Characterization of Neuropeptide Processing by Fast Atom Bombardment Mass Spectrometry

Fast atom bombardment (FAB) was introduced as a new ionization technique (1 ) in 1981 by M. Barber and his coworkers. This was a breakthrough in the analysis of unstable and involatile compounds such as peptides, which were difficult to study by other ionization methods. FAB employs a particle beam consisting of the neutral atoms of xenon or argon. The gas atoms are first ionized and accelerated in the FAB gun and then neutralized on the counter-electrode. The beam, directed toward the sample that is deposited on the probe tip, ionizes and sputters charged particles that are then accelerated in the ion source and analyzed. The sample needs to be added to a viscous, chemically inert, and relatively involatile matrix, which allows a long-term analysis of a target compound. The most commonly used matrix remains glycerol. However, other substances have also been applied for the same purpose (see Section 3.5. ). In contrast to electrospray-ionization mass spectrometry (ES1 MS, see Chapter 13 ), the FAB technique produces molecular ions, corresponding to the mass of intact, charged molecules and designated as [M+H]⁺ or [M-H]⁻ (positive- or negative-ion mode, respectively). Interpretation of the data obtained is simpler than in ES1 because transformation of the spectra is not necessary, and often fragment ions appear along the spectrum, providing sequence information sufficient to reveal the primary structure of the peptide. Compared to other ionization techniques such as ES1 MS or matrix-assisted laser desorption ionization time-of-flight MS (MALDI TOF) (see Chapter 14 ), the major disadvantages of the FAB technique are: limited measuring range at a full accelerating voltage (see Note 1 ); and sensitivity (see Notes 1 and 2 ). Moreover, strong signals belonging to the matrix clusters may obscure analysis. Problems concerning sensitivity and possible connection to liquid chromatography/capillary electrophoresis systems have successfully been overcome by application of continuous-flow FAB (CF FAB, dynamic FAB) where the sample solution is loaded into the instrument via a fused silica capillary (2 ).