Simultaneous Determination of Sesquiterpenes and Pyrrolizidine Alkaloids from Rhizomes of Petasites hybridus (L.) G.M. et Sch. and Dietary Supplements using UPLC-UV and LC-TOF Methods

Common Butterbur (Petasites hybridus (L.) G. Gaertn., B. Mey. & Scherb.) is a herbaceous perennial plant in the family Asteraceae, native to Europe and northern Asia [1]. Petasites hybridus exists in two chemo-varieties: those containing petasins and those with furano-petasins [2]. These two chemotypes of Petasites hybridus cannot be distinguished using classical morphological techniques. The pharmacological active substances of P. hybridus are believed to be sesquiterpene esters of the petasin and the furanopetasin chemotype [2].

UPLC-UV and LC-TOF-MS methods have been developed for the analysis of major sesquiterpenes and pyrrolizidine alkaloids from rhizomes of Petasites hybridus (L.) G.M. et Sch. (Family, Asteracea) and dietary supplements claiming to contain P. hybridus. The best results were obtained with an Acquity UPLC^TM HSS T3 (100mm×2.1mm, i.d., 1.8µm) column using gradient elution with a mobile phase consisting of ammonium formate (50 mM) and acetonitrile (0.05% formic acid) at a constant flow rate of 0.25 mL/min. The newly developed method was validated according to the ICH guidelines with respect to specificity, linearity, accuracy and precision. The limits of detection were found to be 3µg/mL and 0.1µg/mL for pyrrolizidine alkaloids and sesquiterpenes, respectively by the LC-UV method and 1 and 5ng/mL, respectively using the LC-TOF-MS method. The method was successfully used to analyze different P. hybridus market products, as well as to distinguish between other two Petasites species. The total content of petasins was found to be in the range from 0.02–11.6mg/dosage form for 21 dietary supplements analyzed and petasins were not detected in six dietary supplements. The pyrrolizidine alkaloids were detected in seven dietary supplements. LC-mass spectrometry coupled with electrospray ionization (ESI) was used for the identification and confirmation of sesquiterpenes and pyrrolizidine alkaloids from plant extracts, dietary supplements that claimed to contain P. hybridus and different species of Petasites.

Fig.1: UPLC-UV chromatograms of a mixture of compounds (pyrrolizidine alkaloids and sesquiterpenes) and various Petasites samples detected by UV at 220nm. Key to peak identities: 1, Senciophylline N-oxide; 2. Senecionine N-oxide; 3. Senecionine; 4. Senkirkin; 5. S-petasin; 6, 8β-H-eremophilanolide; 7. Isopetasin

Acknowledgements : This research is supported in part by the United States Department of Agriculture, Agricultural Research Service, Specific Cooperative Agreement No. 58–6408–2-0009, and the Global Research Network for Medicinal Plants (GRNMP), King Saud University. The authors would like to thank Annette Ford for extraction of the plant samples. References: [1] Der Marderosian A, Beutler JA, eds. The Review of Natural Products. St. Louis, MO: Facts and Comparisons Publishing Co., 2002. [2] Chizzola R, et al. (2006) Planta Med 72: 1254–1256.