油水双相体系中分枝杆菌降解植物甾醇产9-羟基雄烯二酮的工艺

Biotransformation of phytosterols to 9α-hydroxyandrostenedione by Mycobacterium sp. LY-1 in an oil-water two-phase system

9α-羟基雄甾-4-烯-3,17-二酮(9-羟基-雄烯二酮,9α-OH-AD)是一种重要的甾体药物中间体,主要用于糖皮质激素类和性激素类药物的生产,可由微生物转化法降解植物甾醇而得.植物甾醇在水中的溶解度极低,生物可利用度不高,严重制约了微生物对植物甾醇底物的利用效率.为进一步提高目的产物9α-OH-AD的摩尔得率,以前期诱变筛选的Mycobacterium sp.LY-1作为出发菌株,考察不同HLB值的表面活性剂对9α-OH-AD摩尔得率的影响,筛选得到最适表面活性剂,并建立高效的油水转化体系(油水体系的HLB=7.3).结果表明,HLB值为15的表面活性剂Tween-80有利于提高9α-OH-AD的摩尔得率,当添加4.0 g/L Tween-80时,9α-OH-AD的摩尔得率达到38.5%.在该工作基础上,发现添加促溶剂100.0 g/L的大豆油对分枝杆菌转化植物甾醇生成9α-OH-AD具有促进作用.同时,添加4.0 g/L Tween-80和100.0 g/L大豆油时,当植物甾醇质量浓度为30 g/L时,9α-OH-AD摩尔得率为36.9%,9α-OH-AD质量浓度达到8.17 g/L,较对照提高了324.1%.本研究表明建立的油水转化工艺体系在一定程度上增加了底物质量浓度,提高了菌体的转化能力,结果可为甾醇生物转化体系的研究提供重要的参考信息.

9α-Hydroxyandrost-4-ene-3,17-dione(9α-OH-AD),via the microbial conversion of phytosterols,is an important precursor in the manufacture of numbers of modern glucocorticoid and sex hormone drugs.The solubility of phytosterols was extremely low in water,and the bioavailability was not high,which seriously restricts the utilization efficiency of microorganisms on phytosterols.To increase the molar yield of 9α-OH-AD further,Mycobacterium sp.LY-1 was used as a starting strain by the previous mutagenesis,and the effects of the surfactants with different hydrophilic-lipophilic balance(HLB) values on the biotransformation efficacy of phytosterols to 9α-OH-AD were investigated.The most suitable surfactant was chosen and a highly efficient conversion system of oil-water(HLB=7.3) was established.The results showed that the surfactant Tween-80 with an HLB value of 15 increased the molar yield of 9 a-OH-AD.When 4.0 g/L Tween-80 was added,the molar yield of 9α-OH-AD reached 38.5%.Then,it was found that the addition of the solubilizing agent 100.0 g/L soybean oil promoted the transformation of phytosterol to 9α-OH-AD by Mycobacterium sp.LY-1.When 4.0 g/L Tween-80 and 100.0 g/L soybean oil were added,the molar yield of 9α-OH-AD reached 36.9%,and the quality concentration of 9 a-OH-AD was 8.17 g/L;this was 324.1% higher than the control treatment at a higher phytosterols concentration of 30 g/L.The results indicated that the oil-water conversion process system established in this study could increase the substrate concentration and improved the transformation ability of the Mycobacterium.This might provide a basis for industrial application.