4-O-α-D-glucopyranosyl-D-glucitol

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2012

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference Type:

publication

Title:

The genotoxic and teratogenic effects of maltitol in rats

Author:

Semir Canimoglu and Eyyup Rencuzogullari

Year:

2012

Bibliographic source:

Toxicology and Industrial Health, 29 (10) 935-943

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: Okada et al (2004)

Version / remarks:

The study was performed according to Okada et al (2004) based on OECD 414 guidelines with deviations: the treatment period was from gestation day 1 to gestation day 7 (first trimester)

Deviations:

yes

Remarks:

See "Version/remarks" section for deviations details.

GLP compliance:

no

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

4-O-α-D-glucopyranosyl-D-glucitol

EC Number:

209-567-0

EC Name:

4-O-α-D-glucopyranosyl-D-glucitol

Cas Number:

585-88-6

Molecular formula:

C12H24O11

IUPAC Name:

4-O-α-D-glucopyranosyl-D-glucitol

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: from Sigma, Batch No. M8892
- Expiration date of the lot/batch: not specified
- Purity test date: not specified

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: not specified
- Stability under test conditions: not specified
- Solubility and stability of the test substance in the solvent/vehicle: not specified
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not applicable

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
Maltitol was dossilved in distilled water, no more details.

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: From Medical Sciences, Experimental Research and Application Centre of Cukurova University
- Age at study initiation: 12-16 weeks old
- Weight at study initiation: 210±2.08 g
- Fasting period before study: not specified
- Housing: Plastic Cages (32x46x18 cm)
- Diet (e.g. ad libitum): not detailed
- Water (e.g. ad libitum): not detailed
- Acclimation period:5 days before the beginning of the teratogenicity study

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±1°C
- Humidity (%): Not detailed
- Air changes (per hr): not detailed
- Photoperiod (hrs dark / hrs light): 12h/12h

IN-LIFE DATES: Not detailed

Administration / exposure

Route of administration:

intraperitoneal

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Maltitol was dissolved in distilled water.

VEHICLE
- Justification for use and choice of vehicle (if other than water): no justification was provided
- Concentration in vehicle: not detailed
- Amount of vehicle (if gavage): not applicable
- Lot/batch no. (if required): not specified
- Purity: not specified

Analytical verification of doses or concentrations:

no

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1 male / 2 females
- Length of cohabitation: one night
- Further matings after two unsuccessful attempts: no
- Verification of same strain and source of both sexes: not specified
- Proof of pregnancy: vaginal plug referred to as day 0 of pregnancy

Duration of treatment / exposure:

From gestadion day 1 to gestation day 7 (7 days)

Frequency of treatment:

once daily

Duration of test:

19 Days

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

1 000 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

2 000 mg/kg bw/day (nominal)

Doses / concentrations 4

Dose / conc.:

4 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

8 pregnant dams were used per condition

Control animals:

yes

yes, concurrent vehicle

other: positive control

Details on study design:

- Dose selection rationale: no dose selection rationale was provided
- Rationale for animal assignment (if not random): not detailed, randomly

Examinations

Maternal examinations:

The weight of pregnant rats was scored four times during the gestation period (GD0, GD6, GD12 and GD19). The rats were killed under ether anaesthesia on GD19.

Ovaries and uterine content:

The uterus and ovaries were excised. The number of implantations, live and dead foetuses, were determined. The number of corpora lutea was counted under a binocular digital stereomicroscope.

Fetal examinations:

The number of implantations, live and dead foetuses, were determined. The number of corpora lutea was counted under a binocular digital stereomicroscope. The foetuses were weighed and examined for macroscopic external abnormalities. The foetuses were killed with ether treatment and then they were double-stained for skeletal examination using the double staining method according to Okada et al. (2004) with minor modifications. The skin, muscle, fat and internal organs of the foetuses were gently removed. Skeletons were fixed with acetone for 2 weeks and then stained with the double-staining solution consisting of 0.0005% alizarin red, 0.01% alican blue 8GX and 0.05% glacial acetic acid in 70% ethanol for 2 days. The specimens were macerated in 1% potassium hydroxide for 2 days after rinsing with tap water. Finally, the specimens were treated with 0.8% potassium hydroxide and 20% glycerin solution for 5 days to clean soft tissues. Then, the specimens were preserved in glycerin and 70% ethanol (1:1). Skeletons of double stained-foetuses were examined under a stereomicroscope and digitally photographed with a digital stereomicroscope. The skeletal segments were characterized according to Menegola et al. (2001).

Statistics:

The t test was used for the statistical significance of a number of corpora lutea, total implantation, empty implantation, live and dead foetuses and a number of abnormal foetuses after the one-way statistical analysis of variance.

Historical control data:

No historical data were presented in this study.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Maternal developmental toxicity

Number of abortions:

effects observed, non-treatment-related

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Details on maternal toxic effects:

The weight of dams linearly increased in the control and all the concentrations of maltitol during the gestation periods. The increase in weight of dams between GD0 and GD19 during gestation was 75 g in the control groups, 63, 68 and 58 g in the maltitol concentrations of 1, 2 and 4 mg/kg bw, respectively (Table 2).

Maternal abnormalities

Abnormalities:

no effects observed

Results (fetuses)

Fetal body weight changes:

effects observed, non-treatment-related

External malformations:

no effects observed

Skeletal malformations:

effects observed, treatment-related

Visceral malformations:

not examined

Details on embryotoxic / teratogenic effects:

Maltitol increased the number of corpora lutea when compared with the control; however, it did not increase the number of total and empty implantations and did not induce the number of live foetuses (Table 1). In total, three dead foetuses (0.42 + 0.29) were shown in two dams treated with 2 g/kg bw ofmaltitol; however, there was no statistical significance. Maltitol decreased the weight of the foetuses at all the concentrations. The weight of the foetuses was recorded at 1.97 + 0.05 in dams treated with 4 g/kg bw of maltitol, while growth retardation and a lack of ossify were also observed in this group. Wave ribs as a skeletal abnormality were observed in three foetuses of dams treated with maltitol at 2 g/kg bw.
The authors suggested in the publication that maltitol caused hyperglycemia and caused embryotoxicity by preventing the embryo nutriment transfer and oxygenation via yolk bladder dysfunction dependent on a high blood sugar level (see the publication for review).

Fetal abnormalities

Abnormalities:

no effects observed

Any other information on results incl. tables

Table1. The teratogenic effect of maltitol in rats after intraperitoneal treatment with maltitol during 7 days(fromGD1 toGD7).

 

				Maltitol
	Control	EMS (50 mg/kg)	1 g/kg	2 g/kg	4 g/kg
Number of pregnant dams	8	8	13	12	12
Corpora lutea	9.64+1.74	12.25+1.04	11.23+0.44^a	12.75+0.35^b	11.08+0.45^a
Total implantation	10.62+0.59	10.25+1.12	10.46+0.57	10.00+0.49	10.08+0.55
Empty implantation	0.50+0.26	9.25+0.95	0.50+0.19	0.33+0.14	1.00+0.36
Live foetuses	10.12+0.39	1.00+0.42	10.00+0.55	9.41+0.64	9.08+0.76
Dead foetuses	–	–	–	0.42+0.29	–
Foetusweight(g)	2.60+0.007	1.74+0.10	2.16+0.05^b	2.42+0.83^c	1.97+0.05^b
Litter with abnormal foetuses	–	2	–	3	6
Number of abnormal foetuses	–	3^d	–	3^d	43^e

 EMS: ethylmethansulphonate; ^ap<0.01significantwhen compared with control.; ^bp<0.001significant when compared with control.; ^cp<0.05 significant when compared with control.; ^dSkeletal abnormality (wave ribs); ^eThe foetuses had growth retardation due to the low body weight.

Table 2. The weight of dams during gestation^a.

Gestation period (days)	Control	EMS (50 mg/kg bw)	Maltitol
			1 g/kg bw	2 g/kg bw	4 g/kg bw
0	201.75 ± 5.09	221.62 + 7.36	203.30 + 4.08	216.58 + 2.40	212.41 + 3.25
6	209.50 + 5.61	207.25 + 6.55	212.30 + 4.46	228.25 + 2.57^b	218.75 + 4.08
12	229.50 + 7.05^c	208.37 + 9.30	226.53 + 4.33^b	245.66 + 2.15^b	231.83 + 3.96^b
19	276.50 + 6.33^b	220.00 + 9.98	266.92 + 5.57^b	284.83 + 3.34^b	262.91 + 4.23^b

EMS: ethylmethanesulphonate.; ^aSignificant when compared with weight of dams before gestation (GD0).; ^bp< 0.001.; ^cp< 0.01.

 

 

Applicant's summary and conclusion

Conclusions:

Under the experimental conditions of this study, it was recorded that maltitol was not teratogenic; however, maltitol showed an embryotoxic effect via decreasing the foetuses weight and causing growth retardation at a very high concentration (4 g/kg bw).
This study cannot be used for classification because it does not fully comply with current teratogenicity testing standards: the treatment period was limited to the first trimester of gestation instead of at least from implantation to one day prior to the day of scheduled kill, which should be as close as possible to the normal day of delivery (organogenesis period was not covered during the treatment). In addition, the intraperitoneal route was used instead of the currently recommended oral route and high doses were used (top dose of 4000 mg/kg bw/day). Finally, only 8 pregnant rats were used per condition instead of 16 required in the current guideline.

Executive summary:

In this non GLP compliant in vivo study, the teratogenicity and embryotoxicity of maltitol was investigated in rats according to Okada et al. method (2004).

After mating period with male, the pregnant Sprague-Dawley females rats were intraperitoneally treated with 1, 2 and 4 g/kg bw/day concentrations of maltitol during the first 7 days of gestation (first trimester) to investigate the teratogenicity of maltitol. Ethyl methanesulphonate was used as postive control. The embryos were collected after killing the dams by cervical dislocation under ether anaesthesia on gestation day 19 in order to be evaluated.

The weight of dams linearly increased in the control and all the concentrations of maltitol during the gestation periods. The increase in weight of dams between GD0 and GD19 during gestation was 75 g in the control groups, 63, 68 and 58 g in the maltitol concentrations of 1, 2 and 4 mg/kg bw, respectively.

Maltitol increased the number of corpora lutea when compared with the control; however, it did not increase the number of total and empty implantations and did not induce the number of live foetuses. In total, three dead foetuses (0.42 + 0.29) were shown in two dams treated with 2 g/kg bw ofmaltitol; however, there was no statistical significance. Maltitol decreased the weight of the foetuses at all the concentrations. The weight of the foetuses was recorded at 1.97 + 0.05 in dams treated with 4 g/kg bw of maltitol, while growth retardation and a lack of ossify were also observed in this group. Wave ribs as a skeletal abnormality were observed in three foetuses of dams treated with maltitol at 2 g/kg bw. The authors suggested in the publication that maltitol caused hyperglycemia and caused embryotoxicity by preventing the embryo nutriment transfer and oxygenation via yolk bladder dysfunction dependent on a high blood sugar level (see the publication for review).

Under the experimental conditions of this study, it was recorded that maltitol was not teratogenic; however, maltitol showed an embryotoxic effect via decreasing the foetuses weight and causing growth retardation at a very high concentration (4 g/kg bw).

This study cannot be used for classification because it does not fully comply with current teratogenicity testing standards: the treatment period was limited to the first trimester of gestation instead of at least from implantation to one day prior to the day of scheduled kill, which should be as close as possible to the normal day of delivery (organogenesis period was not covered during the treatment). In addition, the intraperitoneal route was used instead of the currently recommended oral route and high doses were used (top dose of 4000 mg/kg bw/day). Finally, only 8 pregnant rats were used per condition instead of 16 required in the current guideline.