【正文】 zyme sources, reaction media, the molar ratio of substrate, temperature and the structure of substrates, lipase from porcine pancreas, Type II (PPL) catalyzed the FriedelCraft reaction of indole derivative and α, βunsaturated pounds. 15 FridelCraft adducts were synthesized. The control experiments proved that the active site of PPL were responsible for those enzymatic reaction. The mechanisms for the FriedelCraft reaction and tandem reaction catalyzed by PPL were also proposed.We screening the PPL catalyze the tandem Nucleophilic addition/dehydration /Michael addition of aldehyde and indole. The enzyme sources, reaction media, temperature, the structure of substrate that affects this tandem reaction were investigated systemly. In mixture solvent (water/dioxane=4/1), lipase (from porcine pancreas, Type II) can catalyze the tandem Nucleophilic addition/dehydration /Michael addition between aldehydes and indole derivatives. After the stepwise process were optimized, 26 bis(indolyl)alkanes were prepared in moderate to excellent yields by using PPL as catalyst.The influence of Nheterocyclic pounds on the enzymatic reaction was investigated. Though investigating enzyme sources, solvent, additive, the ratio of imidazole and acylase, we found Daminoacylase and imidazole can cocatalyze the ClaisenSchmidt reaction. When the ratio of Daminoacylase and imidazole is 2/1, the activity of Daminoacylase was improved obviously. And 18 tandems Aldol condensation/dehydration adducts were obtained via this method. The control experiments demonstrated that the active site of Daminoacylase was responsible for the enzymatic reaction. The mechanism for the ClaisenSchmidt reaction catalyzed by Daminoacylase and imidazole was also proposed.A single enzyme can cocatalyze the cascade Aldol condensationelimination Michaelintromolecular Aldol condensationelimination for synthesis of conjugate cyclohexanone2ene reaction with imidazole in octane. By investigating enzyme sources, solvent, additive, the ratio of imidazole and acylase, we found Daminoacylase and imidazole can cocatalyze this tandem reaction. After the stepwise process was optimized, 15 conjugate cyclohexanone2enes were prepared. Some control experiments were designed to prove that the enzyme can cocatalyze this cascade reaction with imidazole in octane. The mechanism for the tandem reaction catalyzed by Daminoacylase and imidazole was also proposed.In this thesis, 74 pounds were synthesized and these pounds were characterized by IR, 1H NMR, 13C NMR, GCMS and HRMS.Keywords: lipase。 學(xué)位論文作者簽名： 簽字日期： 學(xué)位論文版權(quán)使用授權(quán)書 本學(xué)位論文作者完全了解 浙江大學(xué) 有關(guān)保留、使用學(xué)位論文的規(guī)定，有權(quán)保留并向國家有關(guān)部門或機(jī)構(gòu)送交論文的復(fù)印件和磁盤，允許論文被查閱和借閱。發(fā)現(xiàn)當(dāng)酶與咪唑質(zhì)量比為2/1時(shí)，?；傅拇呋疌laisenSchmidt活性顯著提高。 酶催化多功能性的研究酶的催化多功能性在最近十年研究的新進(jìn)展，Berglund和 Gotor等人分別報(bào)道了脂肪酶CALB具有催化碳碳鍵[2]和碳氮鍵Michael[3]加成反應(yīng)新功能。 小分子對(duì)酶促反應(yīng)的調(diào)控酶的結(jié)構(gòu)決定了酶的催化功能，一些酶與小分子存在著特殊的作用，從而能引起蛋白構(gòu)象的變化，添加劑的使用能夠調(diào)節(jié)酶催化的功能[23],例如：化學(xué)選擇性調(diào)控、立體選擇性調(diào)控和區(qū)域選擇性調(diào)控。 (b) Ch Li, Feng, N Wang, Y. J Zhou and X. Q Yu, Green Chem., 2008, 10, 616.[6]. C Branneby, P Carlqvist, A Magnusson, K Hult, T Brinck, P Berglund. J. Am. Chem. Soc. 2003, 125, 874. [7]. K Li, T He, Ch Li, X. W Feng, N Wang and X. Q Yu, Green Chem., 2009, 6, 777.[8]. (a) Y Cai, X. F Sun, N Wang, X. F Lin. Synthesis, 2004, 671。與分步反應(yīng)相比, 巧妙的串聯(lián)反應(yīng)不僅能提高反應(yīng)效率，具有高選擇性、原子經(jīng)濟(jì)性,大量減少浪費(fèi)，還能免去了各步后處理和分離帶來的消耗和污染。通過化學(xué)計(jì)算表明絲氨酸會(huì)與組氨酸形成氫鍵，因而絲氨酸的存在會(huì)降低組氨酸與天冬氨酸協(xié)同奪取質(zhì)子的能力。圖 26 水相中脂肪酶催化的羥醛縮合反應(yīng)我們課題組報(bào)道了在DMSO溶劑中，嘧啶類化合物與丙烯酰氧基丙酸乙烯酯的碳氮鍵Michael加成反應(yīng)可以利用Amano Lipase M[11]來催化，合成了一系列的含有嘧啶乙烯酯衍生物（圖27）。這個(gè)機(jī)理中β碳上的負(fù)電荷被活性中心的鋅離子穩(wěn)定，天冬氨酸又作為質(zhì)子傳遞媒介完成反應(yīng)。 圖217 N?；被嵯复呋撍磻?yīng) Janssen課題組[25]報(bào)道了負(fù)離子親核試劑與環(huán)氧化合物的立體選擇性開環(huán)反應(yīng)可以由鹵代醇脫鹵酶（Agrobacterium radiobacter AD1）催化發(fā)生，產(chǎn)物 ee 值范圍在90%~99%。最后在漆酶的作用下，1，3二羰基化合物上的羰基氧對(duì)鄰二苯醌又發(fā)生一個(gè)Michael加成反應(yīng)生成最終的苯并呋喃類衍生物。該課題組還建立了雙醇脫氫酶催化體系。圖 230 串聯(lián)反應(yīng)合成光學(xué)純苯乙烯衍生物的二羥基化Strauss 等使用雙酶體系催化扁桃酸的動(dòng)態(tài)動(dòng)力學(xué)拆分，從而實(shí)現(xiàn)了該過程來合成手性化合物。Ostaszewski等人發(fā)現(xiàn)novozyme 435 能夠催化 3苯基戊二酸酐得到不對(duì)稱的酸酶促水解反應(yīng)與醛、胺和異腈的Ugi反應(yīng)組合的方法[52]，最終合成具有一定光學(xué)純的產(chǎn)物。圖239 加氫還原與酶促?；?lián)Makkee 等人發(fā)現(xiàn)一些金屬試劑不會(huì)引起酶失活[57ab]，甘露醇可以通過組合催化劑異構(gòu)酶和銅催化葡萄糖轉(zhuǎn)化得到。與化學(xué)法相比，該方法的選擇性很高。其次是Lewis酸催化發(fā)生Michael加成得到最終產(chǎn)物（圖 247）。底物3′位的選擇性可以通過加入5%(v/v)[BMIM]PF6的介質(zhì)得到（圖 249）。當(dāng)脂肪酶受到MβCD活化時(shí)，對(duì)R對(duì)映體的選擇性可以提高約17倍，立體選擇性提高約3倍。該立體選擇性是沒有酸存在時(shí)的10倍。 (b) G. H Dijoux, M. M Elenkov, J. H. L Spelberg, B Hauer and D. B Janssen, ChemBioChem., 2008, 9, 1048。 (b) E A. Taylor, J B. Garrett, J B. Thoden, H M. Holden, I Rayment, J A. Gerlt. Biochemistry 2004, 43, 224. [25]. G HasnaouiDijoux, M M. Elenkov, J H L. Spelberg, B Hauer, D B. Janssen. ChemBioChem, 2008, 9, 1048. [26]. (a) T Lange, P Hedden, J E. Graebe. Proc. Natl. Acad. Sci. USA. 1994, 91, 8552。Tsai 課題組研究發(fā)現(xiàn)在異辛烷中外消旋萘普生和三甲基硅甲醇的立體選擇性的酯化反應(yīng)可以通過Candida cyclindracea脂肪酶催化得到[87]，AOT的加入可以明顯改善(S)萘普生酯的產(chǎn)率和立體選擇性。堿絡(luò)合物體積越大，酶的立體選擇性就越高。作者考察了一系列冠醚和幾種聚醚發(fā)現(xiàn)，在添加苯丙冠醚、氮雜冠醚和硫雜冠醚時(shí)，脂肪酶催化的水解反應(yīng)的立體選擇性可以明顯提高。Park和Kazlauskas發(fā)現(xiàn)并證明了Cantarctica 脂肪酶對(duì)葡萄糖具有區(qū)域選擇性[70]，在1(2甲氧基乙基)3甲基咪唑啉四氟硼酸酯([MOEMIM]BF4)中酶催化該反應(yīng)得到的幾乎是6?；a(chǎn)物。該過程操作簡(jiǎn)便，不需要中間體的分離和任何保護(hù)/去保護(hù)的措施（圖 246）。首先，苯甲醛衍生物與Wittig試劑反應(yīng)生成烯酮。圖238 動(dòng)態(tài)動(dòng)力學(xué)立體選擇性合成腈醇 Park 和 Kim等人建立了手性胺的高效合成的方法[56]，它是以酮肟為起始原料一鍋法來合成得到。圖234 三酶一鍋法合成糖類衍生物 Sheldon[49]等人發(fā)現(xiàn)5脫氧乙基D木糖可以利用4種酶來催化丙三醇的磷酸化、氧化、羥醛縮合及去磷酸化四步反應(yīng)來合成。這種方法能夠制備光學(xué)純的苯基1，2二羥基苯乙烷的衍生物。作者發(fā)現(xiàn)當(dāng)在第二步中加入弱酸時(shí)可以明顯地加速該反應(yīng)。圖 221 單酶催化羥基化氧化DielsAlder的串聯(lián)反應(yīng) Beifuss課題組[33]在研究漆酶時(shí)發(fā)現(xiàn)：漆酶可以催化串聯(lián)的氧化Michael加成氧化Michael加成反應(yīng)生成一系列苯并呋喃類衍生物（圖 222）。接著Ohta課題組[23]報(bào)道了芳基丙二酸酯脫羧酶可以催化aldol反應(yīng)，他們?cè)谠囼?yàn)中發(fā)現(xiàn)酶的催化新功能與天然活性都經(jīng)歷一個(gè)烯醇負(fù)離子的中間體。嘧啶類、唑類、嘌呤類等氮雜環(huán)化合物都可以做該反應(yīng)底物，潛在生物活性的氮雜環(huán)衍生物可以利用D氨基?；复呋铣傻玫?。Berglund 和Brinck都報(bào)道了己醛或丙醛的aldol反應(yīng)可以用CAL B在環(huán)己烷中催化進(jìn)行 [9ab]。在異丙醚中，該反應(yīng)可以得到馬氏（Markovnikov）加成產(chǎn)物。最近的研究表明部分酶還具有催化多功能性，即許多酶能夠在主反應(yīng)的活性位點(diǎn)催化第二種反應(yīng)，而且越來越多的研究表明催化多功能性普遍存在。 (c) D. M. Z Schmidt, E. C Mundorff, M Dojka, E Bermudez, J. E Ness, S Govindarajan, P. C Babbitt, J Minshull and J. A Gerlt, Biochemistry, 2003, 42, 8387。Ragauskas和Witayakran[20]也報(bào)道了苯并呋喃衍生物可以通過雙酶體系來合成。 multifunction。論文研究了脂肪酶催化FriedelCrafts烷基化反應(yīng)的新功能，考察了反應(yīng)條件，包括反應(yīng)時(shí)間、酶源、溶劑、底物的比例、酶濃度、溫度、底物結(jié)構(gòu)對(duì)脂肪酶催化FriedelCrafts烷基化反應(yīng)的影響，合成了15種吲哚衍生物。并通過無酶，BSA及失活的PPL來驗(yàn)證酶活性中心的催化作用，提出了可能的