Factors influencing pepsinolysis of methyl-, ethyl- and propyl-ester derivatives of beta-lactoglobulin

beta -Lactoglobulin was esterified with methanol, ethanol or propanol, yielding 100, 60 and 44\% esterification, respectively. The kinetics of peptic hydrolysis of unmodified, methylated-, ethylated- and propylated-beta -lactoglobulin derivatives were followed under various conditions of time, temperature, and enzyme and protein concentrations. The resulting hydrolysates were analyzed by SDS-PAGE and RP-HPLC. Changes of temperature and enzyme concentration influenced strongly the rate of hydrolysis and the RP-HPLC profiles while protein concentration had much smaller impact on reaction rates when tested in the range 2.5-20 mg/mL. Peptic hydrolysis of methylated-beta -lactoglobulin derivatives could proceed at a temperature as low as 4C and at an enzyme/substrate ratio as low as 0.125\%. Peptide profiles of the hydrolysates of methyl-and ethyl-beta -lactoglobulin derivatives were generally quite similar except for the appearance of additional peaks in case of methyl-beta -lactoglobulin. RP-HPLC profile of propyl-beta -lactoglobulin ester was slightly different from that of both the methyl and ethyl-beta -lactoglobulin. The features of the peptide profiles of the different hydrolysates of esterified beta -lactoglobulin did not differ after long pepsinolysis. The population of small hydrophilic peptide increased, accompanied by a decrease of the large hydrophobic peptides (for methyl derivative) and the hydrolysis of intact protein (for methyl- and propyl-derivatives).