Reaction mass of 1-O-α-D-glucopyranosyl-D-fructose and 6-O-α-D-glucopyranosyl-D-fructose and fructose and glucose and sucrose

Toxicity to reproduction

Administrative data

Endpoint:

multi-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: Acceptable, well documented publication/study report which meets basic scientific principles

Data source

Reference

Reference Type:

publication

Title:

MULTIGENERATION REPRODUCTION STUDY OF ISOMALT IN RATS

Author:

Waalkens-Berendsen, D.H. et al.

Year:

1990

Bibliographic source:

Fd Chem. Toxic. 28(1):11-19

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Rats of both sexes were fed with 10% sucrose (test group) or 10% maize starch (control group) throughout 3 successive generations. For each generation, two litters were reared until they were at least 3 weeks old. From the second litter of the F3 generation, 10 animals were used for autospy and histopathology at the age of 4 weeks.

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Sucrose

EC Number:

200-334-9

EC Name:

Sucrose

Cas Number:

57-50-1

Molecular formula:

C12H22O11

IUPAC Name:

beta-D-fructofuranosyl alpha-D-glucopyranoside

Details on test material:

- Name of test material (as cited in study report): sucrose
- Analytical purity: no data

Test animals

Species:

rat

Strain:

other: Cpb:WU (Wistar Random)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Central Institute for the Breeding of Laboratory Animals TNO, Zeist, The Netherlands
- Age at study initiation: newly weaned rats, not further specified
- Housing: 5 animals of the same sex per cage in steel-cages with wire-mesh fronts and floors (premating periods)
- Diet: Institute´s powdered cereal based open-formula diet supplemented with 2.5% casein, 0.1% D,L-methionine and 10% maize starch (control groups) or 10% sucrose (test groups), ad libitum
- Water: ad libitum
- Acclimation period: 9 - 16 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 ± 1
- Humidity (%): approx. 50%
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Mixing appropriate amounts with: stock diet

Details on mating procedure:

- M/F ratio per cage: 1/1
- Length of cohabitation: 3 weeks
- After 1 week, the first male was replaced by another male so that three different males were available for each female.
- After successful mating each pregnant female was caged: individually

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

(P) Males and Females: 12 and 21 weeks before the 1st and 2nd mating, respectively and throughout the study
(F1): Selected rats were kept on the control or test diet of their parents for 12 and 21 weeks before the 1st and 2nd mating, respectively, and throughout the study.
(F2): Selected rats were kept on the control or test diet of their parents for 12 and 21 weeks before the 1st and 2nd mating, respectively, and throughout the study.

Frequency of treatment:

continous

Details on study schedule:

- F1 parental animals not mated until 15 weeks after selected from the F1 litters.
- Selection of parents from F1 generation when pups were at weaning age (not further specified).
- Age at mating of the mated animals in the study: 24 weeks

- F2 parental animals not mated until 15 weeks after selected from the F2 litters.
- Selection of parents from F2 generation when pups were at weaning age (not further specified).
- Age at mating of the mated animals in the study: 24 weeks

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
10%
Basis:
nominal in diet

Doses / concentrations 2

Remarks:

Doses / Concentrations:
P: 7.5 g/kg bw/day (male), 8.0 g/kg bw/day (female);
Basis:
other: mean dose value calculated from the reported body weight and food intake values (see attached Table 1)

Doses / concentrations 3

Remarks:

Doses / Concentrations:
F1: 8.4 g/kg bw/day (male), 9.0 g/kg bw/day (female)
Basis:
other: mean dose value calculated from the reported body weight and food intake values (see attached Table 1)

Doses / concentrations 4

Remarks:

Doses / Concentrations:
F2: 8.5 g/kg bw/day (male), 8.9 g/kg bw/day (female)
Basis:
other: mean dose value calculated from the reported body weight and food intake values (see attached Table 1)

Doses / concentrations 5

Remarks:

Doses / Concentrations:
F3: 12 g/kg bw
Basis:
other: mean dose value calculated from the reported body weight and food intake values (see attached Table 1)

No. of animals per sex per dose:

100 P males and females
20 F1 males and females
20 F2 males and females
10 F3 males and femals

Control animals:

other: yes, diet supplemented with 10% maize starch

Details on study design:

- Other:
Study design:
The P generation consisted of 100 males and females in the control and test group. After 12 weeks on the test and control diets, the first mating occurred. Pups were weaned at day 21 and 50 males and females were used for a 8 week feeding study. The remaining pups were discarded at 3 – 5 weeks of age. The mating procedure was repeated 9 weeks after the first mating by using the same combination of mating partners. From this F1 generation, 20 males and females were used to produce the F2 generation. Therefore, the selected rats were weaned at day 21 and kept on the diets of their parents for further 12 weeks before mating. The offspring of the first mating was discarded and 20 females and males of the second mating were used for the production of the F3 generation. As with the F1-generation, the selected F2 generation rats were fed with the various diets for 12 weeks after weaning and subsequently mated. After 21 weeks of exposure, the second mating procedure started. Pups from the first F3 offsping were discarded 3 weeks after birth. Weanlings from the second mating procedure were used for gross and microscopic evaluation after receiving the diets for 4 weeks (10/sex).

Examinations

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes, daily

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were recorded weekly during the premating and mating period. No weighing was performed during pregnancy and lactation periods.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: NO
- Food consumption for each animal determined as g food/rat/week: YES
For each generation, food consumption was recorded weekly during the premating period of 12 weeks. Food intake was not measured during the mating and lactation periods.

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 1 postpartum: yes
- If yes, maximum of 8 pups/litter.

PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2 / F3] offspring:
number and sex of pups, live births, postnatal mortality, presence of gross anomalies, weight gain, other: viability index at birth and day 4, litter size
F3: haemoglobin level

Postmortem examinations (offspring):

SACRIFICE
- The F1 and F2 offspring not selected as parental animals were sacrificed when they were 3 weeks old.
- The F3 offspring of the first mating was sacrificed at the age of 3 weeks whereas 10 weanlings of the second mating were sacrificed 4 weeks after weaning (males: 28 days after weaning, females: 29 days after weaning) for detailed gross and microscopic examinations.

GROSS NECROPSY
- The following organs were weighed: adrenals, brain, caecum, heart, kidneys, liver, ovaries, spleen, testes, thymus and thyroid.

HISTOPATHOLOGY / ORGAN WEIGTHS
Tissues of all the major systems from the F3 generation including kidneys, liver, lungs, heart, stomach, epididymides, prostate, urinary bladder, pituitary, thyroid were examined microscopically.

Statistics:

Mean values and standard errors were calculated from the examined parameters. Statistical analyses were performed using ANOVA and Dunnett´s test (changes in body weights, food intake, haemoglobin levels and relative organ weights), Student´s t test (mean litter size at birth and pup weight) or chi-square test (viability and lactation indices).

Reproductive indices:

The following parameter were determined in the P, F1 and F2 generation for the first and second mating:
1. fertility index (%): (no. of pregnant females / no. of mated females) x 100
2. gestation index (%): (no. of females with live fetuses / no. of pregnant females) x 100
3. lactation index: (no. of live pups at day 21 / no. of live pups at day 4)

Offspring viability indices:

viability index on day 1 (%): (no. of pups born alive / total no. of born pups) x 100
viability index on day 4 (%) : (no. of live pups at day 4 / total no. of live pups on day 1) x 100%

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

7 out of 100 females died due to delivery problems (non-adverse)

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

F1 females: decreased body weight on day 21; F2 males: increased body weight on day 21 (non-adverse)

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

F1 females: decreased body weight on day 21; F2 males: increased body weight on day 21 (non-adverse)

Organ weight findings including organ / body weight ratios:

not examined

Histopathological findings: non-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):

The fertility index of parental dams was significantly reduced in the first and second mating to 82% and 92% of controls, respectively. As the effect was not reproducible in the F1 and F2 matings, the effect is not considered as treatment-related.

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
No abnormalities of condition or behaviour were observed. As the mortality of 7 out of 100 animals is due to delivery problems, the effect on mortality is considered as not-treatment related.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
A significant but slight decrease in body weight was observed in F1 females at day 21 (control group: 143 ± 1.9; test group: 138.5 ± 2.6) whereas a significant increase in body weight was determined in F2 males (control group: 170.0 ± 2.9; test group: 179.8 ± 3.6). As the effect was not consistent over the observation period and not reproducible in all generations, the effect is not considered to be treatment-related.

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS)
No substance-related changes were observed in food intake.

GROSS PATHOLOGY (PARENTAL ANIMALS)
Gross pathology did not reveal any alterations in the parental animals.

Effect levels (P0)

1

Dose descriptor:

NOAEL

Effect level:

>= 7 455 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: mortality; body weight; food consumption and compound intake; gross pathology; organ weights; histopathology; mating index; fertility index; pregnancy index; pup weight; viability index; lactation index; corresponding to 10%

2

Dose descriptor:

NOAEL

Effect level:

>= 7 998 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: mortality; body weight; food consumption and compound intake; gross pathology; organ weights; histopathology; mating index; fertility index; pregnancy index; pup weight; viability index; lactation index; corresponding to 10%

3

Dose descriptor:

NOAEL

Effect level:

>= 11 816 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: mortality; body weight; food consumption and compound intake; gross pathology; organ weights; histopathology; mating index; fertility index; pregnancy index; pup weight; viability index; lactation index; corresponding to 10%

Remarks on result:

other: Generation: F3 (migrated information)

4

Dose descriptor:

NOAEL

Effect level:

>= 12 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: mortality; body weight; food consumption and compound intake; gross pathology; organ weights; histopathology; mating index; fertility index; pregnancy index; pup weight; viability index; lactation index; corresponding to 10%

Remarks on result:

other: Generation: F3 (migrated information)

Results: F1 generation

General toxicity (F1)

Clinical signs:

no effects observed

Mortality / viability:

mortality observed, treatment-related

Description (incidence and severity):

Significant differences in pup viability were observed (non-adverse).

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Increases in body weights during lactation and after weaning in pups from F0 and F1 (non-adverse).

Sexual maturation:

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Male F3 pups showed a statistically significant increase in relative kidney weight (non-adverse).

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Different abnormalities were observed in one pup each of the F1, F2 and F3 generation (non-adverse).

Histopathological findings:

no effects observed

Details on results (F1)

VIABILITY (OFFSPRING)
Viability index: Several statistically significant differences in pup survival indices were determined including both, increases and decreases in the test group. As the differences from the controls were only small and never appeared to be consistent in successive generations, the effects are considered as non-adverse.
Lactation index: After the 1st and 2nd mating of the P generation, the survival of pups during lactation was generally lower in all groups including controls than in breedings of the following generations. No treatment-related alterations were determined.

CLINICAL SIGNS (OFFSPRING)
No signs of toxicity were observed throughout the study.

BODY WEIGHT (OFFSPRING)
Effects during lactation: Pups from the first mating of the F0 generation showed a significant higher body weight on day 4. Further, pups from both matings of the F1 generation showed a significant higher body weight at day 14 (first mating) and at days 4, 14 and 21 (second mating). In contrast, no alterations were determined in pups from the F2 generation. As the effects are not consistent through the litters and the generations, no effect on body weight of sucrose seems to be apparent during lactation.

Effects after lactation and exposure via feed: Females of the F1 and males of F2 generation showed a significant higher body weight on day 21. As the effect is not consistent in the successive generations, the effect is considered as non-adverse.

SEXUAL MATURATION (OFFSPRING)
No effects on sexual maturation were observed in exposed animals.

ORGAN WEIGHTS (OFFSPRING)
A statistically significant increase in the relative kidney weight was observed in male F3 pups which was not accompanied by histological changes. Thus, the effect is not interpreted as adverse.

GROSS PATHOLOGY (OFFSPRING)
Malformations were observed in one F1 pup (anophthalmia of one eye), one F2 pup (thick and paralysed hind leg) and one F3 pup (missing foot, which was most probably amputated shortly after birth). All these abnormalities were considered fortuitous and not treatment-related.

Effect levels (F1)

1

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

>= 8 377 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: mortality; body weight; food consumption and compound intake; gross pathology; organ weights; histopathology; mating index; fertility index; pregnancy index; pup weight; viability index; lactation index; corresponding to 10%

2

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

>= 8 997 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: mortality; body weight; food consumption and compound intake; gross pathology; organ weights; histopathology; mating index; fertility index; pregnancy index; pup weight; viability index; lactation index; corresponding to 10%

Results: F2 generation

Effect levels (F2)

1

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

>= 8 475 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: mortality; body weight; food consumption and compound intake; gross pathology; organ weights; histopathology; mating index; fertility index; pregnancy index; pup weight; viability index; lactation index; corresponding to 10%

2

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

>= 8 892 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: mortality; body weight; food consumption and compound intake; gross pathology; organ weights; histopathology; mating index; fertility index; pregnancy index; pup weight; viability index; lactation index; corresponding to 10%

Overall reproductive toxicity

Reproductive effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

In the conducted study, exposure to 10% sucrose of 3 successive generations did not induce adverse effects on fertility, reproductive performance or development compared with control animals fed with 10% maize starch. Therefore, sucrose is not considered to induce toxic effects on reproduction.