Simultaneous determination of five marker constituents in traditional Chinese medicinal preparation Le-Mai granule by high performance liquid chromatography.

Abstract An HPLC method for the simultaneous determination of five marker constituents was established for the quality control of traditional Chinese medicinal preparation Le–Mai granule. The marker constituents were danshensu, protocatechuic acid and protocatechualdehyde from Salviae miltiorrhizae bunge ; paeoniflorin from Radix paeoniae rubra and ferulic acid from Rhizoma chuanxiong . Extracted samples were successfully separated on a Diamonsil C 18 column (150 mm × 4.6 mm i.d., 5 μm) at 25 °C. The mobile phase was a mixture of methanol and 1.0% acetic acid employing gradient elution at a flow rate of 1.0 mL/min. Detection was accomplished with a diode-array detector and chromatograms were recorded at 230, 262, 280 and 322 nm. The compounds were identified by comparing their retention times and UV spectra in the 200–400 nm range with authentic standards. Regression equations revealed good linear relationship (correlation coefficients: 0.9993–0.9999) between the peak areas of the constituents and their concentrations. The average recoveries ( n  = 3) were between 96.2 and 102.5%. The proposed method has been successfully applied to the simultaneous determination of the five marker constituents in three lots of Le–Mai granule.