m-phenylenebis(methylamine)

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

No data

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Remarks:

based on test type (migrated information)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1998-12-28 to 1999-03-01

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

Deviations:

yes

Remarks:

temperature or humidity of the animal room deviated from the set range on two days during the study period. However, each deviation was slight and for a short time, and was judged not to have affected the integrity of the study.

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Nippon Charles River Co Ltd (Atsugi-shi, Kanagawa)
- Age at study initiation: 10 weeks
- Weight at study initiation: 354-399 g (males); 216-247 g (females)
- Housing: Barrier sustained animal room (width 5.7 x depth 10.0 x height 2.5 m). Animals were housed individually in a cage with aluminium front and stainless steel mesh floor (width 15.8 x depth 23.8 x height 16.0 cm). Cage specifications were varied during the mating period and, from day 18 of pregnancy, dams were housed individually in a cage with an aluminium front and stainless steel mesh floor (width 36.8 x depth 25.0 x height 16.0 cm) with a lactation tray and nursery material. Cages were exchanged every other week and feeding trays were exchanged weekly.
- Diet: Free access to NMF pellet diet (sterilized by radiation) manufactured by Oriental Industries (Chuo-ku, Tokyo) administered via a nozzle
- Water: Free access to tap water administered via a nozzle
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature: 24 ± 3 degrees Centigrade
- Humidity: 55 ± 20 degrees Centigrade
- Air changes: 15 per hour
- Illuminance (lux): 150-300
- Photoperiod: 12 hours per day (07:00 to 19:00)

Route of administration:

oral: gavage

Type of inhalation exposure (if applicable):

other: Not applicable

Vehicle:

other: Official purified water (Lot 1051)

Details on exposure:

- Pre-treatment observation: 2 weeks before administration began
- Confirmation of childbirth was conducted at 09:00 to 10:00 and day 0 of lactation applied accordingly.
- Post-treatment observation: 1 day for adult animals. Offspring were observed until day 4 of lactation.

Details on mating procedure:

Females whose vaginal smear was examined for 14 days before mating were housed with males of the same group on a 1:1 basis for a maximum of 5 days. Copulation was judged to be successful by confirming the presence of sperms in the vaginal smear the next morning, designated day 0 of pregnancy.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The test substance was dissolved in official purified water (Lot 1051 obtained from Kyouei Pharmaceutical Co Ltd) and adjusted to make 5, 15 and 45 mg/L solutions for each group. After preparation, the solution was kept in the dark under room temperature until use. In addition, the test substance solution at concentrations of 1 and 60 mg/L were confirmed in a pilot study to be stable for 8 days at room temperature after preparation.

The solution for each dosing group was analyzed before the start of administration. The results showed that concentrations were between 99.2 and 106.9 % of indicated concentration. Therefore it was confirmed that each dose solution contained the prescribed quantity of 1,3-benzenedimethanamine. Further analysis, conducted by the manufacturer after the administration period, showed the purity was 99.8% and confirmed that the test substance was stable during the administration period.

Duration of treatment / exposure:

- Males: Duration of administration was 48 consecutive days (14 days before mating, 14 days during mating and 20 days after the end of the mating period.
- Females: Duration of exposure was 14 days before mating, during the mating period (maximum 5 days), throughout the gestation period after copulation and 3 days of lactation after parturation (41-45 days). In females which did not deliver after copulation, administration was completed for 40 days until the day before necropsy.
- Offspring: Not dosed

Frequency of treatment:

Seven days per week

Details on study schedule:

Animals were classified according to their body weight and allocated to each group by random selection. Twelve males and 12 females per group were prepared. All females were observed for estrus cycle for 8 days before allocation, and those animals which had normal estrus cycle were used for allocation. Identification of animals after group allocation was done by tattooing an individual number in the ear auricle and by attaching a card with animal ID number on each cage.

Remarks:

Doses / Concentrations:
0 mg/kg/day
Basis:
actual ingested
12 males and 12 females

Remarks:

Doses / Concentrations:
50 mg/kg/day
Basis:
actual ingested
12 males and 12 females

Remarks:

Doses / Concentrations:
150mg/kg/day
Basis:
actual ingested
12 males and 12 females

Remarks:

Doses / Concentrations:
450mg/kg/day
Basis:
actual ingested
12 males and 12 females

No. of animals per sex per dose:

Twelve animals of each sex per dose level

Control animals:

yes, concurrent vehicle

Details on study design:

In a 28-day repeated dose toxicity study, post administration signs such as salivation, decrease in locomotion activity, piloerection, and abdominal distention were seen in males and females of the 600 mg/kg group, and decrease in food intake and supression of body weight gain were seen in males of the 600 mg/kg group. One male and 4 females out of 12 rats in the group died. In addition, ulceration and epithelial hyperplasia associated with hyperkeratosis were seen in the fore-stomach, and myeloid hyperplasia in the bone marrow, hypertrophy and vacuolation of adrenal cortical cells, and dilation of the cecum were seen in males and females of this group. Based on these results, a two week pilot study (number 4211) was conducted using the same doses as those in the 28-day repeated dose toxicity study (0, 10, 40, 150 and 600 mg/kg). During the pilot study, 2 out of 6 males and 1 out of 6 females died in the 600 mg/kg group. Salivation in males and females, suppression of body weight gain in males, and increase in adrenal weight were also observed in this group after administration of the test substance.

Since male and female animals died in the 600 mg/kg group and planned administration was three times longer than in the pilot study, 450 mg/kg was selected as the highest dose, and 150 and 50 mg/kg, dividing by a common ratio of 3, were applied.

Enforced oral administration was used and the administration volume was set as 1 mL per 100 g body weight. The exact volume was calculated using the latest body weight available from measurements taken during pre-mating and mating periods in males and females. In addition, during the lactation period, volume was calculated based on body weight obtained on day zero of lactation. Enforced oral administration was conducted once daily on seven days per week using a stomach cannula. Purified water of official grade was similarly administered to the control group.

Positive control:

No data

Parental animals: Observations and examinations:

All male and female animals were observed daily during the study period.

Body weight
Males were weighed on days 1 (start of study) 8, 15, 22, 29, 36, 43 and 49 (necropsy day). The body weight gain from days 1 to 49 was calculated. Females were weighed on days 1 (start of study), 8 and 15, and the body weight gain from day 1 to day 49 was calculated. In addition, females with successful copulation were weighed on days 0, 7, 14 and 21 of gestation, and females after parturition were weighed on days 0 and 4 of lactation. Body weight gains from day 0 to day 21 of gestation and from day 0 to day 4 of lactation were calculated.

Food intake
For males, the food was weighed on days 1 (start of study), 8, 15, 22, 29, 36, 43, and 48 (the day before autopsy), and the mean intake was calculated from intake between a certain day and the next measuring day. In addition, cummulative food intakes from day 1 to day 15 and from day 22 to day 48 were calculated. In females, the food was weighed on days 1 (start of study), 8 and 15 and the mean daily food intake was calculated from intake between a certain day and the next measuring day. In addition, cummulative food intake from day 1 to 15 was calculated. In females with successful copulation, food was weighed on days 0, 7, 14 and 21 of gestation. In females with parturition, food was weighed on days 0 and 4, and mean daily food intake was calculated from intake between a certain day and the next measuring day. In addition, cummulative food intake from day 0 to day 21 of gestation was calculated. Food intake of cohabiting animals was not measured during the mating period.

Oestrous cyclicity (parental animals):

Observation of estrus cycle was continued until the day of successful copulation. The number of days between one estrus and the next were taken as duration of estrus cycle, and the mean duration of estrus cycle was calculated. In addition, the incidence [(number of animals with abnormal estrus cycle/number of animals observed)/100] of abnormal estrus cycle (estrus cycle other than 4 or 5 days) was calculated.

Sperm parameters (parental animals):

No data

Litter observations:

Observations were taken daily for gestation period and evidence of delivery.

Postmortem examinations (parental animals):

Organs examined at necropsy (macroscopic and microscopic)
- Organ weights in males: Thymus, adrenal, testes and epididymides
- Organ weights in females: Thymus, adrenal
- Histopathology: Thymus, stomach, adrenal, ovaries, uterus, vagina, testes, epididymides, abnormal site of spleen from two animals, nasal wall from one animal, small intestine from three animals, large intestine from five animals, seminal vesicle from one animal, liver from one animal and lung from one animal.

Other examinations
- Inspected at necropsy: Appearance of the oral cavity, nostril, cranial cavity, skeleton, external appearance and dissected face of brain and spinal cord, thoracic cavity, abdominal cavity and pelvic cavity with viscera, cervial tissues and organs. All abnormalities noted during gross examination were noted together with location, size, hardness and other pertinent details.

Postmortem examinations (offspring):

No histopathology was carried out on offspring

Statistics:

A multiple comparison test was applied to data on body weight, food intake, number of corpora lutea, implantation site, number of offspring, number of offspring dead, sex ratio, mean length of estrus cycle, duration of pregnancy, implantation rate, delivery rate, birth rate, ratio of external abnormalities, ratio of offspring alive on day 4 after birth, organ weight, and organ weight/body weight ratio (relative organ weight).

Firstly homogeneity of variance was analysed by Bartlett's test. When the variance was homogenous, significant differences between the control group and each treated groupwere analysed by Dunnett's multi-comparison test. When variance was heterogenous, significant differences between the control group and each treated group were analysed by Steel's test. For comparison of birth rate, copulation index and fertility index, a chi square test was used. The incidence of abnormal estrus cycle and the incidence of pathology findings were analysed by Fisher's direct probability test. Those histopathological findings which increased in severity in the treated groups were ranked and then analysed by Mann-Whitney's U test. In addition, for the result concerning live offspring during the lactation period, a mean value per one dam was used as one sample. Statistical analysis at 5 % and 1 % significance were performed by a two-tail test.

Reproductive indices:

Copulation index [(number of animals with successful copulation/number of mated animals) x 100] was calculated. A female which died on day 2 of pregnancy was excluded from the summing up of fertility index because it was impossible to judge whether that female was prenant. A male was examined only on histopathology when considering males who failed to impregnate but was included in summing up when considering males who succeeded in impregnation.

Offspring viability indices:

No data

Clinical signs:

effects observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Other effects:

effects observed, treatment-related

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

General observations

In the groups treated with 150 mg/kg or more, salivation and nasal noise was seen in male animals. Ulceration of the fore-stomach and adhesion to surrounding tissue was seen in males from the 450 mg/kg group.

In the 450 mg/kg group, salivation and nasal noise were seen in female animals. In addition, suppression of bodyweight gain, reduced food intake, and ulceration of the fore-stomach were observed during necropsy of females.

Toxic response/effects by dose level

Death and general signs

One male animal from the 150 mg/kg group died on Day 38. Three further males from the 450 mg/kg group died on Days 31, 38 and 39. One female in the 450 mg/kg group died on day 2 of pregnancy.

Dose dependent salivation was the most common observation and was noted in males and females from the 150 mg/kg and 450 mg/kg groups. Salivation observed in a few animals in weeks 5 and 6 and in half the animals in week 7 of administration was a transient change, which appeared immediately after dosing and disappeared 30 minutes later. In males of the 450 mg/kg group, salivation was observed in around half of the animals and most of the animals in week two or later. In week 1, salivation appeared immediately after administration and disappeared after 30 minutes. However, the effect lasted longer as administration continued, being continuously observed at 90 to 180 minutes after administration during the latter stages of the study. In one female of the 150 mg/kg group, salivation was seen on day 2 of pregnancy, with the same transience as observed in males. In females of the 450 mg/kg group, salivation was observed in most animals at pre-mating and mating stages plus the early stages of pregnancy. A gradual decrease in the number of animals exhibiting salivation was found in the middle to late stages of pregnancy. Only one animal showed salivation during the lactation period after delivery. In a few females from the 450 mg/kg group, salivation appeared immediately after administration and lasted for 60 minutes during the gestation period. During other periods, salivation appeared immediately after administration and disappeared 30 minutes later.

In addition, the following observations were made: nasal noise (1 male, 150 mg/kg group and 3 males and females, 450 mg/kg group), irregular respiration (1 male, 150 mg/kg group), abdominal distension, emaciation, staining of fur, piloerection, hypothermia (1 male, 150 mg/kg group and 1 female, 450 mg/kg group), nasal discharge (1 male and 1 female, 450 mg/kg group), pale appearance (1 male, 150 mg/kg group and 2 females, 450 mg/kg group), ptosis (1 male, 150 mg/kg group plus 2 males and 1 female, 450 mg/kg group), prone position (1 female, 450 mg/kg group).

Spontaneous changes such as eye discharge (1 male, control group) and alopecia (1 female, 150 mg/kg group plus 1 male and 1 female, 450 mg/kg group) were also observed.

Bodyweight

Bodyweights of males from the 450 mg/kg group were smaller than those of the control group on Day 8 and thereafter. Bodyweight gain (62 ± 20 g) was also significantly less than the control group (129 ± 26 g).

Females from the 450 mg/kg group had bodyweights on day 14 and 21 of pregnancy that were less than the control group. Bodyweight gain during gestation (140 ± 19 g) was also significantly less than the control group (170 ± 31 g).

Food intake

Food intake in the 450 mg/kg dose group was less than that of the control group and the result was statistically significant. In males, the daily intake between Days 1 to 8, Days 29 to 36, and Days 43 to 48 was smaller. The cumulative intake between Days 1 to 15 was also smaller. The daily intake of female animals between days 1 to 8 and, cumulatively, between Days 1 to 15 was also smaller.

Organ weights

In males from the 450 mg/kg group, absolute weight of the thymus decreased (190 ± 60 mg) compared with controls (277 ± 73 mg). Absolute (90 ± 14 mg) and relative weight (20.526 ± 3.256 %) of the adrenal increased compared with control values of 63 ± 9 mg and 12.382 ± 1.925 %. The relative weight of the testes also increased (0.741 ± 0.029 % compared with 0.0667 ± 0.047 % for the control animals). The relative weight of the thymus was inclined to decrease although no statistical difference was observed (45.090 ± 12.432 % compared with 54.228 ± 13.221 % for the controls).

Necropsy

In males, ulceration of the fore-stomach was seen in 9 animals of the 450 mg/kg group and the incidence was increased with statistical significance in comparison to the control group. Of these 9 animals, six had adhesion of the stomach with surrounding tissue. Other changes were observed in both the control and the test groups.

Females who underwent natural delivery showed ulceration of the fore-stomach (9 animals, 450 mg/kg group) and atrophy of the thymus (6 animals, 450 mg/kg group) and these effects were statistically significant. Three females from the 150 mg/kg group had fore-stomach ulceration with stomach adhesion to surrounding tissues, 3 had mucosal-thickening of the fore-stomach and 1 animal had atrophy of the spleen. Other changes were observed in both the control and test groups.

One female from the control group failed to deliver and a dead foetus was found in the uterus on day 25 of pregnancy.

Histopathology

Male animals showed ulceration of the fore-stomach, squamous epithelium hyperplasia with hyperkeratosis in the fore-stomach and diffuse hyperplasia in the adrenal cortex with statistically higher significance compared to the control group. For ulceration in the fore-stomach, incidence of severe cases also increased with statistical significance compared to control animals.

Females who underwent natural delivery showed ulceration of the fore-stomach, squamous epithelium hyperplasia with hyperkeratosis in the fore-stomach and diffuse hyperplasis in the adrenal cortex. The incidence of these effects was higher than the control group and was statistically significant. As with the male animals, incidence of severity of ulceration increased with statistical significance compared to controls.

Copulation and fertility

The female from the 450 mg/kg group that died on day 2 of pregnancy was excluded from conception observations. All other females conceived. No intergroup differences were observed in the mean duration of the oestrus cycle. No intergroup differences were seen in the incidence of abnormal oestrus cycle.

Delivery and lactation

No abnormalities were seen in delivery. The values of pregnant period, number of corpora lutea, number of implantation sites, number of offspringand number of offspring born alive were similar among each group. No differences among groups were seen in gestation index, implantation index, birth index, sex ratio or viability index on day 4.

Dose descriptor:

NOEL

Effect level:

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

Based on:

test mat.

Sex:

male

Basis for effect level:

other: see 'Details on results (P0)

Dose descriptor:

NOEL

Effect level:

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

Based on:

test mat.

Sex:

female

Basis for effect level:

other: see 'Details on results'

Clinical signs:

not examined

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

effects observed, treatment-related

Histopathological findings:

effects observed, treatment-related

Offspring morphology

Bodyweight and necropsy findings showed dwarfism (bodyweight less than 60 % of mean bodyweight of control group) in only one offspring from the 450 mg/kg dose group. No intergroup differences were observed in bodyweight on day 0 and day 4. Offspring were necropsied on day 4 of lactation and a thymus remnant was seen in 5 males of the control group, 3 males and 1 female of the 50 mg/kg group, and 2 males of the 450 mg/kg group. Other sporadic changes observed in males were whitish spotting, dark spotting, adhesion with diaphragm, yellowish change and anomalous nodule in the liver, pelvic dilation of the kidney, displacement of the kidney, and reddening of the eyeball. In addition, skin bruising was seen in 3 males and females of the smae litter from the 150 mg/kg group.

Dose descriptor:

NOEL

Generation:

F1

Effect level:

450 mg/kg bw/day

Based on:

test mat.

Sex:

not specified

Basis for effect level:

other: see 'Details on results (F1)'

Reproductive effects observed:

not specified

The test substance administration influenced the death of males in the 150 mg/kg group and males and females of the 450 mg/kg group.

Clinical observations were also considered to be caused by substance administration. It was suspected that the substance also caused stimulation of mucus secretion and caused respiratory dysfunction. Other observations such as ptosis and fur staining were considered to be induced by exacerbation of a general condition but not by the substance. Salivation was considered to be an incidental occurance unrelated to the administration of the test substance.

Bodyweight gain was suppressed by substance administration and food intake was also influenced.

The action of the substance appeared to be stronger in males than females. It was therefore concluded that the action of the test substance was sex related.

The substance did not cause any changes in the reproductive organs. The changes in the digestive system were considered to be induced by the corrosive nature of the substance. The substance did not affect any part of the reproductive cycle.

Conclusions:

No influences on fertility or the next generation were observed

Additional information

In a reproductive/developmental toxicity screening test (Ito, 2001), the substance was administered orally by gavage to groups of 12 males and 12 females from 14 days before mating to 20 days after mating in males, and to day 3 of lactation in females. No adverse effects were observed in terms of copulation, fertility, delivery or nursing in the parents and the NOEL was reported as 50 mg/kg bw/day in males and 150 mg/kg bw/day in females. The NOEL for reproductive/developmental toxicity (F1 offspring) was 450 mg/kg bw/day.

Short description of key information:
No influences on fertility or the next generation (OECD 421)

Effects on developmental toxicity

Description of key information

No influences on developmental toxicity (OECD 414). The maternal NOAEL = 100 mg/kg bw/day,  the developmental toxicity NOAEL = 300 mg/kg bw /day.

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 2012-12-13 to 2013-07-09

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline conforming GLP study

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc., Raleigh, NC
- Age at study initiation: 93 days old
- Weight at study initiation: 228 - 295 g
- Fasting period before study: no
- Housing: until pairing individually in clean, stainless steel wire-mesh cages suspended above cage-board
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.7 - 22.4
- Humidity (%): 38.8 - 45.8
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Dosing formulations were prepared at the test substance concentrations indicated below:

Group 1: vehicle control 0 mg/kg/day dose level, 0 mg/mL test substance concentration, pH 6.82
Group 2: 1,3-benzenedimethanamine 30 mg/kg/day dose level, 3 mg/mL test substance concentration, pH 11.19
Group 3: 1,3-benzenedimethanamine 100 mg/kg/day dose level, 10 mg/mL test substance concentration, PH 11.31
Group 4 1,3-benzenedimethanamine 300 mg/kg/day dose level, 30 mg/mL test substance concentration, pH 11.57

The appropriate amounts of vehicle and test substance were transferred to sterile septum vials. The containers were purged with nitrogen, capped with a septum, and inverted to ensure mixing. The formulations were maintained at room temperature, protected from light, until use.

VEHICLE
- Concentration in vehicle: 0, 3, 10, 30 test substance mg/mL
- Amount of vehicle (if gavage): 10 mL/kg

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analyses to demonstrate the stability (following at least 10 days of room temperature storage) of the test substance formulations were conducted. Samples for concentration analysis were collected from the dosing formulations (including the control group) prepared during the first and last week of the study. One set of samples from each collection was subjected to the appropriate analyses. The remaining set of samples was stored at room temperature as back-up. All analyses were conducted using a validated gas chromatography method with flame ionization detection.

Details on mating procedure:

- Impregnation procedure: cohoused
- M/F ratio per cage: no data
- Length of cohabitation: no data
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy

Duration of treatment / exposure:

gestation days 6-19

Frequency of treatment:

once daily

Duration of test:

from 2012-12-18 to 04-04-2013

No. of animals per sex per dose:

25 bred female rats

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dosage levels were selected based on the results of a previous study in which 1,3-benzenedimethanamine was administered orally to female rats at dosage levels of 0, 50, 150, 450, and 650 mg/kg/day from gestation days 6 through 19. One female each in the 450 and 650 mg/kg/day groups was found dead during the treatment period. All other animals survived until the scheduled euthanasia on gestation day 20. Test substance-related reductions in mean body weight gains were noted throughout the treatment period for both the 450 and 650 mg/kg/day groups resulting in statistically significant lower mean body weights for these groups compared to the control group beginning on gestation day 15 and generally persisting for the remainder of the study. Reductions in mean food consumption were also noted for the 450 and 650 mg/kg/day groups when compared to the control group; the changes were generally statistically significant throughout the treatment period. In addition, statistically significant reduced mean fetal weights were noted in the 450 and 650 mg/kg/day groups when compared to the control group. Based on the results of the previous study, a high dose of 300 mg/kg/day was chosen for the current study and was expected to have effects on maternal body weight gain without causing mortality.

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily. Animals were onserved for signs of toxicity approximately 1 hour following dose administration.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily from gestation days 0 through 20.

BODY WEIGHT: Yes
- Time schedule for examinations: on gestation days 0 6-20 (daily)

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day #20
- Organs examined: contents of thoracic, abdominal and pelvic cavities.

Ovaries and uterine content:

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

Fetal examinations:

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

Statistics:

All statistical tests were performed using WTDMS™ unless otherwise noted. Analyses were conducted using two-tailed tests (except as noted otherwise) for minimum significance levels of 1% and 5%, comparing each test substance-treated group to the control group. Each mean was presented with the standard deviation (SD), standard error (SE), and the number of animals (N) used to calculate the mean. Data obtained from nongravid animals were excluded from statistical analyses. Where applicable, the litter was used as the experimental unit.
Maternal body weights (absolute and net), body weight changes (absolute and net), and food consumption, gravid uterine weights, numbers of corpora lutea, implantation sites, and viable fetuses, and fetal body weights (separately by sex and combined) [were subjected to a parametric one-way ANOVA (Snedecor and Cochran, 1980) to determine intergroup differences. If the ANOVA revealed significant (p<0.05) intergroup variance, Dunnett's test (Dunnett, 1964) was used to compare the test substance-treated groups to the control group. Mean litter proportions (percent per litter) of prenatal data (viable and nonviable fetuses, early and late resorptions, total resorptions, pre- and postimplantation loss, and fetal sex distribution), total fetal malformations and developmental variations (external, visceral, skeletal, and combined) and each particular external, visceral, and skeletal malformation or variation were subjected to the Kruskal-Wallis nonparametric ANOVA test (Kruskal and Wallis, 1952) to determine intergroup differences. If the ANOVA revealed significant (p<0.05) intergroup variance, Dunn’s test (Dunn, 1964) was used to compare the test substance-treated groups to the control group.

Indices:

No data

Historical control data:

yes

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects: Moribundity, mean body weight losses, and lower mean body weight gains with corresponding reduced mean food consumption at 300 mg/kg bw/day.

Dose descriptor:

NOAEL

Effect level:

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

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Dose descriptor:

NOAEL

Effect level:

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

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: intrauterine growth and survival or embryo/fetal development

Abnormalities:

not specified

Developmental effects observed:

not specified

Conclusions:

Based on the results of the study, a dosage level of 100 mg/kg bw/day is considered to be no-observed-adverse-effect level (NOAEL) for maternal toxicity. There were no test substance-related effects on intrauterine growth and survival of embryo/fetal development at any dosage level. Therefore, a dosage level of 300 mg/kg bw/day is considered to be the NOAEL for embryo/fetal development when 1,3-benzenedimethanamine was administered orally by gavage to bred Crl:CD(SD) rats.

Executive summary:

The objective of the study was to determine the potential of the test substance to induce developmental toxicity after maternal exposure from implantation to 1 day prior to expected parturition, to characterize maternal toxicity at the exposure levels tested, and to determine a no-observed-adverse-effect level (NOAEL) for maternal and developmental toxicity according to the OECD TG 414 and EPA OPPTS Guideline 870.3700. The test substance, 1,3-benzenedimethanamine, in the vehicle (sterile water for injection, USP) was administered orally by gavage to 3 groups of 25 bred female Crl:CD(SD) rats once daily from gestation days 6 through 19. Dosage levels were 30, 100, and 300 mg/kg bw/day administered at a dosage volume of 10 mL/kg. A concurrent control group composed of 25 bred females received the vehicle on a comparable regimen. The females were approximately 14 weeks of age at the initiation of dose administration. All animals were observed twice daily for mortality and moribundity. Clinical observations, body weights, and food consumption were recorded at appropriate intervals. On gestation day 20, a laparohysterectomy was performed on each surviving female. The uteri, placentae, and ovaries were examined, and the numbers of fetuses, early and late resorptions, total implantations, and corpora lutea were recorded. Gravid uterine weights were recorded, and net body weights and net body weight changes were calculated. The fetuses were weighed, sexed, and examined for external, visceral, and skeletal malformations and developmental variations.

Three females in the 300 mg/kg bw/day group were euthanized in extremis on gestation day 11, 12, or 15 following severe body weight losses and decreased food consumption during the 2-5 days prior to euthanasia. Clinical findings noted for these females included rales, labored respiration, gasping, pale body, decreased defecation, small feces, soft stool, and/or red, clear, brown, and/or yellow material on various body surfaces at the daily examinations and/or approximately 1 hour following dose administration beginning up to 4 days prior to euthanasia. At necropsy, these females were noted with internal findings of distended stomach and/or intestine, dark red areas on the stomach, and dark red discoloration of the lungs. All other females survived to the scheduled necropsy where no remarkable macroscopic findings were observed at any dosage level. Clinical findings of rales, gasping, labored respiration, red and/or clear material on the forelimbs, around the nose, and/or mouth, and excreta-related findings (decreased defecation, soft stool, and small feces) were noted for surviving females in the 300 mg/kg/day group primarily throughout the treatment period at the daily examinations and at approximately 1 hour following administration and were considered test substance-related and adverse. There were no test substance-related clinical findings observed at dosage levels of 30 and 100 mg/kg bw/day. A mean body weight loss or lower mean body weight gains were noted for the 300 mg/kg bw/day group throughout the treatment period, corresponded to decrements in mean food consumption, and were considered test substance-related and adverse. As a result, a lower mean body weight gain was noted for this group when the overall treatment period (gestation days 6-20) was evaluated. These effects resulted in mean body weight for the 300 mg/kg/day group that was up to 7.2 % lower than the control group throughout the study, with correspondingly lower mean net body weight and net body weight gain in this group. Mean gravid uterine weight in the 300 mg/kg bw/day group was unaffected by test substance administration. Mean body weights, body weight gains, net body weights, net body weight gains, and gravid uterine weights, and food consumption in the 30 and 100 mg/kg bw/day groups were unaffected by test substance administration. Intrauterine growth and survival and fetal developmental morphology at 30, 100, and 300 mg/kg kg/day were unaffected by test substance administration.

Moribundity, mean body weight losses, and lower mean body weight gains with corresponding reduced mean food consumption were noted at a dosage level of 300 mg/kg bw/day. Based on these results, a dosage level of 100 mg/kg bw/day was considered to be the no-observed-adverse-effect level (NOAEL) for maternal toxicity.

There were no test substance-related effects on intrauterine growth and survival or embryo/fetal development at any dosage level. Therefore, a dosage level of 300 mg/kg bw/day was considered to be the NOAEL for and embryo/fetal development when 1,3-benzenedimethanamine was administered orally by gavage to bred Crl:CD(SD) rats.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

300 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Only one GLP study according to OECD TG 414 is available.

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

In a prenatal developmental toxicity study (Herberth, 2013), the substance was administered orally by gavage to 3 groups of 25 bred female Crl:CD(SD rats once daily from gestation days 6 through 19. Dosage levels were 30, 100 and 300 mg/kg bw/day. Clinical findings, body weight loss, lower mean body weight gains and decrements in mean food consumption were observed at 300 mg/kg bw/day group. Intrauterine growth and survival and fetal developmental morphology at 30, 100 and 300 mg/kg bw/day were unaffected by test substance administration. Based on these results, a dosage level of 100 mg/kg bw/day was considered to be NOAEL for maternal toxicity and a dosage level of 300 mg/kg bw/day was considered to be the NOAEL for embryo/fetal development when MXDA was administered to rats.

Justification for selection of Effect on developmental toxicity: via oral route:
The developmental toxicity of the substance is characterized from a valid prenatal developmental toxicity study conducted with MXDA. Since the maternal NOAEL = 100 mg/kg bw/day was lower than the developmental toxicity NOAEL = 300 mg/kg bw/day (the highest administered dosis), no adverse developmental effect was observed.

Justification for classification or non-classification

- Reproductive toxicity:

No influences were seen on fertility or the next generation and the test substance does not meet the criteria for classification under the terms of Directive 67/548/EEC or GHS as reflected by Regulation (EC) 1272/2008.

- Developmental toxicity:

The developmental toxicity study only showed maternal toxic effects. Therefore, based on this study, the substance does not have to be classified according to the Directive 67/548/EEC or GHS as reflected by Regulation (EC) 1272/2008.