高效液相色谱-四极杆飞行时间质谱检测中国杨树型蜂胶、巴西绿蜂胶和杨树胶中的酚类化合物及真伪鉴别

Determination of phenolic compounds in Chinese poplar propolis,Brazil green propolis,and poplar gum by high performance liquid chromatography-quadrupole-time-of-flight mass spectrometry and preliminary study of the identification of adulteration

建立了用于检测中国杨树型蜂胶、巴西绿蜂胶和杨树胶中23种酚类化合物的高效液相色谱-四极杆飞行时间质谱(HPLC/Q-TOF MS)法,并进行了差异性分析。蜂胶和树胶样品用甲醇-水(1∶1,v/v)溶解后,过0. 45μm有机相滤膜后进样。采用Agilent Eclipse Plus C18色谱柱分离,以乙腈和0. 1%(体积分数)甲酸水溶液为流动相进行梯度洗脱,电喷雾正离子模式全扫描方式检测,扫描范围为m/z 100～1 000,外标法定量。结果表明,23种酚类化合物在10～200μg/L范围内线性关系良好,相关系数均大于0. 99。桔皮素和刺芒柄花素的检出限和定量限分别为0. 2和1μg/L,其他化合物的检出限和定量限分别为2和10μg/L。在10、25和50 m g/kg 3个添加水平下,23种酚类化合物在3种样品基质中的提取回收率为70. 2%～122. 6%,RSD值均小于10%。水杨苷、肉桂酸、咖啡酸和香豆酸可以作为中国杨树型蜂胶掺假鉴别的特征化合物,咖啡酸、阿魏酸、白杨素、咖啡酸苯乙酯、松属素、高良姜素、香豆酸、异鼠李素、山柰素和阿替匹林C可以作为辨别中国杨树型蜂胶和巴西绿蜂胶的特征化合物。该结果对蜂胶产品的质量控制具有一定的参考意义。

High-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry(HPLC/Q-TOF MS)was used to analyze 23 phenolic compounds in Chinese poplar propolis,Brazil green propolis,and poplar gum.Propolis and poplar gum samples were dissolved in methanol-water(1∶1,v/v)and filtered by 0.45μm organic phase filtration membrane.The separation was carried out in an Agilent Eclipse Plus C18 column with gradient elution using acetoni-trile and 0.1%(volume percentage)formic acid solution as the mobile phases.The compounds were detected using positive ion electrospray ionization in full scan mode(m/z 100-1000).The quantification analysis was performed by the external standard method.The results showed that all the compounds were linear,with correlation coefficients>0.99,in the range of 10-20μg/L.The limits of detection and limits of quantification for tangeretin and formononetin were 0.2 and 1μg/L,respectively,and those for the other compounds were 2 and 10μg/L,respectively.At the spiked levels of 10,25,and 50 mg/kg,the recoveries of all the compounds ranged from 70.2%to 122.6%,with relative standard deviations of less than 10%.Salicin,cinnamic acid,caffeic acid,and coumaric acid could be used as markers for detecting adulteration in Chinese poplar propolis,while caffeic acid,ferulic acid,chrysin,caffeic acid phenethyl ester,pinocembrin,galangin,coumaric acid,isorhamnetin,kaempferide,and arte-pillin C could be used as markers for identifying Chinese poplar propolis and Brazil green propolis.The results presented in this paper may be helpful in quality control of propolis products.