Abstract The biosynthesis of structured lipids (SLs) was carried out by the interesterification of flaxseed oil (FO) and tricaprylin (TC) in an organic solvent medium (OSM), using selected commercial lipases, including Amano DF, Novozym 435, Lipozyme TL-IM and Lipozyme RM-IM. The fatty acyl chains of the synthesized triacylglycerols (TAGs) were identified by atmospheric pressure chemical ionization/mass spectrometric (APCI/MS) analysis, while the fatty acid positional distribution of the MLM- and MML-SLs (M-medium and L-long chain fatty acids) was determined by silver-ion high-performance liquid chromatographic (Ag+/HPLC) analysis. The effects of reaction temperature (Tr, 30–50°C), enzyme concentration (Ec, 0.5–4%, w/v), initial water activity (aw, 0.05–0.43) and reaction time (Rt, 0–72h) on the efficiency of the enzymes, were studied. The bioconversion yield (%) of the synthesized MLM- and MML-SLs was monitored under the established reaction parameters for each lipase. The maximum yield of MLM-SLs was obtained in the order, of Novozym 435>Lipozyme TL-IM>Lipozyme RM-IM>Amano DF. Moreover, considering the ratio of the MLM- to MML-SLs produced by each enzyme, Novozym 435 and Lipozyme TL-IM were selected as the most effective enzymes for interesterification of FO and TC.