本文選題：復合載體 + 固定化酶��； 參考：《蘭州大學》2017年碩士論文

【摘要】：酶作為一種天然催化劑,以其高效、溫和、綠色的獨特優(yōu)勢成為生物、化工、醫(yī)藥等領域的重要研究對象。脂肪酶來源廣泛,能夠在水相和有機相中催化酯化反應、水解反應、酯交換反應等,并且具有化學選擇性、區(qū)域選擇性、對映體選擇性,是一種具有巨大潛力的工業(yè)用酶。但是,在使用過程中游離酶對外界環(huán)境敏感、成本高等缺點限制了其應用,因而推動了酶固定化技術的發(fā)展。經(jīng)過固定化之后,酶的性能得到優(yōu)化,擴大了酶催化的適用范圍,有效降低了生產(chǎn)成本,提高生產(chǎn)產(chǎn)率。反應色譜技術是將化學反應和色譜分離集合在一個單元的技術,不僅能夠簡化化學過程,降低生產(chǎn)成本,而且能夠突破可逆反應化學平衡的限制,提高產(chǎn)率。高效液相色譜能夠很大程度上降低實驗過程的復雜程度。將反應色譜技術和固定化酶技術結合使用后,將固定化酶裝填為具有分離和催化功能的固定化酶色譜柱,實現(xiàn)生物催化與色譜技術的結合,深入反應色譜技術的研究。本論文中選擇硅膠和纖維素組成的復合載體分別用物理吸附法和交聯(lián)法制備了兩種固定化酶,裝填成固定化酶色譜柱后對其在線催化水解反應效果進行了評價。脂肪酶的底物選擇對硝基苯酚棕櫚酸酯,反應體系由正己烷、異丙醇、水組成。吸附法固定化酶的制備中,乙基纖維素涂覆硅膠是最適宜的載體,吸附固定化酶的溫度耐受范圍為15℃-45℃。乙基纖維素硅膠固定化酶色譜柱能實現(xiàn)對硝基苯酚棕櫚酸酯的在線水解和分離,具有反應色譜的性質,穩(wěn)定性很高,在溫度為18℃-50℃范圍內具有較高的催化活性,半衰期長達160 h。交聯(lián)法固定化酶具有良好的操作穩(wěn)定性,對對硝基苯酚具有良好的耐受性,經(jīng)過極性底物抑制或者活化后可恢復活性。交聯(lián)固定化酶色譜柱親水性很強,對對硝基苯酚的吸附強于對對硝基苯酚棕櫚酸酯的吸附。在水含量為10%時,固定化酶色譜柱的催化活性(轉化率50%以上)高于游離酶。流動相中水含量升高時底物轉化率上升,水含量超過20.7%時逐漸達到最大值。正己烷含量的上升會抑制固定化酶色譜柱的催化活性。經(jīng)過160 h反應后交聯(lián)固定化色譜柱活性幾乎沒有衰減,十分穩(wěn)定。
[Abstract]:As a kind of natural catalyst, enzyme has become an important research object in biology, chemical industry, medicine and other fields with its unique advantages of high efficiency, mild and green. Lipases can catalyze esterification, hydrolysis and transesterification in aqueous and organic phases, and have chemical selectivity, regioselectivity and enantioselectivity. Lipase is a kind of industrial enzyme with great potential. However, in the process of using free enzyme is sensitive to the external environment, high cost and other shortcomings limit its application, thus promoting the development of enzyme immobilization technology. After immobilization, the performance of the enzyme was optimized, the application range of enzyme catalysis was expanded, the production cost was reduced effectively, and the production yield was increased. Reaction chromatography is a technique of separating chemical reaction and chromatography into one unit. It can not only simplify chemical process, reduce production cost, but also break through the limit of chemical equilibrium of reversible reaction and increase yield. High performance liquid chromatography (HPLC) can greatly reduce the complexity of the experimental process. After the combination of reaction chromatography and immobilized enzyme technology, the immobilized enzyme was packed into an immobilized enzyme chromatographic column with separation and catalytic functions. The combination of biocatalysis and chromatography technology was realized, and the study of reaction chromatography technology was carried out. In this paper, two kinds of immobilized enzymes were prepared by physical adsorption method and cross-linking method respectively on the composite carrier composed of silica gel and cellulose. The effect of on-line catalytic hydrolysis reaction was evaluated by filling the immobilized enzyme chromatographic column. The lipase substrate was p-nitrophenol palmitate. The reaction system consisted of n-hexane, isopropanol and water. In the preparation of immobilized enzyme by adsorption, ethyl cellulose coated with silica gel is the most suitable carrier, and the temperature tolerance range of adsorption immobilized enzyme is from 15 鈩，

本文編號：1862509