柴胡是一种传统中药，数千年来被用于治疗湿热腹泻、痢疾、白带异常、眼红肿痛、霰肿等症状。它含有多种化学成分，包括香豆素、萜苷、酚酸和黄酮类化合物。香豆素是柴胡的重要活性成分，具有抗菌、抗炎、抗氧化、抗肿瘤和抗病毒活性。角叶素和角叶酮是柴胡中两种主要的香豆素成分，在其质量评价中得到广泛应用。之前用于角叶素和角叶酮测定的HPLC方法存在一些限制，例如分析时间长、溶剂和参比化合物消耗高。本研究建立了一种快速、环保和节省成本的用于柴胡中角叶素和角叶酮测定的HPLC方法，采用核壳柱和等吸收波长(EAW)技术。评估了影响提取过程的不同因素，如提取溶剂、温度和时间，以获取最佳提取条件。结果表明，柴胡样品可以通过超声波提取5分钟，使用25%乙醇水溶液进行良好提取。使用核壳柱，研究了不同流动相和流速，以获得最佳的快速HPLC分离条件。优化的HPLC条件如下：使用Poroshell 120 EC-C18柱（50 mm×4.6 mm，2.7 μm），乙腈-0.1%甲酸水溶液（6∶94，体积比）作为洗脱剂，流速为1.5 mL/min，柱温为25 ℃。角叶素和角叶酮的EAW是使用单个参比化合物进行测定的关键因素。EAW选择分为两个步骤进行。首先，比较两种参比化合物（角叶素和角叶酮）等摩尔浓度溶液的紫外光谱，确定两种分析物的EAW。然后通过参比化合物溶液的HPLC分析验证EAW结果。角叶素和角叶酮的最终EAW为341 nm。在这个EAW下，使用单个参比化合物（即角叶素）通过HPLC-UV实现了角叶素和角叶酮的测定。新开发的HPLC方法显示出两个目标分析物之间的良好线性关系(r=1.0000)。检测限(LODs)和定量限(LOQs)分别为1.5 μmol/L和3.0 μmol/L，角叶素和角叶酮的平均回收率分别为99.0%和97.5%。对样品溶液的稳定性进行了检验，两种分析物在24小时内表现出良好的稳定性。采用所提出的EAW方法和经典外标法(ESM)测定了10批柴胡的目标分析物含量，并获得了可比较的浓度。10批柴胡中角叶素和角叶酮的含量分别为0.26%-2.80%和0.11%-1.47%。对比所提出的EAW技术与中国药典中报道的方法的结果进行了t检验，两种测定方法之间没有显著差异(P>0.05)。将新开发的EAW方法与文献报道的方法进行比较，我们的方法只需10分钟完成，溶剂消耗仅为0.5 mL，只使用一个标准品。因此，所开发的EAW方法是一种快速、简便、环保和经济有效的适用于柴胡及其相关产品中角叶素和角叶酮测定的分析方法。该方法是改进的角叶素和角叶酮测定方法，有助于提高柴胡的质量评价。

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-C18 column (50 mm×4.6 mm, 2.7 μm), 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 solutions. The final EAW of aesculin and aesculetin was 341 nm. The determination of aesculin and aesculetin using only one reference compound (i. e., aesculin) was achieved by HPLC-UV at this EAW. The newly developed HPLC method revealed a good linear relationship between the two target analytes (r=1.0000). The limits of detection (LODs) and limits of quantification (LOQs) were 1.5 μmol/L and 3.0 μmol/L, respectively, and the average recoveries of aesculin and aesculetin were 99.0% and 97.5%. The stabilities of the sample solutions were examined, and the two analytes demonstrated good stability for 24 h. The contents of the target analytes in 10 batches of Fraxini Cortex were determined using the proposed EAW method and the classic external standard method (ESM), and comparable concentrations were obtained. The contents of aesculin and aesculetin in the 10 batches of Fraxini Cortex were 0.26%-2.80% and 0.11%-1.47%, respectively. A t-test was conducted to compare the results of the proposed EAW technique with those obtained via the method reported in the Chinese Pharmacopoeia, and no significant difference between the two assay methods was noted (P>0.05). Comparison of the newly established EAW method with those reported in the literature revealed that our method required only 10 min to complete and used as little as 0.5 mL of the solvent and only one standard. Therefore, the developed EAW method is a rapid, simple, eco-friendly, and cost-effective analytical method that is suitable for the determination of aesculin and aesculetin in Fraxini Cortex and its related products. The proposed technique is an improved method for determining aesculin and aesculetin and contributes to the enhancement of the quality evaluation of Fraxini Cortex.