Characterization and screening of cyclooxygenase-2 inhibitors from Zi-shen pill by affinity ultrafiltration-ultra performance liquid chromatography mass spectrometry

Abstract

Ethnopharmacological relevanceZi-shen pill (ZSP) is a classical Chinese herbal formula used for treatment of benign prostatic hyperplasia (BPH). Aim of the studyTo characterize and screen cyclooxygenase-2 (COX-2) inhibitors from ZSP extract. Materials and methodsThe ZSP extract was incubated with COX-2 and the potential ligands were screened out by affinity ultrafiltration. Celecoxib and glipizide were chosen as positive control and negative control drug, respectively. Affinity ultrafiltration-ultra performance liquid chromatography-mass spectrometry (UPLC-MS) method was used. In addition, in vitro COX-2 inhibitory assay and in silico molecular docking technique were used for further validation. ResultsA total of 20 components were discovered and identified from ZSP ultrafiltrate by high resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS), among which 8 compounds were deduced as potential COX-2 inhibitors by their high specific binding values (>1.5). Inhibitory activities of demethyleneberberine, palmatine, berberine and timosaponin A-I were confirmed by an enzyme assay of COX-2, which validated the reliability of our approach. Molecular docking simulation investigated potential mechanism of action for these compounds. ConclusionThe results revealed that affinity ultrafiltration UPLC-MS could successfully screen out the potential COX-2 inhibitors from complex Chinese herbal formula ZSP extract, indicating that its therapeutic effect on BPH was partly based on the enzyme active ingredients.