旋覆花水提物的TLC和HPLC特征图谱及6种化学成分的含量测定OACSTPCD
Establishment of characteristic chromatograms of TLC and HPLC and determination of six components in water-extract of Inula japonica
目的 建立旋覆花水提物的薄层色谱(TLC)和高效液相色谱(HPLC)的特征图谱,同时建立水提物中6种成分(绿原酸、咖啡酸、异槲皮苷、1,5-二咖啡酰奎宁酸、4,5-二咖啡酰奎宁酸和旋覆花内酯)的含量测定方法,评价11批旋覆花药材水提物的相似性.方法 (1)采用HF254型薄层层析硅胶板、正丁醇-丙酮-吡啶-水-冰醋酸(1∶5∶3∶1∶10µL)为展开剂,在365 nm波长下检视喷洒10%硫酸-乙醇前、后的色谱斑点,建立TLC特征图谱.(2)采用Sino-Chrom ODS-BP(4.6 mm×250 mm,5µm)色谱柱,流动相为0.05%磷酸溶液-乙腈的梯度洗脱程序,流速0.8 mL/min,柱温35℃,检测波长205 nm,建立HPLC特征图谱.(3)6种成分的含量测定条件与(2)相同,但检测波长分别为203 nm(旋覆花内酯)、256 nm(异槲皮苷)和327 nm(咖啡酸、绿原酸、1,5-二咖啡酰奎宁酸、4,5-二咖啡酰奎宁酸).结果 (1)喷洒显色剂前、后的TLC图谱均显示出良好的特征性.(2)HPLC特征图谱显示含有28个共有峰,指认出10个峰(绿原酸、咖啡酸、1,5-二咖啡酰奎宁酸、4,5-二咖啡酰奎宁酸、花旗松素、芦丁、槲皮苷、异槲皮苷、木犀草素和旋覆花内酯).通过相似度评价和聚类分析可知11批旋覆花水提物相似度良好.(3)绿原酸、咖啡酸、异槲皮苷、1,5-二咖啡酰奎宁酸、4,5-二咖啡酰奎宁酸和旋覆花内酯6种成分的线性范围分别为3.25～65.00、4.00～80.00、1.60～32.00、17.75～355.00、2.50～50.00和3.75～75.00µg/mL.11批旋覆花水提物中绿原酸、咖啡酸、异槲皮苷、1,5-二咖啡酰奎宁酸、4,5-二咖啡酰奎宁酸和旋覆花内酯的平均含量分别为3.38～6.66、2.13～7.35、0.51～1.57、15.81～24.87、0.50～1.08和0.77～4.73µg/mg.结论 该方法的TLC和HPLC特征图谱分离效果良好,含量测定方法可靠,可为旋覆花水提物质量控制方法提供实验依据.
Objective To establish the characteristic chromatograms of thin-layer chromatography(TLC)and high-performance liquid chromatography(HPLC)of the aqueous-extract of Inula japonica Thunb.,and to determine the contents of six components(chlorogenic acid,caffeic acid,isoquercitrin,1,5-dicaffeoylquinic acid,4,5-dicaffeoylquinic acid and inulicin)in the aqueous-extract.Methods(1)The HF254 TLC silica gel plate,developing solvents of n-butanol-acetone-pyridine-water-glacial acetic acid(1∶5∶3∶1∶10µL)and 10%sulphuric acid-ethanol were used before the chromatographic spots were examined at 365 nm.(2)The Sino Chrom ODS-BP column(ϕ4.6 mm×250 mm,5µm)and a mobile phase of 0.05%phosphoric acid solution and acetonitrile were used.The flow rate was 0.8 mL/min,the column temperature 35℃and detection wavelength 205 nm.(3)The analytic conditions of the six components were similar to those described in method(2),but the wavelengths were respectively 203 nm(inulicin),256 nm(isoquercitrin)and 327 nm(caffeic acid,chlorogenic acid,1,5-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid).Results(1)The TLC chromatograms showed fine properties before and after the colour developer was sprayed.(2)Twenty-eight common peaks were marked in the HPLC characteristic chromatogram,so were ten components(chlorogenic acid,caffeic acid,1,5-dicaffeoylquinic acid,4,5-dicaffeoylquinic acid,taxifolin,rutin,quercitrin,isoquercitrin,luteolin and inulicin).The aqueous-extracts of eleven batches of materials of Inula japonica shared much similarity based on similarity and cluster analysis.(3)There were good linear relationships,and the linear ranges of chlorogenic acid,caffeic acid,isoquercitrin,1,5-dicaffeoylquinic acid,4,5-dicaffeoylquinic acid and inulicin were 3.25-65.00µg/mL,4.00-80.00µg/mL,1.60-32.00µg/mL,17.75-355.00µg/mL,2.50-50.00µg/mL and 3.75-75.00µg/mL,respectively.The contents of chlorogenic acid,caffeic acid,isoquercitrin,1,5-dicaffeoylquinic acid,4,5-dicaffeoylquinic acid and inulicin in elven batches of water-extract from Inula japonica were 3.38-6.66µg/mg,2.13-7.35µg/mg,0.51-1.57µg/mg,15.81-24.87µg/mg,0.50-1.08µg/mg and 0.77-4.73µg/mg,respectively.Conclusion The specific TLC and HPLC characteristic chromatograms can help ensure accurate quantitative determination of six components of the water-extract of Inula japonica,which can be used for quality control of the water-extract of Inula japonica.
孟凡恒;骆芙瑶;冯婷;杨郁;徐锐;朱帅鸣;单俊杰
军事科学院军事医学研究院毒物药物研究所,北京 100850军事科学院军事医学研究院毒物药物研究所,北京 100850||南京中医药大学,南京 210046
中医学
旋覆花水提物薄层色谱高效液相色谱特征图谱定量分析
Inula japonicaaqueous-extractthin-layer chromatographyhigh-performance liquid chromatographycharacteristic chromatogramquantitative analysis
《军事医学》 2024 (003)
202-212 / 11
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