Identification of Polyunsaturated Fatty Acids Synthesis Pathways in the Toxic Dinophyte Alexandrium minutum Using 13 C-Labelling.

The synthetic pathways responsible for the production of the polyunsaturated fatty acids 22:6n-3 and 20:5n-3 were studied in the Dinophyte Alexandrium minutum. The purpose of this work was to follow the progressive incorporation of an isotopic label ((CO2)-C-13) into 11 fatty acids to better understand the fatty acid synthesis pathways in A. minutum. The Dinophyte growth was monitored for 54 h using high-frequency sampling. A. minutum presented a growth in two phases. A lag phase was observed during the first 30 h of development and had been associated with the probable temporary encystment of Dinophyte cells. An exponential growth phase was then observed after t(30). A. minutum rapidly incorporated C-13 into 22:6n-3, which ended up being the most C-13-enriched polyunsaturated fatty acid (PUFA) in this experiment, with a higher C-13 atomic enrichment than 18:4n-3, 18:5n-3, 20:5n-3, and 22:5n-3. Overall, the C-13 atomic enrichment (AE) was inversely proportional to number of carbons in n-3 PUFA. C-18 PUFAs, 18:4n-3, and 18:5n-3, were indeed among the least C-13-enriched FAs during this experiment. They were assumed to be produced by the n-3 PUFA pathway. However, they could not be further elongated or desaturated to produce n-3 C-20-C-22 PUFA, because the AEs of the n-3 C-18 PUFAs were lower than those of the n-3 C-20-C-22 PUFAs. Thus, the especially high atomic enrichment of 22:6n-3 (55.8% and 54.9% in neutral lipids (NLs) and polar lipids (PLs), respectively) led us to hypothesize that this major PUFA was synthesized by an O-2-independent Polyketide Synthase (PKS) pathway. Another parallel PKS, independent of the one leading to 22:6n-3, was also supposed to produce 20:5n-3. The inverse order of the C-13 atomic enrichment for n-3 PUFAs was also suspected to be related to the possible beta-oxidation of long-chain n-3 PUFAs occurring during A. minutum encystment.