A FRET Approach to Detect Paraoxon among Organophosphate Pesticides Using a Fluorescent Biosensor

The development of faster, sensitive and real-time methods for detecting organophosphate (OP) pesticides is of utmost priority in the in situ monitoring of these widespread compounds. Research on enzyme-based biosensors is increasing, and a promising candidate as a bioreceptor is the thermostable enzyme esterase-2 from Alicyclobacillus acidocaldarius (EST2), with a lipase-like Ser–His–Asp catalytic triad with a high affinity for OPs. This study aimed to evaluate the applicability of Förster resonance energy transfer (FRET) as a sensitive and reliable method to quantify OPs at environmentally relevant concentrations. For this purpose, the previously developed IAEDANS-labelled EST2-S35C mutant was used, in which tryptophan and IAEDANS fluorophores are the donor and the acceptor, respectively. Fluorometric measurements showed linearity with increased EST2-S35C concentrations. No significant interference was observed in the FRET measurements due to changes in the pH of the medium or the addition of other organic components (glucose, ascorbic acid or yeast extract). Fluorescence quenching due to the presence of paraoxon was observed at concentrations as low as 2 nM, which are considered harmful for the ecosystem. These results pave the way for further experiments encompassing more complex matrices

A 3D printable adapter for solid-state fluorescence measurements: the case of an immobilized enzymatic bioreceptor for organophosphate pesticides detection

The widespread use of pesticides in the last decades and their accumulation into the environment gave rise to major environmental and human health concerns. To address this topic, the scientific community pointed out the need to develop methodologies to detect and measure the presence of pesticides in different matrices. Biosensors have been recently explored as fast, easy, and sensitive methods for direct organophosphate pesticides monitoring. Thus, the present work aimed at designing and testing a 3D printed adapter useful on different equipment, and a membrane support to immobilize the esterase-2 from Alicyclobacillus acidocaldarius (EST2) bioreceptor. The latter is labelled with the IAEDANS, a bright fluorescent probe. EST2 was selected since it shows a high specificity toward paraoxon. Our results showed good stability and replicability, with an increasing linear fluorescent intensity recorded from 15 to 150 pmol of labelled EST2. Linearity of data was also observed when using the immobilized labelled EST2 to detect increasing amounts of paraoxon, with a limit of detection (LOD) of 0.09 pmol. This LOD value reveals the high sensitivity of our membrane support when mounted on the 3D adapter, comparable to modern methods using robotic workstations. Notably, the use of an independent support significantly simplified the manipulation of the membrane during experimental procedures and enabled it to match the specificities of different systems. In sum, this work emphasizes the advantages of using 3D printed accessories adapted to respond to the newest research needs. Graphical abstract: [Figure not available: see fulltext.

Fluorescent Chemosensors for Toxic Organophosphorus Pesticides: A Review

Many organophosphorus (OP) based compounds are highly toxic and powerful inhibitors of cholinesterases that generate serious environmental and human health concerns. Organothiophosphates with a thiophosphoryl (P=S) functional group constitute a broad class of these widely used pesticides. They are related to the more reactive phosphoryl (P=O) organophosphates, which include very lethal nerve agents and chemical warfare agents, such as, VX, Soman and Sarin. Unfortunately, widespread and frequent commercial use of OP-based compounds in agricultural lands has resulted in their presence as residues in crops, livestock, and poultry products and also led to their migration into aquifers. Thus, the design of new sensors with improved analyte selectivity and sensitivity is of paramount importance in this area. Herein, we review recent advances in the development of fluorescent chemosensors for toxic OP pesticides and related compounds. We also discuss challenges and progress towards the design of future chemosensors with dual modes for signal transduction. © 2010 by the authors; licensee MPDI, Basel, Switzerland

Biosensors for Environmental Monitoring

Real-time and reliable detection of molecular compounds and bacteria is essential in modern environmental monitoring. For rapid analyses, biosensing devices combining high selectivity of biomolecular recognition and sensitivity of modern signal-detection technologies offer a promising platform. Biosensors allow rapid on-site detection of pollutants and provide potential for better understanding of the environmental processes, including the fate and transport of contaminants.This book, including 12 chapters from 37 authors, introduces different biosensor-based technologies applied for environmental analyses

Applications of Graphene Quantum Dots in Biomedical Sensors

Due to the proliferative cancer rates, cardiovascular diseases, neurodegenerative disorders, autoimmune diseases and a plethora of infections across the globe, it is essential to introduce strategies that can rapidly and specifically detect the ultralow concentrations of relevant biomarkers, pathogens, toxins and pharmaceuticals in biological matrices. Considering these pathophysiologies, various research works have become necessary to fabricate biosensors for their early diagnosis and treatment, using nanomaterials like quantum dots (QDs). These nanomaterials effectively ameliorate the sensor performance with respect to their reproducibility, selectivity as well as sensitivity. In particular, graphene quantum dots (GQDs), which are ideally graphene fragments of nanometer size, constitute discrete features such as acting as attractive fluorophores and excellent electro-catalysts owing to their photo-stability, water-solubility, biocompatibility, non-toxicity and lucrativeness that make them favorable candidates for a wide range of novel biomedical applications. Herein, we reviewed about 300 biomedical studies reported over the last five years which entail the state of art as well as some pioneering ideas with respect to the prominent role of GQDs, especially in the development of optical, electrochemical and photoelectrochemical biosensors. Additionally, we outline the ideal properties of GQDs, their eclectic methods of synthesis, and the general principle behind several biosensing techniques.DFG, 428780268, Biomimetische Rezeptoren auf NanoMIP-Basis zur Virenerkennung und -entfernung mittels integrierter Ansätz

Recent Trends in Monitoring of European Water Framework Directive Priority Substances Using Micro-Sensors: A 2007–2009 Review

This review discusses from a critical perspective the development of new sensors for the measurement of priority pollutants targeted in the E.U. Water Framework Directive. Significant advances are reported in the paper and their advantages and limitations are also discussed. Future perspectives in this area are also pointed out in the conclusions. This review covers publications appeared since December 2006 (the publication date of the Swift report). Among priority substances, sensors for monitoring the four WFD metals represent 81% of published papers. None of analyzed publications present a micro-sensor totally validated in laboratory, ready for tests under real conditions in the field. The researches are mainly focused on the sensing part of the micro-sensors. Nevertheless, the main factor limiting micro-sensor applications in the environment is the ruggedness of the receptor towards environmental conditions. This point constitutes the first technological obstacle to be overcome for any long-term field tests

Fluorescent nanoparticles for sensing

Nanoparticle-based fluorescent sensors have emerged as a competitive alternative to small molecule sensors, due to their excellent fluorescence-based sensing capabilities. The tailorability of design, architecture, and photophysical properties has attracted the attention of many research groups, resulting in numerous reports related to novel nanosensors applied in sensing a vast variety of biological analytes. Although semiconducting quantum dots have been the best-known representative of fluorescent nanoparticles for a long time, the increasing popularity of new classes of organic nanoparticle-based sensors, such as carbon dots and polymeric nanoparticles, is due to their biocompatibility, ease of synthesis, and biofunctionalization capabilities. For instance, fluorescent gold and silver nanoclusters have emerged as a less cytotoxic replacement for semiconducting quantum dot sensors. This chapter provides an overview of recent developments in nanoparticle-based sensors for chemical and biological sensing and includes a discussion on unique properties of nanoparticles of different composition, along with their basic mechanism of fluorescence, route of synthesis, and their advantages and limitations

A Nanoparticle/enzyme System For The Simultaneous Detection And Decontamination Of Organophosphates

The need for a direct visual response system for the detection of organophosphorus compounds stems from the continued threat and use of these toxic agents in military and terrorist conflicts. The development of an enzyme-inhibitor triggered release system allows for direct visual detection with high specificity. Mesoporous silica nanoparticles (MSNs) have physical features that make them attractive as scaffolds for the construction of these systems, such as pore diameters (20-500 Á) that can be synthetically controlled, large surface areas (300-1500 m2g-1), large pore volumes, chemical inertness, stability at elevated temperatures, and surfaces that can be easily functionalized. In our studies, the dye Congo Red was loaded into the pores of MSNs, which were then capped by tethering an enzyme (organophosphorus hydrolase (OPH) or acetylcholinesterase (AChE)) to the external surfaces of MSNs through a competitive inhibitor (diethyl 4-aminobenzyl phosphonate (DEABP) or tacrine, respectively). OPH has been extensively studied for its ability to hydrolyze a wide range of organophosphorus compounds, rendering them non-toxic. AChE has been commonly used for organophosphate detection resulting from its sensitivity to phosphorylation. Upon addition of organophosphorus compounds to suspensions of the modified MSNs, the enzymes detached from the MSN surface, releasing the dye and providing a visual confirmation of organophosphate presence. Enzyme kinetics were studied using 31P NMR or UV-Visible spectroscopy; Congo Red release was also monitored by UV-Visible spectroscopy. The system was sensitive and specific for organophosphorus compounds both in phosphate-buffered saline and in human serum. The rate of dye release directly correlated with the rate of organophosphorus conversion for OPH and the rate of phosphorylation for AChE