Deuterium transfer in the bioconversion of glucose to ethanol studied by specific isotope labeling at the natural abundance level

Site-specific natural isotope fractionation measured by deuterium NMR (SNIF-NMR) was used for investigating the deuterium transfers occurring in the fermentation of sugars into ethanol. In contrast to carbon-13, which is usually assumed to be randomly distributed within the glucose skeleton, very large deviations with respect to a statistical repartition are determined for deuterium. By transforming glucose samples from different origins into acetates and nitrates, the absolute values of the D/H ratios in the nonexchangeable sites were obtained. The hydroxyl sites were considered to contribute to the isotope content of the starting water medium. No direct connection is found between glucose and the methylene site which is only sensitive, with a strong discriminating effect against deuterium, to the isotope content of water. A redistribution coefficient slightly less than unity (0.96 for a concentration of sugar of 100 g L/sup -1/) is found between the isotope ratio of the end and starting water media. The site-specific natural isotope parameters of ethanol constitute a faithful and powerful probe for investigating the physiological biochemical and climatological effects which have governed the photosynthesis of sugars in natural conditions.