fatty acid chloride, C12, reaction product with sodium glutamate and sodium hydroxide

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1992

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The concentration used for the test was selected only based on the cytotoxicity, but not based on the precipitation.The results obtained at concentrations with precipiations are not considered as valid.

Cross-reference

Reason / purpose for cross-reference:

assessment report

Reference

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:

key study

Study period:

1992

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The concentration used for the test was selected only based on the cytotoxicity, but not based on the precipitation.The results obtained at concentrations with precipiations are not considered as valid.

Justification for type of information:

Justification for read-across is provided in chapter 13 as separate statement

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Version / remarks:

adpted July 21, 1997

GLP compliance:

yes (incl. QA statement)

Type of assay:

other: in-vitro chromosome aberration

Species / strain / cell type:

lymphocytes:

Metabolic activation:

with and without

Metabolic activation system:

rat liver S9

Test concentrations with justification for top dose:

The highest dose selected was the dose causing 50% reduction of mitotic indices of control.

Vehicle / solvent:

water

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

ethylmethanesulphonate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in culture medium. The chromosomes were prepared 22 hours after start of treatment with the test item. Evaluation of two cultures per dose group.
DURATION
- Exposure duration: 4 hours (+/- S9 mix) and 22 hours (- S9 mix)
- Fixation time (start of exposure up to fixation or harvest of cells): 22 hours
SPINDLE INHIBITOR (cytogenetic assays): Colcemid
STAIN (for cytogenetic assays): Giemsa
NUMBER OF REPLICATIONS: about 1.5
NUMBER OF CELLS EVALUATED: At least 100 per culture

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index

Species / strain:

lymphocytes:

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Species / strain:

lymphocytes:

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: The author of the study evaluated the results obtained as "positive". The registrant performed re-evaluation, taking account the precipitation in the culture medium as the potential artifical factor for false-positive responce.

Summary of results of the chromosome aberration study: without metabolic activation
Exp	Preparation Interval	Test item concentration [µg/mL]	Mitotic indices in % of control	Aberrant cells		Carrying exchange	Evaluation based on the Guideline adopted July 29, 2016
				Incl. gaps*	Excl. gaps*
Exposure period 4 hrs without S9 mix
IA	22 hrs	Deionized water, 10% (v/v)	100.0	1.5	1.5	0.0	Negative: Precipitation occurred at 99.5 µg/mL, which should be considered as the valid high dose. The dose 304.6 µg/mL is not compliant to the Guideline adopted in 2016.
		EMS, 770 µg/mL	51.7	8.5	8.5^S	0.5
		56.8	82.1	0.0	0.0	0.0
		99.5^P	70.3	0.0	0.0	0.0
		304.6^P	61.8	1.0	1.0	0.0
IB	22 hrs	Deionized water, 10% (v/v)	100.0	0.5	0.5	0.0	Negative: Precipitation occurred at 50.0 µg/mL. The doses 200 and 325 µg/mLis not compliant to the Guideline adopted in 2016.
		EMS, 770 µg/mL	74.8	8.5	8.5^S	3.0
		25.0	101.0	1.5	0.5	0.0
		50.0^P	98.0	0.5	0.5	0.0
		200.0^P	83.0	2.0	2.0	0.0
		325.0^P	45.1	2.5	2.5	0.0
Exposure period 22 hrs without S9 mix
IIA	22 hrs	Deionized water, 10% (v/v)	100.0	0.0	0.0	0.0	Negative: No Precipitation observed, but the highest dose induced clear cytotoxicity.
		EMS, 660 µg/mL	38.0	21.5	20.5^S	9.0
		30.5	96.1	1.0	1.0	0.0
		53.3	82.8	1.0	1.0	0.0
		93.3	62.4	1.0	1.0	0.0
IIB	22 hrs	Deionized water, 10% (v/v)	100.0	2.5	2.0	0.0	Negative: No Precipitation observed, but the highest dose induced clear cytotoxicity.
		EMS, 550 µg/mL	43.1	22.0	21.5^S	9.0
		100.0	86.3	3.0	2.0	0.0
		140.0	56.5	1.5	1.5	0.0
		160.0	37.8	0.5	0.5	0.0
*  Including cells carrying exchanges ^P  Precipitation occurred at the end of treatment ^S  Aberration frequency statistically significant higher than corresponding control values

Summary of results of the chromosome aberration study: with metabolic activation
Exp	Preparation Interval	Test item concentration [µg/mL]	Mitotic indices in % of control	Aberrant cells		Carrying exchange	Evaluation based on the Guideline adopted July 29, 2016
				Incl. gaps*	Excl. gaps*
Exposure period 4 hrs with S9 mix
IA	22 hrs	Deionised water 10.0 % (v/v)	100.0	1.0	1.0	0.0	Negative: Precipitation occurred at 99.5 µg/mL, which should be considered as the valid high dose. The dose 304.6 µg/mL is not compliant to the Guideline adopted in 2016.
		CPA, 15.0 µg/mL	79.6	12.5	12.0^S	1.0
		56.8	100.0	0.0	0.0	0.0
		99.5^P	98.9	0.5	0.5	0.0
		304.6^P	88.9	1.0	1.0	0.0
IIA	22 hrs	Deionised water 10.0 % (v/v)	100.0	2.0	2.0	0.0	Negative: Precipitation occurred at 100 µg/mL. The dose 500 µg/mL is not compliant to the Guideline adopted in 2016.
		CPA, 15.0 µg/mL	69.9	22.5	22.0^S	5.5
		50.0	89.3	1.0	1.0	0.0
		100.0^P	93.2	2.0	2.0	0.0
		500.0^P	87.4	3.5	3.5	1.0
IIB	22 hrs	Deionized water, 10% (v/v)	100.0	1.5	1.5	0.0	Negative: Precipitation occurred at 100 µg/mL. The dose 520 µg/mL is not compliant to the Guideline adopted in 2016.
		CPA, 7.5 µg/mL	78.2	9.5	8.0^S	2.0
		50.0	72.8	2.0	1.5	0.0
		100.0^P	87.9	1.5	1.5	0.0
		520.0^P	56.8	8.0	8.0^S	2.5
IIC	22 hrs	Deionized water, 10% (v/v)	100.0	0.5	0.5	0.0	Negative: Precipitation occurred at 100 µg/mL, and the aberration cells were statistically increased. Taking account the results obtained in other three independent experiments were negative and the incidence is marginally increased, the overall assessment should be negative. The doses 400 and 440 µg/mL are not compliant to the Guideline adopted in 2016.
		CPA, 7.5 µg/mL	54.9	19.5	19.5^S	6.0
		50.0	91.7	2.0	1.5	0.0
		100.0^P^#	99.5	2.8	2.5^S	0.3
		400.0^P^#	61.1	6.3	6.0^S	1.0
		440.0^P^#	37.8	9.0	8.8^S	1.0
*  Including cells carrying exchanges ^P  Precipitation occurred at the end of treatment ^S  Aberration frequency statistically significant higher than corresponding control values ^#   Evaluation of 200 metaphases per culture

Conclusions:

The genotoxicity of the registration substance is assessed via read-across to N-cocoyl glycinate. The genotoxicity of N-cocoyl glycinate was investigated according to the Guideline OECD 473. No genotoxic activity was observed up to the insoluble concentrations. Likewise, no genotoxicity activity can be assigned in the cytogenetic assay in mammalian cells to the registration substance.

Executive summary:

The genotoxicity of the registration substance is assessed via read-across to N-cocoyl glycinate.

The genotoxicity of N-cocoyl glycinate was investigated according to the Guideline OECD 473 (version 1997). Human lymphocytes were treated with the test substance with and without metabolic activation up to the concentration associated with 50% reduction of mitotic indices. In general, reduced cytotoxicity was observed with metabolic activation, leading to the use of higher concentrations with metabolic activation for the genotoxicity investigation. These concentrations were associated with enhanced precipitation.

Total five independent experiments were performed. No increase of aberration frequency was observed without metabolic activation. Significant increase of aberration frequency was obtained with metabolic activation at concentrations associated not only with significant cytotoxicity but also with enhanced precipitation. The author of the study evaluated the results obtained as "positive".

The registrant performed re-evaluation in 2018, taking account the precipitation in the culture medium as potential interfering factor responsible for a false-positve response (according to the current OECD Guideline, version 2016). Based on the re-evaluation, no significant genotoxic activity can be assigned up to the insoluble concentrations for N-cocoyl glycinate.

Likewise, no genotoxicity activity can be assigned in the cytogenetic assay in mammalian cells to the registration substance.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Version / remarks:

adpted July 21, 1997

GLP compliance:

yes (incl. QA statement)

Type of assay:

other: in-vitro chromosome aberration

Test material

Test material information

Constituent 1

Reference substance name:

Sodium cocoyl glycinate (SCG) [INCI]

IUPAC Name:

Sodium cocoyl glycinate (SCG) [INCI]

Details on test material:

Stability of Test Item: Stable under storage conditions
Stability of Test Item Dilution: Unknown in PEG 300; excluded from the statement of compliance.
Storage Conditions: At room temperature (range of 20 ± 5 °C, provided by Harlan Laboratories Ltd.), light protected.
Safety Precautions: Routine hygienic procedures were used to ensure the health and safety of the personnel.
Description: Colorless solid

Method

Species / strain

Species / strain / cell type:

lymphocytes:

Metabolic activation:

with and without

Metabolic activation system:

rat liver S9

Test concentrations with justification for top dose:

The highest dose selected was the dose causing 50% reduction of mitotic indices of control.

Vehicle / solvent:

water

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

ethylmethanesulphonate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in culture medium. The chromosomes were prepared 22 hours after start of treatment with the test item. Evaluation of two cultures per dose group.
DURATION
- Exposure duration: 4 hours (+/- S9 mix) and 22 hours (- S9 mix)
- Fixation time (start of exposure up to fixation or harvest of cells): 22 hours
SPINDLE INHIBITOR (cytogenetic assays): Colcemid
STAIN (for cytogenetic assays): Giemsa
NUMBER OF REPLICATIONS: about 1.5
NUMBER OF CELLS EVALUATED: At least 100 per culture

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index

Results and discussion

Test results

Test results 1

Species / strain:

lymphocytes:

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

lymphocytes:

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: The author of the study evaluated the results obtained as "positive". The registrant performed re-evaluation, taking account the precipitation in the culture medium as the potential artifical factor for false-positive responce.

Any other information on results incl. tables

Summary of results of the chromosome aberration study: without metabolic activation
Exp	Preparation Interval	Test item concentration [µg/mL]	Mitotic indices in % of control	Aberrant cells		Carrying exchange	Evaluation based on the Guideline adopted July 29, 2016
				Incl. gaps*	Excl. gaps*
Exposure period 4 hrs without S9 mix
IA	22 hrs	Deionized water, 10% (v/v)	100.0	1.5	1.5	0.0	Negative: Precipitation occurred at 99.5 µg/mL, which should be considered as the valid high dose. The dose 304.6 µg/mL is not compliant to the Guideline adopted in 2016.
		EMS, 770 µg/mL	51.7	8.5	8.5^S	0.5
		56.8	82.1	0.0	0.0	0.0
		99.5^P	70.3	0.0	0.0	0.0
		304.6^P	61.8	1.0	1.0	0.0
IB	22 hrs	Deionized water, 10% (v/v)	100.0	0.5	0.5	0.0	Negative: Precipitation occurred at 50.0 µg/mL. The doses 200 and 325 µg/mLis not compliant to the Guideline adopted in 2016.
		EMS, 770 µg/mL	74.8	8.5	8.5^S	3.0
		25.0	101.0	1.5	0.5	0.0
		50.0^P	98.0	0.5	0.5	0.0
		200.0^P	83.0	2.0	2.0	0.0
		325.0^P	45.1	2.5	2.5	0.0
Exposure period 22 hrs without S9 mix
IIA	22 hrs	Deionized water, 10% (v/v)	100.0	0.0	0.0	0.0	Negative: No Precipitation observed, but the highest dose induced clear cytotoxicity.
		EMS, 660 µg/mL	38.0	21.5	20.5^S	9.0
		30.5	96.1	1.0	1.0	0.0
		53.3	82.8	1.0	1.0	0.0
		93.3	62.4	1.0	1.0	0.0
IIB	22 hrs	Deionized water, 10% (v/v)	100.0	2.5	2.0	0.0	Negative: No Precipitation observed, but the highest dose induced clear cytotoxicity.
		EMS, 550 µg/mL	43.1	22.0	21.5^S	9.0
		100.0	86.3	3.0	2.0	0.0
		140.0	56.5	1.5	1.5	0.0
		160.0	37.8	0.5	0.5	0.0
*  Including cells carrying exchanges ^P  Precipitation occurred at the end of treatment ^S  Aberration frequency statistically significant higher than corresponding control values

Summary of results of the chromosome aberration study: with metabolic activation
Exp	Preparation Interval	Test item concentration [µg/mL]	Mitotic indices in % of control	Aberrant cells		Carrying exchange	Evaluation based on the Guideline adopted July 29, 2016
				Incl. gaps*	Excl. gaps*
Exposure period 4 hrs with S9 mix
IA	22 hrs	Deionised water 10.0 % (v/v)	100.0	1.0	1.0	0.0	Negative: Precipitation occurred at 99.5 µg/mL, which should be considered as the valid high dose. The dose 304.6 µg/mL is not compliant to the Guideline adopted in 2016.
		CPA, 15.0 µg/mL	79.6	12.5	12.0^S	1.0
		56.8	100.0	0.0	0.0	0.0
		99.5^P	98.9	0.5	0.5	0.0
		304.6^P	88.9	1.0	1.0	0.0
IIA	22 hrs	Deionised water 10.0 % (v/v)	100.0	2.0	2.0	0.0	Negative: Precipitation occurred at 100 µg/mL. The dose 500 µg/mL is not compliant to the Guideline adopted in 2016.
		CPA, 15.0 µg/mL	69.9	22.5	22.0^S	5.5
		50.0	89.3	1.0	1.0	0.0
		100.0^P	93.2	2.0	2.0	0.0
		500.0^P	87.4	3.5	3.5	1.0
IIB	22 hrs	Deionized water, 10% (v/v)	100.0	1.5	1.5	0.0	Negative: Precipitation occurred at 100 µg/mL. The dose 520 µg/mL is not compliant to the Guideline adopted in 2016.
		CPA, 7.5 µg/mL	78.2	9.5	8.0^S	2.0
		50.0	72.8	2.0	1.5	0.0
		100.0^P	87.9	1.5	1.5	0.0
		520.0^P	56.8	8.0	8.0^S	2.5
IIC	22 hrs	Deionized water, 10% (v/v)	100.0	0.5	0.5	0.0	Negative: Precipitation occurred at 100 µg/mL, and the aberration cells were statistically increased. Taking account the results obtained in other three independent experiments were negative and the incidence is marginally increased, the overall assessment should be negative. The doses 400 and 440 µg/mL are not compliant to the Guideline adopted in 2016.
		CPA, 7.5 µg/mL	54.9	19.5	19.5^S	6.0
		50.0	91.7	2.0	1.5	0.0
		100.0^P^#	99.5	2.8	2.5^S	0.3
		400.0^P^#	61.1	6.3	6.0^S	1.0
		440.0^P^#	37.8	9.0	8.8^S	1.0
*  Including cells carrying exchanges ^P  Precipitation occurred at the end of treatment ^S  Aberration frequency statistically significant higher than corresponding control values ^#   Evaluation of 200 metaphases per culture

Applicant's summary and conclusion

Conclusions:

The genotoxicity of N-cocoyl glycinate was investigated according to the Guideline OECD 473. No genotoxic activity was observed up to the insoluble concentrations.

Executive summary:

The genotoxicity of N-cocoyl glycinate was investigated according to the Guideline OECD 473 (version 1997). Human lymphocytes were treated with the test substance with and without metabolic activation up to the concentration associated with 50% reduction of mitotic indices. In general, reduced cytotoxicity was observed with metabolic activation, leading to the use of higher concentrations with metabolic activation for the genotoxicity investigation. These concentrations were associated with enhanced precipitation.

Total five independent experiments were performed. No increase of aberration frequency was observed without metabolic activation. Significant increase of aberration frequency was obtained with metabolic activation at concentrations associated not only with significant cytotoxicity but also with enhanced precipitation. The author of the study evaluated the results obtained as "positive".

The registrant performed re-evaluation in 2018, taking account the precipitation in the culture medium as potential interfering factor responsible for a false-positve response (according to the current OECD Guideline, version 2016). Based on the re-evaluation, no significant genotoxic activity can be assigned up to the insoluble concentrations for N-cocoyl glycinate.