Large-scale purification of glucosinolates by strong ion-exchange centrifugal partition chromatography. From lab-scale CPC to 5L-pilot-scale FCPC

The glucosinolates (GSLs) constitute a large class of hydrosoluble plant secondary metabolites acting as phytoanticipins. They are mainly present in the Brassicaceae, Capparidaceae families in the Capparales order, such as broccoli, mustard, rapeseed, cauliflower, cress, caper and papaya [1]. Naturally occurring GSLs are anionic, β-D-thioglucoside-(Z)-N-hydroxyiminosulfates that contain an amino acid-derived side chain. GSLs and their hydrolysis products are of particular interest owing to their potential benefit to health: they have been described as chemopreventive, chemoprotective, antioxidant and antibiotic substances [2]. However, metabolite fate and bioavailability studies as well as clinical evaluation require large amounts of pure GSLs that are often not commercially available at all. Indeed, due to their particular physicochemical properties, the separation and isolation of glucosinolates from crude plant extracts is an extremely difficult task and only few reference samples can be bought. Therefore, new chromatographic methods have recently been developed in order to obtain the required amounts of pure GSLs (3). Previously, the gram-scale purification of pure sinalbin and glucoraphanin from white mustard and broccoli seed extracts was successfully achieved using strong ion-exchange displacement centrifugal partition chromatography (SIX-CPC) [4]. In this study, the same protocol was successfully applied to the purification of glucotropaeolin from papaya, gluconastrutiin from cress, glucoiberin, glucoiberverin and sinigrin from cauliflower and glucoiberin, sinigrin, gluconapin, glucobrassicanapin and gluconasturtiin from brokoletti seeds. Finally, our protocol was easily scaled up from a 200 mL lab scale CPC column to a 5L FCPC apparatus, in order to prove that our method can be applied to industry-scale GSLs production. In a single run that lasted for 3 hours, 310g of a crude white mustard seeds extract were processed to yield 64g of highly pure sinalbin.

References: 1. Fahey J. W. et al. (2001) Phytochemistry 56:5–51. 2. Holst, B., Williamson, G. (2004) Nat Prod Rep 21:425–447. 3. Fahey J.W. et al. (2003)J. Chromatogr. A 996:85–93. 4. Toribio, A. et al. (2007)J. Chromatogr. A 1170:44–51