微波辅助蛋白酶水解反应工艺和反应器的研究
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摘要
由于常规的酶解反应所需时间比较长,本课题主要研究了微波辐射辅助蛋白酶水解蛋白质的反应。研究发现采用微波辐射可以加快蛋白酶水解反应,提高反应速率。
设计制造了实验室规模的微波辐射-蛋白酶催化反应器,该装置中由夹套反应器、螺旋形管道微波辐射器,流体循环系统和水浴循环控制恒温系统等部分组成。螺旋形管道反应器置于微波炉内,采用循环式的方法解决了蛋白酶在微波辐射过热时使酶失活的问题。并且该反应器实现了微波辐射功率可调,物料反应温度可调。设计计算了夹套式传热反应器,利用循环水浴加热使物料始终保持在一定温度下反应。
本文以木瓜蛋白酶水解大豆蛋白为研究对象,验证反应器系统的优越性。首先研究了在常规实验下木瓜蛋白酶的最佳水解条件,水解的效果用甲醛滴定法测氨基氮含量的高低来确定,通过单因素实验和正交试验得到木瓜蛋白酶对大豆蛋白水解的最佳工艺条件为:固液比是1:8、酶用量5%(酶与大豆蛋白比)、酶解温度50℃、酶解时间5 h、酶解pH值6.7。
由于反应系统中物料是循环式的,固液比是1:8时循环不顺畅,所以取固液比1:10。反应器内循环反应条件为固液比是1:10、酶用量5%、酶解温度50℃、酶解pH值6.7,在不加微波辐射条件下酶解时间5 h与用480 W微波辐射反应120 min试验对比,结果表明用微波辐射60 min氨基氮含量就达到了不用微波辐射下反应5 h的氨基氮含量,反应速率提高了5倍以上。研究了微波功率对水解的影响,功率越高水解速率越快。
用液相色谱分析比较了用微波辐射水解1 h和不用微波辐射水解5 h水解液游离氨基酸的含量,结果表明各种游离氨基酸含量和总氨基酸含量基本相同。
最后对反应器的通用性和放大性进行了研究。
本课题研究表明,微波辐射能辅助蛋白酶水解,设计的反应器有优越性,能提高反应速率。
The conventional hydrolyzation reaction takes long time. This paper has mainly researched the protease-hydrolisis reaction in the assistance of the microwave radiation. The study shows that this method of microwave radiation can speed up the reaction rate.
A laboratory scale microwave radiation reactor for protease-hydrolisis was designed and constructed, and the reactor consists of stirring tank or reactor, helixing pipeline reactor radiated by microwave in cavity, liquid circle system, water circle tank for the controlling the reactant liquid and so on. The over heat effect of microwave radiation can be avoided in this circulation type recator. Moreover, power of microwave radiation and the temperature of reactant liquid can be adjusted in this reactor. The stirring tank was designed and computated, The water circulation tank enables the reactant liquid in certain desireable temperature.
The parameters soy protein hydrolyzed by papayin were optimized in this paper prior to microwave radiation reaction system. Firstly, the best hydrolysis conditions of papayin were studied in regular experiments, the content of nitrogen measured by formaldehyde titrated. the optimizated. craft conditions of hydrolysis analysised by the experiment of single factor and the orthogonal test are: the ratio of solid-liquid ,1:8, the contend of papayin, 5%, the temperature of hydrolysis ,50℃, the hydrolysis time, 5 h, the pH value of hydrolys ,6.7. Because the reactor is a circulating system, And the ratio of solid-liquid, 1:8, the cycle is not smooth, so the ratio of solid-liquid, 1:10. In the reactor the reaction conditions are: the ratio of solid-liquid , 1:10, the content of papain ,5%, the temperature of hydrolysis ,50℃, and the pH value of hydrolysis , 6.7, Contrast the result of 5 hour-hydrolysis experiment without microwave radiation to 120 min hydrolysis experiment under 480 W microwave radiation, the result shows that the content of nitrogen generated by 60 min-hydrolysis experiment under microwave radiation reached the content of nitrogen by 5 hour-hydrolysis experiment without microwave radiation, the rate of reaction increases more than five times. The microwave’s power on the impact of hydrolysis were studied, the higher power,the sooner hydrolysis’s rate.
Compared the content of free amino-acid generated by 5 hour-hydrolysis experiment without microwave radiation to 60 min-hydrolysis experiment under 480W-power-microwave radiation by liquid chromatography analysis, The results showed that the content of each free-amino-acid and the total free-amino-acid are basical the same.
Finally, the common nature and enlargement of the reactor were studied.
The research showed that microwave-radiation can assist hydrolysis, the design of the reactor can increase the rate of action.
设计制造了实验室规模的微波辐射-蛋白酶催化反应器,该装置中由夹套反应器、螺旋形管道微波辐射器,流体循环系统和水浴循环控制恒温系统等部分组成。螺旋形管道反应器置于微波炉内,采用循环式的方法解决了蛋白酶在微波辐射过热时使酶失活的问题。并且该反应器实现了微波辐射功率可调,物料反应温度可调。设计计算了夹套式传热反应器,利用循环水浴加热使物料始终保持在一定温度下反应。
本文以木瓜蛋白酶水解大豆蛋白为研究对象,验证反应器系统的优越性。首先研究了在常规实验下木瓜蛋白酶的最佳水解条件,水解的效果用甲醛滴定法测氨基氮含量的高低来确定,通过单因素实验和正交试验得到木瓜蛋白酶对大豆蛋白水解的最佳工艺条件为:固液比是1:8、酶用量5%(酶与大豆蛋白比)、酶解温度50℃、酶解时间5 h、酶解pH值6.7。
由于反应系统中物料是循环式的,固液比是1:8时循环不顺畅,所以取固液比1:10。反应器内循环反应条件为固液比是1:10、酶用量5%、酶解温度50℃、酶解pH值6.7,在不加微波辐射条件下酶解时间5 h与用480 W微波辐射反应120 min试验对比,结果表明用微波辐射60 min氨基氮含量就达到了不用微波辐射下反应5 h的氨基氮含量,反应速率提高了5倍以上。研究了微波功率对水解的影响,功率越高水解速率越快。
用液相色谱分析比较了用微波辐射水解1 h和不用微波辐射水解5 h水解液游离氨基酸的含量,结果表明各种游离氨基酸含量和总氨基酸含量基本相同。
最后对反应器的通用性和放大性进行了研究。
本课题研究表明,微波辐射能辅助蛋白酶水解,设计的反应器有优越性,能提高反应速率。
The conventional hydrolyzation reaction takes long time. This paper has mainly researched the protease-hydrolisis reaction in the assistance of the microwave radiation. The study shows that this method of microwave radiation can speed up the reaction rate.
A laboratory scale microwave radiation reactor for protease-hydrolisis was designed and constructed, and the reactor consists of stirring tank or reactor, helixing pipeline reactor radiated by microwave in cavity, liquid circle system, water circle tank for the controlling the reactant liquid and so on. The over heat effect of microwave radiation can be avoided in this circulation type recator. Moreover, power of microwave radiation and the temperature of reactant liquid can be adjusted in this reactor. The stirring tank was designed and computated, The water circulation tank enables the reactant liquid in certain desireable temperature.
The parameters soy protein hydrolyzed by papayin were optimized in this paper prior to microwave radiation reaction system. Firstly, the best hydrolysis conditions of papayin were studied in regular experiments, the content of nitrogen measured by formaldehyde titrated. the optimizated. craft conditions of hydrolysis analysised by the experiment of single factor and the orthogonal test are: the ratio of solid-liquid ,1:8, the contend of papayin, 5%, the temperature of hydrolysis ,50℃, the hydrolysis time, 5 h, the pH value of hydrolys ,6.7. Because the reactor is a circulating system, And the ratio of solid-liquid, 1:8, the cycle is not smooth, so the ratio of solid-liquid, 1:10. In the reactor the reaction conditions are: the ratio of solid-liquid , 1:10, the content of papain ,5%, the temperature of hydrolysis ,50℃, and the pH value of hydrolysis , 6.7, Contrast the result of 5 hour-hydrolysis experiment without microwave radiation to 120 min hydrolysis experiment under 480 W microwave radiation, the result shows that the content of nitrogen generated by 60 min-hydrolysis experiment under microwave radiation reached the content of nitrogen by 5 hour-hydrolysis experiment without microwave radiation, the rate of reaction increases more than five times. The microwave’s power on the impact of hydrolysis were studied, the higher power,the sooner hydrolysis’s rate.
Compared the content of free amino-acid generated by 5 hour-hydrolysis experiment without microwave radiation to 60 min-hydrolysis experiment under 480W-power-microwave radiation by liquid chromatography analysis, The results showed that the content of each free-amino-acid and the total free-amino-acid are basical the same.
Finally, the common nature and enlargement of the reactor were studied.
The research showed that microwave-radiation can assist hydrolysis, the design of the reactor can increase the rate of action.
引文
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2. Baghursl D R.J.Chem. Soc.Chem.Commun, 1992, 9:674
3. Plazl I. Acta Chem.sloy, 1994, 41:437
4. Jahngen E G E.J.Org.Chem, 1990, 55:3406
5. Raner K D and.Strauss C R. J Org Chem, 1992, 57:6231
6. Dayal B, Rapole K R, Shefes S .G. S Tint and S R Wilson, synlett, 1995,861
7.乔晓光,李耀先,刘福安.第一届微波化学学术讨论会论文集[C].长春,1996,63
8. Shibata C, Kashima T and Ohchi K.Jpn.J.App.phys.1996, 35:316
9. Gedye R, Smith F, Westawayetc K. Tetrahedron lett, 1986, 27:279
10. Gedye R, Smith F, West away K.The Rapid Synthesis of Organic Compounds in Microwave Ovens. Can J Chem, 1988, 66:17—26
11.李芳良.对羟基苯甲酸丁酯的快速合成.广西化工,2002,31(2):13—15
12.龙立平,钟桐生,文瑞明等.硫酸氢钠催化合成对羟基苯甲酸丁酯.合成化学,2002,10(1):68—70
13. Majetich G, Hicks R.[J].Radiation Physics and Chemistry,1995, 45 (4):567—579
14. Shieh W C,Dell S, Oljan R. [J].Tetrahedron letters, 2002, 43(32):5607—5609
15.吴明书,张想竹.有机化学,2002,22(2):149—151
16. Moghaddam F M, Ghaffarzadeh M, Adbi—Oskoui S H.J.Chem.Rm, Synop.1999,(9):574
17.周金梅,林敏等.微波法合成乙酰苯胺.厦门大学学报:自然科学版,2003,42(5):679—681
18. Chati S, Bortolussi M, Loupy A. Synthesis of diethers derives from dianhydrohexitols.By phase transfer catalysis under microwave [J]. Tetrahedro Let,2000, 41(18):3367—3370
19.李军,庞军,曹国荣等.合成化学,2000,8(4):321—325
20.孟志霞,郭成,戴冲之.微波快速合成1,5-二芳基-1,4-戊二烯-3-酮.化学时刊,2003,17(9):38—40
21. Mohammad M M , Mohammad R S, Mohammad B, et al.Microwave assisted selective oxidation of benzylic alcohols with calcium hypochlorite under solventfree conditions [J]. Monatshefte for Chemie, 2001,1: 655—658
22.张青山,陆宝萍,郭炳南.微波辐射促进腈类化合物水解[J].火炸药学报,2003,03:74—76
23.李伟章,恽榴江,王好山.有机化学,2002,2 (7):511—514
24. Jin J, Wen Z, Long J.One-pot diazo coupling reation under microwave irradiation in theabsence of solvent [J].Synth.Conmmun, 2000, 30(5):829—834
25.刘成峰,翁家宝,郑雪琳,孙萃玉.微波辐射下快速合成聚磷酸双酚A酯[J];福建化工; 2003, 01:1—3
26.张竞清,车曾轮.合成化学,2002,10:175—176
27. Lin G L , Lin W Y.Microwave-Promoted Lipase-Catalyzed Reactions.Tetrhedron Letter, 1998, 39:4333—4336
28. Arker M C,Besson T ,Lamare S,Legoy M D.Microwave Radiation can increase the rate of enzyme-catalysed reactions in organic media.tetrahedron letter, 1996, 37(46);8383—8386
29 .Vacek M,Zarevucka M,Wimmer Z.Selective enzymic esterification of free fatty acids with n-butanol under microwave irradiation and under classicai heating .Biotechnology letter,2000,22;1565—1570
30.蔡汉成,高惠,方银军,夏永梅,蒋陈兰,方云。微波辐射-酶耦合催化(MIECC)戊醇同分异构体与n-辛酸的酯化反应。化学学报,2004,62(9),923—928
31.陈小兵,李丽,王邵,汪华,毕先钧.微波催化技术的应用研究发展,辽宁石油化工大学学报,2006,26(4),15—17
32.吴加根.谷物与大豆食品工艺学.中国轻工出版社,1996:420—422
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