From among a wide variety of protein purification techniques affinity chromatography has proved to be particularly effective for separation of proteolytic enzymes and their inhibitors. In this article, following a general description of affinity adsorbents used for purification of proteinases, we overview a simple separation procedure for some serine proteinases and their inhibitors by way of affinity chromatography in the presence of high NaCl concentration. It has been shown that some highly specific trypsin inhibitors exhibit also antichymotrypsin activity when high concentration of Na(+) but not K(+) or Li(+) ions are present in the reaction mixture. Taking advantage of this phenomenon the virgin forms of trypsin inhibitors from squash seeds, Kazal-type inhibitor from porcine pancreas and alpha(1)-proteinase inhibitor from human and sheep plasma, as an example, were separated using immobilized chymotrypsin or its inactive derivative methylchymotrypsin in the presence of 5 M NaCl.