Quantitative analysis by liquid chromatography-tandem mass spectrometry of glycidamide using the cob(I)alamin trapping method: validation and application to in vitro metabolism of acrylamide.

Glycidamide (GA) is the epoxy metabolite of acrylamide (AA). A sensitive analytical method for quantitative measurement of GA from in vitro metabolism studies is useful in several contexts, e.g. in studies of enzyme kinetics in different species and factors influencing the metabolism of AA to GA. It is however difficult to analyse compounds like GA, mainly due to their inherent reactivity. In the present study cob(I)alamin {Cbl(I)}, a reduced form of vitamin B(12), was used for trapping of GA. Cbl(I) can react with electrophilic species, such as an epoxide, 10(5) times faster than standard nucleophiles. The trapping of GA by Cbl(I) results in the formation of an alkylcobalamin (GA-Cbl) that was used for quantitative analysis of the epoxide. The alkylcobalamin was analysed by LC-MS/MS using an electrospray ionization source in the positive ion mode. The Cbl(I) method was validated for measurement of GA in liver S9 fractions from human and rat. GA levels down to 0.01 μM were measured in the S9 fractions, providing a sensitivity that was ca. 100 times higher than that earlier estimated by the Cbl(I) method for measurement of other (e.g. butadiene) epoxides. Compared to current analytical methods for measurement of GA, the Cbl(I) method was 10-100 times more sensitive. The method was applied to quantify GA formed from the metabolism of AA in liver S9 from human and rat.