Structural characterisation of acetaldehyde adducts formed by a synthetic peptide mimicking the N-terminus of the hemoglobin beta-chain under reducing and nonreducing conditions.

This work was carried out to elucidate the structures resulting from acetaldehyde-induced modifications at the haemoglobin beta-chain N-terminal residues under different experimental conditions. A synthetic peptide (Val-His-Leu-Thr-Pro-Glu-Cys) of m/z 798, which represents the six N-terminal residues of the haemoglobin beta-chain N-terminus with an additional C-terminal cysteine, was used as a model compound. Peptide-acetaldehyde adducts were separated by reverse-phase HPLC. Fast-atom-bombardment MS and linked-scan (B/E) spectra were used to study adduct structures. Under nonreducing conditions at pH 7.4 (1:10 peptide/acetaldehyde molar ratio), two types of adducts of m/z 824 were formed, both with modifications at the N-terminal valine. These two adducts were shown to be a Schiff base, and a cyclic 2-methyl-imidazolidine-4-one. The 2-methyl-imidazolidine-4-one adduct was demonstrated to be formed via the Schiff base and to undergo dissociation gradually after 24 h. Reducing conditions at pH 7.4 (peptide /acetaldehyde molar ratio of 1:1 with 20 mM NaCNBH3) resulted in the formation of an adduct of m/z 826, which was shown to be an N-ethylated adduct of valine. A small amount of nonreduced adducts were also formed under these conditions. Reducing conditions at pH 9.0 (1:10 peptide/acetaldehyde molar ratio with 20 mM NaCNBH3) yielded two secondary, i.e. diethylated (m/z 854), products very rapidly. The cysteine residue of the peptide was not found to form an adduct with acetaldehyde under physiological pH.