Fig. 3

UPLC-MS of phospholipids extracted from E. coli grown in the presence of 20:3 fatty acid. [M-H]⁻ ions were detected by quadrupole mass spectrometry following electrospray ionization. a Overlain extracted ion chromatograms corresponding to the [M-H]⁻ ions of the indicated parent phospholipid. Note that the chromatographic gradient fully resolves species that differ by one double-bond; the extra double-bond increases the polarity such that PE (16:1/20:3) elutes 0.5 min earlier than PE (16:0/20:3). The control culture was analyzed for all of these same peaks, but none were detected. b Mass spectrum of the large peak at 8.5 min. The singly charged parent ion at 740.5 m/z has mass consistent with a PE (36:3) species. Direct observation of the cone fragments at 255.2 and 305.2 m/z, corresponding to 16:0 and 20:3 carboxylate ions respectively, confirm that the parent phospholipid is PE (16:0/20:3). Thus, it is confirmed that cultures grown in the presence of 20:3 fatty acid are capable of incorporating the supplemented PUFA into the membrane phospholipids. All other PUFAs tested, except 22:6, were likewise observed to be incorporated (Supplemental Figure 1)