Enzymatic Hydrolysis of Whey Protein Concentrates: Peptide HPLC Profiles

Abstract

Hydrolysis of whey protein concentrates (WPCs) at different temperatures and pHs, using three enzymes: pepsin, trypsin, and Alcalase^®, was monitored during more than 5 hr by reversed phase HPLC/UV, using a column containing a polystyrene‐divinylbenzene copolymer‐based packing, and an elution gradient from 8% to 80% acetonitrile containing 0.1% TFA. Peptides were separated according to their polarity and size, and degradation of α‐lactalbumin (α‐la) and β‐lactoglobulin (β‐lg) was evaluated. The three proteolytic enzymes (pepsin, trypsin, and Alcalase^®) employed for hydrolysis of WPCs led to different kinetics of degradation of β‐lg. α‐la degradation after 15 min was almost complete for the three enzymes. The hydrolysis catalysed by each enzyme resulted in different peptide profiles by HPLC/UV. Hydrolysates produced by pepsin (HP) were resolved into three main fractions of high retention times, while tripsin hydrolysates (HT) were resolved into nine major peaks and Alcalase^® hydrolysates (HA) were resolved into 12 major peaks, presenting a wide range of polarities and sizes. Although, with different β‐lg hydrolysis extension, chromatographic profiles of the degradation and formation of peptides can be used as a finger print of the type of enzyme used, because peptide profile is not affected either by temperature or pH.

Keywords:

• Whey proteins
• Enzymatic hydrolysis
• RP‐HPLC

Acknowledgment

This work received financial support from Fundação para a Ciência e a Tecnologia (project POCTI/2000/QUI/36452).