxymethyl cellulose

DIET PREPARATION
- Rate of preparation of diet (frequency): No data available
- Mixing appropriate amounts with (Type of food): No data available
- Storage temperature of food: No data available

VEHICLE
- Justification for use and choice of vehicle (if other than water): Test material dissolved in 0.5% Carboxymethyl cellulose
- Concentration in vehicle: 0, 3, 10,50mg/kg/day
- Amount of vehicle (if gavage): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

No data available

Details on mating procedure:

Study 2: Matings were conducted in males and females 1: 1 within the same group for a period of 3 weeks, and confirming the presence of sperm in the vaginal smear and confirming the vaginal plug every morning.

Study 3: Matings were conducted in males and females 1: 1 within the same group 3week period. Confirmation of mating was carried out by confirming the presence of sperm in the vaginal smear

Duration of treatment / exposure:

Study 2 and 3: 10 weeks (71 days of administration)
Study 4: 23 days (from gestation day 6 to 28)

Frequency of treatment:

Study 2, 3 and 4: Daily

Duration of test:

No data available

Remarks:

Study 2: 0 (vehicle control)= 25male and 25female
10mg/kg=25male and 25female
40mg/kg=25male and 25female
160 mg/kg=25male and 25female

Study 3: 0 mg / kg=25 male and 25 female
12.5 mg / kg=25 male and 25 female
50 mg / kg=25 male and 25 female
200 mg / kgmg / kg=25 male and 25 female

Study 4: 0, 3, 10,50mg/kg/day.

No. of animals per sex per dose:

Study 2 and 3:
(vehicle control)= 25male and 25female
10mg/kg=25male and 25female
40mg/kg=25male and 25female
160 mg/kg=25male and 25female

Study 4: Total:100
0mg/kg/day: 25 female
3mg/kg/day:25 female
10mg/kg/day:25 female
50mg/kg/day:25 female

Control animals:

yes, concurrent vehicle

Details on study design:

Study 2: Animals were orally administered from 10 weeks of age including mated up to 3 weeks. The males were necropsied after one week passed after the mating period. Females were allowed to spontaneously deliver after mating and necropsied with their babies on nursing 21th. Both males and females continued to be administered until the day before necropsy, during which the general condition of the parent animal, weight gain and changes in food intake were observed, and at the same time the reproductive ability, including delivery and lactation of the parent animal, and weaning of the infant was observed.

Study 3: oral administration of test chemical in the doses at 12.5, 50 and 200mg/kg in groups of 25/sex was carried out from 10 weeks including matting up to 3 weeks. The males were necropsied after one week passed after the mating period. Females were allowed to spontaneously deliver after mating and necropsied with their babies on nursing 21th. Both males and females continued to be administered until the day before necropsy, during which the general condition of the parent animal, weight gain and changes in food intake were observed, and at the same time the developmental ability was observed

Study 4: - Dose selection rationale: base on the preliminary range-finding study (0, 10, 60 and 300 mg/kg/day).
- Rationale for animal assignment (if not random): No data available
- Other: No data available

Maternal examinations:

Study 2: Parent animals were observed for mortality, general condition, change in weight, food intake, estrus cycle assessment. Animals were sacrificed to determined presence or absence of abnormalities of major organs thoracic abdomen, including the pituitary glands, stomach, testis, epididymis, coagulated glands, seminal vesicles, and prostate in males and pituitary, stomach, ovary, uterus, cervix and vagina in females

Study 3: Parenteral animals were observation for general condition, body weight measurement and food intake. Red blood cell count and white blood cell count were measured. Gross necropsy of major thoracic abdominal organs including the pituitary gland, stomach, adrenal glands, testis, epididymis, coagulated glands, seminal vesicles, prostate,ovaries, uterus and vagina

Study 4: Parental animals observation and examinations
CAGE SIDE OBSERVATIONS: yes
DETAILED CLINICAL OBSERVATIONS: Yes
Time schedule: twice daily
BODY WEIGHT: Yes
Time schedule for examinations: Individual body weights were recorded on gestation days 0, 3, 5, 8, 11, 14, 17, 20, 23, 26, 29 and 30.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes Food consumption per cage of animals were measured over the following periods during gestation: days 0-3, 3- 5, 5-8, 8-11, 11-14, 14-17, 17-20, 20-26, 26-29 and 29-30.
Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes / No / No data: No data available
WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
Time schedule for examinations:
OTHER: No data available

Ovaries and uterine content:

Study 2: Estrus cycle were determined before 2 weeks of start of mating confirmation day, two weeks before the start of administration and two weeks after the start of the administration.

Study 4: The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: No data available

Fetal examinations:

Study 2: Number of births, litter count and weight measurement. Offspings were sacrificed to determined presence or absence of abnormality on the external surface

Study 3: Number of births, general condition and number of deceased offspings, litter count, body weight, sex ratio and survival rate were measured.All surviving infants were necropsied by sacrificing all cases by ether inhalation on the 21th birth. At that time, the organ in which abnormality was observed. The dead neonates was autopsy examined for the presence or absence of abnormality on the external surface

Study 4: - External examinations: Yes: all per litter
- Soft tissue examinations: Yes
- Skeletal examinations: Yes:
- Head examinations: Yes

Statistics:

Study 2 and 3: Fisher's direct probability test 1) was carried out on frequency of type of sex cycle, mating rate, conception rate, morphological abnormality frequency of babiesAccording to histopathological examination findings, the Mann-Whitney U test 2)) shows the grade-separated data, and the total value of the positive grade is obtained by Fisher's direct probability one-sided test 3) between the control group and each test substance administration group Significant difference test was carried out. For the other data, we tested the uniformity of variance of each group by Bartlett method 4) , with the value obtained for each individual, or the average value for each litter as one sample . If the variance is uniform, a one-way analysis of variance 4) was performed, and when significance was observed between the groups, multiple comparisons were performed according to the Dunnett method 5) . On the other hand, Kruskal-Wallis 6) rank test is performed when the variance is 0 in any group and when the variance is not uniform, and if significance is observed between the groups.

Study 4: The test parameters body weight, percent body weight change, feed consumption, prenatal data and foetal data will be analysed using statistical techniques like Bartlett’s test, ANOVA and Dunett’s and Student’s t tests. Foetal examination will be analysed using Chi-square test.

Indices:

Study 2: Conception rate was determined
Study 3: Birth rate and Pregnancy period was observed

Historical control data:

No data available

Clinical signs:

no effects observed

Description (incidence and severity):

Study 2: Nasal discharge, eyelid ptosis or closed eyes, in group administered with 40 mg / kg or more but not in 160mg/kg group , lacrimation in 160 mg / kg administration group was transiently observed Salivation was observed in each administration group of the test substance.

Study 3: Transient salivation was observed after administration in the 200 mg / kg administration group in both males and female

Dermal irritation (if dermal study):

not specified

Mortality:

no mortality observed

Description (incidence):

Study 2: In males, one patient in the 160 mg / kg dose group died on the 60th day of administration. In females, there were no deaths or moribund

Study 3: One case in the control group died due to poor feeding due to irregular occlusion. In addition, each one of 12.5 and 50 mg / kg administration group was moribund slaughtered or died by myeloid leukemia, but neither was caused by administration of the test substance. No death or moribund was observed in other treated animals.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Study 2: No significant difference between the control group and each test group administered group

Study 3: In test chemical treated males, suppression of body weight gain was observed at the end of treatment and in females, suppression of body weight gain was observed between the early stage of administration of 200 mg / kg administration group and the middle stage of pregnancy to postpartum nursing, but the effects of administration was not observed in food intake.

Study 4: There was a significant reduction in the body weight gain during the treatment period in the high dose group (50 mg/kg).The reduction in body weight during the treatment period was considered treatment related.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Study 2: In males no change in food consumption were observed, however in females no change in food consumption uring the period before mating, but significantly lower values than control group, after pregnancy and during the nursing stage

Study 3: No change in food consumption were seen at any dose levels at 12.5, 50 and 200mg/kg/day compared to control animals.

Study 4: The food consumption was comparable to the vehicle control group.

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Description (incidence and severity):

Study 3: The increase in the number of red blood cells(RBC) in the administration group of 50 mg / kg or more, no change related to the administration of the test substance was observed

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

Study 2: Spontaneous locomotor reduction and salivation were observed transiently after administration and no abnormality was observed in both treated males and femlaes. There was a decrease in locomotor activity, eyelid ptosis or closed eyes, and nasal discharge in group administered with 40 mg / kg or more, lacrimation in 160 mg / kg administration group.

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

no effects observed

Description (incidence and severity):

Study 2: There was no change depending on the dose of the test substance in reproductive organsof male and females. Abnormality was not found in the reproductive organs of the 160 mg / kg administration group.

Study 3: At necropsy, hyperplasia of the forestomach mucosa was observed in the 200 mg / kg dose group.. In the 200 mg / kg administration group, attenuation of the degree of periportal fatty liver in the liver was also observed. For the liver, swelling was also observed at autopsy, but no distinct tissue change associated with swelling was observed.

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

Study 2: No abnormality was observed in pituitary gland. In males, in each case in the 40 mg / kg and 160 mg / kg administration groups, the spermatogonia of the spermatogenic cells in the 14 th stage of the spermatogenesis cycle mild degeneration was observed, and cell debris was observed in the lumen of the epididymis in the example of 40 mg / kg administration group. Unilateral seminal vesicle atrophy was observed in each group including the control group. no avnormality were seenon seminal vesicle and Coagulated glands In females, no abnormality was observed in the pituitary gland, stomach, ovary, uterus, cervix and vagina.

Study 3: Histopathological examination of tissues revealed, no change due to administration of the test substance. In addition, changes in the pituitary, testis, epididymis, coagulated glands, seminal vesicles and prostate did not occur in any of the administration groups. In females, hyperplasia of squamous epithelium of forestomachial mucosa, but adrenal gland, pituitary gland, ovary , Uterus, uterine cervix and vagina, no change due to administration of the test substance was observed

Histopathological findings: neoplastic:

not specified

Other effects:

effects observed, treatment-related

Description (incidence and severity):

Study 2: Hair loss was observed in all the treated animals
Study 3: No abnormality was observed in the estrus cycle during study period

Number of abortions:

no effects observed

Description (incidence and severity):

Study 4: One rabbit aborted in the 50mg/kg dose group

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

Study 2: Implantation number in treated animals was equivalent to control animals

Study 4: The maternal data parameters comprising of pre and post-implantation loss in all the treatment groups were comparable to the vehicle control group

Total litter losses by resorption:

effects observed, treatment-related

Description (incidence and severity):

Study 2 :The litter size was significantly low value in 160mg/kg/day group compared to control group.

Study 4: The maternal data parameters comprising of early and late resorptions in all the treatment groups were comparable to the vehicle control group

Early or late resorptions:

not specified

Description (incidence and severity):

Study 4: The maternal data parameters comprising of early and late resorptions in all the treatment groups were comparable to the vehicle control group

Dead fetuses:

not specified

Description (incidence and severity):

Study 3: No dead fetuses were seen at dose levels at 12.5, 50 and 200mg/kg

Changes in pregnancy duration:

no effects observed

Description (incidence and severity):

Study 2: No change in pregnancy duration were seen in treated females.
Study 3: No change in pregnancy duration were observed at any dose levels at 12.5, 50 and 200mg/kg/day

Changes in number of pregnant:

not specified

Description (incidence and severity):

Study 4: There were 2 non pregnant rabbits in control, 4 in low dose group, 3 in mid dose group and 4 in the high dose group. There was one complete resorption in mid dose group.

Other effects:

no effects observed

Description (incidence and severity):

Study 2: No abnormality in labor condition was observed, and there was no effect on birth rate

Dose descriptor:

NOAEL

Effect level:

160 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

behaviour (functional findings)

body weight and weight gain

clinical signs

effects on pregnancy duration

food consumption and compound intake

gross pathology

histopathology: non-neoplastic

maternal abnormalities

mortality

pre and post implantation loss

total litter losses by resorption

other: labor condition was normal no effect on birth rate

Dose descriptor:

NOAEL

Effect level:

200 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

body weight and weight gain

changes in number of pregnant

changes in pregnancy duration

clinical signs

dead fetuses

food consumption and compound intake

gross pathology

haematology

histopathology: non-neoplastic

mortality

Dose descriptor:

NOAEL

Effect level:

10 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

body weight and weight gain

changes in number of pregnant

early or late resorptions

food consumption and compound intake

mortality

number of abortions

pre and post implantation loss

total litter losses by resorption

Fetal body weight changes:

effects observed, treatment-related

Description (incidence and severity):

Study 2: Body weight at the postnatal day 21 was slightly lower in the 160 mg / kg administration group
Study 3: The body weight of the 200 mg / kg administration group decreased from birth date to 21 days after birth.

Reduction in number of live offspring:

effects observed, treatment-related

Description (incidence and severity):

Study 2: Number of live pups examined reduced in highest dosed group
Study 3: The effect of administration of the test substance on the no of pups was not observe

Changes in sex ratio:

no effects observed

Description (incidence and severity):

Study 2: There was no effect of administration on sex ratio

Changes in litter size and weights:

effects observed, treatment-related

Description (incidence and severity):

Study 2: The litter size after adjustment of litter size was decreased in 160mg/kg/day group

Changes in postnatal survival:

effects observed, treatment-related

Description (incidence and severity):

Study 2: In 160mg/kg /day group the postnatal survival declines comapared to control
Study 3: No changes on post natal survivability was observed.

External malformations:

no effects observed

Description (incidence and severity):

Study 2: No morphological changes including malformations where observed
Study 3: External malformations such as the tail, short tail, short bending tail or small eye slightly increased in the 200 mg / kg administration group

Skeletal malformations:

no effects observed

Description (incidence and severity):

Study 2: No morphological changes including malformations where observed

Visceral malformations:

no effects observed

Description (incidence and severity):

Study 2: No morphological changes including malformations where observed
Study 3: Abnormalities in the testis and hypoplasia of the spleen and visceral malformation of the diaphragmatic hernia were also observed in 200mg/kg group.

Other effects:

no effects observed

Description (incidence and severity):

Study 2: No growth inhibition was observed in the administration group of 40 mg / kg or less
Study 3: At the 200 mg / kg administration group growth inhibition ranging from fetal stage to nursing stage was observed

Dose descriptor:

NOAEL

Effect level:

40 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

changes in sex ratio

external malformations

skeletal malformations

visceral malformations

other: No growth inhibition

Dose descriptor:

NOAEL

Effect level:

50 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

reduction in number of live offspring

changes in postnatal survival

other: the mentioned dose level was observed to be safe in change in body weight, growth inhibition, external and visceral malformations

Dose descriptor:

NOAEL

Effect level:

50 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

not specified

Basis for effect level:

changes in sex ratio

fetal/pup body weight changes

Remarks on result:

other: No developmental toxic effects observed

Developmental effects observed:

not specified

Treatment related:

not specified

Conclusions:

From the observation and results, the developmental no observed adverse effect level (NOAEL) is considered to be 160 mg/kg/day for P0 generation and 40mg/kg/day F1 generation respectively.

Executive summary:

Various reproductive/developmental toxicity studies have been investigated to observe the adverse reproductive effects occurring as a result of repeated daily oral dosing with, or exposure, to a substance for a specified period up to the expected lifespan of the test species. Often are the studies based on experiments of 60% and above structurally and functionally similar read across analogues. The studies are summarized as below: 

One-generation reproductive toxicity in rats (MHW Report, 2001) as per OECD TG 415 using the analogue substance. Groups of 25 animals/dose/sex were treated with 0, 10, 40, 160 mg/kg of test chemical by gavage for 14 weeks (98 days). Males were dosed 10 weeks before mating, during mating and they were necropsied one week after the end of the mating period. Females received the test chemical 2 weeks before mating, during 3 weeks of mating and pregnancy till the 21stday of lactation (overall 98 days). Dams were allowed to deliver pups naturally after mating and they were necropsied with their offspring on the 22nd day of lactation. Both males and females continued to be administered until the day before necropsy. General conditions, clinical signs, changes in body weight gain and food intake of parental animals were observed during the administration period. The effects of the test substance on male and female reproductive performance including gonadal function, oestrus cycle, mating behaviour, conception, parturition and lactation and weaning were followed. The early neonatal survival and the growth of offspring were also recorded. Transient salvation and decreased locomotor activity were found in all administration groups in both sexes, although their incidence increased by the dose thus, they were regarded as treatment-related clinical signs. Nasal discharge, lean and prone position, ptosis (eyelid closure) and lacrimation were also observed in both males and females in various dose groups, although these effects did not show dose-dependence in their frequency or severity. In males, there were no significant differences in either body weight or food consumption between the control and the test substance administrated groups. In females, no significant difference was found between the control and any of the treated groups regarding body weight before mating and during pregnancy and only slight decrease in the postpartum body weight were seen in the high-dose (160 mg/kg) group. In females, the food consumption dropped significantly in lactation day 4-21, in the 160 mg/kg dose group. No significant changes were seen in oestrus cyclicity, conception rate and the number of days required from the start of cohabitation to copulation in any of the treatment females in comparison with the controls. As for parturition, single cases of abnormalities were observed in the two highest dose groups and in one case within the 160 mg/kg dose group, all offspring died due to complicated labour. However, no abnormalities were found in the delivery status of the other animals, and there were no significant differences in the birth rate and gestation period between the control and any of the treated groups. The reproductive performance described by the copulation and fertility indexes of both males and females were comparable with controls. Degeneration of seminiferous cells localized to seminiferous tubules in the 14th stage was observed in a few cases in the 40 and 160 mg/Kg dosed animals and in many of the cases epididymal cell debris were observed in the lumen in cases where fertility was confirmed. However, for these findings, no significant difference was found between the control group and each test substance administration group. Similarly, no differences between control and treated groups were seen regarding the rearing conditions. The lack of nursing behaviour was noticed at one female of 160 mg/kg group which led to the death of the litter, but no abnormal nursing behavioural were observed in other animals. Pathological examination of the reproductive organs revealed no treatment-related and dose-dependent changes in male and female gonads at any dose level. Very slight and slight degeneration of spermatids at the 14thstage of spermatogenesis in the seminiferous tubule of testis were observed in all treated groups. However, this impaired spermatogenesis was detected at low frequency of occurrence and severity. Moreover, impaired spermatogenesis was described occurring spontaneously in SD rats used in this study. There was no dose-dependent and test chemical-related changes in female reproductive organs. There was no significant difference in the number of implantations in the uterus between the control and any of the administrated groups. Squamous hyperplasia of the anterior gastric mucosa was observed in the 40 and 160 mg/kg groups were the only significant and treatment-related findings in males during necropsy, although in females, no effect was observed in any tissues. In the F1 generation, skin abnormalities (peeling off the skin) were seen at the high-dose (160 mg/kg) group from Day 0. In addition, the number of live pups was significantly lower in the 160 mg/kg group at lactation day 4 and 7 as compared to controls and the viability index at lactation day 4 was significantly lower than those of control in the same group. Moreover,no significant differences were found between the control and any of the treated groups regarding external and visceral malformations and the total number of the offspring showing morphological abnormalities was comparable with controls in frequency.As the birth weight was equal to that in the control group, it was considered that there was no growth suppression in utero either. In addition, no neonatal growth failure was observed among pups. Thus, it was postulated that the significant increase in death during the early neonatal period either caused by a direct effect of the test chemical on offspring viability or by the slightly suppressed maternal lactating activity due to the mild hypnotic effect of 2-naphthol. In summary, no abnormality was found infertility, reproductive function and parturition of parental animals, but the prenatal exposure to 2-naphthol produced adverse effects on neonatal growth and viability at the concentration of 160 mg/kg or more which occurred in the absence of severe maternal toxicity. Hence, the developmental NOAEL for maternal and F1 generation was concluded as 40 mg/kg due to the decreased neonatal survival rate.

 

This is further supported by one-generation reproductive toxicity in rats (MHW Report, 2012) according to the OECD TG 415 using the analogue substance in male and female Sprague-Dawley rats. The test chemical was given daily at the doses of 0, 12.5, 50 and 200 mg/kg to SD rats (25 animals/group/sex) by gavage for 14 weeks (98 days). Males were dosed 10 weeks before mating, during mating and they were necropsied one week after the end of the mating period. Females received the test chemical 2 weeks before mating, during 3 weeks of mating and pregnancy till the 21stday of lactation (overall 98 days). The dams were allowed to deliver naturally after mating then were necropsied with their offspring on the 21stday of lactation. General conditions, clinical signs, changes in body weight gain and food intake of parental animals were observed during the administration period. The effects of the test substance on male and female reproductive performance including gonadal function, oestrus cycle, mating behaviour, conception, parturition and lactation and weaning of pups were followed. The early neonatal survival and the growth of offspring were also recorded. Post-mortem examinations of parenteral animals were carried out to inspect the major organs in the thoracic-abdominal area including stomach, liver, spleen, kidney, pituitary and adrenal glands, and reproductive organs such as testis, epididymis, coagulated glands, seminal vesicles, prostate, ovaries, uterus and vagina. Necropsy of F1 pups was also performed and the presence or absence of external and visceral abnormalities were observed. Three males from the 0, 12.5 and 50 mg/kg groups died during treatment, but those were not attributable to the test chemical administration. No test chemical-related deaths were recorded in other animals. Salvation and nasal discharge were observed in almost all males and in few females in the 200 mg/kg group and because they occurred concomitantly with the test chemical administration were considered as treatment-related changes. Other clinical signs, such as, red-brown contamination of the eyelid, hair loss, skin scrub formation were noted in various dose levels including controls and their incidence remained low. In males, there was no significant difference in food consumption and body weight between the control and treated groups, but with regard the body weight gain slight suppression in the 200 mg/kg dose group was observed at 64-71, 85-92 and 92-99 days. There was no significant difference in weight gain between the 12.5 mg/kg group and the control group. In females, no significant difference in food consumption between the control and any of the treated group was seen. However, the body weight gain at the beginning of dosing, during late pregnancy and lactation period was suppressed in the 200 mg/kg group. No significant changes in oestrus cycle after treatment were seen in any dose group. The reproductive performance of male and female rats was not affected by the repeated administration of 3-hydroxy-2-naphthoic acid as it was evidenced by the Copulation and Fertility indexes and the number of pairing days until copulation. No abnormal parturition and nursing behaviour were recorded in any dose group and no significant differences in the birth rate and pregnancy period between the control group and treated groups was observed. Prior to necropsy, blood samples were collected from treated males and a significant increase in red blood cell count (RBC) were detected at 50 mg/kg and more. When the white blood cell count (WBC) was compared to control values, significant decreases were seen in all dosed groups. However, the histopathological examinations of bone marrow, liver and spleen tests did not show any changes related to myeloid leukemia, and it was decided that administration of the test chemical was not involved in myelogenous leukemia that occurred in each of the low and medium dose groups. However, the increase in RBC values were depending on the dose of the test substance administered, thus it is considered as treatment-related changes, and concluded that 50 mg/kg or more of test chemical increases the RBC in males rats. In males, a moderately thickened area of the anterior gastric mucosa was observed in the 200 mg/kg dose group at necropsy, and slight hyperplasia of the anterior gastric mucosal squamous epithelium was observed at 50 mg/kg or more by histopathological examination. The swelling of liver was also detected in the highest dose groups, but no clear tissue change associated with swelling was observed. The histopathological examination of bone marrow, spleen, pituitary and adrenal gland showed no change attributable to test substance administration. No changes were observed in male reproductive organs such as testis, epididymis, prostate ventral lobe and adrenal gland in any dose group. In females, a thickened area of anterior gastric mucosa was observed in only one case at 200 mg/kg at necropsy and histopathological examination showed squamous hyperplasia of anterior gastric mucosa in few cases in the high-dose group. However, no histopathological findings attributable to the test chemical administration were seen in adrenal and pituitary gland, uterus and oviduct. It is suggested that the test chemical is irritating to the skin and mucous membrane and it is speculated that tissue change of anterior gastric mucosa is due to the irritant nature of the test chemical. In pups of F1 generation, no alterations in general conditions were seen at any dose level. The number of live pups at day 0, 7,14, and 21 were similar to that of controls. However, the weights of offspring were significantly less in the 200 mg/kg group during the entire nursing period (21 days) for both sexes.The necropsy of pups at lactation day 4 and 21 revealed slightly increased incidence of external malformations (bent-tail, short-tailed, short-curvature tail or small eyes) in the 200 mg/kg group. As the total number of external malformations observed until weaning was significantly higher than those in control, the teratogenic effect of the test chemical at concentration of 200 mg/kg or more could not be ruled out. Only few cases of visceral malformations (malposition of testis, hypoplasia of spleen, diaphragmatic hernia) were observed in the 200 mg/kg group and they remained at low frequency.In summary, no abnormality was found infertility, reproductive function and parturition of parental animals but the prenatal exposure to 3-hydroxy-2-naphthoic acid significantly reduced the weight of offspring from birth till weaning at the concentration of 200 mg/kg. It is possible that the test substance or its metabolite was transferred to milk and it had a direct effect on birth weight of pups. However, it is also possible that the test chemical at the concentration of 200 mg/kg had adverse effect on milk production in the mothers, since the mother body weight gain is also suppressed during the early and middle stages of lactation, when milk production is increasing. Hence, the developmental NOAEL for maternal and F1 generation was concluded as 50 mg/kg due to the decreased weight of pups from birth till the end of the nursing period.

Supporting the above details, A prenatal developmental toxicity study of the test chemical was performed according to OECD guideline 414 on New-Zealand white rabbits. Young adult nulliparous New-Zealand white strain female and male rabbits (Oryctolagus cuniculus) were used in study. The test material was dissolved in 0.5% carboxymethyl cellulose in the concentrations of 0, 3, 10 and 50 mg/kg/day and administered by daily gavage through gestation day 6 to 28 to mated females (25/dose group). Based on a preliminary range-finding study finding, 0, 3, 10 and 50 mg/kg/day were selected as final doses for the main study. Animals were observed twice daily for moribundity and mortality. Individual clinical signs were recorded at least once a day during the treatment period and once daily during the pre- and post-treatment periods. Individual body weights were recorded on gestation days 0, 3, 5, 8, 11, 14, 17, 20, 23, 26, 29 and 30. Food consumption per cage of animals were measured over the following periods during gestation: days 0-3, 3- 5, 5-8, 8-11, 11-14, 14-17, 17-20, 20-26, 26-29 and 29-30. On day 29 of gestation, all the rabbits were sacrificed and examined for gross pathological changes in all the visceral organs. Organs with pathological findings and the kidneys were preserved for further histological evaluation. The ovaries, uteri and uterine contents were removed and examined to determine: the number of corpora lutea, the number of implantations, early and late resorptions, the weight of intact gravid uterus, the number and distribution of live foetuses, the number and distribution of intra-uterine dead fetuses, the individual fetal weight and sex and foetal abnormalities. There was no maternal death at any dose level. There was a significant reduction in the maternal body weight gain during the treatment period in 50 mg/kg group, although the food consumption was comparable to the vehicle control group. This reduction in body weight during the treatment period was considered treatment related. The maternal data parameters comprising of early and late resorptions, pre and post-implantation loss in all the treatment groups were comparable to the vehicle control group. The mean number of corpora lutea, and the mean litter size were significantly lower in the high-dose group when compared with the control group, but the observed decrease in corpora lutea at 50 mg/kg/day is considered as biological variation because the treatment was initiated after the implantation (gestation day 6). Therefore, the decrease observed in the absolute uterine weight, implantation and live foetus reported at this dose level are also considering as biological variation as these observations are directly correlated with the decrease in the number of the corpora lutea. In addition, no changes in foetus weight and in sex ratio were seen at any dose level. In conclusion, no maternal toxicity was observed up to the dose level of 10 mg/kg body weight/day in New Zealand white rabbits following the orally administration of the test chemical during gestation days 6 to 28. Hence, the NOAEL for developmental toxicity was set at 50 mg/kg body weight/day as no treatment-related effects on foetus development were observed.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

40 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

The data is K2 level and provides robbust summary

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Various reproductive/developmental toxicity studies have been investigated to observe the adverse reproductive effects occurring as a result of repeated daily oral dosing with, or exposure, to a substance for a specified period up to the expected lifespan of the test species. Often are the studies based on experiments of 60% and above structurally and functionally similar read across analogues. The studies are summarized as below: 

The study is ongoing and the template will be updated later after the completeion of the study.

One-generation reproductive toxicity in rats (MHW Report, 2001) as per OECD TG 415 using the analogue substance. Groups of 25 animals/dose/sex were treated with 0, 10, 40, 160 mg/kg of test chemical by gavage for 14 weeks (98 days). Males were dosed 10 weeks before mating, during mating and they were necropsied one week after the end of the mating period. Females received the test chemical 2 weeks before mating, during 3 weeks of mating and pregnancy till the 21stday of lactation (overall 98 days). Dams were allowed to deliver pups naturally after mating and they were necropsied with their offspring on the 22nd day of lactation. Both males and females continued to be administered until the day before necropsy. General conditions, clinical signs, changes in body weight gain and food intake of parental animals were observed during the administration period. The effects of the test substance on male and female reproductive performance including gonadal function, oestrus cycle, mating behaviour, conception, parturition and lactation and weaning were followed. The early neonatal survival and the growth of offspring were also recorded. Transient salvation and decreased locomotor activity were found in all administration groups in both sexes, although their incidence increased by the dose thus, they were regarded as treatment-related clinical signs. Nasal discharge, lean and prone position, ptosis (eyelid closure) and lacrimation were also observed in both males and females in various dose groups, although these effects did not show dose-dependence in their frequency or severity. In males, there were no significant differences in either body weight or food consumption between the control and the test substance administrated groups. In females, no significant difference was found between the control and any of the treated groups regarding body weight before mating and during pregnancy and only slight decrease in the postpartum body weight were seen in the high-dose (160 mg/kg) group. In females, the food consumption dropped significantly in lactation day 4-21, in the 160 mg/kg dose group. No significant changes were seen in oestrus cyclicity, conception rate and the number of days required from the start of cohabitation to copulation in any of the treatment females in comparison with the controls. As for parturition, single cases of abnormalities were observed in the two highest dose groups and in one case within the 160 mg/kg dose group, all offspring died due to complicated labour. However, no abnormalities were found in the delivery status of the other animals, and there were no significant differences in the birth rate and gestation period between the control and any of the treated groups. The reproductive performance described by the copulation and fertility indexes of both males and females were comparable with controls. Degeneration of seminiferous cells localized to seminiferous tubules in the 14th stage was observed in a few cases in the 40 and 160 mg/Kg dosed animals and in many of the cases epididymal cell debris were observed in the lumen in cases where fertility was confirmed. However, for these findings, no significant difference was found between the control group and each test substance administration group. Similarly, no differences between control and treated groups were seen regarding the rearing conditions. The lack of nursing behaviour was noticed at one female of 160 mg/kg group which led to the death of the litter, but no abnormal nursing behavioural were observed in other animals. Pathological examination of the reproductive organs revealed no treatment-related and dose-dependent changes in male and female gonads at any dose level. Very slight and slight degeneration of spermatids at the 14thstage of spermatogenesis in the seminiferous tubule of testis were observed in all treated groups. However, this impaired spermatogenesis was detected at low frequency of occurrence and severity. Moreover, impaired spermatogenesis was described occurring spontaneously in SD rats used in this study. There was no dose-dependent and test chemical-related changes in female reproductive organs. There was no significant difference in the number of implantations in the uterus between the control and any of the administrated groups. Squamous hyperplasia of the anterior gastric mucosa was observed in the 40 and 160 mg/kg groups were the only significant and treatment-related findings in males during necropsy, although in females, no effect was observed in any tissues. In the F1 generation, skin abnormalities (peeling off the skin) were seen at the high-dose (160 mg/kg) group from Day 0. In addition, the number of live pups was significantly lower in the 160 mg/kg group at lactation day 4 and 7 as compared to controls and the viability index at lactation day 4 was significantly lower than those of control in the same group. Moreover,no significant differences were found between the control and any of the treated groups regarding external and visceral malformations and the total number of the offspring showing morphological abnormalities was comparable with controls in frequency.As the birth weight was equal to that in the control group, it was considered that there was no growth suppression in utero either. In addition, no neonatal growth failure was observed among pups. Thus, it was postulated that the significant increase in death during the early neonatal period either caused by a direct effect of the test chemical on offspring viability or by the slightly suppressed maternal lactating activity due to the mild hypnotic effect of 2-naphthol. In summary, no abnormality was found infertility, reproductive function and parturition of parental animals, but the prenatal exposure to 2-naphthol produced adverse effects on neonatal growth and viability at the concentration of 160 mg/kg or more which occurred in the absence of severe maternal toxicity. Hence, the developmental NOAEL for maternal and F1 generation was concluded as 40 mg/kg due to the decreased neonatal survival rate.

 

This is further supported by one-generation reproductive toxicity in rats (MHW Report, 2012) according to the OECD TG 415 using the analogue substance in male and female Sprague-Dawley rats. The test chemical was given daily at the doses of 0, 12.5, 50 and 200 mg/kg to SD rats (25 animals/group/sex) by gavage for 14 weeks (98 days). Males were dosed 10 weeks before mating, during mating and they were necropsied one week after the end of the mating period. Females received the test chemical 2 weeks before mating, during 3 weeks of mating and pregnancy till the 21stday of lactation (overall 98 days). The dams were allowed to deliver naturally after mating then were necropsied with their offspring on the 21stday of lactation. General conditions, clinical signs, changes in body weight gain and food intake of parental animals were observed during the administration period. The effects of the test substance on male and female reproductive performance including gonadal function, oestrus cycle, mating behaviour, conception, parturition and lactation and weaning of pups were followed. The early neonatal survival and the growth of offspring were also recorded. Post-mortem examinations of parenteral animals were carried out to inspect the major organs in the thoracic-abdominal area including stomach, liver, spleen, kidney, pituitary and adrenal glands, and reproductive organs such as testis, epididymis, coagulated glands, seminal vesicles, prostate, ovaries, uterus and vagina. Necropsy of F1 pups was also performed and the presence or absence of external and visceral abnormalities were observed. Three males from the 0, 12.5 and 50 mg/kg groups died during treatment, but those were not attributable to the test chemical administration. No test chemical-related deaths were recorded in other animals. Salvation and nasal discharge were observed in almost all males and in few females in the 200 mg/kg group and because they occurred concomitantly with the test chemical administration were considered as treatment-related changes. Other clinical signs, such as, red-brown contamination of the eyelid, hair loss, skin scrub formation were noted in various dose levels including controls and their incidence remained low. In males, there was no significant difference in food consumption and body weight between the control and treated groups, but with regard the body weight gain slight suppression in the 200 mg/kg dose group was observed at 64-71, 85-92 and 92-99 days. There was no significant difference in weight gain between the 12.5 mg/kg group and the control group. In females, no significant difference in food consumption between the control and any of the treated group was seen. However, the body weight gain at the beginning of dosing, during late pregnancy and lactation period was suppressed in the 200 mg/kg group. No significant changes in oestrus cycle after treatment were seen in any dose group. The reproductive performance of male and female rats was not affected by the repeated administration of 3-hydroxy-2-naphthoic acid as it was evidenced by the Copulation and Fertility indexes and the number of pairing days until copulation. No abnormal parturition and nursing behaviour were recorded in any dose group and no significant differences in the birth rate and pregnancy period between the control group and treated groups was observed. Prior to necropsy, blood samples were collected from treated males and a significant increase in red blood cell count (RBC) were detected at 50 mg/kg and more. When the white blood cell count (WBC) was compared to control values, significant decreases were seen in all dosed groups. However, the histopathological examinations of bone marrow, liver and spleen tests did not show any changes related to myeloid leukemia, and it was decided that administration of the test chemical was not involved in myelogenous leukemia that occurred in each of the low and medium dose groups. However, the increase in RBC values were depending on the dose of the test substance administered, thus it is considered as treatment-related changes, and concluded that 50 mg/kg or more of test chemical increases the RBC in males rats. In males, a moderately thickened area of the anterior gastric mucosa was observed in the 200 mg/kg dose group at necropsy, and slight hyperplasia of the anterior gastric mucosal squamous epithelium was observed at 50 mg/kg or more by histopathological examination. The swelling of liver was also detected in the highest dose groups, but no clear tissue change associated with swelling was observed. The histopathological examination of bone marrow, spleen, pituitary and adrenal gland showed no change attributable to test substance administration. No changes were observed in male reproductive organs such as testis, epididymis, prostate ventral lobe and adrenal gland in any dose group. In females, a thickened area of anterior gastric mucosa was observed in only one case at 200 mg/kg at necropsy and histopathological examination showed squamous hyperplasia of anterior gastric mucosa in few cases in the high-dose group. However, no histopathological findings attributable to the test chemical administration were seen in adrenal and pituitary gland, uterus and oviduct. It is suggested that the test chemical is irritating to the skin and mucous membrane and it is speculated that tissue change of anterior gastric mucosa is due to the irritant nature of the test chemical. In pups of F1 generation, no alterations in general conditions were seen at any dose level. The number of live pups at day 0, 7,14, and 21 were similar to that of controls. However, the weights of offspring were significantly less in the 200 mg/kg group during the entire nursing period (21 days) for both sexes.The necropsy of pups at lactation day 4 and 21 revealed slightly increased incidence of external malformations (bent-tail, short-tailed, short-curvature tail or small eyes) in the 200 mg/kg group. As the total number of external malformations observed until weaning was significantly higher than those in control, the teratogenic effect of the test chemical at concentration of 200 mg/kg or more could not be ruled out. Only few cases of visceral malformations (malposition of testis, hypoplasia of spleen, diaphragmatic hernia) were observed in the 200 mg/kg group and they remained at low frequency.In summary, no abnormality was found infertility, reproductive function and parturition of parental animals but the prenatal exposure to 3-hydroxy-2-naphthoic acid significantly reduced the weight of offspring from birth till weaning at the concentration of 200 mg/kg. It is possible that the test substance or its metabolite was transferred to milk and it had a direct effect on birth weight of pups. However, it is also possible that the test chemical at the concentration of 200 mg/kg had adverse effect on milk production in the mothers, since the mother body weight gain is also suppressed during the early and middle stages of lactation, when milk production is increasing. Hence, the developmental NOAEL for maternal and F1 generation was concluded as 50 mg/kg due to the decreased weight of pups from birth till the end of the nursing period.

Supporting the above details, A prenatal developmental toxicity study of the test chemical was performed according to OECD guideline 414 on New-Zealand white rabbits. Young adult nulliparous New-Zealand white strain female and male rabbits (Oryctolagus cuniculus) were used in study. The test material was dissolved in 0.5% carboxymethyl cellulose in the concentrations of 0, 3, 10 and 50 mg/kg/day and administered by daily gavage through gestation day 6 to 28 to mated females (25/dose group). Based on a preliminary range-finding study finding, 0, 3, 10 and 50 mg/kg/day were selected as final doses for the main study. Animals were observed twice daily for moribundity and mortality. Individual clinical signs were recorded at least once a day during the treatment period and once daily during the pre- and post-treatment periods. Individual body weights were recorded on gestation days 0, 3, 5, 8, 11, 14, 17, 20, 23, 26, 29 and 30. Food consumption per cage of animals were measured over the following periods during gestation: days 0-3, 3- 5, 5-8, 8-11, 11-14, 14-17, 17-20, 20-26, 26-29 and 29-30. On day 29 of gestation, all the rabbits were sacrificed and examined for gross pathological changes in all the visceral organs. Organs with pathological findings and the kidneys were preserved for further histological evaluation. The ovaries, uteri and uterine contents were removed and examined to determine: the number of corpora lutea, the number of implantations, early and late resorptions, the weight of intact gravid uterus, the number and distribution of live foetuses, the number and distribution of intra-uterine dead fetuses, the individual fetal weight and sex and foetal abnormalities. There was no maternal death at any dose level. There was a significant reduction in the maternal body weight gain during the treatment period in 50 mg/kg group, although the food consumption was comparable to the vehicle control group. This reduction in body weight during the treatment period was considered treatment related. The maternal data parameters comprising of early and late resorptions, pre and post-implantation loss in all the treatment groups were comparable to the vehicle control group. The mean number of corpora lutea, and the mean litter size were significantly lower in the high-dose group when compared with the control group, but the observed decrease in corpora lutea at 50 mg/kg/day is considered as biological variation because the treatment was initiated after the implantation (gestation day 6). Therefore, the decrease observed in the absolute uterine weight, implantation and live foetus reported at this dose level are also considering as biological variation as these observations are directly correlated with the decrease in the number of the corpora lutea. In addition, no changes in foetus weight and in sex ratio were seen at any dose level. In conclusion, no maternal toxicity was observed up to the dose level of 10 mg/kg body weight/day in New Zealand white rabbits following the orally administration of the test chemical during gestation days 6 to 28. Hence, the NOAEL for developmental toxicity was set at 50 mg/kg body weight/day as no treatment-related effects on foetus development were observed.

In another OECD 422 combined repeated dose toxicity and reproduction/developmental screening study, groups of 10 Sprague Dawley rats/sex/dose were administered acetophenone at doses of 0, 75, 225, or 750 mg/kg/day daily via gavage for a minimum of 14 days before mating and throughout the mating and gestation periods up to lactation day 3. There was a significant increase in the number of stillborn offspring among the high-dose group as compared to controls. There was a significant increase in the number of offspring dying, missing and/or cannibalized, along with an increase in the number of litters with total litter loss among the high-dose group during lactation days 1–4. There was a significant decrease in the total number of live born, viability index, and mean number of live pups per litter on lactation days 1–4. The number of mean live pups per litter was significantly lower on lactation days 1–4, and the live birth index was also reported to be out of the historical control range. Clinical signs among the high-dose group offspring included increased incidences of desquamation, cool to the touch, skin with shiny appearance, skin appearing tight with restricted movement, and a slightly increased incidence of gasping and pale skin color. There was a significant decrease in the pup weight per litter among the high-dose group on lactation days 1 and 4; this was reported to be out of the historical control ranges. During gross pathological examination of offspring, high-dose group pups were reported with incidences of cleft palate and edema, atelectasis, dermal hypoplasia, scabbing, desquamation, and 22 dead pups with observed autolysis. Thus, the NOAEL for the developmental toxicity endpoint was considered to be 225 mg/kg/day, based on effects of treatment on viability of the offspring, alterations in clinical signs, body weight, and gross pathological alterations among the high-dose group offspring

Justification for classification or non-classification

The study is ongoing and the template will be updated later after the completeion of the study.