2-chlorophenol

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:

23 Mar - 2 Apr 1999

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2000

Report date:

2000

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

no data on historical controls provided

GLP compliance:

yes

Remarks:

according to the Japanese GLP Standard (as described on JECDB)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

2-chlorophenol

EC Number:

202-433-2

EC Name:

2-chlorophenol

Cas Number:

95-57-8

Molecular formula:

C6H5ClO

IUPAC Name:

2-chlorophenol

Test material form:

liquid

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: in closed bottle at room temperature
- Solubility and stability of the test substance in the solvent/vehicle: water solubility 2.71%, easily soluble in DMSO and acetone

Method

Target gene:

his operon, trp operon

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Metabolic activation system:

S9 mix, prepared from the livers of rats treated with phenobarbital and 5-6-benzoflavone

Test concentrations with justification for top dose:

TA100 and TA1535, with and without metabolic activation: 39.1, 78.1, 156, 313, 652, 1250 and 2500 μg/mL (tested up to cytotoxic concentrations)
WP2uvrA, TA98 and TA1537, with and without metabolic activation: 156, 313, 652, 1250, 2500 and 5000 μg/mL (tested up to the limit dose)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Due to the limited solubility of the test substance in water, DMSO was selected as the vehicle.

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

sodium azide

other: aminofluorene for TA100 and TA98, N-ethyl-N'-nitrosogunanidine for WP2uvrA, 2-aminoanthracene for strains with S9

Details on test system and experimental conditions:

METHOD OF APPLICATION: preincubation

DURATION
- Preincubation period: 20 minutes
- Exposure duration: 48 h

NUMBER OF REPLICATIONS:3 replications each in 2 independent experiments

Evaluation criteria:

Evaluation:
A test substance was considered positive in the Ames test if:
a) a more than 2-fold increase in the number of revertants was observed
b) the increase was dose-related
c) the positive result was reproducible.

A test substance was considered negative in the Ames test if none of the above criteria were observed under all experimental conditions.

Statistics:

Mean values and standard deviations were calculated.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 98

Remarks:

TA100/ TA1535/ TA1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

starting at 1250 µg/mL +/-S9 in TA 100 and TA 1535; starting at 2500 µg/mL +/- S9 in WP2 uvrA, TA 98 and TA 1537

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
A preliminary test was performed with test concentrations ranging from 1.22 - 5000 μg/mL with/without S9. No increase in the number of revertants was observed. Furthermore, cytotoxicity was observed starting at 1250 μg/mL in TA100 and TA1535, and at 5000 μg/mL in WP2urvA, TA98 and TA1537 with and without metabolic activation.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Measurement of cytotoxicity used: number of revertant colonies

Any other information on results incl. tables

Table 1. Test results for 2 -chlorphenol (Trial 1)

 

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate ± standard deviation
		Base-pair substitution type			Frameshift type
		TA100	TA1535	WP2urvA	TA98	TA1537
–	0	112 ± 7	10 ± 3	24 ± 2	22 ± 5	7 ± 2
–	39.1	123 ± 5	8 ± 1	-	-	-
–	78.1	104 ± 18	12 ± 1	-	-	-
–	156	116 ± 15	11 ± 5	22 ± 3	24 ± 2	6 ± 2
–	313	103 ± 12	8 ± 2	33 ± 3	21 ± 3	7 ± 2
–	625	101 ± 12	12 ± 4	22 ± 2	23 ± 4	7 ± 2
–	1250	84 ± 9*	7 ± 2*	22 ± 2	20 ± 2	5 ± 1
–	2500	0 ± 0*	0 ± 0*	0 ± 0*	0 ± 0*	0 ± 0*
–	5000	-	-	0 ± 0*	0 ± 0*	0 ± 0*
–	Positive controls	AF-2	NaN3	ENNG	AF-2	9-AA
	Mean No. of colonies/plate ± SD	541 ± 38	377 ± 12	821 ± 29	375 ± 29	351 ± 39
+	0	113 ± 4	7 ± 2	27 ± 5	26 ± 4	13 ± 5
+	39.1	111 ± 9	8 ± 1	-	-	-
+	78.1	110 ± 15	11 ± 3	-	-	-
+	156	122 ± 6	9 ± 2	25 ± 4	28 ± 3	10 ± 2
+	313	117 ± 9	7 ± 2	29 ± 9	29 ± 10	12 ± 3
+	625	109 ± 11	9 ± 2	32 ± 7	28 ± 7	14 ± 3
+	1250	85 ± 5*	12 ± 3*	31 ± 4	27 ± 6	12 ± 4
+	2500	0 ± 0*	0 ± 0*	0 ± 0*	0 ± 0*	0 ± 0*
+	5000	-	-	0 ± 0*	0 ± 0*	0 ± 0*
+	Positive controls	2-AA	2-AA	2-AA	2-AA	2-AA
	Mean No. of colonies/plate ± SD	1161 ± 32	168 ± 7	1313 ± 38	443 ± 9	140 ± 6

AF-2= 2 -aminofluorene

NaN3 = sodium azid

ENNG = N-ethyl-N'-nitro-N'-nitrosogunanidine

9-AA = 9-aminoacridine

2AA = 2-aminoanthracene

SD = standard deviation

* = growth inhibition/cytotoxicity

Table 2. Test results for 2 -chlorphenol (Trial 2)

 

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate ± standard deviation
		Base-pair substitution type			Frameshift type
		TA100	TA1535	WP2urvA	TA98	TA1537
–	0	105 ± 4	8 ± 2	20 ± 5	23 ± 3	13 ± 1
–	39.1	106 ± 3	8 ± 2	-	-	-
–	78.1	103 ± 9	7 ± 1	-	-	-
–	156	103 ± 5	8 ± 2	29 ± 2	23 ± 2	15 ± 4
–	313	108 ± 11	11 ± 1	26 ± 4	18 ± 2	12 ± 2
–	625	94 ± 2	8 ± 1	23 ± 4	18 ± 2	15 ± 1
–	1250	87 ± 15*	8 ± 2*	19 ± 3	15 ± 2	14 ± 4
–	2500	0 ± 0*	0 ± 0*	0 ± 0*	0 ± 0*	0 ± 0*
–	5000	-	-	0 ± 0*	0 ± 0*	0 ± 0*
–	Positive controls	AF-2	NaN3	ENNG	AF-2	9-AA
	Mean No. of colonies/plate ± SD	524 ± 14	514 ± 46	843 ± 33	505 ± 21	430 ± 34
+	0	105 ± 3	10 ± 2	25 ± 5	25 ± 5	19 ± 2
+	39.1	101 ± 6	9 ± 1	-	-	-
+	78.1	108 ± 10	8 ± 2	-	-	-
+	156	95 ± 8	11 ± 3	28 ± 5	25 ± 4	18 ± 5
+	313	96 ± 6	10 ± 1	27 ± 4	25 ± 6	16 ± 3
+	625	108 ± 10	9 ± 2	28 ± 6	26 ± 2	21 ± 3
+	1250	96 ± 5*	8 ± 3*	29 ± 3	23 ± 0	19 ± 2
+	2500	0 ± 0*	0 ± 0*	0 ± 0*	0 ± 0*	0 ± 0*
+	5000	-	-	0 ± 0*	0 ± 0*	0 ± 0*
+	Positive controls	2-AA	2-AA	2-AA	2-AA	2-AA
	Mean No. of colonies/plate ± SD	1010 ± 58	210 ± 9	1477 ± 47	424 ± 12	192 ± 7

AF-2= 2 -aminofluorene

NaN3 = sodium azido

ENNG = N-ethyl-N'-nitro-N'-nitrosogunanidine

9-AA = 9-aminoacridine

2AA = 2-aminoanthracene

SD = standard deviation

* = growth inhibition/cytotoxicity