Registration Dossier

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2001-07-18 - 2002-04-17
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guideline and the study was conducted under GLP conditions.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2002
Report Date:
2002

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD Guidelines for Testing of Chemicals, No.415 “One Generation Reproduction Toxicity Study” (adopted 26 May 1983).
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Description : clear amber coloured liquid
Date received : 6/7/2001, provided by Chevron Research and Technology Comapny
Expiry date : 05/01/2002

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
Test Animals
- Source: from Charles River Laboratories, Inc., Kingston, New York,
- Age and weight at study initiation: the males were approximately six weeks of age with body weights ranging from 134 to 166 grams, while the females were approximately seven weeks of age with body weights ranging from 163 to 204 grams.
- Housing: Males were housed two or three per cage for three days to allow the animals to adapt to the automatic watering system. The males were housed individually for the remainder of acclimation and while on study (except during cohabitation) in suspended stainless steel cages. Females were housed individually during acclimation and while on study (except during cohabitation) in suspended stainless steel cages.
During the mating phase, animals were paired on a one male: one female basis within each dose group in polypropylene cages
Following mating the males were re-housed in their original cages and the females were individually housed in solid floor polypropylene cages with stainless steel mesh lids and furnished with softwood flake bedding for the gestation, birth and lactation phases of the study.
- Diet and water: PMI Certified Rodent Chow@ #5002 (Purina Mills, Inc.) and municipal tap water were provided to each animal ad libitum.
- Acclimation period: Males were acclimated to the laboratory conditions for a period of 16 days prior to in-life initiation. Females were acclimated to the laboratory conditions for a period of 22 days prior to in-life initiation.

ENVIRONMENTAL CONDITIONS
- Temperature: 65 to 79 °F (18 to 26 °C)
- Humidity: 30 to 70 %
- Air changes (per hr): 10 to 15 air changes per hour.
- Photoperiod (hr dark / hrs light): 12 hours continuous light and 12 hours darkness

Administration / exposure

Route of administration:
oral: feed
Type of inhalation exposure (if applicable):
other:
Vehicle:
corn oil
Remarks:
Basal laboratory diet
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
- For each test article dose group, a specified amount of HPV-1 (CAS #67124-09-8) was weighed into a pre-calibrated beaker. A sufficient quantity of corn oil was added to the beaker to achieve the desired volume and the mixture was stirred for 30 minutes. Each test article dosing mixture was prepared fresh weekly and stored at ambient conditions for a maximum of one week. During each preparation, corn oil was dispensed as received into daily aliquots for administration to control animals. The physical state of the control and each dosing solution was recorded during each preparation. The vehicle (group 1) was a clear yellow liquid, groups 2 and 3 were clear yellow solutions and group 4 was a clear dark yellow solution. The dosing solutions were stirred prior to dispensation and continuously until dosing was complete. The Study Director visually inspected the first preparation prior to initiation of dosing and found the mixtures acceptable for dosing.

VEHICLE
- Concentration in vehicle: 50, 167 and 500 mg/kg/day groups
- Amount of vehicle (if gavage): 2.5 mL/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Homogeneity, Stability and Concentration Analyses:
Prior to the first day of dosing, samples were collected for homogeneity and stability analyses from the three test article preparations. Three 5 mL samples each were collected from the top, middle and bottom of each concentration (for a total of 27 aliquots). Two of the three sets of samples were sent to Chevron Research and Technology Company, lntegrated Laboratory Technologies, under ambient conditions for analysis of homogeneity and stability over a seven-day period. The remaining set of samples was stored at SLI under ambient conditions. For each dosing preparation, including the vehicle control, two 5 mL samples were taken from each dose level (middle stratum) for concentration analysis. One sample from each dose level was sent to Chevron Research and Technology Company, lntegrated Laboratory Technologies, under ambient conditions for analysis of concentration. The remaining sample from each dose level was stored at SLI under ambient conditions.
Results of homogeneity analyses indicate that the test article was homogeneous in the vehicle at concentrations of 20, 66.8 and 200 mg/mL and stable for more than one week when stored under ambient conditions. Results of concentration analyses verified the appropriate concentration of test article in the dosing mixtures.
Details on mating procedure:
- M/F ratio per cage: 1/1 (Animals were paired on a 1 male: 1 female basis within each dose group)
- Proof of pregnancy: Evidence of mating was determined by the presence of a copulatory plug in the vagina or a sperm positive vaginal smear. The day evidence of copulation was confirmed was designated as day 0 of gestation and the female was separated from her mate and returned to her individual cage.
- After successful mating each pregnant female was caged: Mated females were housed individually during the period of gestation and lactation.
Duration of treatment / exposure:
Oral administration of HPV-1 (CAS #67124-09-8) to F0 male rats for a minimum of 70 days prior to mating and until completion of parturition
F0 females were treated for a minimum of 14 days prior to mating through lactation day 20.
Frequency of treatment:
once daily oral gavage
Duration of test:
Oral administration of HPV-1 (CAS #67124-09-8) to F0 male rats for a minimum of 70 days prior to mating and until completion of parturition
F0 females were treated for a minimum of 14 days prior to mating through lactation day 20.
No. of animals per sex per dose:
28 animals per sex per group
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dosage levels were selected by the Sponsor Monitor in an attempt to produce graded responses to the test article. The high-dose level was expected to produce some toxic effects, but not excessive lethality. The mid-dose level was expected to produce none to minimal observable effects and the low-dose level was expected to produce no observable effects.
- Rationale for animal assignment (if not random): Random
- Section schedule rationale (if not random): Random

Examinations

Maternal examinations:
CAGE SIDE AND DETAILED CLINICAL OBSERVATIONS:
- Yes
- Time schedule: Mortality/general health checks were performed twice daily, in the morning and afternoon. Detailed clinical observations were performed weekly and cage-side observations were performed daily approximately one-half hour to two hours following dosing. During gestation and lactation, detailed clinical observations were performed daily for FO females. Detailed clinical observations were also performed on the day of scheduled euthanasia.

BODY WEIGHT:
- Yes
- Time schedule for examinations: Individual body weights were recorded weekly and on the day of scheduled euthanasia for the males. Individual body weights were recorded weekly for the females until evidence of mating was observed. When positive evidence of mating was detected, the females were weighed on gestation days 0, 7, 14 and 21. Following parturition, the females were weighed on lactation days 1, 4, 7, 14 and 21.

FOOD CONSUMPTION:
- Yes. lndividual food consumption was recorded on the same days as body weights for the males and females, except during cohabitation and lactation

HAEMATOLOGY AND CLINICAL CHEMISTRY: Yes.
Parameters: Erythrocyte count (RBC), Hematocrit (Hct), Hemoglobin concentration (Hgb), Mean corpuscular hemoglobin (MCH), Mean corpuscular hemoglobin concentration (MCHC), Mean corpuscular volume (MCV), Platelet count, Reticulocyte count (reticulocyte slides were prepared but were not examined since no test article changes were noted in the erythrocyte parameters), Total and differential leukocyte counts
- Time schedule for collection of blood: Blood samples for hematology analysis were collected from ten males/group prior to scheduled euthanasia on study day 117/118 and from ten females/group prior to scheduled euthanasia on lactation day 21.
- Anaesthetic used for blood collection: Blood samples were obtained via the ocular orbital plexus while the animals were under light isoflurane anesthesia. H
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included: F0 female reproductive or accessory organs
- Number of implantations: Yes
- Early embryo lethality: Yes

Fetal examinations:
F1 Standardization of Litter Size
Following observations and body weights on lactation day 4, each litter was randomly culled to a maximum of eight pups, four per sex per litter, when possible.
F1 Litter Data
Pup viability was determined daily throughout lactation. A detailed examination of each pup was petformed on lactation days 0, 4, 7, 14 and 21. The sex of each pup was determined on lactation day 0 and verified on lactation days 4, 7, 14 and 21. Individual pup weights were determined on lactation days 1, 4, 7, 14 and 21.
Statistics:
Data, including body weights, body weight gain, food consumption, gestation length, mean live litter size and sperm parameters, were analyzed by ANOVA. If significance was observed with ANOVA, control to treatment group comparisons were performed using Dunnett's test. Count data were analyzed using R x C Chi-Square test followed by Fisher's Exact Test for copulation and fertility indices, pup sex ratios, the number of live and dead pups per group (on lactation day 0) and pup survival (after lactation day 0). Absolute and relative organ weights were analyzed for homogeneity of variance using Bartlett's test. If significance was detected with Bartlett's test (p < 0.01), multiple group comparisons proceeded using the Kruskal-Wallis non-parametric ANOVA, followed by Dunn's test, when p < 0.05. If significance was not detected with Bartlett's test, parametric procedures were used to analyze the data, i.e.,ANOVA followed by Dunnett's test when pc0.05. Hematology data were analyzed for homogeneity of variance using Levene's test (p < 0.01). If significance was detected with Levene's test (p<0.01), multiple group comparisons proceeded using the Kruskal-Wallis non-parametric ANOVA, followed by Dunn's test, when p < 0.05. If significance was not detected with Levene's test, parametric procedures were used to analyze the data, i.e., ANOVA followed by Tukey-Kramer test when p < 0.05. Statistical analyses regarding the hematology data were performed using the SLI Alpha GenTox computer system. All other statistical analyses were performed using the SLI Alpha ReproTox computer system. All analyses were two-tailed with a minimum significance level of 5 % (p < 0.05).
Indices:
Reproduction index
Semen analysis
Mating index
Fertility index
Precoital interval
Gestation length
F1 Put viability


F1 viability index
Still birth
Live birth
Letter size
Viability during lactation
Historical control data:
Historical control data on Reproduction Indices, Gestation Lengths, F1 Pup Viability, Hematology, Semen Analysis were included in the study reports.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
Two F0 females (#I420 and #1423) in the 50 mg/kg/day group had total litter loss and were euthanized on lactation day 0. Total litter loss has been observed in control animals at SLI and was not considered treatment related since it did not occur in the 167 and 500 mg/kg/day groups. In addition, three F0 females (#1379, #I404 and #1421) in the control group, one F0 female (#1400) in the 50 mg/kg/day group, one F0 female (#1434) in the 167 mg/kg/day group and two F0 females (#I353 and #1439) in the 500 mg/kg/day group had evidence of mating but failed to deliver and were euthanized 25 days after evidence of mating was detected. All other F0 females in the control, 50, 167 and 500 mg/kg/day groups survived to scheduled euthanasia.
Prior to death on gestation day 22, clinical signs for F0 female #I347 in the 50 mg/kg/day group included ocular discharge, dark material around the eyes and nose, reddish colored urine stain, fecal stain and piloerection. The death of this animal was not considered to be test article related since no mortality occurred for F0 females in the 167 and 500 mg/kg/day groups. However, a definitive cause of death could not be determined.
The most remarkable clinical signs observed for surviving F0 males included a low incidence of salivation prior to dosing and a dose-related increase in post-dose salivation in the 167 and 500 mg/kg/day groups.
Clinical signs were observed for surviving F0 females in the 50, 167 and 500 mg/kg/day groups. The clinical signs included a low incidence of reddish vaginal discharge, swelling (mammary glands), dark material around the eyes and dark material around the nose in the 50, 167 and 500 mg/kg/day groups; a low incidence of salivation prior to dosing, an increased incidence of urine stain and a dose-related increase in post-dose salivation in the 167 and 500 mg/kg/day groups; and a low incidence of ocular discharge in the 500 mg/kg/day group. A low incidence of clinical signs including soft stools and scab(s) was also observed for F0 females in each study group, including controls.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
There were no toxicologically meaningful differences in mean body weights or body weight change for F0 females in the 50, 167 and 500 mg/kg/day groups prior to mating or during gestation or lactation. With the exception of statistically higher mean body weight change for F0 females in the 500 mg/kg/day group during study days 0-7, mean body weights and body weight change of F0 females in the 50, 167 and 500 mg/kg/day groups were comparable to controls throughout the study.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
The mean precoital interval (number of days until mating) was 3.6 days in the control group, 3.9 days in the 50 mg/kg/day group, 2.6 days in the 167 mg/kg/day group and 3.2 days in the 500 mg/kg/day group.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
The mean precoital interval (number of days until mating) was 3.6 days in the control group, 3.9 days in the 50 mg/kg/day group, 2.6 days in the 167 mg/kg/day group and 3.2 days in the 500 mg/kg/day group.
The mean gestation length was 21.9 days in the control group, 22.1 days in the 50 mg/kg/day group, 21.9 days in the 167 mg/kg/day group and 21.8 days in the 500 mg/kg/day group.
A total of 25 females in the control group, 26 females in the 50 mg/kg/day group, 27 females in the 167 mg/kg/day group and 26 females in the 500 mg/kg/day group completed delivery. Two F0 females in the 50 mg/kg/day group delivered all stillborn pups (total litter loss). Therefore, the number of F0 females with liveborn pups was 25, 24, 27 and 26 in the control, 50, 167 and 500 mg/kg/day groups, respectively.
The total and mean number of F1 pups delivered was comparable between the control, 50, 167 and 500 mg/kg/day groups. However, the number of F1 liveborn pups in the 50 mg/kg/day group was statistically lower than controls. The lower number of liveborn pups in the 50 mg/kg/day group was attributed to a statistically higher number of stillborn pups, primarily from two females in this group with all stillborn pups (total litter loss).
The number of live pups per litter was comparable between the control, 50, 167 and 500 mg/kg/day groups throughout lactation. The sex ratio (male pups to total pups) was comparable between the control, 50, 167 and 500 mg/kg/day groups on lactation days 0 and 21.

ORGAN WEIGHTS (PARENTAL ANIMALS)
Absolute and relative (to final body weight) liver weights of F0 females in the 500 mg/kg/day group were statistically higher than controls at study termination. These differences were not considered to be toxicologically meaningful since they did not correlate with any toxicologically significant test article-related microscopic changes in the liver.
No statistically significant or toxicologically meaningful differences in absolute or relative weights were noted in the F0 female reproductive or accessory organs.

GROSS PATHOLOGY (PARENTAL ANIMALS)
Gross necropsy findings for F0 female #I 347 in the 50 mg/kg/day group found dead on gestation day 22 included mottled lungs (dark red and red), white foam the entire length of the trachea, multiple black foci on the glandular stomach mucosa and yellow mucoid material in the small intestine and colon. A definitive cause of death could not be determined.
Gross necropsy findings for F0 female #I353 in the 500 mg/kg/day group that failed to deliver and was euthanized 25 days after evidence of mating was detected included dark brownish-red fluid in the uterus and vagina and one small placenta and two nonviable pups in the vagina. No remarkable findings were observed at necropsy for the other F0 females that failed to deliver and were euthanized post breeding day 25. Gross necropsy findings for two F0 females in the 50 mg/kg/day group that were euthanized due to total litter loss (#I420 and #1423) included fetal tissue mixed with ingesta in the stomach, red mucoid material in the vagina and two retained pups each (one in each horn of each female).
No remarkable gross necropsy findings were noted for F0 males or females in the control, 50, 167 and 500 mg/kg/day groups that survived to scheduled euthanasia at study termination.

HISTOPATHOLOGY (PARENTAL ANIMALS)
No toxicologically meaningful microscopic lesions were observed in any of the tissues/organs examined from this study.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Effect level:
167 mg/kg bw (total dose)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
167 mg/kg bw (total dose)
Based on:
test mat.
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:not examined

Details on embryotoxic / teratogenic effects:
Embryotoxic and teratogenic effects: no examinedation of caesarean litter data was performed in this study. However, offspring development was assessed by the following post natal examinations:
VIABILITY (OFFSPRING)
A total of 25 females in the control group, 26 females in the 50 mg/kg/day group, 27 females in the 167 mg/kg/day group and 26 females in the 500 mg/kg/day group completed delivery. Two F0 females in the 50 mg/kg/day group delivered all stillborn pups (total litter loss). Therefore, the number of F0 females with liveborn pups was 25, 24, 27 and 26 in the control, 50, 167 and 500 mg/kg/day groups, respectively.
The total and mean number of F1 pups delivered was comparable between the control, 50, 167 and 500 mg/kg/day groups. However, the number of F1 liveborn pups in the 50 mg/kg/day group was statistically lower than controls. The lower number of liveborn pups in the 50 mg/kg/day group was attributed to a statistically higher number of stillborn pups, primarily from two females in this group with all stillborn pups (total litter loss).
The number of live pups per litter was comparable between the control, 50, 167 and 500 mg/kg/day groups throughout lactation. The sex ratio (male pups to total pups) was comparable between the control, 50, 167 and 500 mg/kg/day groups on lactation days 0 and 21.

CLINICAL SIGNS (OFFSPRING)
With the possible exception of a slight increase in the incidence of pups cool to the touch in the 500 mg/kg/day group (i.e., 19 times in 4 pups in the 500 mg/kg/day group vs. 1 time in 1 pup in the control group), there were no remarkable F1 pup observations during lactation. Findings were randomly distributed throughout the groups and included hairloss, scabs and subcutaneous hemorrhages.

BODY WEIGHT (OFFSPRING)
Mean F1 pup weights per litter for the 50, 167 and 500 mg/kg/day groups were not statistically different from the controls on lactation days 1, 4 (pre- and postculling) and 7. Even up to lactation day 21, the mean pup weight for the 50 mg/kg/day group remained comparable to the control. At the time of culling and up to lactation day 7, pup weight for the 167 and 500 mg/kg/day groups were slightly lower than the control (ranging from 5.2-6.7% lower). Mean F1 pup weight in the 167 mg/kg/day group (33.7 ± 4.20 grams) was statistically lower than controls (35.7 ± 2.1 1 grams) on lactation day 14 and mean F1 pup weight in the 500 mg/kg/day group was statistically lower than controls on lactation day 14 (32.6 ± 2.32 grams in the 500 mg/kg/day group vs. 35.7 ± 2.11 grams in controls) and lactation day 21 (51.9 ± 3.81 grams in the 500 mg/kg/day group vs. 56.0 ± 3.85 grams in the control group). However, the mean F1 pups weights in the 167 and 500 mg/kg/day groups remained within the range of SLI historical control data (i.e., 31.2-38.0 grams on lactation day 14 and 49.8-60.2 grams on lactation day 21).

SEXUAL MATURATION (OFFSPRING)
No data

ORGAN WEIGHTS (OFFSPRING)
No data

GROSS PATHOLOGY (OFFSPRING)
No remarkable findings were noted at necropsy for F1 pups stillborn, found dead, culled on day 4 or euthanized on lactation day 21. There were no test article-related malformations or developmental variations observed in the F1 pups.

HISTOPATHOLOGY (OFFSPRING)
No data

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
167 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
fetal/pup body weight changes

Fetal abnormalities

Key result
Abnormalities:
no effects observed

Overall developmental toxicity

Key result
Developmental effects observed:
no

Any other information on results incl. tables

Histopathologic Findings

No toxicologically significant test article-related lesions were noted in the tissues examined.

Although kidney observations are listed separately under left and right kidneys, findings are presented as combined data for both kidneys. Minimal to mild hyaline droplet accumulation was present in 27 Group 4 males (96%). Other prominent renal changes including tubular epithelial degeneration/regeneration, tubular dilatation and inflammation were noted in males and females from Groups 1and 4. The incidences of these lesions varied by sex and dose group.

A nephroblastoma was noted in the left kidney of a Group 4 female (1353). The mass was approximately 5 x 8 mm, was well-delineated and occupied the cortical and medullary space at one pole of the kidney, well within the limits of the normal structure.

A dermal mass observed grossly in a Group 4 female (1403) was diagnosed as a mammary carcinoma.

SUMMARY

The renal lesions noted in the control and treated animals represent changes associated with hyaline droplet nephropathy (HDN) and/or chronic progressive nephropathy (CPN).

Hyaline droplets, as noted in the Group 4 males, can be induced by a variety of chemical compounds in certain strains of male rats and is frequently associated with the accumulation of a2m-globulin within the hyaline droplets. A progressive nephropathy accompanies hyaline droplet accumulation, highlighted early on by tubular epithelial degeneration/regeneration and tubular dilatation with granular casts. Although histological findings in this study are consistent with a2m -globulin, specific identification of a,,-globulin has not been done. Definitive diagnosis of a2m - globulin is academic in that its presence and the associated nephropathy (HDN) In male rats is not considered toxicologically significant for humans according to Environmental Protection Agency Risk Assessment Forum (Baetck K.P, et al 1991).

The renal tubular degeneration/regeneration, tubular dilatation and inflammation, as observed in control animals and Group 4 females, are consistent with the constellation of lesions representing early signs of Chronic Progressive Nephropathy (CPN), a spontaneous, progressive, age-related entity. CPN is most common and prominent in male rats, as noted in this study.

The increased incidence of tubular changes and inflammation in Group 4 males is interpreted and the dual expression of both HDN and CPN.

Nephroblastomas occur rarely in young adult rats, with an incidence of 0 -0.4% (Mesfin, G.M. 1999). This was interpreted as an incidental finding and not considered treatment related.

The most common spontaneous neoplasm in aged Sprague-Dawley females are of mammary gland origin. Incidence of mammary carcinomas is 8.8% according to Chandra, et. al.(Chandra M, 1992). This lesion was interpreted as spontaneous and was not considered test article related.

Other changes noted were considered to be normal findings in rats of this age and strain.

 

Applicant's summary and conclusion

Conclusions:
Based on the results of this study, a dosage level of 167 mg/kg/day was considered to be the NOAEL for parental (F0) toxicity. There were no indications of impaired fertility or other reproductive effects in the F0 males or females at dosage levels up to 500 mg/kg/day. A dosage level of 167 mg/kg/day was considered the NOAEL for developmental effects, since the decreased pup weights noted for the 167 and 500 mg/kg/day group pups during the latter half of lactation were within the range of historical control data.
Executive summary:

Introduction

This study was conducted to evaluate the potential effects of oral administration of HPV-1 (CAS #67124-09-8) on the integrity and performance of the reproductive system in male and female rats, including gonadal function, mating behavior, conception, gestation, parturition, lactation and weaning. This study was also conducted to provide preliminary information concerning the effects of HPV-1 (CAS #67124-09-8) on neonatal moribundity, mortality and postnatal developmental toxicity. The study consisted of a vehicle control and three treatment groups, with 28 males and 28 females in each group. HPV-1 (CAS #67124-09-8) was dissolved in corn oil and administered at dosage levels of 50, 167 and 500 mg/kg/day, by once daily oral gavage, toF0parental animals. All doses were given at a constant volume of 2.5 mL/kg. Control animals were administered corn oil under the same experimental conditions at an equivalent dose volume.F0males were treated for a minimum of 70 days prior to mating and until completion of parturition. F0 females were treated for a minimum of 14 days prior to mating through lactation day 20.

Both F0 parental animals and F1 offspring were closely examined for indications of toxicity. Experimental endpoints for F0 animals included clinical observations, body weights, food consumption, mating, parturition, lactation, hematology determinations and offspring growth and viability. All F0 and F1 animals were subjected to a gross necropsy examination at the time of death or terminal euthanasia.

Results

Oral administration of 1-(tert-dodecylthio)propan-2-ol to F0 male rats for a minimum of 70 days prior to mating and until completion of parturition produced clinical signs including salivation prior to dosing and a dose-related increase in post-dose salivation in the 167 and 500 mg/kg/day groups and lower mean body weights (reduced approximately 5-7% compared to controls) in the 500 mg/kg/ay group.

Mean food consumption of F0 males in the 50, 167 and 500 mg/kg/day groups was comparable to controls throughout the study and there were no toxicologically meaningful differences between the control, 50, 167 and 500 mg/kg/day groups with respect to the F0 male mating and fertility indices, the F0 male hematology parameters evaluated, F0 male gross necropsy observations, F0 male absolute or relative organ weights or the F0 male sperm parameters evaluated. In addition, no toxicologically meaningful microscopic lesions were observed in any of the F0 male tissues/organs examined from this study.

Oral administration of 1-(tert-dodecylthio)propan-2-ol to F0 female rats for a minimum of 14 days prior to mating and throughout lactation produced clinical signs for F0 females in the 50, 167 and 500 mg/kg/day groups. The clinical signs included a low incidence of reddish vaginal discharge, swelling (mammary glands), dark material around the eyes and dark material around the nose in the 50, 167 and 500 mg/kg/day groups; a low incidence of salivation prior to dosing, an increased incidence of urine stain and a dose-related increase in post-dose salivation in the 167 and 500 mg/kg/day groups; and a low incidence of ocular discharge in the 500 mg/kg/day group.

There were no toxicologically meaningful differences between the control, 50, 167 and 500 mg/kg/day groups with respect to F0 female mean body weights, body weight change, food consumption, mating and fertility indices, precoital intervals, gestation lengths or the hematology parameters evaluated. Gross necropsy findings for one F0 female in the 500 mg/kg/day group euthanized on post breeding day 25 included dark brownish-red fluid in the uterus and vagina and one small placenta and two nonviable pups in the vagina. No other remarkable findings were noted in the F0 females at necropsy and no toxicologically meaningful microscopic lesions were observed in any of the F0 female tissues/organs examined from this study.

No toxicologically meaningful differences were noted in F1 pup viability during lactation. Mean F1 pup weights were statistically lower than controls in the 167 mg/kg/day group on lactation day 14 and in the 500 mg/kg/day group on lactation days 14 and 21; however, mean F1 pup weights in these groups remained within the range of testing laboratory historical control data. With the possible exception of a slight increase in the incidence of pups cool to the touch, no remarkable F1 pup observations were noted during lactation. No remarkable findings were noted at necropsy for F1 pups stillborn, found dead, culled on day 4 or euthanized on lactation day 21.

Conclusions

According to the study director, a dosage level of 50 mg/kg/day was considered to be the NOAEL for parental (F0) toxicity. There were no indications of impaired fertility or other reproductive effects in the F0 males or females at dosage levels up to 500 mg/kg/day. A dosage level of 50 mg/kg/ay was considered the NOAEL for developmental effects, as a result of decreased pup weights noted for the 167 and 500 mg/kg/day group pups during the latter half of lactation.

The study results were re-examined and new NOAELs for for parental and developmental toxicity were determined. Comparing to 50 mg/kg/day, the additional clinical observations included salivation and urine stain in females treated with 167 mg/kg/day test substance. Because 1) salivation was considered to be a reflex to administration of the test substance, 2) urine staining did not correlate with urination pattern or kidney gross necropsy, and 3) the severity of the effects is limited. The doses level of 167 mg/kg/days was determined to be the NOAEL for parental toxicity.

For F1 generation, the decreased mean pup weight that was observed at 167 mg/kg/day and 500 mg/kg/day were within historical range of the test laboratory. However, because the decrease in pup weights at the high dose was highly significant, the LOAEL for fetotoxicity was determined to be 500 mg/kg/day. The NOAEL was determined to be 167 mg/kg/day because 1) the decrease in pup weights were within the historical control range and were not highly significant and 2) the pups gained weight at comparable rate comparing to the controls.