Agrobacterium tumefaciens OAH-01产脲酶发酵培养基优化

Optimization of fermentation culture by urease-producing strain Agrobacterium tumefaciens OAH-01

氨基甲酸乙酯(EC)是大多数发酵制品中的潜在致癌物,而尿素是EC的最主要前体物质之一,因此需要采取相关措施控制发酵制品中尿素的含量。向酒体中添加脲酶具有安全、高效和处理条件温和等优点,是FDA推荐的降低EC含量的优先方法。微生物是脲酶的主要来源,可利用其实现脲酶的大规模生产。本文以产脲酶根癌农杆菌Agrobacterium tumefaciens OAH-01为研究对象,考察了该菌株的生长曲线和产酶曲线,证明该菌株产脲酶属生长关联型,且能在较短时间内达到最大产酶量,此外还研究了发酵培养基组成对该菌发酵产酶的影响,通过单因素及正交试验优化确定了最佳发酵培养基组成(g/L):蛋白胨20,酵母粉5,葡萄糖5,FeCl_30.54,Na_2HPO_40.5,KH_2PO_40.5,NiSO_40.1,起始pH 7.0。在最适条件下发酵16 h,菌悬液中脲酶酶活可达1.077U/m L,为优化前的2.4倍。超声破碎后细胞上清中的脲酶活性为0.419 U/m L,为优化前的4.4倍。

Urea as the main precursor of ethyl carbamate（ EC） is a potential carcinogenic component in most of the fermented foods. The methods for elimination of urea must be taken to control the amount of urea. The preferred method for reducing EC content recommended by FDA is adding urease to the wine,because of its advantages such as efficiency,high safety and mild treatment conditions. Microbes are the main source of urease which can be carried out mass production. In this paper,an urease-producing strain Agrobacterium tumefaciens OAH-01 was investigated. Its growth curve and urease-producing curve indicated that the production of enzyme was growth associated model and it could reach the maximum yield in a relatively short time. The effects of fermentation medium composition on urease production by OAH-01 were also investigated. The fermentation medium composition was optimized based on single factor experiment and orthogonal experiment. The optimal medium and conditions were as follows（ g/L） ： peptone 20,yeast extract 5,glucose5,FeCl_30.54,Na_2HPO_40.5,KH_2PO_40.5,NiSO_40.1,pH 7. 0 and cultivation 16 h. Under the optimized conditions,the urease activity of cell suspension was 1. 08 U/m L,it was 2. 4 times than that before optimization. The activity of supernatant after ultrasonic decomposition was 0. 419 U/m L,it was 4. 4 times than that before optimization.