2.3. COX-2 Enzyme Inhibition Study

COX-2 Inhibitor Screening Kit (Fluorometric) (K547-100-BioVision) was used for the assay. COX-2 activity was analyzed by repeating the assay (n=4) [20, 21]. N, N, N, N-tetra methyl-p-phenylenediamine (TMPD) oxidation based chromogenic assay during the reduction of Prostaglandin G2 (PGG2) to Prostaglandin H2 (PGH2) was employed [22]. The assay mixture was pre-incubated at 22°C for 1 min along with the test compounds. Other than compounds Cpd1-10, the assay mixture has EDTA (3 mM), haematin (15 mM), Tris-HCl buffer (100 mM, pH 8.0), COX-2 enzyme (100 mg). Arachidonic acid and TMPD were added together to make it to 1 ml in total volume. The rate of TMPD oxidation in 20 seconds was measured as the enzyme activity at 602 nm absorbance.