發(fā)布時(shí)間：2018-05-12 18:03

  本文選題：生物催化 + 大豆過氧化物酶　； 參考：《大連工業(yè)大學(xué)》2015年碩士論文

【摘要】：生物催化因其高效、專一性強(qiáng)及環(huán)境污染少等優(yōu)點(diǎn)成為綠色合成化學(xué)品的理想催化劑。然而,天然酶常因其在工業(yè)催化條件下的活性和穩(wěn)定性較低而難以使用。納米技術(shù)為構(gòu)建高效酶催化劑提供了新的可能性。如何通過簡便、高效、低成本的方法制備出具有高催化活性和高穩(wěn)定性的納米酶催化劑,并將其應(yīng)用于化學(xué)合成是其中的關(guān)鍵問題。本課題以大豆過氧化物酶為基礎(chǔ),研究了一種新型的以磷酸銅為載體形成的有機(jī)-無機(jī)結(jié)合的納米花固定化酶方法,并初步探索了合成的納米花固定化酶對硝基油酸生成的催化作用,同時(shí)與化學(xué)法合成的硝基油酸進(jìn)行了比較。得到以下結(jié)論:(1)以大豆皮為原料,對大豆過氧化物酶的提取及初步純化進(jìn)行了研究,通過酶液提取的單因素實(shí)驗(yàn)、正交實(shí)驗(yàn),得到酶的最佳提取工藝條件。采用硫酸銨分級(jí)沉淀和丙酮分級(jí)沉淀的方法對大豆過氧化物酶進(jìn)行了初步純化。結(jié)果表明:酶的最適提取條件為料液比1:8,提取溫度35℃,提取時(shí)間24 h;硫酸銨沉淀下限飽和度:30%,上限飽和度:80%;丙酮沉淀下限體積倍數(shù):0.4,上限體積倍數(shù):1.2。(2)利用實(shí)驗(yàn)室自提的大豆過氧化物酶作為研究對象,采用一種新型的固定化酶方法合成了具有花形結(jié)構(gòu)的納米花固定化酶。通過SEM、TEM圖像展示了該納米花固定化酶的形貌特征,研究了時(shí)間和初始酶濃度對該結(jié)構(gòu)產(chǎn)生的影響。通過紅外光譜、X射線衍射、能譜分析手段確定了此種納米花固定化酶的載體為磷酸銅。通過愈創(chuàng)木酚法測定了納米花固定化酶的活性,與游離酶相比納米花固定化酶表現(xiàn)出增強(qiáng)的催化活性,最高酶活能夠提高到446%。重復(fù)性試驗(yàn)證實(shí)了該納米花固定化酶的循環(huán)性良好,經(jīng)過5次循環(huán)后依然保持80%的活性。(3)以油酸為原料,采用化學(xué)法和生物法合成硝基油酸,得到的產(chǎn)物經(jīng)過紅外光譜,核磁共振及液質(zhì)聯(lián)用分析手段加以佐證,考察納米花固定化酶對硝基油酸生成的影響。結(jié)果表明:生物法合成硝基油酸起到了抑制副產(chǎn)物生成的作用。生物法合成硝基油酸是一種新的嘗試,對未來固定化酶與化學(xué)合成結(jié)合的研究奠定了基礎(chǔ)。
[Abstract]:Biocatalysis is an ideal catalyst for green synthetic chemicals because of its high efficiency, high specificity and less environmental pollution. However, natural enzymes are often difficult to use because of their low activity and stability in industrial catalysis. Nanotechnology provides a new possibility for the construction of efficient enzyme catalysts. It is a key problem that how to prepare high catalytic activity and high stability nanometer enzyme catalyst by simple, high efficiency and low cost method, and apply it to chemical synthesis. Based on soybean peroxidase, a novel organic-inorganic binding enzyme immobilization method based on copper phosphate was studied in this paper. The catalytic effect of the immobilized enzyme on the formation of nitrooleic acid was preliminarily explored and compared with that of nitrooleic acid synthesized by chemical method. Conclusion: (1) the extraction and purification of soybean peroxidase were studied by using soybean skin as raw material. The optimum extraction conditions were obtained by single factor experiment and orthogonal experiment. Soybean peroxidase was preliminarily purified by ammonium sulfate fractionation and acetone fractionation. The results showed that the optimum extraction conditions were as follows: the ratio of material to liquid was 1: 8, the extraction temperature was 35 鈩，

本文編號(hào)：1879595