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 delta N-15 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 delta N-15 values of less than 1.0 parts per thousand to be satisfactorily measured, with a standard deviation routinely less than 0.5 parts per thousand. The methodology has been tested by determining the changes in the delta N-15 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 (c) 2005 John Wiley & Sons, Ltd.