Analysis of catechins and condensed tannins by thermally assisted hydrolysis/methylation-GC/MS and by a novel two step methylation

Abstract

The objective of this study is to observe high molecular weight markers of catechins and condensed tannins by thermally assisted hydrolysis/methylation (THM)-GC/MS. Techniques for formation of intact methylated flavanols of catechins using THM in the presence of trimethylsulfonium hydroxide (TMSH), and, of a dimer marker of condensed tannins using a novel two step methylation technique, are presented. The two step methylation procedure involves pre-methylation of the sample with trimethylsilyl diazomethane (TMS-diazomethane) followed by THM. The dimer marker, a methylated product containing the C–C linkage between adjacent flavanol units, has a molecular weight of 540. Intact methylated flavanols of catechins were also successfully observed as high molecular weight compounds including partially methylated catechin and epicatechin (3-flavanol, 3′,4′,5,7-tetramethoxy, cis/trans; m/ z = 346), epigallocatechin and gallocatechin (3-flavanol, 3′,4′,5,5′,7-pentamethoxy, cis/trans; m/ z = 376). These techniques were successfully applied to the analysis of series of condensed tannins isolated from plants, and catechins and other phenolics present in (hot water) extracts of tea leaves. In green tea the major catechins were identified as epicatechin and epigallocatechin along with flavonols and tannin dimers.