【作者】 王龙耀；

【导师】 雷爱祖； 童张法；

【作者基本信息】 广西大学 ， 化学工艺， 2004， 硕士

【摘要】 在5′-三磷酸腺苷(ATP)生产过程中，开发出高效合理的ATP合成方法、ATP的检测方法与ATP的分离提纯方法是获得高质量、低成本ATP产品的关键。本文针对以上三个问题分别进行了理论探讨和实验研究，系统地总结了固定化酶／细胞法合成ATP的方法，建立了ATP的分析方法和ATP的分离与提纯方法。 固定化酶／细胞法具有反应器效能高、主要原材料成本低、收率高及产品的后处理简单等显著优点，并且可以实现连续生产，因此近年来得到较快发展，并成为ATP生产的主要方法。该方法的中心内容包括：适宜的生物催化活性物质；适宜的固定化形式；适宜的反应器及反应操作条件。本文分别对上述内容进行了分析和总结，并对今后的研究开发提出了建议。 ATP的制备、存储过程中，易混入或产生—磷酸腺苷(AMP)、二磷酸腺苷(ADP)等杂质。由于AMP，ADP和ATP的结构相似，许多理化性质相近，因此对三者的分离或选择性检出成为建立ATP分析方法的关键。本文建立了色谱层析法来测定ATP含量，色谱条件为：自制色谱柱(6×500mm，D201树脂)，流动相为pH3.0的乙酸缓冲液，检测波长254nm，流速10ml／min，柱温23±2℃。检测重复性较好，在0.1～8g／L范围内，ATP浓度与峰面积呈良好线性关系，相关系数r=0.9977；平均回收率为98.6％，整个色谱过程可在30min内完成。该方法也可用于AOp和AMp的分离检测。 通过对多种吸附剂和离子交换树脂进行筛选，获得了一种对5’-三磷酸腺昔具有优良交换解析性能的大孔强碱性阴离子交换树脂D201，测定了ATP在该树脂上的离子交换平衡数据，并研究了其对ATp的离子交换特性。在应用0201树脂分离ATP的过程中，cI一型0201树脂具有比较高的吸附容量;洗脱时以PH3.0、lmol/L的Nacl一溶液作为洗脱剂较为合适;并且树脂的ATP高装载量有利于ATP的解吸分离及浓缩。该离子交换树脂对ATP的交换容量达到0.3269/g湿树脂，整个工艺路线对ATP的洗脱收率达到92.1%。 通过对ATP模拟液在丙酮一水、乙醇一水体系中的溶析性能研究，获得了ATP的结晶工艺条件.在3:1(V/V)乙醇一水体系、10℃、ATp浓度409/L条件下，可获得ATP晶体，收率95%。 最后，对ATp生产过程进行了成本与经济核算，对ATP的工业化生产进行了展望。更多还原

【Abstract】 In the production process of adenosine triphosphate (ATP), the method of synthesis, test, separation and purification ATP play an important role. In this paper, the method of ATP biosynthesis has been discussed with theory. The analytic technique and separation and purification technique has been studied with experiment.The biosynthesis of ATP by immobilized enzyme/cells have many advantages, such as high efficiency of reactor, low costing of main raw materials, high yield and so on. So it has been developed in recent years and become the major production method of ATP. In this method, the key techniques are the appropriate enzyme/cells of ATP biosynthesis, the appropriate method of immobilization, the appropriate reactor & operational conditions. The developing trend about it was discussed, and it has advised to the research in the future.In the process of preparation and storage, it is easy to intermingle or creation impurity as AMP, ADP. Because AMP, ADP and ATP has similar structure and property, how to separate or selective detection them is akey technique in the analytic procedure of ATP. A chromatogram method has been founded to assay ATP with conditions as below: chromatographic column(6 X 500mm, D201 resin), mobile phase ( acetic acid buffer solution, pH3.0), detection wavelength 254nm, velocity of flow 10ml/min, column temperature 23 2 C. In the range of 0.1~8g/L, it has well linear relationship between the concentration of ATP and peak area , r=0.9977. Average recovery is 98.6%. The detection process can be finished in 30min. The method has well intra-laboratory reproducibility, and can be used to separate and detect ADP or AMP.From various ion-exchange resins, a kind of macroporous anion-exchange resin D201 was selected to separate ATP. The ion-exchange isotherm of ATP on D201 resin was measured. Types of resin and the choice of washing conditions in the process of ATP separation were investigated in detail. Furthermore, the ion-exchange rate curves and the break-through curves of ATP solution in ion-exchange column were measured. In the process of separate ATP, the D201 resin with Cl- type has high adsorption capacity. It is appropriate with pH3.0, 1mol/L NaCl solution to elute ATP. The higher ATP load on resin is benefit for ATP decoration and concentration. Adsorbance of ATP on D201 resin was up to 0.326g/g (resin) and the yield for ATP in whole process was 92.1%.From the study on ATP dissolution property in acetone-water andalcohol-water, process conditions of ATP crystallization can be acquired. In the process conditions as alcohol-water(3:1, V/V), 10 C, 40g/L ATP solution, the yield of ATP crystallization is 95%.Finally, the costing and economic balance of producing ATP has been evaluated. The developing trend about it was discussed.更多还原