高效脂肪酶产生真菌的筛选、鉴定及产酶条件优化

Screening and identification of efficiently-producing Fungi from lipase and optimization of its lipase-producing condition

研究旨在筛选高产脂肪酶活性的菌株,并鉴定其菌属及确定其最佳产酶条件。采集富含油脂的土壤,通过溴甲酚紫平板法进行菌株分离筛选,并采用橄榄油乳化法测定脂肪酶活力。利用真菌ITS序列通用引物进行PCR扩增,将测序结果Blast比对,并建系统发育树。采用正交试验设计优化产酶条件。通过筛选得到1株产脂肪酶的真菌菌株GW-1,经鉴定为黑曲霉Aspergillus niger。经单因素试验和正交试验得到该菌的最佳产酶条件为:葡萄糖0.5%、黄豆粉2.0%、橄榄油1.0%、MgSO4.7H2O 0.1%、pH值9.0、接种量8%、装液量50 ml/250 ml、发酵4 d。在此发酵条件下,其产酶活力可达39.82 U/(ml.min)。试验结果显示,访菌株在产脂肪酶能力上具有显著优越性,且菌株GW-1产酶活力较优化前高出19.96 U/(ml.min),提高了100.50%。

This experiment was conducted to screen high lipase activity strains, identify the hacterium and define the optimum fermentation conditions. The strain was screened from collected greasy soil through bromocresol purple plate method and the lipase activity was determined with olive oil emulsification method. PCR was performed using the Fungi on ITS sequence universal primers. The sequencing resuhs was Blasted and the development tree was installed. The optimum enzyme production conditions were designed using orthogonal test. A Fungi strain GW-1 producing lipase was screened, which was identified as Aspergillus niger. The optimum fermenta- tion conditions for lipase production were got from the single factor experiments and the orthogohal experiments by strain GW-1 were 0.5% glucose, 2.0% soybean powder, 1.0% olive oil, 0.1% MgSOg＇7H2O, initial pH 9.0, 8% inoculums volume, and bottle filling capacity 50 ml/250 ml, fermentation time 4 d. Under this optimum condition, the lipase activity of strain GW-1 reached 39.82 U/（ml· rain）. These results indicated that the enzyme activity produced by strain GW-1 was increased by 19.96 U/（ml·min） through optimization than before, up 100.50%.