A rapid purification method for soybean Bowman-Birk protease inhibitor using hydrophobic-interaction chromatography.

A protein fraction was isolated from defatted soybean flour by extraction at acid pH, 40% ammonium sulfate precipitation, hydrophobic interaction chromatography, and gel filtration chromatography. SDS-PAGE, under reducing conditions, confirmed it as a homogeneous preparation. This conclusion was consistent with N-terminal amino acid sequence data (20 cycles) which showed a major sequence identical to those reported for soybean Bowman-Birk-type protease inhibitor (BBI), and also indicated a minimum 95% purity based on recoveries of PTH-amino acid residues. The purified fraction inhibited both trypsin and chymotrypsin with average specific activities of 350 and 672 units mg(-1), respectively. Compared with classical BBI purification, this procedure is very rapid requiring only 72-96 h to achieve a yield of 37 mg purified BBI per 200 g starting material.