聚丙烯酰胺-聚乙二醇对L-天门冬酰胺酶的包埋研究

以聚丙烯酰胺，聚乙二醇为载体制备了固定化L-天门冬酰胺酶。对固定化酶载体材料——凝胶的溶胀比、机械强度及其影响因素进行了研究。结果表明，单体及交联剂浓度是影响凝胶性能的重要因素，聚乙二醇作为稀释剂能有效控制聚合速率，提高凝胶的溶涨性能和韧性。此外考察了酶负载量、pH值和温度等因素对固定化酶活性的影响。实验结果还表明分批反应6次或存储42天后，固定化酶仍保持较高的活力。     

超氧化物歧化酶的固定化及其酶学性质

目的制备固定化超氧化物歧化酶(SOD),并分析其酶学性质。方法以壳聚糖为载体,戊二醛为交联剂,制备固定化超氧化物歧化酶。以邻苯三酚自氧化法分别测定溶液酶与固定化酶的活力,并计算回收率。对固定化酶的温度和pH稳定性、半衰期、重复使用的回收率及米氏常数(Km)进行测定。结果固定化超氧化物歧化酶活力为333U/g,酶活回收率为86.32%,半衰期为43.8d;固定化酶室温保存5d后,相对酶活力仍保持在80%以上,最适反应温度为45℃,使用一次后回收率为70.12%,重复使用两次后回收率为51.72%;固定化酶与溶液酶在pH6时活性最强,Km分别为0.18mmol/L和0.16mmol/L。结论该固定化酶较溶液酶的稳定性得到提高,便于贮存,在食品、药品、日用品等领域有良好的应用前景。     

分子筛固定氨基酰化酶Ⅰ的研究

以Y型分子筛为载体,分别采用吸附法、吸附-戊二醛交联法、偶联法和偶联-重氮法4种方法对氨基酰化酶I进行固定化。 研究发现偶联法获得的分子筛固定化酶的活性最高,达0.258 IUmg-1,并对分子筛固定化酶最适pH值、最适反应温度、重复使用性和米氏常数进行了测定;结果表明该载体适合氨基酰化酶I的固定,可以获得较高的酶活力,分子筛固定化酶的最适pH值、最适反应温度都有所拓宽。     

分子筛固定氨基酰化酶I的研究

以Y型分子筛为载体,分别采用吸附法、吸附-戊二醛交联法、偶联法和偶联-重氮法4种方法对氨基酰化酶I进行固定化。 研究发现偶联法获得的分子筛固定化酶的活性最高,达0.258 IUmg-1,并对分子筛固定化酶最适pH值、最适反应温度、重复使用性和米氏常数进行了测定;结果表明该载体适合氨基酰化酶I的固定,可以获得较高的酶活力,分子筛固定化酶的最适pH值、最适反应温度都有所拓宽。     

大孔树脂D380固定化硫酸软骨素裂解酶

目的以大孔树脂D380为载体,戊二醛为交联剂,进行硫酸软骨素裂解酶(ChSase)的固定化,并考察固定化酶的酶学性质。方法分别考察加酶量、吸附温度、吸附时间、吸附pH值、戊二醛交联浓度、交联时间及交联温度对ChSase固定化效果的影响,并分析该固定化酶的最适反应温度、最适反应pH值、米氏常数(Km)及其操作稳定性。结果ChSase的最佳固定化条件为:加酶量150U/g树脂,吸附温度15℃,吸附时间6h,吸附pH值7.0,戊二醛交联浓度0.01%,交联时间3h,交联温度4℃。以此条件制备的固定化酶,其酶结合效率可达79.1%。该固定化ChSase的最适反应温度为45℃;最适反应pH值为7.0;Km达1.46×10-1g/L,较游离酶高;具有较好的操作稳定性。结论以大孔树脂D380为载体固定化ChSase是可行的,所得固定化酶有较高的使用效率和稳定性,适合于工业化生产。     

麦芽根中5′-磷酸二酯酶的分离提纯及其固定化

从麦芽根中分离得到了5′-磷酸二酯酶,采用戊二醛为交联剂,将该酶固定到壳聚糖上,用于酵母RNA的催化水解,以制备5′-核苷酸.研究了酶固定化反应的影响因素,在最适条件下,酶活回收率可达53.6%.进一步研究了固定化5′-磷酸二酯酶的酶学性质,测得固定化酶的米氏常数Km为15.38 mg/mL(以RNA为底物),固定化酶催化水解RNA的最适温度为75℃,最适pH值为5.5,实验发现固定化5′-磷酸二酯酶具有更好的耐热性和更高的纯度.     

固定化漆酶对二氯酚的脱氯作用

采用活性炭吸附与海藻酸钙凝胶包埋相结合的方法使Coriolus versicolor漆酶固定化.利用固定化漆酶对2,4-二氯酚进行脱氯反应,其最适pH值为4.5、最适温度为40℃.与游离酶相比,固定化酶反应的pH值和温度范围更宽,其稳定性得到了明显改善.使用柱式固定化酶反应器处理2,4-二氯酚,在批式反应工艺条件下,当底物浓度为1 mmol•L^-1、反应3～5 h, 2,4-二氯酚的去除率可达99.5%以上（脱除的氯离子浓度达0.5 mmol•L^-1）.连续8批反应的结果表明：固定化漆酶性能稳定、催化效率高,在环境污染废水治理方面具有良好的应用前景.     

重组酪氨酸酚裂解酶全细胞酶法合成L-酪氨酸

利用基因工程手段重组表达了弗氏柠檬酸杆菌来源的酪氨酸酚裂解酶（TPL）,以丙酮酸和L-丝氨酸为底物酶法合成L-酪氨酸,考察了pH、温度、表面活性剂、金属离子、铵盐种类和氯化铵浓度等因素对L-酪氨酸合成的影响,并比较了两种底物合成L-酪氨酸的转化率。结果表明,TPL的最适反应条件是45℃,pH 8.0,4 mmol/l PLP,氯化铵浓度350 mmol/l。1 mmol/l triton-x 100对TPL酶活有促进作用,金属离子无明显影响。质量浓度1%丙酮酸的转化率约是L-丝氨酸的1.8倍。     

腈水合酶固定化方法和催化特性的研究

以一种能够产生腈水合酶诺卡氏菌为研究对象,针对原来的海藻酸盐包埋法存在的固定化细胞强度较小、通透性较差等问题,改进了对酶的固定化方法,并对固定化腈水合酶的催化特性:最适反应温度、pH值、底物丙烯腈浓度、和表面活性剂性质和丙烯酰胺累积浓度对酶活性的影响等五个方面进行了研究。其中固定化腈水合酶最适反应温度在15~25°C;pH 7.0左右;底物丙烯腈浓度为3%~4%;Triton X-100对酶活性基本无影响,而Tween 80和Tween 60对酶活力有抑制作用;固定化腈水合酶在丙烯酰胺累积浓度为15%~20%之间时,酶活性较好。     

海藻酸钙凝胶固定葡萄糖氧化酶的研究

利用海藻酸钙凝胶颗粒固定葡萄糖氧化酶(GOD).确定了固定化条件,并考察了温度、pH值、储存时间对固定化酶和游离酶酶活力的影响,测定了酶活回收率.确定的固定化较优条件为:CaCl2 5.0%(质量体积比),海藻酸钠3.0%(质量体积比);固定化酶的最适催化温度为31℃、最适pH值为6.3,较游离酶分别提高7℃和0.6;固定化酶的平均酶活回收率为61.69%.此外,酶的储存稳定性能也有所提高,可重复多次使用.     

Optimization of L-Asparaginase Immobilization onto Calcium Alginate Beads

In this study, anti-leukemic enzyme L-asparaginase (E.C.3.5.1.1) from Escherichia coli ATCC 11303 was modified by the microencapsulation technique onto calcium alginate beads. Using response surface methodology (RSM), a three-level full factorial design, the values of concentration of sodium alginate, concentration of calcium chloride, and enzyme loading were investigated to obtain the highest residual L-asparaginase (L-ASNase) activity % (immobilized enzyme activity/free enzyme activity). The effects of the studied factors on immobilization were evaluated The predicted values by the model were close to the experimental values, indicating suitability of the model. The results presented that an increase in sodium alginate concentration increased the percent of residual activity of L-ASNase at any given calcium chloride concentration and the moderate amount of enzyme loading increased the percent residual activity. The optimal immobilization conditions were as follows: sodium alginate 1.98% (w/v), calcium chloride concentration 3.70% (w/v), and enzyme load 46.91% (v/v). The highest residual L-ASNase activity % obtained was 34.49%.     

海藻酸钠固定化S-腺苷甲硫氨酸合成酶的制备及其性质研究

研究了以海藻酸钠包埋法制备固定化S-腺苷甲硫氨酸（sAM）合成酶的条件,并考察了固定化酶的酶学性质。结果表明,最适固定化条件为：海藻酸钠质量分数3％、CaCl2质量分数4％、SAM合成酶加量（以每克海藻酸钠计）75mg、固定化时问30min,在此条件下,固定化酶活力回收率达到42％。固定化酶热稳定性较好,在50℃下保温5h仍保留69％的酶活力,而游离酶则完全失活;固定化酶在碱性条件下的稳定性较好,在pH值7．5～9．0的缓冲溶液中4℃下保温10h仍保留84％以上的酶活力;将固定化酶用于SAM的合成,连续反应5批次后,仍保留82％的酶活力。     

Production of fructooligosaccharides by immobilized mycelium of Aspergillus japonicus

β-Fructofuranosidase (EC 3.2.1.26) in Aspergillus japonicus mycelium was immobilized by entrapment in calcium alginate gel. After immobilization, the enzyme was active over a wider pH range, and had improved thermostability. The total amount of fructooligosaccharides produced by immobilized enzyme was similar to that produced by a free enzyme system. A packed-bed reactor was employed for production of fructooligosaccharides at 42°C using the immobilized enzyme. The reactions were continued for 35 days and only 17% of enzyme activity was lost during this period.     

海藻酸钠明胶协同固定化黑曲霉脂肪酶

以海藻酸钠明胶为复合载体,采用包埋法制备固定化黑曲霉脂肪酶,考察了海藻酸钠、明胶浓度等因子对固定化效果的影响,比较固定化酶和游离酶对温度、pH等条件的稳定性。结果表明,制备固定化黑曲霉脂肪酶的最优条件为:海藻酸钠、明胶浓度分别为1.25%和0.5%,CaC l2浓度为10%,给酶量为450 IU/g;固定化酶最适温度为35℃,最适pH为9.0,常见有机溶剂和金属离子对固定化酶的活力影响较小。     

Production of berberine in immobilized plant cell culture ofThalictrum rugosum

Thalictrum rugosum cells were immobilized in calcium alginate, where they continued to live with their biological activity. The immobilized living cells performed the production of berberine in both shake flasks and an airlift bioreactor. Berberine formation was growth associative and most of the berberine produced was stored intracellularly. Rapid hydrolysis of sucrose and preference of glucose over fructose during the growth stage was observed. Phosphate-deficient media increased berberine production and prevented the dissolution of alginate beads. The behavior of immobilized cells grown in an airlift reactor was compared with that of the corresponding shake flask culture with respect to growth and berberine production. The rate of cell growth and berberine production in an airlift reactor operation was higher than those in packedcolumn reactor operation due to a better oxygen transfer.     

Immobilization of Desulfovibrio vulgaris cells with hydrogenase activity

Desulfovibrio vulgaris intact cells immobilized in calcium alginate retained 45% of the hydrogenase activity of free cells. Cell immobilization had no effect on the optimum temperature for the hydrogen uptake rate, whereas the pH optimum was shifted to a higher value. The immobilization conferred on the cells a protection both against oxygen inhibition and thermal denaturation of the hydrogenase system.     

固定化色氨酸合成酶基因工程菌合成L-色氨酸的研究

以海藻酸钠作为包埋剂、戊二醛作为交联剂和氯化钙作为填充剂对色氨酸合成酶基因工程菌进行固定化,同时探究三种物质和菌体负载量对固定化菌影响,响应面法优化色氨酸合成酶基因工程菌合成L-色氨酸。固定化色氨酸合成酶基因工程菌最优制备条件为：海藻酸钠28.92 g/l、戊二醛0.95%、氯化钙19.82 g/l、菌体负载量25 g/l。底物L-丝氨酸浓度为1%、固定化菌8g, L-色氨酸转化率为28.16%。固定化菌可连续使用15批次。     

Hydrolysis of milk lactose in a packed bed reactor system using immobilized yeast cells

BACKGROUND: Hydrolysis of lactose with β‐D‐galactosidase is one of the most promising biotechnological applications in the food industry because of its use in the production of low lactose milk products and whey hydrolysis. To overcome the problem of enzyme extraction from cells due to the intracellular nature of β‐D‐galactosidase and the poor permeability of the cell membrane to lactose, permeabilization of yeast cells was investigated. Permeabilized whole cells have been claimed to have an advantage over more pure enzyme preparations. In view of the advantages of immobilized cell systems over free cell systems, permeabilized cells were immobilized by an entrapment method in calcium alginate gel. A packed bed reactor together with this immobilized cell system has been used for hydrolysis of milk lactose in a continuous system. RESULTS: Different process parameters (temperature, substrate feed rate, biomass load and time‐course) were optimized to maximize lactose hydrolysis. The immobilized yeast cells (300 mg dry wt) resulted in 87.2% hydrolysis of milk lactose at 30 °C and flow rate 7 mL h^?1 in a packed bed reactor system. CONCLUSION: This convenient and relatively inexpensive method of immobilization, resulting in high hydrolysis potential in a continuous system, indicates that permeabilized yeast cells have the potential for the production of low lactose milk and milk products. Copyright © 2010 Society of Chemical Industry     

海藻酸铝固定化糖化酶特性研究

采用海藻酸铝固定化糖化酶,对固定化酶和游离酶的特性进行了比较。结果表明,固定化酶米氏常数Km为13.72 mg/mL,最适温度为65℃,半衰期为100.7 d,固定化糖化酶的活力回收率达61.8%。     

Development of a bacterial preparation based on immobilized cells

A new method for removing thin oil films from a water surface with the use of the oil-degrading microorganism Acinetobacter valentis immobilized in calcium alginate gel has been developed. It has been demonstrated that n-alkanes emulsified into calcium alginate granules impart positive buoyancy to the granules. The parameters of obtaining calcium alginate granules with immobilized oil-degrading cells affect the characteristics of biological preparation. It has been demonstrated that the use of the biological preparation containing emulsified n-alkanes makes it possible to increase the bioremediation rate due to localization of immobilized and free cells in the upper water level. The new form of biological preparations makes it possible to decrease the amount of oil hydrocarbons by as much as 97% for 21 days at a temperature of 10–22°C versus 63% in the control variant.