3-mercaptopropionic acid

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

26 April 2007 to 17 July 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study; GLP, no deviations

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report Date:

2007

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Version / remarks:

adopted 1997

Deviations:

no

Test guideline 2

Qualifier:

according to

Guideline:

EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)

Version / remarks:

adopted 2000

Deviations:

no

GLP compliance:

yes (incl. certificate)

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Reference

Name:

Unnamed

Type:

Constituent

Details on test material:

- Name of test material (as cited in study report): 3-Mercaptopropionic acid
- Physical state: Clear colourless liquid
- Analytical purity: 99.5% (GC)
- Purity test date: 2007-04-18
- Lot/batch No.: 19185
- Expiration date of the lot/batch: 2008-04-17
- Stability under test conditions: no data
- Storage condition of test material: ~4°C in the dark

Method

Species / strain

Species / strain / cell type:

lymphocytes: human

Details on mammalian cell type (if applicable):

- Type and identity of media: MEM
- Properly maintained: not applicable, primary culture
- Periodically checked for Mycoplasma contamination: not applicable, primary culture
- Periodically checked for karyotype stability: not applicable, primary culture
- Periodically "cleansed" against high spontaneous background: not applicable, primary culture

Metabolic activation:

with and without

Metabolic activation system:

S9 was prepared from the livers of male Sprague-Dawley rats weighing ~250 g. These had received three daily oral doses of a mixture of phenobarbitone (80 mg/kg) and ß-naphthoflavone (100 mg/kg), prior to S9 preparation on the fourth day.

Test concentrations with justification for top dose:

4-h exposure: 33.1 to 1060 µg/mL
24-h exposure: 33.1 to 530 µg/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: culture medium

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

without S9

Migrated to IUCLID6: 0.4 and 0.2 µg/mL for 4- and 24-h exposure

Controls 2

Positive control substance:

cyclophosphamide

Remarks:

with S9

Migrated to IUCLID6: 5 µg/mL

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium


DURATION
- Exposure duration: 4, 24 h
- Fixation time (start of exposure up to fixation or harvest of cells): 24 h


SPINDLE INHIBITOR (cytogenetic assays): Colcemid 0.1 µg/mL
STAIN (for cytogenetic assays): 5% Gurrs Giemsa


NUMBER OF REPLICATIONS: 2


NUMBER OF CELLS EVALUATED: 200 per concentration


DETERMINATION OF CYTOTOXICITY
- Method: mitotic index


OTHER EXAMINATIONS:
- Determination of polyploidy: yes
- Determination of endoreplication: yes

Evaluation criteria:

A positive response was recorded for a particular treatment if the % cells with aberrations, excluding gaps, markedly exceeded that seen in the concurrent control, either with or without a clear concentration-relationship. For modest increases in aberration frequency a concentration response relationship is generally required and appropriate statistical tests may be applied in order to record a positive response.

Statistics:

Fisher's Exact test.

Results and discussion

Test results

Species / strain:

lymphocytes: human

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

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

Remarks:

10 mM

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
The concentration range for the Preliminary Toxicity Test was 16.5 to 1060 µg/mL. The maximum concentration was the 10 mM concentration. No precipitate of the test material was observed at the end of the exposure period in any of the three exposure groups. Microscopic assessment of the slides prepared from the exposed cultures showed that metaphase cells were present up to 1060 µg/mL in the 4(20)-hour exposures both in the presence and absence of metabolic activation (S9). The maximum concentration with metaphases present in the 24-hour continuous exposure was 1060 µg/mL, though it should be noted that frequency of metaphases was very low. The test material induced some evidence of toxicity.


COMPARISON WITH HISTORICAL CONTROL DATA: within historical reference range

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

All vehicle (solvent) controls had frequencies of cells with aberrations within the range expected for normal human lymphocytes.  All the positive control materials induced statistically significant increases in the frequency of cells with aberrations indicating the satisfactory performance of the test and of the activity of the metabolising system.  The test material was moderately toxic and did not induce any statistically significant increases in the frequency of cells with aberrations, in either of two separate experiments, using a concentration range that included a concentration level that induced approximately 50% mitotic inhibition.

MPA produced some evidence of toxicity at 1060 µg/mL in the preliminary  toxicity test, as well as in the chromosome aberration test.

Table 1: Experiment 1 - 4 h treatment, 20 h fixation - Without Metabolic Activation

Concentration [µg/ mL]	Mitotic index [%]	Polyploid cells [%]	Aberrant cells (%)
			incl. gaps	excl. gaps
0	100	0.0	0.5	0.0
265	91	0.0	1.5	0.5
530	75	0.5	0.0	0.0
1060	71	0.0	2.0	1.5
MMC, 0.4	60	0.0	35.3	33.3***

MMC: Mitomycin C

*** p<0.001

 

Table 2: Experiment 1- 4 h treatment, 20 h fixation - With Metabolic Activation

Concentration [µg/ mL]	Mitotic index [%]	Polyploid cells [%]	Aberrant cells (%)
			incl. gaps	excl. gaps
0	100	0.0	1.0	0.5
265	100	0.0	0.5	0.5
530	69	0.0	0.5	0.5
1060	70	0.0	1.5	1.5
CP, 5	30	0.0	38.8	33.0***

CP: Cyclophosphamide

*** p<0.001

 

Table 3: Experiment 2- 20 h treatment, 20 h fixation - Without Metabolic Activation

Concentration [µg/ mL]	Mitotic index [%]	Polyploid cells [%]	Aberrant cells (%)
			incl. gaps	excl. gaps
0	100	0.5	0.5	0.5
66.25	104	0.0	2.0	0.5
132.5	82	0.0	2.5	0.5
265	36	0.0	3.0	2.0
MMC, 0.4	50	0.0	17.0	16.0***

MMC: Mitomycin C

*** p<0.001

 

 

Table 4: Experiment 2- 4 h treatment, 20 h fixation - With Metabolic Activation

Concentration [µg/ mL]	Mitotic index [%]	Polyploid cells [%]	Aberrant cells (%)
			incl. gaps	excl. gaps
0	100	0.0	0.0	0.0
265	178	0.0	2.0	1.5
530	142	0.0	0.5	0.5
1060	161	0.0	2.5	0.5
CP, 5	54	0.0	28.0	24.0***

CP: Cyclophosphamide

*** p<0.001

Applicant's summary and conclusion

Conclusions:

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

The test material is non-clastogenic to human lymphocytes in vitro, with and without metabolic activation.