Study On Mutagenic Screening And Fermentation Technology Of Lipase Producting Strains

Rhizopus arrhizus is a common industrial microorganism and a food-safe bacterium certified by the USFDA.Rhizopus arrhizus lipase is used in many fields such as medicine,food,chemical industry and so on.However,the current lipase production of Rhizopus arrhizus is low.In order to obtain the target strain with strong lipase production ability,Rhizopus arrhizus need to be mutated and selected.Its lipase activity was further improved through the optimization of the fermentation process.Based on this purpose,the following works were done by this article based on Rhizopus arrhizus.1.Rhizopus arrhizus were induced by UV+LiCl mutagenesis and DES mutagenesis.First,determine the optimal mutagenesis conditions through experiments.Then,Rhizopus arrhizus was subjected to ultraviolet+LiCl mutagenesis,and finally the enzyme activity of one target strain was 522 U/mL,number Z-3.Using Z-3 as the starting strain for DES mutagenesis,21 strains with higher enzyme activity were screened after the initial screening,and screened according to the above rescreening process to obtain a target strain 55#,which was verified by genetic stability test.The enzyme activity was stable at 620 U/mL,which was 13.5 times the enzyme activity before mutation(46 U/mL).2.In order to improve the lipase yield of Rhizopus arrhizus during fermentation,the paper optimized the culture conditions and medium components during fermentation.First,the morphology and inoculum amount of the bacteria during fermentation were determined,and then the components in the fermentation medium were explored and optimized.The induced oil,nitrogen source,carbon source and inorganic salt ions were studied by shake flask fermentation.The effect and influence of various factors such as the culture conditions on the enzyme production of Rhizopus arrhizus fermentation were determined.The optimal inoculation amount was 15%,and the optimal medium was Tryptone 10 g/L,(NH4)2SO4 3 g/L,Glucose 20 g/L,KH2PO41 g/L,MgSO4·7H2O 0.5 g/L,corn oil 10 g/L,in the form of fungus balls at fermentation pH 7,shaker speed 200 r/min,30? Fermentation was carried out under the condition that the highest enzyme activity was 3068 U/mL,which was 4.95 times before the optimization of the fermentation process.3.The lipase in the fermentation broth was separated by cold acetone method,the pre-treated fermentation and acetone were slowly mixed under ice bath condition,and then centrifuged at 4?,6000 r/min for 6 min to remove the supernatant Recovery of acetone,the remaining precipitate was taken out and placed in a flat dish,allowed to dry naturally in a fume hood and ground into a powder,its enzyme activity was 224800 U/g.The enzyme activity yield was 46.9%.