Interesterified Fat Facilitated the Synthesis of <scp>UPU</scp> Structured Lipid Enriched With α‐Linolenic Acid Through <scp> <i>Rhodococcus opacus</i> </scp> Fermentation

ABSTRACT Rhodococcus opacus has been reported to produce sn ‐1,3‐unsaturated fatty acid‐2‐palmitic acid triglyceride (UPU). Carbon source selection critically impacts fermentation efficiency. This study evaluated four carbon sources, revealing that interesterified fats yielded significantly higher UPU production than fatty acids, ethyl esters, and physically blended fats. The resulting microbial oil contained 22.60% palmitic acid and 36.43% α‐linolenic acid, forming a characteristic 1:1.6 (w/w) ratio. Positional analysis showed palmitic acid constituted 48.52% of sn ‐2 fatty acids, with 71.55% of total palmitic acid selectively esterified at the sn ‐2 position. The principal UPU triglycerides were characterized as OPLn (12.76%), LnPLn (10.75%), and LPLn (8.68%), collectively accounting for 45.40% of total triglycerides. The superior performance of interesterified fats is attributed to their lower melting point, generation of partial glycerides, and homogeneous lipid matrix, facilitating microbial utilization. These findings establish interesterified fats as the optimal carbon source for UPU production, providing a foundational fermentation strategy for structured lipid biosynthesis.