離子液體對固定化纖維素酶水解效率之影響

Effects of ionic liquids on cellulose saccharification using magnetic nanoparticle-bound cellulase

陳冠存(Kuan-Chun Chen)

指導教授 ： 游吉陽

大同大學/工程學院/生物工程學系所/碩士(2009年)

摘要

生質酒精是現今最受矚目的替代能源之一，每年產量約佔全球生質燃料的90%。可用來生產酒精的生質原料主要包含：糖質、澱粉、纖維素等三種，由於前兩種是人類或動物食物來源，易與糧食資源競爭，因此利用纖維素為原料製備纖維酒精是目前的研究重點，而生產纖維酒精最大的問題是如何提高水解反應效率。
纖維素為葡萄糖分子聚合成的直鏈狀聚合物，鏈與鏈之間還有大量的氫鍵與凡德瓦爾力，平行排列後形成的結晶結構，造成cellulase難以充分與纖維素反應，水解速度大幅減緩。
本研究先以FeCl2、FeCl3與NH4OH共沉澱反應生成Fe3O4磁性奈米粒子 (magnetic nanoparticles, MNP)，粒子表面經3-aminopropyltriethoxysilane及glutaraldehyde兩段化學修飾，再將Trichoderma ressei cellulase固定於MNP上 (cellulase-MNP)，並嘗試利用三種離子液體 (Ionic Liquid, IL)，1-n-butyl-3-methyl-
imidazolium chloride (BMIMCl)、1-allyl-3-methylimidazolium chloride (AMIMCl)和1-ethyl-3-methylimidazolium diethylphosphate ([C2mim][(EtO)2PO2])，對纖維素進行預處理，將氫鍵破壞以瓦解纖維素的結晶結構，加快cellulase的水解速度。
經XRD分析結果顯示：(1) 所製備出之粒子組成為Fe3O4；(2) IL可有效破壞纖維素內部的結晶結構；TEM顯示，Fe3O4之粒子大小約20 nm。當固定化時cellulase濃度為0.6 mg/ml，cellulase固定量約0.04 mg/per mg of Fe3O4；活性回收約48%。在利用IL (BMIMCl、AMIMCl和[C2mim][(EtO)2PO2])，以不同比例對纖維素進行預處理部分，三種IL在IL/water比例為1/4時水解效果最好，cellulase於BMIMCl中水解1 h得到之還原糖為未經IL預處理之纖維素的的1.4倍；水解24 h後則為1.5倍；AMIMCl則分別為3.2倍與2倍；[C2mim][(Et)2PO2]則分別為1.4倍與1.5倍。Cellulase-MNP於BMIMCl中水解1 h得到之還原糖為未經IL預處理之纖維素的1.9倍，水解24 h後則為1.3倍；AMIMCl則分別為7.8倍與4.2倍；[C2mim][(Et)2PO2]則皆為2.4倍。當IL/water比例超過3/2時，cellulase與cellulase-MNP兩者幾乎沒有活性。

關鍵字

纖維素 ； 纖維素酶 ； 磁性奈米粒子 ； 生質酒精 ； 離子液體

並列摘要

Bioethanol is perhaps the most important alternative fuel; the annual production of bioethanol accounts for 90% of global production of biofuels.The major feedstock for bioethanol production includes sugars, starch, and cellulose. However, sugars and starch are also food sources for human and animal, using these two feedstocks may lead to potential food crisis. Cellulose appears to be the most appropriate feedstock for bioethanol, nevertheless, hydrolysis rate of cellulose to glucose by cellulase is limited due to high crystallinity in cellulose.
In this work, cellulase was covalently bound onto Fe3O4 magnetic nanoparticles (MNP), synthesized by the co-precipitation of FeCl2 and FeCl3, MNP was activated by 3-aminopropyltriethoxysilane and glutaraldehyde. Furthermore, three different ionic liquids (IL), 1-n-butyl-3-methylimidazolium chloride (BMIMCl), 1-allyl-3-
methylimidazolium chloride (AMIMCl), and 1-ethyl-3-methylimidazolium diethyl-
phosphate ([C2mim][(EtO)2PO2]), were used to pretreat cellulose. The selected IL can accelerate the hydrolysis of cellulose by interrupting its crystalline region.
The XRD analyses revealed that: (1) the structure of the MNP is Fe3O4. (2) IL efficiently disrupts the crystalline region inside cellulose. The TEM micrographs showed that the diameter of the MNP was approximately 20 nm. When cellulase concentration was 0.6 mg/ml, the amount of immobilized cellulase was approximately 0.04 mg/per mg of Fe3O4 with an activity recovery of 48%. Regarding cellulose pretreatment with IL in various IL/water volume ratios, optimal hydrosis efficiency was observed at IL/water ratios of 1/4 for all three ILs; for BMIMCl using cellulase , the reducing sugar concentration was increased by 1.4-fold and 1.5-fold after 1 and 24 h of incubation, respectively, when compared to untreated cellulose; 3.2-fold and 2-fold increase for AMIMCl; 1.4-fold and 1.5-fold for [C2mim][(EtO)2PO2]. For BMIMCl using cellulase-MNP , the reducing sugar concentration was increased by 1.9-fold and 1.3-fold after 1 and 24 h of incubation, respectively, when compared to untreated cellulose; 7.8-fold and 4.2-fold increase for AMIMCl; 2.4-fold and 2.4-fold for [C2mim][(EtO)2PO2]. When IL/water ratio was greater than 3/2, little activity was observed for both cellulase and cellulase-MNP .

並列關鍵字

ionic liquid ； 1-ethyl-3-methylimidazolium diethylphosphate ； bioethanol ； cellulose ； 1-allyl-3-methylimidazolium chloride ； cellulase ； magnetic nanoparticles ； 1-n-butyl-3-methylimidazolium chloride

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