Effect of enzymatic hydrolysis on antigenicity (allergenticity) of casein fractions of cow and buffalo milk

The crude casein (alpha(S1)- and beta-caseins) fractions isolated from cow and buffalo milk were purified on DEAE cellulose ion exchange chromatography by eluting with NaCl gradient. The antisera were raised against these pure protein antigens by immunizing healthy rabbits. The antigenicity was estimated by indirect ELISA using IgG conjugate. beta-casein fraction showed more antigenicity than as alpha(S1)-casein. Buffalo milk beta-casein fraction showed more antigenicity than cow milk fraction. Partial hydrolysis of proteins was carried out at 45degreesC by neutrase enzyme at enzyme to substrate ratio of 1:5000. The casein fractions hydrolysed to 5% degree of hydrolysis analyzed for peptide profiles by SDS-PAGE and RP-HPLC showed peptides of molecular weight < 6 KD, which are mainly of hydrophilic in nature particularly in beta-casein hydrolysate. The residual antigenicity of hydrolysed proteins estimated by inhibition ELISA, indirectly reflected the reduction in allergenicity of milk proteins. Modification of proteins to 5% degree of hydrolysis, reduced the antigenicity by 70-80%. The investigation revealed the usefulness of enzymatic modification in reduction of allergenicity of caseins, as antigenicity could be a measure of allergenicity.