程斌 周爱珍 彭昕 王昕蕾 王娟

摘 要 目的：建立浙貝母超高效液相色谱-四极杆-飞行时间串联质谱（UPLC-Q-TOF-MS/MS）指纹图谱，并确定其抗炎质量标志物。方法：色谱柱为Eclipse Plus C18，流动相为甲醇-0.1%甲酸水溶液（梯度洗脱），流速为0.4 mL/min，柱温为30 ℃，进样量为2 ?L;采用电喷雾离子源，以正、负离子模式检测，在质荷比（m/z）50～1 200范围内进行扫描。采用《中药色谱指纹图谱相似度评价系统》（2004A版）建立10批不同产地浙贝母的UPLC-Q-TOF-MS/MS指纹图谱。以小鼠耳肿胀度和血清中丙二醛（MDA）、一氧化氮（NO）水平为抗炎指标，采用双变量相关分析法和灰色关联度分析法对浙贝母指纹图谱中各共有峰相对峰面积与抗炎指标的相关性进行分析，筛选质量标志物并进行结构鉴定。结果：在正、负离子模式下，10批浙贝母药材分别各有共有峰26、10个。综合双变量相关分析和灰色关联度分析结果，共发现与抗炎作用相关的质量标志物9个，经鉴定分别为贝母素乙、贝母素甲、环巴胺、胡萝卜苷、Polyphyllin Ⅴ、苦鬼臼毒素、植物甾醇、Ent-kaur-15-en-17-ol、Ent-17-norkauran-16-one。结论：所建立的UPLC-Q-TOF-MS/MS指纹图谱可用于评价浙贝母药材的质量;贝母素乙、贝母素甲、环巴胺等可能是浙贝母具有抗炎作用的质量标志物。

关键词 浙贝母;抗炎作用;超高效液相色谱-四极杆-飞行时间串联质谱;双变量相关分析;灰色关联度分析;质量标志物

ABSTRACT   OBJECTIVE： To establish UPLC-Q-TOF-MS/MS fingerprint of Fritillaria thunbergii， and to define its anti-inflammatory quality markers. METHODS： The determination was performed on Eclipse Plus C18 column with mobile phase consisted of methanol-0.1% formic acid solution （gradient elution） at the flow rate of 0.4 mL/min. The column temperature was set at 30 ℃， and injection volume was 2 ?L. The electrospray ion source was used to scan in the range of m/z 50-1 200 with positive and negative ion detection mode. UPLC-Q-TOF-MS/MS fingerprints of 10 batches of F. thunbergii from different habitats were established by using the Similarity Evaluation System of TCM Chromatographic Fingerprints （2004A edition）. With ear swelling degree， the serum levels of MDA and NO as anti-inflammatory indexes， the correlation between the relative area of common peaks in fingerprint and the anti-inflammatory indexes was analyzed by using bivariate correlation analysis and grey correlation analysis， and the quality markers were screened and identified. RESULTS： In positive and negative ion mode， 10 batches of F. thunbergii had 26 peaks and 10 peaks. Based on bivariate correlation analysis and grey correlation analysis， nine quality markers related to anti-inflammatory effect were found， which were identified as peiminine， peimine， cyclobalamine， daucidin， polyphyllin Ⅴ， bitumen podophyllotoxin， phytosterol， ent-kaur-15-en-17-ol， ent-17-norkauran-16-one. CONCLUSIONS： Established UPLC-Q- TOF-MS/MS fingerprint can be used to evaluate the quality of F. thunbergii; peiminine， peimine and cyclobalamine and so on may be the quality markers of anti-inflammatory effect of F. thunbergii.

KEYWORDS   Fritillaria thunbergii; Anti-inflammatory effect; UPLC-Q-TOF-MS/MS; Bivariate correlation analysis; Grey correlation analysis; Quality markers

浙贝母是百合科植物浙贝母（Fritillaria thunbergii Miq.）的干燥鳞茎，具有清热、化痰、止咳、解毒、散结、消痈等功效，主治风热犯肺、痰火咳嗽、肺痈、乳痈、瘰疬和疮毒等症[1]。近年来研究表明，浙贝母的化痰、止咳、松弛气管平滑肌、抗炎等作用是其治疗呼吸系统疾病的药理学基础[2]，其中浙贝母化痰、止咳的活性成分已较为明确，但对其抗炎活性成分的研究相对较少，主要集中在生物碱类成分贝母素甲和贝母素乙[3-5]。……