Comparison of a modified spectrophotometric and the pH-stat methods for determination of the degree of hydrolysis of whey proteins hydrolysed in a tangential-flow filter membrane reactor

A procedure was developed to determine the degree of hydrolysis (DH) of whey protein hydrolysates (WPH) during hydrolysis in either 3 kDa or 10 kDa tangential-flow filter (TIFF) enzymatic membrane reactors (EMR). Protease N Amano G (IUB 3.4.24.28, Bacillus subtilis) was used to hydrolyse an initial 5% (w v(-1)) aqueous solution of whey protein isolate (86.98% protein) at pH 7.0 and 55 degrees C with continuous recirculation and simultaneous removal of hydrolysates through the TIFF, in single- or two-stage operation. The DH in the permeate and the retentate were determined as the concentration of the free alpha-NH(2) using 2.4,6-trinitrobenzene 1-sulphonic acid (TNBS) and compared to the pH-stat method. In the new method, the DH of the permeate, the retentate and for the total EMIR process could be quantified together or independently. The pH-stat method exaggerated the DH in the EMR because of the leakage of the alkali. The TNBS method was more reliable for DH estimation in the EMR. (C) 2008 Elsevier Ltd. All rights reserved.