Chromatographic separation of glycated nucleotides

A high-performance liquid chromatographic method for the separation of nucleotide glycation reaction product(s) has been developed. The product(s) arising during in vitro glycation of a nucleotide (AMP, GMP or CMP) with a reducing sugar (ribose or glucose) were clearly resolved from the non-glycated constituents of the reaction mixture. The peak(s) of the glycated product(s) increased when an amino acid (lysine, arginine, beta-alanine or N epsilon-acetyllysine) was added to the reaction mixture. This increase probably corresponds to the formation of a new product with a different absorption maximum (250 versus 260 nm). Conversely, formation of this product(s) was inhibited by the presence of the metal-chelating agent diethylenetriamine pentaacetic acid and by aminoguanidine.