This study describes the development of Biochemical Oxygen Demand (BOD) sensors that are portable and applicable in fresh and salt-containing wastewater. The fundamental principle of these BOD sensors is the immobilization of living microbial cells both on Clark oxygen electrodes and miniaturized thick film oxygen electrodes. The oxygen consumption by the microbial cells in metabolizing the organic pollutants is easily measured by these sensors....[ Read more ]

This study describes the development of Biochemical Oxygen Demand (BOD) sensors that are portable and applicable in fresh and salt-containing wastewater. The fundamental principle of these BOD sensors is the immobilization of living microbial cells both on Clark oxygen electrodes and miniaturized thick film oxygen electrodes. The oxygen consumption by the microbial cells in metabolizing the organic pollutants is easily measured by these sensors.

The yeast strain Arxula adenininivorans LS3 was chosen because of its salt-tolerance, wide substrate spectrum and robustness. It is immobilized best in a hydrogel, polycarbamoyl sulfonate (PCS). The gel has been found to have high oxygen diffusivity and high water content, both of which are the necessary conditions for the cells. In addition, the gel can attach to a Pt electrode surface and consequently, can be applied as a thick film oxygen electrode. A first BOD sensor prototype was constructed with immobilized Arxula cells in a membrane on a Clark-type oxygen electrode. The BOD values of real domestic wastewater from the sensor and from BOD₅ tests correlate well. The BOD sensor can thus be used for rapid estimation of BOD.

In order to reduce the size of the BOD sensor, an oxygen electrode fabricated by thick film technology was investigated. The oxygen measuring range, working potential and stability proved to be suitable for application in dissolved oxygen measurement. The Arxula cells are directly immobilized in PCS hydrogel on the thick film oxygen electrode. By applying the quasi-kinetic measurement for microbial respiration, the measurement time can be reduced from 12 minutes to 100 seconds. The apparent biocatalyst kinetic constant K_M and V_max are shown to be higher than those of non-immobilized free cells. The linear range and sensitivity are therefore higher for the immobilized cell sensor. The linear range can be increased further by coating extra layers of PCS gel on top of the cell layer to generate a higher diffusion barrier for the oxygen and substrates.

Electrochemical interferences have to be minimized because the thick film sensor is constructed without a gas selective permeable membrane. Cobalt(II) tetra(3-methoxy-4- hydroxyl-phenyl) porphyrin (CoTMHPP) was immobilized in the PCS gel and it performed as a catalyst for oxygen reduction. The oxygen reduction potential can be reduced from -450mV to near 0V against Ag/AgCl. The two rate affecting factors of immobilized mediators: the charge transfer diffusivity and the oxygen reduction rate are found to be higher than the oxygen diffusivity and the apparent V_max of the Arxula cells respectively. These results showed that the kinetics of the sensor is not dependent on the PCS hydrogel immobilized CoTMHPP mediator but on the metabolism of the cells and thus CoTMHPP is a good electrocatalyst. The Arxula cells were immobilized on the top of a CoTMHPP modified thick film electrode. The apparent K_M and V_max are similar to the Arxula sensor without the CoTMHPP mediator. Coimmobilization of Arxula and CoTMHPP results in a less efficient electron transfer but an increase in the linear range of applications.

The BOD values obtained from the thick film sensor with the immobilized CoTMHPP catalyst are comparable to the first BOD sensor prototype in domestic wastewater measurement. For the first time in the literature, both a stationary and a pocket sized microbial sensor have been developed that work under the real site operating conditions of East Asia. The biosensors are sensitive, stable and robust. They give reliable data on Biochemical Oxygen Demand in minutes rather in five days and will be valuable tools in the protection of the environment.