不同陈化时间广陈皮中黄酮类成分的UPLC-Q-Orbitrap HRMS分析

Analysis and Identification of Flavonoids in Citri Reticulatae Pericarpium with Different Aging Time by UPLC-Q-Orbitrap HRMS

目的:应用超高效液相色谱-四极杆-静电场轨道阱高分辨质谱法(UPLC-Q-Orbitrap HRMS)技术分析不同陈化时间广陈皮中的黄酮类成分。方法:采用Agilent Extend-C18色谱柱(3.0 mm×100 mm,1.8μm),流动相0.1%乙酸水溶液(A)-0.1%乙酸甲醇溶液(B)梯度洗脱(0~25 min,5%~95%B;25~30 min,95%B;30~30.1 min,95%~5%B;30.1~35 min,5%B),流速0.4 m L·min^(-1),柱温30℃。高分辨质谱采用电喷雾离子源(ESI),正、负离子切换模式扫描,扫描方式为全扫描/数据依赖二级扫描(Full MS/dd-MS2)。通过多级离子碎片信息结合mz Cloud网络数据库,中药成分高分辨质谱数据库(OTCML),对照品比对及文献数据进行准确定性。结果:共鉴定43种黄酮类成分,其中黄酮类24种,黄酮醇类5种,二氢黄酮类13种,查尔酮类1种。不同陈化时间广陈皮的黄酮类成分在物质种类上较为一致,但峰面积存在差异。依据峰面积比较相对含量,发现30种黄酮类成分的相对含量随陈化时间增加总体呈增加趋势,其中橙皮苷,diosmin,6-去甲氧基橘皮素,川陈皮素,橘皮素等24种黄酮类成分相对含量增加明显;柚皮素、新橙皮苷等其他13种黄酮类成分相对含量随陈化时间增加总体无明显变化。结论:该方法可全面、准确、高效地测定不同陈化时间广陈皮的黄酮类成分及其相对含量变化,可为广陈皮药效物质基础、质量控制研究提供方法学参考,并为阐释陈皮"陈久者良"提供实验依据。

Objective:To analyze and identify the flavonoids of Citri Reticulatae Pericarpium with different aging time by an ultra-performance liquid chromatography-quadrupole-electrostatic field orbitrap high resolution mass spectrometry(UPLC-Q-Orbitrap HRMS).Method:Compounds were separated on Agilent Extend-C18column(3.0 mm×100 mm,1.8μm),mobile phase was 0.1%acetic acid aqueous solution(A)-0.1%acetic acid methanol solution(B)for gradient elution(0-25 min,5%-95%B;25-30 min,95%B;30-30.1 min,95%-5%B;30.1-35 min,5%B),the flow rate was 0.4 m L·min^(-1),and the column temperature was set at 30℃.High resolution mass spectrometry was performed with electrospray ionization(ESI),and scanned in positive and negative ion modes by means of full scan/data dependent secondary scan(Full MS/dd-MS2).The multistage ion fragment information combined with mz Cloud network database,local high resolution mass spectrometry database of traditional Chinese medicine components(OTCML),literature information and relevant reference materials were used for accurate qualitative analysis.Result:Totally 43 flavonoids in Citri Reticulatae Pericarpium were identified,including 24 flavones,5 flavonols,13 dihydroflavones and 1 chalcone.The flavonoids in samples with different aging time were basically consistent in material types,but the peak area was different.According to the comparison of relative content in the peak area,it was found that the relative contents of 30 flavonoids showed an overall increasing trend with the increase of aging time.Among them,the relative contents of 24 flavonoids(such as hesperidin,diosmin,6-demethoxytangeretin,nobiletin and tangeretin)increased significantly.There was no significant change in the relative contents of the other 13 flavonoids(such as naringenin and neohesperidin).Conclusion:An efficient method is established in this paper to identify flavonoids in Citri Reticulatae Pericarpium with different aging time and their relative content changes rapidly and accurately.The findings provide a methodological reference for the study on pharmacodynamic material base and quality control of Citri Reticulatae Pericarpium,and it provides experimental basis that drugs processed long time ago have better effect of Citri Reticulatae Pericarpium.