A simple and sensitive spectrofluorimetric method has been developed for the determination of some selected sulfhydryl-containing drugs namely Acetylcysteine (ACS), Captopril (CAP) and Mesna (MSN).The method is based on the interaction of the drugs with potassium (5-sulfoxino) palladium II in alkaline medium in presence of magnesium ions, where the sulfhydryl group combines with palladium from the non-fluorescent potassium bis (5-sulfoxino) palladium II. The resulting 8-hydroxy-5-quinoline sulfonic acid coordinates with magnesium to form the fluorescent chelate that is a measure of the amount of sulfhydrl containing drug analyzed. The fluorescence intensity was measured at an emission wavelength of 485 nm, by excitation at 345 nm. All the experimental parameters affecting the reaction were studied and optimized. The proposed method was applicable over the concentration range of 0.04-0.44 µg/mL for the three drugs and was applied for their determination in bulk form and in pharmaceutical preparations without interference from common excipients. The assay results were statistically compared with those obtained from previously reported methods where no significant difference was found between them. The selectivity and the stability-indicating aspect of the proposed method were confirmed by preparing the disulphides of the studied drugs and applying the reaction to the parent drugs in presence of their disulphides where no interference was detected from these related substances. By virtue of its high sensitivity, the proposed method was also extended to analyze the drugs in spiked human plasma and urine.