Activity Screening And Antibacterial Mechanism Of Crude Extracts Of Aspergillus Niger

Aspergillus plays a significant role in fermentation,food,food storage,medicine and feed industry.In which Aspergillus niger is an excellent strain for industrial fermentation having advantages of safety and containing many active enzymes.Containing a variety of significantly active enzymes makes Aspergillus niger important strain for industrial fermentation.Aspergillus niger is generally recognized as safe(GRAS)organisms,enzyme preparations produced by using Aspergillus niger is not only safe,reliable,but also growing faster,having shorter fermentation period,thus having obvious advantages.The effect of Aspergillus niger strain on industry has gained more and more attention,however,there are less studies and reports on bacteriostasis of Aspergillus niger.To rationally develop and use the fungus resource,we explored the antagonism and mechanism of an Aspergillus niger xj which has got the proprietary intellectual property rights against Agrobacterium tumefaciens,Erwinia aroideae,Sclerotium rolfsii,Alternaria alternate and other bacteria for common agricultural diseases,which provides theoretical basis for researching and developing new type of environment-friendly broad-spectrum microbial fungicides,main research results are as follows:1.The research took bran as solid fermentation and culture medium for Aspergillus niger,single-factor test was first conducted to preliminarily screen the effect of culture temperature,cultivation temperature,inoculation quantity,pH values,carbon sources,nitrogen sources,other content,light and mixing on sporulation of Aspergillus niger xj.PB test and was conducted next,and finally determine that conditions for response surface analysis to optimize solid fermentation of Aspergillus niger:add MgSO4 1.6‰,glucose 10%,(NH4)2SO41‰,CaCO3 1.6%,CaSO4 1%;illuminate for 22.83h per day,and then agitate after 71.31h culture.Under such conditions,sporulation was 2.636 times higher than that before optimization,and is 4.58 times to PDA culture medium.Then extract mycelium of Aspergillus niger xj by conventional solvent to get ethyl acetate,petroleum ether,n-butyl alcohol,95%alcohol,as well as water extraction and ethanol precipitation,total 5 kinds of polarity isolation productions;gain production of supercritical extraction.Among these,supercritical extraction gained 9 productions through GC-Ms,in all known 5 compounds,hydroxymethyl furfural(5-HMF,90.194%)and 5,5'-(Oxydimethylene)-di-2-furaldehyde(7.843%)had highest contents.2.For target bacteria,median inhibitory concentration(IC50)of four segregated productions to Agrobacterium tumefaciens and Erwinia aroideae was obtained through MTT,it was found that inhibitory effect of isolated productions to two bacteria were,in order,ethyl acetate<n-butanol<water extracting and alcohol precipitation<supercritical extraction.3.For the target fungi,it was found through growth rate method that supercritical extraction and water extracting and alcohol precipitation showed stronger inhibitory ability to two bacteria,inhibitory rate aiming at Sclerotium rolfsii reached 93.22%,79.67%,74.11%respectively on the seventh day.Also,inhibitory rate aiming at Alternaria alternate reached 90.89%,61.89%,57.11%respectively on the seventh day.All polarity extractions showed inhibitory effect on mycelia growth and sporulation of two fungi.4.After treatment of supercritical extraction,water extracting and alcohol precipitation and n-butanol to fungi,bacteria made percentage of solution electrical conductivity highly increase in a short time,and reached the peak after 2h,3h measurements,and solutions with various concentration were higher than percentage of electrical conductivity in control solutions.5.Apart from Agrobacterium tumefaciens,other target fungi and bacteria have leakage of nucleic macromolecule substances at different levels,and leakage increased with time.After treating Agrobacterium tumefaciens,it reached the peak at 4h and then decreased,and it showed that for Agrobacterium tumefaciens,best inhibitory effect was between 3?4h.6.Average activity of succinate dehydrogenase and Malic dehydrogenase in control group of target bacteria was higher than that after drug treatment,it showed that different extractions of Aspergillus niger xj had certain effects on activity of two enzymes of target bacteria.7.In range of experimental concentration,clearance rate of water extracting and alcohol precipitation to DPPH free medium had positive dose-effect relationship with quality concentration,thus with the increase of quality concentration,clearance rate to DPPH free medium increased.When quality concentration was lower than 45mg/mL,clearance rate of supercritical extraction and n-butanol to DPPH free medium had positive dose-effect relationship with quality concentration,then with the increase of quality concentration,the clearance rate hardly changed.When quality concentration was lower than 5mg/mL,clearance rate of Vc,Trolox and BHA to DPPH free medium had positive dose-effect relationship with quality concentration,then with the increase of quality concentration,the clearance rate hardly changed.Clearance rate of all segregation sections to ABTS free medium had positive dose-effect relationship with quality concentration,with the increase of quality concentration,clearance rate to ABTS free medium increased.Section of water extracting and alcohol precipitation linearly increased before 60mg/mL,whose inhibitory rate reached 97.41%at 80mg/mL.