【作者】 柴逸峰；

【导师】 吴玉田； 殷学平；

【作者基本信息】 第二军医大学 ， 药物分析学， 2002， 博士

【摘要】 手性药物的生物活性与其立体构型密切相关。目前新药研究的一个发展趋势是研制和生产光学纯的药物。手性药物的研究已成为国际新药研究的新方向之一，手性药物分析在手性药物研究中作用越来越重要，已成为国际上分析科学中的热点和难点。目前在手性药物分析中主要采用间接分析和直接分析。直接分析简便、快速、准确和可靠，其中HPLC法如今占主导地位，毛细管电泳法呈上升趋势。毛细管电泳是近20年来迅速发展起来的一种高效、快速、微量的分离分析方法。 在毛细管电泳手性分析中，一般是将手性选择剂(又称手性添加剂)加入到分离缓冲液中，提供一个手性环境，由于一对对映体与手性选择剂的作用强弱有差异，导致各自不同的电泳淌度，从而达到手性分离。环糊精是应用最为广泛的一种选择剂，目前的CE的手性拆分研究主要集中于不同类型的环糊精与不同类型的对映体药物之间CE拆分的实验研究，缺少机理和系统理论研究。 本文采用分子对接方法，确定环糊精和对映体包结过程，进而采用分子力学计算结合能，通过计算机化的分子模型技术从微观角度观察环糊精与对映体的包结作用机制，研究手性识别机理，并用对映体之间结合能的差异，结合动态结合的实际情况，预测手性拆分的可能性，并用CE实验验证。研究得到结果1．研究提出环糊精手性识别有别于著名的三点理论，为“多点结合，一点决定”。环糊精识别通过内腔包含和外部结合，与对映体形成不同的结构和性质的可逆性包结物，内腔的包含可以是多点结合，外部的结合可以是一点或多点，若能达到手性分离，至少在外部结合点上有一点作用必须有差异，而此作用点可以是手性碳上直接结合的基团，也可以是整个侧链基团。2．提出可以用环糊精手性拆分的对映体通常具有两个主要结构特征：(1)疏水骨架，芳香族化合物，如苯、丹磺酰、β-萘酰胺、巴比妥酸酯等等，其主要目的在于提高分子与环糊 摘 耍 ＿精的结合能。c）手性原子上连接有大的立体基团，保证有一定的范德华力立体 －作用。这保证了手性分离的异构体之间存在一定的结合能差异，从而揭示手性 分离的本质。此外，还有两个辅助因素：即疏水骨架上存在氢键供受体时，或、侧链上存在可以与环糊精形成氢键的基团，可以辅助性起到手性判别的作用。＿3．研究表明环糊精包结对映体的结合能和结合能的差异值与实验拆分有一定的－－相关性。4．考察了CE手性分析的主要影响因素，实验表明影响大小依次为， CDS种类、pH、CDS的浓度和BGE的离子强度。 同时，从药物分析的角度和按其要求，开展了CE异构体药物分析应用研＿究。建立了混合取代基乃环糊精－－毛细管电泳法分离测定麻黄碱和伪麻黄碱，－对映体L．舒必利中D．型杂质检查，CD－MEKC法测定山莱莫中齐墩果酸和熊果，酸的含量，羟丙基．e．环糊精－毛细管电泳法分离测定日夜百服宁片剂中的伪麻＿黄碱，以及TM顺反异构体的分离分析方法。CE在异构体药物分析应用中分别－选择了对映体麻黄碱、伪麻黄碱、舒必利；同分异构体麻黄碱和伪麻黄碱，熊 果酸和齐墩果酸：顺反异构体TM，系统的进行了分析方法建立和优化的研究，＿＿并注意发挥CE的优势，上述所建立的方法涉及光学纯药物的杂质检查，原料＿＿药、中药、复方西药及化学中间体的含量测定法。为目前CE在复杂对象的药＿”物分析中应用，提供了有益的示例和理论依据。拓宽了CE在异构体药物分析＿中的应用范围。应用研究中舒必利光学纯杂质检查的研究资料通过了药监局的．审评。。－此外，本文合成5种6CD衍生物作为CE手性选择剂，为该类试剂研制－和国产化，提供了有益的尝试。同时，依据本实验和本学科相关工作的积累，＿－提出和建立CE在药物分析中研究和开发的一般策略。更多还原

【Abstract】 Bioactivity of chiral drugs is closely related to the stereo structures. Nowadays the production and usage of optically pure drugs is the trend of new drugs research. The study on chiral drugs has become one of new directions of new drugs research internationally. Analysis of chiral drugs,with increasing importance in pharmaceutical research,is now a hot topic in international analysis science.At present analysis of chiral drugs is mainly achieved by two approaches:direct and indirect enantioseparation. Direct enantioseparation is simple,rapid,accurate and reliable. High performance liquid chromatography (HPLC) plays a leading role in direct enantioseparation,while the applications of capillary electrophoresis (CE) are increasing significantly. CE,developed rapidly hi last two decades,is a fast separation technique with high efficiency and sensitivity.In CE enantioseparation,the chiral selector (or additive) is dissolved in the running buffer to provide chiral environment. So the two enantiomers can be separated owing to the difference of the interactions between chiral selectors and enantiomers and thus the difference of electrophoretic mobilities. Cyclodextrins (CDs) are the most widely used chiral selectors. Presently CE enantioseparation is focused on experimental research between different lands of cyclodextrins and enantiomers,while lacking of mechanism research systematically.In this report,the course of host-guest inclusion was determined by means of molecular docking and thus association energy was calculated by molecular mechanics. With the computer-assisted molecular modeling technique,the inclusion mechanism between CDs and enantiomers was microscopically investigated and the theory about chiral recognition was discussed. The results were as follows. Firstly,the mechanism of chiral recognition by CDs involved combination at multi-point and determination by one point. In chiral recognition by CDs,enantiomers and CDs formed reversible inclusion complexes with different structures and properties by inclusion within cavity and complexation outside of CDs. The inclusion can happen at multi-point,while the complexation at one point or multi-point. There must have difference of interaction at one complexation point outside to achieve chiral separation. This point can be the entire side chain or groups attached directly to chiral carbon. Secondly,compounds which can be chirally separated by CDs always have two major structural features:(l) hydrophobic skeleton such as benzene ring,can increase the association energy between molecules and CDs;(2)the large stereo groups connected with the asymmetric center provide certain van der Waals interactions;(3)auxiliary factors:the H-bond receptors on hydrophobic skeleton mayintroduce H-bond interactions with CDs so as to increase the association energy;(4) when the stereo group attached to the asymmetric center can’t supply all energy for recognition,the group on the side chain which can form H-bond with CDs assisted the chiral recognition. The essence of chiral recognition and separation by CDs is owing to side chains,moreover hydrophobia skeleton can prolong the inclusion time and shorten the inclusion distance. Thirdly,it was demonstrated that there existed a certain correlation between the difference of association energy and chiral separation. Lastly,the main factors influencing the CE enantioseparation were investigated. The results showed the influence reduced according to the order:type of CDs,pH,concentration of CDs and ionic strength.Application research on analysis of isomer drugs by CE was carried out at the same time by the angles and demands of pharmaceutical analysis,such as determination of ephedrine and pseudoephedrine by mixed-substituents-p-CD CE,test of D-isomer impurity hi L-sulpiride,assay of oleanolic acid and malol hi fructus corni by CD-MEKC,determination of pseudoephedrine in BaiFuNing tablets,and separation of cis- and trans- TM. Enantiomer like ephedrine,pseudoephedrine and sulphide,isomer like ephedrine and pseudoephedrin更多还原