College of Chemistry and Pharmaceutical Sciences, Qingdao Agricultural University, Qingdao 266109, China.
College of Veterinary Medicine, Qingdao Agricultural University, Qingdao 266109, China.
J Chromatogr Sci. 2021 Apr 21;59(5):412-418. doi: 10.1093/chromsci/bmab021.
Calycosin and formononetin were efficiently extracted from Astragali Radix and purified by high-speed countercurrent chromatography. Calycosin and formononetin could be hydrolyzed from calycosin-7-glucoside and ononin, respectively. The best extraction conditions were realized by single factor and orthogonal experiments, which were 100% ethanol, 2.5 mol/L hydrochloric acid, 1:40 ratio of solid to liquid, extracted 2 h and one time. The two-phase solvent system of n-hexane-ethyl acetate-ethanol-water (3:5:3:5, v/v) was selected for the purification of calycosin, and 1.3 mg calycosin (the purity was 95.8% and the recovery was 85.9%) was obtained from 264.9-mg crude extraction. The two-phase solvent system of n-hexane-ethyl acetate-ethanol-water (4:5:4:5, v/v) was selected for the purification of formononetin, and 2.0 mg formononetin (the purity was 98.9% and the recovery was 84.4%) was obtained from 248.9-mg crude extraction. Their structures were identified by HPLC, melting points, UV, FTIR, ESI-MS, 1H NMR and 13C NMR spectrum. According to the antioxidant activity assay, the scavenging abilities of calycosin to 1,1-diphenyl-2-picrylhydrazyl and hydroxyl free radicals (·OH) were stronger. The scavenging effect of formononetin was not demonstrated.
毛蕊异黄酮和芒柄花素可从黄芪中高效提取，并通过高速逆流色谱法进行纯化。毛蕊异黄酮苷和芒柄花苷可分别水解为毛蕊异黄酮和芒柄花素。通过单因素和正交实验确定了最佳提取条件，即 100%乙醇、2.5mol/L 盐酸、固液比 1:40、提取 2 小时、提取 1 次。选择正己烷-乙酸乙酯-乙醇-水（3:5:3:5，v/v）两相溶剂系统对毛蕊异黄酮进行纯化，从 264.9mg 粗提物中获得 1.3mg 毛蕊异黄酮（纯度为 95.8%，回收率为 85.9%）。选择正己烷-乙酸乙酯-乙醇-水（4:5:4:5，v/v）两相溶剂系统对芒柄花素进行纯化，从 248.9mg 粗提物中获得 2.0mg 芒柄花素（纯度为 98.9%，回收率为 84.4%）。通过 HPLC、熔点、UV、FTIR、ESI-MS、1H NMR 和 13C NMR 谱对其结构进行了鉴定。根据抗氧化活性测定，毛蕊异黄酮对 1,1-二苯基-2-苦基肼基（DPPH）自由基和羟基自由基（·OH）的清除能力更强。芒柄花素的清除效果则不明显。
