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This work presents a clustered regularly interspaced short palindromic repeat (CRISPR)/Cas-nanopipette nano-electrochemistry (Cas = CRISPR-associated proteins) capable of ultrasensitive microRNA detection. Nanoconfinement of the CRISPR/Cas13a within a nanopipette leads to a high catalytic efficacy of ca. 169 times higher than that in bulk electrolyte, contributing to the amplified electrochemical responses. CRISPR/Cas13a-enabled detection of representative microRNA-25 achieves a low limit of detection down to 10 aM. Practical application of this method is further demonstrated for single-cell and real human serum detection. Its general applicability is validated by addressing microRNA-141 and the SARS-CoV-2 RNA gene fragment. This work introduces a new CRISPR/Cas-empowered nanotechnology for ultrasensitive nano-electrochemistry and bioanalysis.
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MicroRNAs , Nanoporos , Humanos , MicroRNAs/genética , MicroRNAs/análise , Sistemas CRISPR-Cas/genética , RNA ViralRESUMO
Direct single-cell caspase-3 (Casp-3) analysis has remained challenging. A study of single-cell Casp-3 could contribute to revealing the fundamental pathogenic mechanisms in Casp-3-associated diseases. Here, a biomimetic nanochannel capable of single-cell sampling and ionic detection of intracellular Casp-3 is devised, which is established upon the installment of target-specific organic molecules (luc-DEVD) within the orifice of a glass nanopipette. The specific cleavage of luc-DEVD by Casp-3 could induce changes of inner-surface chemical groups and charge properties, thus altering the ionic response of the biomimetic nanochannel for direct Casp-3 detection. The practical applicability of this biomimetic nanochannel is confirmed by probing intracellular Casp-3 fluctuation upon drug stimulation and quantifying the Casp-3 evolution during induced apoptosis. This work realizes ionic single-cell Casp-3 analysis and provides a different perspective for single-cell protein analysis.
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Apoptose , Biomimética , Caspase 3/metabolismo , Apoptose/fisiologiaRESUMO
Plastic degradation by biological systems emerges as a prospective avenue for addressing the pressing global concern of plastic waste accumulation. The intricate chemical compositions and diverse structural facets inherent to polyurethanes (PU) substantially increase the complexity associated with PU waste management. Despite the extensive research endeavors spanning over decades, most known enzymes exhibit a propensity for hydrolyzing waterborne PU dispersion (i.e., the commercial Impranil DLN-SD), with only a limited capacity for the degradation of bulky PU materials. Here, we report a novel cutinase (CpCut1) derived from Cladosporium sp. P7, which demonstrates remarkable efficiency in the degrading of various polyester-PU materials. After 12-h incubation at 55°C, CpCut1 was capable of degrading 40.5% and 20.6% of thermoplastic PU film and post-consumer foam, respectively, while achieving complete depolymerization of Impranil DLN-SD. Further analysis of the degradation intermediates suggested that the activity of CpCut1 primarily targeted the ester bonds within the PU soft segments. The versatile performance of CpCut1 against a spectrum of polyester-PU materials positions it as a promising candidate for the bio-recycling of waste plastics.IMPORTANCEPolyurethane (PU) has a complex chemical composition that frequently incorporates a variety of additives, which poses significant obstacles to biodegradability and recyclability. Recent advances have unveiled microbial degradation and enzymatic depolymerization as promising waste PU disposal strategies. In this study, we identified a gene encoding a cutinase from the PU-degrading fungus Cladosporium sp. P7, which allowed the expression, purification, and characterization of the recombinant enzyme CpCut1. Furthermore, this study identified the products derived from the CpCut1 catalyzed PU degradation and proposed its underlying mechanism. These findings highlight the potential of this newly discovered fungal cutinase as a remarkably efficient tool in the degradation of PU materials.
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Hidrolases de Éster Carboxílico , Cladosporium , Poliuretanos , Poliuretanos/química , Poliuretanos/metabolismo , Cladosporium/genética , Cladosporium/metabolismo , Estudos Prospectivos , Biodegradação Ambiental , Poliésteres/metabolismo , PlásticosRESUMO
The sensorimotor system can recalibrate itself without our conscious awareness, a type of procedural learning whose computational mechanism remains undefined. Recent findings on implicit motor adaptation, such as over-learning from small perturbations and fast saturation for increasing perturbation size, challenge existing theories based on sensory errors. We argue that perceptual error, arising from the optimal combination of movement-related cues, is the primary driver of implicit adaptation. Central to our theory is the increasing sensory uncertainty of visual cues with increasing perturbations, which was validated through perceptual psychophysics (Experiment 1). Our theory predicts the learning dynamics of implicit adaptation across a spectrum of perturbation sizes on a trial-by-trial basis (Experiment 2). It explains proprioception changes and their relation to visual perturbation (Experiment 3). By modulating visual uncertainty in perturbation, we induced unique adaptation responses in line with our model predictions (Experiment 4). Overall, our perceptual error framework outperforms existing models based on sensory errors, suggesting that perceptual error in locating one's effector, supported by Bayesian cue integration, underpins the sensorimotor system's implicit adaptation.
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Adaptação Fisiológica , Teorema de Bayes , Sinais (Psicologia) , Humanos , Masculino , Adulto , Adulto Jovem , Feminino , Desempenho Psicomotor/fisiologia , Aprendizagem/fisiologia , Percepção Visual/fisiologia , Propriocepção/fisiologiaRESUMO
A continuously regenerated cationic impurity removal device (CR-CRD) has been fabricated and applied for ion chromatography (IC). The removal of cationic impurities is realized by electrodialytically replacing the cationic impurities with hydronium ions. The device is configured in a sandwich structure and the central eluent channel is respectively isolated from both electrodes by stacked cation exchange membranes and a bipolar membrane (BPM) plus stacked anion exchange membranes. The eluent channel is packed with cation exchange resins in hydronium form and their continuous regeneration can be achieved by electrodialysis. A desirable feature of the device is gas-free, and no degasser is required. It showed sufficient ability to remove cationic impurities, as indicated by > 99.9 % removal of 10 mL of 1 mM LiOH solution injected (â¼10 µmol) or continuous removal of 1 mM LiOH solution at the flow rate of 1 mL/min (1 µmol/min). A useful application was for sample pretreatment in nuclear power industry, by eliminating strong matrix interference of the sample containing LiOH (1 mM) and boric acid (2000 mg/L) with trace anion analysis.
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Cátions , Cromatografia por Troca Iônica/métodos , Cromatografia por Troca Iônica/instrumentação , Cátions/química , Membranas Artificiais , Resinas de Troca de Cátion/química , Desenho de EquipamentoRESUMO
The degradation kinetics, reaction pathways, and disinfection by-products formation of an organic UV filter, benzophenone-4 (BP4) during UV/persulfate oxidation were investigated. BP4 can hardly be degraded by UV alone, but can be effectively decomposed by UV/persulfate following pseudo-first order kinetics. BP4 degradation rate was enhanced with increasing persulfate dosage and decreasing pH from 8 to 5. However, the degradation rate of BP4 at pH 9 was higher than that at pH 8 because of the presence of phenolic group in BP4 structure. and SO4-â were confirmed as the major contributors to BP4 decomposition in radical scavenging experiments, and the second-order rate constants between HOâ and BP4 as well as those between SO4-â and BP4 were estimated by establishing and solving a kinetic model. The presence of Br- and humic acid inhibited the decomposition of BP4, while NO3- promoted it. The mineralisation of BP4 was only 9.1% at the persulfate concentration of 50 µM. Six degradation intermediates were identified for the promulgation of the reaction pathways of BP4 during UV/persulfate oxidation were proposed as a result. In addition, the formation of DBP in the sequential chlorination was evaluated at different persulfate dosages, pH values, and water matrix. The results of this study can provide essential knowledge for the effective control of DBP formation with reducing potential hazard to provide safe drinking water to the public.
BP4 can be effectively degraded by UV/persulfate process, following pseudo-first order kinetics.
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Soluble suppression of tumorigenicity-2 (sST-2), a marker of myocardial fibrosis and remodeling, has been related to the development of atrial fibrillation (AF). The aim of this meta-analysis was to evaluate the relationship between baseline serum sST-2 levels and the risk of AF recurrence after ablation. Relevant observational studies were retrieved from PubMed, Web of Science, Embase, Wanfang and China National Knowledge Infrastructure (CNKI). A random-effects model was used to combine the data, accounting for between-study heterogeneity. Fourteen prospective cohorts were included. Pooled results showed higher sST-2 levels before ablation in patients with AF recurrence compared to those without AF recurrence (standardized mean difference = 1.15, 95% confidence interval [CI] = 0.67 to 1.63, P < 0.001; I2 = 92%). Meta-regression analysis suggested that the proportion of patients with paroxysmal AF (PaAF) was positively related to the difference in serum sST-2 levels between patients with and without AF recurrence (coefficient = 0.033, P < 0.001). Subgroup analysis showed a more remarkable difference in serum sST-2 levels between patients with and without AF recurrence in studies where PaAF was ≥ 60% compared to those where it was < 60% (P = 0.007). Further analyses showed that high sST-2 levels before ablation were associated with an increased risk of AF recurrence (odds ratio [OR] per 1 ng/mL increment of sST-2 =1.05, OR for high versus low sST-2 = 1.73, both P values < 0.05). In conclusion, high sST-2 baseline levels may be associated with an increased risk of AF recurrence after catheter ablation.
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Polyurethane (PU) is a complex polymer synthesized from polyols and isocyanates. It contains urethane bonds that resist hydrolysis, which decreases the efficiency of biodegradation. In this study, we first expressed the amidase GatA250, and then, assessed the enzymatic characterization of GatA250 and its efficiency in degrading the polyester-PU. GatA250 degraded self-synthesized thermoplastic PU film and postconsumption foam with degradation efficiency of 8.17% and 4.29%, respectively. During the degradation, the film released 14.8 µm 4,4'-methylenedianiline (MDA), but 1,4-butanediol (BDO) and adipic acid (AA) were not released. Our findings indicated that GatA250 only cleaved urethane bonds in PU, and the degradation efficiency was extremely low. Hence, we introduced the cutinase LCC, which possesses hydrolytic activity on the ester bonds in PU, and then used both enzymes simultaneously to degrade the polyester-PU. The combined system (LCC-GatA250) had higher degradation efficiency for the degradation of PU film (42.2%) and foam (13.94%). The combined system also showed a 1.80 time increase in the production of the monomer MDA, and a 1.23 and 3.62 times increase in the production of AA and BDO, respectively, compared to their production recorded after treatment with only GatA250 or LCC. This study provides valuable insights into PU pollution control and also proposes applicable solutions to manage PU wastes through bio-recycling.
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Compostos de Anilina , Hidrolases de Éster Carboxílico , Poliésteres , Poliuretanos , Poliésteres/química , AmidoidrolasesRESUMO
The sharpening switch is one of the most important elements in the trigger generator, which is used to sharpen the front of the trigger pulse. The breakdown performance of the sharpening switch has an important influence on the output and stability of the trigger generator. In this paper, a novel 200 kV capacitance-coupling pre-ionized sharpening switch, which can realize pre-ionization by dividing voltage through its own structural capacitances, is proposed and investigated. In order to obtain the optimal parameters of the pre-ionized sharpening switch, the influences of the electrode structure, electrode material, main gap distance, and pre-ionized gap distance on the breakdown performance are studied experimentally. The experimental results show that the electrode structure with a circular knife-edged cathode and a plate-shaped anode has the smallest breakdown jitter, followed by the needle-plate structure and the tapered ball-head structure. The stainless steel electrode has the smallest jitter, followed by graphite, aluminum, and brass electrodes. When the gap distances of the main gap and the pre-ionized gap are 13 and 1.5 mm, respectively, the breakdown stability of the pre-ionized sharpening switch is the best. Under an input voltage pulse with a rise time of about one microsecond, the jitter of the capacitance-coupling pre-ionized sharpening switch with the optimal parameters is 6.08 ns, which is about 0.6% of the rise time of the input pulse. The jitter decreases by 44.5% compared to the switch without pre-ionization. The rise time of the output pulse is sharpened to 17 ns, corresponding to a voltage rise rate of more than 11 kV/ns.
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Weakly supervised object detection (WSOD) and semantic segmentation with image-level annotations have attracted extensive attention due to their high label efficiency. Multiple instance learning (MIL) offers a feasible solution for the two tasks by treating each image as a bag with a series of instances (object regions or pixels) and identifying foreground instances that contribute to bag classification. However, conventional MIL paradigms often suffer from issues, e.g., discriminative instance domination and missing instances. In this article, we observe that negative instances usually contain valuable deterministic information, which is the key to solving the two issues. Motivated by this, we propose a novel MIL paradigm based on negative deterministic information (NDI), termed NDI-MIL, which is based on two core designs with a progressive relation: NDI collection and negative contrastive learning (NCL). In NDI collection, we identify and distill NDI from negative instances online by a dynamic feature bank. The collected NDI is then utilized in a NCL mechanism to locate and punish those discriminative regions, by which the discriminative instance domination and missing instances issues are effectively addressed, leading to improved object-and pixel-level localization accuracy and completeness. In addition, we design an NDI-guided instance selection (NGIS) strategy to further enhance the systematic performance. Experimental results on several public benchmarks, including PASCAL VOC 2007, PASCAL VOC 2012, and MS COCO, show that our method achieves satisfactory performance. The code is available at: https://github.com/GC-WSL/NDI.
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Current research suggests that mitochondrial dysfunction can be a contributing factor in the development of cardiac arrhythmias. In pursuit of elucidating the causal link between the biological functions of mitochondria and the occurrence of atrial fibrillation/flutter, we conducted a 2-sample Mendelian randomization (MR) study. Mitochondrial proteins were selected for exposure in this study. To enhance the accuracy of our study, we selected data on AF/AFL from the FinnGen study and the UK Biobank for MR analysis, respectively. The inverse variance-weighted method was utilized as the primary analysis technique for MR. In addition, we performed a series of sensitivity analyses to detect heterogeneity and horizontal pleiotropy. MR results indicated a significant positive association between NAD-dependent protein deacylase sirtuin-5 and AF/AFL (odds ratioâ =â 1.084, 95% confidence interval: 1.037-1.133, Pâ =â 3.679â ×â 10-4, Adjusted Pâ =â .024), with consistent outcomes observed in replication analysis (odds ratioâ =â 1.002, 95% confidence interval: 1.001-1.003, Pâ =â 4.808â ×â 10-4, Adjusted Pâ =â .032). NAD-dependent protein deacylase sirtuin-5 can significantly promote the occurrence of AF/AFL, and its specific mechanisms warrant further investigation.
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Fibrilação Atrial , Flutter Atrial , Análise da Randomização Mendeliana , Fibrilação Atrial/genética , Fibrilação Atrial/epidemiologia , Fibrilação Atrial/etiologia , Humanos , Flutter Atrial/genética , Flutter Atrial/epidemiologia , Sirtuínas/genética , Mitocôndrias/genética , Polimorfismo de Nucleotídeo Único , Fatores de RiscoRESUMO
The worldwide dissemination of New Delhi metallo-ß-lactamase-1 (NDM-1), which mediates resistance to almost all clinical ß-lactam antibiotics, is a major public health problem. The global distribution, species, sources, and potential transfer risk of blaNDM-1-carrying bacteria are unclear. Results of a comprehensive analysis of literature in 2010-2022 showed that a total of 6002 blaNDM-1 carrying bacteria were widely distributed around 62 countries with a high trend in the coastal areas. Opportunistic pathogens or pathogens like Klebsiella sp., Escherichia sp., Acinetobacter sp. and Pseudomonas sp. were the four main species indicating the potential microbial risk. Source analysis showed that 86.45 % of target bacteria were isolated from the source of hospital (e.g., Hospital patients and wastewater) and little from surface water (5.07 %) and farms (3.98 %). A plasmid-encoded blaNDM-1Acinetobacter sp. with the resistance mechanisms of antibiotic efflux pump, antibiotic target change and antibiotic degradation was isolated from the wastewater of a typical tertiary hospital. Insertion sequences (IS3 and IS30) located in the adjacent 5 kbp of blaNDM-1-bleMBL gene cluster indicating the transposon-mediated horizontal gene transfer risk. These results showed that the worldwide spread of blaNDM-1-carrying bacteria and its potential horizontal gene transfer risk deserve good control.
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Transferência Genética Horizontal , beta-Lactamases , beta-Lactamases/genética , Bactérias/genética , Antibacterianos/farmacologia , Plasmídeos/genéticaRESUMO
Reinjecting produced methane offers cost-efficiency and environmental benefits for enhances oil recovery. High minimum miscibility pressure (MMP) in methane-oil systems poses a challenge. To overcome this, researchers are increasingly focusing on using surfactants to reduce MMP, thus enhancing the effectiveness of methane injections for oil recovery. This study investigated the impact of pressure and temperature on the equilibrium interfacial tension of the CH4+n-decane system using molecular dynamics simulations and the vanishing interfacial tension technique. The primary goal was to assess the potential of surfactants in lowering MMP. Among four tested surfactants, ME-6 exhibited the most promise by reducing MMP by 14.10% at 373 K. Key findings include that the addition of ME-6 enriching CH4 at the interface, enhancing its solubility in n-decane, improving n-decane diffusion capacity, CH4 weakens n-decane interactions and strengthens its own interaction with n-decane. As the difference in interactions of n-decane with ME-6's ends decreases, the system trends towards a mixed phase. This research sets the stage for broader applications of mixed-phase methane injection in reservoirs, with the potential for reduced gas flaring and environmental benefits.
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In the process of developing tight oil and gas reservoirs, multistage fractured horizontal wells (NFHWs) can greatly increase the production rate, and the optimal design of its fracturing parameters is also an important means to further increase the production rate. Accurate production prediction is essential for the formulation of effective development strategies and development plans before and during project execution. In this study, a novel workflow incorporating machine learning (ML) and particle swarm optimization algorithms (PSO) is proposed to predict the production rate of multi-stage fractured horizontal wells in tight reservoirs and optimize the fracturing parameters. The researchers conducted 10,000 numerical simulation experiments to build a complete training and validation dataset, based on which five machine learning production prediction models were developed. As input variables for yield prediction, eight key factors affecting yield were selected. The results of the study show that among the five models, the random forest (RF) model best establishes the mapping relationship between feature variables and yield. After verifying the validity of the Random Forest-based yield prediction model, the researchers combined it with the particle swarm optimization algorithm to determine the optimal combination of fracturing parameters under the condition of maximizing the net present value. A hybrid model, called ML-PSO, is proposed to overcome the limitations of current production forecasting studies, which are difficult to maximize economic returns and optimize the fracturing scheme based on operator preferences (e.g., target NPV). The designed workflow can not only accurately and efficiently predict the production of multi-stage fractured horizontal wells in real-time, but also be used as a parameter selection tool to optimize the fracture design. This study promotes data-driven decision-making for oil and gas development, and its tight reservoir production forecasts provide the basis for accurate forecasting models for the oil and gas industry.
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Rational design of coordination environment of Fe-based metal-organic frameworks (Fe-MOFs) is still a challenge in achieving enhanced catalytic activity for Fenten-like advanced oxidation process. Here in, novel porous Fe-MOFs with modulated O-Fe-N coordination was developed by configurating amino terephthalic acid (H2ATA) and pyrazine-dicarboxylic acid (PzDC) (Fe-ATA/PzDC-7:3). PzDC ligands introduce pyridine-N sites to form O-Fe-N coordination with lower binding energy, which affect the local electronic environment of Fe-clusters in Fe-ATA, thus decreased its interfacial H2O2 activation barrier. O-Fe-N coordination also accelerate Fe(II)/Fe(III) cycling of Fe-clusters by triggering the reactive oxidant species mediated Fe(III) reduction. As such, Fe-ATA/PzDC-7:3/H2O2 system exhibited excellent degradation performance for typical antibiotic sulfamethoxazole (SMX), in which the steady-state concentration of hydroxyl radical (OH) was 1.6 times higher than that of unregulated Fe-ATA. Overall, this study highlights the role of O-Fe-N coordination and the electronic environment of Fe-clusters on regulating Fenton-like catalytic performance, and provides a platform for precise engineering of Fe-MOFs.
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This work reports the construction of a miniaturized Ag/AgCl nanoelectrode on a nanopipette, which is capable of dual-functions of single-cell drug infusion and chloride detection and is envisioned to promote the study of chloride-correlated therapeutic effects.
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Cloretos , Compostos de Prata , PrataRESUMO
Developing nanoscale ratiometric techniques capable of biochemical response should prove of significance for precise applications with stringent spatial and biological restrictions. Here we present and devise the concept of θ-nanopore ratiometry, which uses ratiometric signals that could well address the serious concerns about device deviation in fabrication and nonspecific adsorption in the detection. As exemplified by a 200 nm θ-nanopore toward miRNA detection, the ±20 nm aperture drift could be mitigated and the issue of nonspecific adsorption could be minimized in the complex cytosolic environment. Practical application of this θ-nanopore ratiometry realizes the measurements of cytosolic miRNA-10b. This work has not only established a nanoscopic ratiometric technique but also enriched the extant armory of nanotools for single-cell studies and beyond.
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Técnicas Biossensoriais , MicroRNAs , NanoporosRESUMO
Data distribution gaps often pose significant challenges to the use of deep segmentation models. However, retraining models for each distribution is expensive and time-consuming. In clinical contexts, device-embedded algorithms and networks, typically unretrainable and unaccessable post-manufacture, exacerbate this issue. Generative translation methods offer a solution to mitigate the gap by transferring data across domains. However, existing methods mainly focus on intensity distributions while ignoring the gaps due to structure disparities. In this paper, we formulate a new image-to-image translation task to reduce structural gaps. We propose a simple, yet powerful Structure-Unbiased Adversarial (SUA) network which accounts for both intensity and structural differences between the training and test sets for segmentation. It consists of a spatial transformation block followed by an intensity distribution rendering module. The spatial transformation block is proposed to reduce the structural gaps between the two images. The intensity distribution rendering module then renders the deformed structure to an image with the target intensity distribution. Experimental results show that the proposed SUA method has the capability to transfer both intensity distribution and structural content between multiple pairs of datasets and is superior to prior arts in closing the gaps for improving segmentation.
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Iodinated X-ray contrast media (ICM) are ubiquitously present in water sources and challenging to eliminate using conventional processes, posing a significant risk to aquatic ecosystems. Ultraviolet light-emitting diodes (UV-LED) emerge as a promising technology for transforming micropollutants in water, boasting advantages such as diverse wavelengths, elimination of chemical additives, and no induction of microorganisms' resistance to disinfectants. The research reveals that iohexol (IOX) degradation escalates as UV wavelength decreases, attributed to enhanced photon utilization efficiency. Pseudo-first-order rate constants (kobs) were determined as 3.70, 2.60, 1.31 and 0.65 cm2 J-1 at UV-LED wavelengths of 255, 265, 275 and 285 nm, respectively. The optical properties of dissolved organic matter (DOM) and anions undeniably influence the UV-LED photolysis process through photon competition and the generation of reactive substances. The influence of Cl- on IOX degradation was insignificant at UV-LED 255, but it promoted IOX degradation at 265, 275 and 285 nm. IOX degradation was accelerated by ClO2-, NO3-and HA due to the formation of various reactive species. In the presence of NO3-, the kobs of IOX followed the order: 265 > 255 > 275 > 285 nm. Photosensitizers altered the spectral dependence of IOX, and the intermediate photoactivity products were detected using electron spin resonance. The transformation pathways of IOX were determined through density functional theory calculations and experiments. Disinfection by-products (DBPs) yields of IOX during UV-LED irradiation decreased as the wavelength increased: 255 > 265 > 275 > 285 nm. The cytotoxicity index value decreased as the UV-LED wavelength increased from 255 to 285 nm. These findings are crucial for selecting the most efficient wavelength for UV-LED degradation of ICM and will benefit future water purification design.
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Clarifying the structure-reactivity relationship of non-noble-metal electrocatalysts is one of the decisive factors for the practical application of water electrolysis. In this field, the anodic oxygen evolution reaction (OER) with a sluggish kinetic process has become a huge challenge for large-scale production of high-purity hydrogen. Here we synthesize a layered quasi-nevskite metastable-phase cobalt oxide (LQNMP-Co2O3) nanosheet via a simple molten alkali synthesis strategy. The unit-cell parameters of LQNMP-Co2O3 are determined to be a = b = 2.81 Å and c = 6.89 Å with a space group of P3Ì m1 (No. 164). The electrochemical results show that the LQNMP-Co2O3 electrocatalyst enables delivering an ultralow overpotential of 266 mV at a current density of 10 mA cmgeo-2 with excellent durability. The operando XANES and EXAFS analyses clearly reveal the origin of the OER activity and the electrochemical stability of the LQNMP-Co2O3 electrocatalyst. Density functional theory (DFT) simulations show that the energy barrier of the rate-determining step (RDS) (from *O to *OOH) is significantly reduced on the LQNMP-Co2O3 electrocatalyst by comparing with simulated monolayered CoO2 (M-CoO2).