In Vivo Anti-Inflammatory Activity Assay

The two lead compounds 18 and 21 were advanced for in vivo anti-inflammatory activity study and celecoxib (1) was used as reference drug. In vivo anti-inflammatory activity was measured using a carrageenan-induced rat paw edema assay. In brief, three to five male Sprague–Dawley rats, 8–11-weeks-old, weighing 180–200 g (Charles-River Canada) were used in each group. Animals were randomized into different treatment groups based on similar paw size and body weight. Test compounds 18 and 21 suspended in water containing 1% methyl cellulose were administered orally for a minimum of four different doses (0.3. 1, 5, 10 mg kg⁻¹) 1 h prior to a 0.05 ml subcutaneous injection of fresh 1% carrageenan in 0.9% NaCl solution under the plantar skin of the hind paw. Control experiments were identical, except that the vehicle did not contain a test compound. The volume of the injected paw was measured at 0, 3, and 5 h using a UGO Basile 7141 Plethysmometer (series no. 43201), each value is mean of 10 measurements. A dose–response curve was constructed using GraphPad Prism 5.0 and ED₅₀were calculated. No unusual change in behavior and toxic effects was noticed in all animals. In vivo anti-inflammatory assays were carried using a protocol approved by the Health Sciences Animal Welfare Committee, University of Alberta, Edmonton, Canada.