The present study was conducted to investigate the effect of active compound (sesamin) found in defatted ethanol extract of Sesamum indicum seeds on the serum levels of lipid in mice. The qualitative and quantitative determination of sesamin bioactive compound in defatted ethanol extract using High Performance Liquid Chromatography (HPLC) analysis was carried out and compared with standard sesamin. It was found that the concentration of sesamin was 79.9% of ethanolic extract according to total peak area. The ffect of active compound (sesamin) in defatted ethanol sesame extract was evaluated in mice blood serum (in vivo) after feeding them high and low fat diet for one month. Defatted Sesame extract at concentration 500 mg/kg of body weight (B.W) showed significant reduction (p#0.05) in the level of total cholesterol (62.26 , 56.14) mg/dl, triglycerides (61.54 , 61.12) mg/dl , and low density lipoprotein LDL (29.97 , 23.21) mg/dl and significant increase in the level of high density lipoprotein HDL (19.39 , 20.70) mg/dl in comparison with both high and low fat diet groups which recorded (80.43 , 68.24) mg/dl, (77.50 , 74.16) mg/dl, (16.80 , 19.32) mg/dl, and (48.09 , 34.08) mg/dL, respectively.