Determination of the natural abundance δ15N of nicotine and related alkaloids by gas chromatography/isotope ratio mass spectrometry

A method is described by which the natural abundance δ15N values of nicotine, analogues, and metabolites can be determined. The alkaloids are extracted from their biological matrix by solid-phase extraction and analysis is conducted using isotope ratio mass spectrometry interfaced to gas chromatography. Repeatability and precision are sufficient to allow differences in the δ15N values of less than 1.0‰ to be satisfactorily measured, with a standard deviation routinely less than 0.5‰. The methodology has been tested by determining the changes in the δ15N values of nicotine, N-methyl-2-phenylpyrrolidine and their respective demethylation products, nornicotine and 2-phenylpyrrolidine, during biotransformation by cell suspension cultures of Nicotiana species. Sufficient precision and reproducibility were obtained to allow the kinetic isotope effects associated with the demethylation reaction to be calculated. Copyright © 2005 John Wiley & Sons, Ltd.