Registration Dossier

Toxicological information

Toxicity to reproduction

Currently viewing:

Administrative data

Endpoint:
two-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: guideline study
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1989
Report Date:
1989

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: TSCA Health Effects Test Guideline for Specific Organ/Tissue Toxicity - Reproduction/Fertility Effects (EPA 1983)
Deviations:
no
Principles of method if other than guideline:
Reproductive toxicity of p-cresol was examined in a two-generation toxicity study for specific organ/tissue toxicity - Reproduction/Fertility effects.
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Test substance: p-cresol, 98.93% pure

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston NY
- Age at study initiation: 6 weeks (P)
- Weight at study initiation: (P) Males: 189-191 g; Females: 141-142 g;

- Housing:
initially 2 /same sex during acclimatisationperiod; and then singly except for the cohabitation and lactation periods
- Diet ad libitum
- Water ad libitum
- Acclimation period: 2 weeks


ENVIRONMENTAL CONDITIONS
- Temperature (°F): 68-74
- Humidity (%): 40-60
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Dosing formulations were prepared by weighing the amount of test chemical into a volumetric flask and diluting to volume with certified corn oil. The resulting solutions were mixed by repeated inversions and stored at room temperature
Details on mating procedure:
- M/F ratio per cage: 1/1
-Length of cohabitation: 21 days
- Proof of pregnancy: vaginal plug referred to as day 0 of pregnancy
- After 7 days of unsuccessful pairing replacement of first male by another male with proven fertility.
- Further matings after two unsuccessful attempts: no
- After successful mating each pregnant female was caged singly
- Any other deviations from standard protocol: no
- M/F ratio per cage:
- Length of cohabitation:
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility.
- Further matings after two unsuccessful attempts: [no / yes (explain)]
- After successful mating each pregnant female was caged (how):
- Any other deviations from standard protocol:
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
A standard stock solution was prepared (1 mg/ml propanol), which was used to prepare standards ranging form 10 to 100 ng/µl. With these solutions standard curve was generated using HPLC. Dosing formulation concentrations were veryfied by preparing aliquots which were injectd on HPLC column. The measured concentration of each dosing solution was then calculated from the equitation for the standared curve developed by linear regression.
Duration of treatment / exposure:
Exposure period: 27 weeks
Premating exposure period (males): 10 weeks
Premating exposure period (females): 10 weeks
Duration of test: 29 weeks
Frequency of treatment:
P- and F1-generation: once per day, 5 days per week
F1 generation producing F2: once per daym 7 days per week
Details on study schedule:
Number of generation studies: 2
At day 28-40 post partum F1 animals were selected to be parents of the F2-generation and were gavaged with their respective formulations for at least 11 weeks on 5 days per week
The F1 animals were approximately 15-17 weeks of age at the initiation of the mating period.
they were dosed from that time point 7 days/week . Mating procedure was performed as done with the P-generation
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 30, 175, 450 mg/kg bw
Basis:

No. of animals per sex per dose:
25 rats/sex/dose
Control animals:
yes, concurrent vehicle
Details on study design:
no further data
Positive control:
no data

Examinations

Parental animals: Observations and examinations:
Mortality:
twice daily
General condition:
daily throughout the course of the study including skin and fur, eyes and mucous membranes, respiratory symptoms, circulatory system, autonomic and central nervous system, somatomotor activity, behavior pattern
Body weight dertermination
male, female: initially and then weekly until mating
female during gestation: day 0, 7, 13, 20 post partum day 0, 4, 7, 14, 21
Food concumption:
measured weekly during pre-breed dosing period for P and F 1 generation;
all other phases of this study determination was made visually
Oestrous cyclicity (parental animals):
vaginal smears were examined to determine pregnancy
Sperm parameters (parental animals):
no data
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of .8 pups/litter (4/sex/litter as nearly as possible); excess pups were killed and discarded.


PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2 / F3] offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities,


GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities; possible cause of death wwas not determined for pups born or found dead.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals after the completion of the mating period
- Maternal animals: All surviving animals after the F1 and F2 litters have been weanded

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.


HISTOPATHOLOGY
Male and female adult rats of the highest doses groups and the controls
The tissues as indicated below were prepared for microscopic examination and weighted, respectively:
Pituitary, vagina, uterus, ovaries, testes, epididmides, seminal vesicles, prostate, and other tissues with gross lesions identified as being potentially treatment-related
A complete histopathological examination was conducted for any parental animal dying on test.
Postmortem examinations (offspring):
All pups dying during lactation are necropsied to investigate the cause of death.
At weaning, postnatal day 21, 1 female and 1 male from each F1 litter is selected on a random basis to become parents of the next generation.
The remaining offspring is examined for gross external abnormalities, euthanized and discarded
Statistics:
Levene's test, ANOVA, t-test corredted by bonferroni method, Kruskal-Wallis test, Mann-Whitney U-test, Fisher's exact test
Reproductive indices:
calculated for p- and F1 animals:
Mating index (%):
---for males: number of males impregnation females devided through the total number of malespaired multiplied by 100
---for females: number of females with copulation plugs devided through the number of females cohabited multiplied by 100

Fertility index(%):
---for males: number of males producing pregnant females dvided through number of maes impregnating females multiplied by 100
---for females: number of females with copulation plugs divided through the number of females cohabited multiplied by 100

Gestational Index (%):
number of females with live litteres devided through number of females pregnant multiplied by 100

Offspring viability indices:
calculated for F1 and F2 animals:
live birth index (%):
number of pups at birth devided through the total number of pups born multiplied by 100
4-day survival index (%):
number of pups surviving 4 days devided through the total number of live pups at birth multiplied by 100
7-day survival index(%):
number of pups surviving 7 days devided through the total number of live pups at birth multiplied by 100
14-day survival index(%):
number of pups surviving 14 days devided through the total number of live pups at birth multiplied by 100
21-day survival index(%):
number of pups surviving 21 days devided through the total number of live pups at birth multiplied by 100

Lactation omdex (%):
number of pups surviving 21 days through total number of live pups at 4 days multiplied by 100

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
not specified
Histopathological findings: non-neoplastic:
not examined
Other effects:
no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed

Details on results (P0)

see section " Remarks on results"

Effect levels (P0)

open allclose all
Dose descriptor:
other: NOAEL (general toxicity)
Effect level:
ca. 30 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: >= 175 mg/kg bw/d: clinical signs of toxicity including hypoactivity, ataxia, twitches, tremors, prostration, urine stains, audible respiration, and perioral wetness 450 mg/kg bw/d: increased mortality, and reduced body weight gain
Remarks on result:
other: Generation: P and F1 (migrated information)
Dose descriptor:
other: NOAEL (offspring)
Effect level:
ca. 175 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: based on toxic effects in the F2 litters of the high dose group animals and no clear evidence of toxic effects in F1 pups.
Remarks on result:
other: Generation: F1 and F2 (migrated information)
Dose descriptor:
other: NOAEL (fertility)
Effect level:
ca. 450 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: No reproductive parameters were affected in either of the two generations.
Remarks on result:
other: Generation: P and F1 (migrated information)

Results: F1 generation

General toxicity (F1)

Clinical signs:
not examined
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
no effects observed
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
not specified
Histopathological findings:
not specified

Details on results (F1)

see section: "Remarks on results"

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

Mortality:  8/28 males and 5/25 females at 450 mg/kg bw; 1/25 females at  30 mg/kg bw
Clinical signs of toxicity
occurred in F0 and F1 males and females at 450 mg/kg bw/day and included  hypoactivity, ataxia, twitches, tremors, prostration,  urine stains,  audible respiration, perinasal encrustation (not in F0 males), and  perioral wetness occurred at >= 175 mg/kg bw.
body weight:
F0 adult males, sign reduced (p<0.01) week1 to week 13 in the 450 mg/kg  bw group; 
F0 adult females:
sign. reduced week 1 (p<0.05) in the 450 mg/kg bw-group,
gestational weight gain not significantly different from control group,  lactational body weight sign. reduced (p<0.05) at d4 at  450 mg/kg bw group

F1 or F2: No reproductive parameters were affected in either of the two  generations (mating index of male and females, fertility index  of males  and females, gestational index. 

Still births in the F1 and F2 generations: 
in F1 pups increased at 175  mg/kg/day (7/13 of one dam), but not at 450 mg/kg bw/day (low, mid, high dose versus control: 4/290 born pups, 13/312 born pups with 7/13 on one dam, 6/193 born pups) in F2 pups increased at 30 and 450 mg/kg bw, but not  at 175 mg/kg/bw (low, mid, high dose versus control: 7/307 born pups, 4/265 born pups,9/163 born pups versus 0/318 born pups)

There was some variability in the number of stillborn in control groups  in F1 and F2 generation (2 versus 0). There was no clear  dose-dependent   effect in both generations (control/low/mid/high dose: F1 pups: 2/4/13/6;  F2 pups: 0/7/4/9). 

F1,F2: Pup survival indices in both generations were not
affected by treatment (4-day survival index, 7-day survival index, 14-day  surval index 21-day survival index and lactation index),  except live  birth indices in F2 (but not F1) which were reduced at 30 and 450 mg/kg  bw, but not at 175 mg/kg/day. Without any  other effects especially in the  30 mg/kg bw-group it is unclear whether the effects on live birth indices  were substance related. 
ross lesions of parental males and females which died prior to scheduled  sacrifice included diffuse, focal or multifocal color changes  in the lung  and stained skin for males and lung congestion and congestion in the  nasal turbinates and erythrocytes on the skin surface  for females.  
There were no treatment related histologic lesions observed in the  examination of organs from parental F0 and F1 adults which survived to  scheduled sacrifice.

Applicant's summary and conclusion

Executive summary:

Reproductive toxicity was examined in a two-generation toxicity study according to TSCA Health Effects Test Guideline for specific organ/tissue toxicity - Reproduction/Fertility effects. Sprague-Dawley rats were given daily 0, 30, 175, or 450 mg/kg bw/day p-cresol in corn oil by gavage. No effects on fertility were detected despite overt general toxicity including increased mortality and reduced body weight gain at 450 mg/kg bw and at >= 175 mg/kg bw/day hypoactivity ataxia, twitches, tremors, prostration, urine stains, audible respiration and perioral wetness. Thus, the NOAEL(fertility) was 450 mg/kg bw/day and the NOAEL(general toxicity) was 30 mg/kg bw/day. The NOAEL(offspring) is 175 mg/kg bw/day due to toxic effects in the F2 litters of the high dose group animals and no clear evidence of toxic effects in F1 pups.