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An optimized matrix‐assisted laser desorption/ionization sample preparation using a liquid matrix, 3‐aminoquinoline/ α ‐cyano‐4‐hydroxycinnamic acid, for phosphopeptides
Author(s) -
Fukuyama Yuko,
Takeyama Kohei,
Kawabata Shinichirou,
Iwamoto Shinichi,
Tanaka Koichi
Publication year - 2012
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.6363
Subject(s) - chemistry , chromatography , matrix assisted laser desorption/ionization , mass spectrometry , phosphopeptide , desorption , analyte , sample preparation , matrix (chemical analysis) , analytical chemistry (journal) , organic chemistry , peptide , biochemistry , adsorption
RATIONALE A liquid matrix, 3‐aminoquinoline (3‐AQ)/ α ‐cyano‐4‐hydroxycinnamic acid (CHCA), introduced by Kolli et al. in 1996 for matrix‐assisted laser desorption/ionization mass spectrometry (MALDI‐MS), has been reported for peptides and proteins, oligonucleotides, oligosaccharides, and glycopeptides. However, it has not been validated for phosphopeptides. METHODS We optimized sample preparation using 3‐AQ/CHCA for phosphopeptides. The sensitivity of six phosphopeptide species as isolated or in digests was systematically evaluated by using MALDI‐quadropole ion trap (QIT)‐time of flight (TOF) MS in positive and negative ion modes, and compared with the conventional methods using a solid matrix, 2,5‐dihydroxybenzoic acid (2,5‐DHB). RESULTS The sensitivity of mono‐ and tetraphosphopeptides was improved 10‐ to 10 000‐fold with the optimized preparation method using 3‐AQ/CHCA compared with the conventional methods using 2,5‐DHB. Improvement by 3‐AQ/CHCA itself was 10‐fold. Adding ammonium dihydrogen phosphate or an analyte solvent composition was also effectively improved the sensitivity. Phosphopeptides in isolated form or in digests were detected at femto‐ or subfemtomole levels. CONCLUSIONS Sensitivity of phosphopeptides was improved by the optimized sample preparation method using 3‐AQ/CHCA compared with the conventional method using 2,5‐DHB. The validation of 3‐AQ/CHCA for phosphopeptides was systematically confirmed, expanding the potential of this matrix to phosphoproteomics. Copyright © 2012 John Wiley & Sons, Ltd.