Separation of chlorogenic acid and concentration of trace caffeic acid from natural products by pH-zone-refining counter-current chromatography

摘要

Chlorogenic acid, a major compound among phenolic acids of Flaveria bidentis (L.) Kuntze and Flos lonicerae, was successfully separated by pH-zone-refining counter-current chromatography. Caffeic acid and chlorogenic acid were selected as test samples, which represent as phenolic acids for separation by the pH-zone-refining counter-current chromatography. The separation of these test samples was performed with two-phase solvent system composed of methyl-tert.-butyl-ether-acetonitrile-water at a volume ration of 4:1:5 (v/v) where trifluoroaceetic acid (8 mM) was added to the organic stationary phase as a retainer and NH4OH (10 mM) to the aqueous mobile phase as an eluter. For separation of crude extract from Flaveria bidentis (L.) Kuntze and Flos lonicerae. a slightly polar two-phase solvent system composed of methyl-tert.-butyl-ether-acetonitrile- n-butanol-water at a volume ratio of 4:1:1:5 (v/v) was selected where trifluoroacetic acid (3 mM) was added to the organic stationary phase as a retained and NH4OH (3 mM) to the aqueous mobile phase as an eluter. pH-zone-refining counter-current chromatography yielded 16.2 mg amount of chlorogenic acid with the purity of 91% from 1.4 g of Flaveria bidentis (L.) Kuntze, and 134 mg of chlorogenic acid at the purity of 99% from 1.3 g of crude extract of Flos lonicerae. These results suggest that pH-zone-refining counter-current chromatography is suitable for the isolation of the chlorogenic acid from the crude extracts of Flaveria bidentis (L.) Kuntze and Flos lonicerae.