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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Under test conditions, HCFC 122a did no have mutagenic activity when tested on standard strains TA 100, TA 98, TA 97, with and without metabolic activation up to cytotoxic doses.

Link to relevant study records
Reference
Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
The data source is the competent authority and it is considered reliable. The test method is well documented and similar to OECD guideline. Although some deviations occur, the results can be considered reliable.
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
other: Guidelines for study of mutagenic activity of drugs_Ministry of Health of the USSR_January 15, 1988
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
yes
Remarks:
3 strains of salmonella t.; a partial activated metabolic system is used for the identication of direct mutagens.
Principles of method if other than guideline:
Plate incorporation method.
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay
Species / strain / cell type:
S. typhimurium, other: TA 97, TA 98, TA 100
Metabolic activation:
with and without
Metabolic activation system:
S9mix FMAM, S9mix PMAM
Test concentrations with justification for top dose:
0.01, 0.1, 1.0, 10, 100 mcg/plate
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
2-nitrofluorene
sodium azide
other: IGR 100 (191); 2-aminoanthracene
Key result
Species / strain:
S. typhimurium TA 97
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at test concentration at 10 mcg/plate and above
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at test concentration at 10 mcg/plate and above
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
at test concentration at 10 mcg/plate and above
Positive controls validity:
valid

Results of the study of mutagenic activity of R122 on the test strains

Test strain

Chemical compound

Concentration % by volume or mcg/dish

Average number of his. revertants per dish in options

FMAM

PMAM

TA 100

Control

102

94

HCFC 122a

0.01

104

100

 

0.1

90

106

 

1.0

82

101

 

10

t. e.

t. e.

 

100

t. e.

t. e.

Sodium azide

2.5

1,786

2AA

2.5

1,932

90

TA 97

Control

116

99

HCFC 122a

0.01

107

104

 

0.1

114

107

 

1.0

87

71

 

10

t. e.

t. e.

 

100

t. e.

t. e.

191

2.5

259

2AA

2.5

1,441

75

TA 98

Control

32

25

HCFC 122a

0.01

32

23

 

0.1

35

27

 

1.0

27

16

 

10

t. e.

t. e.

 

100

t. e.

t. e.

2NF

2.5

1,079

2AA

2.5

3,020

31

t. e. = toxic effect

Starting from the dose of 10 mcg/dish, the substance inhibits growth of bacteria.

Conclusions:
Under test conditions, HCFC 122a did no have mutagenic activity when tested on standard strains TA 100, TA 98, TA 97, with and without metabolic activation up to cytotoxic doses.
Executive summary:

The objective of the reported study was the assessment both of the reprotoxicity potential and of the genotoxicity potential of HCFC 122a. To the latter scope, an in vivo micronucleus test and an Ames test were performed.

Thetest was conducted in accordance with the “Guidelines for study of mutagenic activity of drugs” approved by the Ministry of Health of theon January 15, 1988, using strains of Salmonella typhimurium TA 98, TA 100, TA 97.

The presence of mutagenic effect in the preparation was determined by induction of back mutations from histidine auxotrophy to prototrophy.

HCFC 122a was tested following the plate incorporation method at the concentrations of 0.01, 0.1, 1, 10, 100 mcg/plate. The analysis of the results was performed after 48-72 hours of incubation.

 

Concurrently, the experiment included options with FMAM (S9 Full microsomal activation mixture) and PMAM (S9 Partial microsomal activation mixture).

Options with PMAM can demonstrate action of direct mutagens, i.e. drugs that exhibit mutagenic effect due to the activity of the original structure of the substance. The action of the promutagens, i.e. compounds, the effect of which is associated with the formation of mutagenic metabolites, may be taken into account when comparing the results of analysis of test substances in options FMAM and PMAM.

In the control option, FMAM and PMAM were introduced into the upper layer of a semi-solid agar together with the bacterial suspension and the corresponding volumes of solvent. The experiment was accompanied by positive controls.

3 plates were used in each control and experimental option. The experiment was repeated twice.

The results were registered in case of mutagenic effect in all positive control options.

Analysis was performed using Dunnett’s multiple comparison method.

 

No mutations in S. typhimurium test strains were observed. Positive controls effectively induced mutations in the respective S. typhimurium test strains with and without metabolic activation. Starting from the dose of 10 mcg/dish, the preparation inhibits growth of bacteria.

In conclusion, under test conditions, HCFC 122a did no have mutagenic activity when tested on standard strains TA 100, TA 98, TA 97.

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

Genetic toxicity in vivo

Description of key information

There was no increased incidence in micronuclei in the bone marrow cells of mice administered the substance twice by oral gavage at 24 hour-interval.

Link to relevant study records
Reference
Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Remarks:
Type of genotoxicity: DNA damage and/or repair
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
The data source is the competent authority and it is considered reliable. Although not all the details on materials and results are provided, the test method is similar to official guideline methods.
Reason / purpose for cross-reference:
reference to same study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
yes
Remarks:
no positive control included
GLP compliance:
not specified
Type of assay:
micronucleus assay
Species:
mouse
Strain:
not specified
Sex:
not specified
Route of administration:
oral: unspecified
Duration of treatment / exposure:
2 single administrations.
Frequency of treatment:
Twice with the intervals of 24 hours.
Post exposure period:
24 hours
Remarks:
Doses / Concentrations:
780 mg/kg
Basis:
no data
Remarks:
Doses / Concentrations:
3900 mg/kg
Basis:
no data
No. of animals per sex per dose:
7
Control animals:
yes
Tissues and cell types examined:
Erythrocytes of femoral bone marrow.
Genotoxicity:
negative
Toxicity:
not examined
Negative controls validity:
valid
Positive controls validity:
not examined

Summary of Results

Studied dose

Number of animals

Polychromatocytes

 

 

Polychromatocytes analysed

Polychromatocytes with micronuclei Mav. ± M

% tst

Control

7

7,000

0.31±0.019

 

780 mg/kg

7

7,000

0.37±0.042

1.3

3,900 mg/kg

7

7,000

0.42±0.051

2.02

Conclusions:
Under the test conditions of the reported study, HCFC 122a did not show mutagenic activity following oral administration to mice.
Executive summary:

The objective of the study was the assessment both of the reprotoxicity potential and of the genotoxicity potential of HCFC 122a. To this scope, an in vivo micronucleus test and an Ames test were performed.

 

The micronucleus test was conducted on white mice weighing 18-20 g.

HCFC 122a was administered to mice twice with the intervals of 24 hours, at doses of 1/2 and 1/10 of the LD50 = 7800 mg/Kg. The animals were sacrificed in 24 hours after the last administration.

Femoral bones were taken and the bone marrow was placed into a drop of group IV human serum inactivated at 56 °C for 2 hours and took a smear. After drying of preparations in air, the smear was consequently stained with Main-Grunwald and Giemsa stains.

Analysis of bone marrow smear was performed using the immersion microscope objective Jenaval (magnification H/100x12.5¥0.17-A). Preparations with well expanded erythrocytes, the surface of which did not have protuberances and folds were considered suitable for the analysis. 1000 polychromatocytes were counted per each animal.

HCFC 122a mutagenicity was assessed by the presence or absence of difference in the number of micronuclei between the experimental and control animals. Data are shown as a number of polychromatocytes containing micronuclei in percent. For the statistical evaluation, the original data for each animal were standardized according to the formula:

 

W = arc sin [(r+0.375)/(n+0.75)] 1/2

where r – number polychromatophils with micronuclei

           n – the total number of polychromatocytes,

and then used tst– Student’s t test.

 

The data obtained indicate that the incidence of micronuclei in the bone marrow cells of experimental mice for the studied doses was not statistically different from the control values.

Thus, the studied dosage range of HCFC 122a upon oral administration to mice in micronucleus test did not have mutagenic activity.

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

Additional information

Additional information from genetic toxicity in vivo:

One in vitro and one in vivo studies are reported. The test methods were similar to the current official guideline methods and the results can be considered reliable. Both the two studies show negative results. No genotoxic activity is expected for HCFC 122a.


Justification for selection of genetic toxicity endpoint
Although not all the details on materials and results are provided, the test method is similar to official guideline methods and the results can be considered reliable.

Justification for classification or non-classification

No genotoxic activity is expected for HCFC 122a. According to Regulation (EC) 1272/2008, no classification is required.