Fluorescence assay for three organophosphorus pesticides in agricultural products based on Magnetic-Assisted fluorescence labeling aptamer probe
There has been increasing recent concern about the agricultural use of organophosphorus pesticides. A rapid and sensitive fluorescence assay for the detection of three organophosphorus pesticides has therefore been developed using 6carboxyfluorescein labeling aptamer as the probe and functionalized magnetic nanoparticles as the separation carrier. The aptamer hybridized with complementary DNA conjugated on the surface of the magnetic nanoparticles to form a magnetic aptamercomplementary DNA complex. Upon introducing the target organophosphorus pesticide, the aptamer departed from the complementary DNA, resulting in the fluorescence signal. Under optimized conditions, the limits of detection (LODs, SN 3) for trichlorfon, glyphosate, and malathion were 72.20 ng L1, 88.80 ng L1, and 195.37 ng L1, respectively. The method was applied for the detection of trichlorfon, glyphosate, and malathion in spiked lettuce and carrot samples. The recoveries were in the range of 79.4%118.7%, which were in good agreement with those obtained by gas chromatography, and the relative standard deviations were also acceptable. The method therefore has high sensitivity, so provides a means for the detection of multiple organophosphorus pesticides.
Attenuation of nitrates, antibiotics and pesticides from groundwater using immobilised microalgae-based systems
Groundwater pollution by nitrates and organic microcontaminants (OMCs) such as pesticides and antibiotics has increased in recent years due to the intensification of agriculture and livestock activities. Here we demonstrate, for the first time, the suitability of using microalgae immobilised in different materials (luffa sponge and polyurethane foam) to attenuate nitrates, antibiotics (sulfacetamide, sulfamethazine, and sulfamethoxazole), pesticides (bromacil, atrazine, diuron, bentazone, and mecoprop) from groundwater in two operational modes (batch and continuousfeeding). The results from the batch experiments show that OMC kinetic removal rates ranged from 0.01 to 0.18 d 1 , with halflives from 4 to 69 days. Immobilised microalgae in luffa and foam materials in the batch study was found to enhance the attenuation of selected OMCs from 36% to 51%, on average, after 10 operational days. Microalgae reactors in continuousfeeding operational mode out performed batch mode in terms of OMC removal (65% vs. 50%, on average) at a hydraulic residence time (HRT) of 8 days, whereas nitrate removal was greater in the batch experiments (81 vs. 48%, on average). OMC attenuation showed a high HRT dependence, but immobilised reactors were more resilient to the decrease in HRT. Further studies are needed, including the assessment of transformation products as well as the scaleup of the system to check the feasibility of the technology. Nevertheless, we expect our assay to be the starting point for the applicability of immobilisedmicroalgaebased systems for the treatment of polluted groundwater.
A label-free electrochemical aptasensor based on 3D porous CS/rGO/GCE for acetamiprid residue detection
A novel label-free electrochemical aptasensor was fabricated based on a three-dimensional porous electrode (3D-CS/rGO/GCE) for the detection of acetamiprid residues. The sensing signal was generated by the DNA itself. The porous electrode was prepared by electrodeposition in situ and characterized by scanning electron microscope (SEM), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). These results indicated that the porous electrode has a uniform nanoporous structure, high active area, and excellent conductivity, leading to improve the transmission efficiency of current signals. The 3D-CS/rGO/GCE was used to increase a load of acetamiprid aptamer on the electrode. Meanwhile, DNA self-assembly strategy was used to further increase the DNA amounts. Thus the electrochemical current was amplified significantly due to increased phosphate group amounts by the above synergistic effect. The determination of acetamiprid residues using square wave voltammetry (SWV) showed good sensitivity, with the linear range from 0.1 pM to 0.1 mu M and the detection limit was 71.2 fM. The label-free electrochemical aptasensor was also used to detect acetamiprid residues in tea samples with satisfactory results.
A portable device enabling fluorescent-to-electric resistant transduction for selective Cr3+ detection based on its slow ligand bind kinetics
Using simple and commonly available devices for signal reading has attracted much attention recently for DNA-based sensors. Most previous works focused on electrochemical or colorimetric signals. In this work, a portable device was structured by simple electronic and 3D-printed components to convert fluorescence signal to electric resistance with reading by a multimeter. To demonstrate its application, we took advantage of the slow ligand bind kinetics of Cr3+ to promote DNA adsorption on metal oxide nanoparticles (MONPs). Nanoceria (CeO2), screened from a total of 13 kinds of MONPs, possesses high specificity for Cr3+ from other metal ions. EDTA can mask other metal ions, while it cannot bind Cr3+ quickly so that Cr3+ promotes DNA adsorption and thus fluorescence quenching. The linear range is from 0 to 2 mu M Cr3+ with a LOD of 0.02 mu M. Using common electronic components and the 3D-printed miniaturized device, the sensor to transfer fluorescence to electric resistant signal was also measured by a digital multimeter. In this case, the LOD is about 1 mu M Cr3+. Such a strategy may provide a simple and general approach for fluorescent portable device fabrication and may find applications for other onsite sensing applications.
A novel fluorescent recombinant cell-based biosensor for screening NLRP3 inflammasome inhibitors
Nucleotide-binding oligomerization domain-like receptor pyrin-domain-containing protein 3 (NLRP3) inflammasomes trigger a defense mechanism in response to physiological or pathological reactions caused by various injuries, and they have critical roles in the development of many major diseases. The development of plant source anti-inflammatory agents and the assessment of their anti-inflammatory properties still face many challenges, such as high costs and long testing periods. Exogenous anti-inflammatory factors can be evaluated at the protein level using the NLRP3-green fluorescent protein (GFP) THP-1 cell-based biosensor. To achieve this, THP-1 cells were stably transfected with a plasmid encoding the GFP reporter gene that is under the transcriptional regulation of the NLRP3 promoter. Because the NLRP3 promoter controls the green fluorescent signal, the fluorescence is an indicator of NLRP3 activity. Our results show the fluorescence intensity was dependent on both the degree of inflammation and the effect of the anti-inflammatory agent. Here, we successfully used the biosensor to screen an NLRP3 inhibitor in the presence of polyphenols. Our results demonstrate the potential of this cell-based biosensor in monitoring the development of inflammation and in screening inflammatory inhibitors.
Ratiometric ATP detection on gliding microtubules based on bioorthogonal fluorescence conjugation
An ingenious microtubule functionalization strategy based on the novel bioorthogonal conjugation is developed to construct fluorescent ratiometric probes for ATP sensing. By single excitation, newly synthesized probe RT-1 exhibited green emission (485 nm) from the bioorthogonal fluorescence conjugation part and orange emission (584 nm) from the ATP sensing part. The fluorescence intensity ratio (I-584/I-485) displayed good linear response in ATP ranges of 0-2.5 mM (y = 0.9282x + 0.2398, R-2 = 0.9705, LOD = 0.0354 mM) and 2.0-10.0 mM (y = 2.7153x-3.6234, R-2 = 0.9911, LOD = 0.0121 mM). Moreover, a paclitaxel derivate probe RT-2 was developed to functionalize in vitro polymerized microtubule for in situ ATP detection. This simple and convenient strategy is anticipated to stimulate more microtubule related applications in both in vitro and intracellular studies.
Targeting tumor-intrinsic hexosamine biosynthesis sensitizes pancreatic cancer to anti-PD1 therapy
Pancreatic ductal adenocarcinoma (PDAC) is considered to be a highly immunosuppressive and heterogenous neoplasm. Despite improved knowledge regarding the genetic background of the tumor and better understanding of the tumor microenvironment, immune checkpoint inhibitor therapy (targeting CTLA4, PD1, PDL1) has not been very successful against PDAC. The robust desmoplastic stroma, along with an extensive extracellular matrix (ECM) that is rich in hyaluronan, plays an integral role in this immune evasion. Hexosamine biosynthesis pathway (HBP), a shunt pathway of glycolysis, is a metabolic node in cancer cells that can promote survival pathways on the one hand and influence the hyaluronan synthesis in the ECM on the other. The rate-limiting enzyme of the pathway, glutamine-fructose amidotransferase 1 (GFAT1), uses glutamine and fructose 6-phosphate to eventually synthesize uridine diphosphate N-acetylglucosamine (UDP-GlcNAc). In the current manuscript, we targeted this glutamine-utilizing enzyme by a small molecule glutamine analog (6-diazo-5-oxo-L-norleucine [DON]). Our results showed that DON decreased the self-renewal potential and metastatic ability of tumor cells. Further, treatment with DON decreased hyaluronan and collagen in the tumor microenvironment, leading to an extensive remodeling of the ECM and an increased infiltration of CD8+ T cells. Additionally, treatment with DON sensitized pancreatic tumors to anti-PD1 therapy, resulting in tumor regression and prolonged survival.
Development of a molecularly imprinted polymer electrochemical sensor and its application for sensitive detection and determination of malathion in olive fruits and oils
Malathion (MAL) is an organophosphorus (OP) insecticide. It is a cholinesterase inhibitor, which can pose serious health and environmental problems. In this study, a sensitive and selective molecular imprinted polymer (MIP) based on screenprinted gold electrodes (AuSPE) for MAL detection in olive oils and fruits, was devised. The MIP sensor was prepared using acrylamide as the functional monomer and MAL as the template. Subsequently, the morphology of the electrode surface was studied by scanning electron microscopy (SEM) and atomic force microscopy (AFM). The electrochemical characterization of the developed MIP sensor was performed by cyclic voltammetry (CV), differential pulse voltammetry (DPV), and electrochemical impedance spectroscopy (EIS) techniques. The operational repeatability and stability of the sensor were studied. It was found to have a dynamic concentration range of (0.1 pg mL 1 1000 pg mL 1 ) and a low limit of detection (LOD) of 0.06 pg mL 1 . Furthermore, the sensor was employed to determine MAL content in olive oil with a recovery rate of 87.9% and a relative standard deviation of 8%. It was successfully applied for MAL determination in real samples and promise to open new opportunities for the detection of OP pesticides residues in various food products, as well as in environmental applications.
Use of novel film forming starch complexes to directly and indirectly reduce insect damage to plants
Filmforming polymer technologies can contribute to insectpest control by causing adverse effects directly to insects such as slowing development, causing mortality, andor indirectly as a spray ingredient that improves pesticide efficacy by resisting environmental degradation such as washoff by rain. Amylose complexes produced from either fatty acid amylose sodium palmitate (NaPalm) or fatty ammonium salts amylosehexadecylammonium chloride complex (HexAm), when blended with polyvinyl alcohol (PVOH), dry to form films that were significantly more water resistant then films formed by their constitutive components and adhered strongly to treated soybean Glycine max (L.) Merrill leaves with no impact on photosynthesis or yield in simulated field trials. As an experimental bioinsecticide formulation to resist washoff, AfMNPV baculovirus was incorporated into both amylose complex formulations and sprayed on soybean. After a simulated rain event, soybean treated with the baculovirusNaPalmPVOH (1:1) retained 93% of the original insecticidal activity against Trichoplusia ni (Hbner) larvae as compared with the baculovirus only treatment, 53% activity retained. Curiously, initial insecticidal efficacy of filmforming treatments was significantly lower against T. ni when compared with unformulated baculovirus treatments, thus prompting further investigations. Both amylose complexes reduced larval feeding when sprayed on soybean, corresponding with reduced insect mortality by the baculovirus, which must be ingested to initiate infection. HexAm film coating over artificial diet inhibited the growth and development of T. ni and caused increased T. ni mortality and developmental delay as film thickness of HexAmPVOH increased. These investigations demonstrate the potential of a new modified starch complex formulation to serve as a protective film forming agent and contribute to control of insect pests.
Disinfection by-product formation during UV/Chlorine treatment of pesticides in a novel UV-LED reactor at 285 nm and the mitigation impact of GAC treatment
The UVChlorine process has gained attention in recent years due to the high quantum yield and absorbance of the chlorine species. However, there are still many unknowns around its application as a treatment for drinking water. The potential for the formation of disinfection byproducts (DBPs) is one of them. There are no studies reporting on the formation of trihalomethanes (THMs) or haloacetic acids (HAAs) in complex matrices, such as real source waters, at UV wavelengths tailored to the UVChlorine process, which has been possible thanks to the development of light emitting diodes (LEDs). In addition, consideration of mitigation measures that might be needed after UVChlorine treatment for full scale application have not been previously reported. Specifically, the novelty of this work resides in the use of an innovative reactor using UVLEDs emitting at 285 nm for the removal of three pesticides (metaldehyde, carbetamide and mecoprop), the evaluation of THM, HAA and bromate formation in real water sources by UVChlorine treatment and the mitigation effect of subsequent GAC treatment. A new parameter, the applied optical dose (AOD), has been defined for UV reactors, such as the one in the present study, where the irradiated volume is nonuniform. The results showed the feasibility of using the UVChlorine process with LEDs, although a compromise is needed between pH and chlorine concentration to remove pesticides while minimising DBP formation. Overall, the UVChlorine process did not significantly increase THM or HAA formation at pH 7.98.2 at the studied wavelength. At acidic pH, however, THM formation potential increased up to 30% after UVChlorine treatment with concentrations up to 60 gL. HAA formation potential increased between 100 and 180%, although concentrations never exceeded 35 gL. In all cases, GAC treatment mitigated DBP formation, reducing THM formation potential to concentrations between 3 and 16 gL, and HAA formation potential between 4 and 30 gL.