Primary and beta-secondary deuterium isotope effects in N-deethylation reactions

Lidocaine (1), labeled specifically with deuterium in the alpha-methylene (lidocaine-d4,2) and beta-methyl (lidocaine-d6,3) carbon atoms of the terminal amino group, was used to probe the mechanism of oxidative N-deethylation by rat liver microsomes. The reaction rates were determined by measuring the formation of acetaldehyde colorimetrically. This general assay for oxidative N-deethylation reactions has the advantages of being rapid, producing a relatively stable colored derivative and being linear over the range of 0.25-4 mug of acetaldehyde formed per milliliter of incubate. Deuterium substitution at the methylene carbon atoms, the presumed site of initial oxygen insertion, revealed a kH/kD = 1.49 +/- 0.11 and a KmD/KmH = 1.23. Deuterium substitution on the terminal methyl groups showed a kH/kD = 1.52 +/- 0.10 and a KmD/KmH = 0.92. The results are explained in terms of both primary and secondary isotope effects on a possible rate-determining step in the N-deethylation sequence.