Based on magnetic beads as separating carrier and adenosine as a specific competitor appeared，a method of aptamer-based chemiluminescence(CL) for the determination of adenosine was developed.Experimental parameters including magnetic beads,adenosine-aptamer and report DNA were optimized.Under the optimal conditions，the decreased CL intensity was proportional to the adenosine concentration in the range of 1.0×10-9-1.0×10-4 mol/L(r2=0.99)，with a detection limit of 1.0×10-9 mol/L.The relative standard deviation(n=5) for 0.1 mmol/L adenosine is 5.0%.The present method was successfully applied in the determination of adenosine in real human urine.The method is simple，sensitive and specific for the detection of adenosine，and it is a promising approach for the determination of small biomolecules in the fields of clinical diagnosis，pharmaceutical analysis and so on.