Acrylamid: Bildung von Glycidamid und gentoxische Effekte in genmodifizierten V79 Zellen, die CYP2E1 von verschiedenen Spezies exprimieren
There is some concern about possible health risks from dietary exposure to acrylamide (AA). AA has been shown to be neurotoxic, genotoxic and carcinogenic. In laboratory animals AA is metabolized to glycidamide (GA) catalysed primarily by CYP2E1. AA and its metabolites are rapidly eliminated in urine, mainly as mercapturic acid conjugates of AA and GA. GA is suggested to be much more reactive than AA with DNA. In a recent study we addressed the question if and to which extent GA is formed from AA in human and monkey liver microsomes as well as in genetically modified V79 cells expressing human CYP2E1. We now extended our studies using in addition genetically modified V79 cells expressing CYP2E1 from the rat or the mouse. In addition we investigated the genotoxic potency of AA and GA in these cells by using the COMET Assay. Incubations have been performed according to conventional procedures. Special emphasis was laid on the analytical detection of GA. ¶13 C-Glycidamide and D 3 -Acrylamide were added as external standards. GA and AA were extracted and refined from protein suspensions by using organic solvents and by elution through activated carbon-aluminium oxide-columns. The detection of GA was performed using a gas chromatograph/mass spectrometer (Finnigan Mat SSQ 710) equipment. The results show that AA is metabolized to GA in genetically modified V79 cells expressing CYP2E1 from different species (human, rat, mouse) at various but low amounts. Formation of GA could be inhibited by e.g. MAB-2E1 antibody (monoclonal, raised in mouse, human CYP2E1 selective) to about 80% in cells expressing human CYP2E1. Significant DNA damaging potency could not be demonstrated when cells were incubated with AA despite the fact that GA could be determined. However when GA was added in various concentrations genotoxic effects were clearly detectable.