超高效液相色谱法测定不同产地桑叶中4种主要黄酮含量

Determination of 4 flavonoids in mulberry leaves from different habitats by ultra performance liquid chromatography

目的建立超高效液相色谱(ultra performance liquid chromatography, UPLC)测定桑叶中4种主要黄酮含量的方法,并通过主成分和热图分析比较10个不同产地桑叶中4种黄酮含量的差异。方法选择ACQUITY UPLC BEH C_(18)为色谱柱(2.1 mm×50 mm, 1.7μm),甲醇溶液和0.5%磷酸溶液分别作为流动相A和B进行梯度洗脱。检测波长设置为350 nm;柱温设置为30℃。采用TB tools软件进行主成分分析和热图分析。结果 4种黄酮对照品在相应浓度范围内均呈良好的线性关系(r>0.999);回收率为98%~102%,相对标准偏差为1.12%~1.86%。且10个产地桑叶4种黄酮含量差异较大,具体表现为:安徽地区桑叶中芦丁、异槲皮苷、紫云英苷和槲皮素含量均最高,分别为20.543、6.801、1.328、3.645 mg/g;湖南桑叶中异槲皮苷的含量最低,为3.902 mg/g;其余3种黄酮化合物均在河北地区含量最低,分别为芦丁10.021 mg/g、紫云英苷0.348 mg/g和槲皮素1.784 mg/g。主成分和聚类分析结果显示,桑叶具有明显的地域差异性。结论该方法操作简单,分析时间短,稳定性强,可大大降低分析成本,且不同产地桑叶质量差异明显,可作为桑叶用药来源选择的参考。

Objective To establish a method for the determination of 4main flavonoids in mulberry leaves by ultra performance liquid chromatography(UPLC),and to compare the content of 4 flavonoids in mulberry leaves from 10 different areas by principal component analysis and thermography.Methods ACQUITY UPLC BEH C18 column(2.1 mm×50 mm,1.7μm)was used.Methanol solution and 0.5%phosphoric acid solution were used as mobile phases for gradient elution.The detection wavelength was set at 350 nm and the column temperature was 30℃.TB tools software was used for principal component analysis and heat map analysis.Results The 4 flavonoids showed good linear relationship in the corresponding concentration ranges(r>0.999).The recoveries were 98%‒102%,and the relative standard deviations were 1.12%‒1.86%.The results showed that the content of rutin,isoquercitrin,astragaloside and quercetin in mulberry leaves from Anhui province wasthe highest,which was 20.543,6.801,1.328 and 3.645mg/g,respectively.The content of isoquercitrin in Hunan mulberry leaves was the lowest,which was 3.902 mg/g,andthe other 3 flavonoids were the lowest in Hebei,with rutin 10.021 mg/g,astragaloside 0.348 mg/g and quercetin 1.784 mg/g.The results of principal component analysis and cluster analysis showed that mulberry leaves had obvious regional differences.Conclusion This method has the advantages of simple operation,short analysis time and strong stability,which can greatly reduce the cost of analysis,and the quality of mulberry leaves from different areas has obvious differences,which can be used as a reference for the selection of mulberry leaves.