The use of anhydrotrypsin-Sepharose for isolation of trypsin inhibitors from Vigna unguiculata seeds.

The preparation of anhydrotrypsin-Sepharose 4B for the isolation of trypsin inhibitors by affinity chromatography was described. Soybean trypsin inhibitor and an acetic acid extract of Vigna unguiculata seed meal were used to evaluate the binding of trypsin inhibitors. Both soybean trypsin inhibitor and Vigna trypsin inhibitors were retained at pH 7.6 and desorbed at pH 2.0. The Vigna trypsin inhibitors isolated by the Sepharose-4B anhydrotrypsin did not differ by SDS-polyacrylamide gel electrophoresis and isoelectric focusing from native inhibitors, indicating that partial proteolysis obtained with trypsin affinity columns can be avoided by using anhydrotrypsin, the enzymatically inactive form of the enzyme which retains the capacity to bind inhibitors.