Deep Eutectic Solvent‐Enhanced Counter‐Current Chromatography Coupled With Ultrafiltration Liquid Chromatography for Targeted Separation of Cyclooxygenase‐2 Inhibitors From Magnolia officinalis

Magnolia officinals, owing to its significant anti-inflammatory effect, has been widely utilized in the field of traditional Chinese medicine. This study employed ultrafiltration-LC in conjunction with a deep eutectic solvent-enhanced counter-current chromatography method to achieve the separation and purification of three cyclooxygenase-2 inhibitors from M. officinals plant material. First, cyclooxygenase-2 inhibitors were discovered from M. officinals via ultrafiltration-LC. Response surface methodology was further employed to optimize ultrasonic-assisted extraction parameters. It was found the discovered cyclooxygenase-2 inhibitors could be significantly enriched under the optimized conditions. Using the deep eutectic solvent-enhanced counter-current chromatography, three target cyclooxygenase-2 inhibitors were successfully separated with a solvent system composed of n-hexane/ethyl acetate/DES/water (3:4:5:2, v/v/v/v). Ultimately, three cyclooxygenase-2 inhibitors, including 91 mg of honokiol, 41 mg of 8-obovatol, 95 mg of magnolol were obtained from 500 mg sample. The prioritized magnolol was found to exhibit anti-inflammatory activity via NF-κB pathway phosphorylation cascade activation inhibition and NF-κB/p65 nuclear translocation prevention. It demonstrated the integration of ultrafiltration-LC with deep eutectic solvent-enhanced counter-current chromatography enables the efficient separation of bioactive molecules. It not only advanced the academic understanding of the anti-inflammatory active molecules of M. officinalis, but also created preconditions for the subsequent research, development, and clinical application of M. officinalis.