Xylitol

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Mutagenic effects - bacterial: Ames study, OECD 471; Negative. Reliability = 2.

Clastogenic effects - mammalian: In vitro chromosome aberration test in human lymphocytes, OECD 473; Negative. OECD 473; Reliability = 2.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

3 strains of bacterial used

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Target gene:

histidine

Species / strain / cell type:

S. typhimurium TA 1535

Species / strain / cell type:

S. typhimurium TA 1537

Species / strain / cell type:

S. typhimurium TA 1538

Metabolic activation:

with and without

Metabolic activation system:

S-9

Test concentrations with justification for top dose:

0, 15.6, 31.25, 62.5 and 125 mg/plate

Vehicle / solvent:

dissolved in phosphate buffered saline

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

3-methylcholanthrene

Details on test system and experimental conditions:

NUMBER OF REPLICATIONS: 4

Statistics:

The statistical analysis according to KAPLAN has previously been described.

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

Only in the first experiment, with the highest dose of the compound added to the bacteria and only after metabolic activation a significant (p< 0.01), about twofold increase of the revertants above background could be observed with Salmonella typhimurium TA 1538. As usually in the AMES test only three times more revertant colonies as compared to the background are considered to be a positive result and as this observation could not be reproduced, the authors do not think this finding to be of great relevance. Moreover, even in an experiment with 10 plates per dosage group the result could not be reproduced, while with the positive control 3-MCA a 15-fold increase of the revertant colonies above background could be observed.

Conclusions:

Ames test is negative.

Executive summary:

The Ames test with Salmonella typhimurium TA 1535, TA 1537 and TA 1538 was carried out to assess the potential mutagenic activity of the test substance. Only in the first experiment, with the highest dose of the compound added to the bacteria and only after metabolic activation a significant (p< 0.01), about twofold increase of the revertants above background could be observed with Salmonella typhimurium TA 1538. Although this increase is statistically significant, it is very likely to be an artefact due to the biological variation of the system, as the effect could not be reproduced in further experiments and as usually only a threefold increase of revertant colonies above background or more is considered to be positive. Neither the test substance nor one of its metabolites formed by a rat liver microsomal fraction was able to induce gene mutations.

Reference 2

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

host-mediated assay

GLP compliance:

not specified

Type of assay:

bacterial forward mutation assay

Target gene:

histidine

Species / strain / cell type:

S. typhimurium, other: TA 1530

Species / strain / cell type:

S. typhimurium, other: TA 1532

Species / strain / cell type:

S. typhimurium, other: TA 1964

Metabolic activation:

with

Metabolic activation system:

host-mediated assay; Füllinsdorf Albino mice

Test concentrations with justification for top dose:

0, 1820, 3280 and 5333 mg/kg/body weight of mice

Vehicle / solvent:

distilled water

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

not specified

Details on test system and experimental conditions:

As hosts Füllinsdorf Albino mice (SPF, 4 male and 4 female per dosage group, average weight about 30 g) and as indicator organisms various Salmonella typhimurium strains (TA 1530 = BPS, TA 1532 = FS, TA 1964 = FS), also were used. The test substance was dissolved in distilled water and applicated once per os together with the bacteria which were given intraperitoneally three hours prior to the sacrifice of the mice.

Statistics:

The statistical analysis was performed by means of the Student-t-test.

Species / strain:

S. typhimurium, other: TA 1530

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Species / strain:

S. typhimurium, other: TA 1532

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Species / strain:

S. typhimurium, other: TA 1964

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

No significant increase of the number of revertant colonies above background could be shown in the experiments. Thus, neither the test substance nor one of its metabolites, which may have been formed in the mouse are mutagenic for the Salmonella typhimurium strains used.

Conclusions:

The test substance caused no observable mutagenic effect in the system studied.

Executive summary:

The host-mediated assay in the mouse with Salmonella typhimurium was carried out to assess potential mutagenic activity of the test substance. As hosts Füllinsdorf Albino mice (SPF, 4 male and 4 female per dosage group) and as indicator organisms various Salmonella typhimurium strains (TA 1530, TA 1532 TA 1964), also were used.

No significant increase of the number of revertant colonies above background could be shown in the experiments. Thus, neither the test substance nor one of its metabolites, which may have been formed in the mouse are mutagenic for the Salmonella typhimurium strains used.

Reference 3

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

yes

Remarks:

no positive control or metabolic activation

GLP compliance:

not specified

Type of assay:

mammalian cell gene mutation assay

Species / strain / cell type:

lymphocytes: PHA-stimulated human

Metabolic activation:

not specified

Test concentrations with justification for top dose:

first experiment: 0, 0.45, 0.85, 1.75, 3.5 and 7 mg/mL
second experiment: 0, 3.5, 7 and 14 mg/mL

Vehicle / solvent:

Hanks Balanced Salt Solution

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

no

Details on test system and experimental conditions:

0.25 mL peripheral blood of a 29-year old woman, respectively a 22-year old man in the second experiment, were cultured in 5 mL medium consisting of TC-199 medium supplemented with 25% foetal calf serum, 0.25% liquemine and 1% phytohemagglutinin (PHA, Wellcome), a mitogen for T-lymphocytes. The blood was incubated in plastic tissue culture flasks in a humidified incubator at 37ºC with 5% CO2 in air for three days.

The test substance was dissolved in Hanks Balanced Salt Solution and added to the cultures 24 hours prior to the harvest of the cells.

Evaluation criteria:

Only the cultures with the highest doses and the control cultures of each experiment were evaluated. 100 cells per dosage group and 50 cells for the control group for each experiment were checked for chromosome or chromatid aberrations.

Statistics:

The statistical analysis of the results was performed according to the method of Ustenmum and Bowman.

Species / strain:

lymphocytes: PHA-stimulated human

Metabolic activation:

not specified

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Additional information on results:

No increase of aberration rate in lymphocyte cultures treated with doses of the test substance up to 14 mg/mL as compared to the untreated cultures was shown.

Conclusions:

The test substance caused no observable mutagenic effects in the system studied.

Executive summary:

A cultivated human lymphocyte test was carried out to assess the potential mutagenicity of the test substance. Blood was collected from a young male and female human and the lymphocytes incubated for 3 days. The test substance, at concentrations up to 14 mg/mL, was dissolved in Hanks Balanced Salt Solution and added to the cultures 24 hours prior to the harvest of the cells. The test substance did not cause any chromosome or chromatid aberrations in cultivated human lymphocytes. No increase of the aberration rate in treated lymphocyte cultures was seen as compared to the untreated cultures.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Clastogenic effects - mammalian: In vivo mouse micronucleus, equivalent to OECD 474. Negative. Reliability = 2

Link to relevant study records

Reference

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

no positive controls

GLP compliance:

not specified

Type of assay:

micronucleus assay

Species:

mouse

Strain:

other: Fullinsdorf albino (SPF)

Sex:

male/female

Details on test animals or test system and environmental conditions:

Males: average weight about 38 g
Females: average weight about 33 g

Route of administration:

oral: gavage

Vehicle:

dissolved in PBS

Duration of treatment / exposure:

treated twice, 30 and 6 hours before sacrifice

Frequency of treatment:

two times

Dose / conc.:

1 820 mg/kg bw/day (actual dose received)

Dose / conc.:

3 280 mg/kg bw/day (actual dose received)

Dose / conc.:

5 333 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

3/sex/dose

Control animals:

yes

Positive control(s):

no data

Tissues and cell types examined:

Smears of the bone marrow of both femora were prepared and stained as described by Schuepbach. 4000 erythrocytes per animal were checked for micronuclei.

Details of tissue and slide preparation:

Bone marrow was collected in foetal calf serum from the femur of each animal. The tube was centrifuged at 1000 rev/min for 5 minutes. The supernatant is removed and the cells in the sediment were carefully mixed by aspiration using a pipette. A small drop of the suspension was smeared on a fresh microscope slide, and the slides were air dried. Slides were stained with May-Gruenwald solution, May-Gruenwald diluted with distilled water (1:1), followed by Giemsa diluted with distilled water (1:6). Slides were then rinsed in distilled water, blotted dry and the back was cleaned with methanol. The slide was cleared in xylene and a cover glass was mounted.

Statistics:

The statistical analysis was performed by means of the Student-t-test.

Sex:

male/female

Genotoxicity:

negative

Toxicity:

not specified

Vehicle controls validity:

not specified

Negative controls validity:

not specified

Positive controls validity:

not specified

Additional information on results:

After two-fold application of the test substance (doses up to 5333 mg/kg) no significant increase of micronuclei containing erythrocytes could be shown in the bone marrow of Füllinsdorf Albino mice of either sex.

Conclusions:

The test substance caused no observable mutagenic effect in the system studied.

Executive summary:

The micronucleus test in the mouse was carried out to assess potential mutagenic activity of the test substance. Three male and three female mice were used per dosage group. They were treated twice, 30 and 6 hours before sacrifice, with up to 5333 mg/kg of the test substance dissolved in PBS. Erythrocytes were checked for micronuclei. After twofold applications, no significant increase of micronuclei containing erythrocytes could be shown in the bone marrow of Füllinsdorf Albino mice of either sex. Therefore, the test substance causes neither chromosome breaks nor mitotic non-disjunction in the bone marrow of the mouse.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

Xylitol caused no observable mutagenic effects in any of the systems studied. The analysis included an Ames test with Salmonella typhimurium TA 1535, TA 1537, and TA 1538 with and without S-9 metabolic activation; host-mediated assay in the mouse with Salmonella typhimurium TA 1530, TA 1532, and TA 1964; micronucleus test with Fullinsdorf albino mice; and chromosome analysis of cultured, PHA-stimulated human lymphocytes. Each of these in vivo and in vitro tests for mutagenicity and clastogenicity have consistently yielded negative results. 

Justification for classification or non-classification

The test substance did not produce mutagenicity in vitro or clastogenicity when evaluated in vitro and in laboratory animals. Based on an assessment of the robust genetic toxicity data for this substance, the substance does not need to be classified for germ-cell mutagenicity according to EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.