Determination of flavonol aglycones in Ginkgo biloba dietary supplement crude materials and finished products by high-performance liquid chromatography: single laboratory validation.

In support of the initiative of the National Institutes of Health/Office of Dietary Supplements (NIH/ODS), U.S. Food and Drug Administration (FDA), and AOAC INTERNATIONAL to develop well-validated methods for botanicals and dietary supplements, an expert review panel (ERP) convened to determine the most promising method(s) for further study to determine total flavonol glycosides in Ginkgo biloba (L.) crude material and finished products. After comprehensive review of available and appropriate methods, the ERP chose 2 methods for further study, 1 developed by the NSF International-Institute for Nutraceutical Advancement (NSF–INA; 1), and another developed at Midwest Research Institute (MRI) in support of the National Institute of Environmental Health Sciences (NIEHS) National Toxicology Program (NTP; ntp-server.niehs.nih.gov). The NSF–INA method was chosen as the overall quantitative method, with sample preparation procedures based on the NTP-MRI method. Following protocol development and review, the resulting method was evaluated to conclude suitability for the determination of the flavonol aglycones quercetin, kaempferol, and isorhamnetin in Ginkgo biloba products. The 3 primary aglycone bases (Figure 1) are determined following hydrolysis of the flavonol glycosides with HC1 and heating at 90°C. Crude plant material, dry powder and liquid extracts, and finished product dosage forms are extracted and hydrolyzed with ethanol–water–HCl. The analytes are separated by liquid chromatography (LC) and measured by ultraviolet (UV) absorbance at 370 nm against external standards of quercetin, kaempferol, and isorhamnetin. Total flavonol glycoside concentration is then calculated using individual conversion factors for the 3 aglycones. Figure 1 Structures of predominant flavonol aglycones following hydrolysis of Ginkgo biloba products. For the single laboratory validation (SLV), the method was evaluated using 9 matrixes representative of Ginkgo products found in the marketplace, including crude leaf material, standardized dry powder extract, single and multiple entity tablets and capsules, and ethanol and glycerol tinctures (Table 1). In-depth evaluation of hydrolysis duration, ruggedness, precision, and accuracy testing was conducted on these selected matrixes in order to determine potential suitability of the method for further collaborative study. Table 1 Ginkgo biloba products evaluated for the flavonol aglycone single laboratory validation