Chemical analysis of phenolic glycosides: art, facts, and artifacts.

Phenolic glycosides have been the subject of considerable interest in recent ecological and systematic studies, especially those involving the Salicaceae. But these compounds are markedly labile in aqueous media, and the consequences of spontaneous degradation for valid interpretation of results have been largely ignored by researchers. We found that freeze-drying and oven-drying of leaf samples from several Populus and Salix species produced dramatic changes in the total and relative concentrations of specific phenolic glycosides, when compared to analyses of fresh material. Extraction in aqueous and alcoholic media for extended (24 h) periods also effected changes in glycoside concentrations. Alterations in phenolic glycoside concentrations, interconversions among glycosides, and production of artifactual glycosides result from a series of hydrolytic reactions. These deleterious effects can be best (but not entirely) avoided by the use of fresh plant material, cold, nonaqueous extraction solvents, and short extraction times. Because individual phenolic glycosides exhibit very different biological activities against herbivores, we caution ecologists to use utmost care in the performance and interpretation of phenolic glycoside assays.