Università degli Studi di Milano, DiSTAM - Dipartimento di Scienze e Tecnologie Alimentari e Microbiologiche, Via Celoria 2, 20133 Milan, Italy.
Rapid Commun Mass Spectrom. 2011 Jun 15;25(11):1675-82. doi: 10.1002/rcm.5025.
The purified extract of propolis is used as a traditional remedy for the treatment of several diseases. Its beneficial activities are mainly attributed to the polyphenolic fraction. Nevertheless, propolis can cause allergic dermatitis and the sensitization rate in humans is increasing significantly mainly in younger subjects. The aim of this study was to develop and validate a selective and sensitive ultra-performance liquid chromatography tandem mass spectrometry analysis (UPLC/MS/MS) for the evaluation of the amount of caffeic acid and its esters with allergenic action in raw propolis samples and commercial formulations. The separation was carried out on a 1.7 μm C(18) BEH Shield column and the detection performed by means of electrospray ionization in negative ion mode with multiple reaction monitoring. The confirmation of formulae of the precursor and product ions was accomplished by injection into a high-resolution system (FTICR-MS) using accurate mass measurements. The error was below 1.4 ppm.The range of the standard curves was 0.5-10 μg/mL and dihydrocaffeic acid was used as internal standard (IS). The lower limit of detection (LLOD) for 3-methyl-2-butenyl-(3M2B), 3-methyl-3-butenyl-(3M3B), 2-methyl-2-butenyl-(2M2B), benzyl-(CABE), phenylethylcaffeic acid (CAPE) and for caffeic acid (CA) and the IS was 0.1 and 0.3 μg/mL, respectively. The recoveries were in the range 96-104% and the intra- and inter-day precisions were within 6.2%. In the European (n=8) and Asiatic (n=3) propolis the most abundant allergens were CABE>3M2B>CAPE>3M3B>CA>2M2B. These compounds were not found in the red (n=1) and green (n=1) Brazilian propolis. Hydroalcoholic extracts (n=6) and tablets (n=6) were analyzed by the proposed UPLC/MS/MS method. The results showed that in the commercial products CABE, 3M2B, CAPE and 3M3B were also the most abundant.
蜂胶的提纯提取物被用作治疗多种疾病的传统药物。其有益的活性主要归因于多酚类成分。然而，蜂胶可能引起过敏性皮炎，而且人类的致敏率正在显著上升，尤其是在年轻人群中。本研究的目的是开发并验证一种超高效液相色谱串联质谱分析（UPLC/MS/MS）方法，用于评估生蜂胶样品和商业配方中具有致敏作用的咖啡酸及其酯的含量。分离在 1.7μm C(18) BEH Shield 柱上进行，通过电喷雾电离在负离子模式下进行检测，并采用多反应监测进行检测。通过在使用精确质量测量的高分辨率系统（FTICR-MS）中注入前体和产物离子的公式来完成公式的确证。误差低于 1.4ppm。标准曲线的范围为 0.5-10μg/mL，二氢咖啡酸用作内标（IS）。3-甲基-2-丁烯基-（3M2B）、3-甲基-3-丁烯基-（3M3B）、2-甲基-2-丁烯基-（2M2B）、苄基-（CABE）、苯乙基咖啡酸（CAPE）和咖啡酸（CA）的检测限（LLOQ）和 IS 分别为 0.1 和 0.3μg/mL。回收率在 96-104%范围内，日内和日间精密度均在 6.2%以内。在欧洲（n=8）和亚洲（n=3）蜂胶中，最丰富的过敏原是 CABE>3M2B>CAPE>3M3B>CA>2M2B。这些化合物在红色（n=1）和绿色（n=1）巴西蜂胶中未被发现。水醇提取物（n=6）和片剂（n=6）用所提出的 UPLC/MS/MS 方法进行分析。结果表明，在商业产品中，CABE、3M2B、CAPE 和 3M3B 也是最丰富的。
