Abstract : Colchicine is a medication used to treat gout and Behçet's disease. A rapid, stable and sensitive reversed phase liquid chromatography method coupled with MS/MS detector was developed and validated for determination of Colchicine in human plasma, Colchicine and internal standard (Colchicine-D3) were usefully extracted from human plasma samples by using liquid-liquid extraction technique. Validation parameter was carried out for the chromatographic method used for determining Colchicine in plasma including: accuracy, precision, linearity, selectivity, stability, calibration curve, recovery. The mobile phase consisting of (85% methanol: 15% Ammonium chloride), column ACE C8, (50 X 2.1) mm, 5 µm, flow rate was 0.4 ml/min, retention time was 0.44 minute for colchicine and Colchicine- D3, and the total run time was 0.8 minute. The lower limit of quantitation was 0.05 ng/mL, the calibration curve was linear (R2 = 0.9985) over the range of (0.05 – 4.00) ng/ml. The intra- and inter day precisions for quality control samples were < 10.0%, and the intra- and inter day accuracies were in the range of (89.33-106.33) %. However, the current LC/MS method of colchicine could be applied for pharmacokinetic study in human plasma.