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Toxicity to reproduction

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Administrative data

Endpoint:
one-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 2010-09-17 to 2011-06-10
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP and guideline conform study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report date:
2011

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 415 [One-Generation Reproduction Toxicity Study (before 9 October 2017)]
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Allyl 3-cyclohexylpropionate
EC Number:
220-292-5
EC Name:
Allyl 3-cyclohexylpropionate
Cas Number:
2705-87-5
Molecular formula:
C12H20O2
IUPAC Name:
allyl 3-cyclohexylpropanoate
Details on test material:
- Name of test material (as cited in study report): Allyl cyclohexanepropionate
- Physical state: colorless, clear liquid
- Analytical purity: 99.9%
- Impurities (identity and concentrations): <0.01% allyl alcohol

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc, Portage, MI
- Age at study initiation: (P) x 10 wks
- Weight at study initiation: (P) Males: 320-354 g; Females: 200-230 g
- Housing: individually (P), except during the cohabitation and postpartum periods
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 to 25
- Humidity (%): 30 to 70
- Air changes (per hr): a minimum of 10 changes per hour of 100% fresh air
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
VEHICLE
- Concentration in vehicle: 0, 18.75, 31.25, 62.5 and 125 mg/mL
- Amount of vehicle (if gavage): 4 mL
- Lot/batch no. (if required): J-145
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation: maximum duration of 7 days
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0] of pregnancy
- After successful mating each pregnant female was caged (how): individually
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The results for all dose formulations met the acceptance criteria for concentration and homogeneity.
Duration of treatment / exposure:
P generation male rats were given the test substance and/or vehicle once daily beginning at least 14 days before cohabitation, through cohabitation (maximum duration of 7 days), and continuing through the day before scheduled euthanasia.
P generation female rats were given the test substance and/or the vehicle once daily beginning at least 14 days before cohabitation, through cohabitation (maximum duration of 7 days), and continuing through day 25 of presumed gestation (rats that did not deliver) or day 4 postpartum (rats that delivered a litter).
Frequency of treatment:
daily
Details on study schedule:
not applicable
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 75, 125, 250 and 500 mg/kg/day
Basis:
nominal conc.
No. of animals per sex per dose:
8
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: an acute oral test was conducted for the dose selection: LD50=1.05 g/kg bw
Positive control:
no data

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule:
Aclimation period: at least weekly.
Dosage period: Daily before dosage. On the first day of dosage, postdosage observations is recorded at approximately hourly intervals for the first four hours andat the end of the normal working day.
Postdosage period: daily.

BODY WEIGHT: Yes
- Time schedule for examinations:
Acclimation period: at least weekly
Dosage period: daily
Day of euthanasia: prior to euthanasia

Oestrous cyclicity (parental animals):
The average number of estrous stages per 14 days was reduced at 250 mg/kg/day, in comparison to the vehicle control group value (1.4 stages/14 days vs. 3.4 stages/14 days in vehicle controls). In turn, administration of allyl cyclohexanepropionate increased the number of rats that had 6 of more days of diestrus. Six out of the 8 female rats had 1 estrous stage per 14 days, while the remaining two rats cycled 2 or 3 times within the
14 day evaluation period. The number of estrous stages per 14 days was comparable between the 0 (Vehicle), 75 and 125 mg/kg/day dosage groups during the precohabitation period.
Sperm parameters (parental animals):
No observations reported
Litter observations:
No treatment-related clinical observation occured in the F1 generation pups. All pups that were found dead or that survived to scheduled euthanasia appeared normal at necropsy examination.
Postmortem examinations (parental animals):
GROSS NECROPSY
- Gross necropsy consisted of [external and internal examinations including the cervical, thoracic, and abdominal viscera.]

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated in Table [#] were prepared for microscopic examination and weighed, respectively.
Postmortem examinations (offspring):
Reflecting excessive toxicity and mortality at 500 mg/kg/day and group termination at 250 and 500 mg/kg/day, only data for delivered litters in the vehicle control, 75 and 125 mg/kg/day dosage groups are presented below. No treatment-related clinical observations occurred in the F1 generation pups. Clinical signs that occurred reflected injuries, and included a laceration along the midline, a scab along the lower midline and purple discoloration to the back. No other clinical signs occurred All pups that were found dead or that survived to scheduled euthanasia appeared normal at necropsy examination.
Statistics:
Data generated during the course of this study were recorded either by hand or using the Argus Automated Data Collection and Management System and the Vivarium Temperature and Relative Humidity Monitoring System. All data were tabulated and summarized using the Argus Automated Data Collection and Management System, the Vivarium Temperature and Relative Humidity Monitoring System and Quattro Pro 8.
Reproductive indices:
There were no apparent effects on mating and fertility at 75 and 125 mg/kg/day when compared to the vehicle control group values. However, at 250 mg/kg/day, all mating and fertility parameters, including the number of days in cohabitation, rats that mated, the fertility index and the rats pregnant/rats in cohabitation, reflected the overall pattern of group termination due to excessive toxicity. The two female rats in the 250 mg/kg/day dosage group that were confirmed mated by cohort males of the same dosage group were not pregnant upon examination. However, this finding likely reflects the abnormal estrous cycling (6 or more days in diestrus) that was observed in each of these females. Pregnancy occurred in 7, 6 and 8 rats in the 0 (vehicle), 75 and 125 mg/kg/day dosage groups, respectively. Each of these pregnant rats delivered a litter.
Offspring viability indices:
The average pup body weight on postpartum day 1was reduced by 12 % at 125 mg/kg/day; however, these reductions were transient and had resolved by postpartum day 5.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
250 and 500 mg/kg/day: decreased motor activity, urine-stained abdominal fur, slight excess salivation, mild dehydratation. 250 mg/kg/day: abnormal fecal output, mild and/or moderate dehydratation, thin body condition, hunched posture, etc.
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:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Other effects:
no effects observed
Description (incidence and severity):
Test substance intake: gavage

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
except 2 female rats in 250 mg/kg/day dose group, where abnormal estrous cycling (6 or more days in diestrus) was observed.
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Description (incidence and severity):
all mating and fertility parameters were unaffected by dosages of allyl cyclohexanepropionate as high as 125 mg/kg/day.

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
At 500 mg/kg/day, four male rats were found dead on Day 2 or 3 of Study (DS 2 or 3). As a result, dosage administration was terminated for this dosage group and the remaining rats in the dosage group were euthanized on DS 3. All male rats in the 250 mg/kg/day were euthanized early (DS 12 through 14 or DS 16) because of declining clinical condition. All other male rats survived to scheduled euthanasia. Prior to death or termination of the dosage group, common clinical signs observed in male rats in the 500 mg/kg/day dosage group included, but were not limited to, decreased dehydration (based on skin turgor). These clinical signs were generally observed following the second dose of allyl cyclohexanepropionate and continued until death. Common clinical signs observed at 250 mg/kg/day included, but were not limited to, abnormal fecal output (i.e., scant or soft or liquid feces), mild and/or moderate dehydration (based on skin turgor), thin body condition, hunched posture, decreased motor activity, piloerection, ungroomed coat, slight and/or moderate excess salivation, lacrimation, and chromorhinorrhea. Common gross lesions observed in the 250 and/or
500 mg/kg/day dosage group included a mottled and/or rough appearance to the liver, thickening of the liver lobes, discolored (red or dark red) lymph nodes, and red or dark red areas on the serosal surface of the stomach (one or more regions). Each of the clinical signs and gross lesions were attributed to oral administration of allyl cyclohexanepropionate.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
In general, male rats given 250 or 500 mg/kg/day of allyl cyclohexanepropionate lost body weight following the initiation of dosage administration. Feed consumption values for male rats in the 250 mg/kg/day were reduced to severely reduced in comparison to values for rats in the vehicle control group.

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS)
P generation male rats were given the test substance and/or the vehicle once daily beginning at least 14 days before cohabitation, through cohabitation (maximum duration of 7 days), and continuing through the day before scheduled euthanasia. P generation female rats were given the test substance and/or the vehicle once daily beginning at least 14 days before cohabitation, through cohabitation (maximum duration of 7 days), and continuing through day 25 of presumed gestation (rats that did not deliver) or day 4 postpartum (rats that delivered a litter).

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
The average number of estrous stages per 14 days was reduced at 250 mg/kg/day, in comparison to the vehicle control group value (1.4 stages/14 days vs. 3.4 stages/14 days in vehicle controls). In turn, administration of allyl cyclohexanepropionate increased the number of rats that had 6 of more days of diestrus. Six out of the 8 female rats had 1 estrous stage per 14 days, while the remaining two rats cycled 2 or 3 times within the 14 day evaluation period. The number of estrous stages per 14 days was comparable between the 0 (Vehicle), 75 and 125 mg/kg/day dosage groups during the precohabitation period.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
Reflecting excessive toxicity and mortality at 500 mg/kg/day and group termination at 250 and 500 mg/kg/day, only data for rats in the vehicle control, 75 and 125 mg/kg/day dosage groups are presented below. Pregnancy occurred in 7, 6 and 8 rats in the 0 (Vehicle), 75 and 125 mg/kg/day dosage groups, respectively. Each of these rats delivered a litter. No other natural delivery and litter observations were affected by dosages of allyl cyclohexanepropionate as high as 125 mg/kg/day. Values for the numbers of dams delivering litters, the duration of gestation, averages for implantation sites per delivered litter, the gestation index (number of dams with one or more liveborn pups/number of pregnant rats), the numbers of dams with stillborn pups and of dams with all pups dying, litter sizes, viability and lactation indices, surviving pups per litter, percent male pups per number of pups sexed per litter and live litter size at weighing were comparable among the three remaining dosage groups. All mating and fertility parameters were unaffected by dosages of allyl cyclohexanepropionate as high as 125 mg/kg/day.

ORGAN WEIGHTS (PARENTAL ANIMALS)
Terminal body weights were comparable among the three remaining dosage groups. The weights of the epididymides, testes, seminal vesicles (with and without fluid) and prostate and the ratios of these organ weights to terminal body weight were unaffected by dosages of allyl cyclohexanepropionate as high as 125 mg/kg/day. The weights of the ovaries and the non-gravid uterus (with cervix) and the ratios of these organ weights to terminal body weight were unaffected by dosages of allyl cyclohexanepropionate as high as 125 mg/kg/day. The apparent increase in the absolute and relative weights of the paired ovaries reflected one female rat (no. 1160) in the 125 mg/kg/day dosage group that had a paired ovarian weight of 0.410 g which exceeded the values for other rats in this dosage group (0.092 g to 0.153 g). In addition, this same rat had a uterine (with cervix) weight of 0.52 g which was lower than values for other rats in the same dosage group (0.71 g to 0.94 g). Exclusion of this rat’s data would result in a group mean weight of 0.122 g for the paired ovaries and 0.84 g for the uterus with cervix, which would be comparable to the values of the vehicle control and 75 mg/kg/day dosage groups.

HISTOPATHOLOGY (PARENTAL ANIMALS)
Mortality and morbidity was associated with the early development and severity of the multifocal necrosis and hemorrhage and chronic-active cholangiofibrosis observed in the livers of the treated P generation rats. These lesions appeared to have originated from periportal vacuolation and degenerative changes that developed into pronounced hepatic parenchymal necrosis. This resulted in initial oval cell hyperplasia leading to bile duct hyperplasia surrounded by inflammatory cell infiltrates and connective tissue proliferation. In some P generation rats, these complex changes are associated with the resolving necrosis and hemorrhage or sclerotic changes and early evidence of regenerative hepatocellular hyperplasia. The P generation male rats appeared to be slightly more severely affected than the P generation female rats. These liver changes were seen with increasing severity and/or incidence in all dosages of both sexes and were considered test article-related.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Effect level:
< 75 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see 'Remark'
Dose descriptor:
NOAEL
Remarks:
for reproduction
Effect level:
125 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: The highest survived P dose group

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
litter
Sexual maturation:
not specified
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
no effects observed
Histopathological findings:
not specified

Details on results (F1)

VIABILITY (OFFSPRING): 100, 97.6 and 100 % for vehicle, 75 and 125 g/kg/day group, respectively.

CLINICAL SIGNS (OFFSPRING): purple discoloration on the back in 125 g/kg/day dose group, but it was considered as no treatment-related clinical observation.

BODY WEIGHT (OFFSPRING): litter weight Day 1: 5.8 - 6.6 g, Day 5: 10.0 - 10.2 g

Overall reproductive toxicity

Reproductive effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
The purpose of this study was to provide information for the selection of dosage levels to be used in the oral (gavage) one-generation reproduction study of allyl cyclohexanepropionate in Crl:CD(SD) rats, to provide preliminary data on the reproductive function of P generation male and female rats, and on the growth and development of F1 generation offspring to day 5 postpartum.
Administration of allyl cyclohexanepropionate at dosage levels of 250 or 500 mg/kg/day increased the incidence of adverse clinical signs. In P generation male rats, net loses on body weight occurred early in the dosage period at 250 and 500 mg/kg/day. At 75 and 125 mg/kg/day, body weight gains were reduced for the first two weeks of the dosage period, and overall for the cumulative dosage period. Corresponding reductions in feed consumption occurred overall for the first two weeks of the dosage period at >=75 mg/kg/day, but were most pronounced at 250 mg/kg/day.
Based on the results of this study, the no-observable-adverse-effect-level (NOAEL) for parental toxicity of allyl cycolhexylpropionate is <75 mg/kg/day. The reproductive NOAEL is 125 mg/kg/day.
Executive summary:

The oral (gavage) dose-range reproduction study of allyl cyclohexanepropionate in rats was performed in a GLP test according to principles of OECD 415 guideline. Eight animals per sex per group were exposed to oral (gavage) doses of 0 (vehicle), 75, 125, 250 and 500 mg/kg/day, administered at volume of 4 mL/kg. P generation rats were given the test substance and/or vehicle once daily beginning at least 14 days before cohabitation, through cohabitation (maximum duration of 7 days), and for male rats, continuing through the day before scheduled euthanasia. For female rats, the daily gavage was performed through day 25 of presumed gestation (rats that did not deliver) or day 4 postpartum (rats that deliver a litter). Male rats were euthanized after completion of the cohabitation period. Female rats were allowed to deliver their litters and were euthanized on day 5 postpartum. F1 generation pups were euthanized on day 5 postpartum. Allyl cyclohexanepropionate increased the incidence of mortality and clinical signs (dehydratation, excess salivation, reduced grooming, decreased motor activity, hunched posture, piloerection and/or abnormal fecal output) in P generation male and female rats given 250 or 500 mg/kg/day. In these groups, gross lesions attributed to administration of allyl cyclohexanepropionate were observed primarily in the liver, intestines, stomach, spleen and lymph nodes. At lower dosages of 75 and 125 mg/kg/day, increased incidence of excess salivation was observed, however this clinical sign was not considered an adverse effect of the test substance. In P generation male rats, net losses in body weight, with corresponding reductions in the group men body weight, occurred early in the dosage period at 250 mg/kg/day and 500 mg/kg day (prior to this group termination). At lower dosages (75 and 125 mg/kg/day), body weight gains were reduced for the first two weeks of the dosage period, and overall for the cumulative dosage period. Corresponding reductions in feed consumption occurred overall for the first two weeks of the dosage period at ≥75 mg/kg/day, but were most pronounced at 250 mg/kg/day. In P generation female rats, allyl cyclohexanepropionate reduced body weight gains and feed consumption values at 250 mg/kg/day during premating period. Of the rats that mated in the 250 mg/kg/day dosage group (N=2), there were no successful pregnancies. These effects are consistent with the prominent systemic toxicity in this dosage group. There were no apparent effects on natural delivery parameters in the mated female rats at 75 or 125 mg/kg/day. Reproductive organ weights were unaffected by oral administration of allyl cyclohexanepropionate. Multifocal necrosis, periportal vacuolation of hepatocytes, and colangiofibrosis were observed in both P generation male and female rat livers from all treated groups. Rats that did not survive to scheduled euthanasia also had evidence of hemorrhages in the lungs, stomach and/or intestinal serosa and multifocal erosions in the gastric mucosa. F1 pup body weights were reduced on day 1 postpartum, but this observation was resolved by day 5 postpartum. Based on the results of this study, the no-observable-adverse-effect-level (NOAEL) for toxicity of allyl cyclohexanepropionate is <75 mg/kg/day. The reproductive NOAEL is 125 mg/kg/day.