SIMULTANEOUS SEPARATION AND DETERMINATION OF EPHEDRINE ALKALOIDS AND TETRAMETHYL-PYRAZINE IN EPHEDRA SINICA STAPF BY HPLC

Abstract

A simple, sensitive, and reliable high performance liquid chromatography (HPLC) method was first developed for simultaneous determination of Ephedrine alkaloids and 2,3,5,6-tetramethylpyrazine (TMP) in Ephedra sinica Stapf. The HPLC separation was performed on a reversed phase C18 column (Nova-Pak® C18, 150 mm × 3.9 mm I.D.) with a duplicate gradient composed of methanol and phosphate buffer solution (PBS, 0.03 mol/L KH2PO4-acetic acid-triethyl amine = 400:0.25:0.1, v:v:v, pH = 6.0) to elute the solutes with a flow rate of 0.8 mL/min. The detection wavelength was set at 210 nm. Regression equations revealed linear relationships (correlation coefficients: 0.991 ∼ 0.998) between the peak area of each constituent (E, PE, NE, NPE, TMP) and its concentration. The detection limits for E, PE, NE, NPE, and TMP were 0.4, 0.1, 0.03, 0.02, 0.03 μg/mL, respectively at a signal-to-noise ratio of 3:1, and the recoveries ranged between 96.0–104.5%. The contents of E, PE, NE, NPE, and TMP in Ephedra sinica Stapf extracts were measured 0.480 g/100 g (2.5%, RSD), 0.090 g/100 g (3.0%, RSD), 0.090 g/100 g (2.4%, RSD), 0.060 g/100 g (2.2%, RSD), and 0.022 g/100 g (1.5%, RSD), respectively.