1-[2-(2-chloroethoxy)phenylsulfonyl]-3-(4-methoxy-6-methyl-1,3,5-triazin-2-yl)urea;triasulfuron (ISO)

Toxicity to reproduction

Administrative data

Endpoint:

two-generation reproductive toxicity

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

24 Sept 1985 to 27 Jun 1987

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1987

Report date:

1987

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)

Version / remarks:

1983

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPP 83-4 (Reproduction and Fertility Effects)

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: Proposed U.S. Federal Insecticide, Fungicide and Rodenticide Act Guidelines Draft

Version / remarks:

1982

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2-(2-chloroethoxy)-N-[(4-methoxy-6-methyl-1,3,5-triazin-2-yl)carbamoyl]benzene-1-sulfonamide

EC Number:

617-298-9

Cas Number:

82097-50-5

Molecular formula:

C14H16ClN5O5S

IUPAC Name:

2-(2-chloroethoxy)-N-[(4-methoxy-6-methyl-1,3,5-triazin-2-yl)carbamoyl]benzene-1-sulfonamide

Test animals

Species:

rat

Strain:

Sprague-Dawley

Remarks:

Crl:CD® (SD)BR; albino rats

Sex:

male/female

Details on test animals or test system and environmental conditions:

Test Animal Information
- Age at study initiation: (P) 7 wks; (F1) 21 to 34 days
- Housing: Individual, hanging, stainless steel, wire-bottom cages were used during the acclimation period and all phases of the study, except during the mating, lactation, and gestation periods. During the mating trials, similarly designed galvanized steel cages were used. These cages, which were equipped with solid bottom stainless steel floorplates and nesting material were also used during gestation and lactation.
- Diet: Ad libitum
- Water: filtered tap water. Ad libitum
- Acclimation period: approximately 14 days

Environmental Conditions
- Temperature: 75 +/- 5°F
- Humidity: 50 +/-20%
- Air changes: well ventilated, air-conditioned room was used throughout the study
- Photoperiod: twelve hour light cycle

In-life Dates: From: 24 Sep 1985 To: 16 Jul 1986

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

DIET PREPARATION
- Rate of preparation of diet: Fresh diets were prepared weekly
- The test article was mixed with the basal diet using Hobart mixers to achieve concentrations (w/w) of either 10, 1000, or 5000 ppm of the test item.
- Storage temperature of food: Sufficient diet was offered to each animal to assure ad libitum feeding with the excess stored in sealed containers at room temperature until use or disposal.

Details on mating procedure:

- M/F ratio per cage: 1
- Length of cohabitation: 21 days or until mating was confirmed
- Proof of pregnancy: copulatory plug in the vagina or sperm-positive results of vaginal smears. Gestation day 0 was defined as the day breeding was confirmed
- After successful mating each pregnant female was caged: individually

Mating trials to produce the 'a' litters (F0 generation, F1a litter; F1 generation, F2a litter) were initiated following the pre-mating period exposure to the test diets. Mating trials to produce the F1b litters of the F0 generation were initiated 1 week after the last F1a litter was weaned. Procedures used for the F1a mating trials were followed, however, males were not paired with females they had been assigned to during the F1a mating trials and females that failed to conceive F1a litters were not rebred.

Analytical verification of doses or concentrations:

yes

Remarks:

Analyses of the test article concentrations in diet samples taken from each group were conducted once a month.

Details on analytical verification of doses or concentrations:

The results of the diet assays indicate that the test item was homogeneously distributed in the test diets and that concentrations were within acceptable limits of the target concentrations of 10, 1000, and 5000 ppm

Frequency of treatment:

Continuously

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

10 ppm

Remarks:

Group 1. Dietary equivalent to 0.6 and 0.8 mg/kg bw/day for males and females, respectively.

Doses / concentrations 2

Dose / conc.:

1 000 ppm

Remarks:

Group 2. Dietary equivalent to 64 and 73 mg/kg bw/day for males and females, respectively.

Doses / concentrations 3

Dose / conc.:

5 000 ppm

Remarks:

Group 3. Dietary equivalent to 321 and 380 mg/kg bw/day for males and females, respectively.

No. of animals per sex per dose:

F0, F1 parental animals: 30

Control animals:

yes, plain diet

Details on study design:

First (F0) Generation:
-Selection of F0 Generation Parental Animals
-Pre-mating period: 12 weeks
-F1a mating trials: 3 weeks
-Gestation periods: 3 weeks from mating
-Lactation periods: 3 weeks from delivery
-F1a weanings

Second (F1) Generation from F1a weanings
- Selection of F1 Generation Parental Animals
-Pre-mating period: 14 weeks
-F2a mating trial: 3 weeks
-Gestation periods: 3 weeks from mating
-Lactation periods: 3 weeks from delivery
-F2a weanings

From F1a weaning:
-Rest period: 1 week
-F1b mating trials: 3 weeks
-Gestation periods: 3 weeks from mating
-Lactation periods: 3 weeks from delivery
-F1b weanings

Examinations

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: All animals were observed at least twice each day for mortality, moribundity, and overt signs of toxicity. Additionally, each animal was removed from its cage at least once each week and was thoroughly examined for the detection of physical change.

DETAILED CLINICAL OBSERVATIONS: Each female was observed daily during gestation. Conception was confirmed by the observation of a vascular membrane in the vagina and/or the detection of progeny by palpation. Gravid animals were supplied with nesting material on approximately day 15 of gestation. Starting on gestation day 20, pregnant females were observed twice daily for parturition. The females were allowed to deliver their litters, and daily observations of the females and young were conducted throughout lactation. Litters were weaned at 21 days of age.· Any unusual observations noted during the periods of gestation and lactation were recorded.

BODY WEIGHT: Yes
- Time schedule for examinations: All parental animals were weighed weekly during the pre-mating period (F0 generation, 12 weeks; Fl generation, 14 weeks). At the conclusion of the mating trials, males were then weighed at weekly intervals until their sacrifice. Bred females were weighed on gestation days 0, 7, 14, and 20 and lactation days 0, 4, 7, 14, and 21 (where appropriate). Those females which failed to conceive, deliver viable progeny, or retain progeny throughout the lactation period were weighed at weekly intervals along with the males, until their termination. Final body weights were obtained for each animal at termination.

FOOD CONSUMPTION AND COMPOUND INTAKE: yes
Food consumption determinations were conducted at weekly intervals for parental animals during the pre-mating period. Food consumption was measured for bred females on gestation days 0, 7, 14, and 20 and lactation days 7 and 14. These data, along with the corresponding body weight data, were used to calculate test article consumption as mg of test article per kg of body weight per day

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. If fewer than 4 of either sex survived to day 4, all survivors of that sex were retained along with the appropriate number of pups of the other sex to obtain a total of 8 progeny.

PARAMETERS EXAMINED
The following parameters were examined in the progeny:
sexes and numbers of pups delivered viable or stillborn, or found partially cannibalized, the numbers of pups surviving to lactation days 4, 7, 14, and 21 (weaning)

GROSS EXAMINATION OF DEAD PUPS:
yes, all pups found dead during the lactation period and pups sacrificed on lactation day 4 were subjected to a gross necropsy examination for the detection of gross malformations.

OTHER:
Each pup was also examined thoroughly for developmental anomalies at birth, at each body weight interval, and again at weaning
Body Weights: Individual pup weights and sexes were determined on lactation days 0, 4, 7, 14, and 21 (weaning) for all surviving progeny of each litter. However, pups were not identified within the litter with a unique identification number until weaning.

Postmortem examinations (parental animals):

SACRIFICE
All F0 parental males were sacrificed at 255 days of age having been fed their respective test/control diets for 206 days prior to sacrifice.
-The surviving F0 parental females were sacrificed at 261 days of age having been fed their test/control diets for 212 days prior to sacrifice. In addition, females that failed to conceive F1a progeny were sacrificed 25 days post completion of the mating trials and were 181 days of age after having been fed their test/control diets for 132 days prior to sacrifice.
-The F1 parental males were 173 to 186 days of age at final sacrifice. F1 parental females that conceived F2a litters were 174 to 187 days of age at sacrifice. The F1 females that did not conceive F2a litters were sacrificed at 167 to 172 days of age.

GROSS NECROPSY
F0 and F1 parental animals were necropsied as described below with tissues retained for histopathologic studies. At sacrifice the F0 and F1 parental animals were asphyxiated by carbon dioxide and then exsanguinated. The necropsy included an examination of the external body surface and all orifices; the cranial cavity, external and cut surfaces of the brain and spinal cord; the thoracic, abdominal and pelvic cavities and their viscera; cervical tissues and organs; and the carcass. The kidney(s) and the bladder were examined for the presence of calculi. If calculi were present in the bladder, the bladder was examined for signs of irritation and retained for possible future microscopic examination. All calculi were retained, dry, for possible future chemical analysis. The uteri of parental females were examined for implantation sites. Absolute organ weights were recorded for the testes (with epididymides) or ovaries from all parental animals. Corresponding organ to body weight ratios were calculated. Tissues and organs retained from the parental animals were preserved in 10% neutral buffered formalin. Tissues were retained as deemed necessary by the examining pathologist. All necropsies were conducted under the supervision of a qualified pathologist.
Tissues/organs that were retained are listed below: vagina, uterus, cervix, ovaries, testes (with epididymides), seminal vesicles, prostrate, coagulating gland, pituitary, kidneys, liver, All other tissues/organs that appeared abnormal.

HISTOPATHOLOGY / ORGAN WEIGHTS
Microscopic examinations were conducted upon the vagina, cervix, uterus, ovaries, testes (with epididymides), seminal vesicles, coagulating gland, prostate, and pituitary of all F0 and F1 generation parental animals. In addition, microscopic examinations were conducted upon any tissues which displayed grossly noted changes. Slides were prepared and stained with hematoxylin and eosin for all tissues examined microscopically. Slides were evaluated by a qualified pathologist.

Postmortem examinations (offspring):

SACRIFICE
Thirty males and 30 females were randomly selected from the population of the F1a progeny weaned from each treatment group to serve as F1 parental animals. Ten males and 10 females per treatment group were selected from the F1a, F1b and F2a litters post-weaning for gross necropsy procedures and tissues retained for possible histopathology. The remaining progeny were subjected to complete gross necropsy procedures. All F1a progeny selected as F1 parental animals and the F1a, F1b and F2a progeny selected for possible histopathologic study were ear-tagged for permanent identification with a tag displaying the assigned identification number.

GROSS NECROPSY
Ten male and 10 female F1a, F1b and F2a progeny per treatment group were necropsied with tissues retained for possible future histopathologic studies. All remaining weaned progeny, all progeny found dead during the lactatiori period, and progeny sacrificed on lactation day 4 were necropsied and any tissues that appeared abnormal were saved. At sacrifice the F1a progeny not selected as F1 parental animals, and the F1b and F2a progeny were asphyxiated by carbon dioxide and then exsanguinated. The necropsy included an examination of the external body surface and all orifices; the cranial cavity, external and cut surfaces of the brain and spinal cord; the thoracic, abdominal and pelvic cavities and their viscera; cervical tissues and organs; and the carcass. The kidney(s) and the bladder were examined for the presence of calculi. If calculi were present in the bladder, the bladder was examined for signs of irritation and retained for possible future microscopic examination. All calculi were retained, dry, for possible future chemical analysis. Corresponding organ to body weight ratios were calculated. Tissues and organs retained from 10 male and 10 female, F1a, F1b, and F2a progeny per group were preserved in 10% neutral buffered formalin. Necropsies conducted upon progeny found dead during the lactation period and progeny sacrificed on lactation day 4 included an examination of the external body surface; the thoracic, abdominal, and pelvic cavities for normal appearance of their viscera; and an examination of the brain and palate for the detection of developmental malformations. Tissues were retained as deemed necessary by the examining pathologist. All necropsies were conducted under the supervision of a qualified pathologist. Tissues/organs that were retained are listed below: vagina, uterus, cervix, ovaries, testes (with epididymides), seminal vesicles, prostrate, coagulating gland, pituitary, kidneys, liver, All other tissues/organs that appeared abnormal.


HISTOPATHOLOGY / ORGAN WEIGTHS
Microscopic examinations were conducted upon any tissues which displayed grossly noted changes. Slides were prepared and stained with hematoxylin and eosin for all tissues examined microscopically. Slides were evaluated by a qualified pathologist.

Statistics:

Parental body weights and food consumption data, progeny population and survival data, and progeny body weight data were analyzed using Analysis of Variance. Significant differences between the untreated control group and the treated groups were evaluated using an appropriate multiple comparison test (Tukey's or Scheffe's dependent upon 'N' values). Organ weight ratios were studied using Kruskal-Wallis analyses. Chi-Square analysis and Fisher's Exact Tests were performed where appropriate. All statistical analyses were interpreted using the untreated control group for comparison. Differences were considered significant at the p<0.05 and p<0.01 confidence levels.

Reproductive indices:

Mating Index = (Number of Copulations*)/(Number of Estrus Cycles**) X 100;
Fertility Index = (Number of Pregnancies - Number of Copulations) X 100;
Gestation Index = (Number of Parturitions - Number of Pregnancies) X 100;
Female Fertility Index = (Number of Pregnancies - Number of Females Mated) X 100;
Male Fertility Index = (Number of Sires - Number of Males Mated) X 100;


* Copulation is defined as the presence of a copulatory plug in
the vagina or sperm positive vaginal smears.
** Five days equal 1 estrus cycle.

Offspring viability indices:

Born Viable= (No. Pups Delivered Alive/Total No. Pups Delivered) x 100;
Born Dead= (No. Stillbirths/Total No. Pups Delivered)X 100;
Born and Cannibalized = (No. Pups Found Partially Cannibalized/Total No. Pups Delivered) x 100;
4 Day = (No. Pues Viable at Lactation Day 4/No. Pups Born Alive) x100;
7 Day = (No. Pups Viable at Lactation Day 7/No. Pups Retained at Lactation Day 4) x100;
14 Day = (No. Pues Viable at Lactation Day 14/No. Pups Retained at Lactation Day 4) x100;
21 Day = (No. Pups Viable at Lactation Day 21/No. Pups Retained at Lactation Day 4) x100

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

Clinical findings were observed at approximate equivalent frequency in the treated and control groups and were considered incidental and unrelated to treatment. Observations seen for F0 generation maternal animals were limited to vaginal discharges (amber/yellow) seen in 2 of the 10 ppm animals: AH1435 (gestation day 22, F1a litter) and AH1416 (lactation day 11, F1b litter).

Dermal irritation (if dermal study):

not examined

Mortality:

mortality observed, non-treatment-related

Description (incidence):

A F0 untreated control male died during the F1b mating trials; no deaths occurred among the treated animals.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Although F0 generation animals fed 5,000 ppm of the test item generally weighed slightly less (2 to 8%) than the untreated control animals throughout the pre- and post-mating periods, no statistically significant reductions were noted. Significant reductions (p<0.05) were noted in the pre-mating weight gain of the males and the total weight change of the females, however. The F0 dams fed 5,000 ppm of the test item weighed less than the untreated control dams during the F1a and F1b gestation and lactation periods. Maternal body weights of the 5,000 ppm dams on lactation day 4 of the F1a litter and gestation days 0, 7, 14, and 20 and lactation days 0, 4, and 7 of the F1b litter were significantly less than the untreated control weights.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Statistical analyses of the pre-mating food consumption revealed significant reductions for the 5,000 ppm males during 3 of the 13 pre-mating intervals of the F0 generation. No statistically significant differences were seen in the 10 and 1,000 ppm male food consumption that were considered treatment-related. During week 4 of the F1 generation pre-mating period, a statistically significant reduction in food consumption was noted for both the 10 and the 1,000 ppm males. This difference was not considered an effect of treatment because it was an isolated event with these males generally consuming the same amount or slightly more feed than the untreated control males at all other intervals during the pre-mating period. Sporadic statistical difference in feed intake were noted for the F0 generation treated females, however, these differences also did not reveal a dose response relationship and were not considered treatment-related. Analyses of the maternal food consumption revealed no statistically significant differences.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

No microscopic findings were seen which were considered treatment-related. Uterine hydrometra characterized by dilatation of the uterine horns was seen in all female groups but at a higher incidence in the T-III group. There was not a progressive dose response incidence of the change and in the non-gravid females the change was evident in only the control group. A variety of other diagnoses were made on both control and treated animals, however, all appeared to be unrelated to the test material and were changes expected in a population of experimental rats. No changes in the reproductive tract or other tissues examined that appear to be test article related.

Histopathological findings: neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

No microscopic findings were seen which were considered treatment-related. A neoplastic
diagnosis of the mammary gland was made. A mammary gland fibroadenoma was seen in a single T-III male and a single T-III female rat. This was clearly a benign change typically seen in this species and strain.

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

no effects observed

Description (incidence and severity):

Reproductive performance was not adversely affected in the treated animals; no statistically significant reductions were noted in the reproductive indices.

Reproductive function: sperm measures:

no effects observed

Description (incidence and severity):

Reproductive performance was not adversely affected in the treated animals; no statistically significant reductions were noted in the reproductive indices

Reproductive performance:

no effects observed

Description (incidence and severity):

Reproductive performance was not adversely affected in the treated animals; no statistically significant reductions were noted in the reproductive indices (see Table 1 and 2).

Effect levels (P0)

1

Dose descriptor:

NOAEL

Remarks:

Systemic toxicity

Effect level:

1 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

food consumption and compound intake

Remarks on result:

other: Dietary equivalent to 64 and 73 mg/kg bw/day for males and females, respectively.

2

Dose descriptor:

NOAEL

Remarks:

Toxicity to fertility

Effect level:

> 5 000 ppm

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Remarks:

Dietary equivalent to 321 and 380 mg/kg bw/day for males and females, respectively.

Target system / organ toxicity (P0)

Critical effects observed:

no

Results: P1 (second parental generation)

General toxicity (P1)

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

During the F1 generation (F2a litter), a 10 ppm female (AH6637) exhibited a brown vaginal discharge on lactation day 1. During the F2a gestation period, untreated control female AH6617 exhibited physical changes that apparently were the result of a cage injury on gestation day 6 (crusty nose - red/brown, yellow/brown stained fur - perianal, weak, temperature - cool, did not use posterior appendages) and was sacrificed on gestation day 8. Another untreated control female, AH6613, was sacrificed moribund due to difficulties during parturition (brown vaginal discharge, reddish-brown discharge and matted fur - perianal, weak, cool to the touch, lethargic). This female delivered 6 stillborn pups over a 2-day period, starting 25 days post-copulation. on the second day of this prolonged parturition a tissue substance (possible pup) was found lodged in the vagina. No other noteworthy observations were seen during this study.

Dermal irritation (if dermal study):

not examined

Mortality:

mortality observed, non-treatment-related

Description (incidence):

2 F1 untreated control females were sacrificed moribund during the F2a litters; no deaths occurred among the treated animals.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

During the F1 generation body weight depressions (3 to 8%) were seen for the 5,000 ppm males starting at week 3 of the pre-mating period through final sacrifice. Statistical analyses revealed statistically significant reductions (p<0.05) in body weight at weeks 10 and 12 of the pre-mating period, at final sacrifice, and in the total weight change of the 5,000 ppm males. Weekly (pre- and post-mating) body weight data for the 5,000 ppm F1 females were similar to the untreated control females. Body weight data for the F1 generation dams (F2a litter) fed 5,000 ppm were similar to the untreated control darns. F1 generation animals fed 10 or 1,000 ppm of the test item exhibited body weights and weight gains that were similar to the untreated control animals. No treatment-related depressions were noted.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Statistical analyses of the pre-mating food consumption revealed significant reductions for the 5,000 ppm males during 3 of the 14 pre-mating intervals of the F1 generation. No statistically significant differences were seen in the 10 and 1,000 ppm male food consumption that were considered treatment-related. Sporadic statistical difference in feed intake were noted for the F1 generation treated females, however, these differences also did not reveal a dose response relationship and were not considered treatment-related. Analyses of the maternal food consumption revealed no statistically significant differences.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

Ovaries or testes (with epididymides) weights obtained for the treated animals were similar to the untreated control animals. No statistically significant differences were noted in mean organ weight data. Statistical analyses of the organ to body weight ratios revealed a significant increase in the testes (with epididymides)/to body weight ratios of the 5,000 ppm F1 generation males. This increase was considered a result of the body weight depressions noted for these males at final sacrifice since the testes (with epididymides) weight for these males were similar to the untreated control males.

Gross pathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

Necropsy examination of the F1 generation animals did not reveal any treatment-related lesions.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

No microscopic findings were seen which were considered treatment-related. A neoplastic diagnosis of the subcutaneous tissue was made. A fibroma was seen in a single control male. This was clearly a benign change typically seen in this species and strain in response to chronic traumatic insult.

Histopathological findings: neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

No microscopic findings were seen which were considered treatment-related.

Reproductive function / performance (P1)

Reproductive function: oestrous cycle:

no effects observed

Description (incidence and severity):

Reproductive performance was not adversely affected in the treated animals; no statistically significant reductions were noted in the reproductive indices

Reproductive function: sperm measures:

no effects observed

Description (incidence and severity):

Reproductive performance was not adversely affected in the treated animals; no statistically significant reductions were noted in the reproductive indices

Reproductive performance:

no effects observed

Description (incidence and severity):

Reproductive performance was not adversely affected in the treated animals; no statistically significant reductions were noted in the reproductive indices (see Table 3).

Effect levels (P1)

1

Dose descriptor:

NOAEL

Remarks:

Systemic toxicity

Effect level:

1 000 ppm

Based on:

test mat.

Sex:

male

Basis for effect level:

body weight and weight gain

2

Dose descriptor:

NOAEL

Remarks:

Toxicity to reproduction

Effect level:

> 5 000 ppm

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Remarks:

Dietary equivalent to 321 and 380 mg/kg bw/day for males and females, respectively.

Target system / organ toxicity (P1)

Critical effects observed:

no

Results: F1 generation

General toxicity (F1)

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality / viability:

mortality observed, non-treatment-related

Description (incidence and severity):

Survival indices calculated for the test item progeny were similar to the untreated control progeny. No statistically significant differences were noted

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

Progeny body weights were unaffected by the test item. Statistical analyses of the mean litter weight data did not reveal any consistent, treatment-related differences from the untreated control group. Statistically significant differences noted (5,000 ppm F1a pups on lactation day 7; 10 and 5,000 ppm F1b weaning males) were sporadic in occurrence and were not considered related to treatment.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

not examined

Anogenital distance (AGD):

not examined

Nipple retention in male pups:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment-related findings were noted.

Histopathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment-related findings were noted.

Developmental neurotoxicity (F1)

Behaviour (functional findings):

not examined

Developmental immunotoxicity (F1)

Developmental immunotoxicity:

not examined

Effect levels (F1)

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

1 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

Remarks on result:

other: Dietary equivalent to 64 and 73 mg/kg bw/day for males and females, respectively.

Target system / organ toxicity (F1)

Critical effects observed:

no

Results: F2 generation

General toxicity (F2)

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality / viability:

mortality observed, non-treatment-related

Description (incidence and severity):

Survival indices calculated for the test item progeny were similar to the untreated control progeny. No statistically significant differences were noted

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

Progeny body weights were unaffected by the test item. Statistical analyses of the mean litter weight data did not reveal any consistent, treatment-related differences from the untreated control group. Statistically significant differences noted (5000 ppm F2a pups on lactation day) were sporadic in occurrence and were not considered related to treatment

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

not examined

Anogenital distance (AGD):

not examined

Nipple retention in male pups:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Description (incidence and severity):

No treatment-related findings were noted.

Histopathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment-related findings were noted.

Developmental neurotoxicity (F2)

Behaviour (functional findings):

not examined

Developmental immunotoxicity (F2)

Developmental immunotoxicity:

not examined

Effect levels (F2)

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

1 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

Remarks on result:

other: Dietary equivalent to 64 and 73 mg/kg bw/day for males and females, respectively.

Target system / organ toxicity (F2)

Critical effects observed:

no

Overall reproductive toxicity

Reproductive effects observed:

no

Any other information on results incl. tables

Delivery and population data

Statistical analyses of the delivery and population data revealed no significant differences between the treated groups and the untreated control group. Similar numbers of viable pups were delivered and subsequently weaned by the treated groups and the untreated control group during both the F0 (F1a and F1b litters) and F1 (F2a litter) generations

 

Table 1: reproductive performance F0 generation – F1a litter

Group (PPM)	Mating index %	Fertility index %	Gestation index %	Female fertility index %	Male fertility index %	Average gestation length (days)
U-C (0)	60.0	81.5	100.0	73.3	73.3	22
T-I (10)	85.3	79.3	95.7	76.7	78.7	22
T-II (1000)	93.8	83.3	96.0	83.3	83.3	22
T-III (5000)	80.6	96.6	100.0	93.3	93.3	22

 

Table 2: reproductive performance F0 generation – F1b litter

Group (PPM)	Mating index %	Fertility index %	Gestation index %	Female fertility index %	Male fertility index %	Average gestation length (days)
U-C (0)	67.7	95.2	100.0	90.9	87.0	22
T-I (10)	92.0	91.3	100.0	91.3	91.3	22
T-II (1000)	100.0	96.0	100.0	96.0	96.0	22
T-III (5000)	71.1	77.8	100.0	75.0	75.0	22

 

Table 3: reproductive performance F1 generation – F2a litter

Group (PPM)	Mating index %	Fertility index %	Gestation index %	Female fertility index %	Male fertility index %	Average gestation length (days)
U-C (0)	76.3	89.7	96.2	86.7	86.7	22
T-I (10)	76.3	82.8	95.8	80.0	80.0	22
T-II (1000)	88.2	93.3	100.0	93.3	93.3	22
T-III (5000)	85.7	90.0	100.0	90.0	90.0	22

 

 

Applicant's summary and conclusion

Conclusions:

Exposure to 5000 ppm of the test item resulted in reductions in parental male body weights during both generations and parental female body weights during the first generation. Exposure to 1000 or 10 ppm did not result in any treatment-related effects on parental body weights or parental food consumption. Exposure to 5000, 1000 or 10 ppm did not result in any treatment-related effects on parental reproductive performance, progeny delivery and population data, progeny survival, or progeny body weights. Based on these results it is concluded that the maximum tolerated dose of the test item is 5000 ppm in the rat and the NOAEL is 1000 ppm (dietary equivalent to 64 and 73 mg/kg bw/day for males and females, respectively).

Executive summary:

An OECD TG 416 study was conducted according to GLP to evaluate the potential effects of the test item upon the growth and reproductive processes of two consecutive generations of the Sprague-Dawley derived albino rat. Animals were fed diets containing either 0, 10, 1000, or 5000 ppm (dietary equivalent to 0.6, 64 and 321 mg/kg bw/day for males, and 0.8, 73 and 380 mg/kg bw/day for femalesm respectively). There were no deaths or noteworthy antemortem observations that were treatment-related. Both generations of males fed 5000 ppm of test item weighed less than the concurrent untreated control males. Similar body weight reductions were seen for the 5000 ppm females during the first generation. However, body weights for the 5000 ppm F1 females were similar to the untreated control females. Pre- and post-mating body weights for the 10 and 1000 ppm animals were similar to the untreated control animals during both generations of the study. Exposure to 5000 ppm of the test item resulted in reduced maternal body weights during the F0 generation (F1a and F1b gestation and lactation periods). During the F1 generation (F2a litter), dams fed 5000 ppm weighed the same as the untreated control dams. No treatment-related body weight reductions were noted for either the 10 and 1000 ppm F0 or F1 dams during the gestation and lactation periods of both generations. Pre-mating food consumption data for the 5000 ppm males were statistically less than the untreated control males during 3 out of the 13 intervals evaluated for the F0 generation and 3 of the 14 F1 generation intervals. Food consumption data for the 5000 ppm females and the 10 and 1000 ppm males and females, during the pre-mating periods of both generations revealed no treatment-related reductions. Analyses of maternal food consumption revealed no statistically significant differences. Reproductive performance during the F1a, F1b, and F2a mating trials was not adversely affected by treatment with the test item. The mean numbers of progeny delivered (viable, stillborn, or found partially cannibalized) and viable during the lactation were similar to the untreated control group. Progeny survival was unaffected by treatment. Progeny body weight data were not adversely affected by treatment with the test item. No treatment-related structural anomalies were noted. Necropsy examinations of F0 and F1 parental animals and F1a, F1b, and F2a progeny found dead during lactation, sacrificed on lactation day 4, or sacrificed post-weaning revealed no treatment-related lesions. Ovaries or testes (with epididymides) weights for parental animals fed the test item were similar to the untreated control animal organ weights. Microscopic examination of reproductive organs from the F0 and F1 generation animals revealed no treatment-related findings. In conclusion, exposure to 5000 ppm of the test item resulted in reductions in parental male body weights during both generations and parental female body weights during the first generation. Exposure to 1000 or 10 ppm did not result in any treatment-related effects on parental body weights or parental food consumption. Exposure to 5000, 1000 or 10 ppm did not result in any treatment-related effects on parental reproductive performance, progeny delivery and population data, progeny survival, or progeny body weights. Based on these results it is concluded that the maximum tolerated dose of the test item is 5000 ppm in the rat and the NOAEL is 1000 ppm (dietary equivalent to 64 and 73 mg/kg bw/day for males and females, respectively).