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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Test item, pine tar was tested for their capacity to induce reverse mutation in four Salmonella typhimurium strains and one Escherichia coli strain. This study was performed in the absence and presence of metabolic activation. Two independent assays were carried out. For the two assays, negative and positive controls were carried out in parallel.

Doses (5 000, 1 500, 500, 150 and 50 μg/plate) prepared from solutions of the test item Pine tar / Tar, Pinus sylvestris, do not induce any mutagenic change in Salmonella typhimurium TA 1535, TA 1537, TA 98, TA 100 and in Escherichia coli WP2(uvrA¯) (pKM 101) without, or with metabolic activation, according to the OECD Guidelines n° 471.

Link to relevant study records
Reference
Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
The experiment was conducted between 14.3.-21.3.2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
Bacterial reverse mutation test using Salmonella typhimurium and Escherichia coli
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay
Test concentrations with justification for top dose:
A stock solution for each assay was prepared at 100 mg/mL in DMSO. The highest dose tested was 5 000 μg / plate.
Doses of 5 000, 1 500, 500, 150 and 50 μg/plate were used from solutions of the test item Pine tar / Tar, Pinus sylvestris
Vehicle / solvent:
Vehicle used to solubilize test item ;DMSO
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
sodium azide
Details on test system and experimental conditions:
Salmonella Typhimurium strains: for each strain, 0.1 mL of the bacterial suspension containing 1-9 x109 bacteria/mL and 0.1 mL of each dilution of the original solution and 0.5 mL of sterile phosphate buffer are successively added to 2 mL of overlay agar, maintained supercooled at 45° C, containing 10 % (v/v) of a L-Histidine-D-Biotine solution (0.5 mM).
Escherichia coli strain : in a test tube 0.1 mL of the bacterial suspension containing 1-9 x 109 bacteria/mL and 0.1 mL of each dilution of the original solution and 0.5 mL of phosphate buffer are successively added to 2 mL of overlay agar maintained super cooled in 45° C containing 5% (v/v) of nutrient broth n° 2 to which are added 5 μL of a L-Tryptophane solution at 2 mg/mL.
Plates are incubated at 37° C over a 48-72 hour period. The number of revertant colonies per plate is counted.
Evaluation criteria:
Ensure that the criteria of validity of the study are well respected namely :
-the bacteriostatic activity of the highest concentration tested shall be equal to or less than 75 %,
-the spontaneous reversion rate of the absolute negative control shall comply with the historical values of the laboratory,
-the spontaneous reversion rate of the solvent shall not be statistically different from absolute negative control,
-the mean number of revertant colonies obtained for each strain and the corresponding positive control, with and/or without metabolic activation shall comply with the historical values of the laboratory.
-Negative and positive values should not show significant difference with the historical values of the laboratory (± 2 standard deviations).
Statistics:
After a 48-72 hour incubation period at 37° C, revertant colonies are counted in each plate.
Data are presented as the number of revertant colonies (mean  standard deviation) per plate.
Key result
Species / strain:
S. typhimurium, other: his
Metabolic activation:
with and without
Genotoxicity:
not determined
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Doses (5 000, 1 500, 500, 150 and 50 μg/plate) performed from solutions of the test item Pine tar / Tar, Pinus sylvestris, do not induce any mutagenic change in Salmonella typhimurium TA 1535, TA 1537, TA 98, TA 100 and in Escherichia coli WP2(uvrA-) (pKM 101) without, or with metabolic activation, according to the OECD Guidelines n°471.
Remarks on result:
other: no mutagenic change
Conclusions:
Doses (5 000, 1 500, 500, 150 and 50 μg/plate) prepared from solutions of the test item Pine tar / Tar, Pinus sylvestris, do not induce any mutagenic change in Salmonella typhimurium TA 1535, TA 1537, TA 98, TA 100 and in Escherichia coli WP2(uvrA¯) (pKM 101) without, or with metabolic activation, according to the OECD Guidelines n° 471.
Executive summary:

Test item, pine tar was tested for their capacity to induce reverse mutation in four Salmonella typhimurium strains and one Escherichia coli strain. This study was performed in the absence and presence of metabolic activation. Two independent assays were carried out. For the two assays, negative and positive controls were carried out in parallel.

Doses (5 000, 1 500, 500, 150 and 50 μg/plate) prepared from solutions of the test item Pine tar / Tar, Pinus sylvestris, do not induce any mutagenic change in Salmonella typhimurium TA 1535, TA 1537, TA 98, TA 100 and in Escherichia coli WP2(uvrA¯) (pKM 101) without, or with metabolic activation, according to the OECD Guidelines n° 471.

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

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Justification for classification or non-classification

Doses (5 000, 1 500, 500, 150 and 50 μg/plate) prepared from solutions of the test item Pine tar / Tar, Pinus sylvestris, do not induce any mutagenic change in Salmonella typhimurium TA 1535, TA 1537, TA 98, TA 100 and in Escherichia coli WP2(uvrA¯) (pKM 101) without, or with metabolic activation, according to the OECD Guidelines n° 471.