*Article* **Correlation Relationship between Phase Inversion of Pickering Emulsions and Biocatalytic Activity of Microbial Transformation of Phytosterols**

**Wenyu Zhao <sup>1</sup> , Haisheng Xie <sup>1</sup> , Xuehong Zhang <sup>2</sup> and Zhilong Wang 1,\***


**Abstract:** Microbial transformation of hydrophobic phytosterols into the pharmaceutical steroid precursors AD (androst-4-ene-3, 17-dione) and ADD (androst-4-diene-3, 17-dione) in a water–plant oil two-phase system by *Mycolicibacterium neoaurum* is a paradigm of interfacial biocatalysis in Pickering emulsions stabilized by bacterial cells. In the present work, phase inversion of Pickering emulsions—i.e., Pickering emulsions turning from water-in-oil (W/O) emulsions into oil-in-water (O/W) ones—was observed during microbial transformation in the presence of high concentrations of crystal phytosterols. It was found that there is a correlation relationship between the phase behaviors of Pickering emulsions and the biocatalytic activity of utilizing *M. neoaurum* as a whole-cell catalyst. Efficient microbial transformation under the high crystal phytosterol loadings was achieved due to the formation of O/W emulsions where interfacial biocatalysis took place. Under the optimal conditions (volume ratio of soybean oil to water: 15:35 mL, phytosterols concentration in the soybean oil: 80 g/L, glucose as co-substrate in the aqueous culture medium: 10 g/L), the concentrations of AD and ADD reached 4.8 g/L based on the whole broth (16 g/L based on the oil phase) after microbial transformation for 9 days.

**Keywords:** microbial transformation; phytosterols; Pickering emulsion; phase inversion; plant oil
