Determination of Amino Acids in Rhubarb Seeds with DNFB Pre-column Derivatization by Reversed-phase High Performance Liquid Chromatography

The aim of the study was to determine the contents of amino acids in rhubarb seeds with pre-column derivatization by reversed-phase high-performance liquid chromatography( RP-HPLC). Amino acids extracted by the method of hydrolysis in 6 mol /L HCl were derivatized by 2,4-dinitrofluorobenzene( DNFB) and analyzed by RP-HPLC with gradient elution. The chromatographic separation was performed on a Gemimi-NX C18( 4.6 mm × 250 mm,5 μm) column with 0. 1 M aqueous sodium acetate solution( A) and acetonitrile-water( 1∶ 1)( B) as mobile phases. The column temperature was maintained at 37 ° C,and the detection wavelength was set at 360 nm. The results showed that the three genuine rhubarb seed contained 17 kinds of amino acids,11. 30% to 19. 26% of the total content. The experimental results proved that the developed RP-HPLC method was sensitive,accurate and repeatable.