Bacterial Diversity Analysis In Tobacco Waste Extract And Isolation,Identification And Application Of Nicotine-degrading Bacterium Pseudomonas Sp.JY-Q

Tobacco waste extract(TWE)is an important material used in reconstituted tobacco sheet technology.The content of nicotine in TWE is a critical factor that can influence the quality of reconstituted tobacco.Nicotine,as an alkaloid in Solanaceae plant,is an important component of tobacco and also a kind of environmental toxic substances.Some microbes can use nicotine by different metabolic pathways.In this study,bacterial diversity analysis of tobacco waste extract(TWE)derived from Hangzhou Liqun Environment Protecting Paper Co.Ltd was carried out.The results indicated that there are lots of bacterial genera in TWEs.Pesudomonas and Lactobacillus are the dominant bacteria in different TWEs.The analysis results are useful in studying application in reconstituted tobacco of microbe.Then,a nicotine-degrading bacterium was isolated from the original tobacco waste extract(TWE).The strain can grow with nicotine as sole carbon,nitrogen source and energy.After identification,the strain belonged to Pesudomonas and was named as Pesudomonas sp.JY-Q.Pesudomonas sp.JY-Q could completely degrade 3 g/L and 5 g/L nicotine in basic inorganic salt medium(BSM)within 12 h and 24 h,respectively.Strain JY-Q can grow with the nicotine concentration as high as 10 g/L.The optimal temperature and initial pH for nicotine degrading by strain JY-Q is 37? and 6.5-7.0.Pesudomonas sp.JY-Q could also completely degrade nicotine in 5%TWE(nicotine concentration 1.113 g/L)in 12 h.The addition of(NH4)2SO4 and yeast extract could effectively improve the ability of degrading nicotine in TWE by the strain JY-Q.Immobilized cells of JY-Q also could completely degrade nicotine in BSM(nicotine concentration 2g/L)and degrade 85.47%nicotine in 5%TWE within 24 h.In 30 L fermenter,JY-Q could degrade 87.24%nicotine in 5%TWE within 12 h.The metabolites of nicotine degradation by JY-Q were detected via GC-MS.Three intermediate metabolites were detected by GC-MS as 3-(2,3,4-dihydro-3H-pyrrol-5-yl)-pyridine,nicotyrine and 2,3'-bipyridine,suggesting that Pesudomonas sp.JY-Q might have a novel metabolic pathway using nicotine as substrate.Pseudomonas sp.JY-Q and its immobilized cells could effectively degrade nicotine in BSM and TWE,suggesting that it is potential strain for application to improve the quality of reconstituted tobacco and to reduce pollution of nicotine.