Deuterium isotope effects on carbon-13 chemical shifts in selected amino acids as function of pH

Abstract

The carbon-13 chemical shifts of glycine, alanine, and lysine have been studied as a function of pH, when dissolved in water as well as in deuterium oxide. No correction of the apparent pH measured in the deuterium oxide samples was made. A comparison of the two sets of data thus obtained shows that isotope shifts of up to 0.9 ppm are present. A more detailed analysis of the data reveals that the isotope shifts are caused not only by an isotope effect on the titration shifts but are, in the high pH region, also due to isotope effects on the p K values that correspond to the dissociation of the ammonium groups. Thus relatively small isotope effects on these p K values cause significant changes of the 13C chemical shifts at pH values equal or close to the p K values. While the isotope effect on the titration shifts in the acid and neutral pH regions causes upfield shifts, as normally observed, downfield shifts have been found in the basic pH region. The titration shifts and p K values necessary for the analysis were obtained by a nonlinear least squares fit of the appropriate theoretical model to the shift data lot all carbons in a given amino acid, simultaneously. This procedure allows a precise determination of macroscopic p K values and titration shifts even in the case of strongly overlapping deprotonation reactions, as for lysine.