利用双水相萃取技术分离纯化尿激酶

通过研究双水相萃取系统的各种影响因素：乙醇／（NH4）2SO4的组成比例、pH值、无机盐的加入、粗酶的浓度等,探索以乙醇／（NH4）2SO4组成的双水相萃取体系纯化尿激酶的最佳条件。建立以乙醇／（NH4）2SO4组成的双水相萃取体系分离纯化尿激酶的新途径。结果表明：双水相萃取系统冰乙醇浓度为65％,（NH4）2SO4浓度为10．0％,pH8．0,酶加入量为30％,且不加入任何其他无机盐的条件下,尿激酶的纯化倍数可达到9．2倍,回收率最高达92％。     

双水相体系萃取人参根中人参皂苷的研究

建立稳定的聚乙二醇(PEG)与(NH4)2SO4双水相体系以分离人参根中人参皂苷。通过上下相体积比(R)、分配系数(K)和回收率(Y)分析双水相体系对人参皂苷的萃取效果,研究了PEG分子量、PEG/(NH4)2SO4质量分数、pH值和温度等因素对双水相成相及人参皂苷萃取的影响。结果表明:PEG分子量为3350、PEG3350的质量分数为12%、(NH4)2SO4质量分数为16%、溶液pH为7.0、温度为60℃时,双水相体系对人参皂苷有较高的萃取率,回收率可到达88.94%。     

白地霉Cryytococcus neoformans脂肪酶的双水相萃取

本文研究了不同无机盐的双水相体系对白地霉脂肪酶的萃取分离效果,对PEG/（NH4）2SO4成相系统进行了系统的研究,通过考察体系PEG分子量、不同的无机盐、PEG浓度、（NH4）2SO4浓度、离子强度、pH值及（NH4）2SO4浓度对反萃取的影响,并通过正交实验进一步优化了实验条件,初步确定在PEG浓度15%,（NH4）2SO4浓度22.5%,pH8.0,不加NaCl的条件下进行双水相萃取,脂肪酶分离系数和纯化倍数分别为6.8和7.5,比活力达到40.3 U/mg蛋白。     

双水相萃取法分离竹叶黄酮的工艺研究

以竹叶黄酮水提溶液为原料,采用PEG(聚乙二醇)/(NH4)2SO4双水相体系对竹叶黄酮进行萃取,考察了PEG平均相对分子质量、PEG质量分数、(NH4)2SO4质量分数、pH值、NaCl质量分数、原液质量分数、萃取温度等对双水相及竹叶黄酮萃取效果的影响。双水相萃取法提取竹叶黄酮的最优条件为:PEG 400 31%,(NH4)2SO411%,pH 3.9,NaCl 0.7%,原液51.5%,萃取温度20℃,在此条件下得到的竹叶黄酮萃取率为97.8%。结果说明,双水相萃取法操作简单方便,成本低,不会引起生物质失活或变性,适合于黄酮类化合物的萃取分离。     

Separation of Flavonoids from Bamboo Leaves by Aqueous Two-Phase Extraction

以竹叶黄酮水提溶液为原料,采用PEG(聚乙二醇)/(NH4 )2SO4双水相体系对竹叶黄酮进行萃取,考察了PEG平均相对分子质量、PEG质量分数、(NH4)2SO4质量分数、pH值、NaCl质量分数、原液质量分数、萃取温度等对双水相及竹叶黄酮萃取效果的影响.双水相萃取法提取竹叶黄酮的最优条件为:PEG 400 31％,(NH4)2 SO4 11％,pH3.9,NaCl 0.7％,原液51.5％,萃取温度20℃,在此条件下得到的竹叶黄酮萃取率为97.8％.结果说明,双水相萃取法操作简单方便,成本低,不会引起生物质失活或变性,适合于黄酮类化合物的萃取分离.     

Geotrichum sp．SYBC WU-3脂肪酶的双水相萃取和酶学性质

初步研究双水相体系对Geotrichum sp．SYBC WU-3脂肪酶的萃取分离效果,选用PEC4000／NaH2 PO4作为戍相系统进行系统研究,考察影响脂肪酶萃取的各种因素（如PEG相对分子质量及质量分数、NaH2PO4质量浓度、pH）,并采用正交实验进一步优化实验条件,确定双水相萃取体系为PEG质量分数为30％、NaH2PO4质量分数为20％、体系pH为6,在此条件下Geotrichum sp．SYBC WU-3脂肪酶经硫酸铵沉淀和双水相萃取两步纯化的纯化倍数达到最大,较Geotrichum sp．SYBC WU-3脂肪酶粗酶纯化了22倍。Geotrichum sp．SYBC WU-3脂肪酶纯酶为低温碱性脂肪酶,最适反应温度为15oC,最适pH为9．5,相对分子质量为3．58×10^4。     

双水相萃取技术在白藜芦醇提纯工艺中的应用

论文研究了双水相萃取体系在提纯白藜芦醇工艺中的应用,用乙醇-硫酸胺-水双水相体系使虎杖提取液中的各物质按极性不同在油水两相中得到分离。双水相萃取时,白藜芦醇的含量远高于有机溶剂萃取,达34.29%。可见双水相技术可以完全代替有机溶剂萃取技术提纯白藜芦醇,产品纯度高,具有工艺简单,毒性小,成本低等优点。     

聚乙二醇共沉淀物复溶萃取辣根过氧化物酶的条件优化

为了改进辣根过氧化物酶的提纯方法,在双水相萃取的基础上使用聚乙二醇(PEG6000)在高饱和度(NH4)2SO4中沉淀辣根提取液中的过氧化物酶,利用磷酸盐溶液复溶解共沉淀物形成的双水相萃取体系能高效回收高纯度酶蛋白。研究了pH、PEG浓度和(NH4)2SO4饱和度对酶活性的影响,并考察不同液固比、pH和NaCl浓度对目标酶在双水相体系中的分配行为,并通过响应面法优化出最优萃取条件。结果表明:在液固比0.3 m L/g、pH7.02和NaCl 42 g/L的优化萃取条件下,辣根过氧化物酶回收率达88.1%,酶纯度较优化前提高了21.7倍。该方法的建立对于微量蛋白质的高效率提纯具有重量的参考价值。     

荧光素酶在PEG／盐系统中分配系数的研究

海洋发光细菌产生的荧光素酶在PEG／盐双水相系统中的分配系数(K)的对数值与PEG分子量成线性关系。低分子量PEG亲水性较强,有利于酶分配于PEG上相中,而高分子量PEG疏水性较强,有利于酶分配在下相盐溶液中,PEG／三价盐与PEG／二价盐系统不同,前者有一折点。在使用单一分子量PEG组成的PEG／硫酸铵系统中,K值随系统中使用的硫酸铵浓度增加而增加,硫酸铵达到一定浓度后,其浓度继续增加对K值的增加影响减小;但在两种不同分子量的PEG混合物组成的PEG／硫酸铵系统中,K值随硫酸铵浓度的增加有一最小值,在此硫酸铵浓度之外,增加或减小硫酸铵浓度,K值均会增加。在PEG／硫酸铵系统中,荧光素酶的K值随系统的pH增加略有提高,但变化不大。而在PEG／磷酸盐系统中,在pH6．4时,K值最小,pH高或低于此值,K值均增加。系统的温度变化对K值影响很小。     

1-辛基-3-甲基咪唑二氰铵盐/硫酸铵双水相体系分离纯化银杏黄酮

建立了由亲水性离子液体1-辛基-3-甲基咪唑二氰铵盐([C8mim][N(CN)2])和(NH4)2SO4形成的双水相萃取体系并应用于银杏黄酮的分离纯化研究。研究了盐浓度、体系温度、pH值、NaCl量等因素对银杏黄酮萃取效率的影响;并对下相中无机盐进行回收。体系由18.52%[C8mim][N(CN)2],25.93%(NH4)2SO4构成,加入1.5 mmol NaCl,在室温下进行萃取时萃取效率最佳,在最佳的条件下[C8mim][N(CN)2]/(NH4)2SO4体系对银杏黄酮的萃取效率达96.73%。与传统的双水相体系相比,该体系的萃取效率高,粘度低,同时(NH4)2SO4的回收率达90.54%。[C8mim][N(CN)2]/(NH4)2SO4双水相体系是一种很好的分离纯化银杏黄酮的方法。     

Selection and optimization of an aqueous two‐phase system for the recovery of 1,3‐propandiol from fermentation broth

In this study a suitable alcohol/salt aqueous two‐phase (ATP) system was selected for the recovery of 1,3‐propandiol (1,3‐PD) from fermentation broth. From the different alcohol/salt systems studied the ethanol and dipotassium hydrogen phosphate ATP system appeared to be favorable. To examine the potential of this ATP system the partition coefficient of 1,3‐PD in synthetic solutions was first optimized with the response surface methodology. The parameters studied were concentrations of ethanol (21.99–38.81% w/w), dipotassium hydrogen phosphate (14.99–31.81% w/w) and 1,3‐PD (6.36–73.64 g/L). The optimum conditions were found to be 35.39% w/w for ethanol, 28.40% w/w for dipotassium hydrogen phosphate and 73.6 g/L for 1,3‐PD. Under these conditions the maximum partition coefficient of 1,3‐PD and the extraction yield were determined as 23.14 and 97.82%, respectively. The optimum extraction conditions were then used to guide the recovery of 1,3‐PD from a real fermentation broth. The partition coefficient and extraction yield of 1,3‐PD reached 20.28–97.20% in this case, respectively. A favorable partition of the organic acids lactate, acetate and butyrate in the bottom phase was also achieved. We have also studied the removal of cells and macromolecules from the broth. Removal ratio of cells and proteins were 96.47 and 93.05%, respectively. Thus, the ethanol/dipotassium hydrogen phosphate ATP system appears to be an interesting alternative or can be used as one useful step in the downstream processing of 1,3‐PD from fermentation broth.     

Elimination of overgrowth in delayed-incubation membrane filter test for total coliforms by m-ST holding medium.

Two holding media were compared for their effects on total coliform recovery by the delayed-incubation membrane filter procedure. LES-MF holding medium contains tryptone, m-Endo broth, dipotassium hydrogen phosphate, sodium benzoate, sulfanilamide, para-aminobenzoic acid, and cycloheximide (pH 7.0). m-ST holding medium contains ethanol, sodium monophosphate, dipotassium hydrogen phosphate, sulfanilamide, and Tris (pH 8.6). In tests with natural water and wastewater samples from various sources, recovery with LES-MF and m-ST were similar after a 1-day holding period. With LES-MF, however, after a 2- or 3-day holding period, coliform bacteria frequently were partially or totally overgrown by noncoliforms, causing significant reductions in coliform counts. No significant overgrowth was observed with m-ST. We propose that m-ST be used for all holding periods longer than 1 day.     

Continuous countercurrent salting-out extraction of 1,3-propanediol from fermentation broth in a packed column

The possibility of continuous extraction of 1,3-propanediol in a experimental packed column was investigated using a salting-out extraction system of dipotassium phosphate/ethanol. Mass transfer of 1,3-propanediol takes place from the dispersed phase (salt-rich solution) to the continuous phase (ethanol). The influences of flow rate of dispersed phase and size of packing material on partition coefficient and recovery of 1,3-propanediol were investigated and the results were compared with those obtained in spray column and test tube. Furthermore, the influences of various system compositions on hold up of dispersed phase, mass transfer coefficient, and system stability were also studied in the column packed by stainless steel Dixon 3 × 3 mm. It was found that the packed column showed a good extraction efficiency and stability. Besides, 1,3-propanediol recovery of 90.30% was obtained during a 11 h continuous operation when the real fermentation broth was used. At the same time, 94.4% of phosphate could be recovered when 0.2 volume of anhydrous ethanol was added into the raffinate phase at pH 4.0.     

Elimination of overgrowth in delayed-incubation membrane filter test for total coliforms by m-ST holding medium

Two holding media were compared for their effects on total coliform recovery by the delayed-incubation membrane filter procedure. LES-MF holding medium contains tryptone, m-Endo broth, dipotassium hydrogen phosphate, sodium benzoate, sulfanilamide, para-aminobenzoic acid, and cycloheximide (pH 7.0). m-ST holding medium contains ethanol, sodium monophosphate, dipotassium hydrogen phosphate, sulfanilamide, and Tris (pH 8.6). In tests with natural water and wastewater samples from various sources, recovery with LES-MF and m-ST were similar after a 1-day holding period. With LES-MF, however, after a 2- or 3-day holding period, coliform bacteria frequently were partially or totally overgrown by noncoliforms, causing significant reductions in coliform counts. No significant overgrowth was observed with m-ST. We propose that m-ST be used for all holding periods longer than 1 day.     

Aqueous two-phase extraction of 2,3-butanediol from fermentation broths using an ethanol/phosphate system

Separation of 2,3-butanediol from the complex fermentation broths is a difficult task and becomes a bottleneck in industrial production. Aqueous two-phase systems composed of hydrophilic solvents and inorganic salts could be used to extract 2,3-butanediol from fermentation broths. Aqueous two-phase extraction of 2,3-butanediol from fermentation broths was studied by ethanol and dipotassium hydrogen phosphate system. The influences of phase composition on partition of 2,3-butanediol, removal of cells and biomacromolecules were investigated. The partition coefficient and recovery of 2,3-butanediol reached up to 28.34 and 98.13%, respectively, and the selective coefficient of 2,3-butanediol to glucose was 615.87 when the system was composed of 24% (w/w) ethanol and 25% (w/w) dipotassium hydrogen phosphate. Simultaneously, cells and proteins could be removed from the fermentation broths and the removal ratio reached 99.63 and 85.9%, respectively. This process is convenient and economic, furthermore, the operation is easy to scale-up, that is, this method provides a new possibility for the separation and refining of 2,3-butanediol.     

蓝莓提取物对副溶血性弧菌的抑制作用

本文以北高丛蓝莓中的栽培品种埃利奥特为原料,通过L16(43)正交实验,分析比较了16种提取物对副溶血性弧菌的抑菌效果及各提取物的pH值.结果表明:在30℃下,以65％乙醇(w/w)为溶剂,超声提取25min为最优提取条件.采用此条件得到的提取物具有最低的pH值,在浓度为100和40 mg/mL时,对副溶血性弧菌的抑菌率分别达100％和81.3％.     

微波萃取苦荞壳中总黄酮工艺研究

本文在提取苦荞壳中总黄酮的工艺实验中,优化了提取溶剂的种类,水和乙醇,同时还使用了超声波和微波辅助提取,并分别将不同因素,如溶剂种类及浓度,提取时间,温度,pH,液固比等,对提取结果的影响作了比较,结果表明,当介质为乙醇时的微波萃取效果明显优于其他提取方法。     

Extraction behavior of caffeine and EGCG from green and black tea

A dipping method was developed to extract the catechins (EGCG) and alkaloids (caffeine) from green tea (Korea) and black tea (Sri Lanka). The effects of the solvent composition (water vs. ethanol), extraction time, temperatures, and solvent pH on the amount of catechins (EGCG) and alkaloids (caffeine) extracted from green and black tea were investigated. Reversedphase high-performance liquid chromatography (RP-HPLC) was used to analyze the catechins (EGCG) and alkaloids (caffeine) extracted. The content of EGCG and caffeine in green tea extracts was in the range of 2.04∼0.30 and 10.22∼0.85 mg/g, respectively. The amount of EGCG and caffeine in black tea extracts was in the range of 0.32∼0.24 and 5.26∼1.01 mg/g, respectively. The amount of caffeine extracted from green and black tea was greater than the amount of EGCG. Pure water is the best solvent for extracting EGCG and caffeine from green tea. The amount of caffeine extracted from green and black tea increased as the temperature, extraction time, and hydrogen ion concentration of the solvent increased. Although the amount of EGCG extracted from green tea increased as the temperature increased, the amount of EGCG extracted from black tea was not affected by temperature. The extraction of EGCG from both green and black tea was not affected by the hydrogen ion concentration of the solvent.     

Ultrasonication assistance increases the efficiency of isoflavones extraction from kudzu (Pueraria lobata Ohwi) roots waste

This study assesses the use of ultrasonication to improve the extraction process of classical solvent extraction methods for extracting isoflavones from the kudzu roots waste. The kudzu roots waste was produced after squeezing fresh kudzu roots to make juice. The effects of extraction time, extraction temperature, ultrasonic power, and ethanol concentration in ethanol/water mixtures were investigated. The extraction yield was found to increase with extraction time and temperature. The application of ultrasonication-assisted extraction (UAE) increased the extraction yield of water/ethanol mixture (20:80) at 25°C 3 fold. A maximum amount (7.28 g) of isoflavone was obtained from 100 g of dried kudzu roots waste by UAE with water/ethanol mixture (20:80) for 6 h at 80°C. Combining the use of ultrasonication with conventional vacuum evaporation method also reduced the concentration time for extracts from 45 to 24 min.     

Non-covalent interaction between procyanidins and apple cell wall material: Part I. Effect of some environmental parameters

The adsorption of procyanidins on cell wall material were quantified by bringing into contact a solution of procyanidins and a suspension of cell wall material. The influence of structural features such as degree of polymerisation (DP) and percentage of galloylation (% gall), and of physico-chemical parameters such as pH, ionic strength, temperature and presence of ethanol were investigated. The amount of procyanidins bound to the cell wall increased with the DP, the % gall, and the proportion of (+)-catechin, the last indicating an effect of the stereochemistry of the flavan-3-ols. Complex formation between procyanidins and cell wall material was not affected by pH in the range 2.2-7 but it was decreased by urea, dioxane and ethanol. Adsorption increased with increasing ionic strength and decreased with increasing temperature. This indicated that the bonds which governed the interaction between procyanidins and cell wall material were weak energy bonds of the type hydrogen bond and hydrophobic interaction.