CAPILLARY ELECTROPHORESIS USED TO MONITOR THE ENZYMATIC-HYDROLYSIS OFCASEINS AND THE FRACTIONATION OF HYDROLYSIS PRODUCTS

Alpha(s)-and beta-Casein (CN) was purified from acid casein by anion-e xchange chromatography. The composition of whole casein and of casein fractions was monitored by capillary electrophoresis (CE) with 0.1 M N a-phosphate, pH 7.3, 4M urea as run buffer. The 4 individual caseins w ere well separated and were identified by comparison to pure caseins r un under identical conditions. Purified alpha(s)- and beta-CN were hyd rolysed by chymosin at pH 6.2. Aliquots withdrawn at different interva ls were heated at 85-degrees-C for 10 min and analysed by CE. This sho wed that hydrolysis of alpha(s1)-CN was fast and extensive, all the al pha(s1)-CN was hydrolysed within 4 h, resulting in a number of peptide s. Alpha(s2)-CN, apparently, was not hydrolysed by chymosin. The hydro lysation of beta-CN was much slower than that of alpha(s1)-CN. After t erminating the hydrolysis process (18 h for the alpha(s)-CN, 24 h for the beta-CN preparation) casein and large peptides were precipitated a t pH 4.6 and with 70% ethanol, and peptides in the supernatants were f ractionated by reversed-phase HPLC with an acetonitrile gradient. Comp osition of the fractions collected was monitored by CE using a 0.05 M Na-phosphate buffer, pH 7.3 without urea. This revealed the purity of the fractions. Generally, the same number or a higher number of peptid es were shown by CE than by RP-HPLC. Capillary electrophoresis proved to be a very useful tool for monitoring the fractionation of casein, t he hydrolysis of caseins and the fractionation of hydrolysates. This i mplies that CE may also be used advantageously for monitoring fraction ation of peptides from cheeses and other milk products.