Dehydrated sorbitol, C18 (unsaturated) fatty acid esters, ethoxylated

Toxicity to reproduction

Administrative data

Endpoint:

three-generation reproductive toxicity

Remarks:

based on test type (migrated information)

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Basic data given (no data on test substance purity, limited evaluation and documentation of parameters).

Data source

Reference

Reference 1

Reference Type:

publication

Title:

NUTRITIONAL STUDIES ON RATS OF DIETS CONTAINING HIGH LEVELS OF PARTIAL ESTER EMULSIFIERS. II. REPRODUCTION AND LACTATION

Author:

Oser, B.L. and Oser, M.

Year:

1956

Bibliographic source:

J Nutr. 60(4): 489-505

Reference 2

Reference Type:

publication

Title:

NUTRITIONAL STUDIES ON RATS OF DIETS CONTAINING HIGH LEVELS OF PARTIAL ESTER EMULSIFIERS. I. GENERAL PLAN AND PROCEDURES; GROWTH AND FOOD UTILIZATION.

Author:

Oser, B.L. and Oser, M.

Year:

1956

Bibliographic source:

J Nutr. 60(3): 367-390

Reference 3

Reference Type:

publication

Title:

NUTRITIONAL STUDIES ON RATS OF DIETS CONTAINING HIGH LEVELS OF PARTIAL ESTER EMULSIFIERS. III. CLINICAL AND METABOLIC OBSERVATIONS

Author:

Oser, B.L. and Oser, M.

Year:

1956

Bibliographic source:

J Nutr. 61(1): 149-166

Reference 4

Reference Type:

publication

Title:

NUTRITIONAL STUDIES ON RATS OF DIETS CONTAINING HIGH LEVELS OF PARTIAL ESTER EMULSIFIERS. IV. MORTALITY AND POST-MORTEM PATHOLOGY; GENERAL CONCLUSIONS

Author:

Oser, B.L. and Oser, M.

Year:

1957

Bibliographic source:

J Nutr. 61(2): 235-252

Materials and methods

Principles of method if other than guideline:

Three-generation study with rats dosed for 12 weeks with the test substance prior to mating.

GLP compliance:

no

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Sorbitan monooleate, ethoxylated

EC Number:

500-019-9

EC Name:

Sorbitan monooleate, ethoxylated

Cas Number:

9005-65-6

Molecular formula:

Not applicable as UVCB

IUPAC Name:

Sorbitan, C18-unsat, fatty acid esters, ethoxylated (1 - 6.5 moles ethoxylated)

Constituent 2

Reference substance name:

Tween 80

IUPAC Name:

Tween 80

Details on test material:

- Name of test material (as cited in study report): Tween 80 (polyoxyethylene (20) sorbitan monooleate)
- Analytical purity: no data

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Food Research Laboratory stock colony, Long Island City, NY, USA
- Age at study initiation: (P) approx. 110 days
- Weight at study initiation: (P) Males > 310 g; (P) Females > 210 g
- Housing: for the first 12 weeks on the test diets, animals were housed individually in suspended-wire mesh cages. When matings were started, they were transferred to larger cages, one male being housed with one or two females. When litters were about to be cast, the dams were transferred to separate cages containing shredded paper wads for nesting where they remained until their litters were weaned. During rest period between matings, animals were housed in small groups of the same sex.
- Diet: basal diet prepared by laboratory, ad libitum
- Water: ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23.3-25.6
- Humidity (%): 40-60

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): the final diets were prepared at intervals of not more than two weeks by the addition the test substance and of the vitamin oil mixture. In any case completion of the diet batches by the addition of the test substance, and vitamin mixtures was accomplished about once a week so that no diets were more than 10 days old when fed.
- Mixing appropriate amounts with (Type of food): basal diet

Details on mating procedure:

- M/F ratio per cage: 1/2
- Length of cohabitation: three weeks
- Proof of pregnancy: by palpation or weight increment
- After 21-28 days of unsuccessful pairing replacement of first male by another male with proven fertility.
- After successful mating each pregnant female was caged: individually
- Any other deviations from standard protocol: matings continued in the F0 generation throughout the entire two-year period

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

F0 generation was treated 12 weeks prior to mating, then treatment was performed continuously over 2-years in all generations

Frequency of treatment:

daily, 7 days/week

Details on study schedule:

- F1 parental animals not mated until 1 week after selected from the F1 litters.
- Selection of parents from F1 generation when pups were 21 days of age.
- Age at mating of the mated animals in the study: 110 days
- Other: first F1 litter was discarded; females were allowed one-week rest period before remating; males were rotated among the females within their respective test groups.

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
5, 10 and 20%
Basis:
nominal in diet

Doses / concentrations 2

Remarks:

Doses / Concentrations:
3333, 6666 and 13333 mg/kg bw/day
Basis:
other: calculation based on the assumption of an average food consumption of 20 g/animal per day and an average body weight of 300 g

No. of animals per sex per dose:

F0 generation: 12 males and 20 females
F1, F2, F3 generation: 10 males and 10 females (selected from the second litter of each generation)

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: dose selection was based on the total estimated intake in humans of 0.75 mg/day or 0.125% of the diet. Thus, the lowest feeding level adopted for these investigations, 5%, represented 40 times the maximum human dietary level. Previously reported studies suggested that adverse physiological responses might be expected at the 20% level; hence this was selected as the highest feeding level. The intermediate level, 10%, is the logarithmic mean.

Examinations

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: no data
- Cage side observations included examination of physical appearance and behaviour.

BODY WEIGHT: Yes
- Time schedule for examinations: animals were weighed weekly for 12 weeks (also in each succeeding generation), and then biweekly for the remainder of the study.

FOOD CONSUMPTION: Yes
- Time schedule: records were made of food consumption of 5 rats of each sex, randomly chosen from each group, for the first 12 weeks. Additional checks on food consumption were made on these rats for a two-week period at the 0.5, 1, 1.5, and 2-year stages in the initial generation and for 12 weeks after weaning in succeeding generations.

FOOD EFFICIENCY:
- Body weight gain in g/100 g food consumption (efficiency of food utilisation = EFU) and body weight gain in g per 100 cal (efficiency of caloric utilisation = ECU): Yes

WATER CONSUMPTION: Yes
- Time schedule for examinations: during a two-week period

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no

PARAMETERS EXAMINED
The following parameters were examined in F1 / F2 / F3 offspring: all parameters like for F0 generation (see parental examinations) and number of pups, live births, postnatal mortality, weight at weaning, litter size

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: all surviving animals (except for the first litter of each generation) at the termination of the 2-year study as well as at death or sacrifice of moribund animals
- Maternal animals: all surviving animals (except for the first litter of each generation) at the termination of the 2-year study as well as at death or sacrifice of moribund animals

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated were prepared for microscopic examination and weighed: kidneys, liver, spleen, heart and adrenals of at least 6 animals per sex and dose were weighed, whereas all major organs (e.g. liver, kidneys, bladder, lungs, breast, stomach, gastrointestinal tract, heart, spleen, pancreas, thyroid, gonads, lymph nodes, bone marrow, spinal cord, skin and uterus) were placed in formalin for histopathological investigation. Histopathological examinations were conducted on the livers and kidneys of rats at all dosage levels. Other organs were examined microscopically where indicated by their gross appearance and in at least two apparently normal rats of each sex randomly selected from the 20% group.

Postmortem examinations (offspring):

SACRIFICE
- Male animals: all surviving animals at the termination of the 2-year study as well as at death or sacrifice of moribund animals
- Maternal animals: all surviving animals at the termination of the 2-year study as well as at death or sacrifice of moribund animals

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated were prepared for microscopic examination and weighed: kidneys, liver, spleen, heart and adrenals of at least 6 animals per sex and dose were weighed, whereas all major organs were placed in formalin for histopathological investigation.

The F3 generation was not investigated because the rats in this generation were sacrificed either at weaning or at the end of the 12-week growth period.

Statistics:

Statistical analysis of body weights was performed by the multiple range test of Duncan (1953).

REFERENCE:
DUNCAN, D. B. (1953). Multiple Range and Multiple F Tests. Technical Report no. 6a. Department of Statistics and Statistical Laboratory, Virginia Agricultural Experiment Station, Virginia Polytechnic Institute, Blacksburg, Va. (Sponsored by the Office of Ordnance Research, U. 8. Army)

Reproductive indices:

Fertility index (F.I).: the percentage of matings resulting in pregnancy ((pregnancies/matings)*100)
Gestation index (G.I.): the percentage of pregnancies resulting in the birth of live litters ((litters born/pregnancies)*100)

Offspring viability indices:

Viability index (V.l.): the percentage of rats born that survived 4 days or longer ((pups surviving at Day 4/pups born)*100)
Lactation index (L.I.): the percentage of rats alive at 4 days that survived the 21-day lactation period ((pups weaned/pups at Day 4)*100)

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

20% in the diet: most males showed mild to moderate diarrhoea accompanied by mild/moderate to marked perianal irritation; higher mortality (predominantly in females) compared to controls

Dermal irritation (if dermal study):

not examined

Mortality:

mortality observed, non-treatment-related

Description (incidence):

Mortality statistics indicated no evidence of untoward effect on longevity in any of the 5 or 10% test groups. Only occasional deaths occurred among the males in the test groups during the first year and the few deaths among the females occurred during or after parturition. During the final quarter of the two-year test more deaths occurred among the females on the 20% diets containing the test substance compared to treated males and controls. Whether this was the cumulative stress of the laxative effect or a more basic but obscure manifestation of the test substance remained to be determined. However, median survival time (MST), expressed as the age in weeks when one-half of a group had died, revealed that the MST of the treated animals of all dose groups values fell within the MST of the controls. Therefore, the MST was not affected by either the dietary level of the test substance, nor when compared with those of the controls.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

20% in the diet: reduction in body weights of males of the F0 generation at Week 12

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

20% in the diet: reduction in body weights of males of the F0 generation at Week 12

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

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not examined

Other effects:

no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

effects observed, treatment-related

Description (incidence and severity):

20% in the diet: reduced litter number and fertility index; slightly lower proportion of matings in F1 and F2 generation compared to the F0 generation

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
The external appearance (condition of fur, colour and vascularity of eyes as well as pinkness of the ears and toes) and behaviour (activity) of the rats of all test groups were normal throughout the two-year feeding test, except for those whose diarrheal condition resulted in inflammation and unthrifty appearance around the anal region due to laxative effects of the test substance at the highest dose level. The laxative action was mainly observed in male rats of all generations at the 20% level of the test substance, and to a lesser extent in animals (males and females) at the 10% level. Only in the animals with pronounced diarrhoea, an unthrifty appearance due to the moist stools was observed. Laxation was considered to be probably a cause of longer chain polyols resulting from hydrolysis of the test substance, which were also shown to induce laxation in rats at stoichiometrically equivalent doses to the test substance in the diet.
Mortality statistics indicated no evidence of untoward effect on longevity in any of the 5 or 10% test groups. Only occasional deaths occurred among the males in the test groups during the first year and the few deaths among the females occurred during or after parturition. During the final quarter of the two-year test more deaths occurred among the females on the 20% diets containing the test substance compared to treated males and controls. Whether this was the cumulative stress of the laxative effect or a more basic but obscure manifestation of the test substance remained to be determined. However, median survival time (MST), expressed as the age in weeks when one-half of a group had died, revealed that the MST of the treated animals of all dose groups values fell within the MST of the controls. Therefore, the MST was not affected by either the dietary level of the test substance, nor when compared with those of the controls.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
At 12 weeks, the period when rats normally reach approximately two-thirds of their maximum weight, a significant reduction (-19%) in weight gain was observed only in males treated with a dietary level of 20% of the test substance compared to controls. However, at the end of the study period, no significantly different change in body weight and body weight gain was observed in treated animals at all dose levels compared to the controls (see Table 1 under “Any other information on results incl. tables”).
At 12 weeks, no effects on food consumption were observed in treated animals of all dose group compared to controls (see Table 2 under “Any other information on results incl. tables”). The mean consumption of practically all diets of treated animals was somewhat higher in the descendent generations than in the initial generation, which however, was not correlated with any changes in weight gains of treated animals compared to controls. Regardless of the dose level, the efficiency of food utilization (EFU) as well as the efficiency of caloric utilization (ECU) were somewhat higher in the initial (F0) generation than in the succeeding generations. This may be explained by the difference in nutritional reserves at weaning between the first generation and all the descendent generations. However, EFU values in treated animals did not reach significance when compared to controls.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
In the F0 generation, at all dose levels, no effect on gestation was observed in the treated animals compared to controls (see Tables 3 and 4 under “Any other information on results incl. tables”). However, there was a tendency toward diminished productivity (in terms of numbers of litters) in groups receiving the test substance at the 20% level, which could be due to laxative effects of the test substance at this dose level resulting in lack of interest in copulation.
When the laxative action was of moderate or greater severity, it was commonly associated with irritation and hypersensitivity around the anus sufficient to warrant considering lack of interest in copulation as a possible explanation for the reduced fertility frequently observed in the highest dose group (20% of the test substance in the diet) after 6 matings (see Tables 4 under “Any other information on results incl. tables”).
Comparison of the reproduction data showed that the responses in the successive generations were quite similar (see Tables 3 and 4 under “Any other information on results incl. tables”). The proportion of matings resulting in pregnancy (fertility index) tended to be lower at the 20% dietary level of the test substance. Reproductive performance in general appeared to be inferior in the third generation (F2) rats compared to their progenitors.
The reproductive performance of the F3 generation was not investigated because the rats in this generation were sacrificed either at weaning or at the end of the 12-week growth period.

ORGAN WEIGHTS (PARENTAL ANIMALS)
No changes in relative liver and kidney weights were observed compared to controls. No details were reported on the organ weights of spleen, heart and adrenals.

GROSS PATHOLOGY (PARENTAL ANIMALS)
At gross examination, the lungs appeared to be the most frequently affected organ in the animals. However, neither the frequency, character, nor the degree of the pulmonary lesions were any more marked in the test animals than in the controls.

HISTOPATHOLOGY (PARENTAL ANIMALS)
The more frequent incidence of mammary fibromas in the animals of the 5 and 10% test groups (both 5/20 animals compared to 3/20 animals in the control group) was considered to be without significance in view of the lower incidence (only 1/20 animals) at the 20% dose group, indicating a lack in treatment-related response of this observation.
All histopathological changes observed in liver, kidney and bladder of treated animals were considered to be not treatment-related due to the limited and sporadic incidences among both the control and test groups and the absence of a clear dose-response relationship.
Furthermore, histopathological examination of the major organs and tissues examined in rats (including stomach, gastrointestinal tract, heart, spleen, pancreas, adrenals, thyroid, gonads, lymph nodes, bone marrow and spinal cord, skin and uterus) at the termination of the two-year feeding test, including tumours, mammary or otherwise, showed no lesions which by their frequency or nature could be attributable to treatment with the test substance.

OTHER FINDINGS (PARENTAL ANIMALS)
To determine whether there was any correlation between the degree of laxation, the moisture content of the stools and the voluntary water intake, these parameters were measured in control and all test animals during a two-week period within the study. The faecal moisture was higher in all treated animals suffering from diarrhoea, which was also accompanied by a higher water intake in these animals compared to controls.

Effect levels (P0)

1

Key result

Dose descriptor:

LOAEL

Remarks:

systemic

Effect level:

20 other: % in the diet

Based on:

test mat.

Remarks:

corresponding to 13333 mg/kg bw/day based on the assumption of an average food consumption of 20 g/animal and an average body weight of 300 g

Sex:

male/female

Basis for effect level:

other: most males showed mild to moderate diarrhoea accompanied by mild/moderate to marked perianal irritation; higher mortality in females

Remarks on result:

other: Generation: F0 (migrated information)

2

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Effect level:

10 other: % in diet

Based on:

test mat.

Remarks:

corresponding to 6666 mg/kg bw/day based on the assumption of an average food consumption of 20 g/animal and an average body weight of 300 g

Sex:

male/female

Basis for effect level:

other: overall effects; low incidence of diarrhoea in females

Remarks on result:

other: Generation: F0 (migrated information)

3

Key result

Dose descriptor:

LOAEL

Remarks:

fertility

Effect level:

20 other: % in diet

Based on:

test mat.

Remarks:

corresponding to 13333 mg/kg bw/day based on the assumption of an average food consumption of 20 g/animal and an average body weight of 300 g

Sex:

male/female

Basis for effect level:

other: slight reduction in fertility index (F0: after 6 matings; F1 and F2: after 2 matings)

Remarks on result:

other: Generation: F0, F1, F2 (migrated information)

4

Key result

Dose descriptor:

NOAEL

Remarks:

fertility

Effect level:

10 other: % in diet

Based on:

test mat.

Remarks:

corresponding to 6666 mg/kg bw/day based on the assumption of an average food consumption of 20 g/animal and an average body weight of 300 g

Sex:

male/female

Basis for effect level:

other: no significant effects on reproductive performance

Remarks on result:

other: Generation: F0, F1, F2 (migrated information)

5

Key result

Dose descriptor:

LOAEL

Remarks:

developmental

Effect level:

20 other: % in diet

Based on:

test mat.

Remarks:

corresponding to 13333 mg/kg bw/day based on the assumption of an average food consumption of 20 g/animal and an average body weight of 300 g

Sex:

male/female

Basis for effect level:

other: significant reduction in survival of newborn at 4 days post partum (reduced viability index); slight reduction in average pub weight at weaning (most likely due to reduced maternal care as result of diarrhoea)

Remarks on result:

other: Generation: F0, F1, F2 (migrated information)

6

Key result

Dose descriptor:

NOAEL

Remarks:

developmental

Effect level:

10 other: % in diet

Based on:

test mat.

Remarks:

corresponding to 6666 mg/kg bw/day based on the assumption of an average food consumption of 20 g/animal and an average body weight of 300 g

Sex:

male/female

Basis for effect level:

other: no significant effects on viability of newborn and weight of weaned pubs; no effects on lactation

Remarks on result:

other: Generation: F0, F1, F2 (migrated information)

Results: P1 (second parental generation)

General toxicity (P1)

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality:

not examined

Body weight and weight changes:

not examined

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

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Results: F1 generation

General toxicity (F1)

Clinical signs:

not examined

Dermal irritation (if dermal study):

not specified

Mortality / viability:

mortality observed, treatment-related

Description (incidence and severity):

20% in the diet: significant reduction in survival during days 0-4 post partum

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

20% in the diet: slightly lower pub weights at weaning (associated with reduced maternal care)

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

not specified

Histopathological findings:

not specified

Other effects:

not specified

Details on results (F1)

VIABILITY (OFFSPRING)
In all generations, the size of the litters was substantially reduced at weaning in all groups (treated and controls). From the relative magnitude of the values for the viability and lactation indices, it is apparent that the drop in litter size occurred principally during the few days immediately after birth (see Table 3 and 4 under “Any other information on results incl. tables”).
At the 20% dietary level, a significant reduction in survival of newborns (as shoen in a reduction of viability index) was observed in all generations after 2 matings (see Table 3 under “Any other information on results incl. tables”). The trend toward higher mortality during the 4 days post partum was not as marked in the F1 and F2 as in the F0 generation after 6 matings (see Table 4 under “Any other information on results incl. tables”).

BODY WEIGHT (OFFSPRING)
At the 20% dietary level, slightly lower pub weights at weaning in all generations was observed compared to controls which is normally associated with a possible impairment of lactation. However, neglect of the litters by maternal animals was rather associated with this effect than lactation failure per se, since no significant effects on the lactation indices were observed at this dose level compared to controls. The neglect of litters by maternal animals was considered to be a result of the laxative effect of the test substance at 20% in the diet, resulting in posterior ventral irritation that may have caused adverse influence on the interest of dams in caring for their offspring.

Effect levels (F1)

Key result

Dose descriptor:

NOAEC

Generation:

F1

Effect level:

20 other: %w/w

Based on:

test mat.

Sex:

not specified

Basis for effect level:

body weight and weight gain

Overall reproductive toxicity

Reproductive effects observed:

not specified

Any other information on results incl. tables

Table 1. Body weights of the F0 generation

Dose	Sex	Number of initial animals	Average body weights in g (number of survivors)
			Week 0	Week 3	Week 6	Week 12	Week 24	Week 48	Week 52	Week 96	Week 104
Control	Male	12	57	175	260	326	371	420	417	446 (7)	431 (7)
	Female	20	56	139	176	209	262	307	301	329 (13)	315 (12)
5%	Male	12	56	170	246	310	354 (11)	393 (11)	383 (11)	412 (6)	423 (5)
	Female	20	54	140	179	210	260 (18)	303 (17)	311 (17)	317 (14)	311 (8)
10%	Male	12	58	159	242	305 (10)	375 (10)	409 (10)	414 (9)	438 (8)	387 (3)
	Female	20	57	137	177	209	270 (19)	325 (18)	306 (18)	332 (12)	315 (10)
20%	Male	12	58	141	205	277	351 (10)	388 (8)	388 (8)	426 (6)	422 (6)
	Female	20	57	117	166	200	270 (19)	300 (14)	307 (14)	332 (8)	340 (6)

 

Tabel 2.Food intake and efficiency of four generations of rats (5 of each per sex per group) during the 12-week feeding periods on the test substance

Dose	Generation	Average body weight gain (g)	Average food intake (g)	Efficiency of food utilisation (EFU) (g/100 g food)	Efficiency of calorie utilisation (ECU) (g/100 cal)
Control	F0	201	1248	16.1	4.71
	F1	210	1333	15.8	4.62
	F2	202	1342	15.1	4.42
	F3	226	1394	16.2	4.74
5%	F0	218	1271	17.2	5.12
	F1	198	1256	15.8	4.70
	F2	203	1398	14.5	4.32
	F3	204	1462	14.0	4.17
10%	F0	196	1130	17.3	5.25
	F1	214	1391	15.4	4.67
	F2	202	1379	14.6	4.43
	F3	224	1426	15.7	4.76
20%	F0	190	1243	15.3	4.79
	F1	229	1535	14.9	4.69
	F2	211	1429	14.8	4.65
	F3	235	1681	14.0	4.40

 

Table 3. Summary of reproduction and lactation data of F0, F1 and F2 generation rats for two matings

Dose	Generation	Number of matings	Litters born alive	Pups		Number pups per litter		Average weight of pubs at weaning	Fertility index	Gestation index	Viability index	Lactation index
				Born alive	Weaned	Born alive	Weaned
Control	F0	40	31	276	215	8.9	6.9	41.1	83	94	82	95
	F1	20	18	182	107	10.1	6.0	40.9	90	100	86	69
	F2	19	15	138	101	9.2	6.7	39.1	84	94	84	87
5%	F0	38	28	274	225	9.8	8.0	37.5	76	97	89	92
	F1	20	15	149	126	10.0	8.4	37.7	75	100	93	91
	F2	20	13	126	72	9.7	5.5	35.8	65	100	70	82
10%	F0	39	32	315	242	9.9	7.6	39.0	82	100	79	98
	F1	18	18	191	144	10.6	8.0	35.2	90	100	87	87
	F2	19	13	101	27	7.8	2.1	38.5	68	100	45	60
20%	F0	38	31	308	182	9.9	5.9	37.1	82	100	65	91
	F1	20	12	133	87	11.0	7.2	32.3	60	100	71	92
	F2	20	7	72	44	10.3	6.3	32.0	40	88	64	96

Table 4. Summary of reproduction and lactation data for F0 generation rats for 6 matings

Dose	Generation	Number of matings	Litters born alive	Pups born alive	Average number of pups/litter		Average weight of pubs at weaning	Fertility index	Gestation index	Viability index	Lactation index
					Born alive	Weaned
Control	F0	119	76	654	8.6	6.5	43.3	66	98	83	91
5%	F0	106	75	644	8.6	5.9	41.3	73	97	75	92
10%	F0	112	68	617	9.1	6.6	41.0	63	97	75	96
20%	F0	98	57	504	8.9	4.9	36.1	61	95	55	100