β-methyl-3-(1-methylethyl)benzenepropanal

Endpoint summary

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Effects on fertility

Description of key information

No effects on the reproductive organs or sperm parameters of male rats were found in a 14 days study.

Those findings are supported by the absence of findings in an OECD 407 (28d) in rats.

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14 days

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Remarks:

High quality GLP study

Qualifier:

no guideline available

Principles of method if other than guideline:

A study was conducted to determine the possible adverse effects on the reproductive organs of male rats resulting from repeated oral (gavage) exposure to the test material over a 14 day period. Each male rat was administered the vehicle (corn oil) or test material orally (via gavage) once daily for 14 consecutive days. Dose levels were 25, 75 and 250 mg/kg/day for each of the test materials. The following parameters were evaluated: viability, clinical observations, body weights and body weight gains, feed consumption, necropsy observations, urinalysis, sperm concentration, motility and morphology, organ weights and histopathology. Viability was observed at least twice each day throughout the study. The rats were examined for clinical observations at least weekly during the acclimation period prior to dosage and hourly for the first four hours postdosage and at the end of the business day throughout the study. Rats were also examined for clinical observations prior to sacrifice. Body weights were recorded twice during the acclimation period, daily during the dosage period and prior to sacrifice. Feed consumption values were recorded twice during the acclimation period and on DSs 1, 8 and 14. Rats were fasted following the final dosage on DS 14 to facilitate urine sample collection overnight, beginning on the evening of DS 14. Samples were collected over cold packs, stored refrigerated and subsequently examined for sediment and microscopically analyzed. Fecal samples were collected following dosage administration on DS14. Samples were placed in sealable pouches, labeled and stored frozen for possible future analysis. Surviving animals were sacrificed by CO2 asphyxiation following the last administration of the appropriate test material and/or vehicle (DS 15). All rats were subjected to a complete necropsy examination, which included an evaluation of the carcass and musculoskeletal system; all external surfaces and orifices; cranial cavity and external surfaces of the brain; and the thoracic, abdominal and pelvic cavities with their associated organs and tissues. The lungs were perfused with neutral buffered 10% formalin (NBF). Gross lesions were retained in neutral bufferend 10% formalin for histological evaluation. Specific organs were weighed at necropsy for all animals that survived to scheduled sacrifice. Organ weights were not recorded for rats found dead or sacrificed in poor condition. Paired organs were weighed together. All tissues were preserved in 10% neutral buffered formalin. Sperm concentration and motility were evaluated using computer assisted sperm analysis. Motility was evaluated by the Hamilton Thorne IVOS, using a sample collected from the left vas deferens. A homogenate was prepared from the left or right cauda epididymis for evaluation by the Hamilton Thorne IVOS to determine sperm concentration (sperm per gram of tissue weight). The remaining portion of the left cauda epididymis was used to manually evaluate sperm morphology. Sperm morphology evaluations included the following: 1) determination of the percentage of normal sperm in a sample of at least 200; and 2) qualitative evaluation of abnormal sperm, including such categories as abnormal head, abnormal tail and abnormal head and tail. Tissues identified for microsocpic evaluation were embedded in paraffin. The liver, kidneys and reproductive organs from male rats in the vehicle control groups and the high dosage groups for each test material were sectioned, mounted on glass slides and stained with hematoxylin and eosin. In addition several tissues were processed histologically and examined microscopically. Gross lesions from rats in all dosage groups were processed as described above for microscopic evaluation. All gross lesions from rats in all dosage groups were evaluated microscopically. If lesions attributed to the test material were observed in the rats from a high dosage group, the same organs from rats exposed to lower test material dosages were sectioned and examined microscopically. All remaining tissues were preserved in 10% neutral buffered formalin for possible future evaluation. Rats that died or were sacrificed in poor condition were examined for the cause of death or condition as soon as possible after the observation was made. The rats were examined for gross lesions. The lungs, trachea and esophagus were perfused and saved in neutral bufferend 10% formalin for possible future evaluation. When not precluded by autolysis, specific tissues were retained, processed as described above and evaluated microscopically, but not weighed. Statistical analysis were conducted.

GLP compliance:

yes

Limit test:

no

Specific details on test material used for the study:

Isopropyphenylbutanal

Species:

rat

Strain:

Sprague-Dawley

Remarks:

CRL:CD (SD)

Sex:

male

Details on test animals or test system and environmental conditions:

Room with positive airflow
at least 10 changes of fresh HEPA filtered air per hour
Room temparture between 18 - 26°C
Relative humidity 30-70%
12 hours light/dark cycles
Individual housing in stainless steel wire-bottom cages
Ad libitum standard rodent diet and water
Nesting and enrichment material at disposal

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

Gavage

Details on mating procedure:

N/A

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

14 days

Frequency of treatment:

Once daily

Details on study schedule:

Each male rat was administered the vehicle (corn oil) or test material orally (via gavage) once daily for 14 consecutive days. Dose levels were 25, 75 and 250 mg/kg/day for each of the test materials

Dose / conc.:

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

Dose / conc.:

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

Dose / conc.:

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

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Positive control:

none

Parental animals: Observations and examinations:

mortality, clinical observation, body weight, body weight gain, feed consumption, necropsy observations, urinalysism sperm concentration, motility and morphology, organ weight and histopathology.

Oestrous cyclicity (parental animals):

not observed

Sperm parameters (parental animals):

sperm concentration, motility and morphology

Litter observations:

not observed

Postmortem examinations (parental animals):

Complete necropsy observations, urinalysis, sperm concentration, motility and morphology, organ weight and histopathology

Postmortem examinations (offspring):

none

Statistics:

Clinical observations and other proportional data were analyzed using the Variance test for Homogeneity of the Binomal Distribution.
Continuous data (body weight changes and feed consumption values) were analyzed using Bartlett's Test of Homogeneity of Variances, when appropriate.
If the Analysis of Variance was significant (p<=0.05), Dunnett's test was used to identify the statistical significance of the individual groups.
If the ananlysis of variance was not appropriate, the Kruskal-Wallis test was used (<=75% ties).
In cases where the Kruskal-Wallis test was statistically significant (p<=0.05), Dunn's Method of Multiple Comparisons was used to identify the statistical significance of the individual groups.
If there are greater thant 75% ties, Fischer's Exact Test was used to analyze the data

Reproductive indices:

NA

Offspring viability indices:

Na

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

no effects observed

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

not examined

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

not examined

Key result

Dose descriptor:

NOAEL

Effect level:

>= 250 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: Highest dose tested and no adverse effects were observed

Critical effects observed:

no

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality:

not examined

Body weight and weight changes:

not examined

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

not examined

Remarks on result:

not measured/tested

Critical effects observed:

not specified

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality / viability:

not examined

Body weight and weight changes:

not examined

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

not examined

Anogenital distance (AGD):

not examined

Nipple retention in male pups:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Histopathological findings:

not examined

Other effects:

not examined

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

Remarks on result:

not measured/tested

Critical effects observed:

not specified

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality / viability:

not examined

Body weight and weight changes:

not examined

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

not examined

Anogenital distance (AGD):

not examined

Nipple retention in male pups:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Histopathological findings:

not examined

Other effects:

not examined

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

Remarks on result:

not measured/tested

Critical effects observed:

not specified

Reproductive effects observed:

no

25 mg/kg	no effects
	dose was mg/kg bodyweight. No deaths occurred. All clinical observations were considered unrelated to treatment with the test material. Bodyweights and bodyweight gains were unaffected by treatment with the test material at this dose level. Absolute (g/day) and relative (g/kg/day) feed consumption values were unaffected by treatment with the test material at this dose level. There were no biologically important differences between the vehicle and treated groups in the urinalysis sample collected. There were no test material related necropsy observations. There were no differences compared to the vehicle control group with terminal bodyweight, organ weights and ratios of organ weights to terminal bodyweights at this dose level. The weights of epidiymides, caudal epididymis, testes, seminal vesicle (with and without fluid) and prostate and the ratios of these organ weights to terminal body weight were unaffected by the test material at this dose level. In addition, the absolute and relative weights of the paired kidneys did not differ statistically from the concurrent vehicle control group. All sperm parameters (motility, concentration and morphology) evaluated were unaffected by dosages of test material.
250 mg/kg	liver ,  organ weight changes
	dose was mg/kg bodyweight. No deaths occurred. All clinical observations were considered unrelated to treatment with the test material. Bodyweights and bodyweight gains were unaffected by treatment with the test material at this dose level. Absolute (g/day) and relative (g/kg/day) feed consumption values were unaffected by treatment with the test material at this dose level. At this dose level, the pH of the urine was significantly decreased (p<\=0.01) in comparison to the concurrent vehicle control grup value. The overall significance of this finding could not be determined, but likely reflected excretion of hte test material in the urine. This finding was not considered to be an adverse effect of the test material. There were no other biologically important differences between the vehicle and treated groups in the urinalysis sample collected. There were no test material related necropsy observations. Rats treated at this dose level had statistically significant increases (p<\=0.05 or p<\=0.01) in absolute and relative weights (% terminal body weight) of the liver, as compared to the concurrent vehicle control group values (11% and 12% greater than controls). There were no microscopic changes observed in rats at this dose level that could be correlated with the liver weight differences. Therefore, this finding was not considered to be an adverse effect of the test material. Terminal bodyweighs did not differe statistically from the concurrent vehicle control group value at this dose level. The weights of epidiymides, caudal epididymis, testes, seminal vesicle (with and without fluid) and prostate and the ratios of these organ weights to terminal body weight were unaffected by the test material at this dose level. In addition, the absolute and relative weights of the paired kidneys did not differ statistically from the concurrent vehicle control group. All sperm parameters (motility, concentration and morphology) evaluated were unaffected by dosages of test material.
75 mg/kg	no effects
	dose was mg/kg bodyweight. No deaths occurred. All clinical observations were considered unrelated to treatment with the test material. Bodyweights and bodyweight gains were unaffected by treatment with the test material at this dose level. Absolute (g/day) and relative (g/kg/day) feed consumption values were unaffected by treatment with the test material at this dose level. There were no biologically important differences between the vehicle and treated groups in the urinalysis sample collected. There were no test material related necropsy observations. There were no differences compared to the vehicle control group with terminal bodyweight, organ weights and ratios of organ weights to terminal bodyweights at this dose level. The weights of epidiymides, caudal epididymis, testes, seminal vesicle (with and without fluid) and prostate and the ratios of these organ weights to terminal body weight were unaffected by the test material at this dose level. In addition, the absolute and relative weights of the paired kidneys did not differ statistically from the concurrent vehicle control group. All sperm parameters (motility, concentration and morphology) evaluated were unaffected by dosages of test material.

Conclusions:

The No Observed Adverse Effect Level (NOAEL) of the test material for general toxicity in male rats after oral administration for 14 consecutive days is greater than 250 mg/kg/day.
The No Observed Adverse Effect Level (NOAEL) of the test material for reproductive organs and sperm in male rats after oral administration for 14 consecutive days is greater than 250 mg/kg/day.

Executive summary:

A study was conducted to determine the possible adverse effects on the reproductive organs of male rats resulting from repeated oral (gavage) exposure to the test material over a 14 day period. Two hundred male Crl:CD(SD) rats were randomly assigned to 20 dosage groups, ten rats per group. Animals were approximately 72 days old at arrival and weighed 308-355 at study assignment. Study rooms were maintained under conditions of positive airflow relative to a hallway and independently supplied with a minimum of 10 changes per hour of 100% fresh air that had been passed through 99.97% HEPA filters. Room temperature and humidity were monitored constantly throughout the study. Room temperature was targeted at 64 F to 79 F (18 C to 26 C); relative humidity was targeted at 30% to 70%. Rats were individually housed in stainless steel wire-bottomed cages except during urine sample collection when rats were individually housed in stainless steel metabolism cages. A 12 hour light/dark fluorescent light cycle was maintained. Cage pan liners were changed at least three times weekly. Cages were changed approximately every other week. Bedding was changed as often as necessary to keep the rats dry and clean. Rats were given ad libitum access to Certified Rodent Diet #5002 in individual feeders, except when fasting to facilitate urine sample collection. Local water that had been processed by passage through a reverse osmosis membrane (R.O. water) was available to the rats ad libitum from an automatic watering access system and/or individual water bottles attached to the cages. Chlorine was added to the processed water as a bacteriostat. For rats placed in nesting boxes at the request of veterinary staff, Bed-o'cobs bedding was used as the nesting material. Chewable Nylabones were supplied to all rats during the course of the study. Each male rat was administered the vehicle (corn oil) and/or one of the six test materials orally (via gavage) once daily for 14 consecutive days. Dose levels were 25, 75 and 250 mg/kg/day for each of the test materials. The dosage volume was 4 ml/kg for the vehicle and the test materials. The dosage volume was adjusted for the most recently recorded body weight and given at approximately the same time each day. Suspensions of test material were prepared daily and were stored at room temperature, protected from light. All surviving rats were sacrificed on DS 15 by CO2 asphyxiation. The following parameters were evaluated: viability, clinical observations, body weights and body weight gains, feed consumption, necropsy observations, urinalysis, sperm concentration, motility and morphology, organ weights and histopathology. Viability was observed at least twice each day throughout the study. The rats were examined for clinical observations at least weekly during the acclimation period prior to dosage and hourly for the first four hours postdosage and at the end of the business day throughout the study. Rats were also examined for clinical observations prior to sacrifice. Body weights were recorded twice during the acclimation period, daily during the dosage period and prior to sacrifice. Feed consumption values were recorded twice during the acclimation period and on DSs 1, 8 and 14. Rats were fasted following the final dosage on DS 14 to facilitate urine sample collection overnight, beginning on the evening of DS 14. Samples were collected over cold packs, stored refrigerated and subsequently examined for sediment and microscopically analyzed. Fecal samples were collected following dosage administration on DS14. Samples were placed in sealable pouches, labeled and stored frozen for possible future analysis. Surviving animals were sacrificed by CO2 asphyxiation following the last administration of the appropriate test material and/or vehicle (DS 15). All rats were subjected to a complete necropsy examination, which included an evaluation of the carcass and musculoskeletal system; all external surfaces and orifices; cranial cavity and external surfaces of the brain; and the thoracic, abdominal and pelvic cavities with their associated organs and tissues. The lungs were perfused with neutral buffered 10% formalin (NBF). Gross lesions were retained in neutral bufferend 10% formalin for histological evaluation. Specific organs were weighed at necropsy for all animals that survived to scheduled sacrifice. Organ weights were not recorded for rats found dead or sacrificed in poor condition. Paired organs were weighed together. All tissues were preserved in 10% neutral buffered formalin. Sperm concentration and motility were evaluated using computer assisted sperm analysis. Motility was evaluated by the Hamilton Thorne IVOS, using a sample collected from the left vas deferens. A homogenate was prepared from the left or right cauda epididymis for evaluation by the Hamilton Thorne IVOS to determine sperm concentration (sperm per gram of tissue weight). The remaining portion of the left cauda epididymis was used to manually evaluate sperm morphology. Sperm morphology evaluations included the following: 1) determination of the percentage of normal sperm in a sample of at least 200; and 2) qualitative evaluation of abnormal sperm, including such categories as abnormal head, abnormal tail and abnormal head and tail. Tissues identified for microsocpic evaluation were embedded in paraffin. The liver, kidneys and reproductive organs from male rats in the vehicle control groups and the high dosage groups for each test material were sectioned, mounted on glass slides and stained with hematoxylin and eosin. In addition several tissues were processed histologically and examined microscopically. Gross lesions from rats in all dosage groups were processed as described above for microscopic evaluation. All gross lesions from rats in all dosage groups were evaluated microscopically. If lesions attributed to the test material were observed in the rats from a high dosage group, the same organs from rats exposed to lower test material dosages were sectioned and examined microscopically. All remaining tissues were preserved in 10% neutral buffered formalin for possible future evaluation. Rats that died or were sacrificed in poor condition were examined for the cause of death or condition as soon as possible after the observation was made. The rats were examined for gross lesions. The lungs, trachea and esophagus were perfused and saved in neutral bufferend 10% formalin for possible future evaluation. When not precluded by autolysis, specific tissues were retained, processed as described above and evaluated microscopically, but not weighed. Statistical analysis were conducted. Statistically significant probabilities were reported as either p</= 0.05 or p</=0.01.

The No Observed Adverse Effect Level (NOAEL) of the test material for general toxicity in male rats after oral administration for 14 consecutive days is greater than 250 mg/kg/day. The No Observed Adverse Effect Level (NOAEL) of the test material for reproductive organs and sperm in male rats after oral administration for 14 consecutive days is greater than 250 mg/kg/day.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Effects on developmental toxicity

Description of key information

No developmental study is available on the substance

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Toxicity to reproduction: other studies

Description of key information

No other study was available on the substance

Justification for classification or non-classification

According to the available experimental results, no classification for reproductive or developmental toxicity is needed for this substance under CLP.