DEVELOPMENT OF A HPLC-UV METHOD FOR DETERMINATION OF WHEY PROTEIN OF BOVINE
MILK AND POWDER WHEY PROTEIN
Angela G. S. Correia, Victor H. O. Andrade, Genildo C. F. Júnior, Danilo A. Vieira, Eliane C. Souza, Maria
A. M. Alves, Maria C. D. Silva, Ticiano G. Nascimento. Irinaldo D. B. Júnior. Master Course in Nutrition,
Federal University of Alagoas – UFAL, Av. Lourival Melo Mota s/n, 57072-900, Maceió, Alagoas, Brazil
The whey is an important product generated by the dairy industry and contains bioactive compounds
including proteins, α-lactalbumin (α-la) and β-lactoglobulin (β-lg), with functional and technological
properties for food industry. The HPLC is an excellent analytical method for separation, determination of
compounds present in foods, because high resolution and reproducibility in separation of constituents of
milk and whey. This work developed an analytical method to separate whey protein and to determinate
quantitatively the major proteins of bovine whey in nature and demineralized powder whey. HPLC method
consisted of a chromatographic system containing UV detector to acquire data at 205 nm and column
RP18. The mobile phase consisted of a solvent A (acetonitrile; 0.1% TFA) and solvent B (water, 0.1%
TFA) in gradient system, flow rate of 1.0 mL/min. The run time was 26 minutes. The HPLC method
showed suitable for the separation of the major whey proteins fractions, α-lactoalbumin (α-la) and βlactoglobulin (β-lg), in the retention time, of 12 minutes and 13 minutes with good resolution. The method
presented linearity in the concentration range of 0.025-0.500mg/mL for (α-la) and 0.010 - 1.000mg/mL for
(β-lg), respectively. Using HPLC method, it was possible to determinate the major whey protein in the
samples of whey from bovine milk with concentrations of 1.240g/L for (α-la) and 2.480g/L for (β-lg). In the
demineralized powder whey, the concentrations were 1.230g/L for α-la and 3.261 g/L for β-lg,
respectively. This research demonstrated that the HPLC method presented analytical reproductibility for
determination of the main whey proteins.