Glucosylation of human haemoglobin a in red blood cells studied in vitro. Kinetics of the formation and dissociation of haemoglobin A1c.

We examined the stability of the ketoamine adduct, termed HbA1c, formed between the N-terminal valine of the haemoglobin A (HbA) beta chain and D-glucose using an isoelectric focusing method. Prolonged saline incubation of purified HbA1c followed by renewed isoelectric focusing revealed that the HbA1c concentration decreased while a corresponding increase in the HbA concentration occurred. The emergence of haemoglobin A on saline incubation indicates that the non-enzymatic glucosylation of haemoglobin A to HbA1c is a reversible process. This finding was further substantiated by kinetic studies on the formation and breakdown of the ketoamine adduct during incubation of red cells in glucose and saline. It appeared that the rate constant for the formation k2 was 14.2 X 10(-6) X s-1 at 37 degrees C while the rate constant for dissociation k-2 was 1.7 X 10(-6) X s-1. From these data an equilibrium constant K of 8.4 was calculated. This information should be of importance in interpretation of HbA1c levels during abrupt changes in diabetic control.