Toward Total Structural Analysis of Cardiolipins: Multiple-Stage Linear Ion-Trap Mass Spectrometry on the [M − 2H + 3Li]+ Ions

ESI multiple-stage linear ion-trap (LIT) mass spectrometric approaches for a near-complete structural characterization of cardiolipins (CLs), including identification of the fatty acyl substituents, assignment of the fatty acid substituents on the glycerol backbone, and location of the double-bond(s) or cyclopropyl group along the fatty acid chain are described. Upon collisionally activated dissociation (CAD) on the [M − 2H + 3Li]+ ions of CL in an ion-trap (MS2), two sets of fragment ions (designated as (a + 136) and (b + 136) ions) analogous to those previously reported for the [M − 2H + 3Na]+ ions were observed, leading to assignment of the phosphatidyl moieties attached to 1′- or 3′-position of the central glycerol. Further dissociation of the (a + 136) (or (b + 136)) ions (MS3) gives rise to the (a + 136 − R1(or 2)CO2Li) (or b + 136 − R1(or 2)CO2Li) ion pairs that identify the fatty acid moieties and their position on the glycerol backbone. This is followed by MS4 on the (a + 136 − R1(or 2)CO2Li) (or b + 136 − R1(or 2)CO2Li) ion to eliminate a tricylic glycerophosphate ester residue (136 Da) to yield the (a − R1(or 2)CO2Li) ion, which is then subjected to MS5. The MS5 spectrum contains the structural information that locates the double-bond(s) or cyclopropyl group of the fatty acid substituents. Finally, the subsequent MS6 on the dilithiated fatty acid ions generated from MS5 also yields feature ions that confirm the assignment.

Graphical AbstractThe chemical structure of cardiolipin contains four fatty acyl residues (R1, R2, R1', and R2'), three glycerol backbones (A, B, and C), and two phosphate groups.Figure optionsDownload high-quality image (33 K)Download as PowerPoint slide