A high-performance liquid chromatography with electrochemical detection for the determination of total hypericin in extracts of St. John's Wort.
An HPLC method for the quantitation of hypericin using a new and sensitive amperometric detection is presented. Hypericin was eluted isocratically using a mobile phase consisting of ammonium acetate, methanol and acetonitrile. The oxidation was carried out with a glassy carbon electrode at a potential of + 1.1 V vs. an Ag-AgCl-KCl reference electrode. Under the conditions described, hypericin was separated at a retention time (Rt) of 12 min. Linearity was obtained over the range 0.035-1.30 microg/mL (r = 0.9994). The limit of detection was determined to be 0.010 ng on-column for hypericin. The method was applied to the determination of total hypericin (hypericin, pseudohypericin, protohypericin and protopseudohypericin) in extracts of St. John's wort using hypericin as an external standard. The protoforms were converted into hypericin and pseudohypericin by subjecting the sample to artificial light prior to chromatographic analysis. For the evaluation of total hypericin, the peak areas of pseudohypericin (Rt 3.7 min) and hypericin (Rt 12.0 min) were combined. The relative standard deviation in analysing samples containing Hypericum ranged from 2.5 to 5.4%.
Rückert
Ulla
U
Institute of Pharmaceutical Sciences, Karl-Franzens-University Graz, Schubertstrasse 1, A-8010 Graz, Austria.
Eggenreich
Karin
K
Likussar
Werner
W
Wintersteiger
Reinhold
R
Michelitsch
Astrid
A
D000873:Anthracenes; D000928:Antidepressive Agents / Q000032:analysis* / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D004563:Electrochemistry; D020902:Hypericum / Q000737:chemistry*; D010569:Perylene / Q000031:analogs & derivatives* / Q000032:analysis / Q000737:chemistry; D010936:Plant Extracts / Q000737:chemistry*; D010946:Plants, Medicinal / Q000737:chemistry
D000873:Anthracenes; D000928:Antidepressive Agents; D010936:Plant Extracts; D010569:Perylene; C004965:hypericin