Analysis of phospholipase A(2) glycosylation patterns from venom of individual bees by capillary electrophoresis/electrospray ionization mass spectrometry using an ion trap mass spectrometer

A method based on tryptic digestion, ultrafiltration and capillary electrop horesis/mass spectrometry (CE/MS) has been developed for the analysis of th e glycosylation pattern in the phospholipase A(2) (PLA) of individual honey bees. Without reducing the disulfide bonds, PLA was digested with trypsin a nd filtered with a 3 kDa molecular weight (MW) cut-off membrane. With this procedure, the glycopeptides could be isolated from the nonglycosylated pep tides. After tryptic digestion and ultrafiltration, the disulfide bonds wer e reduced before analysis by CE, To reduce the adsorption, CE separation wa s performed on successive multiple ionic-polymer (SMIL) polybrene (PB) coat ed capillary columns. The SMIL-PB columns allowed partial separation of the glycopeptides and eight glycopeptides were identified by on-line coupling of CE with electrospray ionization (ESI) mass spectrometry, The analysis of phospholipase A(2) from the venom of individual bees indicated that the va riation and relative abundances of different glycopeptides were similar bet ween the younger and the older bees. Copyright (C) 2000 John Wiley & Sons, Ltd.