The characterization of herbal materials is a significant challenge to analytical chemists. Goldenseal (Hydrastis canadensis L.), which has been chosen for toxicity evaluation by NIEHS, is among the top 15 herbal supplements currently on the market and contains a complex mixture of indigenous components ranging from carbohydrates and amino acids to isoquinoline alkaloids. One key component of herbal supplement production is botanical authentication, which is also recommended prior to initiation of efficacy or toxicological studies. To evaluate material available to consumers, goldenseal root powder was obtained from three commercial suppliers and a strategy was developed for characterization and comparison that included Soxhlet extraction, HPLC, GC-MS, and LC-MS analyses. HPLC was used to determine the weight percentages of the goldenseal alkaloids berberine, hydrastine, and canadine in the various extract residues. Palmatine, an isoquinoline alkaloid native to Coptis spp. and other common goldenseal adulterants, was also quantitated using HPLC. GC-MS was used to identify non-alkaloid constituents in goldenseal root powder, whereas LC-MS was used to identify alkaloid components. After review of the characterization data, it was determined that alkaloid content was the best biomarker for goldenseal. A 20-min ambient extraction method for the determination of alkaloid content was also developed and used to analyze the commercial material. All three lots of purchased material contained goldenseal alkaloids hydrastinine, berberastine, tetrahydroberberastine, canadaline, berberine, hydrastine, and canadine. Material from a single supplier also contained palmatine, coptisine, and jatrorrhizine, thus indicating that the material was not pure goldenseal. Comparative data for three commercial sources of goldenseal root powder are presented.

A fast, practical ambient extraction methodology followed by isocratic liquid chromatography (LC) analysis with UV detection was validated for the determination of berberine, hydrastine, and canadine in goldenseal (Hydrastis canadensis L.) root powder. The method was also validated for palmatine, a major alkaloid present in the possible bioadulterants Coptis, Oregon grape root, and bar-berry bark. Alkaloid standard solutions were linear over the evaluated concentration ranges. The analytical method was linear for alkaloid extraction using 0.3–2 g goldenseal root powder/100 mL extraction solvent. Precision of the method was demonstrated using 10 replicate extractions of 0.5 g goldenseal root powder, with percent relative standard deviation for all 4 alkaloids ≤1.6. Alkaloid recovery was determined by spiking each alkaloid into triplicate aliquots of neat goldenseal root powder. Recoveries ranged from 92.3% for palmatine to 101.9% for hydrastine. Ruggedness of the method was evaluated by performing multiple analyses of goldenseal root powder from 3 suppliers over a 2-year period. The method was also used to analyze Coptis root, Oregon grape root, barberry bark, and celandine herb, which are possible goldenseal bioadulterants. The resulting chromato-graphic profiles of the bioadulterants were significantly different from that of goldenseal. The method was directly transferred to LC with mass spectrometry, which was used to confirm the presence of goldenseal alkaloids tetrahydro-berberastine, berberastine, canadaline, berberine, hydrastine, and canadine, as well as alkaloids from the bioadulterants, including palmatine, jatrorrhizine, and coptisine.

An isocratic high-performance liquid chromatography (HPLC) method was developed for the analysis of hydrastinine, palmatine, berberine, hydrastine, and canadine, all alkaloid components known to be present in goldenseal root powder. Optimized separation was achieved on a Zorbax Eclipse-XDB column at 30 • C using a mobile phase of 10 mM ammonium acetate/acetonitrile (70:30, v/v) at a flow rate of 1.0 mL/min with ultraviolet detection at 235 nm. The method showed linearity for palmatine, berberine, hydrastine, and canadine at approximately 4-400 µg/mL, while hydrastinine was linear at approximately 4-80 µg/mL. Method precision and robustness were investigated. An example of an HPLC analysis of goldenseal root powder extract is presented here also. These studies indicate that the method described here is usable for analysis of goldenseal extracts.