Study Of High-efficiency Fermentation Of Surfactin And Its Application In Antibacterial

Surfactin is one of the most powerful biosurfactants produced by various Bacillus subtilis,presenting excellent surface activity,emulsifying activity,antibacterial,antiviral,anti-tumor,hemolytic effects,biodegradability and biocompatibility.It has wide potential applications in biological control,food preservation,cosmetics,biomedical industries,petroleum and environmental remediation.However,the low production level resulted by low yield as well as the lab-scale fermentation caused high procution cost which greatly inhibit the real industrial production of surfactin.Therefore,surfactin is unable to compete with low-cost chemical surfactants in the field of conventional applications fields.In this study,the fermermentation of surfactin was aimed to be imporved by optimizing the fermentation medium,and the optimized fermentation process will be scale-up to a pilot scale of 1000 L.In addition,some modification of surfactin via DCC reaction will be conduted to improve the biological activities of surfactin and the feasibility of application in antibacterial will also be evaluated.The main findings are as followed:(1)In this study,the addition of ferrous ions,ferric ions and metal chelating agents are selected to optimize the fermentation of surfactin by B.subtilis ATCC 21332.As we found,EDTA is for the first time proved to be of great significance in promoting surfactin production in the presence of iron ions.The addition of 0.2 mmol/L EDTA-Fe²⁺got a maximum yield of5.32 g/L which was about 7 times over the regular fermentation.Furthermore,this high efficiency fermentation of surfactin was then scaled up to 1000 L after a seires of optimizations on culture parameters,the corroponding productivity of surfactin was further imporved to 0.123 g/(L·h)which was 66%higher than lab scale.After that,the surfactin was extracted by dichloromethane/methanol system,the recovery as well as purity was 97.5%and90%,resepectively.(2)The carboxyl group on the peptide ring of surfactin was modified by DCC modification method for the first time,then we systematically evaluated the surface activity and biological activity of surfactin and its derivatives.The results showed that arg-surfactin can reduce the surface tension of water to 38.4 m N/m at 20 mg/L,and its HC₅₀ was 50 mg/L.But surfactin exhibit better surface activity(reduce the surface tension of water to 27 m N/m at a concentration of 13 mg/L),and lower hemolytic activity(HC₅₀of 90 mg/L).This may be due to the stronger hydrophilicity of arginine group,and the positive charge of arginine group is conducive to the interaction between surfactin and acid phospholipids with negative charge on cell membrane.(3)Furthermore,we investigated the antibacterial effect of surfactin and arg-surfactin.The results showed that surfactin had a great inhibitory activity against S.aureus,while arg-surfactin can efficient inhibit the cell growth of Moesziomyces sp.This may be related to their different antibacterial mechanisms.Moreover,the antibacterial activity of arg-surfactin was not affected by p H and salinity,while surfactin would lose its antibacterial activity under acidic conditions.In addition,Zn²⁺and Cu²⁺could slightly improve the antibacterial activities of surfactin and arg-surfactin which was at low concentrations.Compared with traditional antibiotics such as gentamicin,erythromycin and tetracycline,arg-surfactin has stronger inhibitory effect on Moesziomyces sp.However the antibacterial effect of their combination is not obvious,which may be due to the insensitivity of antibiotics to Moesziomyces sp.The excellent bioactivity and biocompatibility of surfactin and its derivatives show great potential in the application of pharmaceutical or biological pesticide additives.