Liquid chromatographic techniques in separation of betacyanins and their derivatives from red beet roots

A study on separation of betacyanins and their decarboxylated as well as dehydrogenated derivatives obtained from red beet roots (Beta vulgaris L.) in high pressure liquid chromatography (HPLC) and ion-pair high-speed counter-current chromatography (IP-HSCCC) was performed. The IP-HSCCC process was accomplished in the ‘head-to-tail’ mode in a solvent system composed of butanol — acetonitrile — water (5:1:6 v/v/v, acidifi ed with 0.7% trifl uoroacetic acid). The HPLC separation was performed in a typical reversed phase mode with mass spectrometry (ESI-MS) and diode-array (DAD) detection. The chromatographic profi les of betalains obtained in these two techniques were signifi cantly diff erent. In HPLC, the most polar compounds, like betanin and isobetanin, eluted before less polar decarboxylated and dehydrogenated derivatives (mostly degradation products of betanin). In IP-HSCCC, the dehydrogenated derivatives were eluted faster then their non-dehydrogenated analogues. It was observed for the fi rst time that betanin and neobetanin (14,15-dehydrogenated betanin), which are present in many plants containing betalains, had reversed elution orders during chromatographic separation by these two techniques