有机复合物H对纤维素CMC酶活的影响

采用3,5-二硝基水杨酸(DNS)为显色剂,CMC为底物,测定纤维素酶的酶活。考察了在不同的酶促反应温度、pH值、底物浓度、反应时间等反应条件下,有机复合物H对纤维素酶CMC酶活(CMCA)的影响。结果表明,有机复合物H对纤维素酶的CMCA活性有明显促进作用,且在不同条件下的促进效果有较大差异。     

里氏木霉液体发酵产纤维素酶的研究

在摇瓶试验基础上,采用里氏木霉(Trichoderma reesei)HC-415菌株进行5L自控罐产纤维素酶深层发酵试验。在通气量为 0.2—0.6vvm、搅拌速度为 400r/min、发酵液pH控制在5.8—6.1的条件下,发酵液的羧甲基纤维素(CMC)酶酶活最高为325.0mg糖/ml,滤纸糖酶(FPA)酶活最高达17.9mg糖/ml。发酵周期为108h。所得冻干纤维素酶粉CMC酶活最高3111IU/g,FPA最高135IU/g ,对发酵液得率平均6.7g/L。酶活总收率CMC酶活平均78.2％,FPA酶活平均73.5％。     

秸秆纤维素分解菌的酶活力测定

目的:测定秸秆纤维素分解菌的酶活力。方法:从土壤中分离出具有分解纤维素能力的菌株,采用刚果红染色法进行粗选,得到7株透明圈较大的菌株。将这7株菌株液体发酵培养6d,再分别用滤纸分解度观察、羧甲基纤维素酶活法(CMC)、滤纸酶活法(FPA)和天然纤维素酶活法测定其酶活力。结果:在7株菌株中,F-1、F-2、F-3、F-5的酶活力测定结果与其溶解圈的测定结果、滤纸分解结果基本相同。且天然纤维素酶活力高的菌株,其CMC酶活、FPA酶活也高,滤纸分解效果也比较明显。结论:CMC法、FPA法和天然纤维素酶活法适于测定秸秆纤维素分解菌的酶活力。     

纤维基质对液体培养灵芝产纤维素酶的影响

本研究通过在培养基中添加滤纸、羧甲基纤维素(CMC)、蔗渣和玉米芯等纤维素类基质,观察纤维素基质对液体培养条件下灵芝产纤维素酶和半纤维素酶的影响.结果表明,这些纤维素基质能促进纤维素酶活力的增加,促进作用效果各异,当滤纸的添加量达到1 g/L时,滤纸酶活(FP酶活)达到空白对照的9.04倍,当蔗渣添加量为1 g/L时,灵芝β-1,4-葡聚糖酶酶活(Cx酶活)、FP酶活、半纤维素酶活分别比空白对照样增加了12%、534%、117.3%.     

一株高产纤维素酶菌的筛选与鉴定

利用刚果红脱色从森林腐木中筛选到一株高产纤维素酶菌,结合18S rRNA基因序列分析和菌株形态特性分析,确定该菌株为爪哇正青霉(Eupenicillium javanicum).通过研究粗酶提取时不同离心时间和离心转速对酶活的影响,确定采用3000r/min,15min离心条件下获得的粗酶测定CMC酶活,采用4000r/min,10min的离心条件下获得的粗酶测定FPA酶活和β-葡萄糖苷酶活.酶学性质初步研究显示,纤维素酶反应的最适pH值以5.0为宜;CMC酶最适酶解温度和酶解时间为60℃,15min;FPA和β-葡萄糖苷酶最适酶解温度和酶解时间均为50℃,20min.     

三株丝状真菌分解樟子松凋落物酶活性

该研究以在樟子松凋落物层中高频出现的3株丝状真菌Alternaria sp.、Penicillium sp.和Pestalotiopsis sp.为供试菌株, 以樟子松新鲜落叶为作用底物, 通过发酵纯培养的方法, 测定了底物有机物质质量损失及发酵过程中产生的漆酶(Laccase)、锰过氧化物酶(MnP)、羧甲基纤维素酶(CMCase)和滤纸酶(FPA)酶活性变化, 并验证了酶活性与底物降解的关系。结果表明, Alternaria sp. 引起底物总有机物质质量损失最大, 且产生的漆酶、羧甲基纤维素酶和滤纸酶活性都较高; Penicillium sp. 产生的锰过氧化物酶活性最高。3株丝状真菌同属于真菌功能群中的木质纤维素分解者。     

微波诱变结合化学诱变选育纤维素酶高产菌的研究

以纤维素酶产生菌TP1202为出发菌株,通过微波诱变和硫酸二乙酯、氯化锂诱变剂进行诱变,选育到1株纤维素酶高产菌株AS5。在适宜条件下,其产生的羧甲基纤维素酶活力(CMC),滤纸酶活力(FPA)和棉花糖酶活力分别是出发菌株的107%、152.4%和140.5%。     

发酵床中纤维素降解菌的分离与鉴定

从发酵床垫料中初步分离出43株纤维素降解菌。采用刚果红鉴别培养基及滤纸条培养基初筛,得到5株透明圈较大且使滤纸条产生崩解的菌株,通过进一步液体发酵,测定其CMC酶活、FPA酶活和天然纤维素酶活,获得2株具有较高纤维素降解活性菌株,并分别命名为F7和F21。经16S rRNA基因序列分子生物学鉴定和系统发育分析表明,这2株纤维素降解菌分别归属为枯草芽孢杆菌(Bacillus subtilis)和链霉菌(Streptomyces sp.)。     

纤维素降解菌Aspergillus sp. YN1的产酶条件及酶学特性

从牛羊粪堆肥中筛选出一株纤维素降解菌Aspergillus sp.YN1,主要研究了液体发酵培养基中碳源、氮源、培养温度、起始pH、通气量以及接种菌龄对菌株YN1的羧甲基纤维素酶活(CMC酶活)及滤纸酶活的影响。研究结果表明,在优化条件下,该菌的CMC酶活、滤纸酶活在培养第3天分别达到0.53U/mL和0.15U/mL。在酶学特性研究中,菌株YN1的CMC酶的最适反应温度为70°C,最适反应pH4.0(酶促反应为30min)。用不同温度处理1h或不同pH处理2h,YN1的CMC酶在30°C?50°C或pH3.0?4.0之间仍可保持80%以上的酶活性,对热及酸表现出较高的稳定性。     

产纤维素酶大肠杆菌工程菌的构建和酶的表达

为了使大肠杆菌(Escherichia coli)具备生产纤维素酶的能力,从铜绿假单胞菌(Pseudomonas aeruginosa)PKC-001克隆碱性纤维素酶基因,进行密码子优化后连到载体pET-22b,转入大肠杆菌BL21(DE3)菌株,对工程菌进行表达优化并分析纤维素酶酶活。结果表明大肠杆菌工程菌能生产分子量约39 kD的纤维素酶,在培养温度20℃、摇床转速170 r/min、IPTG终浓度0.01 mmol/L的条件下,纤维素酶的产量最大;在酸性条件下,细胞内和细胞外的滤纸酶活力(FPA)分别为5.78 U/mL和1.13 U/mL,在碱性条件下检测不到滤纸酶活。显然,重组纤维素酶仅在酸性条件下有活力。     

玉米秸秆纤维素分解菌的选育研究

玉米秸秆是纤维组分含量很高的农作物残留物,利用纤维素分解菌生产发酵饲料是当前饲料业的一个发展方向,它可将纤维素分解为牲畜可利用的糖,同时增加饲料中蛋白质的含量.试验从微生物丰富的土壤中分离到10株能分解纤维素的菌株,分别测定滤纸分解度、CMC酶活、FPA酶活和天然纤维素酶活,筛选出6株对天然秸秆纤维素有较强降解能力的菌株.通过改变其培养基中天然纤维素的含量,发现随着培养基中天然纤维素含量的增加,酶活力也随之升高.     

紫外线和60Co-γ联合诱变褐色高温单孢菌及产酶特性研究

本研究以褐色高温单孢菌(Thermomonospora fusca)为出发菌株,通过紫外线和60Co-γ射线联合诱变,获得了一株纤维素酶高产菌株AV8,CMC酶活力达到0.679 IU/mL,与出发菌株相比,其产酶能力提高3.53倍。通过对AV8产纤维素酶的培养条件进行测定,结果显示:产纤维素酶最适应温度为55℃;该菌的最适产CMC酶初始PH值为7.0,最适产FPA酶初始PH值为8.0;当培养到第7天时,产CMC酶达到高峰。当培养到第8天时产FPA酶达到高峰。     

Synergistic Effects of Organic Contaminants and Soil Organic Matter on the Soil-Water Partitioning and Effectiveness of a Nonionic Surfactant (Triton X-100)

Napthalene- and decane-contaminated soils were treated with Triton X-100 (a nonionic surfactant) to characterize the soil-water partitioning behavior of the surfactant in soils with different organic content. Soil samples with different organic content were prepared by mixing sand-mulch mixtures at different proportions. The experimental results indicated that the amount of surfactant sorbed onto soil increased with increasing soil organic content and increasing surfactant concentration. The effective critical micelle concentration (CMC) also increased with increasing organic content in soil. The CMC of Triton X-100 in aqueous systems without soil was about 0.3 mM and the effective CMC values measured for soil-water-surfactant systems (approximately 1:19 soil/water ratio) with 25%, 50%, and 75% mulch content were 0.9, 1.0, and 1.7 mM, respectively. Sub-CMC surfactant sorption was modeled accurately with both the Freundlich and the linear isotherm. The maximum surfactant sorption onto soil varied from 66% to 82% of added surfactant in the absence of contaminant. The effective CMC values for Triton X-100 increased to some extent in the presence of contaminants, as did the maximum surfactant sorption. The maximum surfactant sorbed onto the soil with 75% mulch content increased from 82% for clean soils, to 95% and 96% for soils samples contaminated with naphthalene and decane, respectively.     

一株纤维素降解菌的分离、鉴定及对 玉米秸秆的降解特性

[目的]获得高产纤维素酶细菌菌株,探讨以氨化预处理玉米秸秆为底物时的纤维素酶产酶特性及底物降解特性,探讨纤维素酶作用机理,提高玉米秸秆利用率.[方法]用LB培养基分离并纯化菌株,羧甲基纤维素钠培养基培养、刚果红染色进行初步筛选.考察氨化预处理对底物降解率、产酶能力的影响.通过形态特征观察及16S rRNA、Biolog鉴定菌株.[结果]分离到一株高效纤维素降解菌NH11,经鉴定为枯草芽孢杆菌(Bacillus subtilis). 30℃、发酵5d时,预处理前后玉米秸秆降解率分别为14.24％和24.73％.30℃、pH 7.2时,处理组CMC酶活力峰值处为153.84 U/mL,FPA酶活力为197.24 U/mL,比未处理组分别高出11.45％和10.59％.[结论]NH11具有较高的纤维素酶产酶能力,氨化预处理能够提高菌株对玉米秸秆的降解率.该菌株在秸秆堆肥、制作食用菌培养基和制取反刍动物粗饲料方面具有很高的应用价值.     

Hydrogenase activity of Rhodopseudomonas capsulata growing on organic media

When Rhodopseudomonas capsulata B10 grows in media with different organic compounds, the hydrogenase activity estimated both by the evolution and uptake of H2 is lowest in cells taken from the middle of the exponential growth phase, and highest in cells from the beginning of the stationary phase. Cells grown in a medium containing malate have a higher hydrogenase activity than those cultivated in a medium with lactate or other compounds (900 and 20 nmoles of H2 per 1 min per 1 mg of protein, respectively). In the experiments with chloramphenicol (10(-5) M), organic compounds (not CO2) were shown to repress hydrogenase synthesis. When the cells were incubated in a medium without an organic substrate or in its presence, the exogenous H2 or H2 evolved as the result of nitrogenase action causes an increase in the activity of hydrogenase.     

Lipase-catalyzed cellulose acetylation in aqueous and organic media

Screening for lipases capable of catalyzing acetylation of cellulosic substrates was conducted in aqueous buffer solution using water-soluble carboxymethyl cellulose (CMC) as substrate. Lipase A12 from Aspergillus niger (A. niger) showed the most promising acetylation activity among 11 tested commercial microbial lipases and was further applied to catalyzing acetylation of solid cellulose in aqueous solution. This reaction was shown to be feasible with an acetylation extent of 0.16 wt % achieved compared with no detectable acetylation in the absence of enzyme. Pretreatments on cellulose substrate by ultrasonic irradiation and surfactant solution only slightly improved the acetylation extent by 44 and 27%, respectively. Alternatively, this lipase-catalyzed acetylation was remarkably improved with solubilized cellulose as substrate in the dimethyl sulfoxide/paraformaldehyde solvent system, with an acetylation extent (7.87 wt %) nearly 50 times higher than that achieved in aqueous solution. This improvement was attributed to (1) the absence of bulk water and the increase in substrate solubility by the transition of reaction media from aqueous solution to organic solvents and (2) the ability of lipase A12 to remain catalytically active in highly polar DMSO. This discovery that the A. niger lipase was capable of surviving its contact with polar solvents was further confirmed by its considerably preserved catalytic activity on CMC acetylation in aqueous media after enzyme pretreatments with organic solvents of various polarities and in mixture media with the aqueous phase partially replaced by organic solvents.     

复合菌系RSS-4传代对水稻秸秆腐解的影响

对筛选到的1组稻秆腐解复合菌系RSS-4,以腐解产物的pH值、酶活及稻秆失重率为指标,研究了该复合菌系的性质稳定性和功能稳定性;并用变性梯度凝胶电泳法(DGGE)分析了该复合菌系的菌种组成稳定性。结果表明,多代继代培养过程中各代复合菌系腐解产物的pH值变化趋势基本一致,pH值从起始的6.72左右迅速升至最高点7.75,然后逐渐下降并稳定在7.20;多代继代培养后复合菌系各代之间的酶活动态变化及其对稻秆腐解能力的差异均很小。在连续继代培养的过程中,DGGE图谱揭示第15~23代复合菌系之间的菌种组成没有明显差异。     

固定化光合细菌利用有机物产氢的研究

应用固定化细胞技术包埋荚膜红假单胞菌(Rhodopseudomonas capsulata)菌株386．研究在光照下利用有机物产氢的特性。实验观察到,光照培养120小时,悬浮培养物的产氢量为68．2ml·比产氢速率为104．1ml H2/g(生物量)·h;用琼脂包埋后．其产氢能力得到改善,产氢量和比产氢速率分别达到128．4ml和l 9s．8mlH2/g·h。该菌株除可利用苹果酸外,还可利用葡萄糖、乳酸、丙酸等基质高效地产氢。基质浓度只有控制在适当水平时,才具有较高的基质转化产氢效率。此外．菌体生物量、菌龄、培养液pH、光照强度、光照／黑暗时间比以及温度对产氢过程均有不同程度的影响。