Small molecules are difficult to detect by conventional SPR technique directly because the changes in the refractive index resulting from the binding processes of small biomolecules are often small. In order to extend the application of SPR biosensor in detecting a small molecule, we combine the advantage of aptamer technique with the amplifying effect of Au nanoparticles to design a sensitive SPR sensor for detecting small molecules. The principle of this sensor is based on surface inhibition detection. The aptamer is first immobilized on SPR gold film with its ss-DNA structure. The aptamer possessing this structure can be hybridized with Au nanoparticles-tagged complementary ss-DNA and result in a large change of SPR signal. However, the aptamer will change its structure from ss-DNA to tertiary structure after adenosine is added to the SPR cell. The aptamer possessing tertiary structure could not hybridize with Au nanoparticles-tagged complementary ss-DNA. Thus, the change of SPR signal resulted in the hybridization reaction between aptamer and Au nanoparticles-tagged complementary ss-DNA will decrease with the increase of the number of aptamers possessing tertiary structure, which is proportional to the concentration of the small molecule. Based on this principle, we choose a simple system (antiadenosine aptamer/adenosine) to detect the sensing ability of this SPR biosensor for a small molecule. The experimental results confirm that the SPR sensor we developed possesses a good sensitivity and a high selectivity for adenosine. The detection range for adenosine is from 1 x 10 (-9) to 1 x 10 (-6) M. More significantly, it is fairly easy to generalize this strategy to detect a spectrum of small molecules by SPR spectroscopy using different aptamers. Therefore, it is expected that this method may offer a new direction in designing high-performance SPR biosensors for sensitive and selective detection of a wide spectrum of small molecules.