Registration Dossier

Administrative data

Endpoint:
two-generation reproductive toxicity
Remarks:
based on test type
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-compliance study report, availables as unpublished report, no restrictions, adequate for assessment.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1991
Report Date:
1991

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)
Principles of method if other than guideline:
Other: Monsanto Agricultural Company, Environmental Health Laboratory method
Test conducted in general conformance with the GLP Standards with the following exceptions:
- Charaterization of the test material and analytical reference standard may not have been performed in accordance with the Standard/Principles.
- Determination of the stability of the analytical reference standard may not have been performed in accordance with the Standard/Principles.
- A reserve sample of the analytical reference standard was not retained.
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Physical state: pale, yellow liquid
- Purity: Not applicable (Complex mixture of terphenyl and quaterphenyl isomers)

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratory, Portage, MI
- Age at study initiation: (FØ) Approximately 5 wks
- Weight at study initiation: (FØ) Males: 161.6-225.5 g; Females: 122.2-185.6 g
- Housing for premating and gestation day 0 through 13: Individual. During mating, females were housed with the male in the male's cage.
- Housing from day 14 of gestation through lactation: Females were housed in double wide cages.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: fifteen days

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency): Approximately weekly
- Method of mixing: Neat test material was blended into the basal diet using a high speed mixer.
- Type of food: Purina Mills Certified RODENT CHOW N° 5002
Details on mating procedure:
- M/F ratio per cage: One male and one female (non-littermates) in male's cage.
- Length of cohabitation: 7 days, unless copulatory evidence found sooner. If no evidence of mating was, observed during the initial 7 day cohousing period, the female was co-housed with a male having recorded copulatory activity for an additional 7 days (maximum), or until copulatory evidence was observed. Mating procedures for the F1a adults were the same as for FØ adults except as modified to exclude sibling matings.
- Proof of pregnancy: pulatory plug, or vaginal smear if unable to determine cage from which plug had fallen.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
-Test Material stability: Gas Chromatography with a flame ionization detector; assays against an analytical standard both before and after the study.
- Diet mixture stability: Analysis of 30 and 1000 ppm level samples kept frozen (closed container, 35 days) or at ambient temperature (open container, 7 and 14 days).
- Homogeneity of diet mixtures: Analyses of duplicate samples from top, middle and bottom of mixer.
- Dietary level verification: Extraction of Terphenyl, hydrogenated with hexane; analysis by gas chromatography with a flame ionization detector; all dietary levels the first four weeks; at least one level per week thereafter.
Duration of treatment / exposure:
F0: until 21 days postpartum (F1A litter)
F1: until 21 days postpartum (F2A litter)
F2A: until 21 days postpartum
Duration of test: 275 days
Frequency of treatment:
7 days/week
Details on study schedule:
- Gestation/Lactation Day Designations: Gestation day 0=day on which copulatory evidence found; lactation day 0=day on which delivery of pups was complete, with subsequent days numbered consecutively.
Doses / concentrationsopen allclose all
Dose / conc.:
0 ppm (nominal)
Remarks:
nominal in diet
Dose / conc.:
30 ppm (nominal)
Remarks:
nominal in diet
Dose / conc.:
100 ppm (nominal)
Remarks:
nominal in diet
Dose / conc.:
300 ppm (nominal)
Remarks:
nominal in diet
Dose / conc.:
1 000 ppm (nominal)
Remarks:
nominal in diet
Remarks:
Doses / Concentrations: 1.8, 6.1, 18.5 and 62.0 for males (for the FØ adults)
Basis: analytical conc.
Remarks:
Doses / Concentrations: 2.4, 8.1, 24.3 and 80.6 for females (for the F1a adults)
Basis: analytical conc.
Remarks:
Doses / Concentrations: 1.9, 6.1, 18.2 and 63.1 for males (for the F1a adults)
Basis: analytical conc.
Remarks:
Doses / Concentrations: 2.5, 8.3, 24.4 and 81.2 for females (for the FØ adults)
Basis: analytical conc.
No. of animals per sex per dose:
30 adult rats/sex/group (FØ study start and F1a selected for mating)
Control animals:
yes
Positive control:
None

Examinations

Parental animals: Observations and examinations:
MORTALITY AND MORIBUNDITY: Yes
- Time schedule: Twice daily (am and pm)

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Adult - Once weekly; Offspring - On days of body weight measurement.

BODY WEIGHT: Yes
- Time schedule for examinations: FØ and F1a adult males - Once weekly; FØ and F1a adult females - Weekly until copulation confirmed, then on days 0, 7, 14, 21 of gestation and lactation.

FOOD CONSUMPTION:
- Time schedule for examinations: FØ and F1a adult males - Weekly except during mating; FØ and F1a adult females - Weekly until mating. After copulation was confirmed, maternal food consumption was measured on days 0-7, 7-14 and 14-21 of gestation. Food consumption was not determined for females with no evidence of copulation during cohousing with the male.
Sperm parameters (parental animals):
Parameters examined in FØ and F1a male parental generations:
sperm count in epididymides
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 of lactation: yes
- If yes, maximum of 8 pups/litter; excess pups were killed and discarded.

PARAMETERS EXAMINED
The following parameters were examined in F1a / F2a offspring:
Litter size, mortality, pup weight, survival (%)
Postmortem examinations (parental animals):
UNSCHEDULED NECROPSIES:
Those which died or were sacrificed in extremis were given a gross necropsy and selected tissues were saved.

SACRIFICE
- Male animals: All surviving animals after completion of mating phase.
- Maternal animals: Adult females which produced a litter - On or soon after day 21 of lactation; Nonpregnant adult females - At least 5 days after last expected parturition date

GROSS NECROPSY
- Gross necropsy: External and internal cavities were opened, and organs were examined in place, then removed. Hollow organs were opened and examined.

HISTOPATHOLOGY / ORGAN WEIGHTS
Organs Retained (when present): FØ and F1a adults - Coagulating gland, liver, ovaries, pituitary, prostate, seminal vesicle, skin/mammary gland, testes, epididymides, uterus, vagina and gross lesions;
Tissues Examined: Control and highest dietary levels - All retained tissues from all FØ and F1a adult animals and' all retained tissues from unscheduled adult deaths.
Organs Weighed: None
Postmortem examinations (offspring):
UNSCHEDULED NECROPSIES:
Pups found dead and culled pups had a gross necropsy; however, no tissues were saved and no organs were weighed.

SACRIFICE
- The F1 offspring not selected as parental animals and all F2 offspring were sacrificed on or soon after day 21 of lactation.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:

GROSS NECROPSY
- Gross necropsy: External and internal cavities were opened, and organs were examined in place, then removed. Hollow organs were opened and examined.

HISTOPATHOLOGY / ORGAN WEIGTHS
None
Statistics:
The following statistical procedures were used to detect statistically significant differences between treated animals and their respective controls:
- Dunnett's Multiple Comparison Test (two-tailed)
- EHL decision tree analysis
- Uncorrected Chi-Square Test
- Fisher's Exact Test

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
no effects observed
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 examined
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
not specified

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
There were two adult deaths. A control level FØ female died on day 105, shortly after delivering a litter, with a retained fetus. The other was a T-1 level (30 ppm) F1a female which died on day 132 (29 days after her birth). Neither of these deaths were considered related to treatment.
There were no clinical signs considered, related to administration of the test material.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
The only apparent adverse effects on body weight or weight gain in adult animals occurred during the last three weeks in T-4 level (1000 ppm) males of the FØ generation and in T-4 level F1a dams during gestation. There were no statistically significant differences in group mean body weights at lower dietary levels in males, or at any dietary level in adult females prior to mating. Maternal weight gain of T-4 level F1a dams was slightly reduced (approximately 93% of the control value) during the gestation day 0 to 21 period (regained during lactation), and was increased during the lactation day 0-7 period (1546% of controls). These dams also did not lose as much body weight as controls during the lactation day 14-21 period (lost approximately 12 grams as compared to approximately 23 grams for controls); however, this was not considered an adverse effect.

The only statistically significant decreases in food consumption occurred during the first week of gestation (gm/kg/day) and second week of gestation (gm/day), when T-4 level (1000 ppm) females of the FØ generation consumed slightly less food than did their controls. Food consumption by treated males during the study was similar to, or greater than, that of controls (except for T-2 level (100 ppm) males during week 17).

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
There were no findings in adult animals considered related to treatment. Decreased/absent sperm in the epididymis (unilateral) - was observed in one T-4 level FØ male, and a control male of the F1a generation, and was, therefore, not considered treatment-related.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
Mating, fertility and gestation: There were no statistically significant differences in any of the measured parameters considered related to treatment in either the FØ or F1a generations.

GROSS PATHOLOGY (PARENTAL ANIMALS)
There were no gross necropsy findings in adult animals of either generation considered related to treatment.

HISTOPATHOLOGY (PARENTAL ANIMALS)
Interstitial monocyte/lymphocyte infiltration of the prostate gland (a common lesion in older animals) was somewhat more frequently seen (not statistically significant) in T-4 level males (1000 ppm) of both generations than their respective controls, but was not considered an adverse effect attributable to administration of the test material.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Effect level:
1 000 ppm (nominal)
Sex:
male/female
Basis for effect level:
other: reproductive effects
Dose descriptor:
NOAEL
Effect level:
300 ppm
Sex:
male/female
Basis for effect level:
other: decrease in mean body weights

Results: F1 generation

General toxicity (F1)

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

Details on results (F1)

VIABILITY (OFFSPRING)
Litter size (live pups), dead pups/litter (day 0), and mean pup survival were not adversely affected when compared to controls.

CLINICAL SIGNS (OFFSPRING)
There were no clinical observations in offspring which were considered related to administration of the test chemical.

BODY WEIGHT (OFFSPRING)
There were no body weight effects in offspring considered treatment- related.

GROSS PATHOLOGY (OFFSPRING)
In offspring, distended/dilated stomachs were observed in four males and one female F1a weanlings at the T-4 level (1000 ppm). The cause of this abnormality was not definitively determined, but may have been a result of overeating; this change was not attributed to treatment.

Effect levels (F1)

open allclose all
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
1 000 ppm (nominal)
Sex:
male/female
Basis for effect level:
other: reproductive effects
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
300 ppm
Sex:
male/female
Basis for effect level:
other: decrease in mean body weights

Overall reproductive toxicity

Reproductive effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Non-reproductive effects attributed to treatment in adult rats were limited to a minor decrease in group mean body weights of high level F0 males near the end of the study, and slightly decreased body weights of F1a dams during gestation.
There were no adverse reproductive effects in any of the measured parameter/indices in adult rats or their offspring.
NOAEL reproductive effects = 1000 ppm d (62-81 mg/kg bw/day), and NOAL subchronic toxicity = 300 ppm (18-24 mg/kg bw/day).
Executive summary:

Male and female Sprague-Dawley rats (30 adults/sex/dose) were continuously fed hydrogenated terphenyl through two generations at target levels of 0,30, 100, 300 and 1000 ppm in their diet. Analyses to verify the stability of the test material both neat and when mixed with the diet, the diet homogeneity and concentrations of the test material in the diet were all performed with satisfactory results. Overall study averages for consumption of test material (mg hydrogenated terphenyl/kilogram body weight/day), based on the target concentrations, were as follows: 1.8, 6.1, 18.5 and 62.0 for males and 2.5, 8.3,24.4 and 81.2 for females (for the F0 adults) and 1.9,6.1, 18.2 and 63.1 for males and 2.4, 8.1,24.3 and 80.6 for females (for the Fla adults), at the lowest to highest levels, respectively. Non-reproductive effects attributed to treatment in adult rats were limited to a minor decrease in group mean body weights of high level F0 males near the end of the study, and slightly decreased body weights of F1a dams during gestation. There were no adverse reproductive effects in any of the measured parameter/indices in adult rats or their offspring. On the basis of the above findings, the 1000 ppm dietary level was considered the no-observed-adverse-effect-level (NOAEL) for reproductive effects (corresponding with 62-81 mg/kg bw/day), and the 300 ppm dietary level was considered the NOAEL for subchronic toxicity in this study (corresponding with 18-24 mg/kg bw/day).