前体物质对Glarea lozoyensis发酵产生纽莫康定B0的影响

摘要

纽莫康定B0是棘白菌素类化合物,其半合成衍生物卡泊芬净已被美国食品药品监督管理局(FDA)批准用于治疗无效或无法耐受其他抗真菌药的患者的侵袭性曲霉病.然而,通过发酵产生的纽莫康定B0的效价较低,限制了卡泊芬净的工业生产.本研究将优化好的的培养基进行了改良,添加了4种前体物质.研究结果表明,大豆油、亮氨酸、异丁醇和甲硫氨酸显著影响发酵单位;进一步的正交试验结果表明,大豆油(A)对纽莫康定B0效价的影响极显著(P＜0.05),亮氨酸(B)、异丁醇(C)和甲硫氨酸(D)影响次之,最优发酵条件为A2B2C3D2即大豆油6.0g/L,亮氨酸1.0g/L,异丁醇4.0ml/L,甲硫氨酸0.3g/L时,纽莫康定B0发酵单位达到1295mg/L左右.此外,还应用正交试验验证各因素的显着性.当将kLa作为指导参数,使用A315型叶轮的100L发酵罐进行放大和工业规模生产时,与原始效价相比,纽莫康定B0产量增加了361％,并达到了2250mg/mL的水平.%Pneumocandin B0 is the acylated cyclic hexapeptides of the echinocandin family.Its semisynthetic derivative caspofungin has been approved by FDA for the treatment of invasive aspergillosis in patients who fail to respond or are unable to tolerate other antifungals.However,the titer of pneumocandin B0 by fermentation is low,which would influence the production of caspofungin.Therefore,it is necessary to achieve high-level production in the fermentation culture.In this study,the medium previously developed was modified,in which four precursor substances were added.In addition,orthogonal experiment was applied to verify the significance of each factor.The results of orthogonal experiments revealed that soybean oil significantly affects the titre of pneumocandin B0(P＜0.05) and that the effects of soybean oil were followed by those of leucine (B),isobutanol (C) and methionine (D).The optimized fermentation conditions were A2B2C3D2 (i.e.,soybean oil 6.0g/L,leucine 1.0g/L,isobutanol 4.0mL/L and methionine 0.3g/L).When kL a was selected as an index for scale-up and industrial scale production using a 100L fermenter with the A315 impeller,pneumocandin B0 production has successfully increased by 361％ and has reached 2250mg/mL,comparing to the original titer.