一株纤维素降解菌的分离、鉴定及产酶条件初步研究

目的:筛选具有纤维素降解能力的菌株.方法:在广州近郊自然环境取样,利用CMC固体平板筛选具有纤维素降解能力的茵株.结果:分离获得1株纤维素降解能力较强的菌株OY-01.该菌株最适生长温度和pH分别为35℃和6.0-7.0.形态学观察、生理生化和28S rDNA序列分析表明该菌株属于烟曲霉(Aspergillus fumigatus).结论:菌株OY-01被鉴定为烟曲霉,其最适产酶条件为:氮源为硫酸铵,浓度0.15%,温度为35℃,初始pH6.0-7.0,培养48-60h,Aspergillus fumigatus OY-01具有一定的工业应用潜力.     

青霉生产木质纤维素降解酶系的研究进展

木质纤维素降解酶系的高效生产是实现植物生物质大规模生物炼制的重要支撑。就地生产木质纤维素降解酶,有助于降低其使用成本,提高技术经济效益。青霉是自然界常见的木质纤维素降解真菌,可以合成分泌种类多样、组分齐全的木质纤维素降解酶系,已被应用于纤维素酶制剂的工业生产。文中从就地生产降解酶,为木质纤维素生物炼制构建“糖平台”的角度,综述了青霉木质纤维素降解酶系的性质、菌株遗传改造及发酵工艺的研究进展。     

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。     

纤维素降解菌L-06的筛选、鉴定及其产酶条件的分析

从用于堆肥的水稻秸秆中初筛出一株高效纤维素降解菌L-06, 根据18S rRNA基因序列和菌株形态分析, 初步鉴定该菌为斜卧青霉(Penicillium decumbens)。研究了液体发酵培养基中氮源、碳源、表面活性剂、培养温度、起始pH以及接种量对该菌株各纤维素酶活力的影响。在最适条件下, 该菌的b-葡萄糖苷酶(BGL)、外切纤维素酶(CBH)于培养第3天酶活力分别达到1662 u/mL和2770 u/mL; 内切纤维素酶(EG)、滤纸糖化力(FPase)于培养第4天酶活力分别达到18064 u/mL和4035 u/mL。在产酶优化实验中, 该菌的EG和CBH在pH10的培养条件下分别保持了70%和43%的酶活性; 在50oC培养条件下EG和CBH分别保持了68%和59%的酶活性。表现出了耐热, 耐碱的特性。     

纤维素降解菌L-06的筛选、鉴定及其产酶条件的分析

从用于堆肥的水稻秸秆中初筛出一株高效纤维素降解菌L-06, 根据18S rRNA基因序列和菌株形态分析, 初步鉴定该菌为斜卧青霉(Penicillium decumbens)。研究了液体发酵培养基中氮源、碳源、表面活性剂、培养温度、起始pH以及接种量对该菌株各纤维素酶活力的影响。在最适条件下, 该菌的b-葡萄糖苷酶(BGL)、外切纤维素酶(CBH)于培养第3天酶活力分别达到1662 u/mL和2770 u/mL; 内切纤维素酶(EG)、滤纸糖化力(FPase)于培养第4天酶活力分别达到18064 u/mL和4035 u/mL。在产酶优化实验中, 该菌的EG和CBH在pH10的培养条件下分别保持了70%和43%的酶活性; 在50oC培养条件下EG和CBH分别保持了68%和59%的酶活性。表现出了耐热, 耐碱的特性。     

青霉F10-2固态发酵植物农药残渣产纤维素酶

采用单因素试验观察了不同C、N源、植物提取残渣与麦麸比例、初始pH和水料比对青霉F10-2菌株产纤维素酶的影响;在此基础上根据Box-w ilson的中心组合试验设计原理,选取蛋白胨、初始pH和水料比等影响因素,采用三因素五水平的响应面分析方法,对瓦克青霉F10-2的固态产纤维素酶条件进行了优化。其优化后的产酶条件为m(川楝树皮残渣)∶m(麦麸)∶m(蛋白胨)∶m(KH2PO4)=80∶20∶1.4∶0.4、初始pH 6.2、水料比2∶1、26℃发酵8 d,在此条件下纤维素酶比酶活可达6.47 U/g,较原始培养条件提高了46.38%。     

氯氰菊酯降解菌草酸青霉SSCL-5分离鉴定及降解特性

为解决土壤中残留的菊酯类农药问题,通过富集筛选法从70余份土壤中获得一株高效分解利用菊酯类农药的微生物菌株SSCL-5,该菌株可在含1 000 mg/L的氯氰菊酯无机盐培养基中正常生长。经形态学及ITS测序鉴定,确定该菌为草酸青霉(Penicillium oxalicum),对多种高浓度菊酯类农药耐受。经紫外分光光度法及HPLC验证该菌株草酸青霉SSCL-5在无机盐培养基、28℃、180 r/min摇瓶培养24 h的条件下,对400 mg/L氯氰菊酯的降解率为97%。土壤室内试验证明该菌在土壤中,温度20-34℃、水分含量保藏40%-60%条件下,30 d可将土壤中400 mg/L的氯氰菊酯降解67.6%。     

柚子皮黑曲霉的分离鉴定及产酶特性研究

对柚子皮上自然生长的黑曲霉进行分离鉴定,并探讨其产酶特性。以平板稀释法从柚子皮上分离出一株霉菌菌株,通过观察其形态特征和培养特征,对照《真菌鉴定手册》判定该菌株的种属;采用鉴定培养基法对其产酶特性进行分析。根据柚子皮的成分特性,以干柚子皮为主要原料,该菌为生产菌株,采用固态发酵法探究培养基的成分、柚子皮含量、培养基初始含水量及发酵时间4个因素对纤维素酶活力的影响。结果表明,该菌株为黑曲霉(Aspergillus nige),可产淀粉酶、蛋白酶、纤维素酶、果胶酶;固态发酵培养基中添加柚子皮12g,麸皮0.5 g和(NH_4)_2SO_40.5 g,培养基初始含水量保持在68.5 mL/100 g,培养时间控制在60 h左右时纤维素酶产量较高。     

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。     

大熊猫肠道纤维素分解菌的分离鉴定及产酶性质

【目的】从健康大熊猫新鲜粪便中分离具有纤维素酶活性的菌株,并对其进行菌种鉴定及产酶性质研究。【方法】利用羧甲基纤维素钠培养基分离纯化具有较高纤维素酶活性的菌株,根据形态学特征、生理生化特性以及16S rDNA分析对其进行分类鉴定,研究影响该菌株纤维素酶的产酶条件,以及对不同纤维素底物的降解情况。【结果】分离得到一株纤维素酶产生菌株P2,该菌株为好氧的革兰氏阳性细菌,生长温度范围20-50℃(最适温度37℃),pH范围6.0-9.0(最适pH7.0),NaCl浓度范围0%-15%(最适2%NaCl),培养24h达到产酶高峰。16S rDNA基因序列分析显示,菌株P2与解淀粉芽胞杆菌(Bacillusamyloliquefaciens)NBRC15535相似性为99.66%。该菌株对四种纤维素底物(滤纸、脱脂棉、秸秆、竹纤维)均有不同程度的降解,内切葡聚糖酶、外切葡聚糖酶、β-葡萄糖苷酶和总酶活具有不同的酶活变化。【结论】本研究首次从大熊猫粪便中分离出了好氧纤维素分解菌,并鉴定为解淀粉芽胞杆菌,对上述四种纤维结构均有一定的破坏和分解作用,为进一步研究大熊猫竹纤维消化机制提供了菌源。     

红树植物内生真菌Penicillium sp.中两个甾体化合物的研究

从红树植物内生真菌Penicillium sp.的发酵液中分离纯化了两个甾体类化合物,通过各种波谱实验(1D-NMR,2D-NMR,ESI-MS)确定为:麦角甾-4,6,8(14),22-四烯-3-酮(1)和麦角甾-5,7,22-三烯-3-醇(2),1对于3α-HSD脱氢酶在250μm浓度下有较弱的活性。     

Cellulases from two Penicillium sp. strains isolated from subtropical forest soil: production and characterization

AIMS: To isolate new fungal strains from subtropical soils and to identify those that produce high cellulase activity. To select microbial strains producing thermostable cellulases with potential application in industry. METHODS AND RESULTS: The new strains Penicillium sp. CR-316 and Penicillium sp. CR-313 have been identified and selected because they secreted a high level of cellulase in media supplemented with rice straw. Analysis by sodium dodecyl sulfate polyacrylamide gel electrophoresis, isoelectric focussing and zymography showed that the studied strains secreted multiple enzymes that hydrolyse cellulose. Cellulase activity of Penicillium sp. CR-316, the strain showing higher production, was analysed. Optimum temperature and pH of carboxymethyl cellulase activity were 65 degrees C and pH 4.5, respectively. Activity remained stable after incubation at 60 degrees C and pH 4.5 for 3 h. CONCLUSIONS: Fungal strains that secrete high levels of cellulase activity have been characterized and selected from soil. The isolated strains have complex sets of enzymes for cellulose degradation. Crude cellulase produced by Penicillium sp. CR-316 showed activity and stability at high temperature. SIGNIFICANCE AND IMPACT OF THE STUDY: Two fungal strains with biotechnological potential have been isolated. The strains secrete high levels of cellulase, and one of them, Penicillium sp. CR-316, produces a thermostable cellulase, that makes it a good candidate for industrial applications.     

青霉菌m8产半纤维素酶条件及酶性质研究

半纤维素酶可用于造纸工业的生物制浆和废水处理。通过碳、氮源对产胞外半纤维素酶的影响确定了青霉菌m8的适宜培养基 ,即 4 .5 %麦草粉 ,0 .5 0 % (NH4 ) 2 SO4 ,0 .1%KH2 PO4 ,0 .0 5 %MgSO4 ·7H2 O ,0 .0 3%NaCl,0 .33%Tween80 ,0 .15 %CaCO3。在上述培养基中 ,2 8℃恒温振荡 (12 0r min)培养 4 - 6d ,半纤维素酶活力可达 80u ml左右。用DNS法研究了该酶的性质。结果表明 ,其最适pH值为 4 .5 ,最适反应温度为 5 5℃ ;表观Km值为 4 .6× 10 - 2 g L ;该酶的耐热性比较强 ,可被K+ 、Ca2 + 、Mg2 + 离子激活 ,而被Ag+ 、Fe3+ 和Cu2 + 离子抑制。     

6'-Hydroxyoxosorbicillinol,a New Lipoxygenase Inhibitor and PGD2/LTB4 Release Suppressor from Penicillium sp.

A new lipoxygenase inhibitor, 6'-hydroxyoxosorbicillinol (1, C₁₄H₁₆O₆), was identified from a culture of Penicillium sp. A known compound, oxosorbicillinol (2, C₁₄H₁₆O₅), was also isolated. Compound 1 showed an approximately 10 times greater inhibitory effect on soybean lipoxygenase (IC₅₀, 16 µM) than 2 (IC₅₀, 150 µM), and also showed prostaglandin D₂ (PGD₂) and leucotriene B₄ (LTB₄) release suppression activity (IC₅₀, 10 µM for PGD₂ and 100 µM for LTB₄).     

Penicillium sp.脂肪酶的发酵及催化生成生物柴油的研究

目的：为了提高脂肪酶的产量及更好地应用脂肪酶。方法：采用单因子实验与均匀设计相结合的方法,对青霉Penicil- lium sp．TS414发酵生产脂肪酶的条件进行了优化。结果：在实验优化后的最适产酶培养基中,碳源为1．4%蔗糖,氮源为7．0%豆饼粉,起始pH8．0。均匀设计优化后的产酶水平(315．1U/mL)比优化前(101．5U/mL)提高了约2倍。Penicillium sp．TS414脂肪酶能够有效地催化大豆油转酯化合成脂肪酸甲酯(生物柴油),反应72h后,脂肪酸甲酯的最终得率在96%左右。结论：Penicillium sp．TS414产生的脂肪酶在生物柴油的工业化生产方面,具有潜在的应用前景。     

Influence of cultivating conditions on the alpha-galactosidase biosynthesis from a novel strain of Penicillium sp. in solid-state fermentation

AIMS: The work is intended to achieve optimum culture conditions of alpha-galactosidase production by a mutant strain Penicillium sp. in solid-state fermentation (SSF). METHODS AND RESULTS: Certain fermentation parameters involving incubation temperature, moisture content, initial pH value, inoculum and load size of medium, and incubation time were investigated separately. The optimal temperature and moisture level for alpha-galactosidase biosynthesis was found to be 30 degrees C and 50%, respectively. The range of pH 5.5-6.5 was favourable. About 40-50 g of medium in 250-ml flask and inoculum over 1.0 x 10(6) spores were suitable for enzyme production. Seventy-five hours of incubation was enough for maximum alpha-galactosidase production. Substrate as wheat bran supplemented with soyabean meal and beet pulp markedly improved the enzyme yield in trays. CONCLUSIONS: Under optimum culture conditions, the alpha-galactosidase activity from Penicillium sp. MAFIC-6 indicated 185.2 U g(-1) in tray of SSF. SIGNIFICANT AND IMPACT OF THE STUDY: The process on alpha-galactosidase production in laboratory scale may have a potentiality of scaling-up.     

青霉菌胞外半纤维素酶的固定化研究

青霉菌m8胸外半纤维素酶经过离子交换和分子筛层析两步纯化,纯化的酶固定在戊二醛交联的壳聚精上,残活力为45．6％。固定化酶的最适pH为3．6,最适温度为65℃,且固文化酶在65～75℃活力都较高。该酶的耐热性比较强,固定化酶热稳定性优于原酶;以半纤维素为底物,固定化酶的表观米氏常数为3．58×10－2g／L。     

Sclerotia growth and carotenoid production of Penicillium sp PT95 during solid state fermentation of corn meal

Using corn meal as fermentation substrate, the effect of some factors, fermentation time and supplementation of saccharide and nitrogen sources as well as vegetable oil, on the sclerotia growth and carotenoid production of Penicillium sp PT95 during solid state fermentation were studied. When PT95 strain was grown on the amended medium by supplementing of 3g NaNO3, 10g maltose and 2.5g soybean oil per liter of salt solution to basal medium for 20 days, the dry sclerotia weight and carotenoid yield reached 9.70 g and 5260 g / 100 g of substrate, respectively. Without supplementation only 5.36g dry sclerotia and 2149g carotenoid / 100 g of substrate was attained. © Rapid Science Ltd. 1998     

Enantiomeric Polyketides from the Starfish‐Derived Symbiotic Fungus Penicillium sp. GGF16‐1‐2

One new racemic mixture, penicilliode A ( 1 ) and four pairs of enantiomeric polyketides, penicilliode B and C ( 2 and 3 ) and coniochaetone B and C ( 4 and 5 ), were obtained from the starfish‐derived symbiotic fungus Penicillium sp. GGF16‐1‐2. Interestingly, the strain GGF16‐1‐2 can produce enantiomers. The absolute configuration of 1 was determined by X‐ray diffraction (XRD) analysis, and the absolute configurations of 2 – 4 were determined by the optical rotation (OR) values and electronic circular dichroism (ECD) calculations. Compounds 1 – 5 were firstly isolated from the marine‐derived fungus Penicillium as racemates, and 2 – 5 were separated by HPLC with a chiral stationary phase. All the compounds were evaluated for their antibacterial, cytotoxic and inhibitory activities against PDE4D2.     

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。