纳豆菌固态产酶条件的优化

实验采用单因素试验优化纳豆菌固体发酵条件,通过蛋白凝块溶解时间法测定纳豆激酶活力,筛选出最佳培养条件。固态发酵通过扎孔、添加麦芽糖、蔗糖、葡萄糖和黄豆粉观察它们对纳豆菌产酶活力的影响。试验结果表明：固态发酵最佳条件为,浸泡10 h后,扎孔、添加4%麦芽糖、4%黄豆粉,121℃下蒸煮30分钟,接入纳豆菌2%,在37℃下培养24小时,4℃下后熟24小时。在此条件下培养,测得的纳豆激酶活力相当于尿激酶943.23 IU/g。与先前实验结果相比有了明显的提高：固态发酵由670.15 IU/g提高到943.23 IU/g。     

应用型纳豆菌培养条件的实验研究

对纳豆菌的培养条件进行了旋转摇床对比实验，碳源含量和培养时间对比实验，及不同碳源、不同时间下pH测定。结果显示，进行纳豆菌培养时以摇床恒温(32℃)液体培养，接种量3ml／100ml，碳源含量3g／100ml，液体培养18h进行接种到大豆发酵效果最佳。     

产溶栓酶纳豆菌的筛选及特性的研究

本文主要阐述了产溶栓酶纳豆菌的筛选、鉴定及最适固态发酵条件的初步研究。采用纤维蛋白平板法在经初筛获得 1 5株产溶栓酶菌株基础上 ,经复筛确定一产酶活力最高菌株 -B .N .1 0 ,并对该菌株部分生物学特性进行了研究 ,以B .N .1 0菌株进行固态发酵 ,其产酶最适温度、时间分别为 30℃和 2 4h     

传统纳豆中纳豆菌分离筛选的初步研究

近年来研究显示,纳豆具有特定的生理活性,作为功能性食品而受到人们的喜爱．从古田传统纳豆中分离到具有较高纳豆激酶活性的2个菌株,综合评价认为,编号为N142、N113的菌株具备作为优良生产菌种的特性．     

杏鲍菇产木聚糖酶液态发酵条件优化及酶学性质研究

利用杏鲍菇(pleurotus eyngii)降解啤酒糟液态发酵生产木聚糖酶,运用单因素与正交试验研究了啤酒糟、玉米芯与麸皮的不同组分、氮源、pH值、培养时间、转速、装瓶量等对产木聚糖酶活力的影响,并对其粗酶的酶学性质进行了研究。结果表明:碳源为3%啤酒糟、1%麸皮和1%玉米芯,氮源为0.5%酵母膏,pH值为5,培养时间为8d,转速为160 r/m in,接种量为10%,装瓶量为100 mL/250mL时,酶活力最高。其粗酶液的最适反应温度为50℃,最适反应pH为6,在pH4~6的范围内酶活性较稳定。但粗酶液的热稳定较差,当温度升至60℃时酶活力损失65%以上。     

响应面法优化纳豆激酶液体发酵条件的研究

为了提高纳豆激酶的酶活力,本研究借助Minitab软件,采用Plackett-Burman试验设计方法和响应面分析方法对保存的枯草芽孢杆菌纳豆亚种（Bacillus subtlis natto。简称纳豆菌）-NT207进行了液体发酵条件的优化。首先通过Plackett-Burman方法对11个相关影响因素的效应进行评价,并筛选出有显著正效应的木糖浓度和有显著负效应的接种量、K2HP04浓度等三个因素,然后利用响应面分析方法确定了上述三个因素的最佳工艺参数。实验结果表明,在优化后的发酵条件下,纳豆激酶的产量为1504．51U／mL,比优化前提高了130．3％。     

产β-葡聚糖酶的黑曲霉液态发酵优化的研究

应用β-葡聚糖刚果红营养平板法（GCN平板法），从土壤中筛选产β-葡聚糖酶的丝状真菌。温度、初始pH、平板的厚度都会影响产β-葡聚糖酶的霉菌分解底物所形成水解圈的大小及透明度。选用筛选的黑曲霉做摇瓶实验，通过正交试验，确定最佳培养基：大麦粉2g，麸皮2g，（NH4）2SO40．2g，豆饼粉1g；最佳培养条件是：温度为30℃，初始pH5．0，装样量50mL。     

球孢白僵菌产几丁质酶液态发酵研究

对球孢白僵菌Bb174产几丁质酶的液态发酵条件进行了试验研究。选择麸皮和蚕蛹粉作为培养基的基本成分。以不同比例的麸皮和蚕蛹粉进行试验，结果显示：麸皮和蚕蛹粉分别为5％，培养温度28℃，接种量10％，装液量10mL，起始pH6．0，添加0．3％NaNO3对产几丁质酶有利，最高酶活力为4664U／mL。     

碱性果胶酶生产菌株的产酶条件优化

对自行筛选的碱性果胶酶生产菌培养条件进行了优化。考查了碳源、氮源浓度和接种量对发酵产酶的影响。经单因素实验和正交试验研究,该菌株最佳产酶条件组合为：果胶添加量0．3％、NaNO。添加量0．6％、接种量6％。在此条件下,碱陛果胶酶酶活力达到6079U／mL。     

尖孢镰刀菌产碱性低温脂肪酶发酵条件的优化

通过实验对尖孢镰刀菌NC03摇床发酵产脂肪酶的培养基组成和培养条件进行了优化,得出最佳产酶培养基组成配方为：蛋白胨5 g/L,酵母膏6 g/L,NaH2PO43 g/L;橄榄油250 mL/L,吐温80 25 mL/L。最优发酵条件为250 mL的摇瓶装液量50 mL,培养温度30℃,发酵时间84 h。经过优化后发酵液脂肪酶酶活力最高可达到11.32 U/mL,较优化前提高了3.0倍。     

一种纳豆芽孢杆菌的分子鉴定及纳豆激酶基因克隆

以实验室保藏的一株具有溶解血栓能力的芽孢杆菌（LJQ—NK）为出发菌株,通过形态学分析和16SrRNA比对,对菌株进行鉴定,更进一步利用聚合酶链式反应（PCR）克隆纳豆激酶DNA片段⑤16SrRNA比对。结果表明LJQ-NK与已报道的纳豆激酶16SrRNA序列有99．9％以上的同源性,初步认定为一株纳豆芽孢杆菌。克隆获得1120bp的DNA片段,应用DNAMAN软件分析,与已报道的纳豆激酶编码基因（G129164926）存在两个碱基的差异,编码纳豆激酶开放阅读框区域序列完全相同。     

用十二烷基硫酸钠消除枯草芽孢杆菌中的质粒

为获得宿主菌 ,研究了十二烷基硫酸钠 (SDS)对枯草芽孢杆菌中质粒的消除 .将过夜培养的枯草芽孢杆菌2 4/pMX45接种于含SDS( 0— 0 .0 0 8% )的LB培养基中 ,当SDS浓度 (w /v)大于或等于 0 .0 0 6 %时 ,菌体不能生长 .SDS的亚致死浓度为 0 .0 0 5 % ,其致死率达 99% .菌液稀释后涂LB平板 ,再随机挑选单菌落至抗性平板 ,检测由质粒编码的红霉素抗性是否丢失 .氯化铯 溴化乙锭梯度离心及质粒DNA的电泳图像证实了 2 4/pMX45的衍生菌株A7中的质粒已完全被消除 .A7延迟期较短 ,并且细胞浓度高于 2 4/pMX45 .用SDS处理 8h后 ,2 4/pMX45的遗传标记开始丢失 ,消除率持续增高至 2 2h ,随之消除质粒的菌体量保持恒定 .在未经SDS处理的对照实验中 ,相同条件下培养 2 4h及 48h后 ,没有发现质粒自然丢失的现象 ,因此SDS能消除枯草芽孢杆菌中的质粒     

Purification and characterization of a novel antifungal protein from Bacillus subtilis strain B29"

An antifungal protein was isolated from a culture of Bacillus subtilis strain B29. The isolation procedure comprised ion exchange chromatography on diethylaminoethyl (DEAE)-52 cellulose and gel filtration chromatography on Bio-Gel P-100.The protein was absorbed on DEAE-cellulose and Bio-Gel P-100. The purified antifungal fraction was designated as B29I, with a molecular mass of 42.3 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), pl value 5.69 by isoelectric focusing (IEF)-PAGE, and 97.81% purity by high performance liquid chromatography (HPLC). B29I exhibited in-hibitory activity on mycelial growth in Fusarium oxysporum, Rhizoctonia solani, Fusarium moniliforme, and Sclerotinia scle-rotiorum. The 50% inhibitory concentrations (IC50) of its antifungal activity toward Fusarium oxysporum and Rhizoctonia solani were 45 and 112 μmol/L, respectively. B291 also demonstrated an inhibitory effect on conidial spore germination of Fusarium oxysporum and suppression of germ-tube elongation, and induced distortion, tumescence, and rupture of a portion of the germi-nated spores.     

枯草芽孢杆菌菌株Bv10对大叶黄杨白粉病的防治效果

大叶黄杨白粉病近年来在河南省南阳市发生严重.对大叶黄杨植株春梢进行叶面喷施生防菌枯草芽孢杆菌Bv10菌株的发酵液,进行白粉病小区生防试验,结果显示:喷施7 d后,枯草芽孢杆菌Bv10发酵液对大叶黄杨白粉病的防治效果达70.37%,低于对照药剂粉锈宁;喷施14 d后,该生防菌的防效降低至25.26%.     

Adsorption of Cu^2 ＋, Zn^2＋ and Cd^2＋ on Bacillus subtilis

A process of biosorption of Cu2 , Zn2 and Cd2 on Bacillus subtilis was investigated. The experiments show that the process of biosorption is quite fast. The maximum adsorption was reached after 5 min and hardly changed with time. The experimental data was analyzed using four sorption kinetic models: the pseudo-first-order, the Ritchie second-order, the modified second-order and the Elovich equations, which helped to determine the best-fit equation for the sorption of metal ions onto biomass. The results show that both the Ritchie second-order and modified second-order equations can fit the experimental data. The Langmuir model is able to accurately describe adsorption of Cu2 and Zn2 on B. subtilis. The experimental data points of adsorption Cd2 and Zn2 on B. subtilis are described by Freundlich isotherms model.     

Optimization of cultural conditions for thermostable β-1,3-1,4-glucanase production by Bacillus subtilis ZJF-1A5

The optimization of cultural conditions for β-glucanase production by Bacillus subtilis ZJF-1A5 was investigated in flask trials. Temperature had gre at effect on β-glucanase production which maximized at optimal temperature of 37 ℃ and decreased significantly when temperature was over 37℃.Charge quantity af fected β-glucanase production significantly. Adding oxygen vector N-dodec ane or acetic ether benefited β-glucanase production, but it depended on the conc entrat ion and charge quantity. The results of fractional factorial design showed that age and size of inoculum and shaking speed were the key factors affecting β -gluc anase production and the cultivation time span to reach the highest β-gluc anase activity. The optimal cultural conditions for β-glucanase production obtai ned wi th CCD were as follows: inoculum age and size (16 h, 3.82%(v/v)), shaking speed 2 10 r/min, charge quantity of 30 mL in 250 mL flask and initial pH 7.0, cultured at 37℃ for 50 h. Repeated experimental results accorded with those predicted b y a second-order polynomial model. The amount of β-glucanase, α-amylase and neut ral protease produced by B subtilis ZJF-1A5 was associated partially with c ell g rowth. Those three enzymes' activities increased following the cell growth and i ncreased significantly when cells entered the stationary phase.     

Keratinase production and keratin degradation by a mutant strain of Bacillus subtilis

A new feather-degrading bacterium was isolated from a local feather waste site and identified as Bacillus subtilis based on morphological, physiochemical, and phylogenetic characteristics. Screening for mutants with elevated keratinolytic activity using N-methyl-N'-nitro-N-nitrosoguanidine mutagenesis resulted in a mutant strain KD-N2 producing keratinolytic activity about 2.5 times that of the wild-type strain. The mutant strain produced inducible keratinase in different substrates of feathers, hair, wool and silk under submerged cultivation. Scanning electron microscopy studies showed the degradation of feathers, hair and silk by the keratinase. The optimal conditions for keratinase production include initial pH of 7.5, inoculum size of 2% (v/v), age of inoculum of 16 h, and cultivation at 23 ℃. The maximum keratinolytic activity of KD-N2 was achieved after 30 h. Essential amino acids like threonine, valine, methionine as well as ammonia were produced when feathers were used as substrates. Strain KD-N2,therefore, shows great promise of finding potential applications in keratin hydrolysis and keratinase production.     

Purification and characterization of keratinase from a new Bacillus subtilis strain

The aim of this study was to purify and characterize a keratinase produced by a new isolated Bacillus subtilis KD-N2strain. The keratinase produced by the isolate was purified using ammonium sulphate precipitation, Sephadex G-75 and DEAE (diethylaminoethyl)-Sepharose chromatographic techniques. The purified enzyme was shown to have a molecular mass of 30.5kDa, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The optimum pH at 50℃ was 8.5 and the optimum temperature at pH 8.5 was 55℃. The keratinase was partially inactivated by some metal ions, organic solvents and serine protease inhibitor phenylmethanesulfonyl fluoride (PMSF). Sodium dodecyl sulfate (SDS) and ethylene diamine tetraacetic acid (EDTA) had positive effect on the keratinase activity. Reducing agents including dithiothreitol (DTT),mercaptoethanol, L-cysteine, sodium sulphite, as well as chemicals of SDS, ammonium sulfamate and dimethylsulfoxide (DMSO)stimulated the enzyme activity upon a feather meal substrate. Besides feather keratin, the enzyme is active upon the soluble proteins ovalbumin, bovine serum albumin (BSA), casein and insoluble ones as sheep wool and human hair. Calf hair, silk and collagen could not be hydrolyzed by the keratinase.     

Optimization of Riboflavin Production from ccpA Mutant Bacillus subtilis 24A1/pMX45

Bacillus subtilisis widely used for the productionof vitamins and other products including industrial en-zymes such as amylases, proteases and lipase. Butsyntheses of those products are repressed by the pres-ence of glucose, which is viewed as carbon catabolicrepression (CCR). InBacillus subtilis,CCR is main-ly mediated by the global regulator protein CcpAwhich was encoded byccpAgene[1]. It is therefore of crucial importance to relievingCCR ofBacillus subtilisfor industrial production. I…