SAMPLE PREPARATION FOR THE ANALYSIS OF GLYCEROPHOSPHOLIPIDS BY MATRIX-ASSISTED POSITIVE AND NEGATIVE-ION CF-252 PLASMA DESORPTION TIME-OF-FLIGHT MASS-SPECTROMETRY

A new sample preparation technique for the analysis of 13 different cl asses of glycerophospholipids by positive- and negative-ion Cf-252 pla sma desorption time-of-flight mass spectrometry is described, Until re cently only the direct deposition and the electrospray deposition tech nique, both yielding mass spectra of poor quality, were applied, The i ntroduction of the bifunctional small molecule matrix 3-(3-pyridyl) ac rylic acid (PAA) as sample matrix brings the advantage of selective an alyte adsorption from chloroform solution (in two cases also from pure aqueous solution: phosphatidylinositol-4-phosphate and phosphatidylin ositol-4,5-diphosphate) onto the matrix surface allowing a novel two-s tep rinsing procedure. Prior to this rinsing procedure, abundant alkal i-cationised molecules and an extensive number of diagnostic fragment ions are observed for these compounds, During the first rinsing step e xcess of alkali cations is removed hy cleaning the matrix surface with up to a few mt of water, After the second rinsing step, where chlorof orm is used, the intensity of (de)protonated molecular ions and adduct ions with the matrix increases significantly, Most of the fragment io ns exhibit a significant decrease in relative abundance or even disapp ear due to lowering the internal. energy of desorbed/ionised molecules by removal, of non-adsorbed analyte layers from the matrix surface, I n cases,where analyte molecules do not form protonated and deprotonate d molecular ions ([M + H](+) or [M - H](-)), respectively, intense pea ks due to adduct ions ([M + PAA + H](+) and [M + PAA - H](-)) between matrix (PAA) and analyte (M) molecules are observed, These adduct ions most likely originate via a common mechanism independent of the chemi cal nature of the glycerophospholipid. Choline-containing phospholipid s, which do not form [M-H]ions, most likely form ion-pairs with the ma trix already in the condensed phase yielding [M + PAA - H](-) ions in the negative-ion mass spectra. For the nitrogen-free phospholipids (e. g. phosphatidylinositol), which do not form [M + H](+) ions, ion-pair formation between lipid and matrix in the condensed phase is also assu med to be the major process of [hi + PAA + H](+) ion formation in the postive-ion mode, Applying this sample preparation technique to the an alysis of normal phase HPLC-purified glycerophospholipid-fractions fro m complex biological sources (e.g. human brain, rat lung lavage fluid or coniferous needles) sensitivities in the low ng to mu g range were obtained by positive- and negative-ion plasma desorption time-of-fligh t mass spectrometry.