Studies On Biotransformation Of Phytosterol For Preparing Steroids Intermediates By Marine Derived Microorganisms

Steroid hormone drugs are important in the pharmaceutical industry and have been widely used in clinical applications.Androstenedione is an indispensable key intermediate of the production of steroid hormones drugs.Microbial transformation is an alternative to some reactions that chemical synthesis can not achieve.Androstenedione can be obtained by microbial transformation,and then as a drug intermediate can be used to produce valuable compounds of steroid hormones.In this paper,the advance of microbial production and biotransformation of androstenedione were summarized.It will give reference for further research of steroid hormone drugs.To tranform phytosterol into androstenedione,marine derived microorganism were selected for screening talented strains with phytosterol biotransformation ability.In order to improve transformation rate,microbial transformation process of androstenedione preparation was optimized.Meanwhile,biotransformation research on another biological active compound,20-hydroxymethylpregna-4-en-3-one was also proceeded.In this paper,the main research results are as follows:Enrichment cultivation and oligotrophic cultivation were applied to obtain strains with phytosterol biotransformation ability.62 marine derived strains,including 47 strains isolated from coast water sample collected from East China Sea and 15 strains stored in the laboratory,showed potential biotransformation ability.Eventually,one of the candidates named zqc-12 was used for transforming phytosterol into androstenedione.According to the morphology study and 16S r DNA nucleotide sequence and phylogenetic relationship analysis,active strain was identified as Mycobacterium sp.zqc-12.Based on the single factor and orthogonal experiment,the production process of androstenedione preparation was optimized.The optimal composition of culture medium is glucose 25 g/L,peptone 12 g/L,Na₂HPO₄ 0.7 g/L,Mg SO₄0.5 g/L.The optimal transformation condition is p H 7.5,temperature 30℃,rotate speed 180 r/min,transformation time 72h,substrate concentration 1 g/L,cosolvent is propanediol.Transformation rate of AD is 63.41%,33.4%higher than before.To enhance 20-hydroxymethylpregna-4-en-3-one preparation ability by phytosterol biotransformation,UV and nitrite mutagenesis generated a positive mutagenesis strain zqc-12-22.The optimal composition of culture medium is glucose 20 g/L,peptone 12 g/L.The optimal transformation condition is p H 7.0,the inoculum size is 10%,temperature 30℃,transformation time 48 h,substrate concentration 0.6 g/L.Under the optimal condition,transformation rate is more than 8 fold than before.