利用生防真菌孢子粉生产中的废米生产饲用β-葡聚糖酶和木聚糖酶的黑曲霉固相发酵条件优化

Optimized solid-state fermentation of Aspergillus niger for production of feed-purpose β-glucanase and xylanase using rice residues from mass production of fungal biocontrol agents

为资源化利用生防真菌高纯度孢子粉在生产过程中产生的大量废米,将废米粉按比例混入麦麸作为基础物料,接种黑曲霉固相发酵生产饲用β-葡聚糖酶和木聚糖酶,得到麸米比7∶3的最佳配料;纯麦麸发酵3d后2种酶的产量为353.8、343.8U·g-1干曲.在对发酵时间、配料含水量、补充氮源、微量元素、拌料水的表面活性剂浓度及pH值等单因素分别优化试验的基础上,选取对产酶影响较大的金属离子类型、硫酸铵添加量、拌料水的吐温80浓度和pH进行4因素3水平9处理的正交试验,获得的最优组合为麸米比7∶3的配料中加入0.1%KH2PO4和1%(NH4)2SO4,拌料水为0.3%吐温80和pH5.经3次重复试验验证,用最优组合配料接种后在25℃下发酵54h,β-葡聚糖酶和木聚糖酶产量(±SD)分别达到(716.1±9.2)、(453.0±62.5)U?g-1干曲,远高于优化前相同麸米比配料的酶产量.这一结果为建立环境友好型真菌杀虫剂生产线提供了有用的配套技术,使饲用酶制剂生产节约麦麸30%.

Rice residues from mass production of fungal hiocontrol agents against insect pests were explored for use in the production of feed-purpose β-glucanase and xylanase, which is usually achieved by solid-state fermentation （SSF） of Aspergillus niger with wheat bran. A mixture of 70% wheat bran and 30% milled rice was selected as an acceptable substrate in the trials of 10 ratios of the two components and yielded 353.8 and 343.8 U·g^-1 substrate for β-glucanase and xylanase after 3-day SSF, respectively. Factors found to significantly affect the enzyme yields in the mixture during SSF were the type of metal ion as trace nutrition, the content of （NH4）2SO4 as supplementary nitrogen, and the concentration of Tween-80 and pH in the water used to stir the mixture for SSF. Combinations of the selected factors were further optimized for enzyme production in triple four-factor, three-level orthogonal experiments. The optimized combination of the rice-bran mixture consisted of 0. 1 % KH2PO4 , 1.0% （NH4）2SO4 and the water containing 0.3% Tween-80 with pH 5. When this optimized mixture was used as substrate in three repeated trials of 54-h SSF at 25℃ for confirmation, the mean （±SD） yields of β-glucanase and xylanase reached （716.1±9.2） and （453.0±62.5） U·g^-1, which were greatly enhanced compared to those before optimization. The results highlight an alternative approach to utilizing the rice residues for the enzyme production at the reduced cost of 30% wheat bran and would facilitate to establish a ricebased, environment-friendly technology for bulk production of fungal biocontrol agents.