Simultaneous structural identification of diacylglyceryl‐ N ‐trimethylhomoserine (DGTS) and diacylglycerylhydroxymethyl‐ N,N,N ‐trimethyl‐β‐alanine (DGTA) in microalgae using dual Li + /H + adduct ion mode by ultra‐performance liquid chromatography/quadrupole time‐of‐flight mass spectrometry

Rationale Diacylgycerol‐ N ‐trimethylhomoserine (DGTS) and diacylglycerylhydroxymethyl‐ N,N,N ‐trimethyl‐β‐alanine (DGTA) are structural isomers that are the most commonly described betaine lipids in microalgae. The structural differentiation and precise identification of DGTS and DGTA in microalgae need to be established during mass spectrometry analysis. Methods Total lipid was extracted from Amphora sp. with CHCl 3 /CH 3 OH (1:1, v/v). The qualitative analysis of DGTS and DGTA in Amphora sp. was carried out using Li + /H + dual mode by ultra‐performance liquid chromatography/quadrupole time‐of‐flight mass spectrometry operating in MS E mode (UPLC/QTOF MS E ). Results Characteristic fragment ions [C 10 H 22 O 5 N] + at m/z 236.15 and [C 7 H 14 O 2 N] + at m/z 144.10 from the [M + H] + precursor ion can be used for the qualitative analysis of both DGTA and DGTS, whereas the loss of m/z 87 and 74 from the [M + Li] + precursor ion are specific for DGTS, and the loss of m/z 103 from the [M + Li] + precursor ion is only for DGTA. As a result, 9 DGTSs and 16 DGTAs with different fatty acids were identified simultaneously in Amphora sp. Semi‐quantitative analysis of DGTS and DGTA in Amphora sp. showed that the contents of DGTS ranged from 0.003 to 0.438 nmol mg –1 dw, and that of DGTA from 0.004 to 0.414 nmol mg –1 dw. Conclusions This is the first report to achieve the ambiguous structural identification of DGTS and DGTA by UPLC/QTOF MS E using dual Li + /H + adduct ion mode, which has remained a challenge in the past. It could provide new insights into their phylogeny and be helpful to characterize the natural phytoplankton communities as intact polar lipid biomarkers.