PROTEIN DIGEST ANALYSIS BY PRESSURIZED CAPILLARY ELECTROCHROMATOGRAPHY USING AN ION-TRAP STORAGE REFLECTRON TIME-OF-FLIGHT MASS DETECTOR/

Pressurized capillary electrochromatography (pCEC) has been coupled to an ion trap storage/reflectron time-of-flight mass spectrometer for t he analysis of peptide mixtures and protein digests, Taking advantage of the electroosmotic now, high separation efficiency has been achieve d in pCEC due to a relatively hat now profile and the use of smaller p acking materials, A supplementary pressure was used in these experimen ts which suppressed bubble formation and also allowed the tuning of th e elution of peptides using the electrical field, In this work, a fast separation of a six-peptide mixture has been successfully performed. Using columns only 6 cm long, a tryptic digest of bovine cytochrome c was fully separated in around 14 min by properly tuning the applied vo ltage and the supplementary pressure, In addition, relatively complex protein digests, such as a tryptic digest of chicken ovalbumin, were a nalyzed using this pCEC/MS system, and more than 20 peaks were resolve d in the total ion current chromatogram within 17 min, The use of an i on trap storage/reflectron time-of-flight mass spectrometer as an on-l ine detector further increased the resolving power of the pCEC by unam biguously identifying coeluting components, The nonscanning property o f the time-of-flight mass analyzer and the ion signal integration capa bility of the ion trap were successfully combined to provide rapid and sensitive full-mass range detection in these experiments.