2-chloro-N-[[[4-(trifluoromethoxy)phenyl]amino]carbonyl]benzamide

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

A pre-GLP and non-guideline 3-Generation study with triflumoron in rats is identified as key for this endoint. No adverse effects on fertility were observed.

Link to relevant study records

Reference

Endpoint:

multi-generation reproductive toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

July 1980 to June 1982

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

no guideline followed

Principles of method if other than guideline:

The multi-generation study was based on the recommendations published by the FDA:
Modern Trends in Toxicolop-y. Vol. 1, 1968, 75 - 85
Toxicol. appl. Pharmacol., Vol. 16, 1980, 264-369.
In a three-generation test with two litters per generation Triflumuron was administered in the diet to dose groups made of up to 10 male and 20 female rats at concentrations of 0, 20 200 or 2000 ppm. The F0 (parental generation) animals were exposed to Triflumuron for about 70 days before mating. The rats treated in this way were examined in regard to behaviour, body weight development, mortality, fertility, lactation ability, development of young, and also male/female
relation after two matings in each of three successive generations.
Histopathological examinations of main organs and tissues (including reproductive organs) was done on 4-week old pups of the F3 generation.

GLP compliance:

no

Remarks:

The study was performed before GLP principles were in place.

Species:

rat

Strain:

Wistar

Remarks:

SPF rats strain BOR:WISW.

Sex:

male/female

Details on test animals or test system and environmental conditions:

The animals were approximately 5-6 weeks old at start of study, with a mean starting weight of 80 g (males) and 79 g (females). The adaptation period was seven days. Apart from mating, the animals were kept singly in Makrolon cages type II, at a room temperature of approx. 22° C ± 1° C, atmospheric humidity of approx. 60 %, and with a 12-hour light/dark rhythm (artificial lighting from 7 am to 7 pm MET). During the matings and for F1b and F2b pups aged 4-8 weeks, Makrolon cages type III were used, the other conditions remaining the same. Fresh Altromin R and Ssniff R powdered feed (50:50) was provided once a week (the dams during lactation ad libitum) and tap water was available ad libitum.

Route of administration:

oral: feed

Details on exposure:

The rats received the test substance in the powdered feed. The test item was mixed in the feed weekly. The rats were treated with active ingredient over the entire study period, including mating,
gestation and lactation.

Details on mating procedure:

The 5-6 weeks old animals used for the study were kept singly for about 70 days (up to sexual maturity), and then mated. During mating two female rats were caged constantly with one male in a Makrolon cage type III. During the three-week mating period the males were interchanged, so that each female was confined with three different males, in each case for longer than the length of one cycle. After the matings the animals were recaged singly in type II cages. Five days after birth the litters were reduced, where necessary to ten animals. The animals with which treatment was to be continued were selected by a random list (except for the F1a generation). The young of the F1a, F2a and F3a generations were reared by the dams up to an age of four weeks, and then sacrificed. Each of the FO, F1b and F2b animals were mated for a second time after a two-week waiting period. After each second mating the pups (F1b and F2b) were kept and reared for four weeks like the pups from the first mating, and then separated from the dams and divided into sexes. After the 8th week groups of ten male and twenty female rats in each of the dose groups were chosen randomly for further matings. After reaching a mean age of approx. 100 days, mating took place as described above. The F0, F1b and F2b generation animals were sacrificed after rearing two sets of pups, approx. 1-3 weeks after weaning.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Before start of study stability and homogeneity in the feed were checked by analysis. During the
study the active ingredient content was checked regularly.

Frequency of treatment:

Continuous exposure via feed.

Details on study schedule:

Approximate length of each study phase:

Pre-treatment of F0 generation rats up to 1st mating: 70 days.

Length of 1st mating of F0 generation 20 days.

Length of gestation 21 Lactation of F1a pups up to 4- weeks with subsequent sacrifice: 28 days.

Waiting period: 14 days.

Length of 2nd mating of F0 generation: 20 days.

Length of gestation: 21 days.

Lactation of F1a pups up to age of approximately 100 days (sacrifice of F0 generation): 100 days.

Length of 1st mating of F1b generation: 20 days.

Length of gestation: 21 days.

Lactation of F2a pups up to 4-weeks with subsequent sacrifice: 28 days.

Waiting period: 14 days.

Length of 2nd mating of F1b generation: 20 days.

Length of gestation: 21 days.

Lactation of F2b pups up to age of approx. 100 days (sacrifice of F1b generation): 100 days.

Length of 1st mating of F2b generation: 20 days.

Length of gestation: 21 days.

Lactation of F3a pups up with subsequent sacrifice: 28 days.

Waiting period: 14 days.

Length of 2nd mating of F2b generation: 20 days.

Length of gestation: 21 days.

Lactation of F3b pups up to age of 4- weeks with subsequent sacrifice: 28 days.

Dose / conc.:

20 ppm

Remarks:

Low dose group, equivalent to 1.43 mg/kg bw/d in males and 1.50 mg/kg bw/d in females.

Dose / conc.:

200 ppm

Remarks:

Mid dose group, equivalent to 14.25 mg/kg bw/d in males and 15.04 mg/kg bw/d in females.

Dose / conc.:

2 000 ppm

Remarks:

High dose group, equivalent to 142.5 mg/kg bw/d in males and 150.4 mg/kg bw/d in females.

No. of animals per sex per dose:

In each test group there were ten male and twenty female rats.

Control animals:

yes

Details on study design:

Determination of the weights of parents: The rats were weighed before and after mating in a weekly rhythm. F0 animals were weighed weekly during first mating, and every three days during second mating. F1b and F2b rats were weighed every three days during both matings. Females which gained more than 35 g in weight within three weeks after mating were assumed to be pregnant. Parental F0 (indicated as P0 below), First (F1a, F1b) and second (F2a, F2b) generation results are presented.

In regards to pup data, each litter was weighed immediately after birth, on the 5th day after birth, after reduction to ten pups per birth, and after 1, 2, 3 and 4 weeks. On the date of birth the total number of pups born, the number of living and stillborn animals, and the ratio of males to females were also noted. We appraised the pups grossly immediately after birth, to detect malformations. The pups were also inspected during lactation for malformations.

Parental animals: Observations and examinations:

Appearance and behaviour was observed.

Postmortem examinations (parental animals):

Gross pathology: The 4-week old pups in the F3b generation and their parents (F2b generation) were anaesthetised with ether, sacrificed by exsanguination and then autopsied.

Postmortem examinations (offspring):

The F2b parents' livers, kidneys, testicles or ovaries were weighed.

The following organs of F3B pups were examined histopathologically: brain, eyes, thyroid, thymus, heart, lung, liver, spleen, pancreas, mesenteric lymph nodes, stomach, small intestine, kidneys, adrenals, urinary bladder, testicles, epididymes, sternum (bone marrow) skeletal musculature en bloc with femur, uterus and ovaries.
The organs examined came from one male and one female pup aged four weeks from each of ten dams per dose group. The animals were selected at random.

Statistics:

The following were calculated: arithmetic group means, standard deviations, upper and lower confidence limits at the confidence level of 1 - alpha = 95 % and 1 - alpha = 99 %. The findings for the test groups were compared with the control by the U significance test after MANN, WHITNEY and WILCOXON on the significance level alpha = 5 % and alpha = 1 %. For the indices (numerically derived) the confidence limits were calculated after CLOPPER and PEARSON on the confidence level 1 - alpha = 95 9 and 1 - alpha = 99 %. The test groups were compared with the control group with the exact Fisher test on the significance level alpha = 5 % and alpha = 1 %. To calculate the means of the pups' weights per dose the mean pup weight of each litter was used.

Reproductive indices:

Fertility index: number pregnant females/number mated females *100.

Gestation index: number females with live litters/number pregnant females*100.

Viability index: Number live pups after 5 days/number pups born*100.

Lactation index: Number live pups after 4 weeks/number live pups after 5 days after reduction to 10 *100.

Clinical signs:

no effects observed

Description (incidence and severity):

The rats in the 20 - 2000 ppm dose groups did not differ in appearance and behaviour from the controls during the study period.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

One female in each of the 0, 200 and 2000 ppm groups in the F0 generation died. The death of these
animals was not be attributed to the treatment.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Doses up to 2000 ppm did not result in any significant treatment-related effects on growth either in males or in females (F0 generation).

Reproductive performance:

no effects observed

Description (incidence and severity):

The females treated with up to 2000 ppm of the test material did not differ in respect to fertility and gestation indices to any great extent from the control females. The results are attached below in tabular form.
For the F1a and F1b generation, the total pups, numbers of males and females born, and the ratio in percent of males to females for each dose group. It is apparent that in the F1a and F1b generation, up to the dosage of 2000 ppm there were no differences attributable to the treatment in ratios of males to females and in the total number of animals born in comparison to the control.

Key result

Dose descriptor:

NOAEL

Effect level:

150.4 mg/kg bw/day

Based on:

test mat.

Sex:

male

Basis for effect level:

other:

Remarks on result:

other: Exposure based on calculation by estimation

Key result

Dose descriptor:

NOAEL

Effect level:

150.4 mg/kg bw/day

Based on:

test mat.

Sex:

female

Basis for effect level:

other:

Remarks on result:

other: Exposure based on calculation by estimation

Key result

Dose descriptor:

NOAEL

Effect level:

2 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other:

Remarks on result:

other: Highest dose level tested

Key result

Critical effects observed:

no

Clinical signs:

no effects observed

Description (incidence and severity):

In appearance and behaviour the rats (P1 and P2) in the 20 to 2000 ppm dose groups did not differ from the controls during the study period.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

For the P1, during 1st and 2nd mating one male died in the 20 ppm dose group. Autopsy of this animal did not detect a substance-related cause of death.
No mortality occurred in the P2- generation.

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

Treatment with the test item in doses up to and including 2000 ppm did not have any negative effects on the F1b generation's body weight development. Animals in the 200 ppm group (only females significant) were generally heavier than the controls. there were no treatment-related negative effects on body weight development up to 2000 ppm for males and females.

For the P2-generation, there were no treatment-related negative effects on body weight development up to 2000 ppm for males and females.

Description (incidence and severity):

Liver, kidney and testicles or ovaries from all the parent animals in the F2b generation were weighed on autopsy. No differences in weights were noted in males and females up to 2000 ppm which could not be explained by increased body weights, or which were in correlation to dose. The significantly lower relative kidney weights in females in the 2000 ppm group were within the normal range and not considered toxicologically relevant.

Gross pathological findings:

no effects observed

Description (incidence and severity):

All the parents in the F2b generation were sacrificed and examined two weeks after weaning. Gross appraisal of the dissected animals did not provide any indications of substance-specific organ damage.

Reproductive performance:

no effects observed

Description (incidence and severity):

P1:
Treatment with doses up to and including 2000 ppm after two matings had no negative influence on fertility and gestation. There were no dose related differences from the control either in total of animals born or in the male/female ratio. There were no significant differences between treated and untreated animals in regard to mean litter size.

P2:
The fertility and gestation indices detailed above do not reveal any treatment-induced effects on fertility and gestation in the dose groups up to 2000 ppm after both matings. The male/female ratio for the treated animals up to 2000 ppm did not differ from that of the controls. In regard to the total number of animals born, no dose correlation was apparent after first mating.
In the 2000 ppm group fewer F3b pups were born. This is considered a random result.

Key result

Dose descriptor:

NOAEL

Remarks:

P1 and P2

Effect level:

150.4 mg/kg bw/day

Based on:

test mat.

Sex:

female

Basis for effect level:

other: No adverse effects observed

Remarks on result:

other: Exposure based on calculation by estimation

Key result

Dose descriptor:

NOAEL

Effect level:

142.5 mg/kg bw/day

Based on:

test mat.

Sex:

male

Basis for effect level:

other: No adverse effects observed

Remarks on result:

other: Exposure based on calculation by estimation

Key result

Dose descriptor:

NOAEL

Effect level:

2 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No adverse effects observed up to and including the highest dose level tested

Remarks on result:

other: Highest dose level tested

Key result

Critical effects observed:

no

Clinical signs:

no effects observed

Mortality / viability:

mortality observed, non-treatment-related

Description (incidence and severity):

Viability after five days also did not differ in correlation to dose up to 2000 ppm. Any significant
deviations in the 200 ppm dose group (F1a and F1b) are not of toxicological importance.
The lactation index of treated dams up to 2000 ppm after two matings was about the same as that of the control females.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

The birth weights of the F1a and F1b pups in the groups up to 2000 ppm did not differ significantly from those of the untreated pups. The results are attached below in tabular form.
The F1a and F1b pups' growth in the treated groups up to 200 ppm was the same as that of the
controls. At 2000 ppm the F1b pups had significantly lower body weights in the 2nd, 3rd and 4th study weeks. As these deviations were only slight, they are not considered the result of a toxic effect on growth.

Gross pathological findings:

no effects observed

Description (incidence and severity):

All the pups were without grossly apparent abnormalities at birth and during lactation.

Key result

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

2 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No adverse effects observed

Remarks on result:

other: Highest dose level tested

Key result

Critical effects observed:

no

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

Description (incidence and severity):

No deaths occurred. In addition, compilation of the viability indices of the F3a and F3b pups did not reveal any dose correlation.
For the F3-generation, the female rats' lactation performances after the first mating up to 2000 ppm did not differ to any great extent from that of the controls. After the second mating there was a significant reduction in lactation index in the 20 ppm group. This figure is to be considered random and toxicologically insignificant, as better lactation indices were noted at 200 and 2000 ppm.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Administration of up to a dosage of 2000 ppm did not produce any significant and dose-related effects on the birth weights of F2a and F2b pups. The results are attached below in tabular form.

The F3a rats' body weights in the 20 to 2000 ppm dose groups were slightly but significantly lower than those of the controls, and this may be explained by the relatively high birth weight in the control group, so that the finding should not be attributed to the treatment. This is confirmed by the fact that birth weights corresponding to those of the controls were recorded after the second mating up to and including 2000 ppm.

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Description (incidence and severity):

No deformed pups were noted in the F2 and F3 generations, either immediately after birth or during the lactation period.

The pathological anatomical examination of F3 pups did not reveal any alterations attributable to dosage with the test item.

Histopathological findings:

no effects observed

Description (incidence and severity):

No indications of organ alterations were noted in any of the animals examined (F3-generation).

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Generation:

other: F2 and F3

Effect level:

2 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no adverse effects observed

Remarks on result:

other: Highest dose level tested

Key result

Critical effects observed:

no

Key result

Reproductive effects observed:

no

Treatment related:

no

Conclusions:

A 3-generation study (two litters per generation) was performed using dietary concentrations of 20, 200 and 2000 ppm. Based on the results of this study a NOAEL for reproduction of 2000 ppm (corresponding to 142.5 and 150.4 mg/kg bw/d in males and females, respectively) is established.

Executive summary:

Triflumuron was examined in a multigeneration study in rats for its potential effect on reproduction. Triflumuron was administered to male and female rats at dietary concentrations of 0 (controls), 20, 200 and 2000 ppm. The rats treated in this way were examined in regard to behaviour, body weight development, mortality, fertility, lactation ability, development of young, and also sex ratio after two matings in each of three successive generations.
Behaviour, mortality, body weight development in parent and young animals were not negatively affected by administration of Triflumuron up to 2000 ppm. In regard to fertility, viability, lactation, litter size and birth weight no dose-related  differences between treated and untreated animals were recorded up to the dosage of 2000 ppm. In the dose groups up to and including 2000 ppm no malformations were noted in the young. The sex ratio did not show substance-specific abnormalities in any generation up to 2000 ppm. Autopsies on parents which died and were sacrificed provided no indications of treatment-related alterations. Gross and microscopic appraisal of the F3b young provided no indications of organ damage up to 2000 ppm. Triflumuron was tolerated under the conditions described in the multigeneration study, in concentrations up to 2000 ppm in the feed, without damage to reproduction. Food consumption was not measured in this study. For estimation of the compound intake at the NOAEL of 2000 ppm, the lowest food intake values out of four reference studies were taken into account. The NOAEL for reproduction was therefore established retrospectively at 142.5 and 150.4 mg/kg bw/d for males and females, respectively.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

133 mg/kg bw/day

Study duration:

chronic

Species:

rat

Quality of whole database:

A reliable multigeneration study with triflumuron in rat is available for this endpoint (RL2).

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Effects on developmental toxicity

Description of key information

The key studies for developmental toxicity endpoints are as follows:
rat (US EPA 83-3, GLP, RL2), NOAEL (maternal and developmental) = 300 mg/kg bw/day (M-086940-01-1, 1986)
rabbit (US EPA 83-3, GLP, RL2), NOAEL (maternal and developmental) = 300 mg/kg bw/day (M- 0865549-01-1, 1987)
The following supporting studies are available:
rat (no guideline, non-GLP, RL4), NOAEL (maternal and developmental) = 100 mg/kg bw/day (highest dose tested) (M- 087602-01-1, 1981)
rabbit (no guideline, non-GLP, RL4), NOAEL (maternal and developmental) = 100 mg/kg bw/day (highest dose tested) (M-087600-01-1, 1981)

Link to relevant study records

Reference

Reference 1

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

April 18 1986 - August 25 1986

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EPA OPP 83-3 (Prenatal Developmental Toxicity Study)

Version / remarks:

1984

Deviations:

no

Principles of method if other than guideline:

The study does not fully comply with the current version of the test guideline (OECD 414) related to the dosing period (gestation day 6-18).

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

Himalayan

Remarks:

CHBB:HM

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: THOMAE, Biberach a.d. Riss, Germany
- Age at study initiation: Sexually mature
- Weight at study initiation: 2056 -3319 g (females)
- Housing: Individually housed in perforated sheet metal cages under conventional conditions
- Diet: Ssniff K4 rabbit feed (Ssniff Spezialfutter GmbH, Soest, Germany)
- Water: tap water
- Acclimation period: Appr. 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ±3
- Humidity (%): 60 (average)
- Air changes (per hr): at least 10 times
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: April to August 1986

Route of administration:

oral: gavage

Vehicle:

other: aqueous 0.5% Cremophor EL emulsion

Details on exposure:

The test substance was administered orally via intubation tube. The volume administered was 5 mL/kg body weight and was the same for the animals of all test groups

Analytical verification of doses or concentrations:

yes

Details on mating procedure:

One female was placed together with one male; copulation was verified by observation. This day was designated Gestation Day 0.

Duration of treatment / exposure:

Males were not treated. Females were treated from gestation day 6-18.

Frequency of treatment:

Once daily

Duration of test:

Females were sacrificed on Gestation Day 29.

Dose / conc.:

100 mg/kg bw/day

Remarks:

Low dose group

Dose / conc.:

300 mg/kg bw/day

Remarks:

Mid dose group

Dose / conc.:

1 000 mg/kg bw/day

Remarks:

High dose group

No. of animals per sex per dose:

The main study group comprised of groups of 20 rats. The satellite groups comprised of two groups of 5 rabbits.

Control animals:

yes

Details on study design:

The dose levels were established on the basis of a range-finding test on rats using a dose of 1000 mg/kg bw, in which the conventional parameters of embryotoxicity studies revealed no indications of maternal toxicity.

Maternal examinations:

The appearance and behavior of the animals were observed at least once each day from Gestation Day 0 to 29. At cesarean section dams were grossly examined.

For the animals in the satellite group (sacrificed GD 19) the spleen weight was determined.

Ovaries and uterine content:

The following examinations were performed:
- Determination of the number of implantations
- Determination of the number of corpora lutea
- Determination of the uterus weight

Blood sampling:

Blood samples of the rabbits of the satellite groups were obtained on GD 19.
Parameters: Leukocytes, erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, thrombocyte count.

Fetal examinations:

The following examinations were performed:
- Determination of the number of live and dead fetuses/embryos (dams without live fetuses are classified as not pregnant)
- Determination of the sex of all live fetuses
- Determination of the individual fetus weights and the stunted fetuses
- Determination of the placental weight
- Determination of the crown-rump length of the fetuses
- Thorough inspection of all fetuses for external malformations
- Examination of the fetal cranium for visceral malformations by the modified WILSON technique
- Evisceration of the fetuses and evaluation of the abdominal and thoracic organs with subsequent clearing of the fetuses with dilute potassium hydroxide, staining of the skeletal system with alizarin red S, and evaluation of the skeletal system (DAWSON)

Statistics:

The following methods were used to test for statistical significance:

a. Nonparametric rank sum test of WILCOXON (WILCOXON-MANN-WHITNEY U-test), e.g. for body weight gains, number of implantations, number of fetuses, and number of resorptions.

b. Chi-square test (correction of YATES), e.g. for the number of stunted fetuses.

c. Chi-square test (correction of YATES or as the so-called exact test of FISHER, depending on the frequency anticipated) for the indices of fertilized and pregnant animals.

Unless otherwise indicated, a difference is regarded as significant if, in the statistical calculation, the probability of error is less than 5%.

Clinical signs:

no effects observed

Description (incidence and severity):

No changes in the appearance and behavior of the dams that could be regarded as a result of treatment were found during the daily observations.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

One rabbit in the high dose group died during the study. The cause of death was not determinable, a bloody nose was noted , presumably following fight with animal in adjacent cage. This animal was pregnant.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Decreased body weight gain was noted for rabbits exposed to the test item. The results are attached below in tabular form.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

There were significant reductions in the hemoglobin, erythrocyte count, and hematocrit, and the leukocyte count was increased as a result of the damage to the red blood profile (results from satellite group). The results are attached below in tabular form.

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

There was a significant increase in spleen weight of the rabbits of the 1000 mg/kg bw satellite group.

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

effects observed, non-treatment-related

Description (incidence and severity):

There was a significant increase in the resorption index in the 1000 mg/kg bw group. This increase in the index was caused by a single dam, which resorbed completely. Without this animal, the resorption index would be 1.0. Moreover, the resorption index in the control group was very low. The mean resorption index of historical control data was 1.04 per dam.

Dead fetuses:

no effects observed

Changes in number of pregnant:

no effects observed

Description (incidence and severity):

The number of fertilized animals was 13/20 in the control group and 18/20, 16/20 and 13/20 in the low, mid and high dose groups, respectively. The number of pregnant animals of the fertilized animals at cesarean section was 13/13 in the control group and 17/18, 16/16 and 11/13 in the low, mid and high dose groups, respectively. As the pregnancy rate was in the range of the historical control values, the test item was concluded to have no effect on the pregnancy rate.

Details on maternal toxic effects:

Results are attached below in tabular form.

Key result

Dose descriptor:

NOAEL

Effect level:

300 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

haematology

organ weights and organ / body weight ratios

Remarks on result:

other: Based on results satellite group

Key result

Abnormalities:

no effects observed

Fetal body weight changes:

no effects observed

Changes in sex ratio:

effects observed, non-treatment-related

Description (incidence and severity):

A significantly lower number of male fetuses in the highest dose group was observed but this was regarded as a random occurrence.

Changes in litter size and weights:

no effects observed

External malformations:

no effects observed

Description (incidence and severity):

The type and frequency of the malformations are consistent with the spontaneously occurring malformations of the strain employed. The results are attached below in tabular form.

Skeletal malformations:

no effects observed

Description (incidence and severity):

No fetuses with slight skeletal changes in the form of delayed ossification were observed. The results are attached below in tabular form.

Visceral malformations:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

300 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Increased number of resorptions at 1000 mg/kg bw/day

Key result

Abnormalities:

no effects observed

Key result

Developmental effects observed:

no

Conclusions:

A prenatal developmental toxicity study was performed in the rabbit (20/group) using gavage dose levels of 100, 300 and 1000 mg/kg bw/d. An increased proportion of resorptions was seen at the highest and maternally toxic dose level (1000 mg/kg bw/day).

Executive summary:

The test substance triflumuron was administered orally to at least 20 inseminated Himalayan rabbits per group daily from Gestation Day 6 to Gestation Day 18 at dose levels of 0, 100, 300, and 1000 mg/kg bw/d.  The dams were examined with respect to body weight, appearance, and behavior. The fetuses obtained via cesarean section on Gestation Day 29 were examined for detrimental effects by weighing and by external and internal morphological evaluation.

In addition, satellite groups of 5 rabbits each were treated at 0 or 1000 mg/kg body weight; treatment was comparable to that of the main groups. Blood samples were obtained from these rabbits on Gestation Day 19 for haematological examinations. Moreover, their spleens were weighed. There were no treatment-related mortalities. No treatment-related changes in appearance or behavior were observed. There was no effect on the pattern of body weight gain of the dams. In the satellite group treated at 1000 mg/kg bw/d, the spleen weights were increased; there were reductions in erythrocyte count, haemoglobin, and haematocrit. Dose levels of up to and including 300 mg/kg bw/day had no effect on embryonic/fetal development. There was a significant increase in the number of resorptions at 1000 mg/kg/ bw/d. This dose was toxic to the dams. No teratogenic effects were observed up to and including 1000 mg/kg bw/day.

Reference 2

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

April 1986 - September 1986

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EPA OPP 83-3 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

Additional satellite groups consisting of five animals were treated with doses of 0 or 1000 mg/kg bw/day and used for hematological and gravimetric spleen examinations on post-insemination day 16.

Route of administration:

oral: unspecified

Details on exposure:

Relative to the immediate body weight, doses were administered to the female rats orally at the same time each day from Gestation Day 6 - 15.
The volume administered was 10 mL/kg body weight and was the same for the animals of all test groups.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The concentration, stability and homogenecty of the administration formulations were analyzed.
Stability of the test item in the formulation was verified over 7 days (98-101% of nominal after 7 days). Homegenicity of the formulations was confirmed analytically (89 - 97% of nominal, 3 samples tested from 10 and 200 mg/mL formulations each).
The test item concentration in the formulations was determined (83% - 94% of nominal concentration for the three dose formulations).

Details on mating procedure:

The mating was performed overnight; one male was placed together with two females in a Type III Makrolon cage. If sperm was detected in a vaginal smear on the morning following mating, this day was designated Gestation Day 0.

Duration of treatment / exposure:

10 days (Gestation Day 6 - 15)

Frequency of treatment:

Once daily

Duration of test:

Rats were sacrificed on gestational day 20.

Dose / conc.:

100 mg/kg bw/day

Remarks:

Low dose group

Dose / conc.:

300 mg/kg bw/day

Remarks:

Mid dose group

Dose / conc.:

1 000 mg/kg bw/day

Remarks:

High dose group

No. of animals per sex per dose:

25 females

Control animals:

yes, concurrent vehicle

Details on study design:

Satellite groups of 5 female rats were dosed at 0 and 1000 mg/kg bw. Blood samples were obtained
from these rats on the 16th day p.c. for hematological examinations. Moreover, the spleens were weighed.
The mean feed consumption was determined from Day 0 to 6, 6 to 11, 11 to 16, and 16 to 20 p.c.

Blood sampling:

Blood samples of the rats of the satellite groups were obtained by cardiac puncture on gestational day 16.
Parameters: Leukocytes, erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, thrombocyte count.
Spleen weights were determined.

Fetal examinations:

The following examinations were performed at cesarean section:
a. Gross examination of the dams
b. Determination of the number of implantations
c. Determination of the number of corpora lutea
d. Determination of the uterus weight
e. Determination of the number of live and dead fetuses /embryos (dams without live fetuses are classified as not pregnant)
f. Determination of the sex of all live fetuses
g. Determination of the individual fetus weights and the stunted fetuses
h. Determination of the placental weight
i. Determination of the crown-rump length of the fetuses
j . Thorough inspection of all fetuses for external malformations
k. Examination of a number of fetuses for visceral malformations by the modified WILSON technique
l. Evisceration of the remaining fetuses and evaluation of the abdominal and thoracic organs with subsequent clearing of the fetuses with dilute potassium hydroxide, staining of the skeletal
system with alizarin red S, and evaluation of the skeletal system (DAWSON)

Statistics:

The following methods were used to test for statistical significance:

a. Nonparametric rank sum test of WILCOXON (WILCOXON-MANN-WHITNEY U-test), e.g. for body weight gains, number of implantations, number of fetuses, and number of resorptions.

b. Chi-square test (correction of YATES), e.g. for the number of stunted fetuses.

c. Chi-square test (correction of YATES or as the so-called exact test of FISHER, depending on the frequency anticipated) for the indices of fertilized and pregnant animals.

Clinical signs:

no effects observed

Description (incidence and severity):

No changes in the appearance and behavior of the dams that could be regarded as a result of treatment were found during the daily observations. Gross examination of the dams revealed no treatment-related changes

Mortality:

no mortality observed

Description (incidence):

None of the animals died during the test period.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

The mean body weight gains in the groups of the main experiment during the administration period and during the entire gestation period were considered comparable. The results are attached below in tabular form.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

The feed consumption was comparable in all groups.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

There were significant reductions in the hemoglobin, erythrocyte counts, and hematocrit of the rats treated at 1000 mg/kg bw/day. The results are attached below in tabular form below.

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Spleen weights were elevated in the exposed groups, however the effect was not statistically significant.

Gross pathological findings:

no effects observed

Description (incidence and severity):

No treatment-related findings were noted.

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

The results are attached below.

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Changes in number of pregnant:

no effects observed

Description (incidence and severity):

No effect of the test item on pregnancy rate was found. The number of fertilized rats was 24, 25, 24 and 25 for the control group and the low, mid and high dose group, respectively. All the fertilized rats were pregnant at sacrifice.

Key result

Dose descriptor:

NOAEL

Remarks:

Maternal

Effect level:

300 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

haematology

Key result

Abnormalities:

no effects observed

Fetal body weight changes:

no effects observed

Description (incidence and severity):

Details on the results are included in the attachment.

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

no effects observed

Skeletal malformations:

effects observed, treatment-related

Description (incidence and severity):

The type and frequency of the findings were consistent with the spectrum of spontaneous malformations for the strain employed.
A significant increase in the number of fetuses with slight skeletal changes in the 1000 mg/kg bw/day group was observed. The results are attached below in tabular form. There was no relevant increase in the number of fetuses with skeletal changes per litter up to and including 300 mg/kg bw/day.

Visceral malformations:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

300 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Delayed skeletal ossification

Remarks on result:

other: adverse effect associated with maternal toxicity

Key result

Abnormalities:

no effects observed

Key result

Developmental effects observed:

yes

Lowest effective dose / conc.:

1 000 mg/kg bw/day

Treatment related:

yes

Relation to maternal toxicity:

developmental effects as a secondary non-specific consequence of maternal toxicity effects

Dose response relationship:

yes

Relevant for humans:

yes

Conclusions:

A prenatal study was performed in the rat (25/group) at dose levels of 100, 300 and 1000 mg/kg bw/d. An increased proportion of fetuses with skeletal developmental delays was seen at the highest and maternally toxic dose level.

Executive summary:

Triflumuron was administered orally to 25 inseminated Wistar rats per group daily from Gestation Day 6 to Gestation Day 15 at dose levels of 0, 100, 300, and 1000 mg/kg bw/d. The dams were examined with respect to body weight, appearance, and behavior. The fetuses obtained via cesarean section on Gestation Day 20 were examined for detrimental effects by weighing and by external and internal morphological evaluation. In addition, satellite groups of 5 rats each were treated at 0 or 1000 mg/kg bw/d/. Blood samples were obtained from these rats on Gestation Day 16 for hematological examinations. Moreover, their spleens were weighed. There were no treatment-related mortalities. There was no effect on the pattern of body weight gain of the dams. In the satellite group treated at 1000 mg/kg bw/d, there were reductions in erythrocyte count, hemoglobin, and hematocrit. The spleen weights were increased. Thus the dose level of 1000 mg/kg bw/d had a toxic effect on the dams. As a sign of  embryotoxicity, there was an increase in the number of fetuses with delayed skeletal development in the 1000 mg/kg bw/d group. No indications of potential teratogenicity were found.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

300 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

GLP- and guideline- (OECD 414) compliant studies of developmental toxicity in the rat and rabbit are available for triflumuron (both RL2).

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Justification for classification or non-classification

The results of the available studies (multigeneration study in rats and developmental toxicity studies in rats and rabbits) do not indicate that Trimflumuron induces reproductive and/or developmental toxicity. Therefore, Triflumuron does not meet the criteria for classification for fertility and developmental toxicity according to Regulation (EC) No. 1272/2008.