Development of a triple hyphenated HPLC-radical scavenging detection-DAD-SPE-NMR system for the rapid identification of antioxidants in complex plant extracts

A. Pukalskas, T.A. van Beek, P. de Waard

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A rapid method for the simultaneous detection and identification of radical scavenging compounds in plant extracts was developed by combining an HPLC with on-line radical scavenging using DPPH as a model radical and an HPLC¿DAD¿SPE¿NMR system. Using this method a commercial rosemary extract was investigated. All major compounds present in the extract were collected on SPE cartridges after their separation. Advantages of on-line SPE peak trapping are the possibility to perform HPLC with non-deuterated solvents, a concentration effect and being able to record NMR spectra in pure 100% deuterated solvents. After comparing DAD and DPPH scavenging chromatograms, 1H NMR spectra of compounds having radical scavenging activities were recorded. Afterwards all compounds were collected and infused into an ESI-MS. The five main active compounds ¿ carnosol, carnosic acid carnosaldehyde, 12-methoxycarnosic acid and epiisorosmanol ¿ could be identified from the combined UV, NMR and mass spectral data without actually isolating them. It was possible to record on-line an HMBC spectrum of carnosic acid. Also one compound was tentatively identified as epirosmanol methyl ether
Original languageEnglish
Pages (from-to)81-88
JournalJournal of Chromatography. A, Including electrophoresis and other separation methods
Issue number1-2
Publication statusPublished - 2005



  • rosmarinus-officinalis l
  • carnosic acid
  • phenolic diterpenes
  • salvia-officinalis
  • rosemary extracts
  • online detection
  • sage extracts
  • flow-probe
  • constituents
  • oil

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