1,4-diisopropylbenzene

Endpoint summary

Description of key information

Repeated dose toxicity oral

In a study according to the OECD guideline 422 under GLP compliance the NOAEL for systemic toxicity was considered to be 1000 mg/kg/day.

 

Repeated dose toxicity inhalation

No study available

 

Repeated dose toxicity dermal

No study available

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

28 JUL 2015 - 01 JUN 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

see below

Principles of method if other than guideline:

- Short description of test conditions: Estrous cycles were not monitored before treatment started to select for the study females with regular cyclicity. In addition, vaginal smears were not monitored daily from the beginning of treatment period until evidence of mating. However, all females assigned to the study had evidence of mating, mean pre-coital intervals for all groups were within 1 normal estrous cycle (less than 4-5 days), and 39 of the 40 females assigned to the study were gravid. Therefore, the females assigned to this study were cycling normally and the lack of estrous cycle monitoring had no impact on the study.

GLP compliance:

yes

Limit test:

no

Specific details on test material used for the study:

TEST MATERIAL
- name as cited in test report: 1,4-diisopropylbenzene
- Molecular weight: 162 g/mol
- Molecular formula: C12H18
- CAS no. 100-18-5
- Physical state: clear, colorless liquid
- Date of receipt: 27-Aug-2015

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: stored at room temperature, protected from light
- Stability with H+; OH-; Heat, Light, H2: considered stable under these conditions

Species:

rat

Strain:

Sprague-Dawley

Details on species / strain selection:

Sexually mature male and virgin female Sprague Dawley [Crl:CD(SD)] rats were used as the test system on this study. The animal model, the Crl:CD(SD) rat, is recognized as appropriate for reproductive toxicity studies and has been proven to be susceptible to the effects of reproductive toxicants. In addition, WIL Research has reproductive historical control data in the Crl:CD(SD) rat.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc., Raleigh, NC on 20-Aug-2015.
- Age at study initiation: approximately 10 weeks old.
- Weight at study initiation: Male body weights ranged from 326 g to 396 g and female body weights ranged from 223 g to 278 g
- Housing: Following receipt and until pairing, all F0 animals were housed 2-3 per cage by sex in clean, solid-bottom cages with bedding material (Bed-O'Cobs®; The Andersons, Cob Products Division, Maumee, OH).
- Diet: ad libitum; Feeders were changed and sanitized once per week.
- Water: ad libitum
- Acclimation period: 12 days prior to the first day of treatment.

DETAILS OF FOOD AND WATER QUALITY:
The basal diet used in this study, PMI Nutrition International, LLC Certified Rodent LabDiet® 5002, was a certified feed with appropriate analyses performed by the manufacturer and provided to WIL Research. Feed lots used during the study were documented in the study records. Municipal water supplying the facility was sampled for contaminants according to WIL Research SOPs.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.1 °C to 22.2 °C
- Humidity (%): 45.1 % to 60.5 %
- Air changes (per hr): Air handling units were set to provide a minimum of 10 fresh air changes per hour.
- Photoperiod (hrs dark / hrs light): Fluorescent lighting provided illumination for a 12-hour light (0600 hours to 1800 hours)/12-hour dark photoperiod.

Route of administration:

oral: gavage

Details on route of administration:

The vehicle and test item formulations were administered orally by gavage, via an appropriately sized flexible, Teflon®-shafted, stainless steel ball-tipped dosing cannula once daily. The males were dosed during study days 0-27 or 0-28 (14 days prior to pairing through 1 day prior to scheduled euthanasia) for a total of 28-29 doses. The females were dosed during study days 0 through the day prior to euthanasia (14 days prior to pairing through lactation day 13) for a total of 49-59 doses. The female that failed to deliver was dosed through the day prior to euthanasia (post-mating day 25) for a total of 39 doses. The dose volumes for Group 1, Group 2, Group 3, and Group 4 were 1.18, 0.12, 0.35, and 1.18 mL/kg, respectively. Individual dosages were based on the most recently recorded body weights to provide the correct mg/kg/day dose. All animals were dosed at approximately the same time each day. Dosage levels were determined from the results of previous studies and were provided by the Sponsor after consultation with the Study Director. Additionally, the dosage levels selects for the current study test up to the limit dose of 1000 mg/kg/day.
The selected route of administration for this study was oral (gavage) because this a potential route of exposure for humans. Historically, this route has been used extensively for studies of this nature.

Vehicle:

water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
VEHICLE
For the control group, an appropriate volume of deionized water was divided into aliquots for daily dispensation approximately weekly and stored at room temperature, protected from light.
- Amount of vehicle (if gavage): 0.12, 0.35 and 1.18 mL/kg; The volume of the neat test item to be administered was determined from the weight of the test item divided by the specific gravity (0.85 g/mL).
The vehicle and test item formulations were administered orally by gavage, via an appropriately sized flexible, Teflon®-shafted, stainless steel ball-tipped dosing cannula once daily.

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

The test item was a liquid and was administered as-is, with no dilution. The test item was aliquoted for daily dispensation approximately weekly, and stored at room temperature, protected from light.

Duration of treatment / exposure:

Males 28-29 days, Females 49-59 days.

Frequency of treatment:

Daily

Dose / conc.:

100 mg/kg bw/day (nominal)

Dose / conc.:

300 mg/kg bw/day (nominal)

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dosage levels were determined from the results of previous studies and were provided by the Sponsor after consultation with the Study Director. Additionally, the dosage levels selects for the current study test up to the limit dose of 1000 mg/kg/day.
- Rationale for animal assignment (if not random): based on body weight stratification in a block design.

Individual dosages were based on the most recently recorded body weights to provide the correct mg/kg/day dose.
The selected route of administration for this study was oral (gavage) because this a potential route of exposure for humans. Historically, this route has been used extensively for studies of this nature.

Positive control:

no

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: All rats were observed twice daily, once in the morning and once in the afternoon, for appearance, behavior, moribundity, mortality, and signs of overt toxicity.

DETAILED CLINICAL OBSERVATIONS: Yes

BODY WEIGHT: Yes
- Time schedule for examinations: Individual male body weights were recorded weekly.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
Food consumption was recorded on the corresponding weekly body weight days until pairing. Food consumption was measured on a per cage basis for the corresponding body weight intervals.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Not specified

OPHTHALMOSCOPIC EXAMINATION: Yes

HAEMATOLOGY: Yes
- Time schedule for collection of blood:
- Anaesthetic used for blood collection: Yes, isoflurane anesthesia.
- Animals fasted: Yes

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Blood samples for clinical pathology evaluations (hematology and serum chemistry) werecollected from 5 animals/sex/group at the scheduled necropsies (study day 28 or 29 for males and lactation day 14 for (females).
- Animals fasted: Yes

PLASMA/SERUM HORMONES/LIPIDS: Yes
- Time of blood sample collection: Blood samples for clinical pathology evaluations (hematology and serum chemistry) werecollected from 5 animals/sex/group at the scheduled necropsies (study day 28 or 29 for males and lactation day 14 for females).
- Animals fasted: Yes

URINALYSIS: Not specified

NEUROBEHAVIOURAL EXAMINATION: Not specified

IMMUNOLOGY: Not specified

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Other examinations:

THYROID HORMONE ANALYSIS
Blood samples for thyroid hormone analysis were collected as follows. Blood (approximately 1.0 mL) was collected via the retro-orbital sinus following isoflurane anesthesia from all F0 males and females on the day of scheduled euthanasia (study day 28 or 29 for males and lactation day 14 for females). On PND 4, blood (as much as possible; pooled by litter) was collected via cardiac puncture under isoflurane inhalation from 1 culled pup/sex/litter. On PND 13, blood (approximately 1.0 mL) was collected via cardiac puncture under isoflurane inhalation from 1 pup/sex/litter. Blood was collected into tubes containing no anticoagulant and allowed to clot at room temperature. Serum was isolated in a refrigerated centrifuge and stored frozen (approximately -70 °C).

Thyroxine (T4) concentrations were evaluated for F0 males and F1 PND 13 males and females. The samples from the F0 females and F1 culled pups (PND 4) were not analyzed and were discarded.

Statistics:

Each mean was presented with the standard deviation (S.D.), standard error (S.E.), and the number of animals (N) used to calculate the mean. Due to the use of significant figures and the different rounding conventions inherent in the types of software used, the means, standard deviations, and standard errors on the summary and individual tables may differ slightly. Therefore, the use of reported individual values to calculate subsequent parameters or means will, in some instances, yield minor variations from those listed in the report data tables. Data obtained from nongravid females were excluded from statistical analyses following the matingperiod. Statistical analyses were not conducted on F0 weekly female body weight data after 1 or more animals had entered the gestation phase. Where applicable, the litter was used as the experimental unit.

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 item-treated group to the control group by sex. Parental mating, fertility, conception, and copulation indices were analyzed using the Chi-square test with Yates’ correction factor (Hollander and Wolfe, 1999). Parental body weights (weekly, gestation, and lactation), body weight changes, and food consumption, offspring body weights and body weight changes, pre-coital intervals, gestation length, numbers of former

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Test item-related clinical observations of red and/or clear material around the mouth were noted for males and females in all test item-treated groups approximately 1 hour following dose administration. These observations were noted throughout the treatment period and occurred in a dose-related manner. However, the material observations did not persist to the weekly examinations and were considered nonadverse in the absence of other signs of systemic toxicity. There were no test item-related clinical observations noted at the weekly examinations. Observations noted in the test item-treated groups, including hair loss on various body surfaces and yellow material in the urogenital area, occurred infrequently, at similar frequencies in the control group, and/or in a manner that was not dose-related.

Mortality:

no mortality observed

Description (incidence):

All males and females survived to the scheduled necropsy.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

MALES
Mean body weight gain in the 1000 mg/kg/day group males was similar to the control group during the pre-mating treatment period (study days 0-13) and during study days 13-21. A test item-related, significantly (p < 0.01) lower mean body weight gain was noted in this group during study days 21-27 compared to the control group. As a result, a lower mean body weight gain was noted in the 1000 mg/kg/day group when the entire treatment period (study days 0-27) was evaluated; the difference from the control group was significant (p < 0.05). The effect on mean body weight gain during the last week of treatment was not of sufficient magnitude to affect absolute mean body weights for this group, and therefore, was not considered adverse. Mean body weights and body weight gains in the 100 and 300 mg/kg/day group males were unaffected by test item administration throughout the study. None of the differences from the control group were statistically significant.

FEMALES (pre-mating)
Mean body weights and body weight gains in the 100, 300, and 1000 mg/kg/day group females were unaffected by test item administration during the pre-mating period. None of the differences from the control group were statistically significant.

FEMALES (gestation)
Mean body weights and body weight gains in the 100, 300, and 1000 mg/kg/day group females were unaffected by test item administration during the pre-mating period. None of the differences from the control group were statistically significant.

FEMALES (lactation)
Mean body weights and body weight gains in the 100, 300, and 1000 mg/kg/day groups were unaffected by test item administration during lactation. None of the differences from the control group were statistically significant.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

MALES
Mean food consumption, evaluated as g/animal/day, in the 1000 mg/kg/day group males was significantly (p < 0.01) lower (2 g/animal/day) than the control group during the first week of treatment (study days 0-7). This decrement was noted in the absence of an effect on mean body weight gain during this same interval. Mean food consumption in the 1000 mg/kg/day group was similar to the control group during study days 7-13, and therefore the effect on mean food consumption noted during the first week of test item administration was considered transient and not test item-related. Mean food consumption in the 100 and 300 mg/kg/day group males was similar to that in the control group throughout the study. None of the differences from the control group were statistically significant.

FEMALES (pre-mating)
Mean food consumption, evaluated as g/animal/day, in the 100, 300, and 1000 mg/kg/day group females was unaffected by test item administration during the pre-mating period. None of the differences from the control group were statistically significant.

FEMALES (gestation)
Mean maternal food consumption, evaluated as g/animal/day and g/kg/day, in the 100, 300, and 1000 mg/kg/day groups was unaffected by test item administration during gestation. None of the differences from the control group were statistically significant.

FEMALES (lactation)
Mean maternal food consumption, evaluated as g/animal/day and g/kg/day, in the 100, 300, and 1000 mg/kg/day groups was unaffected by test item administration during lactation. Differences from the control group were not statistically significant, occurred in a manner that was not dose-related, and/or were transient.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not specified

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

There were no test item-related changes in hematology and coagulation parameters.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Test item-related higher cholesterol, alkaline phosphatase (ALP), alanine aminotransferase (ALT), and bile acids (1000 mg/kg/day group females), lower glucose (1000 mg/kg/day group males and females), higher phosphorus (300 and 1000 mg/kg/day group males, 1000 mg/kg/day group females) and higher urea nitrogen (1000 mg/kg/day group females) values were noted at study week 4 (males) or lactation day 14 (females). Minimal to mild elevations of cholesterol, ALP, ALT, and bile acids in 1000 mg/kg/day group females were coincident with test item-related hepatocellular hypertrophy; differences from the control group were significant (p < 0.05 or p < 0.01) for cholesterol, ALT, and bile acids. Minimally lower glucose was not associated with lower food consumption. Minimally to mildly higher phosphorus (significant at p < 0.01 or p < 0.05, males and p < 0.05, females) and urea nitrogen (significant at p < 0.01, females) were suggestive of dehydration but urinalysis was not performed in the present study. Glucose, phosphorous and urea changes were considered nonadverse given the magnitude of change. There were no other test item-related effects on serum chemistry parameters. However, some statistically significant differences were observed when the control and test item-treated groups were compared. Lower urea nitrogen in the 100 mg/kg/day group males was not considered test item-related given the lack of a dose related response and a high control group mean and individual values. There were no test item-related changes in T4 levels in males.

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

effects observed, non-treatment-related

Description (incidence and severity):

HOME CAGE OBSERVATIONS: Home cage parameters were unaffected by test item administration. There were no statistically significant differences for the test item-treated groups when compared to the control group during study week 3 (males) or on lactation day 13 (females).

HANDLING OBSERVATIONS: Handling parameters were unaffected by test item administration. There were no statistically significant differences for the test item-treated groups when compared to the control group during study week 3 (males) or on lactation day 13 (females).

OPEN FIELD OBSERVATIONS: Open field parameters were unaffected by test item administration. A significantly (p < 0.05) higher mean rearing count was noted in the 1000 mg/kg/day group males during study week 3 compared to the control group. However, in the absence of effects on any other effects on CNS activity, the higher rearing count was not considered to be test item-related. There were no other statistically significant differences for the test item-treated groups when compared to the control group during study week 3 (males) or on lactation day 13 (females).

SENSORY OBSERVATIONS: Sensory parameters were unaffected by test item administration. There were no statistically significant differences for the test item-treated groups when compared to the control group during study week 3 (males) or on lactation day 13 (females).

NEUROMUSCULAR OBSERVATIONS: Neuromuscular parameters were unaffected by test item administration. A significantly (p < 0.05) lower mean hindlimb grip strength value was noted in the 300 mg/kg/day group males during study week 3 compared to the control group value. However, no dose-response was evident, and therefore this difference was not considered test item-related. There were no other statistically significant differences for the test item-treated groups when compared to the control group during study week 3 (males) or on lactation day 13 (females).

PHYSIOLOGICAL OBSERVATIONS: Physiological parameters were unaffected by test item administration. There were no statistically significant differences for the test item-treated groups when compared to the control group during study week 3 (males) or on lactation day 13 (females).

MOTOR ACTIVITY: Motor activity patterns (total activity as well as ambulatory activity counts) in F0 animals were unaffected by test item administration at all concentrations when evaluated during study week 3 (males) and on lactation day 13 (females). Significantly (p ≤ 0.008) lower mean total counts were noted in the 300 and 1000 mg/kg/day group males during the 11-20 minute subinterval. However, the differences were the result of a greater habituation response in these groups relative to the control group during the first 20 minutes of testing. When rats are placed in a novel environment, exploratory behavior (resulting in a high level of activity counts) followed by habituation (resulting in a reduced level of activity counts) is an expected response. In addition, there were no statistically significant differences in the cumulative total counts at these dosage levels. Therefore, the lower activity counts in the 300 and 1000 mg/kg/day group males during the 11-20 minute subinterval were attributed to biologic variability and were not considered test item-related. All other values obtained from the 6 subintervals evaluated (0-10, 11-20, 21-30, 31-40, 41-50 and 51-60 minutes) and the overall 60-minute test session values were comparable to the concurrent control values and the WIL Research historical control data. Differences from the control group were slight, not statistically significant when analyzed by a repeated measures analysis, within the WIL Research historical control data ranges, and/or did not occur in a dose-related manner. Other than the greater habituation response described previously for the 300 and 1000 mg/kg/day males during the 11-20 minute subinterval, no remarkable shifts in the pattern of habituation occurred in any of the test item-treated groups when the F0 animals were evaluated during study week 3 or lactation day 13.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Test item-related higher liver weights were noted in the 1000 mg/kg/day group males and females. Higher liver weights (absolute, relative to body and brain weight) were associated with test item-related hepatocellular hypertrophy and the weight relative to body weight was significant (p < 0.01) in males. Some organ weight differences were statistically significant when compared to the control group but were considered to be a result of a non-test item-related higher body weight. The thyroid/parathryoid weight relative to brain weight was higher in 1000 mg/kg/day group females. There were no other test item-related effects on organ weights. However, some statistically significant differences were observed when the control and test item-treated groups were compared. Higher mean Cowper’s gland weights were noted in the 100 (absolute, relative to brain and body weight), 300 (absolute and relative to body weight), and 1000 (relative to body weight) mg/kg/day group males but were not considered test item-related given the lack of a dose-related response and lack of correlating microscopic changes.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Test item-related dark red discoloration of the liver was noted in 100, 300, and 1000 mg/kg/day group males and correlated with angiectasis in single males in the 300 (no. 2827) and 1000 (no. 2797) mg/kg/day groups. Dark red discoloration in remaining animals in the 100, 300, and 1000 mg/kg/day groups correlated with sinusoidal congestion or was not associated with a relevant gross finding in 1 male (no. 2813) in the 1000 mg/kg/day group. Descriptions of test item-related histologic alterations are discussed in Section 6.1.9.3.
The mean numbers of unaccounted-for sites and implantation sites in the 100, 300, and 1000 mg/kg/day groups were similar to the control group values.

Neuropathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Test item-related microscopic findings were noted in the adrenal cortex of the 1000 mg/kg/day group males and liver of the 1000 mg/kg/day group males and females.

Liver: Minimal to mild hepatocellular hypertrophy, characterized by increased amounts of cytoplasm, was noted in the centrilobular region of the 300 and 1000 mg/kg/day group males and 1000 mg/kg/day group females. Angiectasis, characterized by focally extensive dilation of sinuosoids, was observed in a single male each in the 300 and 1000 mg/kg/day groups (male nos. 2827 and 2797, respectively) and correlated with dark red discoloration in the liver. Angiectasis was differentiated from routine terminal congestion that was noted in the control and all test item-treated groups and was characterized by blood in the sinusoids.

Adrenal cortex: There was an increased incidence and severity of cytoplasmic vacuolation of the adrenal cortex in the 1000 mg/kg/day group males. Vacuolation was characterized by variably sized, well demarcated vacuoles in the zona fasciculata and was not considered adverse. Vacuolation was also noted in females but vacuoles were uniform and there was no test item-related difference.

There were no other test item-related histologic changes. Remaining histologic changes were considered to be incidental findings. There was no test item-related alteration in the prevalence, severity, or histologic character of those incidental tissue alterations. Female no. 2852, which had a total litter loss, had mild myeloid hyperplasia of the bone marrow and a moderate increase in splenic extramedullary hematopoeisis which were both related to moderate uterine subacute inflammation with endometrial hyperplasia and mild mixed cell infiltrate in the vagina. Female no. 2846, which failed to conceive, was in early diestrus at the time of necropsy (study day 39).

Histopathological findings: neoplastic:

no effects observed

Description (incidence and severity):

REPRODUCTIVE PERFORMANCE
No test item-related effects on reproductive performance were observed at any dosage level. No statistically significant differences were noted between the control and test item-treated groups. One male in the 1000 mg/kg/day group did not sire a litter, as the female that this male was paired with was nongravid. The mean numbers of days between pairing and coitus in the test item-treated groups were similar to the control group value. None of these differences were statistically significant.

GESTATION LENGTH AND PARTURITION
The mean gestation length in the 1000 mg/kg/day group (22.3 days) was significantly (p < 0.05) longer than the concurrent control group (21.7 days). The 1000 mg/kg/day group gestation length value was within the WIL Research historical control range (21.3 to 22.3 days) and was primarily attributed to a single female (no. 2852) that delivered on gestation day 24 (and subsequently had a total litter loss on lactation day 0). Therefore, the longer mean gestation length at 1000 mg/kg/day was not considered test item-related. Mean gestation lengths in the 100 and 300 mg/kg/day groups (21.7 and 22.0 days, respectively) were similar to those in the concurrent control group. No signs of dystocia were noted in any group.

Details on results:

LITTER DATA

POSTNATAL SURVIVAL
The mean number of pups born, live litter size, the percentage of males at birth, and postnatal survival between birth and PND 0 (relative to number born), PND 0-1, 1-4 (pre-selection), 4-7 (post-selection), 7-10, 10-13, and from birth to PND 4 (pre-selection) and PND 4 (post-selection) to PND 13 were unaffected by test item administration at all dosage levels. Differences form the control group were slight, were not statistically significant, and/or did not occur in a dose-related manner. Slightly lower postnatal survival was noted during PND 0 relative to birth and when birth to PND 4 was evaluated in the 1000 mg/kg/day group; however, these differences were not statistically significant compared to the control group and were attributed to 1 female (no. 2852) in this group that had a total litter loss on PND 0.

GENERAL PHYSICAL CONDITION
The general physical condition (defined as the occurrence and severity of clinical findings) of all F1 pups in this study was unaffected by parental administration of the test item. Pups (litters) that were found dead numbered 5(5), 9(5), 3(2), and 14(5) in the control, 100, 300, and 1000 mg/kg/day groups, respectively. One, 2, and 2 pups in the control, 300, and 1000 mg/kg/day groups, respectively, were missing and presumed to have been cannibalized.

ANOGENITAL DISTANCE
The anogenital distances (absolute and relative to the cube root of pup body weight) in the 100, 300, and 1000 mg/kg/day groups were similar to the control group values. Differences from the control group were slight and not statistically significant.

OFFSPRING BODY WEIGHTS
Mean male and female pup body weights and body weight changes during PND 1-13 in the 100, 300, and 1000 mg/kg/day groups were unaffected by parental administration of the test item. No statistically significant differences from the control group were noted.

AREOLAE/NIPPLE ANLAGEN
Areolae/nipple anlagen in the F1 male pups was not affected by parental administration of the test item. The test item group values for males were not statistically significantly different from the control group.

THYROID HORMONE ANALYSIS (PND 13)
There were no test item-related changes in T4 levels in F1 males or females on PND 13.

ORGAN WEIGHTS (PND 13)
There were no test item-related changes in thyroid/parathyroid weights for F1 pups at 100, 300, and 1000 mg/kg/day.

NECROPSIES OF PUPS FOUND DEAD
The numbers of pups (litters) found dead during PND 0-13 numbered 5(5), 9(5), 3(2), and 14(5) in the control, 100, 300, and 1000 mg/kg/day groups, respectively. No internal findings that could be attributed to parental test item administration were noted at the necropsies of pups that were found dead. Aside from the presence or absence of milk in the stomach, the following internal findings were noted. Pup no. 2882-10 in the 1000 mg/kg/day group was noted with a malformation consisting of a right-sided aortic arch (aortic arch and descending aorta coursed to the right of the vertebral column; the right carotid and subclavian arteries arose independently from the aortic arch [no brachiocephalic trunk]; and the left carotid and subclavian arteries arose from a common vessel from the aortic arch). Dark red subcutis contents were noted for pup no. 2871-01 in the 1000 mg/kg/day group. In the 100 mg/kg/day group, pup no. 2873-15 was noted with clear fluid in the abdominal cavity and renal papilla(e) not developed.

Key result

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical signs

food consumption and compound intake

gross pathology

histopathology: non-neoplastic

organ weights and organ / body weight ratios

Remarks on result:

other: NOAEL is the highest dose (limit dose) of 1000 mg/kg bw/day.

Critical effects observed:

no

Conclusions:

There were no adverse test item-related clinical observations or effects on mean body weights, body weight changes, and food consumption noted at any dosage level. Furthermore, there were no adverse effects on organ weights, clinical pathology parameters, thyroid hormones, or macroscopic/microscopic alterations in F0 males and females at any dosage levels. Therefore, the NOAEL for systemic toxicity was considered to be 1000 mg/kg/day. The NOAEL for postnatal toxicity was 1000 mg/kg/day based on the absence of effects on F1 offspring at all dosage levels.

Executive summary:

A screening study according to the OECD guideline 422 under GLP compliance was performed with the test substance. Under the conditions of this screening study, no test item-related effects were observed on reproductive performance at any dosage level. Therefore, a dosage level of 1000 mg/kg/day was considered to be the no-observed-adverse-effect level (NOAEL) for reproductive toxicity of 1,4-diisopropylbenzene when administered orally by gavage to Crl:CD(SD) rats. There were no adverse test item-related clinical observations or effects on mean body weights, body weight changes, and food consumption noted at any dosage level. Furthermore, there were no adverse effects on organ weights, clinical pathology parameters, thyroid hormones, or macroscopic/microscopic alterations in F0 males and females at any dosage levels. Therefore, the NOAEL for systemic toxicity was considered to be 1000 mg/kg/day. The NOAEL for postnatal toxicity was 1000 mg/kg/day based on the absence of effects on F1 offspring at all dosage levels.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Repeated dose oral toxicity

A screening study according to the OECD guideline 422 under GLP compliance was performed with the test substance. Under the conditions of this screening study, no test item-related effects were observed on reproductive performance at any dosage level. Therefore, a dosage level of 1000 mg/kg/day was considered to be the no-observed-adverse-effect level (NOAEL) for reproductive toxicity of 1,4-diisopropylbenzene when administered orally by gavage to Crl:CD(SD) rats. There were no adverse test item-related clinical observations or effects on mean body weights, body weight changes, and food consumption noted at any dosage level. Furthermore, there were no adverse effects on organ weights, clinical pathology parameters, thyroid hormones, or macroscopic/microscopic alterations in F0 males and females at any dosage levels. Therefore, the NOAEL for systemic toxicity was considered to be 1000 mg/kg/day. The NOAEL for postnatal toxicity was 1000 mg/kg/day based on the absence of effects on F1 offspring at all dosage levels.

Justification for classification or non-classification

According to the criteria of Classification, Labelling and Packaging of Substances and Mixtures Regulation (EC) No 1272/2008 (CLP Regulation), a classification of 1.4-diisopropylbenzene is not warranted.