Wang Caiyun, Hu Shuang, Chen Xuan, Bai Xiaohong
School of Pharmacy, Shanxi Medical University, Taiyuan, China.
J Sep Sci. 2016 May;39(10):1814-24. doi: 10.1002/jssc.201600103. Epub 2016 May 2.
Hollow fiber cell fishing, based on HepG-2, SKOV-3, and ACHN cancer cells, and hollow fiber liquid/solid microextraction with HPLC were developed and introduced for researching the anticancer activity of Rhizoma Curcumae Longae, Radix Curcumae, and Rhizoma Curcumae. The structures of curcumin, demethoxycurcumin, and bisdemethoxycurcumin screened were identified and their contents were determined. The compound target fishing factors and cell apoptosis rates under the effect of the three medicines were determined. The binding sites (cell membrane and cell organelle) and binding target (phospholipase C) on the cell were researched. Hollow fiber liquid/solid-phase microextraction mechanism was analyzed and expounded. Before the application, cell seeding time, growth state and survival rate, compound nonspecific binding, positive and negative controls, repeatability in hollow fiber cell fishing with high-performance liquid chromatography; extraction solvent, sample pH, salt concentration, agitation speed, extraction time, temperature and sample volume in hollow fiber liquid/solid-phase microextraction with high-performance liquid chromatography were investigated. The results demonstrated that the proposed strategy is a simple and quick method to identify bioactive compounds at the cellular level as well as determine their contents (particularly trace levels of the bioactive compounds), analyze multicompound and multitarget entirety effects, and elucidate the efficacious material base in traditional medicine.
基于HepG-2、SKOV-3和ACHN癌细胞的中空纤维细胞垂钓法以及结合高效液相色谱的中空纤维液/固微萃取法被开发并引入,用于研究姜黄、郁金和莪术的抗癌活性。对筛选出的姜黄素、去甲氧基姜黄素和双去甲氧基姜黄素的结构进行了鉴定,并测定了它们的含量。测定了三种药物作用下的化合物靶点垂钓因子和细胞凋亡率。研究了细胞上的结合位点(细胞膜和细胞器)以及结合靶点(磷脂酶C)。对中空纤维液/固微萃取机制进行了分析和阐述。在应用前,考察了高效液相色谱中空纤维细胞垂钓法中的细胞接种时间、生长状态和存活率、化合物非特异性结合、阳性和阴性对照、重复性;以及高效液相色谱中空纤维液/固微萃取法中的萃取溶剂、样品pH值、盐浓度、搅拌速度、萃取时间、温度和样品体积。结果表明,所提出的策略是一种在细胞水平上鉴定生物活性化合物及其含量(特别是痕量水平的生物活性化合物)、分析多化合物和多靶点整体效应以及阐明传统药物有效物质基础的简单快速方法。