Scaling up of high-speed countercurrent chromatographic apparatus with three columns connected in series for rapid preparation of (-)-epicatechin.

In the present study, compact high-speed countercurrent chromatographic apparatus was constructed with three columns connected in series. Two sets of columns were prepared from 10 mm and 12 mm I.D. tubing to form 12 L and 15 L capacities, respectively. Performance of these columns was compared for the separation of (-)-epicatechin gallate (ECG) from a tea extract by flash countercurrent chromatography (FCCC). In each separation, 200 g of the tea extract in 1600 mL of mobile phase was loaded onto the column. The 12 L column gave 7.5 L (35 g of ECG) and the 15 L column gave 9 L (40 g of ECG) of ECG solution without impurities. The ECG solution was directly hydrolyzed by tannase into (-)-epicatechin. The hydrolysate was purified by flash chromatography on AB-8 macroporous resin to give 52 g of EC (purity 99.1%). This scaled up apparatus could be used for the industrial separation of natural products.