A creatinase-producing bacterium was isolated in this work. Based on its morphological and physiological characteristics, G + C mol% of DNA and 16S rDNA sequence, it was identified to be a Paracoccus sp. Creatinase produced from it was purified to electrophoretic homogeneity with recovery of 17.4% by ion exchange chromatography, hydrophobic chromatography and gel filtration. The molecular mass of the enzyme was estimated to be 48,000 Da by SDS-PAGE. The enzyme showed maximum activity at pH 7.5 and was stable at pH 5.5-9.5. The K-m of the enzyme was 24.6 mM with creatine as substrate at 37 degrees C. Metal ions such as Cu2+, Hg2+ and Ag+ inactivated its activity completely. When the creatinase was used to measure creatinine, the concentration of the creatinine and the absorbance of the dye at 500 nm were linearly related up to 2000 mu mol& This study demonstrates that Paracoccus is a new source for producing creatinine-analyzing enzyme. (c) 2006 Elsevier Ltd. All rights reserved.