Pentan-3-one

Endpoint summary

Additional information

Data available for diethyl ketone

The diploid Saccharomyces cerevisiae strain D61.M was used to study the induction of mitotic chromosomal malsegregation, mitotic recombination and point mutation. Diethyl ketone was tested at concentrations of 0.99, 1.23 or 1.48 % in the absence of a metabolic activation system. The highest test concentration induced aneuploidy and a low level of (Zimmermann et al., 1985) recombination and point mutation. However, the treatment protocol included a storage of the exposed cells on ice for 17 hours and thus, deviates significantly from treatment protocols specified in common guidelines on genotoxicity tests using Saccharomyces cerevisiae, e.g. OECD test guideline 480 and 481. Moreover, the test concentration leading to genotoxic effects (1.48 %) corresponds to 12.1 mg/mL, which greatly exceeds the limit concentration of 5 mg/mL generally used in in vitro genotoxicity testing. Thus, the study is not reliable. The positive outcome of the test is a result of artificial test conditions and does not reflect a biologically relevant genotoxic potential of the test item.

In a further test by Zimmermann et al. (1988), Saccharomyces cerevisiae strain D61.M was exposed to DEK at concentrations of 0, 1.28, 1.38, 1.48, 1.57, 1.67, 1.77 % without metabolic activation. The purpose of the study was to examine the influence of incubation temperature on the potential of DEK to induce aneuploidy. An induction of aneuploidy was noted only at incubation temperatures below the range of 28 to 37°C specified in the OECD test guidelines for the evaluation of genetic toxicity in Saccharomyces cerevisiae (OECD test guideline 480 and 481). When the test was performed at 28°C no induction of aneuploidy was found even though all test item concentrations were clearly above the limit concentration of 5 mg/mL generally used in in vitro genotoxicity testing. Thus, DEK did not induce aneuploidy in Saccharomyces cerevisiae strain D61.M when incubation conditions were comparable with the conditions described in OECD test guideline 480 or 481.

 

There are no further studies available for diethyl ketone. However, the experimental data presented below on the structural analogue methyl ethyl ketone can be used in a read-across approach. A justification of the read-across is given in section 13 of the IUCLID file.

 

In vitro data available for methyl ethyl ketone

 

Ames Test

In a reverse gene mutation assay the bacteria strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 of S. typhimurium as well as the E. coli strains WP2 and WP2 uvrA were exposed to methyl ethyl ketone (MEK) up to concentrations of 4000 µg/plate (3 plates/dose) in the presence and absence of mammalian metabolic activation applying the plate incorporation method (Brooks et al., 1988). The positive controls induced the appropriate responses in the corresponding strains. There was no evidence of induced mutant colonies over background in all strains with and without metabolic activation.

 

In a reverse gene mutation assay the bacteria strains TA 1535, TA 1537, TA 97, TA 98 and TA 100 of S. typhimurium were exposed to methyl ethyl ketone concentrations of 0, 100, 333, 1000, 3333, or 10000 µg/plate in the presence and absence of mammalian metabolic activation (10 % or 30 % induced male Sprague Dawley rat liver S9 and 10 % or 30 % induced male Syrian hamster liver S9) applying the preincubation method (NTP, 1986 ). Neither cytotoxicity nor precipitation was seen in any of the tester strains up to the highest test concentration. The positive and solvent (water) controls induced the appropriate responses in the corresponding strains. There was no evidence of induced mutant colonies over background in all strains with and without metabolic activation. Thus, methyl ethyl ketone was not mutagenic in this study.

 

The negative result of both assays is supported by another reverse gene mutation assay reported by O'Donoghue et al. (1988).The bacteria strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 of S. typhimurium were exposed to methyl ethyl ketone at concentrations of 0.05, 0.1, 0.5, 2.0, 16.0, or 32.0 µL/plate (at least 3 plates/dose) in the presence and absence of mammalian metabolic activation applying the preincubation method. Concentrations were based on the results of a preliminary cytotoxicity assay. The positive and solvent (DMSO) controls induced the appropriate responses in the corresponding strains. There was no evidence of induced mutant colonies over background in all strains with and without metabolic activation when tested up to cytotoxic concentrations.

 

Mouse Lymphoma Assay

In a mammalian cell gene mutation assay for the TK locus L5178Y cells were exposed to methyl ethyl ketone at concentrations of 0, 0.67, 0.89, 1.2, 1.6, 2.1, 2.8, 3.8, 5.0, 6.7, 8.9, 12.0, 16.0 or 21.0 µL/mL in the presence and absence of mammalian metabolic activation (S9 -mix) (O'Donnoghue et al., 1988). Concentrations above 8.9 µg/mL in the absence and 12 µg/mL in the presence of metabolic activation resulted in no cell survival. Relative total growth was 46 % and 30 % at the highest evaluable concentration in the absence and presence of metabolic activation, respectively. There was no evidence of induced mutant colonies over background. The positive controls induced the appropriate response. In conclusion, methyl ethyl ketone was not mutagenic in this assay in the absence and presence of metabolic activation.

 

Unscheduled DNA Synthesis

In an unscheduled DNA synthesis assay (O'Donoghue et al., 1988) primary rat hepatocyte cultures were exposed to methyl ethyl ketone at concentrations of 0, 0.1, 0.5, 1.0, 2.5, 5.0 µL/mL for 18 hours. MEK was tested up to cytotoxic concentrations. At the highest test concentration of 5 µg/mL the relative survival rate was reduced to 12.7 %. There was no evidence that unscheduled DNA synthesis, as determined by radioactive tracer , was induced. The positive control substance 2 -AAF induced an appropriate response.

 

Chromosome aberration

An in vitro assay for chromosomal damage was conducted in cloned Chinese hamster ovary cells (CHO-W-B1) to identify the potential of methyl ethyl ketone (MEK) of inducing chromosomal aberrations (NTP, 1985). CHO cells were exposed to MEK at concentrations of 0, 2500, 3750 or 5000 µg/mL with and without metabolic activation. The positive (Mitomycin C and Cyclophosphamide) controls induced an appropriate response.

The trial with no activation was evaluated by the NTP assessors as questionable since a positive trend (P < 0.003) was observed in the absence of a statistically-significant increase in chromosomal aberrations at any one dose point when compared with controls. The result of the second trial employing S-9 activation was negative since there was no positive dose response or significant differences compared to the vehicle control. Overall, the assay was assessed as negative.

 

This result is supported by the result of a chromosome damage assay where rat liver cells of the cell line RL4 were exposed to MEK for 22 hours up to concentrations of 1000 µg/mL (Brooks, 1988). There was no evidence of induced chromosome damage.

 

Sister Chromatide Exchange

An in vitro assay for sister chromatide exchange (SCE) was conducted in cloned Chinese hamster ovary cells (CHO-W-B1) to identify the potential of methyl ethyl ketone (MEK) of inducing chromosomal damage (NTP, 1985). CHO cells were exposed to MEK at concentrations of 0, 500, 1666.7 or 5000 µg/mL with and without metabolic activation in trial 1 and 2. The trial with no activation gave a weak positive response. The SCE frequency in the highest dose (5000 µg/mL) was about 25 % above the concurrent controls. Therefore, another trial without metabolic activation was conducted, employing doses of 0, 3000, 4000 or 5000 µg/mL. The results of the repeated trial were negative since there was no positive response compared to the vehicle control. This demonstrates that the findings in trial 1 were not biologically relevant. The positive (Mitomycin C and Cyclophosphamide) controls induced an appropriate response. Overall, MEK gave a negative response in the SCE assay with and without metabolic activation.

 

In vivo data available for methyl ethyl ketone

 

In a CD-1 mouse bone marrow micronucleus assay 5 animals/sex were treated by a single intraperitoneal injection with MEK at a dose of 0 or 1.96 mL/kg bw MEK in corn oil (O'Donoghue et al., 1988). The test item dose was selected as the maximum tolerated dose (LD20) based on a preliminary toxicity study. Bone marrow cells were harvested at 12, 24 or 48 hours post-treatment, stained and scored for the presence of micronuclei. There was not significant increase in the frequency of micronucleated polychromatic erythrocytes in bone marrow in males or female mice. The positive control (TME) induced the appropriate response.

 

In a Chinese hamster bone marrow micronucleus assay groups of 5 male and female animals were treated by intraperitoneal injection with methyl ethyl ketone at a dose of 0 or 411.0 mg/kg bw in corn oil (Basler, 1986). Bone marrow cells were harvested at 12, 24, 48 or 72 hours post-treatment, stained and scored for the presence of micronuclei. Methyl ethyl ketone did not increase the number of micronuclei in polychromatic erythrocytes. No mutagenic response was observed in the samples of the bone marrow taken 12, 24, 48 or 72 h after the treatment. The positive control (Vindesine) induced the appropriate response.

Short description of key information:
In a yeast gene mutation assay without metabolic activation system diethyl ketone exhibited no genotoxic effects at culture conditions as specified in OECD test guideline 480 and 481.
The read-across to the analogue substance methyl ethyl ketone (MEK) revealed no genotoxic potential in in vitro (Ames, chromosomal aberration, unscheduled DNA synthesis, mouse lymphoma, sister chromatide exchange assay) and in vivo (micronucleus test) test systems.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Based on the availabe data for diethyl ketone and the read-across to data on methyl ethyl ketone, it is concluded that diethyl ketone is not subject for classification and labelling according to Directive 67/548/EEC and Regulation 1272/2008/EC regarding mutagenicity.