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

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

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: Comparable to guideline study with acceptable restrictions.

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

1997

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

6-O-α-D-glucopyranosyl-D-fructose

EC Number:

237-282-1

EC Name:

6-O-α-D-glucopyranosyl-D-fructose

Cas Number:

13718-94-0

Molecular formula:

C12H22O11

IUPAC Name:

6-O-alpha-D-glucopyranosyl-D-fructose

Details on test material:

- Name of test material (as cited in study report): Isomaltulose (6-O(α-D-glucopyranosyl)-D-fructofuranose)
- Physical state: white crystalline powder
- Analytical purity: 98.5%

Test animals

Species:

rat

Strain:

other: Wistar (Hsd/Cpb:WU)

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan-CPB, Zeist, The Netherlands
- Age at study initiation (at mating): 13 weeks
- Housing: mated females were housed individually in suspended stainless-steel wire-screen cages
- Diet: institute's cereal-based powdered stock diet (during treatment period supplemented with 2.5% casein, 0.1% DL-methionine and 0-10% maize starch), ad libitum
- Water: ad libitum
- Acclimation period: 5 days before mating

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22-24
- Humidity (%): 50-60
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

DIET PREPARATION
- Mixing appropriate amounts with (Type of food): institute's cereal-based powdered stock diet supplemented with 2.5% casein, 0.1% DL-methionine and 10% maize starch; 0, 2.5, 5 or 10% test substance was incorporated at the expense of maize starch.
- Storage temperature of food: freezer

Fresh portions of diet were provided to the rats every 4-6 days.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

All diets were analysed for test substance content by HPLC. The adequacy of the mixing procedure was investigated by analysis of 5 samples of each test diet from the first batch. The stability of the test substance in this batch of diets was examined after storage for 7 days at room temperature.
The test substance was found to be stable in the diet under the experimental conditions applied.
The actual levels of the test substance were as follows:
low-dose diet: 87-96% of the intended level
mid-dose diet: 98-102% of the intended level
high-dose diet: 98-103% of the intended level
The analyses of the 5 samples of each of the diets from the first batch showed coefficients of variation of 4% or less, indicating satisfactory homogeneity.

Details on mating procedure:

- Impregnation procedure: cohoused
- M/F ratio per cage: 1:2
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy

Duration of treatment / exposure:

Day 0 - 21 of gestation

Frequency of treatment:

continuously

Duration of test:

21 days of gestation

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
2.5, 5 and 10%
Basis:
nominal in diet

Doses / concentrations 2

Remarks:

Doses / Concentrations:
1760, 3520 and 6930 mg/kg bw/day
Basis:
other: overall means of periodical calculations from data on body weight, food intake and nominal levels of test substance

No. of animals per sex per dose:

24 females

Control animals:

yes, plain diet

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Body weight gain during days 0-6, 6-16 and 16-21 of gestation was determined.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Water intake (g/rat/day) during days 0-6, 6-16 and 16-21 of gestation was determined.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: major abdominal and thoracic maternal organs

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: Number of live and dead foetuses; sex of live foetuses; foetal and placental weights; foetal length;

Fetal examinations:

- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [half per litter]
- Skeletal examinations: Yes: [half per litter]
- Head examinations: No

Statistics:

Body weight gain, food and water intake were evaluated by ANOVA/Dunnett's test (P < 0.05). Maternal survival and pregnancy status, maternal and foetal external macroscopic observations, and skeletal and visceral examinations were evaluated by Fisher's exact probability test. The percentages of pre- and post-implantation loss were subjected to Kruskal-Wallis one-way ANOVA followed by the Mann-Whitney U test.

Indices:

Fertility index (%) = (No. of pregnant females/No. of mated females) x 100
Preimplantation loss (%) = [(No of corpora lutea-No. of implantation sites)/No. of corpora lutea] x 100
Postimplantation loss (%) = [(No of implantation sites-No. of live foetuses)/No. of implantation sites] x 100

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
No mortality occured and no clinical signs attributabe to the treatment were noted. Body weight gain, food and water consumption did not reveal treatment-related differences during pregnany. There were no severe macroscopic abnormalities at gross pathology of the maternal organs and tissues.
No significant differences in fertility and gestation indices, the numbers of corpura lutea and implantations, live and dead foetuses, foetal sex ratios and early and late resorptions were noted. There were no differences in ovaries, gravid and empty uterus weights in any of the dose groups compared to control animals.

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

>= 6 930 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: No adverse effects were noted up to the highest dose tested.

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Placental weight, foetal weight and foetal length were comparable between treatment and control groups.
Examination of foetal soft tissues did not reveal treatment related changes. There were no visceral malformations in any of the groups. The few visceral anomalies or visceral variations (Table 1) were spread about equally among the treatment and control groups or occurred occationally in one or a few foetuses. No significant differences between treated and control animals were found at examination of the placentas during foetal screening.
No foetal skeletal malformations occurred. Among skeletal variations, the incidence of foetuses with a supernumerary rudimentary 14th rib was higher in the controls than in any of the treatment groups (Table 2). The incidence of crooked toes varied among the treatment groups and was slightly increased in foetuses of the top-dose group. Similar variations have been noted regularly in control rats in previous studies and therefore the occurence of crooked toes in the present study is considered to reflect a processing artefact rather than a treatment-related effect. A statistically significant increased incidence of foetuses showing incomplete ossification of the skull bones was found in the low-dose group and was considered to be accidental, since it was not confirmed in the higher dose groups (table 2). Almost every foetus, including those in the control group, showed delayed ossification of the phalanges (Table 2). The delayed ossification was more prominent in the treatment groups. However, there was no dose-response relationship and no relevant effects on ossification of other bones were observed. Thus, the delayed ossification of the phalanges was not considered treatment-related.

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

>= 6 930 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No adverse effects were noted up to the highest dose tested.

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

Table 1: Visceral alterations in foetuses

Parameters	Dose groups (dietary levels)
	0	2.5%	5%	10%
No of foetuses examined	126	120	119	130
Visceral anomalies
Eyes: folded retin	1	0	0	1
Brain: meningeal haemorrhage	3	1	1	2
Heart: pericard filled with haemorrhagic fluid	3	2	2	3
Spleen: heamorrhage	0	1	0	0
Stomach: heamorrhage	2	2	5	5
Thoracic activity: filled with haemorrhagic fluid	0	0	0	1
Abdominal cavity: containing haemorrhagic fluid	1	2	0	1
Extremeties: flexure of one limb	0	0	0	1
Pancreas: haemorrhage	5	10	7	11
Skin subcutaneous haemorrhage	0	0	1	0
Total incidence of visceral anomalies	13	16	15	21
Visceral variations
Oesophagus: oesophagectasia	7	8	9	3
Thymus: small haemorrhage(s), petechiae	3	1	0	0
Stomach: hypertrophy	13	5	5	12
Stomach: hypotrophy	0	1	0	0
Kidneys: increased renal pelvic cavitation	11	11	19	14
Urether: hydrourether	1	2	0	4
Urether: bent urether	0	1	0	0
Tail: crooked	3	5	4	2
Tail: curly	1	2	2	1
Tail: ringtail	0	1	0	0
Extremeties: crooked	0	2	0	0
Extremeties: bent	0	1	0	0
Salivary gland: haemorrhage	0	0	0	2
Total incidence of visceral variations	30	34	31	32

 

 

Table 2: Skeletal alterations in foetuses

 

Parameters	Dose groups (dietary levels)
	0	2.5%	5%	10%
No of foetuses examined	137	134	137	145
Skeletal variations
Sternum: dislocated sternebra(e)	37	31	30	51
Sternum: supernumerary sternebra(e)	0	1	1	1
Sternum: absent	0	0	1	0
Ribs: 14^thrib present	9	3	4	1*
Ribs: irregular shaped ribs	0	1	0	0
Ribs: wavy rib(s)	1	0	0	1
Ribs: 13^thrib rudimentary	0	0	1	0
Forelimbs: crooked fingers	1	2	1	2
Hindlimbs: crooked toes	1	7	4	10*
Total skeletal variations	44	43	38	61
Skeletal ossification
Skull bones: two or more bones incompletely ossified	36	58***	48	49
Skull bones: hyoid bone absent	0	1	0	2
Sternum: sternebra(e) reduced or incompletely ossified	59	70	63	63
Phalanges front: 10-20 digits incompletely ossified	4	1	3	5
Phalanges front: 10-20 digits not ossified	136	118***	132	133**
Phalanges front: more than 20 digits not ossified	1	15***	4	10*
Phalanges hind: 10-20 digits incompletely ossified	3	1	2	0
Phalanges hind: 10-20 digits not ossified	136	111	129*	127***
Phalanges hind: more than 20 digits not ossified	1	19**	8*	18***
Total skeletal retardations	137	134	137	145

 */**/***: values differ significantly from the corresponding control value (* P<0.05; ** P<0.01; *** P<0.001) using Fisher's exact probability test

Applicant's summary and conclusion

Conclusions:

No maternal toxicity occurred and no effects on reproductive performance, nor on embryonic or foetal development including visceral and skeletal examination were noted up to and including a dietary level of 10% isomaltulose equivalent to 6930 mg/kg bw/day.