碱性普鲁兰酶产生菌选育和发酵条件的研究

Studies on screening and breeding of alkaline pullulanase producing bacterium and regulation of nutrition

从儿童食品中分离筛选到 1株产碱性普鲁兰酶活性较高的菌株 ,编号为 SX- 1 2 ,初步鉴定为芽孢杆菌 Bacillussp.。研究了 SX- 1 2原生质体制备与再生最佳条件。其原生质体经紫外线诱变处理 ,选育出产碱性普鲁兰酶的高产菌株 SX- 1 2 C67,酶活由出发菌株的 2 .42 U/m L提高到 6.87U/m L ,提高了约 1 .8倍。在此基础上对产酶条件进行了优化 ,优化后的最佳发酵培养基为 :可溶性淀粉 3% ,蛋白胨 1 .0 % ,酵母膏 0 .5% ,K2 HPO40 .2 % ,Mg SO4.7H2 O0 .0 5% ,Mn Cl2 0 .0 0 0 1 % ,最适 p H9.5,最适温度 40℃。初步研究了酶的部分性质 ,酶反应的最适p H、温度分别为 1 0 .0～ 1 0 .5和 55℃。在 55℃反应条件下 ,酶在 p H6.0～ 1 1 .0的范围内都具有一定的活性。 Ca2 +、Mn2 +、Mg2 +等离子是酶的激活剂 ,Zn2 +、Hg2

Bacillus sp. SX 12,a strain producing alkaline pullulanase,was isolated from baby food.The optimal condition for preparating of the protoplast and its regenerating were studied.Protoplast of Bacillus sp. SX 12 were induced by ultraviolet ray.One High producing strain (SX 12C 67 ) were obtained from process of screening.The activity of alkaline pullulanase in strain SX 12C 67 increased from 2.42 U/mL in strain SX 12 to 6.87 U/mL,About 1.8 times to which in strain SX 12.The fermentation medium and culture condition of the strain were investigated,the optimum medium for fermentation is that Soluble Starch 3%,pepton 1.0%,Yeast extract 0.5%,K 2HPO 4 0.2%,MgSO 4.7H 2O 0.05%,MnCl 2 0.0001%.The optimum temperature is 40℃,The optimum pH is 9.5,The optimum temperature for enzyme is 55℃,The optimum pH is 10.0-10.5.It shows a high stability during pH 6.0-11.0 at 55℃.The activity of alkaline pullulanase could be promoted by Ca 2+ ?Mn 2+ ?Mg 2+ and inhibited by Zn 2+ ?Hg 2+ .