(S)-2-chloropropionic acid

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:

17th - 24th July 1992

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study to GLP.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1992

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

(S)-2-chloropropionic acid

EC Number:

411-150-5

EC Name:

(S)-2-chloropropionic acid

Cas Number:

29617-66-1

Molecular formula:

Hill formula: C3H5ClO2 CAS formula: C3H5ClO2

IUPAC Name:

(2S)-2-chloropropanoic acid

Details on test material:

- Name of test material (as cited in study report): L-2-Chloropropionic acid
- Physical state: Liquid
- Analytical purity: 90.4% w/w
- Purity test date: 30th April 1992
- Lot/batch No.: Bx 0071, ADH 05138
- Expiration date of the lot/batch: Not specified

Method

Species / strain

Species / strain 1

Species / strain / cell type:

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

Species / strain 2

Species / strain / cell type:

E. coli WP2

Species / strain 3

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

S9 Fraction

Test concentrations with justification for top dose:

100, 200, 500, 1000, 2500 and 5000 micrograms/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Sterile deionised water for the test compound, negative control and some positive controls.
DMSO was used for the remaining positive controls.

Controls

Controls 1

Untreated negative controls:

yes

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

WP2P uvrA

Controls 2

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

WP2P strain

Controls 3

Positive controls:

yes

Positive control substance:

other: Acridine Mutagen ICR191

Remarks:

TA1537 strain

Controls 4

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

TA1535 and TA100

Controls 5

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene

Remarks:

All strains

Controls 6

Positive controls:

yes

Positive control substance:

other: Daunomyc in HCl

Remarks:

TA98 Strain

Details on test system and experimental conditions:

METHOD OF APPLICATION: The initial assay was performed using the standard plate incorporation protocol.
The re-test was performed as above using the standard plate incorporation protocol for the -S9 assays but the pre-incubation protocol for the +S9 determinations.

DURATION
- Pre-incubation period: 60 minutes
- Exposure duration: 3 days

NUMBER OF REPLICATIONS: Test compound – 3 plates per dose
Negative controls – 5 plates
Positive controls – 3 dose levels per strain, 2 plates per dose.


DETERMINATION OF CYTOTOXICITY
- Method: Number of revertant colonies

Evaluation criteria:

A positive response in a (valid) individual experiment is achieved, when one or both of the following criteria are met:
a) a statistically significant dose-related increase in the mean number of revertant colonies is obtained;
b) a two-fold or greater increase in the mean number of revertant colonies which is statistically significant, is observed at at least one dose level.

Results and discussion

Test results

Test results 1

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Any other information on results incl. tables

Test data for Experimental Phase 1.

 

Strain	Activation	Dose Levels (micrograms/plate)	No revertants/plate
			Plate 1	Plate 2	Plate 3
TA 1535	+S9	5000	26	13	13
		2500	8	14	25
		1000 *	18	22	13
		500	12	C	19
		200	14	12	16
		100	6	10	11
	-S9	5000	1	0	0
		2500	6	1	5
		1000	11	6	8
		500	11	6	9
		200 *	11	14	16
		100	16	8	11
TA 1537	+S9	5000	4	3	2
		2500	6	2	2
		1000	5	2	3
		500	3	2	5
		200	2	8	3
		100	3	4	10
	-S9	5000	0	0	0
		2500	0	1	1
		1000	2	1	3
		500	1	1	2
		200	1	1	3
		100	4	1	1
TA 98	+S9	5000 *	30	45	36
		2500 *	41	33	38
		1000 *	37	36	40
		500	37	26	26
		200	17	35	30
		100	34	37	25
	-S9	5000	4	2	0
		2500	11	8	6
		1000	11	19	13
		500	14	16	18
		200	16	13	13
		100	16	14	12
TA 100	+S9	5000 *	113	88	94
		2500	78	98	119
		1000 **	119	97	103
		500	104	92	81
		200 *	100	91	94
		100	86	81	86
	-S9	5000	1	0	0
		2500	6	5	5
		1000	64	72	65
		500	84	80	81
		200	86	65	84
		100	75	78	85
WP2P	+S9	5000	44	41	42
		2500	36	41	35
		1000	41	35	41
		500	37	38	35
		200	38	40	36
		100	32	33	34
	-S9	5000	19	16	10
		2500	27	28	25
		1000	30	33	27
		500	27	27	22
		200	29	24	24
		100	28	17	21
WP2P uvrA	+S9	5000 *	159	145	143
		2500**	166	162	179
		1000 *	140	164	154
		500 *	160	C	139
		200 *	152	162	135
		100	130	155	128
	-S9	5000	62	57	51
		2500	77	50	75
		1000	81	57	73
		500	101	85	76
		200	81	70	104
		100	115	77	80

 

C            denotes contaminated plate

*             Statistically significant

**          Indicative of a possible effect  

 

 

 

Test Data for Experimental Phase 2 (+S9). 

 

Strain	Activation	Dose Levels (micrograms/plate)	No revertants/plate
			Plate 1	Plate 2	Plate 3
TA 1535	+S9	5000	14	11	17
		2500*	18	17	12
		1000	13	6	11
		500	14	9	8
		200	6	8	11
		100	8	8	11
TA 1537	+S9	5000	1	1	2
		2500	4	2	1
		1000	3	1	4
		500	4	3	2
		200	1	4	4
		100	2	2	3
TA 98	+S9	5000	19	19	5
		2500	12	34	29
		1000	21	28	29
		500	34	24	28
		200	34	21	29
		100	18	16	19
TA 100	+S9	5000	87	72	81
		2500	62	68	76
		1000	96	57	79
		500	68	80	73
		200	73	61	72
		100	79	69	82
WP2P	+S9	5000	36	22	22
		2500	24	33	27
		1000	44	40	30
		500	28	34	28
		200	36	42	28
		100	22	33	34
WP2P uvrA	+S9	5000	119	156	136
		2500	132	111	129
		1000	136	139	132
		500	141	132	130
		200	132	106	114
		100	146	137	125

*             Statistically significant

 

 

Test Data for Experimental Phase 2 (-S9). 

 

Strain	Activation	Dose Levels (micrograms/plate)	No revertants/plate
			Plate 1	Plate 2	Plate 3
TA 1535	-S9	5000	0	3	0
		2500*	12	9	6
		1000	8	6	9
		500	6	10	6
		200	6	10	8
		100	11	5	6
TA 1537	-S9	5000	0	0	0
		2500	0	0	1
		1000	1	1	2
		500	2	1	2
		200	1	2	2
		100	2	2	3
TA 98	-S9	5000	0	0	2
		2500	11	8	3
		1000**	22	13	25
		500	12	20	9
		200	13	6	9
		100**	21	16	17
TA 100	-S9	5000	0	0	0
		2500	4	16	18
		1000	74	71	76
		500	65	72	87
		200	72	76	84
		100	76	67	79
WP2P	-S9	5000	16	21	16
		2500	17	24	20
		1000	28	22	19
		500	36	44	37
		200	36	37	36
		100	29	27	34
WP2P uvrA	-S9	5000	64	73	54
		2500	70	75	67
		1000	120	123	79
		500	97	115	106
		200	109	128	113
		100	99	103	103

**           Statistically significant

**          Indicative of a possible effect  

Applicant's summary and conclusion

Conclusions:

Interpretation of results:
negative with metabolic activation
negative without metabolic activation

Under the conditions of this assay, L-2-Chloropropionic acid did not induce any significant, reproducible increases in the number of revertant colonies. Therefore it is concluded that the substance gave a negative i.e. non-mutagenic response in the presence and absence of an auxiliary metabolising system (S9).

Executive summary:

A study was conducted to evaluate the effect of L-2-Chloropropionic acid in a guideline bacterial mutagenicity assay. An initial assay was performed using the standard plate incorporation protocol over a dose range of 5000 – 100 micrograms/plate, both with and without the presence of S9 metabolic activation. Four Salmonella strains (TA1535, TA1537, TA98 and TA100) and two E.coli strains (WP2P and WP2P uvrA) were tested.

The substance was re-tested with all six strains over the same dose range. The +S9 phase of the second assay was conducted using the pre-incubation protocol, the incubation period being 3 days at 37^oC.

For each experiment positive controls were tested to validate the bacterial strains and confirm the activity of the S9 mix.

Under the conditions of the assay, L-2-Chloropropionic acid did not induce any significant, reproducible increases in the number of revertant colonies. It was concluded that the substance gave a negative i.e. non-mutagenic response with S. typhimurium strains TA1535, TA2537, TA98 and TA100 and E.coli strains WP2P and WP2P uvrA, in the presence and absence of an auxiliary metabolising system (S9).