title: High-speed counter-current chromatography in separation of betacyanins from flowers of red Gomphrena globosa L. cultivars.
abstract: Antioxidant and possible chemopreventive properties of betacyanins, natural plant pigments, contribute to a growing interest in their chemistry and separation. Mixtures of betacyanins from fresh red Gomphrena globosa L. cultivar flowers were separated in three highly polar solvent systems by high-speed counter-current chromatography (HSCCC) for a direct comparison of their separation effectiveness. Three samples of crude extract (600mg) were run on semi-preparative scale in solvent system (NH4)2SO4soln - EtOH (2.0:1.0, v/v) (system I) and the modified systems: EtOH - ACN - 1-PrOH - (NH4)2SO4satd.soln - H2O (0.5:0.5:0.5:1.2:1.0, v/v/v/v/v) (system II) and EtOH - ACN - (NH4)2SO4satd.soln - H2O (1.0:0.5:1.2:1.0, v/v/v/v) (system III). The systems were used in the head-to-tail (system I) or tail-to-head (systems II and III) mode. The flow rate of the mobile phase was 2.0ml/min and the column rotation speed was 860rpm. The retention of the stationary phase was 52.0% (system I), 80.2% (systems II) and 82.0% (system III). The betacyanins in the crude extract as well as HSCCC fractions were analyzed by LC-MS/MS. System I was applied for the first time in HSCCC for the separation of betacyanins and was quite effective in separation of amaranthine and 17-decarboxy-amaranthine (αI=1.19) and very effective for 17-decarboxy-amaranthine and betanin (αI=2.20). Modification of system I with acetonitrile (system III) as well as acetonitrile and propanol (system II) increased their separation effectiveness. Systems II-III enable complete separation of 17-decarboxy-amaranthine (KD(II)=2.94,KD(III)=2.42) and betanin (KD(II)=2.46,KD(III)=1.10) as well as betanin and gomphrenin I (KD(II)=1.62, KD(III)=0.74). In addition, separation of amaranthine and 17-decarboxy-amaranthine is the most effective in system II, therefore, this system proved to be the most suitable for the separation of all polar betacyanins.
authors.affiliation: Department of Analytical Chemistry, Institute C-1, Faculty of Chemical Engineering and Technology, Cracow University of Technology, ul. Warszawska 24, Cracow 31-155, Poland. Electronic address: anetasporna@chemia.pk.edu.pl.
authors.forename: Aneta
authors.initials: A
authors.lastname: Spórna-Kucab
authors.affiliation: Department of Analytical Chemistry, Institute C-1, Faculty of Chemical Engineering and Technology, Cracow University of Technology, ul. Warszawska 24, Cracow 31-155, Poland.
authors.forename: Ewelina
authors.initials: E
authors.lastname: Hołda
authors.affiliation: Department of Analytical Chemistry, Institute C-1, Faculty of Chemical Engineering and Technology, Cracow University of Technology, ul. Warszawska 24, Cracow 31-155, Poland.
authors.forename: Sławomir
authors.initials: S
authors.lastname: Wybraniec
mesh_terms: D027703:Amaranthaceae / Q000737:chemistry*; D050859:Betacyanins / Q000737:chemistry / Q000302:isolation & purification*; D002851:Chromatography, High Pressure Liquid; D003377:Countercurrent Distribution / Q000379:methods*; D035264:Flowers / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry
chemical_list: D050859:Betacyanins
keywords: Amaranthine; Betacyanins; Betanin; Gomphrena globosa L.; HSCCC