Formation of nitri- and nitrosylhemoglobin in systems modeling the Maillard reaction

Nitric oxide (NO) and its metabolites can nitrosylate hemoglobin (Hb) through the heme iron. Nitrihemoglobin (nitriHb) can be formed as result of porphyrin vinyl group modification with nitrite. However, in those with diabetes the non-enzymatic glycation of Hb amino acids residues (the Maillard reaction) can take place. The objectives of this study were to investigate effects of the Maillard reaction on the interaction of methemoglobin (metHb) with S-nitrosoglutathione (GSNO) and nitrite.

Nitrosylhemoglobin production was registered using increasing optical density at 572 nm and compared with 592 nm, and with EPR spectroscopy. Formation of nitriHb was determined using an absorbance band of reduced hemochromogen (582 nm) in the alkaline pyridine solution. Accumulation of fluorescent advanced glycation end-products of Hb was measured through increasing of fluorescence at 385-395 nm (excitation λ=320 nm).

We determined that NO metabolites such as GSNO and nitrite at physiological pH values and aerobic conditions caused modification of metHb porphyrin vinyl groups with nitriHb formation. It was ascertained that this formation was inhibited by superoxide dismutase. In microaerobic conditions metHb was nitrosylated under the action of GSNO or GSNO with methylglyoxal. Nitrite nitrosylated metHb only in the presence of methylglyoxal. It was shown that GSNO inhibited accumulation of fluorescent products which formed during Hb glycation with methylglyoxal.

The assumption was made that intermediates of the Hb glycation reaction play an important role both in vinyl group nitration and in heme iron nitrosylation. Oxygen content in reaction medium is an important factor influencing these processes. These effects can play an important role in pathogenesis of the diseases connected with carbonyl, oxidative and nitrosative stresses.