Promoting laccase sensing activity for catechol detection using LBL assemblies of chitosan/ionic...

The performance of electrochemical laccase-based biosensors can be improved by immobilizing the enzyme on composite Layer-by-Layer (LbL) supports in which materials with complementary functions are combined. LbL films are formed by layers combining an electrocatalytic material which favors electron transfer (sulfonated copper phthalocyanine, CuPc^S(-)), an ionic liquid which enhances the electrical conductivity of the layers (1-butyl-3-methylimidazolium tetrafluoroborate, IL⁽⁺⁾) and a material able to promote enzyme immobilization (chitosan, CHI⁽⁺⁾). Composite films with different structures have been demonstrated to be efficient electrocatalysts, producing an increase in the magnitude of the responses towards catechol. The most intense and reproducible electrocatalytic effect was observed when a layer of the CuPc^S(-) was placed on top of a layer formed by a mixture of CHI⁽⁺⁾ + IL⁽⁺⁾ to obtain [CHI⁽⁺⁾ + IL⁽⁺⁾|CuPc^S(-)]₂ films. Biosensors with laccase immobilized on the surface of the LbL layers [CHI⁽⁺⁾ + IL⁽⁺⁾|CuPc^S(-)]₂|Lac showed mediated electron transfer between the redox enzyme and the film and a reproducibility of device-to-device performance of 4.1%. The amperometric biosensor showed a sensitivity of 0.237 A·M^-1 and a linear detection range from 2.4 μM to 26 μM for catechol. The excellent Limit of detection (LOD) of 8.96·10^-10 M (3·σ /m) is one order of magnitude lower than that obtained in similar studies. A Michaelis-Menten constant of 3.16 μM confirms excellent enzyme-substrate affinity.