Abstract: A kind of lipase derived from Rhizopus oryzae was used to study its characteristics of hydrolysis,esterification and interesterification,and applied in preparation of TG fish oil with high-content omega-3 fatty acid.Using the total contents of EPA and DHA in TG of transesterification products as index,the effects of the temperature,time,dosage of added enzyme,and substrate mass ratio on the transesterification of fish oil were analyzed.The oxidation properties of products were determined by NMR analysis.The results showed that when the additions of enzyme was 0.9%,the substrate mass ratio was 1∶1,and the reaction was conducted at 30℃ for 18 h.The total contents of EPA and DHA in TG was increased from 29.39% of raw fish oil to 44.83%.NMR analysis showed that TG fish oil has low oxidation degree when the enzymatic transesterification temperature was 30℃.Compared to this,TG fish oil products showed higher oxidation when Novozyme 435 was conducted at 50℃ of transesterification temperature.Therefore,the temperature condition of this cold-active lipase could guarantee the quality of fish oil.