酶法拆分N-乙酰-D,L-蛋氨酸转化条件优化

重组大肠杆菌BL21/pET22b-argE表达的N-乙酰鸟氨酸脱酰基酶(NAOase)可用于脂肪族氨基酸的手性拆分.水解N-乙酰-D,L-氨基酸中的L-型底物,N-乙酰-D,L-蛋氨酸为其最合适的底物.为了确定NAOase拆分N-乙酰-D,L-蛋氨酸合适的转化条件,考察了反应温度、pH值、Co2+浓度、转化时间、底物浓度和加酶量对产物的影响.结果表明,合适的反应条件为37℃,pH值7.0,Co2+ 1 mmol/L,转化时间20 min,底物浓度150 mmol/L,菌泥5 g/L.在上述反应条件下,N-乙酰-D,L-蛋氨酸的转化率可达81.4%.     

氨基酰化酶拆分制备手性蛋氨酸工艺条件研究

以N-乙酰DL-蛋氨酸(N-Ac-DL-Met)为拆分底物,对氨基酰化酶法拆分工艺参数如反应温度、pH、激活剂Co2 浓度以及底物浓度等因素进行了初步考察。结果表明,在反应温度37℃,缓冲体系pH7.5,Co2 浓度为0.5 mmol/L,初始底物N-Ac-DL-Met的浓度为0.06 mol/L,氨基酰化酶加入量为5μg/mL的较佳条件下,L-蛋氨酸(L-Met)的产率可达45%;产物L-Met结晶的光学纯度(用对映体过量值%e.e表示)为95.4%。实验中首次采用多组分标准曲线法,提高了茚三酮显色法在氨基酰化酶法拆分制备手性氨基酸的检测精确度     

固定化米曲霉氨基酰化酶反应动力学及其连续拆分DL–蛋氨酸实验

探讨了用明胶-戊二醛固定化米曲霉菌球氨基酰化酶反应动力学,对固定化菌体连续光学拆分DL-蛋氨酸进行了研究. 当底物浓度小于200 mmol/L时,没有底物抑制现象,此时拆分反应速率符合Michaelis-Menten方程. 在37℃时, 米氏常数和最大反应速率分别为11.9 mmol/L和1.3 mmol/L. 在连续拆分反应中反应物体积流量为7.5 ml/h,底物浓度为200和400 mmol/L时,L-蛋氨酸的转化率分别为93%和78%. 随体积流量的增加,L-蛋氨酸转化率降低. 固定化菌体的操作半衰期为82 d.     

酶法动态动力学拆分制备R-(-)-乙酰基邻氯扁桃酸

采用假单胞菌脂肪酶Pseudomonas sp. ECU1011催化乙酰基邻氯扁桃酸进行不对称水解,利用突变后的扁桃酸消旋酶(V29I)对拆分后的产物S-(-)-邻氯扁桃酸进行消旋,消旋后的邻氯扁桃酸经过酰化重新被利用到水解反应中,实现了酶法动态动力学拆分制备R-(-)-乙酰基邻氯扁桃酸。通过对拆分反应、拆分混合物的分离回收以及消旋反应的工艺优化,最终获得光学纯度ee>99.9%的R-(-)-乙酰基邻氯扁桃酸,其收率达80%。本研究建立的R-(-)-乙酰基邻氯扁桃酸的动态动力学拆分工艺,对其工业化应用具有重要的指导意义。     

重组氨基酰化酶1与N-乙酰鸟氨酸脱乙酰酶酶学性质比较

利用基因工程手段重组表达了牛肾来源的氨基酰化酶1(ACY1),并与实验室之前构建的N-乙酰鸟氨酸脱乙酰酶(NAO)进行比较,以N-乙酰-DL-蛋氨酸为底物,研究了其酶学性质,考察了pH、温度、反应时间、表面活性剂、金属离子等因素对拆分DL-蛋氨酸的影响。结果表明,重组NAO和ACY1拆分300 mmol/L的N-乙酰-DL-蛋氨酸所需时间分别是2 h和6 h;重组NAO的酶活及N-乙酰-L-蛋氨酸的转化率分别为1 139.41 U/g和98%,高于重组ACY1的671.24 U/g和58.78%,约是重组ACY1的1.7倍。     

固定化米曲霉氨基酰化酶反应动力学及其连续拆分DL-蛋氨酸实验

探讨了用明胶－戊二醛固定化米曲霉菌球氨基酰化酶反应动力学，对固定化菌体连续光学拆分DL－蛋氨酸进行了研究。当底物浓度小于200mmol/L时，没有底物抑制现象，此时拆分反应速度符合Michaelis-Menten方程。在37℃时，米氏常数和最大反应速率分别为11.9mmol/L和1.3mmol/L。在连续拆分反应中反应物体积流量为7.5ml/h，底物浓度为200和400mmol/L时，L－蛋氨酸的转化率分别为93％和78％。随体积流量的增加，L－蛋氨酸转化率降低。固定化菌体的操作半衰期为82d。     

雷美替胺的合成路线改进

以1,2,6,7-四氢-8H-茚并[5,4-b]呋喃-8-酮为原料,经缩合、氢化两步得到外消旋中间体。再通过N-乙酰基-L-苯丙氨酸拆分获得S-型异构体中间体,最后通过丙酰氯试剂酰化生成雷美替胺原料药。改进后的工艺操作简便,条件温和,中间体和产物反应终点易于监控,总收率为13%,适合工业化生产。     

L-脯氨酸不对称转换的研究

L-脯氨酸在正丁醛或水杨醛催化作用下,可在羧酸中消旋。对L-脯氨酸在不同溶剂、温度、催化剂、反应时问下的消旋作了研究。采用不对称转换的方法,S-酒石酸作拆分剂,正丁醛作催化剂,正丁酸作溶剂,经过D-脯氨酸酒石酸盐,合成D-脯氨酸。在最佳工艺条件下的收率为92．4％,光学纯度为100％。     

L-二苯甲酰酒石酸为拆分剂结晶拆分L,D-薄荷醇

以L-酒石酸为原料,合成并纯化了结晶拆分剂L-二苯酒石酸;考察了L-二苯酒石酸结晶拆分过程的动力学控制,L-薄荷醇的收率提高23%。同时,研究了结晶拆分过程的溶剂影响。结果表明,用氯化锌代替氯化铁作为催化剂,L-二苯酒石酸的合成温度可降低12℃,重结晶纯化只需一次,可达到拆分剂的要求;L-二苯酒石酸结晶拆分消旋体薄荷醇喜好弱极性的组合溶剂（环己烷/水=80：20,V/V）,单一的高极性和非极性溶剂对结晶拆分不利。同时,展望了未来研究方向。     

L-氮杂环丁烷-2-羧酸合成工艺研究进展

L-氮杂环丁烷-2-羧酸作为一种重要的医药中间体,被广泛应用于制备医药产品、有机手性化合物等。本文综述了通过对外消旋体进行消旋或拆分的方法制备L-氮杂环丁烷-2-羧酸的合成路线,并对以上工艺路线的特点进行了分析评价。     

常压CO2气流法制D,L-丙交酯

在ＣＯ２气流中用Ｄ，Ｌ－乳酸为原料，硫酸亚锡为催化剂，包括内外消旋体的粗丙交酯产率可达８２％，外消旋Ｄ，Ｌ－丙交酯约占其中的１／２。作为对比，用Ｎ２气流时的粗丙产酯产率为８３．２％，二者产率相差无几，可用ＣＯ２代替Ｎ２用于丙交酯的制备。     

常压CO2气流法制D，L-丙交酯

在CO     

Simulierte Gegenstromchromatographie

Simulated Countercurrent Chromatography. This paper reviews simulated countercurrent chromatography, a separation process that has received only little attention. In contrast to standard batchwise operated chromatography, a periodical mode of operation is used. The feed and collection lines are shifted along the column, thus simulating countercurrent operation of the liquid phase and the stationary solid phase. As a result, higher product yield and higher product purity are obtained. At the same time the amount of solid phase and solvent phase required in comparison to discontinuous chromatography is reduced. Industrial applications have been developed since 1964 by Universal Oil Products (UOP). Large scale separation processes in the petrochemical and the sugar industries are already in operation. On the bench scale this process is used to separate biotechnological products and pharmaceuticals. The need to solve difficult separation problems (isomeric or racemic mixtures, biomolecules, polymers) will encourage further developments. Together with improved process-simulation tools and process control, simulated countercurrent chromatography will be established as a powerful separation technique with widespread industrial applicability.     

酶促酯化拆分-化学消旋串联法制备S-布洛芬

以消旋布洛芬为反应底物，探究了酶促酯化拆分布洛芬的工艺。利用南极假丝酵母脂肪酶B的对映体选择性催化布洛芬选择性酯化，并对醇的种类、反应介质、底物摩尔比、酶量、干燥剂种类和添加量、温度及时间等反应条件进行了优化，以56%的收率得到了光学纯的S-布洛芬，接下来对酯化产物进行化学法消旋。以二甲亚砜为反应介质，利用二异丙基氨基锂催化布洛芬发生外消旋反应，完全消旋后再进行酶促拆分仍能达到之前的效果。如此循环7次布洛芬的利用率可达到90%以上。     

动力学控制结晶法拆分1,1''''-联二萘酚工艺研究

对外消旋1,1'-联二萘酚(BINOL)的拆分进行了研究.采用动力学控制结晶的方法改进了(S)-2-吡咯烷酮-5-羧酰苯胺拆分BINOL的工艺,得到一种实用有效的获得光学纯BINOL的方法.采用该方法可以同时得到BINOL的R和S两种异构体,它们的ee值均大于99%.采用正交实验的方法研究了反应时间、拆分剂用量、溶剂组成以及溶剂体积在拆分过程中对(R)和(S)-BINOL的产率的影响.在最佳条件下,(R)-BINOL和(S)-BINOL的收率分别为18.0%和35.2%,外消旋BINOL的回收率为58.8%,拆分剂的回收率为68.6%.     

Enantiomer-selective activation of racemic catalysts

Asymmetric catalysts can be evolved into highly activated catalysts by association with chiral activators. This asymmetric activation process is particularly useful in racemic catalysis through selective activation of one enantiomer of the racemic catalysts. Recently, a strategy whereby a racemic catalyst is selectively deactivated by a chiral additive has been reported to yield nonracemic products. However, we have reported a strategy that is an alternative to asymmetric catalysts but is conceptually opposite, in which a chiral activator selectively activates rather than deactivates one enantiomer of a racemic chiral catalyst. The advantage of this activation strategy over the deactivation counterpart is that the activated catalyst can produce a greater enantiomeric excess (x(act)% ee) in the products than the ee attained by the enantiomerically pure catalyst on its own. Therefore, 'asymmetric activation' could provide a general and powerful strategy for the use of not only atropisomeric and, hence, racemic ligands but also chirally flexible and 'pro-atropisomeric' ligands without enantiomeric resolution!     

C1—5醇乳酸酯及丙交酯制备的初步研究

用Ｃ１－５醇酯化Ｄ,－Ｌ乳酸制得相应的乳酸酯,其中用Ｃ４－５醇酯化乳酸不用带水剂,产率可达８０％以上,用Ｄ,Ｌ－乳酸酯制丙交酯会生成等量的外消旋与内消旋体。外消旋丙交酯为目标产物,内消旋丙交酯可回收作原料。由此计算外消旋丙交酯单程产率将不大于５０％,实验中除乳酸甲酯外,乳酸酯单程消耗和丙交酯单程产率分别为２６％－３５％和２２％－３５％。     

Diastereomeric salt crystallization of chiral molecules via sequential coupled-Batch operation

A novel process, sequential coupled-batch diastereomeric crystallization is presented for the resolution of racemic mixtures. This strategy utilized a preferential crystallization of diastereomeric salt followed by sequential coupled diastereomeric salt resolution of racemic ibuprofen with the resolving agent (S)-lysine utilizing a novel small scale pneumatic liquid exchange design to couple two batch crystallizers. Results were compared to conventional sequential resolution via seeded isothermal batch crystallization. Diastereomeric salt resolution was developed using the ternary phase diagram of the isolated (R)-ibuprofen-(S)-lysine and (S)-ibuprofen-(S)-lysine salts and optimized empirically with the aid of process analytical technology. Sequential coupled-batch crystallization was found to be capable of increasing the yield of both salts while maintaining purity. This gave an increase in both the productivity and yield (20.5% for (S)-ibuprofen-(S)-lysine) compared to the equivalent sequential batch (17.7% for (S)-ibuprofen-(S)-lysine) crystallization methodology. Single crystals of the (S)-ibuprofen-(S)-lysine were isolated, and its single crystal structure determined. © 2018 American Institute of Chemical Engineers AIChE J, 65: 604–616, 2019     

重组大肠杆菌全细胞用于苯乙酮酸的绿色生物合成

苯乙酮酸是化学合成中重要的合成砌块,可用于合成多种药物中间体,探索苯乙酮酸的绿色合成工艺具有重要的意义。以包含D-扁桃酸脱氢酶LhDMDH编码基因的重组大肠杆菌全细胞为催化剂,考察了它在无辅酶和辅底物添加的条件下对D-扁桃酸生物转化的效果,并对催化产物进行了纯化和鉴定。结果表明,本研究成功实现了在无辅酶和辅底物添加条件下苯乙酮酸的生物合成,产物的得率和纯度分别为45%和99%左右。成果也为外消旋扁桃酸的手性拆分及苯乙酮酸的生物合成奠定了基础。     

Employing racemic precursors in directed biosynthesis of 6‐dEB analogs

Substitution of racemic diketide thioesters for optically pure compounds in precursor‐fed fermentations was investigated. These substrates were shown to be as effective as optically pure materials in diketide‐fed fermentation processes for producing analogs of 6‐deoxyerythronolide B. However, since half of the racemic mixture is not utilizable for polyketide biosynthesis, higher total levels of diketide are required. Toxicity to cells was evident at high diketide concentrations. In fermenters, exhaust gas analysis was used to indicate the optimal time for diketide addition. While both enantiomers were shown to disappear from cultures at similar rates, the presence of unincorporated enantiomer had minimal effect on polyketide production within a range of feed concentrations. Use of racemic diketide thioesters was successful and dramatically reduced the cost of the fermentation process. © 2002 Society of Chemical Industry