Benzaldehyde, 3-ethoxy-4-(.beta.-D-glucopyranosyloxy)-

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

July - September 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

21 July 1997

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

3-ethoxy-4-{[(2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy}benzaldehyde

EC Number:

602-769-3

Cas Number:

122397-96-0

Molecular formula:

C15H20O8

IUPAC Name:

3-ethoxy-4-{[(2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy}benzaldehyde

Test material form:

solid: particulate/powder

Method

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Details on mammalian cell type (if applicable):

CELLS USED
- Source of cells: Molecular Toxicology Inc., 157 Industrial Park Dr. Boone, NC 28607, U.S.A.
- Suitability of cells: standard cells, genotype confirmed by the laboratory

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254 induced rat liver S9 fraction

Test concentrations with justification for top dose:

Based on initial experiment: 156.25, 312.5, 625, 1250 , 2500, and 5000 µg/plate both in the absence and presence of metabolic activation system (10% v/v S9 mix) for all the tester strains.
In the initial experiment, normal bacterial lawn was observed up to the tested concentration of 5000 µg/plate both in the absence and presence of the metabolic activation system (5% v/v S9 mix) in the tester strains of TA1537, TA1535, TA98, TA100 and TA102.

Vehicle / solvent:

dimethyl sulfoxide (DMSO)

Controls

Untreated negative controls:

yes

Remarks:

DMSO

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

sodium azide

mitomycin C

other: 2-aminoanthracene

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Incubation period: 48 hrs
- Expression time (cells in growth medium):
- Selection time (if incubation with a selection agent):
- Fixation time (start of exposure up to fixation or harvest of cells):

SELECTION AGENT (mutation assays): crystal violet for rfa wall mutation, biotin for growth requirement (except TA 102), UV exposure sensitivity (except for TA102), histidine for growth requirement, resistance to ampicillin (TA98, 100, 102) for pKM101 plasmid, resistance to tetracycline (TA102) for pAQ1 plasmid

NUMBER OF REPLICATIONS: 3 per each concentration

NUMBER OF CELLS EVALUATED: plate density 1 - 2 x 10^9 bacteria/mL


DETERMINATION OF CYTOTOXICITY : decrease in the number of revertant colonies per plate and/or by a thinning of the bacterial background lawn

Rationale for test conditions:

Guideline requirements

Evaluation criteria:

A result is considered positive if a concentration-related increase over the range tested and/or a reproducible increase at one or more concentrations in the number of revertant colonies per plate in at least one strain with or without metabolic activation system.

Strains TA1535, TA1537
Data sets were judged positive, if the increase in mean revertants at the peak of the dose response was equal to or greater than 3.0 times the mean negative control value.

Strain TA98, TA100 and TA102
Data sets were judged positive, if the increase in mean revertants at the peak of the dose response was equal to or greater than 2.0 times the mean negative control value.

Statistical analysis was used as an aid in the evaluation of the dose response.
A response that did not meet all three of the above criteria (magnitude, concentration-responsiveness, reproducibility) was not judged as positive.
Negative results obtained in the initial toxicity-mutation test were confirmed by a second trial, using the same method as specified above, with an alteration in concentration spacing

Statistics:

Simple linear regression analysis was performed for TA1537, TA1535, TA98, TA100 and TA102, separately, to assess the dose dependent nature of any increase in revertant colonies.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

S. typhimurium TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Normal bacterial background lawn was observed up to the tested concentration of 5000 µg/plate both in the absence and presence of the metabolic activation system (10% v/v S9 mix) in tester strains. Results revealed that there was no positive mutagenic effect in tester strains TA1537, TA1535, TA98, TA100, and TA102, at the tested concentration levels 156.25, 312.5, 625, 1250, 2500, and 5000 µg/plate of ethyl vanillin glucoside, in the absence and presence of the metabolic activation system (10% v/v S9 mix), when compared with the concurrent negative control. Statistical analysis did not reveal any significant effect.

Applicant's summary and conclusion

Conclusions:

From results of this study, it is concluded that ethyl vanillin glucoside is non-mutagenic to any strains of Salmonella typhimurium , viz., TA1537, TA1535, TA98, TA100, and TA102 when tested under the specified experimental conditions.

Executive summary:

From results of this study, it is concluded that ethyl vanillin glucoside is non-mutagenic to any strains of Salmonella typhimurium , viz., TA1537, TA1535, TA98, TA100, and TA102 when tested under the specified experimental conditions.