An economical and environmentally friendly whey protein fractionation process was developed using supercritical carbon dioxide (sCO(2)) as an acid to produce enriched fractions of alpha-lactalbumin (alpha-LA) and beta-lactoglobulin (beta-LG) from a commercial whey protein isolate (WPI) containing 20% alpha-LA and 55% beta-LG, through selective precipitation of alpha-LA. Pilot-scale experiments were performed around the optimal parameter range (T = 60 to 65 degrees C, P = 8 to 31 MPa, C = 5 to 15% (w/w) WPI) to quantify the recovery rates of the individual proteins and the compositions of both fractions as a function of processing conditions. Mass balances were calculated in a process flow-sheet to design a large-scale, semi-continuous process model using SuperproDesigner(R) software. Total startup and production costs were estimated as a function of processing parameters, product yield and purity. Temperature, T, pressure, P, and concentration, C, showed conflicting effects on equipment costs and the individual precipitation rates of the two proteins, affecting the quantity, quality, and production cost of the fractions considerably. The highest alpha-LA purity, 61%, with 80% alpha-LA recovery in the solid fraction, was obtained at T = 60 degrees C, C = 5% WPI, P = 8.3 MPa, with a production cost of $8.65 per kilogram of WPI treated. The most profitable conditions resulted in 57%-pure alpha-LA, with 71% alpha-LA recovery in the solid fraction and 89% beta-LG recovery in the soluble fraction, and production cost of $5.43 per kilogram of WPI treated at T = 62 degrees C, C = 10% WPI and P = 5.5 MPa. The two fractions are ready-to-use, new food ingredients with a pH of 6.7 and contain no residual acid or chemical contaminants.