Determination of green tea catechins in human plasma using liquid chromatography–electrospray ionization mass spectrometry

Abstract

A method for the sensitive and specific determination of eight green tea catechins, consisting of catechin (C), epicatechin (EC), gallocatechin (GC), epigallocatechin (EGC), catechin-3-gallate (CG), epicatechin-3-gallate (ECG), gallocatechin-3-gallate (GCG) and epigallocatechin-3-gallate (EGCG), in human plasma was established. For optimization of conditions for LC-ESIMS, the separation of the eight catechins was achieved chromatographically using Inertsil ODS-2 column combined with a gradient elution system of 0.1M aqueous acetic acid and 0.1M acetic acid in acetonitrile. Detection using a mass spectrometer was performed with selected ion monitoring at m/z=289 for E and EC, 305 for GC and EGC, 441 for CG and ECG, and 457 for GCG and EGCG under negative ESI. A preparative procedure, consisting of the addition of perchloric acid and acetonitrile to the plasma for deproteinizing and the subsequent addition of potassium carbonate solution to remove excess acid, was developed. In six different plasma with the eight catechins spiked at two different concentrations, the average recoveries were in the range between 72.7 and 84.1%, which resulted from the matrix effect and preparative loss, with coefficients of variance being 8.2-19.8% among individuals. The levels of the catechins in prepared plasma solutions that were kept at 5 degrees C within 24h were stable, which allows us to simply analyze many prepared plasma solutions using an autosampler overnight. When using this method to analyze the eight catechins in human plasma after oral ingestion of a commercial green tea beverage, we detected all the catechins absorbed into human blood for the first time. This also suggested that extremely small amounts of the eight catechins orally ingested may be absorbed based on each absorptive property for the catechins. The method should enable pharmacokinetic studies of green tea catechins in humans.