弹性蛋白酶产生菌的筛选及其发酵条件的初步研究

Screening of elastase-producing strains and primariry atudies on fermentation conditions.

从肉联厂和屠宰场附近的土壤和污水中采集到的近100个样品中,经富集培养分离到123株能产生弹性蛋白酶的菌株,反复初筛、复筛后,分离到一株产胞外弹性蛋白酶活较高且稳定的细菌,经初步鉴定属芽孢杆菌属(Bacillussp.EL3)。同时研究了其产酶最适条件,产酶最适碳源为葡萄糖,最适氮源为豆饼粉。并对碳源、氮源和生长因子进行了正交实验,研究发现碳源对产酶的影响最大,其次为氮源。当葡萄糖的用量为6%、豆饼粉为5%、玉米提取液为0.2%时,其具有较高的产酶水平。最适发酵初始pH为7.5,250mL最适摇瓶装液量为20mL。在该发酵培养基中培养,35h时达最高的产酶水平。同时还对离心后的粗酶液进行温度和pH试验,发现该酶最适作用温度和pH分别为50℃和7.4,该酶在40℃以下保温30min酶活仍较高,在60℃以上酶活全部丧失。在pH7.4～9.0的范围内比较稳定,而在pH7.0以下时,酶活下降很快。金属离子Cu2+、Zn2+、Al3+能抑制弹性蛋白酶的活性,而Fe3+、Li2+能激活弹性蛋白酶活性,螯合剂EDTA则严重抑制弹性蛋白酶活性。

Strains capable of secreting extracellular elastase were isolated from the soil and wastewater samples that were near the processing factory of pig meat Of them 123 strains produced elastase Strain NoEL3 with the highestyield elastase was identified as Bacillus sp In addition, the experimental results revealed that the most suitable carbon and nitrogen sources for elastase production were glucose 6% and soybean flour 5%, respectively The ANOVA results demonstrated that when glucose was 6%, soybean flour 5% and corn steep flour 02%, it had the highest elastase production When the initial pH of the medium was 75, and 20 mL broth in 250 mL flask it was optimal for elastase production The culture maintained maximum elastase activity at 35 ℃ for 35 hours under the optimized conditions The elastiolytic activity was the highest at pH74 and 50 ℃ This enzyme was stable over the range of pH 7490 and below 40 ℃, and its enzyme activity was inhibited by Cu2+, Zn2+, Al3+ and EDTA Fe3+ and Li2+ were beneficial to enzyme activity