基于水凝胶3D细胞培养的靶向α1A受体细胞仿生色谱法筛选姜黄中活性成分

细胞膜色谱法（CMC）已得到广泛应用和良好评价。但细胞膜固定相是由破碎的细胞膜重新融合而成，与原细胞膜显著不同，难以模仿药物与细胞的实际作用情况。申请人前期研究发现姜黄提取物可舒张家兔前列腺平滑肌。针对CMC的问题和发现，本项目提出基于活细胞的仿生色谱构想-细胞仿生色谱法。应用细胞生物学技术和色谱技术，利用高表达α1A受体细胞结合琼脂糖凝胶3D培养技术，利用琼脂糖凝胶的温敏效应，制备保持细胞生理活性的细胞仿生固定相；建立靶向α1A受体细胞仿生色谱法，模拟药物与细胞的相互作用。以其为核心技术与LC-MS结合建立在线二维分析系统，提高活性筛选的定性与定量水平。并应用于姜黄中可特异性作用于高表达α1A受体细胞的活性成分筛选；结合分子生物学、离体药理学等技术对活性成分作用进行验证。项目的完成将丰富生物亲和色谱内容；为中药活性成分筛选分析提供新技术和方法。

Cell membrane chromatography (CMC) has been widely used and well evaluated. However, the stationary are reassembled with cell membrane fragments, which causes the receptors distribution on cells distinctly different from the original. So it is difficult to simulate the actual interaction of drugs and cells in vivo. In my previous study, turmeric extract has been found the effect of relaxing the smooth muscle of prostate in rabbits. Aiming at the problem and discovery of CMC, this project proposes a bionic chromatography-cell chromatography method based on living cells. By using cell biology technology and chromatographic technology, the biomimetic cell stationary phase that maintaining the physiological activity of the cells will be prepared with high-expression α1A AR cells and agarose gel three dimensional culture technology. Then the high-expression α1A AR-targeted cell bionic chromatography method will be developed to simulate drug-cell interactions in vivo. To improve the qualitative and quantitative level of cell chromatography for the screening efficiency, the cell chromatography will be hyphenated with LC/MS to develop an online two-dimensional analysis system. Moreover, the method will be used to screen the active components specifically acting on high-expression α1A AR cells from Curcuma longa extract. In combination with molecular biology, the effects of active ingredients were verified using in vitro pharmacology. The completion of the project will enrich the content of bio-affinity chromatography, and provide a new technology and method for the screening and analysis of the active ingredients from traditional Chinese medicine.