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氨基树脂固定胃蛋白酶的方法及性质研究 总被引:1,自引:0,他引:1
采用氨基树脂作为载体,戊二醛作为交联剂,对胃蛋白酶的固定化进行了研究,并对固定化条件和固定化胃蛋白酶的部分酶学性质进行了分析。确定固定条件为:戊二醛浓度为5%,载体处理温度为室温(25℃),处理时间为5h,m(胃蛋白酶):m(氨基树脂)为1:25,pH为3.0,固定时间为12h。此条件下固定化的胃蛋白酶活力为30U/g,酶的活力回收率为60%。与非固定化相比最适水解温度由50℃升高到60℃,最适pH值由2.0升高到4.0,游离酶米氏常数3.08g/L,固定化酶米氏常数1.2g/L,固定化胃蛋白酶的储存半衰期约为25天。对珠蛋白的操作半衰期为9天。 相似文献
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海藻酸钠-明胶协同固定化S-腺苷甲硫氨酸合成酶的研究 总被引:1,自引:0,他引:1
以海藻酸钠和明胶为载体,对S-腺苷甲硫氨酸合成酶进行固定化。再用戊二醛对其进一步交联,增强固定化酶的稳定性。考察了海藻酸钠和明胶质量分数、CaCl2质量分数、酶和载体比例以及交联剂戊二醛体积分数等因素对固定化酶的影响。结果表明,最佳固定化条件为:海藻酸钠质量分数2.0%、明胶质量分数1.0%、CaCl2质量分数4.0%、固定化酶量为2.5 g/L凝胶、戊二醛体积分数0.6%。交联固定化酶热稳定性得到大幅度提高,在50℃下保温5 h仍保留72%的活力,而游离酶则完全失活。交联固定化酶在碱性溶液中的稳定性较高,在pH=8.0~9.0的缓冲液中4℃保温10 h酶活性仍保留87%以上。将交联固定化酶用于S-腺苷甲硫氨酸的合成,连续反应8批次后酶活性仍保留65%。 相似文献
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以氨基树脂为载体对S-腺苷甲硫氨酸(SAM)合成酶进行固定化,优化了酶的固定化条件并对固定化酶的性质进行了研究。优化的固定化条件为:戊二醛体积分数5%、SAM合成酶添加量20mg·g-1、固定化时间5h。所制备的固定化SAM合成酶的酶活力为476.8U·g-1,酶活力回收率为74.5%。与游离SAM合成酶相比,固定化SAM合成酶的稳定性大幅提高,在50℃孵育5h酶活力仍保留61.2%,而游离SAM合成酶则完全失活;在pH值为6.0~6.5、8.0~9.5的缓冲溶液中,固定化SAM合成酶也更加稳定;固定化SAM合成酶连续催化反应10批次,酶活力保留86.3%;固定化SAM合成酶在4℃储存30d,酶活力保留81.4%。固定化SAM合成酶米氏常数KATPm=0.14mmol·L-1,KLm-Met=0.28mmol·L-1。 相似文献
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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 总被引:1,自引:0,他引:1
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。 相似文献
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研究了以中空球形壳聚糖固定As1.398中性蛋白酶的方法,分析了戊二醛浓度、给酶量对固定化的影响,并对固定化酶的性质进行了讨论。实验结果显示:戊二醛质量浓度为4%、每克载体给酶量25mg时,酶活力回收为65.4%,固定化酶的热稳定性、pH稳定性及乙醇的稳定性均高于游离酶。 相似文献
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采用包埋法对重组大肠杆菌E.coli BL21进行了固定化研究,确定海藻酸钙是较好的固定化载体。考察了海藻酸钠质量分数、CaCl2质量分数、钙化时间、细胞包埋量以及固定化颗粒直径对固定化细胞活性和机械强度的影响,确定优化的制备条件为:海藻酸钠的质量分数为2.0%,CaCl2的质量分数为2%,钙化10 h,包埋菌体的质量浓度为0.05 g/mL,颗粒直径为2.7 mm。与游离细胞相比,固定化细胞的热稳定性和pH稳定性有明显提高。利用固定化重组大肠杆菌细胞催化拆分体积分数为1.5%的外消旋环氧氯丙烷,得到(R)-环氧氯丙烷的产率和光学纯度(对映体过量)分别为36.3%和100%。固定化细胞重复4批仍保持80%以上的活力,显示了良好的操作稳定性。 相似文献
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反胶束体系中青霉素酰化酶活性及稳定性研究 总被引:6,自引:0,他引:6
反胶束体系是一种新型的酶反应介质体系。构建合适的反应体系,使青霉素酰化在该体系中保持较高的酶活性,是实现反胶束体系中酶法合成头孢类抗生素的关键所在。本文分别考察了青霉素酰化酶在CTAB/正己醇和CTAB/异辛烷/正己醇两种反应胶束体系中不同水含量和pH值对酶活力的影响。结果表明:包埋于CTAB/异辛烷/正己醇反胶束体系中的青霉素酰化酶在W0为14,pH为7.2时表现出较高的酶活性。试验中还考察了不同温度对包埋于CTAB/异辛烷/正己醇反胶束体系中的青霉素酰化酶活力的影响,并与游离酶液进行了比较,该酶在所采用的反胶束体系中的最适水解反应温度为39℃,较游离酶液的最适水解温度升高了2℃。同时酶的热稳定性亦较游离酶液有所提高,表现在CTAB/异辛烷/正己醇反胶束体系中,经50℃保温240min,酶活保留率为76.0%,而在同样条件下游离酶液仅为36.5%。 相似文献
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为提高酶的催化水解活力和稳定性,将青霉素酰化酶组装于介孔泡沫二氧化硅(MCFs)中,并应用于水/有机混合体系催化水解。分别考察了有机介质种类和体积分数、葡聚糖(Dex10k)修饰对固定化酶活力的影响,研究了不同条件下固定化酶的稳定性。实验结果显示:体积分数20%石油醚中,添加Dex10k的介孔泡沫硅固定化酶比活力达209.5U/mg,是缓冲液中MCFs固定化酶活力的196.2%。20%石油醚中,经25次连续操作,固定化酶保持初始活力的71.5%。结果表明:石油醚等烷烃形成的水/有机体系是适合青霉素酰化酶催化的二相体系,且添加Dex10k能提高固定化酶在二相体系中的催化活力及稳定性。 相似文献
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Pepsin [EC, 3.4.23.1] from Porcine stomach mucosa was immobilized in silicone elastomers utilizing condensation-cure room temperature vulcanization (RTV) of silanol-terminated
poly(dimethylsiloxane) (PDMS). Two network precursor chain molar masses were used in this investigation: in pepsin–silicone
(A), M
n ∼26,000 g mol−1 and in pepsin–silicone (B) M
n ∼750 g mol−1. Tetraethyl orthosilicate (TEOS) was used as the cross-linking agent and dibutyltin dilaurate was used as the catalyst. The
activity and stability of free pepsin and pepsin immobilized in PDMS were studied with respect to pH, temperature, cross-link
density, solvents and storage time using a hemoglobin assay. A notable finding is that free pepsin has zero activity in neutral
buffer solution (pH 7) after incubation for 5 h, while pepsin immobilized in the silicone elastomers was found to retain more
than 70% of its maximum normalized activity. There was no marked improvement in the thermal stability of the PDMS immobilized
pepsin when compared to free pepsin and all the three systems showed no activity at and above 70 °C. From the Lineweaver–Burk
kinetic analyses, the apparent K
m (g L−1 hemoglobin) for free pepsin was 4.5, for pepsin–silicone (A) was 5.1, and for pepsin–silicone (B) was 3.9, the V
max (U/mg of pepsin) for free pepsin was 14,000, for pepsin–silicone (A) was 11,710, and for pepsin–silicone (B) was 8,510, respectively
after incubation in buffer solution at pH 2 and 37 °C. The activity of the free and the PDMS immobilized pepsin in six different
organic solvents was also studied. The pepsin retained high activity in non-polar solvents such as n-hexane, isooctane and toluene, but the enzyme performed poorly in methanol, ethanol and tetrahydrofuran. The degree of swelling
of the pepsin immobilized silicone elastomers in these solvents had no impact on the activity of the pepsin. When stored at
room temperature for time periods up to 6 months, pepsin immobilized in silicone elastomers was observed to retain its full
activity. The results reported herein demonstrate that cross-linked PDMS is a promising support material for the immobilization
of hydrolytic enzymes such as pepsin. 相似文献
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Synthesis of Fatty Acid Ethyl Ester from Acid Oil in a Continuous Reactor via an Enzymatic Transesterification
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Nakyung Choi Yangha Kim Jeom-Sig Lee Jieun Kwak Junsoo Lee In-Hwan Kim 《Journal of the American Oil Chemists' Society》2016,93(3):311-318
Synthesis of a fatty acid ethyl ester via the lipase‐catalyzed transesterification of acid oil and ethanol was investigated in a continuous reactor. Lipozyme TL IM was employed as the immobilized lipase. This immobilized lipase derived from Thermomyces lanuginosus was purchased from Novozymes (Seoul, Korea). The acid oil was prepared by the acidification of soapstock formed as a by‐product during the refining of rice bran oil. The parameters investigated were water content, temperature, and molar ratio of substrates. The relative activity of Lipozyme TL IM was assessed during the repeated use of the immobilized lipase. The water content of the substrate had a considerable effect on the yield and the optimum water content was 4 %. The optimum temperature and molar ratio of acid oil to ethanol were 20 °C and 1:4, respectively. The maximum yield of approximately 92 % was achieved under the optimum conditions. The corresponding compositions were 92 % fatty acid ethyl esters, 3 % fatty acids, and 5 % acylglycerols. When glycerol formed during the reaction was removed by intermittent washing with ethanol, the relative activity of lipase was maintained over 82 % for a total usage of 27 cycles. For a mean residence time of 4 h, the half‐life times of Lipozyme TL IM on the control (unwashed) and treatment (washed) were 39 and 45 cycles, respectively. 相似文献
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Starch‐poly(acrylamide‐co‐2‐acrylamido‐2‐methylpropanesulfonic acid) graft copolymers prepared by reactive extrusion
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Graft copolymers of starch with acrylamide and 2‐acrylamido‐2‐methylpropanesulfonic acid (AMPS) were prepared by reactive extrusion in a twin‐screw extruder. The weight ratio of total monomer to starch was fixed at 1 : 3, while the molar fraction of AMPS in the monomer feed ranged from 0 to 0.119. Monomer to polymer conversions were 85% or greater, with grafting efficiencies of 68% (highest AMPS content) to 85% (no AMPS). Absorbency in distilled water at pH 7 increased linearly with the mole fraction AMPS in the grafted polymer, while absorbencies in 0.9% NaCl were independent of AMPS content. When swollen in water/ethanol mixtures, swelling decreased gradually with increasing ethanol volume fraction, followed by a large decrease over a narrow ethanol concentration. This behavior is similar to that observed for AMPS‐acrylamide gels. The swelling properties suggest these graft copolymers may have applications as responsive materials. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42405. 相似文献
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固定化Candida sp.99-125脂肪酶催化大豆油合成脂肪酸乙酯 总被引:1,自引:0,他引:1
探讨了酶法合成脂肪酸乙酯作为生物柴油的可行性. 以大豆油和乙醇为原料,利用本实验室自制的固定化Candida sp. 99-125脂肪酶催化反应,深入研究水含量、溶剂量、脂肪酶量及反应温度等因素对酶法合成脂肪酸乙酯的影响. 结果表明,以大豆油质量为基准,在水含量为 12.5%(w)、溶剂正己烷为3 mL/g、脂肪酶量为20%(w)、温度40℃的优化反应条件下,3次流加乙醇,170 r/min摇瓶反应,12 h后可以达到96.8%的最高酯得率. 进一步研究表明,在此优化反应条件下,连续使用14批脂肪酶酯得率可保持70%以上. 相似文献