Phytochemical Profiling and Anti-Inflammatory Activity of Leaf Extract from Cleome gynandra Linn. and Melicope ptelefolia Champ. ex Benth

Herbal remedies are increasingly studied for their potential as safer alternatives to conventional anti-inflammatory drugs. However, the anti-inflammatory efficacy and interaction of many herbal combinations remain poorly understood. This study addresses this gap by investigating the anti-inflammatory activity, herb-herb interactions, and LC-ESI-MS/MS metabolite profiles of Cleome gynandra and Melicope ptelefolia leaf extracts. Fresh leaves of both plants were extracted and tested for cytotoxicity and nitric oxide (NO) inhibition using the murine RAW 264.7 macrophage cell line. Cell viability was assessed via MTT assay, while NO inhibition was measured for individual and combined extracts at varying ratios. LC-ESI-MS/MS analysis was performed to identify the major bioactive metabolites. Cleome gynandra extract showed higher cell viability compared to M. ptelefolia, indicating lower cytotoxicity. Among all tested combinations, the 70:30 (w/w) ratio of C. gynandra to M. ptelefolia exhibited the highest cell viability. Although C. gynandra demonstrated limited NO inhibition at 0.5 mg/mL, it still more potent than M. ptelefolia and their formulations. The LC-MS/MS analysis revealed that M. ptelefolia contains flavonoids like kaempferol neohesperidoside and derivatives with sugars such as rhamnose and arabinose, known to inhibit nitric oxide synthase and reduce NO production. Isorhamnetin derivatives also support its anti-inflammatory potential by modulating signaling pathways. In C. gynandra, the presence of quercetin rutinoside (rutin), kaempferol rutinoside, and phenolic acids like caffeic acid contribute to its anti-inflammatory effects. Quercetin rutinoside inhibits enzymes like COX-2 and iNOS and suppresses cytokines such as TNF-α and IL-6. Caffeic acid, a potent antioxidant, inhibits NF-κB activation, collectively supporting its anti-inflammatory capacity. In conclusion, C. gynandra demonstrates promising potential as a safer anti-inflammatory agent compared to M. ptelefolia, although its efficacy remains lower than that of conventional drugs such as diclofenac sodium.