Creation of Environmentally Friendly Super "Dinitrotoluene Scavenger" Plants.

Pervasive environmental contamination due to the uncontrolled dispersal of 2,4-dinitrotoluene (2,4-DNT) represents a substantial global health risk, demanding urgent intervention for the removal of this detrimental compound from affected sites and the promotion of ecological restoration. Conventional methodologies, however, are energy-intensive, susceptible to secondary pollution, and may inadvertently increase carbon emissions. In this study, a 2,4-DNT degradation module is designed, assembled, and validated in rice plants. Consequently, the modified rice plants acquire the ability to counteract the phytotoxicity of 2,4-DNT. The most significant finding of this study is that these modified rice plants can completely degrade 2,4-DNT into innocuous substances and subsequently introduce them into the tricarboxylic acid cycle. Further, research reveals that the modified rice plants enable the rapid phytoremediation of 2,4-DNT-contaminated soil. This innovative, eco-friendly phytoremediation approach for dinitrotoluene-contaminated soil and water demonstrates significant potential across diverse regions, substantially contributing to carbon neutrality and sustainable development objectives by repurposing carbon and energy from organic contaminants.

Separation and identification of the DL-forms of short-chain peptides using a new chiral resolution labeling reagent.

There are several reports of D-amino acids being the causative molecules of serious diseases, resulting in the formation of, for example, prion protein and amyloid ß. D-Amino acids in peptides and proteins are typically identified by sequencing each residue by Edman degradation or by hydrolysis with hydrochloric acid for amino acid analysis. However, these approaches can result in racemization of the L-form to the D-form by hydrolysis and long pre-treatment for hydrolysis. To address these problems, we aimed to identify the DL-forms of amino acids in peptides without hydrolysis. Here, we showed that the DL-forms in peptides which are difficult to separate on a chiral column can be precisely separated by labeling with 1-fluoro-2,4-dinitrophenyl-5-D-leucine-N,N-dimethylethylenediamine-amide (D-FDLDA). Additionally, the peptides could be quantitatively analyzed using the same labeling method as for amino acids. Furthermore, the detection sensitivity of a sample labeled with D-FDLDA was higher than that of the conventional reagents Nα-(5-fluoro-2,4-dinitrophenyl)-L-alaninamide (L-FDAA) and Nα-(5-fluoro-2,4-dinitrophenyl)-L-leucinamide (L-FDLA) used in Marfey's method. The proposed method for identifying DL-forms of amino acids in peptides is a powerful tool for use in organic chemistry, biochemistry, and medical science.

Quantitative vapor delivery for improved canine threshold testing.

The canine olfactory system is a highly efficient and intricate tool often exploited by humans for detection for its many attributes, including impressive sensitivity to trace analyte vapors. Canine detectors are often touted as having lower limits of detection, or olfactory detection threshold (ODT), than other field-relevant detection technologies; however, previous attempts to quantify canine ODTs have resulted in reported estimates spanning multiple orders of magnitude, even for the same analyte. A major contributor to these discrepancies is the vapor delivery method used for testing, where losses due to adsorption and dilution are often unaccounted for, and the presence of unattended compounds in the vapor stream due to carryover may go unnoticed. In this research, a trace vapor generator (TV-Gen) was used to deliver quantitatively accurate amounts of vapor reproducibly over time for canine testing. Analyte losses due to adsorption to surfaces in the flow path, dilution in the sniff port at the outlet, and analyte carryover were considered. Computational fluid dynamic (CFD) modeling was used to visualize analyte vapor spread throughout the port. CFD simulations revealed the need for a diffuser to encourage the diffusion of the analyte throughout the port. As a result, the modified vapor generator provides analyte air as a diffuse flow that is evenly distributed through the custom sampling orifice, as opposed to a narrow stream of air at the chosen concentration which exits directly into the environment. Laboratory validations were carried out for three analytes, amyl acetate, 2,4-dinitrotoluene (DNT), and methyl benzoate. A linear response across more than two orders of magnitude vapor concentration range was achieved for all analytes. These efforts will be applied in further research utilizing this TV-Gen vapor delivery system for canine ODT testing, eliminating many quantitative changes seen previously. Graphical abstract.

One-Step Instantaneous Detection of Multiple Military and Improvised Explosives Facilitated by Colorimetric Reagent Design.

Although colorimetric detection based on reagents has been widely used in the fields of practical trace analysis, its versatility for detecting multitargets remains the most challenging problem. As a proof of concept, a general colorimetric reagent based on potassium isopropanol (C3H7KO) and dimethyl sulfoxide for one-step instantaneous detection and discrimination of typical military and improvised explosives was designed. Vivid colors from none to purple red, blue green, yellow green, and green were shown, respectively, when detecting 2,4,6-trinitrotoluene (TNT), 2,4-dinitrotoluene (DNT), elemental sulfur (S), and potassium permanganate (KMnO4). The unique design including the specific nucleophilic addition reaction and the base-catalyzed oxidation-induced electron transfer ensures perfect selectivity even upon facing more than 20 interferents. It is further experimentally demonstrated that the confinement effect introduced by Tween-20 plays an essential role in enhancing the color signal on the surface and thus boosts the detection performance even with a mass as low as 1.45 ng. The applicability of this versatile colorimetric reagent was further verified by integrating the reagent onto paper strips for the in-field identification of TNT, DNT, S, and KMnO4 with the help of a portable smartphone-based microscope apparatus, and a practical detection mass of 10.3 ng could be realized. We expect the present colorimetric reagent design strategy would pave a way for one-step instantaneous visual detection toward trace multianalytes.

Analytical methodology to screen UV-filters and synthetic musk compounds in market tomatoes.

A Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) methodology followed by gas chromatography-tandem mass spectrometry (GC-MS/MS) analysis was developed to extract thirteen synthetic musk compounds (SMCs: cashmeran, celestolide, phantolide, traseolide, galaxolide, tonalide, musk ambrette, musk xylene, musk ketone, musk tibetene, musk moskene, ethylene brassylate and exaltolide) and six ultraviolet-filters (UVFs: 2-ethylhexyl 4-dimethylaminobenzoate, 3-(4'-methylbenzylidene) camphor, 2-ethylhexyl 4-methoxycinnamate, 2-ethylhexyl 2-cyano-3,3-diphenylacrylate, benzophenone and drometrizole trisiloxane) from tomatoes. The proposed methodology was optimized: 2 g of freeze-dried tomato was extracted with 4 mL of water and 10 mL of ethyl acetate, adding 6 g of MgSO4 and 1.5 g of NaCl, then a dispersive solid-phase extraction was performed using 3 g of MgSO4, 300 mg of primary-secondary amino adsorbent (PSA) and 300 mg of octadecyl-silica (C18). Validation delivered recoveries between 81 (celestolide) and 119% (musk tibetene), with relative standard deviations <10%. The instrumental limit of detection varied from 0.02 (2-ethylhexyl 4-methoxycinnamate) to 3.00 pg (exaltolide and musk xylene). Regarding the method quantification limits, it ranged between 0.4 (celestolide) and 47.9 ng g-1 dw (exaltolide). The method was applied to different varieties of tomatoes (Solanum lycopersicum), revealing UVFs and SMCs between 1 and 210 ng g-1 dw. Higher concentrations were found for benzophenone (29-210 ng g-1 dw) and galaxolide (9-53 ng g-1 dw). The risk associated to the ingestion of contaminated tomatoes has also been estimated, showing that a potential health risk is unlikely.

Implementation of Combinatorial Genetic and Microenvironmental Engineering to Microbial-Based Field-Deployable Microbead Biosensors for Highly Sensitive and Remote Chemical Detection.

Bioreporters, microbial species genetically engineered to provide measurable signals in response to specific chemicals, have been widely investigated as sensors for biomedical and environmental monitoring. More specifically, the bioreporter encapsulated within a biocompatible material, such as a hydrogel that can provide a suitable microenvironment for its prolonged activity as well as efficient scalable production, has been viewed as a more broadly applicable mode of biosensors. In this study, alginate-based microbeads encapsulated with the bacterial bioreporter capable of expressing green fluorescence protein in response to nitro compounds (e.g., trinitrotoluene and dinitrotoluene) are developed as biosensors. To significantly enhance the sensitivity of the microbial-based microbead biosensors, "multifaceted" modification strategies are simultaneously employed: (1) multiple genetic modifications of the bioreporter, (2) tuning the physicomechanical properties of the encapsulating microbeads, (3) controlling the initial cell density within the microbeads, and (4) enrichment of nitro compounds inside microbeads via functional nanomaterials. These microbial and microenvironmental engineering approaches combine to significantly enhance the sensing capability, even allowing highly sensitive remote detection under a low-vapor phase. Thus, the strategy developed herein is expected to contribute to various cell-based biosensors.

Ti-Catalyst Biomimetic Sensor for the Detection of Nitroaromatic Pollutants.

An electrode modified by a Ti complex in Nafion was used in the electrochemical detection of nitroaromatic compounds (NACs). The catalyst reduced nitroaromatic groups by a six-electron process with a high apparent rate constant, k1, which was estimated to be at least 19 000 mol-1 L s-1 by simulation of the first step of the cathodic reduction of 4-nitrophenylacetic acid, which was used as model molecule. The modified electrode was prepared and optimized in terms of Nafion and catalyst concentrations. In the chosen analytical medium, the Ti-catalyst-containing electrode showed good stability in the presence of nitro species. A standard deviation around 15% was calculated on the analysis performed with 28 different modified electrodes, which could be expected for a drop-coating process. We show that the modified electrode detects several nitroaromatic compounds such as nitrophenols, mononitroaniline, and dinitrotoluene with detection limits ranging from 1 × 10-4 to 9 × 10-4 g L-1 (0.2 × 10-6 to 5.1 × 10-6 mol L-1). As an interesting feature, the sensor exhibits good selectivity toward NAC detection because titanocene does not catalyze the reduction of nitroaliphatic compounds.

Electrochemiluminescence on smartphone with silica nanopores membrane modified electrodes for nitroaromatic explosives detection.

Silica nanopores have electron channels and ion channels interpenetrating each other, which prompt the use of this structure for creating efficient electronic devices. In this study, silica nanopores membrane modified screen printed electrodes were applied in a smartphone-based electrochemiluminescence system for nitroaromatic explosives detection. Universal serial bus-on the go (USB-OTG) and camera on smartphone were used as the electrical stimulation and luminescence capture, respectively. ⎕Multimode methods including (red-green-blue) RGB, (hue-saturation-brightness) HSB, and Gray were proposed for luminescence analysis. Specific polypeptides were immobilized on the nanopores modified electrodes for nitroaromatic explosives sensing. With positive-charged tris(2,2'-bipyridyl)ruthenium(II) (Ru(bpy)32+) as electrochemiluminescence label, the increase in luminescence was associated with the selective ion channels and the well-conductive electron channels in the negative-charged nanopores. Besides, on account of the large specific surface area, nanopores modified screen printed electrodes showed stable and uniform luminescence. Results showed that the nanopores-enhanced electrochemiluminescence on smartphone covered a linear dynamic range from 10-7 mg/mL to 10-3 mg/mL for nitroaromatic explosives detection with the detection limit of 2.3 × 10-9 mg/mL. Therefore, high-efficient photo-electricity conversion capabilities of nanopores made it a kind of promising platform for sensitive and stable electrochemiluminescence. Furthermore, smartphone-based electrochemiluminescence with disposable screen printed electrodes could facilitate the mobile monitoring of biochemical analytes in the fields of environment, security, and health.

Electrochemical Determination of TNT, DNT, RDX, and HMX with Gold Nanoparticles/Poly(Carbazole-Aniline) Film-Modified Glassy Carbon Sensor Electrodes Imprinted for Molecular Recognition of Nitroaromatics and Nitramines.

Since nitroaromatic- and nitramine-type energetic materials, mostly arising from military activities, are persistent pollutants in soil and groundwater, on-site sensing of these hazardous chemicals has gained importance. A novel electrochemical sensor was designed for detecting nitroaromatic- and nitramine-type energetic materials, relying on gold nanoparticles (Aunano), modified glassy carbon (GC) electrode coated with nitro-energetic memory-poly(carbazole-aniline) copolymer (Cz- co-ANI) film (e.g., TNT memory-GC/P(Cz- co-ANI)-Aunano modified electrode). Current was recorded against concentration to build the calibration curves that were found to be linear within the range of 100-1000 µg L-1 for 2,4,6-trinitrotoluene (TNT) and 2,4-dinitrotoluene (DNT): 50-1000 µg L-1 for 1,3,5-trinitro-1,3,5-triazacyclohexane (RDX) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX). The corresponding limits of detection were 25 µg L-1 for TNT, 30 µg L-1 for DNT, and 10 µg L-1 for both RDX and HMX, using nitro-energetic memory-GC/P(Cz- co-ANI)-Aunano electrodes. These electrodes were used separately, and specific determinations were made in various mixtures of nitro-energetic materials. The developed method could be efficiently used in electroanalyzing nitroaromatics and nitramines in military explosives (i.e., comp B, octol, and comp A5). The sensor electrodes were specific for the tested nitro-energetic compounds and did not respond to paracetamol-caffeine-based analgesic drug, acetylsalicylic acid (aspirin), sweetener, and sugar that can be used as camouflage materials in passenger belongings. The developed method was statistically validated against the standard LC-MS reference method in contaminated clay soil samples containing TNT and RDX explosives.

In situ pilot test for bioremediation of energetic compound-contaminated soil at a former military demolition range site.

Bioremediation was performed in situ at a former military range site to assess the performance of native bacteria in degrading hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) and 2,4-dinitrotoluene (2,4-DNT). The fate of these pollutants in soil and soil pore water was investigated as influenced by waste glycerol amendment to the soil. Following waste glycerol application, there was an accumulation of organic carbon that promoted microbial activity, converting organic carbon into acetate and propionate, which are intermediate compounds in anaerobic processes. This augmentation of anaerobic activity strongly correlated to a noticeable reduction in RDX concentrations in the amended soil. Changes in concentrations of RDX in pore water were similar to those observed in the soil suggesting that RDX leaching from the soil matrix, and treatment with waste glycerol, contributed to the enhanced removal of RDX from the water and soil. This was not the case with 2,4-DNT, which was neither found in pore water nor affected by the waste glycerol treatment. Results from saturated conditions and Synthetic Precipitation Leaching Procedure testing, to investigate the environmental fate of 2,4-DNT, indicated that 2,4-DNT found on site was relatively inert and was likely to remain in its current state on the site.

Energetic contaminants inhibit plant litter decomposition in soil.

Individual effects of nitrogen-based energetic materials (EMs) 2,4-dinitrotoluene (2,4-DNT), 2-amino-4,6-dinitrotoluene (2-ADNT), 4-amino-2,6-dinitrotoluene (4-ADNT), nitroglycerin (NG), and 2,4,6,8,10,12-hexanitrohexaazaisowurtzitane (CL-20) on litter decomposition, an essential biologically-mediated soil process, were assessed using Orchard grass (Dactylis glomerata) straw in Sassafras sandy loam (SSL) soil, which has physicochemical characteristics that support "very high" qualitative relative bioavailability for organic chemicals. Batches of SSL soil were separately amended with individual EMs or acetone carrier control. To quantify the decomposition rates, one straw cluster was harvested from a set of randomly selected replicate containers from within each treatment, after 1, 2, 3, 4, 6, and 8 months of exposure. Results showed that soil amended with 2,4-DNT or NG inhibited litter decomposition rates based on the median effective concentration (EC50) values of 1122 mg/kg and 860 mg/kg, respectively. Exposure to 2-ADNT, 4-ADNT or CL-20 amended soil did not significantly affect litter decomposition in SSL soil at ≥ 10,000 mg/kg. These ecotoxicological data will be helpful in identifying concentrations of EMs in soil that present an acceptable ecological risk for biologically-mediated soil processes.

Evaluation of NfsA-like nitroreductases from Neisseria meningitidis and Bartonella henselae for enzyme-prodrug therapy, targeted cellular ablation, and dinitrotoluene bioremediation.

OBJECTIVES: To characterize the activities of two candidate nitroreductases, Neisseria meningitidis NfsA (NfsA_Nm) and Bartonella henselae (PnbA_Bh), with the nitro-prodrugs, CB1954 and metronidazole, and the environmental pollutants 2,4- and 2,6-dinitrotoluene. RESULTS: NfsA_Nm and PnbA_Bh were evaluated in Escherichia coli over-expression assays and as His6-tagged proteins in vitro. With the anti-cancer prodrug CB1954, both enzymes were more effective than the canonical O2-insensitive nitroreductase E. coli NfsB (NfsB_Ec), NfsA_Nm exhibiting comparable levels of activity to the leading nitroreductase candidate E. coli NfsA (NfsA_Ec). NfsA_Nm is also the first NfsA-family nitroreductase shown to produce a substantial proportion of 4-hydroxylamine end-product. NfsA_Nm and PnbA_Bh were again more efficient than NfsB_Ec at aerobic activation of metronidazole to a cytotoxic form, with NfsA_Nm appearing a promising candidate for improving zebrafish-targeted cell ablation models. NfsA_Nm was also more active than either NfsA_Ec or NfsB_Ec with 2,4- or 2,6-dinitrotoluene substrates, whereas PnbA_Bh was relatively inefficient with either substrate. CONCLUSIONS: NfsA_Nm is a promising new nitroreductase candidate for several diverse biotechnological applications.

Bioconcentration factors and plant-water partition coefficients of munitions compounds in barley.

Plants growing in the soils at military ranges and surrounding locations are exposed, and potentially able to uptake, munitions compounds (MCs). The extent to which a compound is transferred from the environment into organisms such as plants, referred to as bioconcentration, is conventionally measured through uptake experiments with field/synthetic soils. Multiple components/phases that vary among different soil types and affect the bioavailability of the MC, however, hinder the ability to separate the effects of soil characteristics from the MC chemical properties on the resulting plant bioconcentration. To circumvent the problem, this work presents a protocol to measure steady state bioconcentration factors (BCFs) for MCs in barley (Hordeum vulgare L.) using inert laboratory sand rather than field/synthetic soils. Three MCs: 2,4,6-trinitrotoluene (TNT), 2,4-dinitrotoluene (2,4-DNT), and 2,4-dinitroanisole (2,4-DNAN), and two munition-like compounds (MLCs): 4-nitroanisole (4-NAN) and 2-methoxy-5-nitropyridine (2-M-5-NPYNE) were evaluated. Approximately constant plant biomass and exposure concentrations were achieved within a one-month period that produced steady state log BCF values: 0.62 ± 0.02, 0.70 ± 0.03, 1.30 ± 0.06, 0.52 ± 0.03, and 0.40 ± 0.05 L kgplant dwt-1 for TNT, 2,4-DNT, 2,4-DNAN, 4-NAN, and 2-M-5-NPYNE, respectively. Furthermore, results suggest that the upper-bounds of the BCFs can be estimated within an order of magnitude by measuring the partitioning of the compounds between barley biomass and water. This highlights the importance of partition equilibrium as a mechanism for the uptake of MCs and MLCs by barley from interstitial water. The results from this work provide chemically meaningful data for prediction models able to estimate the bioconcentration of these contaminants in plants.

3D Printed Electrodes for Detection of Nitroaromatic Explosives and Nerve Agents.

Three-dimensional (3D) printing has proven to be a versatile and useful technology for specialized applications in industry and also for scientific research. We demonstrate its potential use toward the electrochemical detection of nitroaromatic compounds 2,4,6-trinitrotoluene (TNT), 2,4-dinitrotoluene (DNT), and fenitrothion (FT). The detection of these compounds is of utmost importance in military and forensic applications. Stainless steel electrodes were fabricated by 3D printing, and the surface was electroplated with gold. The electrochemical performance of the 3D printed electrodes was compared to that of the conventionally employed glassy carbon electrode (GCE) and proved to be more sensitive toward the detection of all three nitroaromatic compounds. 3D printing of customizable electrodes provides a viable alternative to traditional electrodes for the analysis of samples with electrochemical methods.

Use of Mass Spectrometric Vapor Analysis To Improve Canine Explosive Detection Efficiency.

Canines remain the gold standard for explosives detection in many situations, and there is an ongoing desire for them to perform at the highest level. This goal requires canine training to be approached similarly to scientific sensor design. Developing a canine training regimen is made challenging by a lack of understanding of the canine's odor environment, which is dynamic and typically contains multiple odorants. Existing methodology assumes that the handler's intention is an adequate surrogate for actual knowledge of the odors cuing the canine, but canines are easily exposed to unintentional explosive odors through training material cross-contamination. A sensitive, real-time (∼1 s) vapor analysis mass spectrometer was developed to provide tools, techniques, and knowledge to better understand, train, and utilize canines. The instrument has a detection library of nine explosives and explosive-related materials consisting of 2,4-dinitrotoluene (2,4-DNT), 2,6-dinitrotoluene (2,6-DNT), 2,4,6-trinitrotoluene (TNT), nitroglycerin (NG), 1,3,5-trinitroperhydro-1,3,5-triazine (RDX), pentaerythritol tetranitrate (PETN), triacetone triperoxide (TATP), hexamethylene triperoxide diamine (HMTD), and cyclohexanone, with detection limits in the parts-per-trillion to parts-per-quadrillion range by volume. The instrument can illustrate aspects of vapor plume dynamics, such as detecting plume filaments at a distance. The instrument was deployed to support canine training in the field, detecting cross-contamination among training materials, and developing an evaluation method based on the odor environment. Support for training material production and handling was provided by studying the dynamic headspace of a nonexplosive HMTD training aid that is in development. These results supported existing canine training and identified certain areas that may be improved.

Assessing seasonal variation of synthetic musks in beach sands from Oporto coastal area: A case study.

Synthetic musk compounds are widely used in the formulation of several cosmetics, personal care and household products. Due to their massive and widespread use, together with some health concerns, they are considered emerging pollutants and have been detected in different environmental compartments. This study focused on the evaluation of the concentration of synthetic musks (five nitro, five polycyclic and one macrocyclic musks) in beach sands, from Oporto coastal area (Portugal), contributing to the enhancement of the knowledge of levels, trends and behaviour of these compounds in this particular matrix. To accomplish this task, a QuEChERS methodology ("Quick, Easy, Cheap, Effective, Rugged, and Safe") coupled to gas chromatography-mass spectrometry (GC-MS) was successfully used to determine synthetic musks from beach sand. The chosen methodology proved to be suitable, achieving satisfactory results for precision (relative standard deviation values below 15%), accuracy (average recovery of 97%) and limits of detection (below 38 pg g-1). Synthetic musks were detected in all 45 analysed samples, in concentrations ranging from 0.01 to 27 ng g-1dw. Tonalide (93%), exaltolide (89%) and galaxolide (76%) were the most commonly detected compounds, but also those detected in higher concentrations (up to 27 ng g-1dw). Musk ambrette, moskene, tibetene and xylene were not detected in any of the samples. Higher concentrations were as expected detected in the Summer (total average concentration of 9.21 ng g-1dw), namely in samples from Valadares Sul (29 ng g-1dw), Francelos (25 ng g-1dw) and Castelo do Queijo (25 ng g-1dw). The preliminary environmental risk assessment study based on the determination of hazard quotients revealed that the presence of analysed compounds (tonalide, galaxolide and musk ketone) seems to pose no risk to the studied environmental compartment.

In-situ functionalized monolithic polysiloxane-polymethacrylate composite materials from polythiol-ene double click reaction in capillary column format for enantioselective nano-high-performance liquid chromatography.

This work reports on the proof-of-principle of preparation of novel one step in-situ functionalized monolithic polysiloxane-polymethacrylate composite materials in capillary columns for enantioselective nano-HPLC using a thiol-ene click reaction. Quinine carbamate as functional monomer and ethylene dimethacrylate as crosslinker were both used as ene components in a thermally initiated double click-type polymerization reaction with poly(3-mercaptopropyl)methylsiloxane as thiol component in presence of 1-propanol as porogenic solvent. Elemental analysis and on-capillary fluorescence measurement proved the successful incorporation of the functional chiral monomer into the polymer. Scanning electron microscopy images revealed a macroporous polymer morphology which is typical for a nucleation and growth mechanism of pore formation. The individual microglobules appear relatively spherical and smooth indicating a non-porous nature. Nano-HPLC experiments of the chiral monolithic capillary column provided successful enantiomer separation of N-3,5-dinitrobenzoylleucine as test compound in polar organic elution mode clearly documenting the successful implementation of the proposed concept towards new functionalized monolithic composite materials.

Thread-based microfluidic chips as a platform to assess acetylcholinesterase activity.

In this paper, a microfluidic thread-based analytical device (µTAD) to assess the activity of acetylcholinesterase (AChE) via colorimetric analylsis is described. Fabrication of the device consists of two platforms, both with a nylon thread trifurcated into three channels terminating at open analysis sites at the end of the thread. 5,5'-Dithiobis-(2-nitrobenzoic acid) (DTNB) was spotted and dried on the analysis sites. Acetylthiocholine iodide (ATC) (or cysteine, Cys) is transported through an inlet channel of the nylon thread by capillary action due to the hydrophilic nature of nylon. AChE is transported through the other inlet channel and mixes with the ATC (or Cys) as they travel up to the analysis sites. As the solution reaches the analysis sites, an intense yellow color change occurs indicating the reaction of the thiol with DTNB to produce the yellow anion TNB2- . The sites are then dried, scanned, yielding a linear range of inverse yellow mean intensity versus substrate concentration. An IC50 value (1.74 nM) with a known inhibitor, neostigmine bromide (NB), is obtained on the device. The multiplex design enables triplicate data collection in a device that is easy to use. µTADs have great potential to be employed in a myriad of tests including point-of-care diagnostic devices for resource-challenged settings.

A microfluidic paper-based device to assess acetylcholinesterase activity.

Neurotransmitters play key roles in cell-to-cell communication. These chemical messengers are involved in many functional processes, including growth, reproduction, memory, and behavior. In this communication, we describe a novel microfluidic paper-based analytical device (µPAD) to detect acetylcholinesterase (AChE) activity and inhibitor screening through a colorimetric analysis. The µPAD is easily fabricated via a wax printing process whereby wax is deposited onto the surface of chromatographic paper, and heated to create a hydrophobic barrier. Separate solutions of 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) and samples containing AChE and acetylthiocholine iodide (ATC) (or cysteine, Cys), respectively, are directly spotted onto the µPAD. DTNB and AChE/ATC (or Cys) flow towards each other where a reaction occurs to form the yellow colored 2-nitro-5-thiobenzoic acid anion (TNB2- ). The device is dried, scanned, and analyzed yielding a linear range of average inverse yellow intensities versus substrate concentration. An IC50 value (0.045 nM) with a known inhibitor, neostigmine bromide (NB), is obtained on the device. µPADs are low cost and easy to fabricate and have great potential to quantify neurotransmitter activity.

Acute toxicity assessment of explosive-contaminated soil extracting solution by luminescent bacteria assays.

Explosive-contaminated soil is harmful to people's health and the local ecosystem. The acute toxicity of its extracting solution was tested by bacterial luminescence assay using three kinds of luminescent bacteria to characterize the toxicity of the soil. An orthogonal test L 16 (45) was designed to optimize the soil extracting conditions. The optimum extracting conditions were obtained when the ultrasonic extraction time, ultrasonic extraction temperature, and the extraction repeat times were 6 h, 40 °C, and three, respectively. Fourier transform infrared spectroscopy (FTIR) results showed that the main components of the contaminated soil's extracting solution were 2,4-dinitrotoluene-3-sulfonate (2,4-DNT-3-SO3-); 2,4-dinitrotoluene-5-sulfonate (2,4-DNT-5-SO3-); and 2,6-dinitrotoluene (2,6-DNT). Compared with Photobacterium phosphoreum and Vibrio fischeri, Vibrio qinghaiensis sp. Nov. is more suitable for assessing the soil extracting solution's acute toxicity. Soil washing can remove most of the contaminants toxic to luminescent bacterium Vibrio qinghaiensis sp. Nov., suggesting that it may be a potential effective remediation method for explosive-contaminated soil.