新型有机-硅胶杂化分子印迹整体柱电色谱特异识别酪氨酸磷酸化多肽的研究

Tyrosine phosphorylated protein could be used to indicate several disease. The establishment of a specific identification for tyrosine-phosphorylated peptide (pTyr-P ), could be used to monitor the health data of the representative species for the risk assessment of the environment pollution. The aim of this project is to prepare a function monolith and develop the specific recognition of pTyr-P by capillary electrochromatography (CEC). By means of the technologies such as epitope imprinting, affinity interaction of phosphate titanium and organic-silica hybrid monolith, a novel functional molecular imprinting monolith could be obtained and the corresponding capillary electrochromatography with high performance could be established. Many advantages may be found on the prepared hybrid silica monolith, such as high rigidity, stable structure of molecular imprinted site, strong interaction between titanium ion and phosphate group, and less competition of the solvent. The problems existing in the conventional organic polymer such as easy swelling and high interference cause by water will be well avoid. It is favorable for the highly specific recognition of pTyr-P. Via the resultant monolith, CEC with on-line enrichment function is studied by using electrostacking and chromatographic retention on imprinted stationary phase. Coupled with electrochemiluminescence (ECL) detection, a sensitive CEC-LIF hyphenated technology would be established and a further selective enrichment and effective analysis of trace pTyr-P would be implemented. It is meaningful to offer an alternative technology for monitoring the environment pollution and health hazards. Finally, satisfactory result including 5-8 research paper will be expected.

酪氨酸磷酸化蛋白与疾病发生密切相关。建立高效的酪氨酸磷酸化多肽（pTyr-P）特异识别方法，藉此快速分析生物中酪氨酸磷酸化蛋白水平，对污染暴露下生物的健康数据或污染危害风险评估十分重要。项目拟致力于pTyr-P的功能整体柱及其电色谱特异识别方法的研究。通过抗原决定基印迹识别、磷酸酯钛亲和作用和有机-硅胶杂化聚合技术的系统应用，创新研发高稳定性的有机-硅胶杂化抗原决定基印迹识别整体柱，所得聚合整体柱骨架刚性强、印迹位点结构稳定、磷酸酯钛与模板分子磷酸基作用强，溶剂干扰小，避免了有机聚合物水相中易溶胀、水分子干扰大的问题，对强极性pTyr-P水相识别特异性、稳定性高；在此基础上，创新研发了集电场堆积、固定相印迹吸附和电色谱分析的新型在线富集技术，联用高灵敏电致化学发光检测技术，实现了低丰度pTyr-P的选择性富集和高效分析，为环境污染危害健康的风险监测提供新技术。拟发表论文5-8篇。

酪氨酸磷酸化蛋白与疾病发生密切相关。建立高效的酪氨酸磷酸化多肽（pTyr-P）特异识别方法，藉此快速分析生物中酪氨酸磷酸化蛋白水平，对污染暴露下生物的健康数据或污染危害风险评估十分重要。项目从“功能化整体柱制备”-“作用机制研究”-“特异识别分析”着手，研发了磷酸基修饰聚合整体柱、金属离子螯合苯基磷酸的抗原决定基分子印迹整体柱，研制了磷酸氢二（甲基丙烯酰氧乙基）酯（BMEP）或磷酸甲基丙烯酸乙二醇酯（EGMP）修饰的磷酸化聚合整体柱，以Ti4+/Zn2+离子螯合苯基磷酸，制备了poly（TMOS-VTMS-co-EGMP-Ti-PPA）、poly（TMOS-VTMS-co- ZnA-PPA）苯基磷酸的分子印迹杂化硅胶整体柱，研制了高稳定性的有机-硅胶杂化抗原决定基印迹识别整体柱，开展了磷酸化肽、酪氨酸磷酸化肽的选择性识别分析，并探索了分子印迹整体柱对模板分子识别作用的预测性和交互选择性，分析了苯基磷酸抗原决定基识别特性，实现了对磷酸化肽的选择性分离和酪氨酸磷酸化肽（pTyr-P）的有效富集，部分相关成果发表了SCI论文6篇、获得国家发明专利授权2项，为酪氨酸磷酸化肽亲和识别整体柱的制备及作用机制累积了相关的科学数据。