Effect of β-Glucuronidase on Extraction Efficiency of Silymarin from Human Plasma Samples Using Validated HPLC-UV Analysis

Purpose: To investigate the effect of β-glucuronidase on the extraction efficiency of silymarin (mainly as silybin) from spiked human plasma using a sensitive and reproducible high performance liquid chromatography (HPLC) method. Methods: The importance of β-glucuronidase was evaluated by comparing the extraction efficiency of silymarin in β-glucuronidase-treated and untreated plasma samples. Isocratic HPLC with simple UV detection (288 nm) was applied to analyze the major silymarin components using Thermo-Electron C18 column (200 mm, 4.6 mm I.D., 5μm particle size). The mobile phase, consisting of methanol and 20 mM potassium dihydrogen phosphate buffer (50:50 v/v pH 2.8), was pumped at 1 ml/min. Results: The mean extraction efficiency was 98.97 % (CV = 1.69 %) for treated and 40.88 % (CV = 2.77 %) for untreated plasma samples, compared with nominal concentrations. Conclusion: The studied method showed 60 % reduced extraction efficiency of untreated samples compared to treated samples.