本論文係有關於氧化鐵磁性奈米粒子在酵素固定化及分離上之應用研究。前者主要將脂肪酶(Lipase)固定化在四氧化三鐵(Fe3O4)磁性奈米粒子上，探討製備變因、產品特性、及其在水相系統中之操作性能。後者主要將聚丙烯酸(PAA)共價鍵結在Fe3O4磁性奈米粒子上，作為一種新型磁性奈米吸附劑，探討製備變因、產品特性、及其在鳳梨酵素吸附分離上的應用。
關於Lipase固定化在Fe3O4磁性奈米粒子上之研究，首先以化學共沉法製備出Fe3O4磁性奈米粒子，然後將Lipase利用carbodiimide活化的方式直接固定化在磁性奈米粒子上，並再探討其操作在水相系統中的操作性能。由穿透式電子顯微鏡(TEM)、X射線繞射儀(XRD)和超導量干涉磁量儀(SQUID磁量儀)分析得知，磁性奈米粒子在Lipase固定化後，其大小、結構和超順磁性並無明顯改變。傅立葉轉換紅外線光譜儀(FTIR)分析可確認Lipase確實已固定化在磁性奈米粒子上，並推測其一可能的反應機構。固定化Lipase的殘餘活性為游離態酵素的1.41倍且其貯存穩定性為游離態酵素的31倍。在pH值方面，固定化Lipase較游離態酵素具有較高的容忍度，而在相關的動力行為在本研究中皆有探討。
關於磁性奈米吸附劑之製備與應用，首先以化學共沉法製備出Fe3O4磁性奈米粒子，然後將PAA藉carbodiimide活化直接共價鍵結在磁性奈米粒子上，形成具有離子交換功能之磁性奈米載體，並再探討其在水相鳳梨酵素吸附分離上的應用。由TEM、XRD和SQUID磁量儀分析得知，磁性奈米粒子在共價鍵結PAA後，其大小、結構和超順磁性並無明顯改變。由FTIR、熱重分析儀(TGA)、熱差分析儀(DTA)和化學分析電子光譜儀(XPS)分析可確認PAA已共價鍵結在磁性奈米粒子上。本研究所得的磁性奈米吸附劑，其離子交換容量為1.64 meq/g，遠較一般商業化的吸附劑為高。而磁性奈米吸附劑在pH 3~5、磷酸鹽濃度為0.1M、鳳梨酵素濃度為6 mg/ml時，在1分鐘內能將鳳梨酵素完全吸附。並且在pH 7時當KCl濃度超過0.6 M就能將鳳梨酵素完全脫附。至於水相鳳梨酵素在磁性奈米吸附劑上的吸附，可以Langmuir恆溫吸附模式描述，而其最大吸附量(qm) Langmuir吸附常數分別為0.476 mg/mg 和58.4 ml/mg。另外鳳梨酵素經過吸附/脫附程序後，其殘餘活性仍達87.4 %。

This thesis concerns the applications of iron oxide magnetic nanoparticles in enzyme immobilization and separation. In the former, Lipase was immobilized on Fe3O4 magnetic nanoparticles. The preparation conditions, product properties, and the performances in the water systems were investigated. In the latter, polyacrylic acid (PAA) was covalently bound onto Fe3O4 magnetic nanoparticles to be a novel nano-adsorbent. The preparation conditions, product properties, and the application in the adsorption of Bromelain in aqueous solution were investigated.
Lipase was covalently bound onto Fe3O4 magnetic nanoparticles (12.7 nm) via carbodiimide activation. The Fe3O4 magnetic nanoparticles were prepared by co-precipitating Fe2+ and Fe3+ ions in an ammonia solution and treating under hydrothermal conditions. The analyses of transmission electron microscopy (TEM) and X-ray diffraction (XRD) showed that the size and structure of magnetic nanoparticles had no significant changes after enzyme binding. Magnetic measurement revealed the resultant lipase-bound magnetic nanoparticles were superparamagnetic with a saturation magnetization of 61 emu/g (only slightly lower than that of the naked ones (64 emu/g)), a remanent magnetization of 1.0 emu/g, and a coercivity of 7.5 Oe. The analysis of Fourier transform infrared (FTIR) spectroscopy confirmed the binding of lipase onto magnetic nanoparticles. Compared to the free enzyme, the bound lipase exhibited a 1.41-fold enhanced activity, a 31-fold improved stability, and better tolerance to the variation of solution pH . The kinetic behavior of bound Lipase was also determined in aqueous solution.
A novel magnetic nano-adsorbent was prepared by covalently binding polyacrylic acid (PAA) on Fe3O4 magnetic nanoparticles (13.2 nm) via carbodiimide activation. From the analyses of TEM, XRD and magnetism, the magnetic nanoparticles showed no change in size, structure and superparamagnetic characteristics after binding PAA. The analyses of FTIR, thermogravimetric analysis (TGA), differential thermal analysis (DTA) and X-ray photoelectron spectroscopy (XPS) confirmed the binding of PAA to magnetic nanoparticles and suggested the binding mechanism of PAA. The ionic exchange capacity of the resultant magnetic nano-adsorbents was estimated to be 1.64 meq/g, much higher than those of the commercial ionic exchange resins. The full recovery of bromelain was achievable within 1 min onto the nano-adsorbents at pH 3-5 and 0.1 M phosphate when the concentration of bromelain was 6 mg/ml. Moreover, the complete desorption of bromelain from the nano-adsorbents was attained within 1 minute at pH 7 when the concentration of KCl was above 0.6 M. The isothermal adsorption indicated that the adsorption behavior of bromelain followed the Langmuir adsorption isothermal and the values of the maximum amount of adsorbed bromelain (qm) and Langmuir constant (K) were 0.476 mg/mg and 58.4 ml/mg, respectively. In addition, bromelain retained 87.4% activity after adsorption/desorption.

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