The aim of this study is to investigate stereoisomeric behavior of penicillamine and the effect of temperature on labeling. In addition, it was explored how stereoisomerism affected biological behavior of them. In the present work, D- and L-enantiomers of penicillamine(D-PA, L-PA) were labeled with Tc-99m using SnCl2 as reducing agent and their radiopharmaceutical potentials were investigated. Quality control procedures were carried out using thin layer radiochromatography (TLRC), electrophoresis and high performance liquid radiochromatography (HPLRC). HPRLC chromatograms showed two peaks for Tc-99m-D-PA, while a single peak was observed for Tc-99m-L-PA at room temperature. However, the single peak was observed at 90 A degrees C for both isomers. Labeling yields of each isomer were found to be over 98%. Biological activity of these complexes was determined on male Albino Wistar rats by biodistribution studies. While the biodistribution result of Tc-99m-D-PA showed high uptake in the liver, maximum uptake of Tc-99m-L-PA was observed in the kidneys. Both two complexes were cleared rapidly from the blood, mainly by the renal system. Since the activity concentration of Tc-99m-D-PA at the 30th minute in the kidneys and the liver reached a maximum value and at the 120th minute, it was removed by renal and hepatobiliary excretion. As a result, it can be concluded that stereoisomerism affect not only the chemical behavior, but also differs their biological behavior of these compounds.