Quadrupole time-of-flight versus triple-quadrupole mass spectrometry for the determination of phosphopeptides by precursor ion scanning

An API 3000 triple-quadrupole instrument and a QSTAR Pulsar quadrupole time -of-flight (TOF) mass spectrometer were compared for the determination of p hosphopeptides by precursor ion scanning in both the positive and negative nanoelectrospray ionization modes. The limits of detection for synthetic ph osphopeptides were similar (500 amol mul(-1)) for both types of instruments when monitoring precursors of -79 Da (PO3-). However, the quadrupole TOF s ystem was approximately fivefold more sensitive (1 fmol mul(-1)) than the t riple-quadrupole instrument (5 fmol mul(-1)) when monitoring precursors of 216 Da (immonium ion of phosphotyrosine). The recently introduced Q(2)-puls ing function, which enhances the transmission of fragment ions of a selecte d m/z window from the collision cell into the TOF part, improved the sensit ivity of precursor ion scans on a quadrupole TOF instrument. The selectivit y of precursor ion scans is much better on quadrupole TOF systems than on t riple quadrupoles because the high resolving power of the reflectron-TOF ma ss analyzer permits high-accuracy fragment ion selection at no expense of s ensitivity. This minimizes interferences from other peptide fragment ions ( a-, b-, and y- type) of the same nominal mass but with sufficient differenc es in their exact masses. As a result, the characteristic immonium ion of p hosphotyrosine at m/z 216.043 can be utilized for the selective detection o f tyrosine phosphorylated peptides. Our data suggest that, in addition to t heir superior performance for peptide sequencing, quadrupole TOF instrument s also offer a very viable alternative to triple quadrupoles for precursor ion scanning, thus combining high sensitivity and selectivity for both MS a nd MS/MS experiments in one instrument. Copyright (C) 2001 John Wiley & Son s, Ltd.