Electrochemical biosensor for creatinine based on the immobilization of creatininase, creatinase and sarcosine oxidase onto a ferrocene/horseradish peroxidase/gold nanoparticles/multi-walled carbon nanotubes/Teflon composite electrode

•A novel electrochemical multienzyme biosensor for creatinine is presented.•CA, CO and SO enzymes are immobilized onto HRP/Fc/AuNPs/MWCNTs/Teflon electrodes.•The high selectivity allows the rapid determination of creatinine in human serum.

A composite electrode consisted of gold nanoparticles (AuNPs), multi-walled carbon nanotubes (MWCNTs) and Teflon, to which peroxidase (HRP) and ferrocene (Fc) were incorporated as auxiliary enzyme and redox mediator, respectively, was constructed. The enzymes creatininase, creatinase and sarcosine oxidase were then co-immobilized onto the surface of the resulting HRP/Fc/AuNPs/MWCNTs/Teflon electrode for the preparation of a creatinine biosensor. Amperometry in stirred solutions using a detection potential of 0.0 V vs Ag/AgCl allowed a linear calibration plot to be obtained in the 0.003–1.0 mM creatinine concentration range with a limit of detection of 0.1 μM (S/N = 3). The apparent Michaelis-Menten constant for creatininase was KMap = 4.1 ± 0.4 mM. The developed biosensor was validated with good results by determining creatinine in human serum and correlating with the spectrophotometric Jaffe's method.