Isolation and identification of unstable components from Caesalpinia sappan by high-speed counter-current chromatography combined with preparative high-performance liquid chromatography

L. ( . L.), a traditional Chinese medicine, has been widely used to treat bruises and dysmenorrhea, performing pharmacological activities such as anti-inflammatory and anti-tumor. . L. has been reported to contain compounds such as protosappanins, brazilins, and homoisoflavones. In the pre-experiments, we discovered that there were many unstable components in the characteristic chromatogram of . L. Here, silica-gel column chromatography, high-speed counter-current chromatography, and preparative high-performance liquid chromatography were combined and applied to isolate the unstable components from alcohol extract of . L. The results showed that four unstable compounds were collected with the purity higher than 95.0%, characterized as episapponal, brazilin, sapponal, and 4- -methylsapponal by hydrogen-1 and carbon-13 nuclear magnetic resonance. Based on the above results, the characteristic chromatogram of . L. was established, and the characteristic peaks were identified. These results provided a theoretical basis for the quality assessment of . L.