Purification and Characterization of Three Alkaline Endopolygalacturonases from a Newly Isolated Bacillus gibsonii

A newly isolated Bacillus gibsonii, designated as S-2 (CGMCC1215), was cultivated for production of alkaline pectinases utilizing sugar beet pulp as growth substrate. Purification of three alkaline endopolygalacturonases (endoPGs) from the crude pectinases extract was carried out by ultra-filtration, ammonium sulphate fractionation and ion-exchange chromatography, and their enzyme activities characterized. The three purified alkaline endoPGs, designated as S-I, S-II, and S-III, had a molecular weight about 38 kDa as determined by SDS-PAGE. The Km value and optimal temperature for optimal enzyme activities of S-I, S-II and S-III were 1.2 mg/mL and 60℃, 0.9 mg/mL and 55℃, 1.1 mg/mL and 60℃, respectively. Their best performances were given at an optimal pH 10.5, and sodium polygalacturonate was found to be the best substrate. The isoelectric points of S-I, S-II and S-III were 5.4, 7.4, and 8.2, respectively. Surfactants of Tween-80 and Tween-20 and metal ions such as Mg2+ and Ca2+ stimulated the activity of S-I, S-II and S-III, whereas S-III was inhibited by Ca2+, and Mn2+ and Zn2+ ions inhibited the activity of the three enzymes.     

Fractionation of Lignocellulosic Materials for the Biorefinery: Separation and Characterization of Lignin from Calamagrostis angustifolia Kom

Dewaxed Calamagrostis angustifolia Kom was pretreated with hot water at 60 and 90°C for 3 h, and then the residue obtained was successively treated with 70% ethanol, and 70% ethanol containing 0.2%, 1.0%, 2.0%, 4.0%, and 8.0% NaOH at 80°C for 3 h. The dissolved components were subjected to further separation to get eight lignin fractions, which were characterized by gel permeation chromatography, Fourier transform infrared, and sugar analysis. All the lignin fractions had small weight-average molecular weights between 810 and 2580 g/mol. Two typical lignins, L₃ (prepared with 70% ethanol) and L₅ (prepared with 70% ethanol containing 1.0% NaOH), were further analyzed using ¹H, ¹³C NMR and HSQC spectroscopy. Signals from guaiacyl (G), syringyl (S), and p-hydroxyphenyl (H) units observed in aromatic/olefinic region of HSQC spectra indicated that the lignin from Calamagrostis angustifolia Kom could be classified as “GSH” lignin. In aliphatic-oxygenated region, β-O-4′ together with small amounts of β-5′, β-β′, and p-hydroxycinnamyl alcohol end group were the main interunit linkages observed. Aqueous ethanol, which could avoid the cleavage of ether bonds in lignin at neutral condition, was more effective than water on lignin extraction.     

魔芋生物碱的初步分离及结构表征

研究了用酸性醇提取魔芋中生物碱的方法,通过氧化铝干柱层析法初步分离出了魔芋生物碱样品Ⅰ、Ⅱ和Ⅲ,样品Ⅰ的结构经1H NMR、IR和MS表征。     

巴斯德梭菌甘油脱水酶基因的克隆表达、纯化及酶学性质的研究

用聚合酶链反应(PCR)技术,从巴斯德梭菌(C lostridium pasteurianum)扩增出甘油脱水酶基因(dhaBCE),在大肠杆菌中高效表达,SDS-PAGE(聚丙烯酰胺凝胶)电泳结果显示,分别有相对分子质量为66、21、16 kD三条特异性蛋白表达条带。用金属镍亲和层析及S-300H凝胶层析将重组蛋白进行分离纯化,所得纯酶的比活为4.26 U/mg。重组酶的最适反应温度42~44℃,最适作用pH=9.10~9.50,它对三个底物结构类似物的米氏常数(Km)值分别是:1,2-丙二醇0.38 mmol/L,甘油0.29 mmol/L,1,2-乙二醇2.0 mmol/L;对辅酶B12的Km值为0.22μmol/L。     

Separation and Purification of Polyhydroxyalkanoates from Newly Isolated Comamonas sp. EB172 by Simple Digestion with Sodium Hydroxide

A simple, mild, and effective process for the recovery of intracellular polyhydroxyalkanoate from a newly isolated gram-negative wild-type bacteria Comamonas sp. EB172 was developed using sodium hydroxide. Various parameters such as sodium hydroxide concentration, digestion time, and temperature were examined for their effect on polyhydroxyalkanoate recovery. The results showed that polyhydroxyalkanoate with 88.6% purity and 96.8% recovery yield were obtained by incubating the dried cells with 0.05 M sodium hydroxide at 4°C for 1 h, followed by purification steps using ethanol and water. Removal of non-polymeric cellular materials from the Comamonas sp. EB172 was increased under alkaline solution as a result of enhanced cell wall permeability. In addition, the presence of glycerol in the polymer suspension proved that saponification of the lipid layer in the bacterial cell wall occurred due to sodium hydroxide reaction.     

海洋青霉菌壳聚糖酶的分离纯化与性能

以从福建崇武海边的海螺、贝壳及其附着物中经过筛选分离得到海洋青霉菌(marine Penicillium oxalicum)为出发菌株,对固态发酵海洋青霉菌所产壳聚糖酶进行分离提取工艺条件的探索.分别采用不同相对分子质量的PEG(相对分子质量分别为1000,4000,6000,10000,20000)、AB-8大孔树脂...     

Purification and Characterization of Iso-Ribonucleases from a Novel Thermophilic Fungus

A thermophilic fungus previously isolated from composted horse manure was found to produce extracellular iso-RNases that were purified 127.6-fold using a combination of size exclusion chromatography and a novel affinity membrane purification system. The extent of purification was determined electrophoretically using 4%–15% gradient polyacrylamide gels. RNase activity was dependent on the presence of a metal co-factor with significantly more activity with Zn²⁺ or Mn²⁺ than Mg²⁺. The RNases exhibited maximum activity at both pH 3.0 and pH 7.0 with no activity at pH 2.0 or 10.0. The optimal temperature for the iso-RNase was 70 °C. The molecular weight of the iso-RNase was determined to be 69 kDa using a Sephadex G-75 column.     

姜黄色素3种主要成分的分离纯化研究进展

近年来,针对从姜黄中提取分离得到姜黄素的研究综述文章很多,但对分离得到3种单体姜黄色素的综述还未见报道。本文就近年来文献报道的对3种姜黄色素:姜黄素、脱甲氧基姜黄素、双脱甲氧基姜黄素的分离纯化研究作综述。     

Biosorption of Reactive Yellow 145 Dye by Dried Penicillum restrictum: Isotherm,Kinetic, and Thermodynamic Studies

Removal of dyes from wastewaters causes a big concern from the environmental point of view due to their extreme toxicity towards aquatic life and humans. Commonly used traditional methods to treat these effluents are ineffective because dyes show resistance to many chemicals, oxidizing agents, and light. In this context, the biosorption process has attracted great attention in recent years since they utilize not only cheap plant materials but also a wide variety of microorganisms as biosorbing agents, displaying a high dye-binding capacity. In this study, biosorption potential of dried Penicillum restrictum (DPR) for Reactive Yellow 145 (RY 145) was studied with respect to pH, equilibrium time, and temperature to determine equilibrium and kinetic models. The most suitable pH and equilibrium time were determined as 1.0 ± 0.05 and 75 min respectively, at a biomass dosage of 0.4 mg L^?1 and 20 ± 0.5°C. Data obtained from batch studies fitted well with the Dubinin-Radushkevich (D-R) followed by the Freundlich and Langmuir isotherm models. Maximum uptake capacities (qm) of DPR for the dyestuff (RY 145) were 109.7, 115.2, and 116.5 mg g^?1 biomass at temperatures of 20, 30, and 40 ± 0.5°C, respectively. The overall biosorption process was best described by the pseudo-second-order kinetic model. Gibbs free energy changes were calculated as ?384.6, ?273.5, and ?245.9 J mol L^?1 at 20, 30, and 40 ± 0.5°C, respectively.     

大肠杆菌L- 天冬酰胺酶的分离纯化及其特性

目的 建立简单有效的L- 天冬酰胺酶分离纯化工艺，研究L-天冬酰胺酶的特性。方法 采用冷丙酮破壁处理，经稀碱液抽提、离子交换层析和亲和层析进行纯化。通过温度试验、pH试验、SDS－PAGE图谱、紫外吸收光谱等研究酶的特性。结果 建立了简单高效的纯化工艺，可使L-天冬酰胺酶的纯度达94．4％，比活为520u/mg蛋白，总收率为65．7％。L-天冬酰胺酶的相对分子质量分别为143900和166700，均为有抗癌活性的EC－2组分，在270nm有最大吸收。最适作用温度为40－50℃，最适作用州为8.0-9.0在40℃以下能保持稳定。结论建立了大肠杆菌L-天冬酰胺酶的分离纯化工艺，并研究了L-天冬酰胺酶的酶学特性。     

Production, Purification and Partial Characterization of Four Lipases from a Thermophile Isolated from Deception Island

Four lipases were purified from ID17, a thermophilic bacterium belonging to Geobacillus genus isolated from Deception Island, Antarctica. Lipase activity was detected by opacity test and p-nitrophenyl laurate methods. Lipase production was better in a medium containing tryptone as the carbon and nitrogen source, without non-ionic detergents and pH 7.5. Proteins were ultrafiltered from supernatant and separated using anion exchange and size exclusion chromatography resulting in four distinct fractions with lipase activity (called Lip1–4). Purified lipases showed an optimal pH at 9.0, 9.5, 10.0 and 8.0 and temperature at 65, 70, 75 and 80 °C for Lip1–4, respectively. Lip1 and Lip2 showed higher activity using p-nitrophenol decanoate as substrate, whereas Lip3 and Lip4 prefer p-nitrophenol laurate. Based on their molecular weight Lip1 and Lip2 are trimeric and pentameric proteins, respectively, whereas Lip3 and Lip4 are monomeric proteins. Lip1 was exceptionally thermostable maintaining 70 % of its activity after incubating it at 70 °C for 8 h. Based on their characteristics, the four lipases obtained from ID17 are good candidates to understand the mechanisms of lipase stability and to be used in different types of industrial applications.     

煤焦油的净化、分离及应用浅析

阐述了国内外近年来煤焦油加工的研究状况,列举了我国煤焦油的净化技术、分离技术及其应用;并指出煤焦油的研究应注重系统化和可行性,对促进我国煤焦油加工技术的发展具有参考价值。     

异丙醇生产副产物中异丙醚的分离精制技术

采用萃取精馏和共沸精馏的试验方法,从异丙醇装置副产品中分离精制异丙醚．结果表明,以甘油为萃取剂,甲醇为夹带荆,可分别得到质量分数大于99％的异丙醚和质量分数大于99．5％的异丙醇,异丙醚收率为70．35％,具有明显的经济效益。     

烟草过氧化物酶(TOPI)的分离与纯化的研究

A new isoenzyme of tobacco peroxidase(TOP) I was purified from tobacco (K326) by using acetone powder, ammonium sulfate precipitation and column chromatography on DEAF-52 cellulose, Sephadex G-75 and DEAE-Sephadex A-50. It is an iron-protein containing haemachrome, whose molecular weight is 21888.5 and the isoelectric point is 3.5. The optimum pH value and temperature of this enzyme is 6.0 and 45℃ respectively. The enzyme is stable in the pH range from 3.0 to 10.0 and has a favorable thermostability.     

甘草甙的分离纯化及鉴定

甘草甙是甘草黄酮类化合物中重要的单体活性成分,具有抗氧化、抗H IV等多种药理作用。该文在无标样条件下对甘草甙的分离纯化进行了研究,并首次采用模拟移动床色谱技术提纯甘草甙。实验表明,原料液固萃取的最佳条件为:水作萃取溶剂,原料质量浓度为40 g/L,90℃恒温水浴中萃取2.0 h。模拟移动床色谱在进样流速0.1 mL/m in;洗脱流速1.5 mL/m in;萃取流速1 mL/m in;切换时间20~21 m in;样品质量浓度0.2 g/mL;流动相V(乙醇)∶V(水)=15∶85;工作模式:1-1-2条件下(即三带模拟移动床色谱中洗脱带、精馏带、吸附带色谱柱数分别为1,1,2),可实现甘草甙的精细分离。重结晶所得产品通过质谱、紫外可见光谱、红外及核磁共振谱图对其结构进行了表征。     

猪肺肝素钠提取工艺优化、分离与初步鉴定

该文研究了中心组合设计和响应面分析优化猪肺肝素钠的提取工艺,通过D-254树脂交换吸附、沉淀剂除蛋白质、乙醇沉淀以及DEAE-Sephasoe fast flow离子交换柱层析纯化得到高纯度的猪肺肝素钠,最后采用琼脂糖凝胶电泳、红外光谱、紫外光谱和核磁共振波谱等对其进行初步鉴定。猪肺肝素钠的最佳提取工艺参数为:提取温度53℃,时间4 h,pH=8.5。粗品和纯化后产品的提取率分别为0.81%和0.05%,羊血浆法测定粗品和纯化后产品效价分别为6.5 U/mg和108.3 U/mg。红外光谱显示,纯化后样品在890 cm-1和940 cm-1处出现肝素钠特征吸收峰,紫外光谱最大吸收波长为205 nm,结合琼脂糖凝胶电泳和核磁共振波谱证明用上述方法制备的纯化后的样品为肝素钠。     

草酸青霉Penicillium oxalicum菌体吸附活性染料的性能及微观过程

研究了草酸青霉(Penicillium oxalicum)在模拟染料废水中以边生长边吸附的方式对活性翠蓝KN-G, M-GB, K-GL,活性黑K-BR、活性艳蓝M-BR、活性红紫K-3R和活性深蓝K-R 7种水溶性活性染料的脱色性能及吸附过程. 结果显示,生长菌体对7种活性染料具有良好的脱色性能,染料初始浓度为200 mg/L时,平均脱色率可达93.0%;染料初始浓度为400 mg/L时,活性翠蓝KN-G和M-GB的脱色率仍达到了99.7%和99.9%. 上清液的紫外光谱图及染料分子中铜离子浓度的检测结果表明,染料通过吸附方式从废水中去除. 通过SEM, TEM观察发现,生长菌体在吸附过程中,菌丝发生肿胀膨大,细胞壁发生结构重组,厚度增加10~15倍. 细胞壁的结构变化是生长菌体吸附染料的重要机制,为染料吸附提供位点和进入细胞内部的通道.     

生物油性质及分离纯化的研究进展

生物质通过热化学转化可获得一种多相复杂的液体产品——生物油,其中包括了数百种有机化合物和功能化学品。本文以生物质热化学转化生物油的理化性质为基础,针对国内外生物油分离纯化技术的研究现状进行系统评述,主要侧重于蒸馏、溶剂分离、超临界CO2萃取技术以及色谱分离等相关技术,同时指出柱层析和溶剂分离技术相结合可以对生物油中的复杂成分进行有效的分离,并能得到高纯度的化学品,其在生物油的分离方面具有较好的发展前景。     

经亚氯酸钠和白腐菌处理后蔗渣半纤维素的分离

蔗渣经亚氯酸钠脱木素后,木素含量降到了3.4%,木素脱除率达到85.0%,抽提出的半纤维素纯度达到84.8%;将亚氯酸钠和白腐菌处理结合起来,能将蔗渣木素脱除92.1%,抽提的半纤维素纯度达到87.4%,聚戊糖抽提率也有较大提高。     

2,6-二异丙基萘的分离精制

研究了从萘与丙烯合成的异丙基萘混合物中分离、精制2,6 二异丙基萘(2,6 DIPN)的方法. 考察了精馏操作条件对精馏分离结果的影响,并研究了结晶温度及溶剂对产品纯度和收率的影响. 实验结果表明,以自行合成的烷基化产品混合物为粗产品,在1000 Pa真空度下减压精馏,收集165℃的馏出物可得到含量约50 的2,6 DIPN,将此馏分在9℃下结晶,所得晶体以乙醇为溶剂洗涤2次后,可精制出纯度为99.42 的2,6 DIPN.