基于超高效液相色谱-紫外检测定量指纹图谱结合化学模式识别的复方金钱草颗粒质量评价

复方金钱草颗粒具有利尿、抑制泌尿系结石形成、抗炎、抗氧化作用,且具有较大的市场需求。因此,采用超高效液相色谱-紫外检测(UPLC-UV)法建立定量指纹图谱,并结合化学模式识别技术对不同年份的复方金钱草颗粒进行质量评价,可为其质量控制提供依据。采用聚类分析(HCA)和主成分分析(PCA)等化学模式识别技术对35批复方金钱草颗粒样品的指纹图谱数据进行分析,筛选出质量差异标志物芒果苷和异芒果苷,并对二者进行含量测定。在复方金钱草颗粒指纹图谱中共指认出12个共有峰,且35批样品的相似度均在0.952以上。在HCA中,将35批样品分为了两类,其中2018年和2019年的样品为一类,2020年和2021年的样品为一类。此外,PCA结果显示了与聚类分析相同的聚类趋势。在此基础上,进一步通过正交偏最小二乘法分析 (OPLS-DA)筛选出了导致2018年、2019年与2020年、2021年的样品产生差异的差异标志物芒果苷和异芒果苷。以两个差异标志物芒果苷和异芒果苷为指标进行含量测定,结果显示色谱峰的分离度良好,线性关系良好,平均加标回收率分别为101.7%~105.6%和103.4%~105.5%,且相对标准偏差(RSD)均低于1.43%。在35批样品中,2020年、2021年的样品与2018年、2019年的样品相比,芒果苷与异芒果苷含量更高且波动范围更小。该研究建立了准确、可靠的复方金钱草颗粒质控方法,实现了对不同年份的复方金钱草颗粒样品合理、有效的质量评价,可为建立更系统、更全面的质量控制标准提供借鉴与参考。

Quality evaluation of Fufang Jinqiancao granules based on ultra performance liquid chromatography-ultraviolet detection quantitative fingerprint combined with chemical pattern recognition

Fufang Jinqiancao granules have a large market demand due to the fact that they contain diuretics, inhibit urinary calculi formation, and exhibit both anti-inflammatory and antioxidant effects. In the current study, a fast and efficient quantitative ultra performance liquid chromatography-ultraviolet detection (UPLC-UV) fingerprinting method was established to analyze the Fufang Jinqiancao granules, while a chemical pattern recognition technology was used to evaluate the quality of the granules over different years. More specifically, the UPLC-UV system consisted of a Waters Acquity UPLC BEH C18 column (100 mm×2.1 mm, 1.7 μm), acetonitrile (mobile phase A), and a 0.1% formic acid aqueous solution (mobile phase B), wherein a gradient elution protocol was followed. Ultra performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF-MS, Agilent Infinity Ⅱ 1290-6545) was used in combination with reference substances and literature comparisons to identify the common peaks present in the quantitative fingerprint. The fingerprints of 35 batches of Fufang Jinqiancao granules were established by means of the quantitative UPLC-UV fingerprinting method, and the fingerprint data obtained for these samples were further analyzed by chemical pattern recognition techniques, including hierarchical cluster analysis (HCA) and principal component analysis (PCA). The quality difference markers, namely mangiferin and isomangiferin, were screened, and their contents were determined. It was found that 12 common peaks existed in the fingerprint of the Fufang Jinqiancao granules, and the similarities of all 35 batches of samples were greater than 0.952. In addition, for the purpose of HCA, the 35 batches were divided into two categories, of which sample years 2018 and 2019 belonged to one category, and sample years 2020 and 2021 belonged to another category. Notably, PCA gave the same clustering trends as HCA. Based on the obtained results, the mangiferin and isomangiferin components responsible for the differences between the 2018, 2019 and 2020, 2021 samples were further screened by orthogonal partial least squares discriminant analysis (OPLS-DA). Moreover, the contents of the 35 batches of samples were determined using the two differential markers mangiferin and isomangiferin as indicators. The obtained results indicated that the chromatographic peaks of all 35 batches had acceptable resolutions, with mangiferin exhibiting a good linear relationship in the range of 5.3291-133.2276 mg/L, and isomangiferin exhibiting a similar linear relationship in the range of 4.1847-104.6170 mg/L. The average recovery of mangiferin was 101.7%-105.6%, with a relative standard deviation (RSD) of 0.63%-1.43%, while that of isomangiferin was 103.4%-105.5%, with an RSD of 0.60%-1.18%. Importantly, all RSD values were less than 1.43%, thereby indicating that our method meets the requirements of the Chinese Pharmacopoeia (2020 Edition). Among the 35 batches of samples, the contents of mangiferin and isomangiferin were higher in the 2020 and 2021 samples, and the content fluctuation range was smaller. Overall, the development of an accurate and reliable quality control method for Fufang Jinqiancao granules, and a reasonable and effective quality evaluation of Fufang Jinqiancao granule samples from different years was realized. We therefore expect that this study will provide a reference for establishing a more systematic and comprehensive quality control system.