| 生物采油解堵剂中产蛋白酶菌株的筛选及培养条件优化 |
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| 作者姓名： | 左佳敏 何连芳 张玉苍 刘飞 |
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| 作者单位： | 大连工业大学资源开发利用研究所;海南大学材化学院 |
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| 基金项目： | 辽宁省工业攻关计划项目(2009223003) |
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| 摘 要： | 为研究生物酶采油解堵剂中产蛋白酶菌株的初、复筛选及培养条件优化,从大庆原油样品中筛选菌种,通过水解酪素的透明圈实验及福林酚测蛋白酶酶活的方法进行菌株的初、复筛选;以蛋白酶酶活为优化指标,采用单因素实验对筛选的产蛋白酶菌株的培养基及培养条件进行优化,优化最适培养基:可溶性淀粉为15g/L,蛋白胨为20g/L,酵母膏为20g/L,NaCl为1.0g/L,CaCl2为0.02g/L,Na2HPO4为0.2g/L,NaH2PO4为0.1g/L;在初始pH为6.0、接种量为5%(体积分数)、温度为31℃、摇床转速为160r/min的条件下,培养72h后,菌株的蛋白酶酶活为551.0U/mL,为复筛选菌株的蛋白酶酶活的22.92倍,即为菌株生长繁殖及代谢的最佳条件,能够获得更高的蛋白酶酶活,有利于后续实验的进行.结果表明:菌株产蛋白酶对原油作用效果为发酵液表面张力从作用前的56.2mN/m降低到作用后的30.5mN/m,表面张力显著降低,还有降解降黏原油等效果,具有一定的研究价值.
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| 关 键 词： | 生物采油 蛋白酶 菌株筛选 培养条件 优化 |
Screening and optimizing culture conditions of strains producing protease for bio-extraction plug removal agent in petroleum |
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| Authors: | ZUO Jia-min HE Lian-fang ZHANG Yu-cang LIU Fei |
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| Institution: | 1(1.Research Institute of Resource Exploitation and Utilization,Dalian Polytechnic University,Dalian,Liaoning 116034,China;2.School of Chemistry and Material,Hainan University,Haikou,Hainan 570228,China) |
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| Abstract: | Screening,rescreening and optimizing culture conditions of bacterial strains producing protease was studied for biological enzyme plug removal agent in petroleum.Protease producing strains were screened from Daqing crude oil samples.Namely the use of hydrolyzed casein transparent circle experiment and forint phenol method of protease activity for screening and rescreening strains.Then,to satisfy the size of protease activity indicator,by single factor test,the culture mediums and culture conditions of the strain were optimized.The strain that showed protease activity had grown well with soluble starch(15 g/L),peptone +yeast extract(20 g/L,20 g/L),NaCl(1.0 g/L),CaCl2(0.02 g/L),Na2HPO4(0.2 g/L),NaH2PO4(0.1 g/L),and the optimum culture conditions were initial pH(6.0),vaccination(5%),temperature(31 ℃),shaker speed(160 r/min),culture time(72 h).The activity of protease in the optimal conditions was up to 551.0 U/mL which was found to be 22.92 times more than that produced by the rescreened strains.At this time,the best condition that growth reproduction and metabolism of the strain and higher protease activity were obtained,which is favorable for conducting subsequent experiments.Protease produced by the strain have effects in reducing surface tension of fermentation liquid to crude oil,from 56.2 mN/m reduced to 30.5 mN/m,surface tension decreased significantly.What’s more,it can play an important role in degrading crude oil and reducing viscosity,having certain research value. |
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| Keywords: | bio-extraction of petroleum protease screening strains culture conditions optimization |
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