新型核/壳型手性固定相评价体系的建立及其在氨基酸衍生物药物对映体分离与质量控制中的应用

Chiral drugs and human biological molecules mutual recognition are stereoselectivity, leads to differences in the efficacy and side effects of different enantiomers in the human body. For example, R-thalidomide has sedative, sleep effect, but the S-enantiomer and its metabolites have serious teratogenic effects. Therefore, the separation of enantiomers of chiral drugs has been one of the most important and necessary topics in the pharmaceutical research field. The chiral stationary phase (CSP) method can be used for both analysis and separation,as well as for the preparation and semi-preparation of enantiomers, which is the most striking method for the separation of drug enantiomers, and more than 100 CSPs have been commercialized. However, the chiral recognition mechanism of commercially available chiral columns is mostly unclear and the quantity evaluation system is absent. Our group focused on the study of chiral stationary phase over 20 years. In former experiment, we developed two different configurations of the homochiral polyaniline/silica shell complexes, which have obvious chiral recognition ability for amino acids. Compared with polyaniline, the silica coated core/shell composite got chiral recognition ability enhancement. Base on these results, the program will study a novel category of hybrid core/shell HPLC chiral stationary phase (CSP), which consisting of a unique organic core (homochirality helical polyaniline nanofibers) and functional inorganic shell for enantioseparation. The core of CSP with stereo regularity is homochiral polyaniline nanofibers, which providing a stereo space for chiral recognition. The inorganic shell with functional silica, protecting the core, as well as providing interaction sites of π-π, dipole-dipole, hydrogen-bonding and so on. The enantiosepation ability of different configuration CSP will be evaluated and compared.These new CSPs will be used for the separations of chiral drugs, such as amino acids, β-blocks and other racemates with different mobile phases. Based on the enantioseparation results, 4-6 solutes will be selected as standard substances to build these CSPs assessment model, using selectivity and resolusion as creterion. The chiral recognition mechanism is also investigated. And then the suitable CSPs will be rapidly screened out for the application of enantioseparation and quality control, including quality analysis in vitro and metablism and transfer in vivo of four amino acid derivative drugs, levodopa, levodopa methyl ester, methotrexate and melphalan. The investigation above-mentioned can provide reference for the systematic study of the assessment system and chiral recognition mechanism of other kinds of CSP.

手性固定相（CSP）法是手性药物对映体分离中最引人注目的方法，但CSP手性识别机理大多不明确，且CSP评价缺乏量化标准。本课题组长期聚焦CSP研究，近来针对氨基酸研制出具有明显手性识别能力的两种不同构型单一手性聚苯胺内核/二氧化硅外壳复合物。与聚苯胺相比，经二氧化硅包覆后的核/壳复合物手性识别能力增强。在此基础上，拟将复合物外壳衍生化制备系列核/壳CSP，在不同流动相模式中拆分手性药物对映体。根据手性拆分结果，选择4-6个手性药物为“标准物质”，以“标准物质”的选择因子或分离度值为指标，建立CSP量化评价模型，探讨手性识别机理。基于此新型手性识别机理研究模型，建立左旋多巴等4种氨基酸衍生物药物质量控制方法，应用于它们的对映体分离、制备、手性杂质检查以及体内手性转化研究。本项目为新型CSP的制备、量化评价体系建立及机理研究提供借鉴，也为手性药物体内、外手性分离分析研究提供参考。

立体规整的聚合物是目前应用最多的手性固定相（CSP）。手性聚苯胺(PANI)多采用手性掺杂剂诱导掺杂，主链优先以单一螺旋构型排列而制备得到，且螺旋的手性是由诱导分子的手性决定的。具有规整结构的手性PANI在手性分离中具有潜在的应用价值。.本项目以樟脑磺酸作为诱导酸及掺杂剂，分别在水、有机溶剂、水-有机溶剂中，采用界面聚合法、自组装法、低聚物辅助法、二次掺杂法、化学氧化法等对手性PANI的合成进行研究，通过UV、 FTIR、CD、SEM和XRD等技术手段表征，发现不同方法制备出的手性PANI的形貌、光学活性类似，但溶剂溶剂体系会影响最终产物的光学活性：水溶剂和有机溶剂体系中，得到的聚苯胺纳米纤维的螺旋方向、光学活性相反。.以天冬氨酸和谷氨酸为模板分子PANI分子印迹聚合物，两种氨基酸分子印迹PANI均可用于与模板分子结构相差较大的氨基酸分离。.以D/L-PANI为核，硅胶为壳制备出2种核壳化合物，对丙氨酸和亮氨酸对映体可产生选择性结晶，经不同构型的核壳手性选择剂诱导，优先结晶的氨基酸构型不同。以D/L-PANI为核，3,5-二甲基苯基异氰酸酯衍生化硅胶、3,5-二甲基苯基氨基甲酸酯纤维素分别为壳，制备出3种手性固定相（CSP），对半胱氨酸等10种手性化合物表现出一定程度的对映体分离能力；以硅胶为内核，PANI为中间层，多糖衍生物为外壳，制备出3种“三明治”CSP，对反式氧化吡烯、2,2,2-三氟-1-(9-蒽基)乙醇等手性化合物实现了基线分离；以低聚苯胺（OAIN）和大环抗生素制备出3种复合型CSP，对7种氨基酸显示出了手性识别。.本项目筛选出3种分离效果好的CSP，分别选择2、4种手性化合物为衡量这些CSP手性识别能力的“标准物质”；确定了医药中间体黄烷酮等5种手性物质的基线分离条件，可应用于这些化合物生产过程中的质量控制。.本项目为立体规整聚合物用作手性固定相提供依据，为新型CSP的制备、评价体系建立以及应用提供借鉴。本项目发表论文13篇，申请发明专利2项。获云南省卫生科技成果奖二等奖1项。培养硕士研究生6人，培养在读博士研究生3人、硕士研究生1人。参与课题研究的4名教师获得职称提升。