Isotope effects in enzymatic N-demethylation of tertiary amines

The N-demethylation of 1-(N-methyl-N-trideuteriomethylamino)-3-phenylpropane (1) by rodent liver homogenates was studied. The ratio of 1-trideuteriomethylamino-3-phenylpropane (2)/1-methylamino-3-phenylpropane (3) was determined by gc-ms. The ratio of 2/3 in the product of N-demethylation of 1 by liver homogenate from phenobarbital-treated rats was 1.31 and from untreated rats was 1.32, and the differences between the two groups were not statistically significant. The ratio of 2/3 in the product obtained from N-demethylation of 1 by liver homogenates from mice was 1.45. The ratio of 2/3 of greater than unity indicates the presence of a kinetic primary isotopes effect in the enzymatic N-demethylation reaction. This effect is exclusive of the secondary isotope effects on the amino nitrogen. The presence of this primary isotope effect indicates that the cleavage of the C-H bond of the N-methyl group is a rate-limiting step in the N-demethylation of tertiary amines by rodent microsomal enzymes.