DNA-damaging potential and glutathione depletion of 2-cyclohexene-1-one inmammalian cells, compared to food relevant 2-alkenals

2-Cyclohexene-1-one (CHX) occurs as a natural ingredient in some tropical f ruits and has been detected as a contaminant in certain artificially sweete ned soft drinks. To elucidate its cytotoxic/genotoxic effectiveness, CHX wa s tested in mammalian cell lines (V79 and Caco-2) and in primary human colo n cells in comparison to structurally related 2-alkenals. Inhibition of cel l growth (IC50) and cytotoxicity (LC50) were determined by protein staining with sulforhodamin B (SRB) and by trypan blue exclusion, respectively. DNA damage - both strand breaks and oxidised purines - was quantified by comet assay. Depletion of glutathione was measured in a kinetic assay, based on 5-thio-2-nitrobenzoate (TNB) formation. For CHX, a moderate cytotoxicity wa s observed after 1 h incubation in V79 cells (LC50: 4.75 mM). The 2-alkenal s ((E)-2-octenal (OCTE), (2E,4Z)-2,4-hexadienal (HEXDI), (E)-2-nonenal (NON E), (2E,6Z)-2,6-nonadienal (NONDI)) exhibited a distinctly higher cytotoxic ity, except for (E)-2-hexenal (HEX) (LC50: 3.67 mM) and cinnamaldehyde (CA) (LC50: 4.45 mM). If the incubation time was prolonged to 24 h, an IC50 of 15 muM was obtained for CHX which is well within the range obtained for the 2-alkenals (4 and 17 muM). Concentration-dependent DNA damage was observed after I h incubation with CHX. The respective DC50 values (concentration i nducing DNA damage in 50% of cells) were 272 muM (V79) and 455 muM (Caco-2) . All 2-alkenals were more active under these conditions, except for CA. In primary human colon cells, CHX (800 muM, 30 min) exhibited a weak, but sti ll significant DNA-damaging potential. Glutathione levels in V79 cells were effectively depleted (down to approximate to 20%) by CHX concentrations no t yet inducing DNA damage (c less than or equal to 50 muM). Incubation with CHX or 2-alkenals (50 and 100 muM, 1 h), followed by H2O2 treatment (5 min , 25 muM) resulted in increased levels of oxidised purines in the modified comet assay. CHX and HEX, additionally tested in primary human colon cells, depleted glutathione and increased the sensitivity towards oxidative stres s. (C) 2001 Elsevier Science B.V. All rights reserved.