A redox mediated UME biosensor using immobilized chromobacterium violaceum strain R1 for rapid biochemical oxygen demand measurement

An effective ferricyanide-mediated microbial biochemical oxygen demand (BOD) biosensor was constructed and used for BOD determination in a water system. This BOD sensor uses ultramicroelectrode (UME) technology in which the tip of the sensor consists of a two-electrode system (10-µm Pt working electrode and Pt counter electrode). Because of their small size, UMEs have relatively large diffusion layers and small overall currents enabling rapid achievement of useful steady-state conditions with very high scan rates. Living Chromobacterium violaceum R1 cells (isolated from pineapple industry wastewater) were immobilized on the surface of the UME working electrode using a calcium alginate gel and further enclosure by a layer of polyamide membrane. Glucose-glutamic acid (GGA) solution was used as the standard solution. The amperometric measurement was optimized at +450 mV operating potential and 30 mM ferricyanide in a 0.1 M phosphate buffer (pH 7.0) at 26 °C. The sensor exhibited a linear response ranging from 20 to 225 mg O2 L-1 BOD5 for standard GGA solution and 25 to 230 mg O2 L-1 BOD5 for OECD synthetic wastewater with a response time of 30 min. Repeatability and reproducibility of the biosensor were within the limits set by the APHA; i.e., less than 15.4%. The rapid BOD estimation of the biosensor is applicable for measuring samples with a high content of fast and easily assimilated compounds. When used to estimate the BOD of various wastewaters, the developed biosensor provided values comparable to those obtained using the conventional BOD5 method