RESUMO
The development of sensitive methodologies for detecting agrochemicals has become important in recent years due to the increasingly indiscriminate use of these substances. In this context, nanosensors based on atomic force microscopy (AFM) tips are useful because they provide higher sensitivity with operation at the nanometer scale. In this paper we exploit specific interactions between AFM tips functionalized with the enzyme acetolactate synthase (ALS) to detect the ALS-inhibitor herbicides metsulfuron-methyl and imazaquin. Using atomic force spectroscopy (AFS) we could measure the adhesion force between tip and substrate, which was considerably higher when the ALS-functionalized tip (nanobiosensor) was employed. The increase was approximately 250% and 160% for metsulfuron-methyl and imazaquin, respectively, in comparison to unfunctionalized probes. We estimated the specific enzyme-herbicide force by assuming that the measured force comprises an adhesion force according to the Johnson-Kendall-Roberts (JKR) model, the capillary force and the specific force. We show that the specific, biorecognition force plays a crucial role in the higher sensitivity of the nanobiosensor, thus opening the way for the design of similarly engineered tips for detecting herbicides and other analytes.
Assuntos
Técnicas Biossensoriais/métodos , Enzimas Imobilizadas/química , Herbicidas/análise , Microscopia de Força Atômica/métodos , Nanotecnologia/métodos , Acetolactato Sintase/química , Acetolactato Sintase/metabolismo , Sulfonatos de Arila/análise , Sulfonatos de Arila/metabolismo , Enzimas Imobilizadas/metabolismo , Herbicidas/metabolismo , Imidazóis/análise , Imidazóis/metabolismo , Quinolinas/análise , Quinolinas/metabolismoRESUMO
The use of agrochemicals has increased considerably in recent years, and consequently, there has been increased exposure of ecosystems and human populations to these highly toxic compounds. The study and development of methodologies to detect these substances with greater sensitivity has become extremely relevant. This article describes, for the first time, the use of atomic force spectroscopy (AFS) in the detection of enzyme-inhibiting herbicides. A nanobiosensor based on an atomic force microscopy (AFM) tip functionalised with the acetolactate synthase (ALS) enzyme was developed and characterised. The herbicide metsulfuron-methyl, an ALS inhibitor, was successfully detected through the acquisition of force curves using this biosensor. The adhesion force values were considerably higher when the biosensor was used. An increase of ~250% was achieved relative to the adhesion force using an unfunctionalised AFM tip. This considerable increase was the result of a specific interaction between the enzyme and the herbicide, which was primarily responsible for the efficiency of the nanobiosensor. These results indicate that this methodology is promising for the detection of herbicides, pesticides, and other environmental contaminants.
Assuntos
Sulfonatos de Arila/análise , Técnicas Biossensoriais/métodos , Microscopia de Força Atômica/instrumentação , Nanopartículas/química , Acetolactato Sintase/antagonistas & inibidores , Acetolactato Sintase/metabolismo , Sulfonatos de Arila/farmacologia , Colorimetria , Ensaios Enzimáticos , Inibidores Enzimáticos/farmacologia , Herbicidas/toxicidade , Humanos , Proteínas Recombinantes/metabolismo , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
The increasing importance of studies on soft matter and their impact on new technologies, including those associated with nanotechnology, has brought intermolecular and surface forces to the forefront of physics and materials science, for these are the prevailing forces in micro and nanosystems. With experimental methods such as the atomic force spectroscopy (AFS), it is now possible to measure these forces accurately, in addition to providing information on local material properties such as elasticity, hardness and adhesion. This review provides the theoretical and experimental background of afs, adhesion forces, intermolecular interactions and surface forces in air, vacuum and in solution.