Using HPTLC/DESI-MS for peptide identification in 1D separations of tryptic protein digests

• Published in: Analytical and bioanalytical chemistry 2008-05, Vol.391 (1), p.317-324
• Main Authors: Pasilis, Sofie P, Kertesz, Vilmos, Van Berkel, Gary J, Schulz, Michael, Schorcht, Susanne
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Analytical Chemistry; Animals; BASIC BIOLOGICAL SCIENCES; Biochemistry; CELLULOSE; Characterization and Evaluation of Materials; Chemistry; Chemistry and Materials Science; Chromatography, Thin Layer - methods; CYTOCHROMES; Cytochromes c - chemistry; DESORPTION; Food Science; HPTLC/DESI-MS; IONIZATION; Laboratory Medicine; MASS SPECTRA; mass spectrometry; MASS SPECTROSCOPY; Molecular Sequence Data; Monitoring/Environmental Analysis; MYOGLOBIN; Myoglobin - chemistry; Original Paper; Peptide Fragments - analysis; Peptide Fragments - chemistry; PEPTIDES; PLATES; Pressure; PROTEINS; SILICA GEL; Spectrometry, Mass, Electrospray Ionization - methods; Trypsin - chemistry; Trypsin - metabolism; Water - chemistry
• ISSN: 1618-2642; 1618-2650
• DOI: 10.1007/s00216-008-1874-6

Desorption electrospray ionization mass spectrometry (DESI-MS) was investigated as a method to detect and identify peptides from tryptic digests of cytochrome c and myoglobin separated on ProteoChrom® HPTLC Silica gel 60 F₂₅₄s plates and ProteoChrom® HPTLC Cellulose sheets. Full-scan mass spectra and data-dependent tandem mass spectra were acquired in separate plate scans and used to identify peptide ions. Peptide distributions along the development lane were mapped for each separated protein digest. Signal levels ranged over several orders of magnitude. In general, highest signal levels were obtained for the peptides with the highest R f values on a plate, while peptides with very low R f values were often not detected. Sequence coverages for cytochrome c were 58% for the digest separated on the silica gel plate and 72% for the separation on the cellulose sheet; myoglobin sequence coverages were 62% and 68% on silica gel and cellulose, respectively. Weak correlations between peptide hydrophilicity and R f values on the silica gel and cellulose plates were found, with the more hydrophilic peptides having lower R f values.