Separation of Steviol Glycosides by Hydrophilic Liquid Interaction Chromatography

Institut Prof. Dr. Kurz GmbH, Eupener Str. 161, 50933 Cologne, Germany; Corporate Analytical Services–Development, Weleda AG, Möhlerstraße 3, 73525 Schwäbisch Gmünd, Germany
Food Analytical Methods (Impact Factor: 1.97). 01/2012; 5(2):266-271. DOI: 10.1007/s12161-011-9229-x

ABSTRACT Stevia rebaudiana Bertoni contains several steviol glycosides with sweet flavour. They all are sweeter than sucrose (up to factor 450). The
various steviol glycosides are difficult to separate by reversed-phase chromatography. In this paper, five different hydrophilic
liquid interaction chromatography columns are characterized using isocratic elution (5–20% water in acetonitrile with buffer
or formic acid). Separation of the steviol glycosides is possible with all but one of the tested columns, but the robustness
of the separation against changes of buffer concentration and percentage of water differ. Aqueous percentage and ion strength
of the eluent are the main factors to be optimized in method development.

Stevia rebaudiana
–Hydrophilic liquid interaction chromatography–Steviol glycosides–HPLC–Mass spectrometry

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    ABSTRACT: Stevia rebaudiana contains several steviol glycosides that have a sweet flavor. They are up to 450 times sweeter than sucrose, but some have an undesirable aftertaste. Up to 2010, ten different steviol glycosides have been described from the leaves or purified extracts of S. rebaudiana. In this paper, the tandem mass spectrometric fragmentation patterns of these ten compounds are compiled, along with a scheme for structural elucidation. This scheme is then applied to 12 steviol glycosides that have not yet been described. The proposed structures of five steviol glycosides have been confirmed by other authors.
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