Simultaneous extraction and fractionation of omega-3 acylglycerols and glycolipids from wet microalgal biomass of Nannochloropsis gaditana using pressurized liquids

Microalgae are primary producers of eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids in marine environment and so they are considered promising alternative sources of omega-3 long-chain polyunsaturated fatty acids (LC-PUFAs). Nannochloropsis gaditana is an oleaginous cultivated microalga with an important content of triglycerides and polar lipids which include interesting EPA amounts. In this work, new methods using pressurized liquids for simultaneous extraction and fractionation of neutral and polar lipids from we microalgal biomass are proposed. Pressurized liquid extraction (PLE) using different solvents, combined or not with pre-treatment of the alga with enzymes, allows one to enrich and fractionate neutral and polar lipids such as triacylglycerols, diacylglycerols, monoacylglycerols, free fatty acids and glycolipids. The use of enzymatic pre-treatment did not show a significant increase in the lipid yield, though a different fractionation of the microalgal lipids was achieved. PLE with a single solvent (hexane at 120 degrees C) rendered a lipid yield of 17.6%, slightly higher than the traditional Folch method (16.9%), but with a less-toxic solvent and faster extraction technique. In addition, PLE with hexane attained a high triacylglycerols fraction (17.2% +/- 3.0). The optimal four-step sequential method developed using pressurized liquids reached a good fractionation characterized by a high content of glycolipids (29.2% +/- 2.0) and monoacylglycerols (17.3% +/- 1.9), with a lipid recovery of 14.0% +/- 1.3 for the original biomass. The analysis by GC/MS of fractions obtained with each method and solvent combination, showed a different composition in fatty acids of fractionated neutral and polar lipid species, especially in saturated and omega-3 fatty acids. Therefore, the proposed fast PLE methods allow the simultaneous extraction and fractionation of the neutral and polar lipids of Nannochloropsis gaditana and produce enriched EPA fractions (up to 53%).