Fluometuron

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records

Reference

Endpoint:

three-generation reproductive toxicity

Remarks:

based on test type (migrated information)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 August 1978 to 10 April 1980

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study with acceptable restrictions. The experiment was performed and compiled to a great extent to then in force EPA guidelines (proposed guidelines for registering pesticides in the US, Federal Register. Vol 43, no 163, August 22,1978).

Qualifier:

equivalent or similar to guideline

Guideline:

EU Method B.35 (Two-Generation Reproduction Toxicity Test)

Deviations:

yes

Remarks:

The study is a three-generation reproduction toxicity test.

Principles of method if other than guideline:

The experiment was performed and compiled to a great extent to then in force EPA guidelines (proposed guidelines for registering pesticides in the US, Federal Register. Vol 43, no 163, August 22,1978). The method used differs from the prescribed method EU/88/302, B.35:
The study is a three-generation reproduction toxicity test rather than the prescribed two-generation reproduction toxicity test.
The number of males (8) and females (16) used for mating is lower than the males (10) and females (20) as recommended however, as the ratio maintained was 1:2 and three generations were used for reproduction an assessment of the reproductive effects of fluometuron can be made
The deviations do not limit or impair the scientific validity of the study.

GLP compliance:

yes

Limit test:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

- thirty-two male and sixty-four female weanling albino Sprague-Dawley rats
- Source: Charles River Breeding Laboratories, Inc., Wilmington, Massachusetts, received 19. July 1978
- weight: males: 149-210 g, females: 99-187 g
- Tap water and the appropriate diet (compound blended in Purina Rodent Laboratory Chow® ) were available ad libitum
- room temperature

Route of administration:

oral: feed

Vehicle:

other: administered in the diet

Details on mating procedure:

Upon completion of weaning of the F1b and F2b litters, eight males and sixteen females per treatment level were selected as the P2 and P3 generations, respectively, by a random card draw method (at least one pup from every available litter) and assigned individual identification numbers. The animals remained in their original dietary group.
The P1 generation was fed for 9 weeks before being cohabited (1 male with 2 females from the same dose group) for mating over a maximum of 21 days. Vaginal smears were taken daily during co-habitation to establish the date of mating. The day on which a vaginal plug was found or sperm were observed in the smear was designated Day 0 of gestation. At mating, the females were individually housed and the male returned to its original cage. Any female failing to mate after 1 week of co-habitation was re-cohabited with a different male from the same dose level. A maximum of 3 males was co-habited with each female during one breeding cycle, each being co-habited for up to one week.
On Day 5 after birth (post partum), the F1a litters were reduced (standardised) to 5 males and 5 females, where possible, by random selection. These pups were then reared to weaning and killed. Ten days after weaning, the P1 animals were re-cohabited using the same male:female ‘pairings’ for a further 21 days to produce the F1b litters, which were also standardised on Day 5 post partum, then reared to Day 21 post partum when 8 males and 16 females per group was selected to form the P2 generation. Remaining offspring were discarded. Parental animals were killed once the second litter had been weaned.
The P2 generation was fed for a 9-week growth period then bred as described above to derive the F2a and F2b litters which were standardised to a randomly selected 5/sex, where possible, on Day 5 post partum. The P3 generation (8 males and 16 females/group), similarly derived from the standardised F2b litters, was reared for 9 weeks prior to breeding to produce the F3a and F3b litters.
Fluometuron technical was administered continously in the diet through three successive generations of male and female albino rats.
Appropriate amounts of the test material required for each test level were individually weighed on a Mettler balance (beginning with week 53 of the study, an Arbor balance was utilized) and premixed with a small amount of feed in a Waring blender. The pre-mix was then added to the total amount of feed required on a weight-per-weight basis, and thouroughly mixed in a twin shell blender for minute/kg. The control animals received only the basal diet. Fresh diets were prepared and presented weekly.

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

The P1 generation was fed for 9 weeks before being cohabited (1 male with 2 females from the same dose group) for mating over a maximum of 21 days. The P2 generation was fed for a 9-week growth period then bred as described above to derive the F2a and F2b litters which were standardised to a randomly selected 5/sex, where possible, on Day 5 post partum. The P3 generation (8 males and 16 females/group), similarly derived from the standardised F2b litters, was reared for 9 weeks prior to breeding to produce the F3a and F3b litters.
The study was initiated on 10. August 1978 and was terminated on 10. April 1980 with the sacrifice of the F3b generation.

Frequency of treatment:

continously with the diet, 7 days each week

Remarks:

Doses / Concentrations:
0 ppm (Control: group 1)
Basis:
nominal in diet
0, 10, 300 or 1000 ppm of fluometuron technical for three generations m/f (P1, P2 and P3)

Remarks:

Doses / Concentrations:
10 ppm (Low: group 2)
Basis:
nominal in diet
0, 10, 300 or 1000 ppm of fluometuron technical for three generations m/f (P1, P2 and P3)

Remarks:

Doses / Concentrations:
300 ppm (Mid: group 3)
Basis:
nominal in diet
0, 10, 300 or 1000 ppm of fluometuron technical for three generations m/f (P1, P2 and P3)

Remarks:

Doses / Concentrations:
1000 ppm (High: group 4)
Basis:
nominal in diet
0, 10, 300 or 1000 ppm of fluometuron technical for three generations m/f (P1, P2 and P3)

No. of animals per sex per dose:

Group 1 (Control): 8 males, 16 females, dietrary level: 0 ppm
Group 2 (Low): 8 males, 16 females, dietrary level: 10 ppm
Group 3 (Mid): 8 males, 16 females, dietrary level: 300 ppm
Group 1 (High): 8 males, 16 females, dietrary level: 1000 ppm

Control animals:

yes, concurrent no treatment

Details on study design:

Groups of 8 male and 16 female Sprague Dawley CD rats were given dietary concentrations of either 0, 10, 300 or 1000 ppm of fluometuron technical for three generations (P1, P2 and P3). Each generation was bred twice. The first litters (F1a, F2a and F3a) were killed at weaning on Day 21 post partum whilst the second litters (F1b, F2b and F3b) were used to derive the P2 and P3 generations, respectively. Treatment was continuous from initiation of the P1 generation to necropsy of the last surviving animals (P3 parental animals and F3b offspring). Parental animals were housed individually except during the breeding periods.

Parental animals: Observations and examinations:

Parental animals were observed daily for mortality and moribundity. Individual parental body weights and food consumption were recorded weekly during the 9-week growth period. Mated females were observed daily during gestation. Maternal body weights were noted on Days 0, 7, 14 and 21 of gestation.

Oestrous cyclicity (parental animals):

P1 generation: Vaginal smears were taken daily during co-habitation to establish the date of mating. The day on which a vaginal plug was found or sperm were observed in the smear was designated Day 0 of gestation

Litter observations:

All litters were examined 24 hours after birth (on Day 1 post partum) and the number of live and dead pups, their sex and any macroscopic abnormalities recorded. Pups were also observed and individual weights noted on Day 5 post partum (after the litters had been standardised) and on Day 14 when the total number of live pups of each sex was documented.

Postmortem examinations (parental animals):

Parental animals were killed after the second litter had been weaned. They examined for macroscopic abnormalities, the weights of the brain, heart, liver, spleen, kidneys, testes and epididymides, and ovaries recorded. A comprehensive range of tissues was preserved and those from 5 males and 5 females from each of the control and highest dose group (1000 ppm) examined histopathologically.
In addition, the liver and spleen from 5 rats/sex from the low and intermediate dose groups from each parental generation selected for necropsy were examined microscopically

Postmortem examinations (offspring):

The liver and spleen from 5 rats/sex from the low and intermediate dose groups from 10 pups/sex from the same dose groups of the F3b litters selected for necropsy were examined microscopically.

Clinical signs:

no effects observed

Description (incidence and severity):

No treatment related effects were noted with respect to parental survival, clinical observations, maternal body weight or food consumption values during gestation.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Slightly higher mean body weights were generally noted in the P1 male and female and P2 male treated animals. In the P3 females, the mean body weight values were generally higher than control values in the low-dose group.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Slightly higher mean body weights were generally noted in the P1 male and female and P2 male treated animals. In the P3 females, the mean body weight values were generally higher than control values in the low-dose group.

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

(P1, P2, P3; f/m; mddle (300m ppm)/high (100 ppm) doses: increased extramedullary haematopoiesis and haemosiderin-laden macrophages, Increased extramedullary haematopoiesis

Other effects:

effects observed, treatment-related

Description (incidence and severity):

Test substance intake: The food consumption values of the treated groups in each parental generation during growth were generally higher than respective control values.

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

no effects observed

Description (incidence and severity):

Evaluation of the reproduction indices revealed no treatment related trends through the three generations.

Slightly higher mean body weights were generally noted in the P1 male and female and P2 male treated animals when compared to the respective control groups. However the P1 initial mean body weights were also slightly higher in the treated groups compared to control. The mean body weight values of the P2 high dose females were generally lower than those of the respective control, while the mean body weight values of the P2 low dose females were significantly higher than those of the respective control animals at Weeks 1, 2 and 5. During the P3 growth period the mean body weight values were generally higher from Weeks 2 through 9 in the male treated groups than in the control group. In the P3 females, the mean body weight values were generally higher than control values in the low-dose group, and lower than control values in the mid and high dose groups (significantly lower at Week 0 in the mid-dose group).
See Tables 7.8.1-01 to 7.8.1-06
The food consumption values of the treated groups in each parental generation during growth were generally higher than respective control values. With several significantly higher than control values noted at various weeks.
See Tables 7.8.1-01 to 7.8.1-06.
No treatment related effects were noted with respect to parental survival, clinical observations, maternal body weight or food consumption values during gestation.
Enlarged, dark red, and/or dark brown spleens were the most common necropsy observations noted in the parental animals. Enlarged spleens were noted in greater incidence in the high-dose P1 females and the P3 treated males and females than in the respective control groups. Dark red or dark brown spleens were also noted in the high-dose P2 females in greater incidence than in the respective control group, see
Table 7.8.1-07.
Other necropsy findings appeared to be incidental in nature and not related to treatment.
Evaluation of the organ weights and the organ/body weight ratios revealed a treatment-related increase in the mean absolute and relative spleen weights in each parental generation. Significantly higher than respective control values were noted in the following spleen weights: P1 females – mean absolute spleen weight in the mid and high dose groups and the mean relative spleen weight in the high dose group; P2 males - mean absolute spleen weight in all treated groups and mean relative spleen weight in the high dose groups, P2 females - mean absolute spleen weight in the mid and high dose groups and mean relative spleen weight in the mid and high dose groups; P3 males - mean absolute and relative spleen weight in the high dose group; P3 females - mean absolute spleen weight in the high dose group and mean relative spleen weight in the mid and high dose groups. Occasional significant differences from control values were noted in organ weights and organ body/weight ratios other than the spleen. These are believed not to compound related.
See Tables 7.8.1-08 to 7.8.1-13.
Similar findings in the histopathological evaluation were noted in the three parental generations. In the P1 animals compound-related histomorphical changes were present in the spleen in the high-dose animals of both sexes and mid-dose females characterised by increased extramedullary haematopoiesis and haemosiderin-laden macrophages (confirmed by Gomori’s iron strain). Spleen sections of the low-dose animals and mid-dose males were comparable to untreated animals. Compound related histomorphologic alterations in the P2 generation were present in the spleen of the high and mid-dose animals of both sexes as observed in the P1 generation. Increased extramedullary haematopoiesis was observed in the high dose animals of both sexes and mid-dose females. The mid-dose males also had slightly increased numbers of pigment-laden macrophages. Congestion of the spleen was present in nine out of ten high-dose animals and one mid-dose female. Similar compound related histomorphologic alterations in the P3 generation were present in the spleens of the high and mid-dose rats of both sexes, characterised by increased extramedullary haematopoiesis and pigment - laden macrophages. Spontaneous disease lesions and incidental findings were comparable in incidence and severity in the control and treated animals of each parental generation.
See Tables 7.8.1-14 to 7.8.1-16

Dose descriptor:

NOEL

Effect level:

>= 0.8 - <= 0.9 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

effects were observed, but considered negligible

Mortality / viability:

mortality observed, treatment-related

Description (incidence and severity):

P1-F1b generation revealed a possible treatment-related decrease in survival.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

effects were observed, but considered negligible

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

P3-F3b, m/f 1000 ppm: absolute spleen weights; P3-F3b f/m 1000 ppm, f 300 ppm: relative spleen weights

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

offspring of the P3-F3b, f/m, 1000 ppm: enlarged spleens, dark or dark red spleens

Histopathological findings:

effects observed, treatment-related

Description (incidence and severity):

F3b, f/m 1000 ppm: increased extramedullary haematopoiesis, pigment laden macrophages and congestion

Evaluation of the reproduction indices revealed no treatment related trends through the three generations. Although some slight differences were observed between the control and treated groups, no statistical differences were noted.
The lactation survival indices, the mean neonatal survival indices and the mean weaning survival indices of the P1-F1b generation revealed a possible treatment-related decrease in survival. Except for the neonatal index of the low-dose group, the treated groups showed a decrease from control in each case, with the high dose group showing a significant decrease in the mean weaning survival index when compared to the control group. Decreases in the survival indices were also noted in the P3-F3b generation, however the decrease was also noted in the control as well as the treated groups. The survival indices of the P1-F1a, P2 and P3 generations did not reflect any abnormal or consistent differences between the respective control and treated groups.
Significantly lower than control values were noted in the P1-F1a sex ratios on Days 1 and 5 (pre and post-cull) and in the P1-F1b sex ration on Day 1; however the sex ratios of each filial generation demonstrated normal biological variation.
No consistent or treatment-related effects were noted in the offspring body weight data. Lower than control values were generally noted in the P1-F1a female and P1-F1b male and female body weight values; and higher than control values (including significantly higher values in the P2-F2a low-dose males and females on Day 21, and in the P3-F3a high-dose males on Day 21) were noted occasionally in the P2 and P3 filial generations.
See Tables 7.8.1-17 to 7.8.1-19.
Offspring clinical observations revealed no apparent treatment-related effects. A number of incidental findings are believed to be natural occurrences and not thought to be compound related.
Gross pathology findings in the offspring of the P3-F3b generation included enlarged spleens in the high-dose males (five out of ten animals) and high-dose females (one out of ten animals). Dark or dark red spleens were noted in the both the high dose male (three out of ten animals) and females (one out of ten animals) groups with slightly greater incidence than control. Other F3b findings appeared to be incidental in nature and did not appear to be compound related.
Evaluation of the offspring in the P3-F3b generation organ weights and organ/body weight ratios revealed a treatment-related increase in the mean absolute and relative spleen weights in the males and females, consistent with the parental findings. Significantly higher than respective control values were noted in the offspring mean absolute spleen weights of the high-dose males and females and in the mean relative spleen weights of the high-dose-males and of the mid and high-dose females.
See Tables 7.8.1-20 to 7.8.1-21
Compound-related histomorphologic findings were present in the spleens of the high-dose rats of both sexes of the F3b generation, and were characterised by increased extramedullary haematopoiesis, pigment laden macrophages and congestion.
See Tables 7.8-22.
A variety of other spontaneous disease lesions and incidental findings were occasionally observed.

Reproductive effects observed:

not specified

Table 7.8.1-01: Summary of male rat mean body weight and food intake values for P1 generation growth period 

Week after study initiation	Dose level
	0 ppm		10 ppm		300 ppm		1000 ppm
	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)
0	169.5	-	181.4	-	181.0	-	180.5	-
1	223.0	156.3	235.8	160.9	236.4	153.8	232.0	149.6
2	269.1	172.9	283.9	177.6	282.3	176.1	286.0	168.8
3	303.4	181.5	318.9	171.3	315.0	177.1	317.0	172.1
4	333.4	169.6	346.3	176.6	341.9	185.1	342.1	172.6
5	354.8	171.1	372.3	181.6	363.8	182.3	367.5	174.0
6	371.9	166.0	395.5	185.9	385.1	171.5	392.1	171.4
7	386.1	188.8	406.3	193.0	397.3	189.8	405.3	185.0
8	401.3	182.0	422.4	203.1	409.6	185.4	411.9	185.6
9	415.1	171.5	439.9	198.0*	423.0	189.6	429.1	188.5

*p < 0.05%

 

Table 7.8.1-02: Summary of male rat mean body weight and food intake values for P1 generation growth period 

Week after study initiation	Dose level
	0 ppm		10 ppm		300 ppm		1000 ppm
	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)
0	142.4	-	148.0	-	145.6	-	152.6	-
1	160.0	109.0	168.3	115.4	166.8	107.4	174.9	111.5
2	176.1	116.9	184.9	123.1	185.9	117.7	190.4	129.4
3	187.5	121.3	196.3	115.3	196.1	117.8	200.3	129.6
4	198.0	120.0	205.3	123.2	207.7	123.6	210.6	134.0
5	201.6	120.8	212.4	128.6	215.5	136.8*	216.2	139.3
6	214.8	119.3	221.9	123.6	224.9	122.1	227.8	129.7
7	219.4	131.3	226.2	141.3	229.6	138.9	230.3	144.1
8	227.3	137.3	230.4	143.3	235.5	143.8	233.2	142.7
9	233.4	131.0	236.6	138.1	241.1	139.3	242.2	158.6*

 

Table 7.8.1-03: Summary of male rat mean body weight and food intake values for P2 generation growth period

Week after study initiation	Dose level
	0 ppm		10 ppm		300 ppm		1000 ppm
	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)
0	112.3	-	124.1	-	110.8	-	110.9	-
1	171.3	135.5	187.9	137.4	177.0	141.4	167.1	124.1
2	214.5	155.4	237.0	165.4	226.4	169.5	213.9	162.1
3	248.6	163.1	278.1	174.5	266.9	173.4	253.4	169.5
4	276.5	178.1	308.9	183.8	300.8	181.9	280.9	187.1
5	306.6	161.4	330.4	166.5	326.9	170.3	313.9	181.0
6	329.4	180.5	355.4	194.9	353.4	193.5	333.9	181.3
7	350.9	161.6	378.9	179.9	378.8	177.5	357.5	183.4
8	381.0	176.6	410.0	186.4	407.8	189.5	386.1	187.4
9	394.8	181.5	423.9	191.9	419.4	191.1	395.0	191.3

 

 

Table 7.8.1-04: Summary of female rat mean body weight and food intake values for P2 generation growth period 

Week after study initiation	Dose level
	0 ppm		10 ppm		300 ppm		1000 ppm
	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)
0	90.8		114.3		98.7		100.1
1	139.6	111.5	160.7*	126.7	145.3	111.1	140.5	111.3
2	163.2	132.1	182.8*	149.4*	169.3	141.9	166.6	140.8
3	182.4	131.6	200.4	150.8	184.5	148.7	179.9	146.4
4	193.4	157.6	209.3	181.9	196.1	195.9	189.6	183.1
5	207.9	131.3	227.3*	141.8	211.4	167.3*	202.4	168.4*
6	213.4	154.9	233.2	166.1	221.4	174.2	211.9	161.8
7	225.1	146.0	244.9	142.4	229.1	145.5	218.5	142.2
8	236.3	134.4	254.6	147.9	237.2	144.5	225.4	141.8
9	240.3	141.8	262.4	156.9	245.6	157.5	233.6	150.9

*P < 0.05%

 

Table 7.8.1-05: Summary of male rat mean body weight and food intake values for P3 generation growth period

Week after study initiation	Dose level
	0 ppm		10 ppm		300 ppm		1000 ppm
	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)
0	130.6	-	136.9	-	122.8	-	122.8	-
1	202.5	147.8	206.0	171.8	199.6	165.9	191.9	154.9
2	254.5	164.4	257.3	168.3	257.8	164.3	249.1	166.4
3	283.6	155.8	292.8	178.5	290.5	182.9*	286.4	185.3*
4	318.1	163.1	329.4	180.5	324.9	178.4	327.0	182.1
5	344.8	168.6	359.6	173.9	350.5	177.5	346.0	187.6
6	370.6	165.6	384.3	180.4	377.3	187.9*	385.9	199.0
7	391.3	174.1	404.9	178.3	397.3	176.9	414.5	187.0
8	410.8	170.4	424.5	177.3	415.9	178.8	436.0	191.3
9	431.1	176.4	443.5	183.3	429.0	179.8	454.9	197.4

*P < 0.05%

 

Table 7.8.1-06: Summary of female rat mean body weight and food intake values for P3 generation growth period

Week after study initiation	Dose level
	0 ppm		10 ppm		300 ppm		1000 ppm
	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)	Body weight (g)	Food intake (g/week)
0	115.9	-	112.8	-	97.2*	-	101.1	-
1	158.8	127.2	165.8	136.1	147.1	127.5	148.3	123.5
2	186.3	116.1	192.1	127.6	176.3	119.9	175.3	115.9
3	197.4	121.9	206.2	137.7*	192.7	139.3*	186.5	129.8
4	213.3	140.9	233.8	141.9	208.8	155.3	204.6	144.1
5	224.1	127.6	237.1	137.6	219.8	144.9*	216.6	144.7*
6	234.4	127.4	251.1	138.7	227.4	133.2	229.0	141.3
7	241.4	128.6	259.8	144.4	238.2	131.5	239.1	133.1
8	249.8	132.7	266.7	148.7*	245.8	130.7	243.8	136.1
9	258.4	135.8	275.5	147.2	252.4	142.3	252.0	143.8

*P < 0.05%

for further tables of results (7.8.1 -07 to 7.8.1 -22) look at attached background material

 

Conclusions:

Under the conditions of the study the administration of fluometuron technical to rats at dose levels of 300 and 1000 ppm resulted in treatment related morphometric and histomorphologic changes in the spleen. The no observed effect level (NOEL) for the study is proposed to be 10 ppm equivalent to 0.8 and 0.9 mg/kg bw/day for male and female rats respectively
A NOEL was not achieved in terms of maternal reproduction indices, postnatal offspring survival and growth under the conditions of the study.

Executive summary:

This report presents the findings from a three-generation reproduction study of Fluometuron Technical in albino rats when administered continuously in the diet at levels of 0, 10, 300, and 1000 ppm (Groups 1, 2, 3, 4; control, low-, mid-, and high-dose, respectively). Eight males and sixteen females per group were used for each parental generation (P₁, P₂and P₃). The three reproduction cycles consisted of two full-term litter generations (F_1a, F_1b; F_2a, F_2b; and F_3a, F_3b). The P₂and P₃animals were selected from the F_1band F_2blitters, respectively.

Criteria evaluated for compound effect in the animals were parental mortality, clinical signs, body weight and food consumption data (during growth and gestation periods), gross and microscopic pathology and organ/body weight data; reproduction indices; full-term offspring survival, growth, clinical signs, and sex ratios; and F_3bgross. and microscopic pathology and organ/body weight data.

No treatment-related effects were noted with respect to parental survival, clinical observations, maternal body weight, or food consumption values during gestation.

Evaluation of the mean body weights during the growth period of each generation revealed generally slightly higher than control values in the P₁and P₂male and female values (low- and mid-dose, groups only) and P₃male values.· The P₁mean body weights, however, were also slightly higher at Week 0 in the treated groups than in the control groups. Significantly higher than control values were noteed in the mean body weight values of the P₂low-dose females at Weeks 1, 2, and 5. In the P₃females the low-dose values were generally higher than control values, and the mid- and high-dose values were lower than control values (significantly lower at Week 0 in the mid-dose group).

 Food consumption values of the treated groups in each parental generation during growth were generally slightly higher than respective control values, with several significantly higher than control values noted at various weeks.

Enlarged, dark red, and/or dark brown spleens were the most common necropsy observations noted in the parental animals. Enlarged spleens were noted in greater incidence in the high-dose P₁female

and the P3 treated males and females than in the respective control groups. Dark red or dark brown spleens ware also noted in the high-dose P₂females in greater incidence than in the respective control group.

Evaluation of the organ weights and the organ/body weight ratios revealed a treatment-related increase in the mean absolute and relative spleen weights in each parental generation. Significantly higher than respective control values were noted in the following spleen weights: P₁females - mean absolute spleen weight in the mid- and high-dose groups (by log₁₀transformation) and the mean relative spleen weight in the high-dose group (by log₁₀transformation); P₂males - mean absolute spleen weight in all treated groups and mean relative spleen weight in the high-dose group; P₂females - mean absolute spleen weight in the mid- and high-dose groups am mean relative spleen weight in the mid- and high-dose groups (by log₁₀transformation); P₃males – mean absolute and relative spleen weights in the high-dose group; P₃-females - mean absolute spleen weight in the high-dose group (by log₁₀transformation) and mean relative spleen weight in the mid- and high-dose groups (by log₁₀transformation). Occasional significant differences from control values were noted in organs and ratios other than the spleen, but are not thought to be compound-related.

Similar findings in the histopathological evaluation were noted in the three parental generations. In the P₁animals, Fluometuron Technical at a dietary level of 1000 ppm (high-dose) resulted in increased extramedullary hematopoiesis and hemosiderin-laden macrophages in the spleen of high-dose rats of both sexes and a dietary level of 300 ppm (mid-dose) resulted in slightly increased extramedullary hematopoiesis and hemosiderin-Iaden macrophages in the mid-dose females. Spleen sections of the low-dose rats and mid-dose males were essentially camparable to the control.

In the P₂animals, Fluometuron Technical at dietary levels of 300 (mid-dose) and 1000 ppm (high-dose) resulted in changes in the spleen consisting of increased extramedullary hematopoiesis in high-dose rats of both sexes and mid-dose females, increased pigment-(hemosiderin) laden macrophages in the high- and mid-dose rats of both sexes and congestion of the spleen in nine of ten high-dose rats and one mid-dose female.

In the P₃animals, Fluometuron Technical at dietary levels of 300 (mid-dose) and 1000 ppm (high-dose) resulted in changes in the spleen of both sexes, characterized by increased extramedullary hematopoiesis and pigment-laden macrophages.

Spontaneous disease lesions and incidental findings were essentially comparable in incidence and severity in the control and treated rat of each parental generation.

Evaluation of the reproduction indices revealed no treatment-related trends through the three generations; and although some slight differences did occur between the control and treated values, no statistical differences were noted.

 The lactation survival indices, the mean neonatal survival indices, and the mean weaning survival indices of the P1-F_1bgeneration revealed a possible treatment-related decrease in survival. Except for the mean neonatal index of the low-dose group, the treated groups showed a decrease from control in each case, with the high-dose group showing a significant decrease in the mean weaning survival index when compared to that of the control group.

Decreases in the survival indices were also noted in the P₃-F_3bgeneration, but the decrease was noted in the control as well as the treated groups. The survival indices of the P1-F_1a, P₂, and P₃generations did not reflect any abnormal or consistent differences between the respective control and treated groups.

Significantly lower than control values ware noted in the P₁-F_1asex ratios on Days 1 and 5 (pre- and postcull) and in the P₁-F_1bsex ratio on Day 1; however, the sex ratios of each filial generation demonstrated normal biological variation. No consistent or treatment-related effects were noted in the offspring body weight data. Lower than control values were generally noted in the P₁-F_1afemale and P₁-F_1bmale and female body weight values; and higher than control values (including significantly higher values in the P₂-F_2alow-dose males and females on Day 21, and in the P₃-P_3ahigh-dose males on Day 21) were noted occasionally in the P₂and P₃filial generations.

Offspring clinical observations revealed no apparent treatment-related effects. Three P₂mid-dose male pups (two F_2aand one F_2bfrom the same mother) had similar observations of a dome-shaped head. The F_2apups were also small, weak, and appeared to be under stress. Grass pathology observations of the F_2aheads included doming of the skull, a thin layer of brain tissue, and a considerable amount of fluid filling the cavity. The medulla portion of the brain was recognizable, but the cerebral hemispheres were extremely dilated with fluid. These findings appear to be natural occurrences and are not thought to be compound-related.

Offspring grass pathology (P₃-F_3b) findings appearing to be treatment-related included enlarged spleens (noted in five high-dose males and one high-dose female of the ten animals/sex/ group that were examined) and dark or dark red spleens (noted in three high-dose males and one high-dose female).

Evaluation of the offspring (P₃-F_3b) organ weights and the organ/body weight ratios revealed a treatment-related increase in the mean absolute and relative spleen weights in the male and females which is consistent with the parental finding. Significantly higher than respective control values were noted in the offspring mean absolute spleen weights of the high-dose males and females and in the mean relative spleen weights of the high-dose males (by log₁₀transformation) and of the mid- and high-dose females (by log10 transformation).

Fluometuron Technical at dietary levels of 300 and 1000 ppm resulted in changes in the spleens of rats of both sexes of the F_3bgeneration. These changes included increased extramedullary hematopoiesis and pigment-laden macrophages.

Spontaneous disease lesions and incidental findings were essentially comparable in incidence and severity in the control and treated rats.

It is concluded from the data presented in this report, that administration of Flumeturon Technical to rats under conditions herein described and at dose levels of 300 and 1000 ppm, resulted in treatment related morphometric and histomorphologic changes in the spleen. Therefore, in terms of toxicity, a no effect level with Fluometuron Technical was achieved in this study at 10 ppm. It should be noted however, that in terms of maternal reproduction indices, postnatal offspring survival and growth, an effect level with Fluometuron Technical was not achieved in this study.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

0.8 mg/kg bw/day

Study duration:

subacute

Species:

rat

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

The effects of fluometuron on reproduction were evaluated in a rat three-generation toxicity study with two litters per generation.

In the three generation study with two litters per generation, treatment (by diet with 0, 10, 30 and 1000 ppm equivalent to 0.8, 23.8 and > 85 mg/kg bw/day in males and 0.9, 32 and >100 mg/kg bw/day in females) was for three generations (P1, P2 and P3). The first litters (F1a, F2a and F3a) were killed at weaning on Day 21 post partum whilst the second litters (F1b, F2b and F3b) were used to derive the P2 and P3 generations, respectively. Treatment was continuous from initiation of the P1 generation to necropsy of the last surviving animals (P3 parental animals and F3b offspring). The P1, P2 and P3 generations were fed for 9 weeks before pairing. The spleen was the target organ. Spleen weights were increased in both sexes of all generations at the highest dose level, in both sexes of P2 generation and in P3 females fed 32 mg/kg/day. In addition increased extramedullary haematopoiesis and haemosiderin-laden macrophage was observed in the spleen in the mid and high dose groups in the P1, P2 and P3 generations, and in the spleens of male and female weanlings from the F3b generation fed > 100 mg/kg/day. The increase in haemosiderin is probably due to haemolysis. Evaluation of the reproduction indices, lactation, neonatal survival and weaning survival showed slight differences between the control and treated groups however none of the changes were considered statistically significant.

The No Observed Effect Levels (NOELs) for parental toxicity were 0.8 and 0.9 mg/kg/day for males and females respectively and for reproductive toxicity were > 85 and 100 mg/kg/day for males and females respectively.

The principal findings in the study are summarised below:

Type of study (route of administration)	Species	Dose levels (mg/kg/day)	NOAEL/NOEL	LOEL/LOAEL	Reference
Three-generation (dietary).	Rat	0, 10, 300 or 1000 ppm	NOEL   Parental toxicity 10 ppm ≈ to 0.8 and  0.9 mg/kg bw/day for males and females respectively Reproductive toxicity > 1000 ppm ≈ > 85 and 100 mg/kg bw/day for males and females respectively	LOEL   Parental toxicity 300 ppm ≈ 23.8 and 32.0 mg/kg bw/day for males and females respectively   Increased mean absolute spleen weights in P1 and P2 in mid and high dose females; P3 females high dose group; P2 males all treated groups and P3 males high dose group. Increased mean relative spleen weights in P1, P2 and P3 high dose group females; P2, P3 high dose males as well as P2 and P3 mid dose group females. In the spleen increased extramedullary haematopoiesis and haemosiderin-laden macrophage in mid and high dose groups in P1, P2 and P3 generations, and also spleen of male and female weanlings from the F3b generation	Burdock, 1982 (903176V1-2)

Effects on developmental toxicity

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

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

Administration of fluometuron to rats at dose levels of 300 and 1000 ppm resulted in treatment related morphometric and histomorphologic changes in the spleen. The no observed effect level (NOEL) for the study is proposed to be 10 ppm equivalent to 0.8 and 0.9 mg/kg bw/day for male and female rats respectively.

A NOEL was not achieved in terms of maternal reproduction indices, postnatal offspring survival and growth under the conditions of the study.

In respect to regulation 1272/2008, Annex 1, 3.7.2 Fluometuron is not classified as being toxic to reproduction.