Preparation In EPA And DHA-Enriched Triglycerides By Enzymatic Approaches

Eicosapentaenoic acid(EPA) and docosahexaenoic acid(DHA) are mainly present as triglycerides in the fish oil. The current market fish oil product in the form of triglycerides often has low contents of EPA and DHA, with only about 18% and 12%, respectively. Therefore enhancing their content in the fish oil is of interest. This thesis employs enzymes to prepare fish oil product in triglyceride form with a total EPA and DHA content above 45%. This will provide scientific support for industrial-scal production of fish oil with high EPA and DHA content. The major contents of this research are as follows:(1) Using refined deep-sea fish oil as the starting material, lipase Amano AY“Amano”400SD was selected to effectively conduct the partial hydrolyze fish oil and hence to enrich EPA and EPA. Reaction conditions optimized include: the enzyme loading(0.2‰ of substrate weight), p H(6), mass ratio of water and oil(1 : 1), reaction temperature(35 ℃), reaction time(10 h) and agitation rate(200 r/min). In case of separating the formed glycerides and fatty acids by molecular distillation, the impact of distillation temperature was investigated and confirmed that the optimal one is 160 ℃. After two-stage molecular distillation, the heavy phase obtained was mainly glycerides that are rich in EPA and DHA, with contents of mono-, di-, and triglycerides at 0.85%, 28.3% and 70.7%. This phase has an acid value of 0.92 mg KOH/g. Compared with raw fish oil, EPA and DHA content were increased from 19.3% to 25.3% and 13.0% to 21.6% respectively. The recovery of EPA and DHA were 68.3% and 87.2%, respectively. Molecular distillation also allows the recycle of hydrolysis byproducts(fatty acids).(2) Through enzymatic transesterification, glycerides derived from the hydrolysis of fish oil were reacted with concentrated fish oil in the form of ethyl esters to prepare triglycerides. The reaction parameters optimized were: the concentrated fish oil Ethyl Ester and mixed free Oxhydryl of Glyceride mole ratio of 1:1, the loading of Lipozyme 435(2% of substrate mass), reaction temperature(60 ℃), reaction time(6 h), absolute pressure(20 Pa), stirring speed(1000 r/min). Under these conditions, the glycerides product obtained after molecular distillation contains 28.2% of EPA and 21.4% of DHA, with a triglyceride content of 97.6%.(3) Urea adduction was employed to enrich EPA and DHA in byproducts fatty acids derived from hydrolysis. The optimized operational conditions were: the mass ratio of urea to fatty acids(2.5 : 1), the mass ratio of ethanol and urea(3 : 1), adduction temperature 5 ℃, adduction time 1.5 h. After processed under these conditions, the content of EPA in fatty acids received raised from 11.4% to 47.8% and that of DHA increase from 2.10% to 11.9%, and the recovery of EPA and DHA were 70.3% and 90.4%. The fatty acids were converted into ethyl esters and reacted with the glycerides products from molecular distillation. The final glyceride product was composed of 95.9% of triglycerides, with EPA and DHA contents of 25.6% and 17.4% in the product.