Enrichment And Separation Of ω-3 PUFAs And Glyceride In Microalgae

Deep-sea fish’s main food microalgae have been regarded as the origin of polyunsaturated acids in fish. The oil extracted from microalgae is rich in DPA and DHA.ω-3 PUFAs such as DPA and DHA have been proved to show important physiological significance in respects of preventing disease of heart head blood-vessel, regulating blood glucose and blood lipid metabolism, promoting the cranial and optic nerves deveopment and recovery, improving immunity, againsting cancer and so on. Period researches indicated that ω-3 PUFAs-triglycerides are more easily to be absorbed than ω-3PUFAs-ethyl esters and free ω-3 acids. So it should be remained the nature form when extracting and enriching.The paper presents the comparison of enrichment of DPA-glyceride and DHA-glyceride from microalgae oil by frozen solvent crystallization and by High Performance Liquid Chromatography(HPLC). The paper also presents Chemical hydrolysis process optical conditions of enriched microalgae oil under frozen solvent crystallization optimization, and founding a best proportion of mobile phase to separate free DPA and DHA from hydrolysate. The themes of the paper consist of the following subjects:(1) physical-chemical properities of microalgae oil: acid value is 1.08 mgKOH/g,iodine value is 236.75 g I2/100 g, peroxide value is 4.91mmol/kg,unsaponifiable matter content is 0.66%,Water and volatile matter contents are 0.20%,saponification value is174.22 mgKOH/g. The molecular structures mainly contains cis-bonds. The main content of fatty acids includes DHA(44.64%), DPA(11.65%), C16:0(31.75%), ARA(0.70%) and EPA(0.75%).(2) DPA and DHA-glyceride enriched by frozen solvent crystallization, The optimal conditions were as follows: acetone as solvent, 1:6 volume ratio of oil to solvent, freezing time at 8h, crystallization temperature at-40 ℃. Under the optimal conditions, the contents of DPA and are DHA 14.36% and 57.47%. Physical-chemical properities of enriched microalgae oil: acid value is 0.79 mgKOH/g, iodine value is 279.845 g I2/100 g, peroxide value is 5.99mmol/kg,unsaponifiable matter content is 0.97%,Water and volatile matter contents are 0.18%. The molecular structures has not changed, high quality has been remained.(3) Separating the target tri-DHA from others by HPLC. The optimal conditions are UV detection wavelength at 205 nm, 65% methanol-35% isopropanol as a mobile phase,1mL/min of flow rate, column temperature of 25 ℃. The retention time of tri-DHA is21.83 min, and resolution is 0.96. The collection determined by GC/MS contains ten kinds of acids. The DHA content is 56.27% while the DPA’s only 3.81%. Although the effect of enrichment is not as good as frozen solvent crystallization, the DHA-glyceride can be better separated form DPA-glyceride in this way.(4) Chemical hydrolysis process optical conditions of enriched microalgae oil under frozen solvent crystallization optimization are NaOH as Hydrolysis reagent, the solution consisting of ethanol and water(10:1, v/v), Saponification time as 60 min, Saponification temperature at 50℃. The hydrolysis degree can reach as high as 99.23%, percent recovery of DPA and DHA are 71.71% and 71.39%. Separating free DHA and DPA from hydrolysate under the optical mobile phase of 65% acetonitrile-35% water by HPLC. The mass spectrum results show that they can be separated well.