Rapid Determination of Theophylline, Theobromine and Caffeine in Dietary Supplements Containing Guarana by Ultra-Performance Liquid Chromatography

A rapid and simple method for determination of theophylline, theobromine and caffeine in dietary supplements containing guarana by ultra-performance liquid chromatography (UPLC) has been developed. Theophylline, theobromine and caffeine were extracted from finely powdered samples with water in a boiling water bath for 20 min, then the extracts were filtered and the filtrates were subjected to UPLC. Liquid samples were diluted with water and filteres, and the filtrates were subjected to UPLC. UPLC separation was performed on an AQUITY UPLC BEH C 18 column (2.1 mm i.d. X 50 min, 1.7 mu m, Waters) with 10 mmol/L ammonium acetate buffer (pH 4.0)-acetonitrile gradient and eluates were monitored at 275 nm. The recoveries of theophylline (spiked at 200 mu g/g [tablet] and 50 mu g/mL [liquid]), theobromine (spiked at 200 mu g/g [tablet] and 50 mu g/mL [liquid]) and caffeine (spiked at 1,000 mu g/g [tablet] and 250 mu g/mL [liquid]) were 97.6-98.7%, 97.3-99.7%, 97.1-105.4%, respectively. The quantitation limits of theophylline, theobromine and caffeine were 10 mu g/g (seed, seed powder, tablet and capsule) and 2.0 mu g/mL (liquid) each. When this analytical method was applied to commercial dietary supplements, theophylline, theobromine and caffeine were found at concentrations of 4.45 mg/tablet, 5.48 mg/tablet, 139 mg/tablet, respectively. Taking 4 tablets of this product according to the directions on the package could be dangerous to consumers because of possible overdosing of these ingredients.