In this study, acetylcholinesterase (AChE; EC 220.127.116.11) was purified from sheep liver by affinity chromatography. The purification rate was found 3541.7 fold. The purification control of enzyme was done with sodium dodesilsulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The optimal pH, ionic strength and temperature of enzyme were determined. The optimal pH and the optimum temperature were 7.5-8.5 and 35 degrees C respectively. The highest activity was seen in concentration of 0.2 M (NH4)(2)SO4 as ionic strength. On the other hand, the inhibition effects of some painkiller, (paracetamol + caffein), (neostigmin methylsulfate) and (paracetamol + propifenazon + caffein) were investigated in this study. According to results, the I-50 values are 1.27 X 10(-3), 1.02 x 10(-4) and 1.236 M respectively. Also, K-i values are determined as 1.246 x 10(-3), 4.326 x 10(-5) and 1.646 x 10(-3) mol(-1) min(-1) respectively.