N,N'-bis(1,4-dimethylpentyl)-p-phenylenediamine

Endpoint summary

Additional information

In vitro data

The mutagenic potential in bacteria of the test substance 77PD was evaluated in a GLP study (Monsanto Co. 1986). Here, the tester strains Salmonella typhimurium TA 98, TA 100, TA 1535, TA and 1537 were used. A dose range finding test was conducted using tester strain TA100 with and without metabolic activation. Toxicity was indicated at levels of 10 µg/plate and above without metabolic activation and 200 µg/plate with metabolic activation. A second dose range finding test was performed using a maximum level of 10µg/plate without metabolic activation. In this repeat toxicity test, only the maximum level tested, 10 µg/plate, was toxic. Based on these findings the maximum levels tested in the plate incorporation tests were 200µg/plate in presence of metabolic activation, and 10µg/plate in absence of metabolic activation. The maximum levels tested, were toxic in the plate incorporation assay. In this study no biologically relevant and dose dependent increases in revertants was observed in any of the tester strains evaluated with and without metabolic activation. The authors concluded that the test sample was not mutagenic towards any of the Salmonella typhimurium test strains used with and without metabolic activation.

In addition, in earlier bacterial mutation assays the test substance also indicated a non-mutagenic potential (Monsanto Co. 1976, 1977a, 1977b).

The negative findings from the bacterial mutation assays are confirmed in a mammalian cell mutation assay. The test substance 77PD was negative in a HGPRT assay, done with CHO cells (Monsanto Co. 1986). Initial cytotoxicity experiments were conducted with 77PD in CHO cells at different S9 concentrations. In the absence of S9, 77PD was found to be significantly cytotoxic (>50 % cell killed) at 10 µg/ml. In the presence of various levels of S9 activation, 77PD was observed to be significantly cytotoxic at levels of 3 and 7 µg/ml with 1% and 2% S9, whereas at 5% and 10 % S9 cytotoxicity was significant at 30µg/ml.

An initial experiment to determine the potential mutagenicity of the test material was conducted using a range of S9 concentrations. In this experiment without metabolic activation 77PD was significantly cytotoxic at concentrations of 5µg/ml and above. In the presence of S9 activation, significant cytotoxicity was demonstrated for treatment levels of 10µg/ml in presence of 1% and 2% S9 and at 20µg/ml in presence of 5% S9. No significant cytotoxicity was observed when 77PD was tested at a maximum level of 30µg/ml in presence of 10% S9, but the relative survival was 0.51 compared to control. No statistically significant increases in mutation frequency were observed in any of the 77PD treated cultures.

The non-mutagenicity of 77PD was confirmed by a subsequent experiment. 77PD was tested at 1, 3, 5, 6, and 7.5µg/ml in the absence of exogenous metabolic activation and at 5, 10, 15, 20, and 30µg/ml in the presence of 5% S9. In the absence of S9 activation 77PD was observed to be significantly cytotoxic at levels of 6µg/ml and above. In the presence of 5% S9 the test sample was found to be significantly cytotoxic at levels of 20µg/ml and higher. No statistically significant increases in mutant frequency were observed in this experiment.

The authors concluded that the test substance 77PD was not mutagenic in CHO cells under the experimental conditions used.

Moreover, no genotoxicity was indicated in an in vitro rat hepatocyte DNA repair assay (Monsanto Co. 1986).

A chromosomal aberration test with CHO cells was performed to determine the clastogenic potential of 77PD (NTP 1989). CHO cells were treated with 5, 10 and 16 µg/ml test substance with and without metabolic activation. Harvest times were 12 hours in absence of metabolic activation. In presence of metabolic activation cells were treated with the test substance for two hour and harvest after 13 hours. Cells were harvested by mitotic shake-off, fixed and stained with Giemsa. Two hundred first-division metaphase cells were scored at each dose level, except the highest dose level without metabolic activation. Chromosomal aberration data are presented as the percentage of cells with aberrations. Statistical analysis was performed for trend analysis and statistically significance compared to solvent control. No statistically significant increase of aberrant cells was seen neither in presence nor absence of metabolic activation. The test substance 77PD was assessed to be negative in the in vitro chromosomal aberration assay under the experimental conditions used.

In addition, 77PD was stated to be negative in a sister chromatid exchanges assay in CHO cells (NTP 1989).

In vivo data

No data available.

Short description of key information:
The data from the bacterial mutation assays indicated no genotoxic potential of 77PD (Monsanto Co. 1986, Monsanto Co. 1976, 1977). This negative finding is confirmed by the results from a mammalian cell mutation assays (Monsanto Co. 1986). In addition, no genotoxicity of 77PD was indicated in an in vitro chromosomal aberration assay (NTP 1986) and sister chromatid exchange assay (NTP 1986).

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

No classification is required according to the classification criteria 67/548/EWG and regulation no. 1272/2008 (GHS).