Targeted Screening of Cyclooxygenase-2 Inhibitors From Dendropanax dentiger Root Using Affinity Ultrafiltration Coupled With UHPLC-MS.

Abstract

Cyclooxygenase-2 (COX-2) plays a critical role in the pathogenesis of rheumatoid arthritis (RA), whereas Dendropanax dentiger root (DDR) is a traditional Chinese medicine (TCM) widely used to treat RA. However, the specific bioactive compounds responsible for its therapeutic effect remain unidentified. In this study, affinity ultrafiltration coupled with liquid chromatography and mass spectrometry (AUF-LC-MS) was employed to screen for COX-2 inhibitors in DDR. Targeted compounds were identified by analyzing MS data and comparing them with reference standards. Molecular docking, molecular dynamics simulation, and enzyme inhibition assays were conducted to validate the COX-2 inhibitory effects of the identified compounds. As a result, 10 phenylpropanoids, including neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, liriodendrin, isochlorogenic acid B, isochlorogenic acid A, eleutheroside E1, isochlorogenic acid C, 3,5-di-O-caffeoylquinic acid methyl ester, and 4,5-di-O-caffeoylquinic acid methyl ester, were identified as potential COX-2 inhibitors in DDR using AUF-LC-MS. Mass spectrometric fragmentation patterns of these compounds were analyzed, revealing consistent and logical fragmentation profiles. Molecular docking results revealed that all 10 compounds exhibited strong binding affinities for COX-2, with binding energies ranging from -8.0 to -9.8 kcal/mol. The results of molecular dynamics simulations further supported molecular docking's findings. Further experimental validation confirmed that these compounds exhibited potent COX-2 inhibitory activity, with IC₅₀ values ranging from 5.2 to 10.3 µM. These compounds are likely to represent the primary anti-inflammatory components of DDR. Additionally, this study systematically identifies chlorogenic acids within the Dendropanax genus and investigates their mass spectrometric fragmentation patterns. The findings contribute to the scientific basis for the clinical application of DDR.