一株中温淀粉酶菌株的分离鉴定及其产酶条件分析

Isolation,Identification and Analysis on Enzyme Producing Condition of a Strain of Medium-Temperature Amylase

利用固体淀粉筛选培养基,从安阳市郊区面粉厂附近的土壤里分离筛选出1株产淀粉酶的菌株,编号为MF-3-2。经过菌株形态、革兰氏染色、16S rDNA鉴定及系统进化树分析,初步确定其为枯草芽孢杆菌(Bacillus subtilis)。摇瓶培养后对其酶学性质研究发现,该菌株淀粉酶的最适温度为65℃,最适pH值为6.0,在pH值4.8~6.0范围内仍能残余70%以上的酶活力。该菌株的最适生长温度为40℃,最适生长pH值为6.5。产酶条件优化结果表明:最适碳源为马铃薯淀粉,最适氮源为豆粕粉,最适碳氮比为1:15,发酵温度30℃,发酵pH值6.0,装液量10%,种龄10 h,接种量5%,转速200 r/min,48 h达到产酶高峰。通过发酵产酶条件优化,其淀粉酶活性达到86.8 U/mL,是优化前的35倍。另外,在酸性条件下还具有较好的活性。因此,该菌株的淀粉酶具有潜在的工业应用前景。

One strain(MF-3-2)producing amylase was isolated on starch screening medium from the soil around the flour factory,Anyang City.By observing the form of the morphology,gram straining and amplification of its 16S rDNA sequence,the strain MF-3-2 was identified as Bacillus subtilis.After shaking culture,the enzymatic properties were studied by using the crude enzyme.The amylase exhibited maximal activity at pH 6.0 and 65℃,and was pH adaptability(>70%activity at pH 4.8~6.0).Optimal growth of the strain occurred at 40℃and pH 6.5.The optimization of fermentation conditions were used in this study.The results showed that the optimal medium composing of potato starch,soya bean meal,C/N ratio(1:15),initial cultural pH 6.0,culture volume 10%,inoculum age 10 h,inoculum size 5%,revolving speed 200 r/min,30℃fermentation 48 h.On the optimal culture condition,the maximal enzyme production was 86.8 U/mL,which was 35 folds of that before optimization.Medium-temperature amylase produced by this bacterium showed high acid adaptability.So the medium-temperature amylase of the strain has great development potential and application in industrialization.