Urpi-Sarda Mireia, Monagas Maria, Khan Nasiruddin, Lamuela-Raventos Rosa M, Santos-Buelga Celestino, Sacanella Emilio, Castell Margarida, Permanyer Joan, Andres-Lacueva Cristina
Nutrition and Food Science Department, XaRTA, INSA, Faculty of Pharmacy, University of Barcelona, 08028 Barcelona, Spain.
Anal Bioanal Chem. 2009 Jul;394(6):1545-56. doi: 10.1007/s00216-009-2676-1. Epub 2009 Mar 31.
Proanthocyanidins, flavonoids exhibiting cardiovascular protection, constitute a major fraction of the flavonoid ingested in the human diet. Although they are poorly absorbed, they are metabolized by the intestinal microbiota into various phenolic acids. An analytical method, based on an optimized 96-well plate solid-phase extraction (SPE) procedure and liquid chromatography tandem mass spectrometry (SPE-LC-MS/MS) for the analysis of 19 phenolic microbial metabolites and monomeric and dimeric flavanols in urine samples, was developed and validated. Human urine samples were obtained before and after ingestion of an acute consumption of 40 g of soluble cocoa powder and rat urines before and after the prolonged administration (2 weeks) of different diets composed of natural cocoa powder. The mean recovery of analytes using the new SPE-LC-MS/MS method ranged from 87% to 109%. Accuracy ranged from 87.5% to 113.8%, and precision met acceptance criteria (<15% relative standard deviation). Procyanidin B2 has been detected and quantified for the first time in human and rat urine after cocoa consumption. Changes in human and rat urinary levels of microbial phenolic acids and flavanols were in the range of 0.001-59.43 nmol/mg creatinine and of 0.004-181.56 nmol/mg creatinine, respectively. Major advantages of the method developed include reduction of laboratory work in the sample preparation step by the use of 96-well SPE plates and the sensitive measurement of a large number of metabolites in a very short run time, which makes it ideal for use in epidemiological studies.
原花青素是一类具有心血管保护作用的黄酮类化合物，是人类饮食中摄入的黄酮类化合物的主要成分。尽管它们的吸收率很低，但它们会被肠道微生物群代谢成各种酚酸。本文开发并验证了一种基于优化的96孔板固相萃取（SPE）程序和液相色谱串联质谱（SPE-LC-MS/MS）的分析方法，用于分析尿液样本中的19种酚类微生物代谢物以及单体和二聚体黄烷醇。在摄入40克可溶性可可粉后，采集了人类尿液样本；在长期（2周）给予由天然可可粉组成的不同饮食后，采集了大鼠尿液样本。使用新的SPE-LC-MS/MS方法分析物的平均回收率在87%至109%之间。准确度在87.5%至113.8%之间，精密度符合验收标准（相对标准偏差<15%）。食用可可后，首次在人类和大鼠尿液中检测并定量了原花青素B2。人类和大鼠尿液中微生物酚酸和黄烷醇水平的变化范围分别为0.001 - 59.43 nmol/mg肌酐和0.004 - 181.56 nmol/mg肌酐。所开发方法的主要优点包括使用96孔SPE板减少了样品制备步骤中的实验室工作量，以及在非常短的运行时间内对大量代谢物进行灵敏测量，这使其非常适合用于流行病学研究。
