Characterization of cardiolipin as the sodiated ions by positive-ion electrospray ionization with multiple stage quadrupole ion-trap mass spectrometry.

The application of multiple-stage ion-trap (IT) mass spectrometric methods for the structural characterization of cardiolipin (CL), a 1,3-bisphosphatidyl-sn-glycerol that consists of four fatty acyl chains and three glycerol backbones (designated as A, B, and central glycerol, respectively), as the sodiated adduct ions in the positive-ion mode was evaluated. Following collisionally activated dissociation (CAD), the [M - 2H + 3Na]+ ions of CL yield two prominent fragment ion pairs that consist of the phosphatidyl moieties attached to the 1'- and 3'-position of the central glycerol, respectively, resulting from the differential losses of the diacylglycerol moieties containing A and B glycerol, respectively. The results are consistent with those previously described for the [M - H]- and [M - 2H + Na]- ions in the negative-ion mode, thus permitting assignment of the two phosphatidyl moieties attached to the 1'- or 3'-position of the central glycerol. The identities of the fatty acyl substituents and their positions on the glycerol backbones (glycerol A and B) are deduced from further degradation of the above ion pairs that give the fragment ions reflecting the fatty acid substituents at the sn-1 (or sn-1') and sn-2 (or sn-2') positions. The ions that arise from losses of the fatty acid substituents at sn-1 and sn-1', respectively, are prominent, but the analogous ions from losses of the fatty acid substituents at sn-2 and sn-2', respectively, are of low abundance in the MS2 product-ion spectra. This feature further confirms the assignment of the positions of the fatty acid substituents. The similar IT multiple-stage mass spectrometric approaches including MS2 and MS3 for structural characterization of CL using its [M + Na]+ and the [M - H + 2Na]+ ions are also readily applicable. However, their uses for structural characterization are less desirable because formation of the [M + Na]+ and the [M - H + 2Na]+ ions for CL is not predictable.