Methyl (R)-amino(4-hydroxyphenyl)acetate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

end on 21-MAR-1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: study performed according to OECD guideline and GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Methyl (R)-amino(4-hydroxyphenyl)acetate

EC Number:

253-657-2

EC Name:

Methyl (R)-amino(4-hydroxyphenyl)acetate

Cas Number:

37763-23-8

Molecular formula:

C9H11NO3

IUPAC Name:

methyl (2R)-2-amino-2-(4-hydroxyphenyl)acetate

Details on test material:

- Name of test material (as cited in study report): FGHM
- Molecular formula (if other than submission substance): C9H11NO3
- Structural formula attached as image file (if other than submission substance): see Fig.
- Substance type: monoconstituent substance
- Physical state: powder
- Stability under test conditions: stability in dimethylsulphoxide not indicated
- Storage condition of test material: in refrigerator in the dark

Method

Species / strain

Species / strain / cell type:

other: Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 ; Escherichia coli strain WP2uvrA

Metabolic activation:

with and without

Metabolic activation system:

Rat liver microsomal enzymes were routinely prepared from adult male Wistar rats, which were obtained from Charles River, Sulzfeld, Germany

Test concentrations with justification for top dose:

Experiment 1 & 2: 100, 333, 1000, 3330, 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: data not available

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: -S9mix: sodium azide (SA, 1µg, TA1535), 9-aminoacridine (9AC, 60µg, TA1537), daunomycine (DM, 4µg, TA98), methylmethanesulfonate (MMS, 650µg, TA100), 4-nitroquinoline-N-oxide (4-NQO, 10µg, WP2uvrA) : +S9mix: 2-aminoanthracene (2AA, all strains, 0.5-5µg)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: after solidification of the top agar, the plates were turned and incubated in the dark at 37°C for 48 h.

NUMBER OF REPLICATES: triplicate in each strain.

DETERMINATION OF CYTOTOXICITY
- Method: the reduction of the bacterial background lawn, the increase in the size of the microcolonies and the reduction of the revertant colonies were observed

OTHER: the revertant colonies (histidine independent c.q. tryptophan independent) were counted automatically with a Artek model 880 colony counter or manually, if less than 40 colonies per plate were present.

Evaluation criteria:

A test substance is considered negative (not mutagenic) in the test if:
a) The total number of revertants in any tester strain at any concentration is not greater than two times the solvent control value, with or without metabolic activation.
b) The negative response should be reproducible in at least one independently repeated experiment.

A test substance is considered positive (mutagenic) in the test if:
a) It induces a number of revertant colonies, dose related, greater than two-times the number of revertants induced by the solvent control in any of the tester strains, either with or without metabolic activation.
However, any mean plate count of less than 20 is considered to be not significant.
b) The positive response should be reproducible in at least one independently repeated experiment.

The preceding criteria were not absolute and other modifying factors might enter into the final evaluation decision.

Statistics:

not applicable

Results and discussion

Test results

Species / strain:

other: Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 ; Escherichia coli strain WP2uvrA

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:

not examined

Positive controls validity:

valid

Additional information on results:

Precipitate:
HPGM (FGHM) did not precipitate in the top agar. Precipitation of HPGM (FGHM) on the plates was not observed at the start or at the end of the incubation period in all tester strains.

Toxicitv of the test substance:
The bacterial background lawn was not reduced at all concentrations tested and no decrease in the number of revertants was observed.

Number of revertants:
All bacterial strains showed negative responses over the entire dose range, i.e. no dose-related, two-fold, increase in the number of revertants in two independently repeated experiments.
The negative and strain-specific positive control values were within the laboratory background historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table 1 : results of experiment 1

Dose (µg/plate)	Mean number of revertant colonies/3 replicate plates with different strains of Salmonella typhimurium and one Escherichia coli strain
	TA 1535	TA 1537	TA 98	TA 100	WP2uvrA
	WITHOUT S9-MIX
3				66	16
10				70	17
33				73	19
100	16	8	26	66	18
333	18	7	32	69	18
1000	17	7	27	73	17
3330	13	7	26	69	20
5000	17	7	31	79	17
Solvent control	14	10	23	93	14
Positive control	251	325	325	720	520
	WITH S9-MIX
3				65	17
10				60	21
33				68	20
100	16	7	35	64	20
333	15	5	36	79	16
1000	15	7	31	77	13
3330	13	8	29	70	16
5000	15	5	28	76	16
Solvent control	17	4	30	76	16
Positive control	79	397	915	740	261

 

Table 2 : results of experiment 2

Dose (µg/plate)	Mean number of revertant colonies/3 replicate plates with different strains of Salmonella typhimurium and one Escherichia coli strain
	TA 1535	TA 1537	TA 98	TA 100	WP2uvrA
	WITHOUT S9-MIX
100	8	3	21	96	8
333	9	4	16	90	9
1000	7	7	21	78	11
3330	9	4	19	95	13
5000	6	5	20	91	9
Solvent control	8	5	21	93	9
Positive control	157	301	350	908	269
	WITH S9-MIX
100	7	5	22	96	16
333	6	3	20	88	13
1000	5	5	18	75	9
3330	6	5	19	82	14
5000	7	5	18	81	15
Solvent control	7	6	26	88	10
Positive control	336	686	822	997	240

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative without metabolic activation

Based on the results of this study it is concluded that HPGM (FGHM) is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Executive summary:

HPGM (FGHM) was tested in the Salmonella typhimurium reverse mutation assay with four histidine requiring strains of Salmonella typhimurium (TA1535, TA1537, TA100 and TA98) and in the Escherichia coli reverse mutation assay with a tryptophan-requiring strain of Escherichia coli WP2uvrA in two independent experiments. (OECD 471, GLP).

HPGM (FGHM) was tested up to concentrations of 5000 µg/plate in the absence and presence of S9-mix. HPGM (FGHM) did not precipitate on the plates at this dose-level. The bacterial background lawn was not reduced at all concentrations tested and no decrease in the number of revertants was observed.

HPGM (FGHM) did not induce a dose-related, two-fold, increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9- metabolic activation. These results were confirmed in an independently repeated experiment.

Based on the results of this study it is concluded that HPGM (FGHM) is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.