土壤中产木聚糖酶菌株的筛选及发酵条件优化

Screening and fermentation conditions optimization of xylanase-producing strain from soil

摘要:

[背景] 木聚糖广泛存在于木质纤维类生物质中，是世界上含量最丰富的半纤维素，利用产酶微生物对木质纤维类生物质进行发酵处理是木质纤维类生物质资源化和能源化的有效手段。[目的] 通过产木聚糖酶菌株的筛选鉴定、酶学特性分析和发酵条件优化，获得开发多纤维农林废弃物生产新型多元化饲料添加剂的材料。[方法] 利用青藏高原土壤筛选产木聚糖酶菌株，通过形态学观察和rDNA ITS区域序列分析鉴定菌株XC70种属，对其所产酶的酶学特性及该菌的生长规律和产酶规律进行分析，并利用单因素法和正交试验法优化其发酵条件。[结果] 菌株XC70经形态学和分子生物学方法鉴定为草酸青霉（Penicillium oxalicum）。菌株XC70所产木聚糖酶的最适反应条件为：pH 5.0，70℃，温度低于50℃时稳定性较好，具备一定的耐酸性，Na⁺和K⁺对木聚糖酶活力具有促进作用（P<0.05），在发酵54 h后菌体量和上清液酶活力大小均达到高峰。经过单因素法和正交试验法优化后确定了该菌的最优发酵条件为：蛋白胨7 g/L，玉米秸秆50 g/L，KCl 4 g/L，培养基初始pH 4.0，28℃，摇床转速200 r/min，接种量2%。在此发酵条件下，木聚糖酶活力可达到1 489.33 U/mL，与优化前相比提高了3倍多。[结论] 从青藏高原土壤中筛选获得的菌株XC70具有一定的产木聚糖酶能力，其所产生的酸性木聚糖酶可用于降解多纤维物质开发新型饲料添加剂，具有一定的应用潜力和开发价值。

Abstract:

[Background] Xylan which widely exists in lignocellulosic biomass is the most abundant hemicellulose in the world. It is an effective way for the resource and energy utilization of lignocellulosic biomass to utilize enzyme producing microorganisms to ferment lignocellulosic biomass. [Objective] The aim of this experiment was to obtain materials for producing multiple feed additives from multi fiber agricultural and forestry wastes by screening and identifying xylanase producing strains, analyzing enzymatic characteristics and optimizing fermentation conditions. [Methods] The xylanase producing strains were screened from the soil of Qinghai-Tibet Plateau. The strain XC70 were identified by morphological observation and rDNA ITS sequence analysis. The enzymatic characteristics, growth and enzyme production attributes of the strain were analyzed, and the fermentation conditions were optimized by single factor method and orthogonal test method. [Results] The strain XC70 was identified as Penicillium oxalicum by morphological and molecular biological methods. The results showed that the optimal reaction conditions of xylanase produced by XC70 were pH 5.0, 70℃, the xylanase had good stability when the temperature was lower than 50℃, and it showed some acid resistance. Na⁺ and K⁺ promoted the xylanase activity (P<0.05). After 54 h of fermentation, the biomass and enzyme activity of supernatant reached the peak. Through single factor method and orthogonal test method, the optimal fermentation conditions were determined as follows:peptone 7 g/L, corn straw 50 g/L, KCl 4 g/L, initial pH 4.0, 28℃, shaking speed 200 r/min, inoculum size 2%. Under these conditions, the xylanase activity reached 1 489.33 U/mL, which was three times higher than that before optimization. [Conclusion] The strain XC70 screened from the soil of Qinghai-Tibet Plateau has certain acidic xylanase-producing capacity, which can be used to degrade multi fiber materials and develop new feed additives, and this has certain application potential and development value.