Abstract High sn ‐2 docosahexaenoic and arachidonic acid oils (DHAO m and ARAO m , respectively) were produced independently via enzymatic interesterification of DHA‐rich and ARA‐rich single cell oils (DHASCO and ARASCO, respectively) using a mix of immobilized lipases, Lipozyme ® TL IM and Novozym ® 435 (weight ratio 1:1) as the biocatalyst system. Response surface methodology (RSM) was employed to model and optimize the reactions conditions. Three independent variables, namely reaction time, reaction temperature, and enzyme load, were investigated in DHAO m and ARAO m models. The prediction power of the model was further confirmed by solvent‐free scale‐up reactions of 100 g per batch. Final results showed that DHAO m contained 46.53 mol% of total DHA (49.70 % at the sn ‐2 position), while ARAO m contained 47.25 mol% of total ARA (36.08 % at the sn ‐2 position). This represents a significant increment in the amount of DHA and ARA at the sn ‐2 position when compared to DHASCO (47.8 mol%; 30.30 % at the sn ‐2) and ARASCO (47.79 mol%; 28.50 % at the sn ‐2), respectively. These products have potential as additions to infant formulas where DHA and ARA supplementation is required.