Human milk fat substitute (HMFS) is a structured lipid designed to resemble human milk fat. It contains 60-70 % palmitic acid at the sn-2 position and unsaturated fatty acids at the sn-1,3 positions in triacylglycerol structures. HMFS is synthesized by the enzymatic interesterification of vegetable oils, animal fats, or blend of oils. The efficiency of HMFS synthesis can be enhanced through the selection of appropriate substrates, enzymes, and reaction methods. This review focuses on the synthesis of HMFS by lipase-catalyzed interesterification. This work provides a detailed overview of biocatalysts, substrates, synthesis methods, factors influencing the synthesis, and purification process of HMFS. Major challenges and future research in the synthesis of HMFS are also discussed. This review can be used as an information for developing future strategies in producing HMFS.