Registration Dossier

Diss Factsheets

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

For the assessment of the potential to induce genetic toxicity no studies are available for the target substance, D-(-)-lactic acid. Therefore, data from suitable read-across partners, L(+)-lactic acid and lactic acid was used to assess genetic toxicity of the target substance. For details and justification of read-across please refer to the report attached in section 13 of IUCLID.

Data from an in vitro genetox testing battery conducted in accordance with OECD guidelines with the suitable read-across partner L(+)-lactic acid was used in a weight of evidence approach. L(+)-lactic acid was tested negative in a bacterial reverse gene mutation test conducted according to OECD 471, in an in vitro chromosome aberration assay conducted according to OECD 473 and in a mammalian cell gene mutation assay conducted according to OECD 476 (nowadays OECD 490). In supporting studies, the second source substance lactic acid showed no potential to induce any mutagenic or clastogenic effect. Based on results, D-(-)-lactic acid is considered to not induce genetic toxicity.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
For details and justification of read-across please refer to the read-across report attached to IUCLID section 13.
Reason / purpose for cross-reference:
read-across source
Key result
Species / strain:
lymphocytes: peripheral human lymphocytes
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
based on determination of the mitotic index.
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
True negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: At the highest concentration of the test substance (10 mM equal to 901 µg/ml) the pH was 7.1 compared to a pH of 7.8 in the solvent control.
- Effects of osmolality: At the highest concentration of the test substance (10 mM equal to 901 µg/mL) the osmolarity was 275 mOsm/kg compared to an osmolarity of 269 mOsm/kg in the solvent control.
- Water solubility: miscible
- Precipitation: No

RANGE-FINDING/SCREENING STUDIES: In the dose range finding test blood cultures were treated with 10, 33, 100, 333, 901 µg/mL L(+)-lactic acid/mL culture medium (equal to concentrations of 0.1, 0.4, 1.1, 3.7 and 10 mM) with and without S9-mix.

COMPARISON WITH HISTORICAL CONTROL DATA: The number of cells with chromosome aberrations found in the solvent control cultures was within the laboratory historical control data range.

For individual results see box 'Any other information on results incl. tables'.

Mitotic Indices:

Table1: Mitotic index of human lymphocyte cultures treated with L(+)-lactic acid in the dose range finding test

L(+)-lactic acid concentration

(µg/ml)                                     

Number of

metaphases:

 

Absolute

Number of

metaphases:

 

Number of cells scored

Number of

metaphases:

 

Percentage

of control

Without metabolic activation (-S9-mix)

 

 

 

3 h exposure time, 24 h fixation time

 

 

 

Control a)

96

1004

100

10

99

1007

103

33

80

1045

83

100

60

1008

63

333

66

1009

69

901

63

1003

66

24 h exposure time, 24 h fixation time

 

 

 

Control a)

65

1007

100

10

62

1042

95

33

71

1041

109

100

66

1016

102

333

68

1048

105

901

36

1028

55

48h exposure time,48h fixation time

 

 

 

Control a)

68

1017

100

10

64

1026

94

33

51

1013

75

100

65

1010

96

333

57

1017

84

901

33

1033

49

With metabolic activation (+S9-mix)

 

 

 

3 h exposure time, 24 h fixation time

 

 

 

Control a)

85

1044

100

10

70

1013

82

33

71

1008

84

100

68

1006

80

333

66

1020

78

901

71

1007

84

a.) culture medium

Table 2: Mitotic index of human lymphocyte cultures treated with L(+)-lactic acid in the first cytogenetic assay

L(+)-lactic acid concentration (µg/ml)

Number of metaphases a)

 

Absolute 

Number of metaphases a)

 

Number of cells scored

Percentage of control

 

Without metabolic activation (-S9-mix)

 

 

 

3 h exposure time, 24 h fixation time

 

 

 

Control b)

3-36

1009-1028

100

10

3-35

1002-1012

103

100

3-28

1008-1033

90

901

1-9

1002-1040

34

MMC-C; 0.5 µg/ml

4-7

1026-1031

16

MMC-C; 0.75 µg/ml

7-5

1029-1004

17

With metabolic activation (+S9-mix)

 

 

 

3 h exposure time, 24 h fixation time

 

 

 

Control b)

3-48

1040-1028

100

10

3-27

1013-1007

79

100

4-35

1035-1001

101

901

4-32

1008-1007

93

CP; 10 µg/ml

21-14

1005-1025

43

a)     Duplicate cultures

b)     Culture medium

Table 3: Mitotic index of human lymphocyte cultures treated with L(+)-lactic acid in the second cytogenetic assay 

L(+)-lactic acid concentration (µg/ml)

Number of

metaphasesa

 

Absolute

Number of

metaphasesa

 

Number

of cells scored

Percentage

of control

Without metabolic activation (-S9-mix)

 

 

 

24 h exposure time, 24 h fixation time

 

 

 

Control b)

90-85

1000-1000

100

100

75-83

1000-1003

90

333

67-65

1008-1000

75

666

73-66

1001-1000

79

901

39-42

1002-1000

46

MMC-C; 0.2 µg/ml

24-34

1000-1003

33

MMC-C; 0.3 µg/ml

21-33

1003-1000

31

48 h exposure time, 48 h fixation time

 

 

 

Control b)

93-88

1005-1000

100

100

71-87

1001-1000

87

333

66-51

1000-1000

65

666

34-37

1000-1002

39

901

22-24

1003-1000

25

MMC-C; 0.1 µg/ml

18-20

1002-1003

21

MMC-C; 0.15 µg/ml

17-19

1000-1004

20

With metabolic activation (+S9-mix)

 

 

 

3 h exposure time, 48 h fixation 

time

 

 

 

Control b)

88-87

1000-1000

100

10

66-75

1000-1045

81

100

62-64

1003-1005

72

901

71-63

1000-1000

77

CP; 10 µg/ml

22-18

1005-1000

-c.)

a) Duplicate cultures

b) Culture medium

c) CP was fixed after 24 hours. Therefore, the mitotic index could not be calculated as percentage of control.

Chromosome aberrations in human lymphocyte cultures treated with L(+)-lactic acid

1. Cytogenetic Assay:

Conc Exposure Time [h] Fixation Time [h] Metabolic Activation Culture Mitotic Index [%] Cells 
scored
Cells
 + gaps
Cells
- gaps
Culture medium 3 24 without A+B 100 200 0 0
10 µg/ml 3 24 without A+B 103 200 5 5
100 µg/ml 3 24 without A+B 90 200 0 0
901 µg/ml 3 24 without A+B 34 200 4 4
MMC-C 0.5 µg/ml 3 24 without A+B 16 150 76*** 76***
Culture medium 3 24 with A+B 100 200 2 2
10 µg/ml 3 24 with A+B 79 200 3 3
100 µg/ml 3 24 with A+B 101 200 2 2
901 µg/ml 3 24 with A+B 93 200 2 2
CP 10 µg/ml 3 24 with A+B 43 200 59*** 59***

*) Significantly different from control group (Chi-square test), p < 0.001

2. Cytogenetic Assay:

Conc ExposureTime [h] FixationTime [h] Metaboic Activation Culture Mitotic Index 
[%]
Cells 
scored
Cells
 + gaps
Cells
- gaps
Culture medium 3 48 with A+B 100 200 2 1
10 µg/ml 3 48 with A+B 81 200 1 1
100 µg/ml 3 48 with A+B 72 200 0 0
10 µg/ml 3 48 with A+B 77 200 3 3
CP 10 µg/ml 3 48 with A+B n.d.b 100 53*** 53***
Culture medium 24 24 without A+B 100 200 1 1
100 µg/ml 24 24 without A+B 90 200 1 1
666 µg/ml 24 24 without A+B 79 200 0 0
901 µg/ml 24 24 without A+B 46 200 3 3
MMC- 0.1 µg/ml 24 24 without A+B 33 100 53*** 53***
Culture medium 48 48 without A+B 100 200 1 1
100 µg/ml 48 48 without A+B 87 200 3 3
333 µg/ml 48 48 without A+B 65 200 2 2
666 µg/ml 48 48 without A+B 39 200 2 2
MMC- 0.1 µg/ml 48 48 without A+B 21 100 53*** 53***

bCP was fixed after 24 hours. Therefore, the mitotic index could not be calculated as percentage of control.

*) Significantly different from control group (Chi-square test), p < 0.001

Conclusions:
L(+)-lactic acid is considered to be not clastogenic in the in vitro mammalian chromosomal aberration test using human lymphocytes, with and without metabolic activation.
Executive summary:

In an in vitro cytogenicity assay conducted according to OECD guideline 473, peripheral human lymphocyte cultures were exposed to L(+)-lactic acid (90% purity), solved in RPMI 1640 cell culture medium. In the first experiment, the doses were 0, 10, 100, 901 µg/mL with and without metabolic activation. In the second experiment doses were 0, 100, 333, 666, 901 µg/mL without metabolic activation and 0, 10, 100, 901 µg/mL with metabolic activation (rat liver S9-mix).

L(+)-lactic acid was tested up to 901 µg/mL, which was cytotoxic based on determination of the mitotic index after an exposure time of 24 and 48 hours. The percentage of the mitotic index after 24 hours of 901 µg/mL was 55%, that after 48 hours of 901 µg/mL 49%. Concentrations lower than 901 µg/mL did not cause a dose-dependent decrease in the percentage of the mitotic index after 24 and 48 hours of exposure. The mitotic index after 3 hours of exposure was lower compared to control (66% in experiment 1, 84% in experiment 2) but did not reach the threshold value of 45 ± 5% according to OECD guideline 473 for cytotoxicity. Positive controls induced the appropriate responses. There was no evidence for a concentration related positive response of chromosome aberration induced over background.

This study is classified as acceptable and satisfies the requirement for the in vitro mammalian chromosomal aberration test according to OECD 473.

This information is used in a read-across approach in the assessment of the target substance. For details and justification of read-across please refer to the read-across report attached to IUCLID section 13.

Endpoint:
in vitro gene mutation study in mammalian cells
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
For details and justification of read-across please refer to the read-across report attached to IUCLID section 13.
Reason / purpose for cross-reference:
read-across source
Key result
Species / strain:
mouse lymphoma L5178Y cells
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:
valid
True negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: At the highest concentration of test substance (0.01 M equal to 901 µg/mL) the pH was 6.84 compared to a pH of 7.31 in the solvent control.
- Effects of osmolality: At the highest concentration of test substance (0.01 M equal to 901 µg/mL) the osmolarity was 0.319 Osm/kg compared to an osmolarity of 0.299 Osm/kg in the solvent control
- Water solubility: miscible
- Precipitation: No

RANGE-FINDING/SCREENING STUDIES:
Cytotoxicity data were obtained by treating 8 x 10^6 cells (10^6 cells/mL for 3 hours treatment) or 5 x 10^6 cells (1.25 x 10^5 cells/mL for 24 hours treatment) with 0, 17, 52, 164, 512 and 901 µg of test substance for 3 hours in the presence of S9-mix and for 3 and 24 hours in the absence of S9-mix.
After exposure, the cells were separated from treatment solutions centrifugation steps and re-suspended in RPM 1640 medium supplemented with 10% (v/v) inactivated horse serum (R10 medium). Cells were counted with the coulter particle counter.
For determination of the cytotoxicity, the surviving cells of the 3 hours treatment were subcultured twice. After 24 hours of subculturing, the cells were counted (day 1) and subcultured again for another 24 hours, after which the cells were counted (day 2). The surviving cells of the 24 hours treatment were subcultured once. After 24 hours of subculturing, the cells were counted. If less than 1.25 x 105 cells/mL were counted no subculture was performed.
The suspension growth expressed as the reduction in cell growth after approximately 24 and 48 hours or only 24 hours cell growth, compared to the cell growth of the solvent control, was used to determine an appropriate dose range for the mutagenicity tests.

COMPARISON WITH HISTORICAL CONTROL DATA:
Spontaneous mutation frequencies in the solvent-treated control cultures were between the minimum and maximum value of the historical control range.

Experiment 1 & 2: For individual results see Tables 3-5 in box 'Any other information on results incl. tables'.

Table 1: Dose-rangefindingtest: Cytotoxicity of L(+)-lactic acid (3 hours treatment)

Dose
(µg/mL)
Cell count after 3 hours of treatment (cells/mL x 10^5) Cell count after 24 hours of subculture (cells/mL x 10^5) Cell count after 48 hours of subculture (cells/mL x 10^5) SG(1)
x 10^5 cells/mL)
RSG (2)
%

without metabolic activation

SC 6.9 5.0 6.9 152 100
17 6.3 5.0 7.4 149 98
52 7.0 5.2 6.9 161 106
164 7.3 5.2 7.1 173 113
512 7.6 5.2 6.8 172 113
901 6.8 5.4 7.0 166 109

with metabolic activation

SC 5.3 4.9 7.8 130 100
17 5.2 5.2 7.5 130 100
52 4.2 5.2 7.6 106 82
164 4.1 5.3 7.2 100 77
512 5.0 5.1 7.5 122 94
901 4.3 5.1 7.4 104 80

Note: all calculations were made without rounding off

SC = solvent control = exposure medium

(1) = suspension growth

(2) relative suspension growth

SG= (Cell count after 3 h treatment) x (Cell count after 24 h subculture)/(Cells subcultured (at t=3 h)(1.25x10^5 c/mL)) x (Cell count after 48 h subculture)/(Cells subcultured (at t=24 h) (1.25 x 10^5 c/mL))

RSG = [SG(test)/SG(control)] x 100

Table 2: Dose-rangefindingtest: Cytotoxicity of L(+)-lactic acid (24 hours treatment)

Dose
(µg/mL)
Cell count after 24 hours of treatment (cells/mL x 10^5) Cell count after 24 hours of subculture (cells/mL x 10^5) SG(1)
x 10^5 cells/mL)
RSG (2)
%

without metabolic activation

SC

9.5 5.9 45 100
17 8.9 5.9 42 93
52 9.3 5.7 42 93
164 9.1 5.2 39 85
512 8.8 5.5 39 87
901 7.2 4.6 26 58

Note: all calculations were made without rounding off

SC = solvent control = exposure medium

(1) = suspension growth

(2) relative suspension growth

SG = (Cell count after 24 h treatment) x (Cell count after 24 h subculture)/(Cells subcultured after treatment (1.25 x 10^5 c/mL)

RSG = [SG(test)/SG(control)] x 100

Cytotoxic and mutagenic response of L(+)-lactic acid in the mouse lymphoma L5178Y test system

Abbreviations:

RSG: Relative Suspension Growth

CE: Cloning Efficiency

RS: Relative Survival

RTG: Relative Total Growth

MF: Mutation Frequency per 10^6 Survivors

SC: Solvent Control (= Exposure Medium)

MMS: Methylmethanesulfonate

CP: Cyclophosphamide

Experiment 1

Table 3: 3 h treatment, without metabolic activation

Dose
[µg/mL]
RSG
[%]
CEday2
[%]
RSday2
[%]
RTG
[%]
MF
total
MF
small
MS
large
SC1 100 97 100 100 89 70 16
SC2 100 80 100 100 86 66 18
0.54 107 86 98 105 98 71 25
1.7 118 79 89 106 98 72 23
5.4 126 83 93 117 94 62 28
17 129 77 87 112 122 91 26
52 108 75 84 91 124 97 23
164 112 78 88 99 104 66 34
512 106 72 82 87 147 99 41
901 101 88 99 100 116 89 23
MMS 79 41 47 37 1149 870 191

Table 4: 3 h treatment, with metabolic activation

Dose
[µg/mL]
RSG
[%]
CEday2
[%]
RSday2
[%]
RTG
[%]
MF
total
MF
small
MS
large
SC1 100 68 100 100 51 26 23
SC2 100 64 100 100 55 30 25
0.54 92 78 118 108 53 31 20
1.7 78 111 168 132 26 17 8
5.4 56 93 140 79 38 28 10
17 60 97 146 88 31 22 9
52 65 66 100 65 62 47 14
164 92 74 111 102 53 45 7
512 64 77 116 74 45 12 32
901 93 81 123 114 45 25 19
CP 37 29 43 16 849 647 167

Experiment 2

Table 5: 24 h treatment, without metabolic activation

Dose
[µg/mL]
RSG
[%]
CEday2
[%]
RSday2
[%]
RTG
[%]
MF
total
MF
small
MS
large
SC1 100 98 100 100 57 26 30
SC2 100 102 100 100 50 19 30
0.54 92 84 84 77 63 19 42
1.7 91 86 86 78 71 40 28
5.4 99 89 89 88 65 25 38
17 90 89 89 80 49 11 38
52 86 98 98 84 51 16 34
164 85 88 87 74 72 34 36
512 78 90 90 71 50 22 26
901 64 107 107 68 53 9 43
MMS 80 61 61 49 621 198 368
Conclusions:
In conclusion, L(+)-lactic acid is considered to be non-mutagenic in the in vitro mammalian cell gene mutation test (OECD 476, nowadays OECD 490) in the presence and absence of mammalian metabolic activation.
Executive summary:

In a mammalian cell gene mutation assay conducted in accordance to OECD guideline 476 (nowadays OECD 490), L5178Y mouse lymphoma cells cultured in vitro were exposed to L(+)-lactic acid (90% purity), solved in RPMI 1640 medium. In the first experiment, L(+)-lactic acid was tested up to concentrations of 901 µg/mL (0.01 M, the highest concentration recommended in the guidelines) in the absence and presence of S9-mix. The incubation time was 3 hours. In the second experiment, L(+)-lactic acid was again tested up to concentrations of 901 µg/mL in the absence S9-mix. The incubation time was 24 hours. No toxicity was observed at this dose level in the absence and presence of S9 mix. The induced mutation frequency with and without metabolic activation was not increased compared to control in all tested concentrations. The positive controls did induce the appropriate response. Based on the results, it can be concluded, that L(+)-lactic acid is not mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions described.

This information is used in a read-across approach in the assessment of the target substance. For details and justification of read-across please refer to the read-across report attached to IUCLID section 13.

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
For details and justification of read-across please refer to the read-across report attached to IUCLID section 13.
Reason / purpose for cross-reference:
read-across source
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
True negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
True negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
True negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
True negative controls validity:
not examined
Positive controls validity:
valid
Key result
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:
not examined
True negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No

RANGE-FINDING/SCREENING STUDIES: Dose range finding test performed with TA100 and WP2uvrA. Doses tested: 0, 3, 10, 33, 100, 333, 1000, 3330, 5000 µg/plate. Based on the results, the following doses were selected for the main experiment with TA1535, TA1537 and TA98 in the presence and absence of S9-mix: 100, 333, 1000, 330 and 5000 µg/plate.

COMPARISON WITH HISTORICAL CONTROL DATA: The negative control values were within the laboratory historical control data ranges, except for TA100 in the absence of S9-mix, second experiment. Evaluation: The mean plate count (146) was just outside the limit of the range (144) and clear negative results are observed in all experiments. Therefore, this deviation in the mean plate count of the solvent control had no effect on the results of the study. The strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly, except the response for TA1535 in the absence of S9-mix, first experiment. Evaluation: The value (257) was just below the limit of the range (262). The purpose of the positive control is as a reference for the test system, where a positive response is required to check if the test system functions correctly. Since the value was more than 3 times greater than the concurrent solvent control values, this deviation in the mean plate count of the positive control had no effect on the results of the study.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
No reduction of the bacterial background lawn and no biologically relevant decrease in the number of revertants were observed.

MUTAGENICITY:
Experiment 1: No increase in the number of revertants was observed upon treatment with the test item under all conditions tested.
Experiment 2: Based on the results from the first experiment, the test item was tested up to 5000 µg/plate. No increase in the number of revertants was observed.
Conclusions:
In conclusion, L(+)-lactic acid is not genotoxic in the bacterial reverse gene mutation assay (OECD 471) in the presence and absence of mammalian metabolic activation.
Executive summary:

In a reverse gene mutation assay in bacteria conducted according to OECD guideline 471, Salmonella typhimurium strains TA1535, TA1537, TA98, TA100 and E. coli strain WP2uvrA were exposed to L(+)-lactic acid (90% purity) at concentrations of 0, 100, 333, 1000, 3330 and 5000 µg/plate in the presence and absence of mammalian metabolic activation.

L(+)-lactic acid was tested up to the limit concentration of 5000 µg/plate. The positive controls induced the appropriate responses in the corresponding strains. There was no evidence of induced mutant colonies over background. Based on the results, the test item can be considered to be non-mutagenic.

This study is classified as acceptable and satisfies the requirement for Test Guideline OECD 471 for in vitro mutagenicity (bacterial reverse gene mutation assay).

This information is used in a read-across approach in the assessment of the target substance. For details and justification of read-across please refer to the read-across report attached to IUCLID section 13.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

For the assessment of the potential to induce genetic toxicity no studies are available for the target substance, D-(-)-lactic acid. Therefore, data from suitable read-across partners, L(+)-lactic acid and lactic acid was used to assess genetic toxicity of the target substance. For details and justification of read-across please refer to the report attached in section 13 of IUCLID.

L(+)-lactic acid was tested in an in vitro test battery, which were performed in accordance to the OECD testing guidelines 471 (Ames test), 473 (chromosome aberration) and 476 (MLA, nowadays OECD 490).

In the bacterial reverse mutation test four strains of Salmonella typhimurium, TA1535, TA1537, TA100 and TA98 plus the Escherichia coli strain WP2uvrA were treated with L(+)-lactic acid up to the limit concentration of 5000 µg/plate. No increase in the rate of revertant colonies was observed in all tested concentrations (0, 100, 333, 1000, 3330 and 5000 µg/plate) with and without addition of S9-mix. Thus, L(+)-lactic acid can be considered as not mutagenic in the bacterial reverse mutation assay.

In a gene mutation assay conducted according to OECD 476 (nowadays OECD 490), L5178Y mouse lymphoma cells were exposed for 3 hours (with and without S9 mix) and 24 hours (without S9 mix). Eight doses between 0.54 and 901 µg/mL L(+)-lactic acid were tested, not resulting in an increase of mutation frequency per 10^6 survivors in the presence or absence of metabolic activation with 3 or 24 hours exposure time. Thus, L(+)-lactic acid is considered as not mutagenic.

The third in vitro test was a chromosome aberration test conducted according to OECD 473. Five doses of L(+)-lactic acid were tested ranging from 10 to 901 µg/mL in peripheral human lymphocytes. Assessment of the results showed no significant increase over control levels for all exposure and fixation times tested in the presence and absence of metabolic activation.

In addition to these three OECD guideline studies with the source substance L(+)-lactic acid, five publications report further results on the genotoxic potential of the second source substance lactic acid. No positive results were reported, with exception of Morita (1990), where some chromosome aberrations had been found under non-physiological conditions, i.e. low pH. The same study reports negative findings under physiological conditions, whereas the positive finding is of no toxicological relevance.

Therefore, in view of all genotoxicity tests, the target substance D-(-)-lactic acid is considered as not genotoxic.

Justification for classification or non-classification

Based on the available results, the target substance D-(-)-lactic acid is not considered to be genotoxic and no classification is warranted in accordance with CLP Regulation 1272/2008.