Based on the hydroxypropyl-β -cyclodextrin-resting cells reaction system, high substrate concentration and improved reaction rate were achieved in a bioprocess of microbial side-chain cleavage of phytosterols to 9-OH-AD. Investigation of the kinetics of biotransformation showed that the initial reaction rate was strongly inhibited by a substrate concentration greater than 8 g/L. The oxygen transfer rate was also a critical parameter that could infl uence the rate of reaction. The results highlighted the importance of dissolved oxygen, particularly at high substrate concentration. Finally, analysis of metabolites showed that the main loss of process yield was due to by-product (16.3%) and nucleus degradation (8 -11%) in the biotransformation. Investigation of the kinetics of metabolite formation provided fundamental knowledge on the biochemical reaction which was critical for understanding how to develop the biotransformation into an industrial application.

Nucleus degradation of products is one of the main problems encountered during phytosterol biotransformation. To solve this problem, the effect of temperature on nucleus degradation was investigated in the industrial production of steroid intermediates. The results are also helpful to the genetic modification of sterol-producing strains.

isms. 9 Hydroxypropyl-b-cyclodextrin (HP-b-CD), a CD derivative, has a greater capacity for complex formation than CD, and it has been recently used for the bioconversion of phytosterol to ADD and AD. 5,10 Recently, it was demonstrated that the bacterial strain Mycobacterium neoaurum NwIB-yV can be used in transforming phytosterol to 9-OH-AD. 11 The production of 9-OH-AD and the associated space-time yield reached 7.33 g L -1 and 1.22 g L -1 d -1 , respectively, from 15 g L -1 phytosterol in an aqueous system using growing M. neoaurum NwIB-yV cells, indicating a high potential for industrial appli-The steroid metabolites 9-hydroxy-androstenedione (9-OH-AD), androstadienedione (ADD) and androstenedione (AD) are important steroidal pharmaceuticals. In order to raise the production of steroid metabolites, an efficient resting cell phytosterol bioconversion process was developed to produce 9-OH-AD in the presence of hydroxypropyl-b-cyclodextrin (HP-b-CD). Cell growth medium containing phytosterol as an inducer positively improved cell activity. Under aerobic conditions, bioconversion proceeded at 70 g L -1 phytosterol in the presence of HP-b-CD (the optimized molar ratio of HP-b-CD/phytosterol was 1:1) with 30 g L -1 resting Mycobacterium neoaurum NwIB-yV cells (cell dry mass) in a 5-L bioreactor, where 9-OH-AD production and space-time yield reached 36.4 g L -1 and 9.1 g L -1 d -1 , respectively. The recycling of cells and HP-b-CD enables cost-saving and industrial applications. This bioprocess was also applied for the production of ADD and AD. The production of these steroid metabolites was much higher than that reported in previous studies.

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