Determination of Molecular Weight and Partial Purification of Cyclooxygenase-2 Enzyme and its Kinetic Study in Male Colorectal Cancer Patients

Abstract

Objective: To purify cyclooxygenase-2 from the serum of male colorectal cancer patients.

Study Design: Combined clinical and experimental in vitro study
Place and Duration of Study: This study was conducted at the College of Science, Kirkuk University, Kirkuk, Iraq from 15th December 2025 to 30th April 2026.

Methods: Sedimentation techniques, hemodialysis, ion-exchange chromatography column separation, and gel filtration were used. The resin material (DEAE cellulose) was positively charged.

Results: Two analogs of the enzyme were obtained. Cyclooxygenase-2 activity was higher in peak A than in peak B, with specific activity values of 4.94 U/mg and 2.17 U/mg respectively. The approximate molecular weight of the cyclooxygenase-2 from (peak A) was estimated by gel filtration using Sefadex G-100 gel and was 71302 dalton. Optimal conditions were estimated after kinetic study of purified cyclooxygenase-2: velocity maximum (Vmax=1 U/mL), Michaelis-Menten constant (Km=2.5 mM, pH=7.5), temperature 38 °C and time period 70 seconds.

Conclusion: The success of the proposed purification protocol in isolating highly purified cyclooxygenase-2 enzyme and maintaining its kinetic stability from complex clinical samples. The inferred kinetic parameters (K m, Vmax) contribute to a deeper understanding of the catalytic behavior of the human enzyme, supporting future research into the mechanisms of pharmacological inhibition.