QUANTIFICATION OF THE MAJOR BOVINE WHEY PROTEINS USING CAPILLARY ZONEELECTROPHORESIS

Bovine whey comprises four major protein groups [alpha-lactalbumin, be ta-lactoglobulin variants A and B, bovine serum albumin and immunoglob ulin (specifically IgG)] which have a diverse range of molecular masse s, pI values, number of phenotypic variants and subunit compositions. The development of a capillary zone electrophoresis method to separate these whey proteins is described. Initially separation of the individ ual whey proteins was evaluated using a number of different buffer sys tems with pK(a) values above pH 7. At buffer pH values greater than 7, protein-capillary wall interactions were minimized as the majority of the whey proteins had a net negative charge because their pI values a re in the range pH 4-6. A wide range of buffer additives (organic modi fiers and surfactants) were also added to alter the chemistry of the s eparation, to further block protein-capillary wall interactions and to thus optimize the resolution of the different protein peaks. A sample buffer/separation buffer system was developed which eliminated an ini tial solvent trough that coincided with the IgG peak. This made it pos sible to quantify the IgG protein. Optimum resolution and analysis tim e (10 min) for the four whey proteins was achieved with a sample buffe r consisting of 10 mM phosphate, pH 7.4 and a separation buffer consis ting of 150 mM sodium berate, pH 8.5 containing 0.05% Tween 20. This m ethod was successfully used to separate a mixture of commercially puri fied whey proteins and to separate and quantitate the individual whey proteins in an acid whey sample.