高效液相色谱紫外等吸收波长法快速测定秦皮中秦皮甲素和秦皮乙素  

Rapid determination of aesculin and aesculetin in Fraxini Cortex by high performance liquid chromatography-ultraviolet at equal absorption wavelength

作　　者：钱正明 吴梦奇 谭国英 金李玲 李宁 谢菊英[1] QIAN Zhengming;WU Mengqi;TAN Guoying;JIN Liling;LI Ning;XIE Juying(College of Medical Imaging Laboratory and Rehabilitation,Xiangnan University,Chenzhou 423000,China;Dongguan HEC Cordyceps R&D Co.,Ltd.,Dongguan 523850,China;College of Traditional Chinese Medicine,Yunnan University of Chinese Medicine,Kunming 650500,China)

机构地区：[1]湘南学院医学影像检验与康复学院,湖南郴州423000 [2]东莞市东阳光冬虫夏草研发有限公司,广东东莞523850 [3]云南中医药大学中药学院,云南昆明650500

出　　处：《色谱》2023年第8期690-697,共8页Chinese Journal of Chromatography

基　　金：广东省药品监督管理局科技创新项目(2021ZDB05)。

摘　　要：研究建立了一种快速、环保和节约对照品的秦皮化学成分含量测定方法。秦皮样品采用25%(v/v)乙醇水溶液超声提取5 min。样品提取溶液采用Poroshell 120 EC-C18色谱柱(50 mm×4.6 mm, 2.7μm)进行分析,0.1%甲酸水溶液-乙腈(94∶6, v/v)等度洗脱,流速1.5 mL/min。采用紫外分光光度仪对等浓度的秦皮甲素和秦皮乙素进行紫外光谱扫描,初步筛选得到2个对照品的等吸收波长,并用高效液相色谱仪对筛选得到的等吸收波长进行确证,从而获得2个对照品的等吸收检测波长341 nm。新建立的高效液相色谱紫外等吸收波长法的方法验证结果显示,秦皮甲素和秦皮乙素在各自的浓度范围内具有良好的线性关系(r=1.000 0),检出限和定量限分别为1.5μmol/L和3.0μmol/L,平均加标回收率分别为99.0%(RSD=1.4%)和97.5%(RSD=0.9%)。通过比较高效液相色谱紫外等吸收波长法和传统高效液相色谱外标法对10批秦皮样品中秦皮甲素和秦皮乙素的含量测定结果,显示两种方法测定结果一致。此外还采用t检验对该方法与《中国药典》2020版方法的含量测定结果进行比较,对比结果表明两种测定方法无显著性差异(P>0.05)。该方法耗时10 min,有害溶剂消耗0.5 mL,只使用1个对照品,具有快速、简便、环保和对照品消耗少的特点,适用于秦皮中秦皮甲素和秦皮乙素的含量测定,为秦皮质量评价的技术提升提供了依据。Fraxini Cortex is a traditional Chinese herbal medicine that has been used for thousands of years to treat dampness-heat diarrhea,dysentery,red or white vaginal discharge,painful swelling or redness of the eyes,and nebula.It contains various chemical components,including coumarins,iridoids,phenolic acids,and flavonoids.Coumarins are important active ingredients in Fraxini Cortex and have antibacterial,anti-inflammatory,antioxidant,antitumor,and antiviral activities.Aesculin and aesculetin are two major coumarin components of Fraxini Cortex that are widely used in its quality evaluation.Previous HPLC methods for determination of aesculin and aesculetin present several limitations,such as long analysis times and high solvent and reference compound consumption.In this study,a rapid,eco-friendly and cost saving HPLC method for the determination of aesculin and aesculetin in Fraxini Cortex was established by using the core-shell column and equal absorption wavelength(EAW).Different factors influencing the extraction process,such as the extraction solvent,temperature,and time,were assessed to obtain the optimal extraction conditions.The results showed that Fraxini Cortex samples could be well extracted by ultrasonic extraction for 5 min with a 25%ethanol aqueous solution.A core-shell column was used,and different mobile phases and flow rates were investigated to obtain the best rapid-HPLC separation conditions.The optimized HPLC conditions were as follows:a Poroshell 120 EC-C 18 column(50 mm×4.6 mm,2.7μm),acetonitrile-0.1%formic acid aqueous solution(6∶94,v/v)as the eluent,a flow rate of 1.5 mL/min,and a column temperature of 25℃.The EAW of aesculin and aesculetin was a key factor in their determination using a single reference compound.EAW selection was performed in two steps.First,the UV spectra of two equimolar concentrations of the reference compounds(aesculin and aesculetin)were compared to determine the EAW of the two analytes.The EAW results were then verified by the HPLC analysis of the reference compound

关 键 词：高效液相色谱 等吸收波长 核壳型色谱柱 秦皮甲素 秦皮乙素 秦皮 

分 类 号：O658[理学—分析化学]