降解秸秆的纤维素酶产生菌的筛选及产酶条件研究

从土壤、霉变的农产品和实验室保存的真菌中筛选到15株产纤维素酶的菌株,其中绿色木霉T206产酶能力最强。利用液体发酵,研究了碳源、氮源、接种量、起始pH值、培养时间等对菌株产酶的影响,以及该菌株所产纤维素酶的种类。在最适条件下菌株培养96h后,羧甲基纤维素酶活(CMCA)最高达到7654.33U,是优化前酶活的1.7倍,滤纸酶活(FPA)达到1675.12U。     

普鲁兰酶产生菌的筛选鉴定及其发酵条件优化与酶学性质研究

从淀粉厂附近土壤中筛选得到5株产普鲁兰酶的菌株,与实验室已有保藏菌株做活性对比,结果筛选得到的S1117产酶活性较高,根据其形态特征及16S rRNA序列分析,初步鉴定为芽孢杆菌。对其产酶条件进行了优化,确定了该菌株的最适发酵培养条件,其发酵54 h时酶活达到最大2.5 U/mL。初步研究了其酶学性质,其所产普鲁兰酶反应最适温度与pH分别为40℃,5.2;在60℃仍保持有80%以上酶活。     

枯草芽孢杆菌产烟酰胺酶的发酵工艺条件优化

对已筛选出的菌株枯草芽孢杆菌N-17进行发酵条件优化,利用单因素实验方法,对烟酰胺酶的发酵培养基的组分及发酵条件进行优化,确定了各因素的最适值.结果表明:该菌株在温度28℃,起始pH值7.0,己内酰胺浓度为0.2％,葡萄糖1.2％,酵母粉0.6％,Mg2+0.02％时,产烟酰胺酶活性最大,酶活比之前提升了213.8％.     

一株产木聚糖酶菌株的筛选及发酵产酶条件优化

以木聚糖为唯一碳源,从含有大量腐烂枯枝树叶土壤中筛选到一株高产木聚糖酶菌株,经形态学分析和分子学鉴定,确定其为链霉属(Streptomyces)。通过单因素实验考察了碳源、氮源、初始pH值、发酵温度和发酵时间对酶活的影响;在单因素实验的基础上,利用Design-Expert软件对碳源、氮源和发酵时间进行响应面分析。单因素实验确定适宜发酵产酶条件为:复合碳源为玉米芯粉+蔗糖(1∶2)、氮源为硝酸铵、初始pH值4.0、发酵时间2.5d、发酵温度30℃,此时,所产木聚糖酶酶活为511U·mL~(-1);响应面分析确定最优发酵产酶条件为:复合碳源(玉米芯粉∶蔗糖=1∶2)添加量4.09%、氮源硝酸铵添加量1.16%、初始pH值4.0、发酵时间3.14d、发酵温度30℃,此时,所产木聚糖酶酶活达到641U·mL~(-1),较单因素实验提高了25.44%。     

磷脂酶A1高产菌株的筛选及其等离子体诱变

目的从富油土壤中筛选磷脂酶A1高产菌株,优化其发酵条件,并经等离子诱变提高其产酶能力。方法用筛选培养基和发酵培养基分别对富油土壤中磷脂酶A1产生菌进行初筛和复筛,经气相色谱鉴定产酶类型;对获得的高产菌株进行形态、生理生化特征和分子生物学鉴定;优化该菌株的发酵时间,温度,pH值及摇床转速;从该菌株出发进行等离子体诱变,绘制致死率曲线,进行平板初筛及摇瓶复筛,并检测获得菌株的遗传稳定性。结果经初筛和复筛,获得1株磷脂酶A1高产菌株W22,经气相色谱鉴定其产酶为磷脂酶A1;该菌株在普通琼脂培养基上呈圆形,质地软,表面光滑、扁平,乳白色,晕圈明显,为芽孢杆菌属蜡样芽孢杆菌;最佳发酵条件为:发酵时间12 h,初始pH 8.0,温度32℃,摇床转速200 r/min,优化后酶活为9.12 U/ml,较优化前(6.74 U/ml)提高了35.3%;等离子体诱变后,经平板及摇瓶筛选,获得的菌株W22-5酶活最高,为12.80 U/ml,较诱变前(9.12 U/ml)提高了89.91%,且具有良好的遗传稳定性。结论成功获得1株磷脂酶A1高产菌株W22,优化了该菌的发酵条件,且经等离子诱变进一步提高了菌株的产酶能力,为磷脂酶A1的生产和应用奠定了基础。     

嗜温耐乙醇β-葡萄糖苷酶高产菌株及其性质

从生物质资源丰富的武夷山中采样天然腐烂枯枝、烂叶、土壤等材料中分离筛选,采用对硝基酚-β-葡萄糖苷(pNPG)法对复筛菌株进行酶活测定,得到一株高产β-葡萄糖苷酶菌株,并对其酶学性质进行研究,结果表明:该酶的液体发酵最高酶活高达482.1U/mL,最适反应pH值为4.8,最适反应温度为65℃;乙醇浓度为10%对酶活有最大促进作用,对β-葡萄糖苷酶酶活提高将近1倍,乙醇耐受能力高达30%。将该酶应用于同步糖化发酵中,发酵至120h得乙醇最高产量,所产乙醇含量高达41.25g/L,与阴性、阳性对照相比,乙醇产量提高近2倍。该菌株所产的β-葡萄糖苷酶酶活力较高,应用于同步糖化发酵过程具有明显的促进效果,对于促进纤维素乙醇的产业化进程具有广阔的发展前景。     

N~+注入诱变选育β-葡萄糖苷酶高产菌株及其产酶条件的优化

以筛选得到能够有效利用木糖渣作为碳源生产β-葡萄糖苷酶的菌株为目的,利用N+注入棘孢曲霉L22,筛选得到一株β-葡萄糖苷酶活提高1倍的突变株NIP35。通过单因子实验结合正交实验,对其培养基组成如碳源、氮源、表面活性剂等进行了优化,使其产酶水平最终提高了近2倍。     

高产纤维素酶菌株的筛选及产酶条件的研究

植物纤维是自然界最丰富的资源,筛选高产纤维素酶菌株,降低纤维素酶的生产成本,让植物纤维变废为宝一直是研究热点。从腐烂植物叶片中分离到一株高产纤维酶菌株CXYJ-1。对CXYJ-1菌株进行发酵培养条件的研究,结果表明,以豆饼粉为碳源,(NH_4)_2SO_4为氮源,培养温度为35～37℃,初始pH=5,培养96 h,产酶活力最高,CMC酶活为4.8 U/mL,FP酶活为4.5 IU/mL。     

拟青霉E7蛋白酶发酵条件及酶性质研究

针对产蛋白酶的淡紫拟青霉E7菌株发酵条件进行优化,得到利于产酶发酵培养基配方:以2%大豆粉为碳源、2%麦麸及1%硝酸铵为氮源,pH值7.0,28℃,150 mL三角瓶装液量为30 mL,培养时间72h.对该酶性质研究结果表明:蛋白酶在40 ℃热稳定性较好,最适pH值为10.5,在pH值9～11范围内具有较高的酶活与耐碱性.     

香蕉假茎中产碱性果胶酶细菌的筛选发酵优化与酶学性质

利用果胶酶处理天然纤维对环境污染小,然而筛选高产量的菌株仍是降低成本获得果胶酶的关键。本文利用以果胶为唯一碳源的培养基,从香蕉假茎中分离筛选出了一株具有较高果胶酶活性的菌株。经16S rDNA序列分析表明,该菌株与枯草芽孢杆菌(Bacillus substilis)具有99%的相似性,将该菌株命名为Bacillus sp. ZLXH-5。该菌株最佳发酵时间为2天,最佳发酵温度是37℃,最佳初始pH值是5,最佳转速是180r/min,最佳葡萄糖浓度为15g/L。通过发酵条件优化,果胶酶产量提高了56.6%。对粗酶液的性质进行分析,其在55℃下达到最佳反应速率,最适催化pH值是8.5,不同的离子对于果胶酶的活性起到不同的效果。由于该菌株具有较高的果胶酶活性,可以利用该菌株对香蕉假茎进行生物脱胶。     

产木聚糖酶优良菌株的选育

对荧光假单孢杆菌野生株（Pseudomonasflu）0－01进行紫外（UV）诱变和化学诱变后,得诱变株0-96,其在自落形态上稍有改变,产酶能力得到显著提高,本聚糖酶活可达325．45IU／mL,而纤维素酶的含量却比较低,只有0．5IU／mL左右。该菌株可用廉价的农副产品作为培养基,而获得较高的防产量。     

直接降解木质素的漆酶/木聚糖酶体系的合成

对一株高产漆酶及伴有木聚糖酶和少量纤维素酶的菌株,用不同的碳源和氮源对其调控培养,合成漆酶/木聚糖酶体系。实验结果表明,最佳的碳源是可溶性淀粉,用它作碳源,漆酶活性高达730IU/mL,木聚糖酶活性是4.49IU/mL,纤维素酶活性只有0.23IU/mL;最佳的氮源是蛋白胨,用其作氮源,合成漆酶活性可达812IU/mL,木聚糖酶活性是4.68IU/mL,纤维素酶活只有0.09IU/mL。     

氮源对里氏木酶合成木聚糖酶的影响

分别以（ＮＨ４）２ＳＯ４、酵母浸膏及蛋白胨为产酶氮源制备木聚糖酶。试验结果表明,酵母浸膏的产酶效果最好,其次是（ＮＨ４）２ＳＯ４。当以酵母浸膏为氮源产酶时,酶活力最高达到２５．４７ＩＵ／ｍＬ,酶得率和酶产率分别为３６３８．６ＩＵ／ｇ木聚糖和８４９０．０ＩＵ／Ｌ·ｄ,酶活力分别是以（ＮＨ４）２ＳＯ４及蛋白胨为氮源时的１．５倍和２．０倍     

Peach-palm (Bactris gasipaes Kunth.) waste as substrate for xylanase production by Trichoderma stromaticum AM7

This work investigated the xylanase production by fungi isolates from tropical agroforestry systems using peach-palm industrial waste as a substrate. Trichoderma stromaticum AM7 was the best xylanase producer and there was a 160% increase in xylanase activity after optimizing by the Box–Behnken statistical design. The optimization process demonstrated that the maximum activity occurred at 0.95% nitrogen concentration after 6 days of cultivation at 32°C, which achieved a yield of 1440?U?g^?1 of dry initial substrate. Analysis by scanning electron microscopy showed degradation of the fibers after cultivation. The optimum pH and temperature for xylanase activity were 5.0 and 50°C, respectively. The extensive degradation of the peach-palm waste and xylanase production by T. stromaticum AM7 was due to the combination of a good physicochemical composition of the culture medium and the characteristics of the selected fungus.     

产纤维素酶细菌的筛选及其紫外诱变

采用刚果红脱色圈法初筛得到143株有纤维素酶活性的细菌。然后采用DNS法复筛得到纤维素酶酶活最高的6株细菌,进一步进行紫外诱变处理,获得酶活提高最大且具有遗传稳定性的菌株E140’,酶活为0．90IU·mL^-1,较出发菌株（0．68IU·mL^-1）提高了32．35％。表明采用刚果红脱色圈法和DNS法联合筛选并结合紫外诱变。可以获得纤维素酶活性高的细菌。     

微波辐照选育木聚糖酶高产菌宇佐美曲霉

A high yield xylanase producing strain, A. usamii L336-23, was screened out from its parent strain A.usamii L336 after microwave irradiation. Its productivity of xylanase activity increased by 35.7% from 21000μu·ml-1 to 28500μu·ml-1 and was stable after frequent subcultures and storage for more than two months. The mechanism of microwave mutation was also discussed.     

Optimization of Xylanase Production from Penicillium sp.WX-Z1 by a Two-Step Statistical Strategy: Plackett-Burman and Box-Behnken Experimental Design

The objective of the study was to optimize the nutrition sources in a culture medium for the production of xylanase from Penicillium sp.WX-Z1 using Plackett-Burman design and Box-Behnken design. The Plackett-Burman multifactorial design was first employed to screen the important nutrient sources in the medium for xylanase production by Penicillium sp.WX-Z1 and subsequent use of the response surface methodology (RSM) was further optimized for xylanase production by Box-Behnken design. The important nutrient sources in the culture medium, identified by the initial screening method of Placket-Burman, were wheat bran, yeast extract, NaNO(3), MgSO(4), and CaCl(2). The optimal amounts (in g/L) for maximum production of xylanase were: wheat bran, 32.8; yeast extract, 1.02; NaNO(3), 12.71; MgSO(4), 0.96; and CaCl(2), 1.04. Using this statistical experimental design, the xylanase production under optimal condition reached 46.50 U/mL and an increase in xylanase activity of 1.34-fold was obtained compared with the original medium for fermentation carried out in a 30-L bioreactor.     

云芝木聚糖酶的培养和Tween80对其产酶的促进作用

本文研究了不同碳源和吐温80（Tween80）对云芝木聚糖酶产醇的影响，当以1％滤纸为碳源时，木聚精酶活最高可达11U／mL，用微晶纤维素为碳源时也能产生很高的木聚精酶．而以1％木聚糖为碳源时，只能产生很低的木聚精酶活．Tween80对木聚糖酶的产生具有明显的促进作用，分别可达3倍（微晶纤维素为碳原）和30％以上（滤纸为碳源），这主要由于Tween80能促进木糖耷醉的分泌，提高胞外木糖苷酶的活性。     

Crystallographic and mutational analyses of an extremely acidophilic and acid-stable xylanase: biased distribution of acidic residues and importance of Asp37 for catalysis at low pH

Xylanase C from Aspergillus kawachii has an optimum pH of 2.0 and is stable at pH 1.0. The crystal structure of xylanase C was determined at 2.0 A resolution (R-factor = 19.4%). The overall structure was similar to those of other family 11 xylanases. Asp37 and an acid-base catalyst, Glu170, are located at a hydrogen-bonding distance (2.8 A), as in other xylanases with low pH optima. Asp37 of xylanase C was replaced with asparagine and other residues by site-directed mutagenesis. Analyses of the wild-type and mutant enzymes showed that Asp37 is important for high enzyme activity at low pH. In the case of the asparagine mutant, the optimum pH shifted to 5.0 and the maximum specific activity decreased to about 15% of that of the wild-type enzyme. On structural comparison with xylanases with higher pH optima, another striking feature of the xylanase C structure was found; the enzyme has numerous acidic residues concentrated on the surface (so-called 'Ser/Thr surface' in most family 11 xylanases). The relationship of the stability against extreme pH conditions and high salt concentrations with the spacially biased distribution of charged residues on the proteins is discussed.      

响应面法优化壳聚糖酶产生菌Mitsuaria sp.K1的产酶发酵条件

壳聚糖（chitosan）及其降解产物因具有优良的物理特性和生物活性而被广泛关注。通过平板透明圈初筛、摇瓶复筛方法,从青岛海岸土壤中分离筛选到1株产壳聚糖酶活性较高的细菌Mitsuaria sp.K1,并对其产酶发酵条件进行了单因素试验和响应面优化分析试验。结果表明：在最适培养基组成（1%粉末壳聚糖、0.5%硝酸钾、0.22%KH2PO4、0.1%Na2HPO4、0.15%KCl、0.05%MgSO4?7H2O）和最佳培养条件（培养温度25.2 ℃,培养时间25.4 h,起始pH值6.5,接种量3%,装液量100 mL/500 mL摇瓶,160 r/min）下,Mitsuaria sp.K1的发酵粗酶液最高酶活平均达11.56 U/mL,比优化前的2.17 U/mL提高了4.32倍。与前人研究结果相比,该菌发酵产酶温度降低了5～10 ℃,产酶周期缩短了23～47 h,因此具有工业发酵应用价值。