摘要
目的建立黄精芡实汤复合多糖(HQD-P)的质量控制方法并验证其药理活性。方法提取纯化15批HQD-P,采用水提醇沉法提取复方总多糖,进行相关方法学研究,确定HQD-P含量。液相色谱法测定HQD-P的单糖组成,柱温:40℃,流动相:磷酸盐缓冲溶液(A)-乙腈(B);洗脱程序:0~60 min,85%A;Cell Counting Kit-8法对多糖作用后的细胞体外存活率测定,优选HQD-P溶液中的最佳浓度。结果HQD-P中由D-甘露糖、D-核糖、鼠李糖、D-半乳糖醛酸、D-无水葡萄糖、D-半乳糖和L(+)阿拉伯糖7种单糖组成。CCK-8法测定HQD-P的0~500μg/ml各浓度的试验组均不影响肝细胞存活率,其中500μg/ml浓度组促进细胞活力的效果最明显。流式细胞技术分析结果显示HQD-P明显降低细胞内ROS水平,抑制肝细胞凋亡(P<0.05);荧光显微镜分析结果显示HQD-P明显改善线粒体膜电位水平(P<0.05)。结论探讨优选的测定方法稳定可靠,可以作为一种HQD-P的质量控制方法。
Objective To establish a quality control method for Huangjing Qianshi Decoction polysaccharides(HQD-P)and verify its pharmacological activity.Methods Fifteen batches of HQD-P were extracted and purified,and the total compound polysaccharides were extracted using water extraction and alcohol precipitation methods.Relevant methodological studies were conducted to determine the content of HQD-P.Determination of monosaccharide composition of HQD-P by liquid chromatography.column temperature:40℃;mobile phase:phosphate buffer solution(A)-acetonitrile(B)as mobile phase;Elution procedure:0-60 min,85%A;the Cell Counting Kit-8 method was used to determine in vitro survival rate of cells after polysaccharide treatment,and the optimal concentration in HQD-P solution was selected.Results HQD-P was composed of seven monosaccharides:D-mannose,D-ribose,xylose,D-galactouronic acid,D-anhydrous glucose,D-galactose,and L(+)arabinose.The CCK-8 method was used to determine the survival rate of liver cells in the experimental groups of HQD-P at concentrations ranging from 0 to 500μg/ml.Among them,the 500μg/ml concentration group showed the most significant effect on promoting cell viability.Results of flow cytometry analysis showed that HQD-P significantly reduced intracellular ROS levels and inhibited hepatocyte apoptosis(P<0.05);Results of fluorescence microscopy analysis showed that HQD-P significantly improved the mitochondrial membrane potential level(P<0.05).Conclusion This article explores the stability and reliability of the preferred determination method,which can be used as a quality control method for HQD-P.
作者
朱贻霖
蔡嘉洛
李晓屏
周芳
肖望重
欧阳林旗
孙相如
邓桂明
ZHU Yi-lin;CAI Jia-luo;LI Xiao-ping;ZHOU Fang;XIAO Wang-zhong;OUYANG Lin-qi;SUN Xiang-ru;DENG Gui-ming(First Hospital Afiliated to Hunan University of Chinese Medicine,Changsha 410000,China;Hunan University of Chinese Medicine,Changsha 410208,China)
出处
《时珍国医国药》
北大核心
2025年第2期280-288,共9页
Lishizhen Medicine and Materia Medica Research
基金
湖南省临床医疗技术创新引导项目(2021SK51408)
湖南省中医药科研计划一般项目(D2022091)
湖南省自然科学基金科药联合项目(2023JJ60486)
湖南省教育厅优秀青年项目(23B0374)
湖南省临床医学研究中心(基地)项目(2023SK4061)
湖南省卫生健康委科研计划项目(B202303066210)。
关键词
黄精芡实汤复合多糖
水提醇沉法
液相色谱法
质量控制
Huangjing Qianshi Decoction polysaccharides
Water extraction and alcohol precipitation method
Liquid chromatography method
Quality control
