Rhodanese plays a critical role in cyanide detoxification by catalyzing the conversion of toxic cyanide into the less harmful thiocyanate. This comprehensive review examines recent studies on the purification and characterization of rhodanese from two distinct biological species such as plants and bacteria. In this study, different rhodanese assay techniques and purification methods are evaluated. The results showed that bacterial species, particularly Bacillus cereus and Klebsiella oxytoca, indicated the highest specific activity of 25.30 and 52.7 RU/mg, respectively, outperforming plant sources under similar purification conditions. Factors affecting enzyme activity such as temperature, pH, metal ion inhibition, and substrate specificity were also reviewed to minimize their effects on comparison by selecting nearly optimum temperature and pH, the highest affinity to the same substrate, and minimum effect of metal ion. The study concluded that bacterial species have greater rhodanese specific activity, and Lee modified by Agboola and Okonji and Bradford methods were the most used for rhodanese assay and protein quantification, respectively. Ion exchange chromatography is also the most effective rhodanese purification technique.
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