High lipid heterotrophic microalgae (HM) Schizochytrium limacinum is a good dietary source of long-chained polyunsaturated fatty acids. HM biomass has successfully been used in aquafeeds. However, the high saturated fatty acid content of the triacylglycerol (TAG) could limit the applications of HM as a main feed lipid source. Enzymatic interesterification of HM oil with unsaturated oils may increase the utilization efficiency and remove the technical challenges in using such oils. In the present study, extracted oil from HM biomass (Alltech Inc.) with rapeseed oil are mixed, and interesterified enzymatically with either Lipozyme RM IM or Lipozyme 435 in reactions with no, addition or 5% addition, of distilled water. The experimental oil mixes are formulated to target a total fatty acid profile similar to fish oil. No addition of water to the reaction mixture leads to more efficient TAG recovery after interesterification. Overall, enzymatic interesterification of HM and rapeseed oils with Lipozyme RM IM produces oils with lower levels of tri-saturated TAG isomers, higher content of TAG isomers with unsaturated fatty acids and a lower slip melting point. Animal studies need to be performed to evaluate the biological effects of interesterified against unprocessed highly unsaturated oils. Practical Applications: Heterotrophic microalgal oil as a pure product could have limited application in feeds and foods due to its structural content of triacylglycerol (TAG) with saturated fatty acids in all three positions, due to both nutritional and practical reasons. Enzymatic interesterification of HM oil with rapeseed oil is in the present study shown to be an efficient technology to produce oils with more desirable TAG composition and reduced melting point for feed and food applications compared to the original raw material. Content of selected triacylglycerol (TAG) isomers in rapeseed oil (RO), extracted heterotrophic microalgae (HM) oil, and oil mixtures following enzymatic interesterification in reaction with no addition of water (Trial 2) by the use of Lipozyme 435 or Lipozyme RM IM.