Anal. Chem. 1995, 67, 4170-4177
On=LineMicroreactors/Capillary Electrophoresis/ Mass Spectrometry for the Analysis of Proteins and Peptides Larry Licklider,t Wemer G. Kuhr,*-t Martin P. Lacoy,* Thomas Keough,* Mkhael P. Purdon,@ and Ray Takigiku*s
Department of Chemistry, Universiiy of Califomia, Riverside, Califomia 92521, and Corporate Research Division, Corporate Professional & Regulatory Division, and Procter and Gamble Phatmaceuticals, The Procter and Gamble Company, Miami Valley Labs, P.0. Box 538707, Cincinnati, Ohio 45253-8707
We report the use of trypsin and carboxypeptidase YmodiEied capillary microreactors (50 pm i.d. and 20 nL volume per centimeter length) in combinationwith mass spectrometry for peptide molecular mass mapping of various peptides and proteins. Advantages of immobilized enzyme capillary microreactors include picoliter to nanoliter volume requirements, longer enzyme lifetimes, h@er stability, and the ability to reuse enzymes conveniently. Additionally, extremely efficient sample handling modes are used, and the reaction products are easily separated from the enzyme reagents. Plasma desorptionand ma&assisted laser desorptiodionizationwere used for off-line analyses of digestion products. The use of MassMap for the identificationof proteins is also discussed. Finally,a trypsin microreactor was integrated on-linewith capillary electrophoresis/ion spray mass spectrometry for fast peptide mapping. Digestion of the oxidized insulin Bchain in an on-line trypsin microreactor and electromigration of aliquots from the capillary microreactor into the CE separation capillary allowed the entire peptide mapping procedure to be completed in
