HPLC coupled on-line to ESI-MS and a DPPH-based assay for the rapid identification of anti-oxidants in Butea superba

Phytochem Anal. 2005 Nov-Dec;16(6):422-8. doi: 10.1002/pca.865.


A reversed-phase HPLC coupled on-line to a radical scavenging detection system and MS/MS was developed in order to combine separation, activity determination and structural identification of anti-oxidants in complex mixtures in one run. The sample was separated by HPLC and the eluate split into two flows. The major portion was fed into an electrospray ionisation MS/MS system, while the minor part was mixed with a free radical, 2,2'-diphenyl-1-picrylhydrazyl (DPPH), and the reaction determined spectrophotometrically. The negative peaks, which indicated the presence of anti-oxidant activity, were monitored by measuring the decrease in absorbance at 517 nm. The developed method was successfully applied to the identification of anti-oxidant compounds in a fraction, obtained by solid-phase extraction, of an extract of a Thai medicinal plant, Butea superba Roxb. The anti-oxidant compounds were separated and identified as procyanidin B2, (-)-epicatechin and procyanidin B5.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antioxidants / analysis*
  • Antioxidants / chemistry
  • Biphenyl Compounds*
  • Butea / chemistry*
  • Chromatography, High Pressure Liquid / methods*
  • Hydrazines*
  • Picrates
  • Spectrometry, Mass, Electrospray Ionization / methods*


  • Antioxidants
  • Biphenyl Compounds
  • Hydrazines
  • Picrates
  • 1,1-diphenyl-2-picrylhydrazyl