Fluorometric enzymatic autoindicating biosensor for H2O2 determination based on modified catalase

Ortega, Estefania; de Marcos, Susana; Galbán, Javier

Fluorometric enzymatic autoindicating biosensor for H2O2 determination based on modified catalase

2013

Ortega, Estefania | de Marcos, Susana | Galbán, Javier

Our general aim is to develop reversible optical biosensors which can be used for continuous monitoring. In this paper we propose a biosensor for H₂O₂ determination. The bioreceptor is catalase (Cat) previously linked to a Ruthenium O₂-sensitive fluorophore (Cat–Ru). It is based on the reversible H₂O₂ disproportionation into O₂ and H₂O.First, the fluorescent-enzymatic system was optimized for batch measurements (linear response ranges from 1×10⁻⁴ to, at least, 1×10⁻³M H₂O₂). Because of its reversibility, the same enzyme aliquot can be used for performing the whole calibration step (and the subsequent determination). Secondly, the optical sensor was prepared by Cat–Ru immobilization in a polyacrylamide film. The sensor permits H₂O₂ determination in a similar concentration range as in batch mode and can be used during at least 1 month. A mathematical model has also been developed which permits the effect of the experimental parameters to predict. The model also explains the sensor behavior if different fluorophores are used, and shows that the analytical signal only slightly depends on the initial concentration of the O₂ in the sample.Finally an alternative sensor is presented based on a commercially available O₂ fluorescence sensor linked to catalase. This system gives an analytical behavior similar to that shown for the Cat–Ru sensor.

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