N-Terminal pyrazinones: a new class of peptide-bound advanced glycation end-products

The reaction of peptide Gly-Ala-Phe with the alpha-dicarbonyl compounds glyoxal and methylglyoxal was studied under physiological conditions (pH=7.4, 37 degrees C). Using HPLC with UV and fluorescence detection, a rapid derivatization of the peptide and the concomitant formation of well-defined products were observed. The products, which showed characteristic UV absorbance (lambda(max)=320 to 340 nm) and fluorescence (lambda(ex)=330 to 340 nm, lambda(em)=395 to 405 nm), were identified by ESI-MS and NMR spectroscopic analysis as the N-terminally pyrazinone-modified peptides I (N-[2-(2-oxo-2H-pyrazin-1-yl)-propyl]-phenylalanine) and II (N-[2-(5-methyl-2-oxo-2H-pyrazin-1-yl)-propionyl]-phenylalanine). Model experiments revealed that the reactivity of the N-termini of peptides towards a derivatization by glyoxal is in the same order of magnitude as that of arginine, which generally is attributed as main target for alpha-dicarbonyl compounds in proteins. Incubation of insulin with glyoxal proved the protein-bound formation of pyrazinones, with the N-terminus of the B-chain as the main target. According to these results, we conclude that N-terminal pyrazinones represent a new type of advanced glycation end-products (AGEs) with significance for biological systems and foods.