N+注入选育产蛋白酶益生菌及其生物学效应研究

试验研究了N+注入选育产蛋白酶益生菌及其生物学效应.经N+反复注入诱变筛选,突变高产菌株的酶活由出发菌株的75.8IU.g-1提高到631.6IU.g-1;利用中心组合设计原理摸索出了突变菌株最佳产酶条件即麸皮24.83%,豆粕23.68%,水50%,硫酸铵1.49%,发酵温度30℃,发酵时间66h,培养基pH 5.5.生物学效应研究发现真空处理对活细胞有明显伤害;孢子存活率先是随着注入剂量加大而迅速降低,当注入剂量加大到50×2.6×1013 ions/cm2时存活率出现平缓回升,注入剂量超过100×2.6×1013ions/cm2时存活率又再次开始下降;注入剂量越大,孢子突变率越高,中等注入剂量30×2.6×1013N+/cm3～80×2.6×1013N+/cm2可引起较高的正突变率;ESR谱研究结果表明,N+注入前和注入后都有自由基ESR波谱单一峰,随着注入剂量的增大,孢子内的自由基ESR波谱的强度也逐渐增大;SEM观察发现,未经N+注入的孢子较为完整、饱满且表面光滑,而经注入后的孢子表面粗糙,可见明显的刻蚀损伤和破壁现象,并且注入剂量越大,伤痕也越深宽.试验结果提示N+注入是一种有效的动物益生菌选育方法.

Studies on Breeding of Probiotic Aspergillus niger and its and Biological Effects by N+ Implantations

Breading of probiotic Aspergillus niger and its biological effects by N~+ implantations were studied. The activities of protease of mutated strain Aspergillus niger AN02 was raised from 75.8 IU.g~(-1) to 631.6 IU.g~(-1) IU.g~(-1); the optimum factors of mutated strain Aspergillus niger AN0 was tested by the theory of center combination; That is medium bran 24.83 %, bean cake 23.68 %,water 50 %, (NH_4)_2SO_4 1.49 %, fermentation temperature, 30 ℃,time period, 66 h, pH, 5.5. The biological effects showed: the treatment of vacuum harm live cell; The survival rate of spores was decreased with increasing of implantation dose and increased gradually when the dose of implantation was 50×2.6×10~(13) ions/cm~2; but that of spores decreased when the implantation ion dose was 100×2.6×10~(13)　ions/cm~2; There was high positive mutation rate when 30×2.6×10~(13) N~+/cm~2～80×2.6×10~(13) N~+/cm~2.The lager the implantation dose was, the higher the spore mutation rate was. The ESR results showed that free radicals have existed in the control and increased with added implantation dose. The SEMindicated that the control were integrity ,while the surface of spores was coarseness , moreover the damage of ionic etching and phenomenon of dilapidating cell wall could be seen , When dose increased continually , the damage to cells became more and more serious. Thus a conclusion may be drawn that it is a effective way of breeding probiotics by N+ implantation.