Registration Dossier

Diss Factsheets

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Ames-test: negative (BASF SE, 2009)
In vitro chromosome aberration test: negative (BASF SE, 2010)
HPRT-Assay: negative (Harlan, 2013)
In vivo micronucleus assay: negative (BASF SE, 2010)

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

Genetic toxicity in vivo

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

Additional information

In vitro:

The test substance was not mutagenic in a Ames test with and without metabolic activation according to OECD test guideline 471 (tested up to 250-5000 μg/plate in Salmonella typhimurium TA1535, TA 1537, TA 98 and TA 100 as well as E. coli WP2 uvrA; metabolic activation: S-9 mix; BASF SE 2009). The test was performed as independent standard plate test and preincubation test.

A bacteriotoxic effect was observed depending on the strain and test conditions from about 125 ug/plate onward using Salmonella

strains. With E. coli WP2 uvrA bacteriotoxicity was occasionally observed from 2500 ug/plate onward.

The test substance was analysed in a in vitro chromosome aberration test according to OECD 473 guideline using V79 cells in the presence and absence of metabolic activiation (BASF SE, 2010).

In Experiment I in the absence of S9 mix clear reduced cell numbers (25.6 % of control) were observed at the highest applied concentration (50 µg/mL). In the presence of S9 mix concentrations showing clear cytotoxicity were not evaluable for cytogenetic damage. However, at the highest evaluable concentration (312.5 µg/mL) the mitotic index was reduced to 59.2 % of control.

In Experiment II in the absence and presence of S9 mix concentrations showing clear cytotoxicity were not evaluable for cytogenetic damage. However, in the absence of S9 mix at preparation interval 18 hours the cell numbers were reduced to 61 % of solvent control at a concentration of 50 µg/mL At a preparation interval of 28 hours the cell numbers were reduced to 66.2 % of the solvent control in absence and to 71.3 % of the solvent control in presence of metabolic activation at concentrations of 50 µg/mL and 312.5 µg/mL, respectively.

In both independent experiments no clastogenicity was observed at the concentrations evaluated for cytogenetic damage either with of without metabolic activation. The aberration rates after treatment with the test item were within the laboratory’s historical solvent control data range (0.0 – 4.0 % aberrant cells excluding gaps).

In conclusion, it can be stated that under the experimental conditions reported, the test item did not induce structural chromosome aberrations in V79 cells in vitro.

The study was performed to investigate the potential of Kerocom FM 38 to induce gene mutations at the HPRT locus in V79 cells of the Chinese hamster according to OECD 476 guideline and GLP (Harlan, 2013).

The assay was performed in two independent experiments, using two parallel cultures each. The first main experiment was performed with and without liver microsomal activation and a treatment period of 4 hours. The second experiment was performed with a treatment time of 4 hours with and 24 hours without metabolic activation.

The maximum concentration of the pre-experiments (5.0 μL/mL) was chosen with respect to the current OECD guideline 476 regarding the solubility properties of the test item. The concentration range of the main experiments was limited by cytotoxic effects. DMSO was used as solvent.

No substantial and reproducible dose dependent increase of the mutation frequency was observed up to the maximum concentration with and without metabolic activation.

Appropriate reference mutagens (EMS and DMBA), used as positive controls, induced a distinct increase in mutant colonies and thus, showed the sensitivity of the test system and the activity of the metabolic activation system.

In conclusion it can be stated that under the experimental conditions reported the test item did not induce gene mutations at the HPRT locus in V79 cells. Therefore, Kerocom FM 38 is considered to be non-mutagenic in this HPRT assay.

In vivo:

The potential of the test substance to induce micronuclei in polychromatic erythrocytes (PCE) in the bone marrow of the mouse was inverstigated (OECD 474 guideline).

The test item was formulated in corn oil, which was also used as vehicle control. The volume administered orally was 10 mL/kg b.w.. 24 h and 48 h after a single administration of the test item the bone marrow cells were collected for micronuclei analysis.

Seven males per test group (except the vehicle and positive control groups with 5 males each) were evaluated for the occurrence of micronuclei. Per animal 2000 polychromatic erythrocytes (PCEs) were scored for micronuclei.

To describe a cytotoxic effect due to the treatment with the test item the ratio between polychromatic and normochromatic erythrocytes was determined in the same sample and reported as the number of PCEs per 2000 erythrocytes.

The following dose levels of the test item were investigated:

24 h preparation interval: 500, 1000, and 2000 mg/kg b.w..

48 h preparation interval: 2000 mg/kg b.w..

The highest dose (2000 mg/kg; maximum guideline-recommended dose) was estimated

by a pre-experiment to be suitable.

Since no gender specific differences concerning signs of toxicity in the pre-experiment were observed, the main study was performed using males only.

After treatment with the test item the number of PCEs was not substantially decreased as compared to the mean value of PCEs of the vehicle control thus indicating that the test substance did not exert any cytotoxic effects in the bone marrow.

In comparison to the corresponding vehicle controls there was no biologically relevant or statistically significant enhancement in the frequency of the detected micronuclei at any preparation interval after administration of the test item and with any dose level used.

40 mg/kg b.w. cyclophosphamide administered orally was used as positive control which showed a substantial increase of induced micronucleus frequency.

Justification for classification or non-classification

No classification and labeling for mutagenic effects is necessary according to 67/548/EWG and 1272/2008/EG (CLP)

as the test substance was not mutagenic in an Ames Test and HPRT Test, was not clastogenic in an in vitro chromosome aberration test and did not induce micronuclei in bone marrow cells of male NMRI mice.