Chemical Profiling of Anti‐Inflammatory Constituents for Hongteng Baijiang Decoction Using Comprehensive Two‐Dimensional Countercurrent Chromatography × High Performance Liquid Chromatography Coupled With Quadrupole Time‐Of‐Flight Mass Spectrometry

Comprehensive two-dimensional reverse phase countercurrent chromatography (CCC)×high performance liquid chromatography (HPLC) was investigated for chemical profiling and identification of bioactive components with anti-inflammatory activities for a traditional Chinese medicine formula, Hongteng Baijiang decoction (HBD), for the first time. Anti-inflammatory activity of different extractions from HBD were evaluated. Two-dimensional contour plots for CCC×HPLC were obtained after comprehensive separation was completed. The total peak capacity of the two-dimensional system was evaluated, yielding approximately 782 peaks through comprehensive two-dimensional separation, which demonstrated a high degree of orthogonality and large separation space occupancy of 90.16%. Totally 207 chemical components were identified in ethyl acetate extract from HBD using liquid chromatography-quadrupole time-of-flight mass spectrometry based on the two-dimensional separation, with additional 70 compounds identified compared to that in one-dimensional HPLC separation. It was found that terpenoids were major bioactive components contributing to anti-inflammatory bioactivity in HBD. These findings highlight the potential of comprehensive two-dimensional chromatography (CCC×HPLC) as a powerful and promising method for separating and chemical profiling for complex traditional Chinese herbal formula.