Abstract
Suitably validated analytical methods that can be used to quantify medicinally active
phytochemicals in natural health products are required by regulators, manufacturers,
and consumers. Hawthorn (Crataegus) is a botanical ingredient in natural health products used for the treatment of cardiovascular
disorders. A method for the quantitation of vitexin-2″-O- rhamnoside, vitexin, isovitexin, rutin, and hyperoside in hawthorn leaf and flower
raw materials and finished products was optimized and validated according to AOAC
International guidelines. A two-level partial factorial study was used to guide the
optimization of the sample preparation. The optimal conditions were found to be a
60-minute extraction using 50 : 48 : 2 methanol : water : acetic acid followed by
a 25-minute separation using a reversed-phased liquid chromatography column with ultraviolet
absorbance detection. The single-laboratory validation study evaluated method selectivity,
accuracy, repeatability, linearity, limit of quantitation, and limit of detection.
Individual flavonoid content ranged from 0.05 mg/g to 17.5 mg/g in solid dosage forms
and raw materials. Repeatability ranged from 0.7 to 11.7 % relative standard deviation
corresponding to HorRat ranges from 0.2 to 1.6. Calibration curves for each flavonoid
were linear within the analytical ranges with correlation coefficients greater than
99.9 %. Herein is the first report of a validated method that is fit for the purpose
of quantifying five major phytochemical marker compounds in both raw materials and
finished products made from North American (Crataegus douglasii) and European (Crataegus monogyna and Crataegus laevigata) hawthorn species. The method includes optimized extraction of samples without a prolonged
drying process and reduced liquid chromatography separation time.
Key words
Flavonoids -
Crataegus
- single-laboratory validation - liquid chromatography - hawthorn - vitexins - Rosaceae
