Abstract
As an advanced stage of glycation, glycated human serum albumin (G-HSA; glucose content, 2mol of 5-hydroxymethylfufural equivalent/mol of HSA) was incubated at 37°C up to 30d in 0.2M phosphate buffer, pH 7.4, with 100 μM Fe3+. G-HSA incubated for 30d (G-HSA-30(Fe)) was subsequently hydrolyzed at 110°C for 24h in 6N HCl. In the hydrolysate, N2-carboxymethyllysine (CML) was identified by cochromatography with synthesized CML on an amino acid analyzer. pI of HSA (4.8) shifted to 4.5 in G-HSA. A more acidic fraction, pI 4.3, appeared in G-HSA-30(Fe). CML content (mol of CML/mol of HSA) of HSA and G-HSA was as follows; 0 in HSA, 0.2 in HSA-30(Fe), 0.4 in G-HSA and 1.5 in G-HSA-30(Fe) pI 4.3. The amino acid compositions also changed in lysine, arginine and tyrosine at the advanced stage of the reaction.
