Development and validation of HPLC-DAD-CAD-MS(3) method for qualitative and quantitative standardization of...

A reversed phase high performance liquid chromatography method (HPLC) coupled with a diode array, charged aerosol detector or mass spectrometer was developed for the quantitative and qualitative standardization of Agrimoniae eupatoriae herba (Ph. Eur). Twenty four constituents comprising phenolic acids, flavan-3-ol derivatives, ellagitannin and flavonoids were fully or partially identified. Eight of detected compounds were reported from common agrimony for the first time. Fourteen major polyphenols were quantified using validated HPLC method with UV-vis and corona charged detection. Both detectors were shown to be equal for the quantification of selected polyphenols. Some limitations of universal response of corona charged detector for phenolic compounds were discussed. Using obtained data for DAD detector the sum of tannins, flavonoids and phenolic acids quantified in examined plant material was determined. Investigated samples contained 8.2-10.9mg/g of flavonoids, 6.3-10.9mg/g of tannins (among which agrimoniin was dominating constituent 2.6-5.4mg/g) and 0.6-0.9mg/g of phenolic acids proving that flavonoids should be considered as second major group of constituents in common agrimony.