Phenolic compounds are important environmental contaminants due to their high toxicity and persistence in the environment. The use of enzyme-based electrochemical biosensors is a simple, sensitive and low-cost alternative for the determination of these pollutants in contaminated waters. However, in most cases, it is impossible to detect specific compounds in a mixture of phenols due to signal-overlap, as the instruments operate at very close potentials, given that the system is based on a single enzyme to detect similar structures. In order to overcome this problem, in the present work we have successfully used multivariate calibration with partial least squares (PLS) for the simultaneous determination of hydroquinone and guaiacol by a tyrosinase-based biosensor that was assembled using an enzyme extract from yam. The use of PLS allowed us to work with a large number of voltammograms, leading to a single mathematical model for the simultaneous determination of phenols of similar structure in real samples with concentration values of mmol L-1.

In this work, a simple electrochemical biosensor for 4-chlorophenol was developed based on laccase immobilized on a hybrid nanocomposite (ZnO nanoparticles/chitosan), and incorporated in a carbon paste electrode. There are few biosensors in the literature for this specific pollutant because it tends to form polymeric films on the electrode, causing surface passivation or even enzyme inactivation. The carbon paste allowed the surface to be easily renewed by polishing, which amends this limitation. To optimize the experimental conditions, we used cyclic voltammetry and hydroquinone as a representative of phenolic compounds due to the high toxicity of chlorophenol, thus avoiding the generation of hazardous residues. After optimization, a calibration curve was constructed for 4-chlorophenol using differential pulse voltammetry, and a linear response was obtained from 1 to 50 µM, with a lower detection limit of 0.7 µM. The obtained biosensor showed high accuracy when employed in the analysis of industrial wastewater.

O presente trabalho de revisão aborda os mais recentes avanços na tecnologia de biossensores alcançados através da montagem de biomoléculas associada com nanopartículas de ouro na construção de dispositivos analíticos. Esta revisão está dividida de acordo com a biomolécula empregada no desenvolvimento de biossensores: (i) compostos imunológicos; (ii) DNA/RNA funcionais; e (iii) enzimas e proteínas Heme. A fim de facilitar a compreensão, cada seção foi subdividida de acordo com o modo de transdução. Os imunossensores contendo nanopartículas de ouro têm uma ampla gama de aplicações nos campos alimentício, ambiental, farmacêutico, químico e de diagnósticos clínicos. As nanopartículas foram empregadas para melhorar o sinal analítico ou a imobilização dos imunocompostos. Em outra seção, os biossensores DNA/RNA empregando nanoestruturas de ouro como labels e biossensores label-free associados a nanoestruturas de ouro como transdutores foram sistematicamente relatados para a rápida identificação de patógenos, espécies de interesse ambiental e diagnóstico clínico. A inclusão de nanopartículas de ouro em eletrodos modificados aumenta a transferência de elétrons entre o transdutor e a biomolécula proporcionando um melhor desempenho quando enzimas e proteínas redox heme são usados. Biossensores para a detecção e quantificação de glicose e peróxido de hidrogênio foram também discutidos.

The present review discusses the latest advances in biosensor technology achieved by the assembly of biomolecules associated with gold nanoparticles in analytical devices. This review is divided in sections according to the biomolecule employed in the biosensor development: (i) immunocompounds; (ii) DNA/RNA and functional DNA/RNA; and (iii) enzymes and Heme proteins. In order to facilitate the comprehension each section was subdivided according to the transduction mode. Gold nanoparticles based immunosensors have a wide range of applications in food, environmental, pharmaceutical, chemistry and clinical diagnostics. The nanoparticles were employed to improve whether the analytical signal or the immunocompounds immobilization. In another section, biosensors based on DNA/RNA biomolecules employing gold nanostructures as labels and label-free funtional DNA/RNA biosensors associated to gold nanostructures as tranducers were systematically reported for rapid identification of pathogens, species of environmental interest and clinical diagnostics, respectively. The inclusion of gold nanoparticles in modified electrodes itself enhances the electron transfer between the transducer and biomolecules leading to improved bioanalytical devices when redox enzymes and heme proteins are used. Biosensors for the detection and quantification of glucose and hydrogen peroxide are discussed as well.

Este trabalho descreve a construção de um filme de multicomponentes via método layer-by-layer (LbL) e a análise cinética da interação da enzima peroxidase de raiz forte (HRP) sobre a camada de ds-DNA de timo de bezerro formada numa superfície de ouro modificada. As técnicas de ressonância de plásmon de superfície (SPR) e a espectroscopia de impedância eletroquímica (EIE) foram usadas para caracterizar a construção do filme sobre a superfície de ouro. A ressonância de plásmon de superfície forneceu informações essenciais para o estudo e caracterização da interação proteína e ácido nucléico. Este método não necessita de marcador e monitora as interações em tempo real. Os estudos cinéticos obtidos por SPR da formação do filme de HRP sobre o ds-DNA apresentaram valores de 24,7 L mol-1 s-1 e 1,2×10-3 s-1 para k a e k d, respectivamente. A energia livre de Gibbs obtida para o sistema foi de - 23,1 kJ mol-1. Os resultados obtidos mostram que a interação das moléculas da enzima sobre o ds-DNA é cinética e termodinamicamente favoráveis e a interação entre as camadas ocorre principalmente pela atração de cargas opostas.

This paper describes the construction of a multicomponent film via layer-by-layer (LbL) method and the kinetic analysis of the interaction between horseradish peroxidase (HRP) enzyme on calf thymus double-stranded DNA layer on a modified gold surface. Surface plasmon resonance (SPR) and electrochemical impedance spectroscopy (EIS) are used to characterize the successful construction of the film on the gold surface. Surface plasmon resonance provided essential information for the study and characterization of protein and nucleic acid interaction and this method is label-free and monitors the interactions in real time. The kinetic studies determined by SPR of the horseradish peroxidase film formation on ds-DNA layer showed values of 24.7 L mol-1 s-1 and 1.2×10-3 s-1 for k a and k d, respectively. The Gibbs free energy obtained for the system was -23.1 kJ mol-1. The results obtained show that the interaction of the enzymes molecules on ds-DNA is kinetically and thermodynamically favourable and the interaction among the layers probably occurs mainly by attraction of opposite charges.

Tióis de cadeias carbônicas com tamanhos diferentes e contendo grupo terminal COOH foram usados para formar monocamadas auto-organizadas sobre a superfície de eletrodos de ouro. A transferência de elétrons do par Fe(CN)6(3-/4-) para o eletrodo foi estudada em diferentes pH usando-se as técnicas de voltametria cíclica (CV) e de espectroscopia de impedância eletroquímica (EIS). Mudanças no pH da solução resultaram em variações na carga do grupo terminal das monocamadas auto-organizadas e, conseqüentemente, em alterações na interação eletrostática da SAM com as espécies eletroativas em solução.

Thiols with different alkyl chain length and containing COOH terminal group were self-assembled on gold electrodes. The electron transfer of Fe(CN)6(3-/4-) couple to the electrode was studied at different pH by means cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). Changes in solution pH resulted in the charge variation of the self-assembled monolayer (SAM) terminal group and, consequently, the electrostatic interaction of SAM with the electroactive species in the solution.