Position-specific N Isotope Analysis in Organic Molecules: A High-precision N NMR Method to Determine the Intramolecular N Isotope Composition and Fractionation at Natural Abundance

The position-specific N isotope content in organic molecules, at natural abundance, is for the first time determined by using a quantitative methodology based on N Nuclear Magnetic Resonance (NMR) spectrometry. N NMR spectra are obtained by using an adiabatic "Full-Spectrum" INEPT sequence in order to make possible N NMR experiments with a high signal-to-noise ratio (>500), to reach a precision with a standard deviation below 1‰ (0.1%). This level of precision is required for observing small changes in N content associated to N isotope effects. As an illustration, the measurement of an isotopic enrichment factor ε for each N isotopomer is presented for 1-methylimidazole induced during a separation process on a silica column. The precision expressed as the long-term repeatability of the methodology is good enough to evaluate small changes in the N isotope contents for a given isotopomer. As observed for C, inverse and normal N isotope effects occur concomitantly, giving access to new information on the origin of the N isotope effects, not detectable by other techniques such as isotope ratio measured by Mass Spectrometry for which bulk (average) values are obtained.