A NEW SPECTROPHOTOMETRIC ASSAY FOR NATIVE BETA-LACTOGLOBULIN IN RAW AND PROCESSED BOVINE-MILK

The new method includes three selective steps for isolation of native P-lactoglobulin (beta-lg) in milk: (i) the isoelectric precipitation o f caseins; (ii) a preconditioning step using a selective buffer to imp rove the solubility of native beta-lg required to withstand denaturati on and (iii) precipitation in presence of 2% (w/w) trichloroacetic aci d (TCA). The TCA-filtrates were desalted on Econo Pak 10 DG. Eluates c ontaining the pure native beta-lg were measured spectrophotometrically at 280 nm against a blank prepared with 12% (w/w) TCA. Chromatographi c separations were accomplished using FPLC and a Superose 12 column. T he new assay shows a high selectivity for native beta-lg, is fast and requires no high-cost equipment. The coefficient of variation was calc ulated to be < 1.9%. To monitor the loss of native beta-lg, kinetic an d thermodynamic parameters for the heat-induced denaturation were exam ined over the temperature range of 70-95 degrees C (90-300 s) and a re action order of 1.5 was found for beta-lg denaturation. (C) 1998 Elsev ier Science Ltd. All rights reserved.