The use of glycated molecules as indicators of the glycaemic levels of diabetics and for screening:
By Harris Ramuth
Diabetes is characterised by an overproduction of glucose by the liver and an under utilisation by other organs. There is a set of causes to diabetes mellitus and there is an intricate relationship between the vascular and neuropathic components of the metabolic syndrome. Recently Compiled data showed that between 120 to 140 million people suffer from diabetes mellitus worldwide and that number is projected to double by the year 2025. Much of this increase will occur in developing countries and will be due to an ageing population, unhealthy diets, obesity and a sedentary life style.
Glycation of molecules is a natural process occurring in every individual. Molecules that are glycated are mostly proteins namely haemoglobin and albumin. In adults, circulating haemoglobin typically consists of 3 main portions namely Haemoglobin A1, Haemoglobin A2 ( 0.5%) and Haemoglobin F (2.5%). HbA is by far the most important fraction of haemoglobin and it is composed of minor fractions like HbA1a, HbA1b and HbA1c, collectively called haemoglobin A1 or fast haemoglobin, glycosylated haemoglobin, glycated haemoglobin or simply glycohaemoglobin.
There is a condensation reaction between the N- terminal Valine amino acid of each b - chain of the haemoglobulin molecule. An unstable product called Aldimine is formed. This is also known as pre-HbA1c. the unstability of this product is such that it dissociates back to glucose and haemoglobin. Nevertheless, a small portion is converted to haemoglobin A1c, which is a ketoamine. This conversion occurs slowly and by a molecular re-arrangement known as Amadori. This step is non enzymatic, mostly irreversible and proportional to the glucose concentration to which the red cell is exposed to and continuous over the life span of the red cell. Therefore, the determination of this fraction of haemoglobin and its estimation as a percentage of total haemoglobin can be indicative of the mean blood glucose concentration over a period of 2 to 3 months prior to measurement. It is a retrospective index of the glycaemic state of the patient in that preceding period. Sophisticated methods now exist to make measurements of both HbA1 and HbA1c. there exist autoanalysers like the Hi-autoA1c, for example, which is able to dilute, haemolyse and remove labile fractions of Hba1 within the instrument.
High performance liquid chromatography( HPLC) provides excellent results with high degree of reproducibility, can be automated and eliminate the risks of falsely high or low values, usually associated with haemoglobinopathies or due to other haemoglobin.
Low performance liquid chromatography techniques are relatively much cheaper. Commercial kits consisting of microcolumns with ion exchange resins and buffer are readily available easy to manipulate and can be introduced into routine work without great difficulty. Electrophoresis on agarose gel, iso electric focussing on polyacrylomide gels, immunoassays and colorimetric estimation of HbA1 using thiobarbituric acid can also be used for similar effects.