APPLICATION OF REVERSED-PHASE HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY AND CAPILLARY ZONE ELECTROPHORESIS TO THE PEPTIDE-MAPPING OF PEPSIN ISOENZYMES

A combination of reversed-phase high-performance liquid chromatography (RP-HPLC) and capillary zone electrophoresis (CZE) was used for the c haracterization of peptide maps of swine pepsin after its digestion wi th alpha-chymotrypsin. Peptide maps obtained by both methods were comp ared and five selected chromatographic peaks were identified on an ele ctrophoreogram. The different order of peaks found in RP-HPLC compared to CZE confirmed the complementarity of these two methods. More pepti de fragments were resolved by RP-HPLC, which was also found to be less sensitive to salt content in peptide mixtures, than by CZE, but only CZE was able to separate and identify phosphorylated and dephosphoryla ted peptide fragments of swine pepsin digest. CZE provides faster sepa ration than RP-HPLC, however, the salts have to be removed by ultrafil tration or by RP-HPLC pre-separation prior to CZE analysis. Combined u se of RP-HPLC and CZE for peptide mapping makes it possible to disting uish between the phosphorylated and dephosphorylated forms of swine pe psin. This is important from a diagnostic point of view, because pepsi n phosphorylation may be associated with gastric cancer.