基于理论计算的膝沟藻毒素分子印迹固相萃取机理研究

Recently, poisoning incidents from paralytic shellfish toxins caused by the consumption of shellfish were often reported in China. Gonyautoxins have posed a serious hazard to public health and threatens shellfish aquaculture in the coastal region. Molecularly imprinted solid-phase extraction (MISPE) presents the high specificity, selectivity and sensitivity of the molecular recognition mechanism and the high resolving power of separation methods. However, there has been no reports on the isolation study of gonyautoxins based on MISPE. Molecular recognition and isolation mechanism also need to be investigated. So, solid-phase extraction of gonyautoxins and mechanism study using molecular imprinting technology have important significances to monitor seafood safety and protect the health of consumers.. In this study, a highly selective sample cleanup procedure combined with MISPE will be developed for the isolation of gonyautoxins from Alexandrium minutum extract samples. Molecularly imprinted polymer microspheres will be prepared using the dummy template molecule. The polymer microspheres will be used as a selective sorbent for the solid-phase extraction of GTXs and the MISPE protocols were optimized. An innovative and effective extraction method based on MISPE will be established. In order to get a deeper insight into the recognition mechanism of imprinted polymers adsorbing gonyautoxins and to explain solid-phase extraction process, theoretical calculations based on Artificial Force Induced Reaction of Global Reaction Route Mapping will be employed in the research. The results will offer a sound basis for the high selective isolation of gonyautoxins from the interference matrix. Moreover, the study will open up new ways to effective isolation of marine toxins and promote further study in the field of micro-algae toxins.

我国近海的麻痹性贝毒中毒事件，以膝沟藻毒素居多，分子印迹固相萃取技术在分离纯化领域已被广泛应用，在微藻毒素的提纯中则研究较少，其对毒素的固相萃取机理有待研究。因此，开展膝沟藻毒素的提取纯化研究，阐明其分离机理，对保障我国水产品安全和保护人类健康具有重要的理论和实际意义。.本项目拟以分子印迹技术为基础，制备对膝沟藻毒素具有特异性选择吸附能力的分子印迹微球，结合固相萃取技术，利用自制固相萃取柱，对微小亚历山大藻（Alexandrium minutum）有毒藻株中的膝沟藻毒素GTX1,4和GTX2,3进行提取纯化，通过实验优化固相萃取条件，建立一套有效的膝沟藻毒素分子印迹固相萃取方法，同时运用理论计算全局反应路径图方法中的假想力诱导反应揭示该提纯方法的分子识别机制和固相萃取机理，为分子印迹固相萃取微藻毒素提供理论依据，为微藻毒素的分离纯化开辟新的途径，以推动我国微藻毒素领域的深入研究。

麻痹性贝毒是海洋微藻毒素的一类，分布最广、危害最大，我国近海的PSP中毒事件，以膝沟藻毒素（Gonyautoxins, GTXs）居多。分子印迹固相萃取技术在分离纯化领域已被广泛应用，在微藻毒素的提纯中则研究较少，其对毒素的固相萃取机理有待研究。因此，开展膝沟藻毒素的提取纯化研究，阐明其分离机理，对保障我国水产品安全和保护人类健康具有重要的理论和实际意义。.本项目以分子印迹技术为基础，采用咖啡因和己酮可可碱为虚拟双模板，以聚乙烯醇为分散剂，通过悬浮聚合法，制备对膝沟藻毒素具有特异性选择吸附能力的分子印迹微球；对印迹材料进行表征，考察其吸附性能，优化合成条件，得到分子特异性识别能力较高的印迹材料。然后结合固相萃取技术，对微小亚历山大藻（Alexandrium. minutum）有毒藻株中的膝沟藻毒素GTX1,4和GTX2,3进行分离提取，利用制备的分子印迹微球，经过两次分子印迹固相萃取过程，分别建立GTX1,4和GTX2,3在微藻粗提液中的纯化方法；同时运用全局反应路径图中的假想力诱导反应c初步揭示固相萃取机理，阐明印迹聚合物与膝沟藻毒素之间的分子识别机制，为分子印迹固相萃取技术提供理论依据，为微藻毒素的分离纯化开辟新的途径，以推动我国微藻毒素领域的深入研究。