Solid-State Fermentation Process for Production of Antimicrobial Lipopeptide by Bacillus Amyloliquefaciens ES-2-4
|School||Nanjing Agricultural College|
|Keywords||lipopeptide solid-state fermentation in triangular flask koji plate fermentation|
Antimicrobial Peptides are a kind of bioactive little peptides which exist in biosphere widely. They are amphoteric molecules, produced by induction in organism always with positive charge. According to the difference of the resources, Antimicrobial Peptides can be divided into three kinds:animal origin, plant origin and microbial origin. Antimicrobial lipopeptides belong to the latter, produced by Bacillus sp. with the function of antibacterial, antifungal. antivirus and antimycoplasmal without normal cells’ damage. So they are safe for human and have an important application value in pharmaceutical and biomedicine. In addition, lipopeptides can be used as a biosurfactant that widely applied in the fields of petroleum industry, food, cosmetic and agriculture. Since the output of lipopeptide by liquid fermentation is low and the cost is pricy, industrial production of lipopeptide is quite difficult. Therefore, the paper focused on the solid state fermentation condition of lipopeptide. to prepare for the industry production. This paper contained three parts as follows:1. Firstly, the study was focused on using Bacillus amyloliquefaciens ES-2-4 as a parent strain to improve the solid-state fermentation condition in triangular flask. Bran, bean, medium account, initial ration of water and medium and inoculum size were chose as the key factors after single factor experiments. When the composition of medium was 70% bran and 30% bean, medium account in a triangular flask of 250 mL was 12.5g, mineral salt solution was 10%, initial ration of water and medium was 1:1, pH was 7, inoculum size was 10%, the temperature was 32℃, and fermentation time was 72 h, the conditions were most suitable for Bacillus amyloliquefaciens ES-2-4 to produce lipopeptide in triangular flask.2. Based on the optimal flask culture medium composition and fermentation conditions, koji plate fermentation for Bacillus amyloliquefaciens ES-2-4 was studied comprehensively. According to the results of single factor experiments, orthogonal experiments and response surface methodology were used to optimize the cultivation medium and the fermentation parameters, respectively. The optimal levels of the main factors were as follows:bran and bean 25%(w/w 3:1) as solid C/N resources, sucrose and ammonium sulfate as liquid C/N resources, mineral salt solution 10%. initial ration of water and medium 5:6, inoculum size7.1%, height of medium 2 cm, humidity 76.3%, temperature 33℃, fermentation time 48h. Under this condition, the lipopeptide yield could get to 6.98g/kg.3. Extraction and concentration of lipopeptide after koji plate fermentation were also studied. The best processing condition were as follows:the solid medium per 1 kg was first soaked in 6L alcohol, and then alcohol was concentrated by frozen concentration. The lipopeptide recovery could get to 90%.