This is the first research to show that 3-chloro-1,2-propanediol (3-MCPD) was formed upon the action of 0.1% pancreatin at the early stage of soy sauce fermentation with Aspergillus oryzae and mixtures of soy flakes and wheat flour. A series of experiments were then performed to gain insight into such 3-MCPD formation. Results showed that adding 0.1% pancreatin causes slightly decreased reducing sugar content but significantly increased 3-MCPD, without significant changes in amino nitrogen, total nitrogen, and total acid contents. 3-MCPD was formed by the action of different exogenous lipases (pancreatic phospholipase, Candida antarctica lipase B [CALB], and phospholipase A2). Further, soybean oil (not lecithin) could facilitate the 3-MCPD formation upon the action of CALB, phospholipase A2, and pancreatic lipase due to the substrate specificity. Without adding such enzymes, all soy sauce koji with soybean oil had detectable 3-MCPD, indicating that lipase may bind to the substrate via covalent bonding and chloride ions exert nucleophilic attack to form 3-MCPD esters (which were hydrolyzed to release 3-MCPD). Further studies are required to clarify precise mechanisms underlying the 3-MCPD formation in the presence of exogenous lipase especially phospholipases. Practical applications To avoid the prevalence of 3-MCPD brewing in soy sauce, this manuscript gives the theoretical basis.