固态混合发酵提高木聚糖酶和纤维素酶活力的研究

研究了接种比例、接种时间、碳源、氮源等因素对木霉和黑曲霉混合发酵产木聚糖酶和纤维素酶的影响。试验结果表明,当木霉和黑曲霉按4:6同时接种,以玉米芯3．75g、麸皮3．75g、葡萄糖37．5mg为混合碳源,Mandels营养盐11．5mL、添加NH_4NO_37．5mg为氮源,在84h产纤维素酶活力达到230IU/g干物质,木聚糖酶活力达到1308IU/g干物质,与两菌纯培养相比,纤维素酶活力提高163％,木聚糖酶活力提高79．5％。     

Rhizopus sp.PW358菌脂肪酶固态发酵生产

研究了Rhizopus sp.PW358菌的固态生长和产脂肪酶条件。结果表明：黄豆饼粉为培养基的基本成分,用来生产脂肪酶。培养基中可加入淀粉和蛋白胨作为碳源和源,有利于脂肪酶的合成,培养基的含水量以及金属离子Ca^2 ,Mg^2 的浓度也影响Rhizopus sp.PW358菌和脂肪酶 产生。在优化条件下,12g豆粉中含1.0g淀粉及0.5g蛋白胨、15ml营养盐中Ca^2 ,Mg^2 离子浓度分别为8．0和4．0g/L,培养基含水量为55．6％,在接种后培养48h,酶活力可达最大值320IU／g干培养基。脂肪酶的基本性质研究表明,酶的最适反应温度和PH分别为35℃和7．0,酶的半失活温度为53．5℃,不同的PH环境中,30℃保温1h后酶在PH6．5－8．5范围内较为稳定。     

长梗木霉纤维素酶的产生及提取

对长梗木霉(Trichoderma Longibrachiatum)ANU_3-958纤维素酶的产生及提取进行了研究。结果表明,固体曲培养144小时,固体曲与浸提液比例为1:7(W/V),采用硫酸铵分级沉淀法时所得纤维素酶活力最高。所得冻干纤维素酶粉经测定:羧甲基纤维素(CMC)酶活最高为2788.89IU/g,滤纸糖酶活(FPA)最高为79.44IU/g。相对固体曲得率平均为13.28%。     

微生物酶在不同压实度生物垃圾好氧处理中的影响

研究了农贸市场的生物垃圾不同压实度的好氧生物处理,对发酵初期的生物学指标如相关的纤维素酶及木聚糖酶活力进行测定与研究。实验表明:处理初期微生物酶活力大小对生物有机物的降解具一定作用。较适宜的压实度是0.247kg/L,其有机物降解度最大,达到41.9%;相关酶活力较高,纤维素酶活力达20.4567 IU/g.DM,木聚糖酶活力达40.28 IU/g.DM,并与温度变化曲线呈现出一致,与降解度也有一定相关性。     

细菌木聚糖酶高产菌的选育及发酵工艺研究

经筛选和诱变 ,选育出 1株产 β 木聚糖酶活力较高 ( 1 2 4.45IU/mL) ,而羧甲基纤维素酶活力较低( 1 .0 8IU/mL)的突变株L1 0 IGA。得出该菌产酶的最适发酵工艺为 1 .5 %半纤维素 ,0 .1 2 7%蛋白胨 ,0 .1 0 5 %K2 HPO4,0 .0 5 5 %MgSO4·7H2 O ,0 .45 %酵母膏 ,0 .2 2 5 %Twen40 ,装量 3 2 .5mL/1 5 0mL三角瓶 ,起始pH为 7.2 5 ,3 7℃发酵 48h后 45℃发酵 2 4h。     

斜卧青霉纤维素酶和木聚糖酶高产菌株的选育

以纤维素酶高产菌株斜卧青霉A50为出发菌株,通过紫外诱变原生质体获得1株木聚糖酶活力提高80%而纤维素酶活力没有改变的6号菌。蛋白质电泳和酶谱检测结果显示,纤维素酶谱基本无差别,而木聚糖酶谱显示6号菌比A50多了一条带。6号菌优化后的产酶培养基组成为:麸皮7%、葡萄糖0.1%,该条件下,纤维素酶活为19.7IU/mL,木聚糖酶活力为215.4IU/mL。     

利用天然纤维废弃物发酵生产L-乳酸的研究

为了降低L-乳酸的生产成本,更好的实现生物质秸秆的资源化,利用天然纤维素依次接种经离子注入诱变处理的木聚糖酶高产菌黑曲霉P602和米根霉RL6041高产菌进行固、液体二次发酵的方法,将其转化成用于工业生产的L-乳酸。结果表明:本实验条件下,未经过任何化学预处理的秸秆等物质接种黑曲霉P602进行固体发酵,产生的木聚糖酶活力为6 320 IU/g干(培养)基,纤维素酶活力为29 IU/g干基;加入100 mL水浸提后,产生的还原糖浓度为14.07 g/L,纤维物质糖化率为79.45%。取滤液接入米根霉RL6041进行液体发酵后,生成乳酸的量为7 g/L,糖酸转化率为47.6%,以(NH4)2SO4作为氮源时,最佳氮源浓度为3 g/L。     

纤维素降解真菌A25-2的分离、鉴定及其产纤维素酶的酶学特性

本研究以Avicel-刚果红选择培养基为初筛培养基,从云南哀牢山国家级自然保护区和广西猫儿山国家级自然保护区的土壤样品中分离筛选得到4200株真菌,从中筛选出透明圈与菌落直径比较大、透明程度较为清晰的12个菌株。通过液体培养发酵,测定其上清液中的羧甲基纤维素酶活力、滤纸酶活力和Avicel酶活力,最终筛选出一株产该三种酶且其活力均最高的真菌菌株A25-2。通过对菌株A25-2形态学观察和其内转录间隔区(internal transcribed spacer,ITS)序列同源性比对分析,将菌株A25-2鉴定为哈茨木霉(Hypocrea lixii)。酶活测定结果表明菌株A25-2产纤维素酶的酶活力较高,在最适作用pH4.5和最适作用温度55℃下,其羧甲基纤维素酶活力为2.26IU/mL,滤纸酶活力为0.58IU/mL,Avicel酶活力为0.39IU/mL。薄层层析实验表明A25-2具有完整的纤维素酶系统。因此,真菌A25-2可作为饲料加工等生产和纤维素酶相关研究的备选菌株。     

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%.     

利用黑曲霉固态发酵啤酒糟生产饲料复合酶的研究

以啤酒糟为主要基质,利用黑曲霉固态发酵生产酸性蛋白酶、木聚糖酶和纤维素酶等多种饲料复合酶,研究了黑曲霉固态发酵培养基组成对复合酶酶活的影响,确定最优培养基配方为:啤酒糟75%,麸皮25%,硫酸铵1%,KH_2PO_4 0.2%,MnSO_4 0.1%、ZnSO_4 0.2%,料水比1:2。在适宜的发酵条件下,经30℃发酵5 d,烘干后得到的复合酶制剂中,具有多种酶活性(以干基计)。其中酸性蛋白酶活力3 800 U/g,木聚糖酶活力12 00 U/g和纤维素酶活力18 U/g。     

根霉TC1653产纤维素酶提纯及酶学性质研究

对根霉所产纤维素酶酶系进行了分析并研究了部分酶学性质。实验选择超滤和凝胶柱分离相结合的方式提纯纤维素酶,结果显示根霉TC1653纤维素酶系是一个完全酶系,具有一个较为明显的内切葡聚糖酶组分。β-葡萄糖苷酶组分的最适反应温度为70℃,温度高于70℃时,活性迅速下降,但在这种高温下具有最高反应活性的酶很少见,很可能又是一种新的β-葡萄糖苷酶。     

A comparison of raw starch-digesting glucoamylase production in liquid and solid cultures of Rhizopus strains

Maximum growth for Rhizopus sp. A-11 was obtained at a zinc ion concentration of 0.7 ppm in a liquid medium. Glucoamylase (GA, EC 3.2.1.3) production in Rhizopus sp. A-11 was maximized at 710 U/ml, at the presence of 75 ppm for calcium and 0.7 ppm of zinc ions in liquid medium. Zinc ion is known as an essential biometal for Rhizopus growth; however, growth was inhibited by the zinc ion concentration, not maximized. Although calcium ion was not necessary to Rhizopus growth, GA production using Rhizopus sp. A-11 was markedly stimulated by calcium ion concentration over 75 ppm in the liquid medium. The GA productivity of the present liquid culture was about 4.4 times higher than that of the solid state culture, based on the unit starch amount in the liquid and solid media carbon source. The characteristics of the GA produced by the Rhizopus sp. A-11 liquid culture were interesting; that is, almost all the GA produced was classified as raw starch-digesting GA (GA-I). Secreted protein in the culture liquid after 30 h was nearly GA, and had a limited amount of impure protein. As a result, it was found that using a Rhizopus culture in a specified metal-ion regulated medium was an effective method for producing GA. Thus the present culture method was renamed the "metal-ion-regulated liquid culture method".     

黑根霉对人参皂苷Re的生物转化

利用菌种黑根霉Rhizopus sp.对人参皂苷Re进行生物转化,并对人参皂苷Re及其发酵产物进行HPLC系统分析比较,经液相色谱-质谱分析得出人参皂苷Re转化率为92.16%,并制备出人参皂苷Re发酵产物中峰值升高的成分,转化后的人参皂苷发酵产物中化合物1确定为人参皂苷Rg2,化合物2为Rg2的同分异构体,得率为10.13%;化合物3和化合物4确定为人参皂苷Rg5/Rk1,得率为29.23%。从结果初步推测得出人参皂苷Re被黑根霉转化为人参皂苷Rg2的机理,人参皂苷Re转化成人参皂苷Rg5/Rk1的机理还有待于进一步研究。     

诱变选育脂肪酶高产菌株及其脂肪酶固定化

以紫外和微波复合诱变选育脂肪酶产生菌 Rhizopus sp. RXF12,获得高产突变株RZ13,其脂肪酶摇瓶发酵单位是出发株的2.62倍。菌株经多次传代,遗传性状稳定。对RZ13菌株的发酵条件进行了正交优化,在25 ℃、pH 8.0的条件下,接入5 %(v/v)的RZ13菌株单孢子悬液 (107个/ml) 振荡培养84 h,达到RZ13菌株最佳产酶状态,脂肪酶活可达95.08 U/ml。考察了脂肪酶性质,在低于40 ℃,pH 7.0~9.0范围内脂肪酶活稳定。经载体筛选及固定化过程优化,选用镁铝水滑石25℃吸附4 h,对RZ13脂肪酶进行了固定化。结果表明,固定化酶的最适作用温度为35~55℃,pH为7.5~9.0,较游离酶的均有较大扩展。     

聚球藻7942高效泌氨突变种的获得及其泌氨、谷氨酰胺合成酶活性、光合和生长

将含有Ａｎａｂａｅｎａｓｐ．ＰＣＣ７１２０反义ｇｌｎＡ基因片段的穿梭表达质粒ｐＤＣ－ＡＴＧＳ转化单细胞蓝藻聚球藻Ｓｙｎｅ－ｃｈｏｃｏｃｃｕｓ　ｓｐ．ＰＣＣ７９４２,通过同源重组,外源ＤＮＡ定位整合到染色体上。经过抗菌素筛选,获得一种高效泌氨的Ｓｙｎｅｃhｏｃｏｃｃｕｓ　ｓｐ．７９４２突变种。将此突变种固定化在聚氨脂泡沫中后,定量测定其谷氨酰胺合成酶（ＧＳ）活性。结果表明,固定化后的突变藻培养９ｄ后泌氨活性比自由生活的野生藻高１５６倍,ＧＳ活性降低７３．６％;其生长速度与同条件下野生藻相近,７７Ｋ荧光光谱表明突变种固定化后光系统Ⅱ活性提高４４％。     

Establishment of a Highly Efficient Ammonia Secreting Mutant of Synechococcus sp. PCC 7942 and the Glutamine Synthetase Activity,Photosynthesis and Growth in Its Immobilized Cells

A recombinate plasmid pDC-ATGS was constructed, which contained the antisense fragment of glnA gene from Anabaena sp. PCC 7120 and transformed the unicellular cyanobactefium Synechococcus sp. PCC 7942. The foreign DNA was inserted into the site of glnA locus of the chromosome through the homologous recombination. By using neomyisin, a highly efficient ammonia secretion mutant was selected. After immobilized, the cells of the mutant within polyurethane (PU) foams, glutamine synthetase (GS) and NIt4+ secretory activity of GS, and its growth and photosynthesis were measured. It was shown that NH4+ secretion of the immobilized mutant was enhanced 156 folds which was much higher than that of free-living cells of the wild type. The activity of GS was decreased by 73.6%. Growth of the mutant was the same as that of the wild type. The activity of photosystem Ⅱ in the immobilized mutant cells increased by 44% with 77 K fluorescence spectrum measurement.     

血栓溶解酶产生菌及其培养条件的研究

通过对根霉１２号发酵液的分析及发酵条件的研究,发现其发酵液中含有能溶解血栓的物质,但不能分解血细胞。它在麸皮胰蛋白胨培养基（ｐＨ５．１）上３０℃振荡培养４８～６０ｈ,ｐＨ值达到７．４左右时产生血栓溶解酶的活力最高。     

Conversion of zearalenone to zearalenone glycoside by Rhizopus sp

The microbial conversion of zearalenone by various species of fungi was studied. Among them, Rhizopus sp. was the sole fungus which produced a new metabolite from zearalenone in addition to alpha- and beta-zearalenol. The structure of the new metabolite was determined to be zearalenone 4-beta-D-glucopyranoside on the basis of mass, infrared, and nuclear magnetic resonance spectroscopies. The results suggest that the mycelium of Rhizopus sp. catalyzes the glycosidation at the C-4 position of zearalenone.     

Conversion of zearalenone to zearalenone glycoside by Rhizopus sp.

The microbial conversion of zearalenone by various species of fungi was studied. Among them, Rhizopus sp. was the sole fungus which produced a new metabolite from zearalenone in addition to alpha- and beta-zearalenol. The structure of the new metabolite was determined to be zearalenone 4-beta-D-glucopyranoside on the basis of mass, infrared, and nuclear magnetic resonance spectroscopies. The results suggest that the mycelium of Rhizopus sp. catalyzes the glycosidation at the C-4 position of zearalenone.     

不同地区根霉分离株的RAPD多样性分析

为了研究微生物的分子多样性,在获得系列米根霉分离物的基础上,应用经筛选的13条随机引物,对来自不同地区的18株米根霉分离物进行了RAPD分析,聚类结果表明,18株分离物在0.75的相异水平上可以聚为6个群体,其分子多样性同地理来源之间具有一定的相关性。