Study On The Nevel Blocking Remover Of Biological Enzyme And The Simulating Expenments In Laboratery

Biological enzyme extracting and deplugging technology developed in recent years is a new technology,which has good effect and high recovery rate in oil field development.Biological enzyme blocking remover have good effects in production,injection,water control,water plugging,profile control,sand control and other aspects,applied to oil well,can change the wettability,reduce interfacial tension and the viscosity of crude oil.Biological enzyme blocking remover mainly contain protease,cellulase,hemicellulase and surface active agents,in which the protease plays a certain role.In order to develop the new type of Biological enzyme blocking remover,this paper has done the following research work and come to conclusions:1.Five protease producing strains were screened from Daqing crude oil samples in preliminary.After rescreening,a protease strain S-H-5 with high production was obtained.By morphological observation,observation of growth property and 16SrDNA sequence analysis,the strains were identified.The result of identification showed that the strain S-H-5 was Pseudomonas aeruginosa.2.Through single factor test and orthogonal test,the culture mediums and culture conditions of the strain were optimized.After optimization,the optimal mediums were soluble starch 15.00g/L,peptone 20.00g/L,yeast extract 20.00g/L,NaCl1.50g/L,CaCl20.02g/L,Na2HPO40.20g/L,NaH2PO40.10g/L.The optimal culture conditions:the initial pH6.5,inoculation amount 5%(v/v),temperature 31?,shaker speed 160r/min,incubation time 72h.At this time,the protease activity of the strain was up to 628.03U/mL which was found to be 26.12 times more than that produced by the rescreened strains.3.Under the optimal culture mediums and culture conditions of the strain,growth curve and enzyme production curve were drawn.Then,separation and purification steps were used for the strain S-H-5 with fermentation liquid to centrifugal removing thallus,saturated ammonium sulfate precipitation,dialysis,concentration and Sephadex-100 gel hromatography.Purification effects were purification multiples 3.68 and the recovery rate 30.08%.By SDS-PAGE electrophoresis,the protease molecular weight was about 35.5kDa.4.Compound experiments:The protease and cellulase,xylanase and fermentation centrifugal supernatant were mixed,as a new type of biological enzyme blocking remover.Then compound effects of the novel biological enzyme blocking remover were observed and tested by shake flask culture and washing oil experiment.The novel biological enzyme blocking remover can play an important role in degrading crude oil and reducing viscosity.It can reduce the surface tension and has a certain effect of washing oil.What's more,it will have good potential application and have certain research value.