1,1-dimethoxy-2-phenylethane

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:

30 Jan - 23 Feb 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

adopted 21. Jul 1997

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Ministerium für Umwelt und Verkehr Baden-Württemberg, Stuttgart, Germany

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

1,1-dimethoxy-2-phenylethane

EC Number:

202-945-6

EC Name:

1,1-dimethoxy-2-phenylethane

Cas Number:

101-48-4

Molecular formula:

C10H14O2

IUPAC Name:

(2,2-dimethoxyethyl)benzene

Method

Target gene:

his operon

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

cofactor supplemented post-mitochondrial fraction (S9 mix), prepared from the livers of male Sprague-Dawley rats treated with Aroclor 1254 (500 mg/kg bw)

Test concentrations with justification for top dose:

Experiment 1
15, 50, 150, 500 and 1500 µg/plate with and without metabolic activation for TA100 and TA102
50, 150, 500, 1500 and 5000 µg/plate with and without metabolic activation for TA98, TA1535 and TA1537

Experiment 2
50, 150, 500, 1500 and 5000 µg/plate without metabolic activation for TA98, TA100, TA102 and TA1537
500, 1000, 1500, 3000 and 5000 µg/plate without metabolic activation for TA1535
50, 150, 500, 1500 and 5000 µg/plate with metabolic activation for all strains

Vehicle / solvent:

- Vehicle/solvent used: DMSO

Controls

Untreated negative controls:

yes

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 (2-AA)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 to 72 h

NUMBER OF REPLICATIONS: 3 replications each in 2 independent experiments

DETERMINATION OF CYTOTOXICITY
- Method: reduction in the number of revertant colonies and/or a diminution of the background lawn

Statistics:

X²-test was used to estimate the statistical significance of the difference between the mean number of revertants in the negative controls and the plates at each dosage level. Mean values and standard deviation were calculated.

Results and discussion

Test results

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

positive

Remarks:

in TA1535 without S9 mix

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

in TA102 at 1500 µg/plate without S9 mix and at 5000 µg/plate with S9 mix

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No precipitation occurred up to the highest investigated dose.

Remarks on result:

other:

Any other information on results incl. tables

Table 1. Test results of Experiment 1 (plate incorporation).

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate (average of 3 plates ± Standard deviation)
		Base-pair substitution type			Frameshift type
		TA100	TA1535	TA102	TA98	TA1537
–	0	99 ± 8	25 ± 2	293 ± 31	31 ± 6	10 ± 5
–	0 (DMSO)	86 ± 2	24 ± 3	298 ± 19	26 ± 3	12 ± 2
–	15	89 ± 14	-	305 ± 13	-	-
–	50	73 ± 8	25 ± 6	296 ± 21	32 ± 3	12 ± 3
–	150	76 ± 5	32 ± 3	303 ± 14	42 ± 5	12 ± 4
–	500	83 ± 5	38 ± 7	248 ± 31	33 ± 8	11 ± 5
–	1500	97 ± 8	43 ± 8	221 ± 6^T	39 ± 3	12 ± 4
–	5000	-	47 ± 3**	-	36 ± 4	10 ± 4
Positive controls, –S9	Name	NaN3	NaN3	MMC	2-NF	9-AA
	Concentrations (μg/plate)	0.7	0.7	0.15	2.5	50
	Mean No. of colonies/plate (average of 3 ± SD)	503 ± 58	905 ± 41	755 ± 51	614 ± 15	606 ± 63
+	0	97 ± 10	10 ± 3	343 ± 28	21 ± 4	17 ± 5
+	0 (DMSO)	88 ± 8	11 ± 3	340 ± 14	21 ± 1	14 ± 3
+	15	88 ± 3	-	349 ± 20	-	-
+	50	84 ± 15	9 ± 2	342 ± 37	25 ± 3	11 ± 4
+	150	81 ± 5	10 ± 3	348 ± 12	22 ± 6	14 ± 5
+	500	81 ± 3	10 ± 2	324 ± 17	25 ± 4	15 ± 3
+	1500	85 ± 8	12 ± 1	305 ± 21	23 ± 4	17 ± 3
+	5000	-	14 ± 3	-	22 ± 3	15 ± 2
Positive controls, +S9	Name	2-AA	2-AA	2-AA	2-AA	2-AA
	Concentrations (μg/plate)	0.8	0.8	0.9	0.8	1.7
	Mean No. of colonies/plate (average of 3 ± SD)	627 ± 90	216 ± 47	655 ± 61	741 ± 94	374 ± 103

DMSO: Dimethylsulphoxide

NaN3: Sodium azide

2-AA: 2-aminoanthracene

9-AA: 9-aminoacridine

2-NF: 2-nitrofluorene

MMC: Mytomycin C

^T: bacteriotoxic

**: Significantly different from control (p < 0.01)

 

Table 2. Test results of Experiment 2 (plate incorporation).

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate (average of 3 plates ± Standard deviation)
		Base-pair substitution type			Frameshift type
		TA100	TA1535	TA102	TA98	TA1537
–	0	201 ± 16	19 ± 3	326 ± 20	25 ± 4	8 ± 4
–	0 (DMSO)	158 ± 6	20 ± 2	292 ± 18	23 ± 5	10 ± 2
–	50	140 ± 22	40 ± 3**	348 ± 14	23 ± 3	8 ± 4
–	150	174 ± 21	35 ± 6	304 ± 22	24 ± 3	10 ± 3
–	500	140 ± 15	44 ± 5**	303 ± 19	26 ± 5	10 ± 3
–	1500	173 ± 18	43 ± 5**	248 ± 20	21 ± 3	6 ± 3
–	5000	132 ± 21	38 ± 10	138 ± 8^T	24 ± 3	9 ± 4
Positive controls, –S9	Name	NaN3	NaN3	MMC	2-NF	9-AA
	Concentrations (μg/plate)	0.7	0.7	0.15	2.5	50
	Mean No. of colonies/plate (average of 3 ± SD)	471 ± 91	844 ± 12	916 ± 68	772 ± 26	493 ± 174
+	0	171 ± 23	9 ± 6	365 ± 5	29 ± 3	15 ± 6
+	0 (DMSO)	174 ± 6	9 ± 3	360 ± 13	25 ± 3	11 ± 3
+	50	176 ± 23	6 ± 2	353 ± 37	25 ± 3	11 ± 5
+	150	206 ± 14	10 ± 3	369 ± 3	30 ± 1	12 ± 5
+	500	215 ± 15	8 ± 3	328 ± 30	26 ± 3	12 ± 5
+	1500	189 ± 21	8 ± 3	305 ± 48	25 ± 6	13 ± 4
+	5000	210 ± 29	9 ± 4	253 ± 8^T	25 ± 4	12 ± 5
Positive controls, +S9	Name	2-AA	2-AA	2-AA	2-AA	2-AA
	Concentrations (μg/plate)	0.8	0.8	0.9	0.8	1.7
	Mean No. of colonies/plate (average of 3 ± SD)	647 ± 113	129 ± 11	457 ± 23	833 ± 43	248 ± 24

DMSO: Dimethylsulphoxide

NaN3: Sodium azide

2-AA: 2-aminoanthracene

9-AA: 9-aminoacridine

2-NF: 2-nitrofluorene

MMC: Mytomycin C

^T: bacteriotoxic

**: Significantly different from control (p < 0.01)

In the concentration range investigated the test substance induced a significant and dose related increase in the mutation frequency of the tester strain TA1535 in the absence of metabolic activation.

Applicant's summary and conclusion

Conclusions:

Under the conditions of this Ames test the test substance was mutagenic in TA1535 without metabolic activation.

Executive summary:

A bacterial gene mutation assay with the test substance was performed in accordance with OECD Guideline 471 and in compliance with GLP (2001). In two independent experiments, the Salmonella typhimurium strains TA98, TA100, TA102, TA1535 and TA1537 were exposed to the test substance using the plate incorporation method. Based on the results of a pre-experiment, test substance concentrations of 15 to 5000 µg/plate were selected for the incubation with and without metabolic activation in the first experiment. In the second experiment, concentrations of 50 to 5000 µg/plate were selected for the incubation with and without metabolic activation. No precipitation occurred up to the highest investigated dose. In the absence and presence of metabolic activation the test substance was bacteriotoxic towards the strain TA102 at 1500 and 5000 µg/plate, respectively. In the concentration range investigated the test substance induced a significant and dose related increase in the mutation frequency of the tester strain TA1535 in the absence of metabolic activation. The results with the positive control substances confirmed the known reversion properties and specificity of the tester strains as well as the full activity of the metabolizing system. Under the conditions of this experiment, the test substance was mutagenic in the S. typhimurium strain TA1535 without metabolic activation.