Structural elucidation of co-eluted triglycerides in the marine diatom model organism Thalassiosira pseudonana by ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry.

Rationale: The precise identification of fatty acids at the sn-2 position of triacylglycerols (TAGs), especially for positional regioisomers (AAB/ABA), needs to be established during mass spectrometry analysis. The detailed structural information about TAGs is significant not only for the assessment of biofuel quality, but also for the tracing of biosynthetic precursors.

Methods: Total lipid was extracted from T. pseudonana by a modified Bligh and Dyer method. The qualitative analysis of TAGs in T. pseudonana was carried out using ultra-performance liquid chromatography/electrospray ionization-quadrupole time-of-flight mass spectrometry (UPLC/ESI-Q-TOF-MS). The raw LC/MS data were analyzed using MassLynx software (version 4.1, Waters).

Results: The acyl group at the sn-2 position of the TAGs has been identified unequivocally by [M + Li-R1/3COOH-R2CH=CHCOOH](+) and the abundance of [M + Li-R1/3COOH-R2CH=CHCOOH](+) can be used to confirm whether the TAG isomers are co-eluted. In total, twelve TAGs were identified in T. pseudonana based on the fragmentation patterns discussed above. The data indicated that only C16 fatty acids were located at the sn-2 position, which was important to trace the biosynthetic precursors of TAGs.

Conclusions: We put forward a hypothesis that TAGs in T. pseudonana are only derived from lipids in chloroplasts through prokaryotic biosynthesis pathway based on the precise information of sn-2 fatty acids, which is significant not only for the assessment of biofuel quality, but also for the tracing of biosynthetic precursors.