Studies On The Breeding Of Xylanase Overproduction Strain And The Conditions Of Xylanase Production From Bacillus Sp.

In this paper, the extracting methods and conditions of xylan from corncob, the breeding of xylanase overproduction strain from Bacillus sp., the conditions of the enzyme production, and the characteristics of the xylanase were investigated.Based on the study of xylanase, the extracting methods and conditions of xylan from corncob were improved. The virtue of the cooking method was integrated with the alkali extracting method. The main process of extracting consists of cooking, alkaline soak, and depositing. The conditions of pretreatment, alkali extracting and depositing were researched individually. After being smashed and screened, the corncob was cooked at 120 for 60 minutes(100g corncob in 600ml water) and extracted by ethanol(100g corncob in 300ml ethanol).Then the corncob slurry was soaked in 10%NaOH at 100 for lhour(100g corncob in 1500ml 10%NaOH). At last hemicellulose A was deposited at pH4.5 and hemicellulose B was deposited in ethanol (100ml liquor obtained by centrifugal in 300ml 95% ethanol). The content of hemicellulose in the product is 72.6% and the ratio of extracting is 40.7%Aw-26 (Bacillus sp.) was chosen as an original strain for screening high-producing strain of xylanase. The strain was treated with UV and EMS. The dosage of UV was irradiated at 30cm for 2.5min with 15w ultraviolet lamp. The dosage of EMS was soaked in 0.5% EMS for 45min. After screening, a mutant strain named WAL-105 was selected. The xylanase activity of WAL-105 is as high as 532.43IU/ml and it is 3.65 times of xylanase activity of Aw-26.The culture conditions that can influence the growth and enzyme production were investigated. When WAL-105 was cultured in complex medium containing wheat bran as sole carbon source, urea as nitrogen source, the enzyme production was optimized. Tween 80 and yeast extract were helpful. It hasn't an obvious effect on xylanase activity when corncob, xylan and cellulose were used as the complex carbon source. The culture conditions for WAL-105 optimized by the orthogonal test were 7% wheat bran, 0.7% urea, 0.2%yeast extract, 0.2%Tween 80, 0.5%NaCl, 0.1%KH2PO4, 0.02% MgSO4.7H2O. Variance analysis to the results of the orthogonal test showed that wheat bran had the very obvious effect onxylanase, Tween 80 and urea had the obvious effect on xylanase and MgSO4.7H2O had the effect on xylanase activity. The initial pH is 7.0. The optimized stirring rate is 1 SOrpm and the optimized temperature is 30. In the best-case conditions, xylanase activity could reach 764.62IU/ml. Based on the study of fermentation in erlenmeyer flask, the fermentation in 5L fermentor was experimented. In the fermentation process in fermentor, the stability of Xylanase production by WAL-105 was well and the highest xylanase activity was 568.27IU/ml.In the fermenting process of erlenmeyer flask, Xylanase activity is 764.62IU/ml, CMCase activity is 0.36IU/ml. The production of xylanase is enhanced by Zn2+, A13+, Ca2+, Li+, Mg2+and inhibited by Ag+ Mn2+> Fe3+, Cu2+. Xylanase activity is induced by xylan and CMC-Na. It is hindered by glucose and galactose. The effect on xylanase activity is not obvious when xylan and CMC-Na are used together as the complex carbon source and the xylanase activity is lower than that using xylan and cellulose individually. The optimum temperature of xylanase is 52 and the optimum pH is 7.0. It is stable in a broad pH range and shows well stability below 50. Its xylanase activity is increased dramatically by Ca Mg and strongly inhibited by Fe3+, Ag+, Pb2+.