蛹虫草菌深层液体发酵耦合大孔树脂吸附高效生产虫草素

doi:10.11949/0438-1157.20190076

化工学报 ›› 2019, Vol. 70 ›› Issue (7): 2675-2683.DOI: 10.11949/0438-1157.20190076

蛹虫草菌深层液体发酵耦合大孔树脂吸附高效生产虫草素

关海晴¹(),李鸿宇¹,李倩²,刘冰南¹,王际辉^1,³(),王亮¹()

1. 大连工业大学生物工程学院，辽宁大连 116034
2. 大连大学生命科学与技术学院，辽宁大连 116622
3. 东莞理工学院化学工程与能源技术学院，广东东莞 523808

收稿日期:2019-01-25 修回日期:2019-04-17 出版日期:2019-07-05 发布日期:2019-07-05
通讯作者: 王际辉,王亮
作者简介:关海晴（1994—），女，硕士研究生，<email>1446757116@qq.com</email>
基金资助:
国家重点研发计划项目(2018YFC1406800);国家自然科学基金项目(31500040);辽宁省科技厅自然科学基金计划重点项目(20170520102)

Efficient production of cordycepin during submerged liquid fermentation by Cordyceps militaris coupled with macroporous resin adsorption

Haiqing GUAN¹(),Hongyu LI¹,Qian LI²,Bingnan LIU¹,Jihui WANG^1,³(),Liang WANG¹()

1. School of Biological Engineering, Dalian Polytechnic University, Dalian 116034, Liaoning, China
2. School of Life Sciences and Biotechnology, Dalian University, Dalian 116622, Liaoning, China
3. School of Chemical Engineering and Energy Technology, Dongguan University of Technology, Dongguan 523808，Guangdong, China

Received:2019-01-25 Revised:2019-04-17 Online:2019-07-05 Published:2019-07-05
Contact: Jihui WANG,Liang WANG

摘要/Abstract

摘要：

虫草素是一种核苷类抗生素，具有抗癌、抗肿瘤等活性，主要由蛹虫草菌液体发酵生产，然而低产量限制了其应用。考虑到高浓度虫草素积累对其生物合成过程产生强烈反馈抑制，提出了应用发酵分离耦合技术移除部分虫草素、弱化反馈抑制作用以提高虫草素产量的策略。通过静态吸附实验，筛选出虫草素吸附和解吸性能良好的大孔树脂NKA-Ⅱ，并优化了大孔树脂使用量（60 g·L^-1）、吸附温度（37℃）和解吸剂种类（100%乙醇）。在30 g·L^-1葡萄糖的分批发酵体系中，经过两次树脂吸附，发酵21 d后虫草素产量达到644.50 mg·L^-1，较对照组提高32.07%；随后，将树脂吸附分离策略应用于补料分批发酵体系，虫草素产量达到787.50 mg·L^-1，较对照组提高35.78%。这些结果表明，将虫草素液体发酵与大孔树脂吸附分离耦合，有效弱化了虫草素反馈抑制，显著提高虫草素产量，为虫草素高效生产分离提供了新思路。

关键词: 发酵, 生物分离, 生物过程, 虫草素, 蛹虫草菌

Abstract:

Cordycepin, as a kind of nucleoside antibiotic, has various biological and pharmaceutical activities, such as anti-cancer and anti-tumor activities. Cordycepin is mainly obtained from liquid culture of Cordyceps militaris, whose low yield restricted its further application though. Considering the high concentration of cordycepin accumulation to produce strong feedback inhibition on its biosynthesis process, a strategy to remove some cordycepin and weaken the feedback inhibition to increase the production of cordycepin by fermentation separation coupling technique was proposed. Next, static adsorption test was taken and macroporous resin NKA-II was screened, followed by optimization of adsorption conditions such as resin usage (60 g·L^-1), temperature (37℃), and desorption solvent (100% ethanol), giving 644.50 mg·L^-1 cordycepin in fermentation under 30 g·L^-1 initial glucose with repeated adsorption processing (twice), 32.07% higher than the control. To further improve the cordycepin yield, fed-batch fermentation coupled with resin adsorption separation was performed, giving a final yield of 787.50 mg·L^-1, which was 35.78% higher than the control. It was found that the strategy of submerged liquid fermentation coupled with macroporous resin adsorption could efficiently weaken the feedback inhibition of cordycepin, and dramatically enhance the productivity of product, providing novel ideas for industrial cordycepin production.

Key words: fermentation, bioseparation, bioprocess, cordycepin, Cordyceps militaris

中图分类号:

Q 815

关海晴, 李鸿宇, 李倩, 刘冰南, 王际辉, 王亮. 蛹虫草菌深层液体发酵耦合大孔树脂吸附高效生产虫草素[J]. 化工学报, 2019, 70(7): 2675-2683.

Haiqing GUAN, Hongyu LI, Qian LI, Bingnan LIU, Jihui WANG, Liang WANG. Efficient production of cordycepin during submerged liquid fermentation by Cordyceps militaris coupled with macroporous resin adsorption[J]. CIESC Journal, 2019, 70(7): 2675-2683.

图/表 5

图1 蛹虫草菌液体发酵表型参数及动力学参数

Fig.1 Fermentation parameters and kinetics of submerged liquid fermentation by C. militaris FFCC 5111

图2 外源虫草素对蛹虫草菌生长及产物合成的影响

Fig.2 Effect of exogenous cordycepin on cell growth and cordycepin production of C. militrais FFCC 5111

表1 大孔树脂对虫草素静态吸附、解吸结果比较

Table 1 Comparison of performances of various types of macroporous resin on static adsorption and

Resin	Polarity	Pore size/μm	Adsorption capacity Q/(mg·g^-1)	Adsorption ratio E/%	Desorption ratio D/%
XAD-16	strong polarity	0.15	12.26±0.28	59.75±1.71	97.45±3.42
NKA-Ⅱ	strong polarity	85—95	24.65±0.32	98.15±0.06	85.68±0.71
AB-8	weak polarity	145—155	6.71±0.29	23.32±0.79	81.58±2.10
H103	nonpolarity	130—140	22.65±0.34	96.35±0.29	81.07±3.02

图3 吸附温度、树脂用量及解吸溶剂对大孔树脂吸附虫草素的影响

Fig.3 Adsorption and desorption capacities of cordycepin under different adsorption temperatures (a), resin dosages (b), and desorption solvents (c)

图4 大孔吸附树脂吸附分离耦合蛹虫草菌深层液体发酵

Fig.4 In situ separation and production of cordycepin during submerged liquid fermentation by C. militaris FFCC 5111 using macroporous resin adsorption

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蛹虫草菌深层液体发酵耦合大孔树脂吸附高效生产虫草素

Efficient production of cordycepin during submerged liquid fermentation by Cordyceps militaris coupled with macroporous resin adsorption

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