Development of a Whole-Cell Bioprocess for Ursodeoxycholic Acid Production from Lithocholic Acid Using Fusarium equiseti HG18

Abstract

Ursodeoxycholic acid (UDCA) is the first-line therapy for primary biliary cholangitis and an important active constituent of bear bile. Whole-cell microbial conversion of inexpensive lithocholic acid (LCA) offers a promising alternative to conventional chemical synthesis owing to its high regioselectivity and mild reaction conditions, yet the fermentation process of Fusarium equiseti HG18 (CCTCC M2023160), a natural fungal catalyst for LCA 7β-hydroxylation, has not been systematically engineered for scalable production. This study developed a whole-cell process for UDCA production by F. equiseti HG18. Systematic optimization by single-factor experiments, Plackett–Burman design and Box–Behnken response surface methodology raised the shake-flask UDCA titer from 0.19 to 0.59 mg mL−1. Scale-up in a 3 L stirred-tank bioreactor shortened the fermentation time from 144 h in shake-flask cultivation to 96 h in the bioreactor under optimized operating conditions (pH 8.5, aeration 2.5 L min−1, and agitation 200 rpm). Experiments across a range of initial LCA loadings revealed a progressive decline in UDCA molar yield at elevated substrate concentrations, consistent with substrate-related inhibitory effects, and this loading-dependent behavior was used to design a two-stage fed-batch feeding strategy. Under the optimized condition (2.0 mg mL−1 LCA fed at 0 and 48 h), UDCA titer reached 1.71 mg mL−1, corresponding to a volumetric productivity of 17.8 mg L−1 h−1 and a UDCA molar yield of 41%. This study establishes a laboratory-scale whole-cell bioprocess for UDCA production from LCA using a wild-type fungal catalyst, integrating statistical medium optimization, bioreactor process development, and substrate-feeding strategies. The findings provide a practical framework for improving fungal whole-cell steroid biotransformation and highlight the potential of wild-type fungal platforms for scalable biocatalytic production.