Structural elucidation of catechin and epicatechin in sorrel leaf extracts using liquid-chromatography coupled to diode array-, fluorescence-, and mass spectrometric detection.

Flavonoids are natural compounds in medicinal herbs and posses several biological activities important in plant drug design. Especially strong antioxidant effects play an important role against radical oxidative stress causing pathological processes, such as arteriosclerosis or cancer. The aim of this work was to investigate unknown analytes found in sorrel leaf (Rumex acetosa) extracts in order to discover new leading compounds to enable quality control in phytopharmaceuticals made thereof. Therefore compounds of interest were separated after methanolic extraction by reversed-phase liquid chromatography (RP-LC) employing silica-C18 as stationary phase using gradient elution with water (10 mM H3PO4) and acetonitrile as mobile phase. Structural elucidation was carried out by diode array, fluorescence, and mass spectroscopic detection. Photodiode-array detection allowed the extraction of UV-absorbance spectra from the peaks of interest. Absorbance maxima were found at 203 and 279 nm with a shoulder at 230 nm. Additionally fluorescence emission and excitation spectra were recorded from the analytes using a fluorescence detector (FLD) after stop flow (lambdaex = 279 nm, lambdaem = 307 nm). The chromatographic reversed-phase system was coupled to an ion-trap mass spectrometer (LC-MS) by using an electrospray ionization interface (ESI). After optimization processes the separation was carried out using an ammonium acetate buffer at pH 5.5. Mass spectrometric detection turned out to be more sensitive in negative mode. Collisionally induced dissociation (CID) was used to obtain fragment ions of structural relevance (LC-MS/MS). Finally, compounds of interest coming from sorrel leaf (Rumex acetosa) eluting at low acetonitrile concentrations were confirmed to be catechin and epicatechin.