降解黄曲霉毒素B_1菌株发酵工艺的研究

OPTIMIZATION OF FERMENTATION PROCESS OF KLEBSIELLA SP. XY1 FOR DEGRADING AFLATOXIN B_1

对黄曲霉毒素B1(AFB1)降解菌株克雷伯氏菌(Klebsiella sp.)XY1降解AFB1进行发酵工艺的优化,主要包括培养基成分和培养条件的优化。对培养基成分的优化主要包括:碳源,如葡萄糖、可溶性淀粉、甘露醇、玉米芯粉、蔗糖、α-乳糖、D-果糖;氮源,如牛肉膏、硝酸铵、硝酸钠、蛋白胨、尿素、胰蛋白胨、酵母浸粉;金属离子,如CaCl_2、MgCl_2、FeCl_2、ZnSO_4、CuSO_4、MnCl_2;培养条件主要包括温度、起始pH、接种量、装液量等。选出最佳碳源、氮源和金属离子,设计新的培养基组成,根据单因素试验结果,运用Box-Behnken设计原理,设计响应面试验,对培养基配方进行最优化分析。结果表明:模型(P=0.001 4)在1%水平上具有极显著性,失拟项不显著(P=0.065 2>0.05),其R2=0.943 1,R2adj=0.870 1,说明该模型拟合程度良好,可以模拟94.31%的AFB1降解率变化,其中玉米芯粉对AFB1降解率的影响最大。确定最优发酵培养基配方:玉米芯粉添加量1.06%;胰蛋白胨添加量1.32%;Ca Cl2添加量0.036 mol/L。最佳发酵条件:起始p H值为7.0、温度34℃、接种量6%(V/V)、装液量25 m L/250 m L,降解时间72 h。经发酵工艺优化,Klebsiella sp.XY1对AFB1降解率由71.91%升高到95.13%。

In this study,fermentation process of aflatoxin B1(AFB1)degradation strain Klebsiella sp.XY1 preserved in our lab was optimized.The optimization of fermentation process consisted the fermentation medium and conditions including carbon source(glucose,soluble starch,corncob,α-Lactose,D-Fructose,sucrose,mannitol),nitrogen source(yeast extract,beef extract,peptone,tryptone,urea,ammonium nitrate,sodium nitrate)and metal ion(CaCl2,MgCl2,FeCl2,ZnSO4,CuSO4,MnCl2).Additionally,the fermentation conditions including initial pH,temperature,inoculation and medium volume were also optimized.Based on single-factor experiment,the best carbon source,nitrogen source and metal ion were confirmed.According to the single factor test results,response surface test using the principle of Box-Behnken was designed to optimize the degradation conditions.The results of variance analysis showed that the response surface model(P=0.001 4)was significant at the 1%level and lack of fit was not significant(P=0.065 2>0.05).R2=0.943 1,R2Adj=0.870 1,indicating that the model fitting degree was suitable to simulate the change of AFB1 degradation rate of 94.31%,wherein the effect of corncob on the degradation rate of AFB1 was most significant.The results indicated that the optimal medium contained 1.06%corncob,1.32%tryptone and 0.036 mol/L calcium chloride.The optimum fermentation conditions were as follows:initial pH 7.0,temperature 34℃,inoculation amount 6%(V/V),medium volume 25 mL in a 250 mL erlenmeyer flask.After optimization of the fermentation process,the degradation rate of AFB1 increased from 71.91%to 95.13%.