Label-Free Direct Identification of MicroRNAs Based on a Narrow Constant-Inner-Diameter Emitter Mass Spectrometry Analysis.

MicroRNAs (miRNAs) are a class of endogenous noncoding small RNAs that play important roles in various biological processes and diseases. Direct determination of miRNAs is a cost-efficient and accurate method for analysis. Herein, we established a novel method for the analysis of miRNAs based on a narrow constant-inner-diameter mass spectrometry emitter. We utilized the gravity-assisted sleeving etching method to prepare a constant-inner-diameter mass spectrometry emitter with a capillary inner diameter of 5.5 µm, coupled it with a high-voltage power supply and a high-resolution mass spectrometer, and used it for miRNA direct detection. The method showed high sensitivity and reproducibility for the analysis of four miRNAs, with a limit of detection of 100 nmol/L (170 amol) for the Hsa-miR-1290 analysis. Compared with commercial ion sources, our method achieved higher sensitivity for miRNA detection. In addition, we analyzed the total miRNAs in the A549 cells. The result indicated that both spiked and endogenous miRNAs could be quantified with high accuracy. As a result, this method offers a promising platform for highly sensitive and accurate miRNA analysis. Furthermore, this approach can be extended to the analysis of other small oligonucleotides and holds the potential for studying clinical samples and facilitating disease diagnosis.

Monolithic 3D nanoelectrospray emitters based on a continuous fluid-assisted etching strategy for glass droplet microfluidic chip-mass spectrometry.

Glass microfluidic chips are suitable for coupling with mass spectrometry (MS) due to their flexible design, optical transparency and resistance to organic reagents. However, due to the high hardness and brittleness of glass, there is a lack of simple and feasible technology to manufacture a monolithic nanospray ionization (nESI) emitter on a glass microchip, which hinders its coupling with mass spectrometry. Here, a continuous fluid-assisted etching strategy is proposed to fabricate monolithic three-dimensional (3D) nESI emitters integrated into glass microchips. A continuous fluid of methanol is adopted to protect the inner wall of the channels and the bonding interface of the glass microfluidic chip from being wet-etched, forming sharp 3D nESI emitters. The fabricated 3D nESI emitter can form a stable electrospray plume, resulting in consistent nESI detection of acetylcholine with an RSD of 4.5% within 10 min. The fabricated 3D emitter is integrated on a glass microfluidic chip designed with a T-junction droplet generator, which can realize efficient analysis of acetylcholine in picoliter-volume droplets by nESI-MS. Stability testing of over 20 000 droplets detected by the established system resulted in an RSD of 9.1% over approximately 180 min. The detection of ten neurochemicals in rat cerebrospinal fluid droplets is achieved. The established glass droplet microfluidic chip-MS system exhibits potential for broad applications such as in vivo neurochemical monitoring and single-cell analysis in the future.

Ultra-Large Two-Dimensional Metal Nanowire Networks by Microfluidic Laminar Flow Synthesis for Formic Acid Electrooxidation.

Despite the great research interest in two-dimensional metal nanowire networks (2D MNWNs) due to their large specific surface area and abundance of unsaturated coordination atoms, their controllable synthesis still remains a significant challenge. Herein, a microfluidics laminar flow-based approach is developed, enabling the facile preparation of large-scale 2D structures with diverse alloy compositions, such as PtBi, AuBi, PdBi, PtPdBi, and PtAuCu alloys. Remarkably, these 2D MNWNs can reach sizes up to submillimeter scale (~220 µm), which is significantly larger than the evolution from the 1D or 3D counterparts that typically measure only tens of nanometers. The PdBi 2D MNWNs affords the highest specific activity for formic acid (2669.1 mA mg-1) among current unsupported catalysts, which is 103.5 times higher than Pt-black, respectively. Furthermore, in situ Fourier transform infrared (FTIR) experiments provide comprehensive evidence that PdBi 2D MNWNs catalysts can effectively prevent CO* poisoning, resulting in exceptional activity and stability for the oxidation of formic acid.

Enhanced recognition of G-quadruplex DNA oxidative damage based on DNA-mediated charge transfer.

G-quadruplex (G4) DNA is present in human telomere oligonucleotide sequences. Oxidative damage to telomeric DNA accelerates telomere shortening, which is strongly associated with aging and cancer. Most of the current analyses on oxidative DNA damage are based on ds-DNA. Here, we developed a electrochemiluminescence (ECL) probe for enhanced recognition of oxidative damage in G4-DNA based on DNA-mediated charge transfer (CT), which could specifically recognize damaged sites depending on the position of 8-oxoguanine (8-oxoG). First, a uniform G4-DNA monolayer interface was fabricated; the G4-DNA mediated CT properties were examined using an iridium(III) complex [Ir(ppy)2(pip)]PF6 stacked with G4-DNA as an indicator. The results showed that G4-DNA with 8-oxoG attenuated DNA CT. The topological effects of oxidative damage at different sites of G4-DNA and their effects on DNA CT were revealed. The sensing platform was also used for the sensitive and quantitative detection of 8-oxoG in G4-DNA, with a detection limit of 28.9 fmol. Overall, these findings present a sensitive platform to study G4-DNA structural and stability changes caused by oxidative damage as well as the specific and quantitative detection of oxidation sites. The different damage sites in the G-quadruplex could provide detailed clues for understanding the function of G4-associated telomere functional enzymes.

Kinetically Controlled Nucleation Enabled by Tunable Microfluidic Mixing for the Synthesis of Dendritic Au@Pt Core/Shell Nanomaterials.

The nucleation stage plays a decisive role in determining nanocrystal morphology and properties; hence, the ability to regulate nucleation is critical for achieving high-level control. Herein, glass microfluidic chips with S-shaped mixing units are designed for the synthesis of Au@Pt core/shell materials. The use of hydrodynamics to tune the nucleation kinetics is explored by varying the number of mixing units. Dendritic Au@Pt core/shell nanomaterials are controllably synthesized and a formation mechanism is proposed. As-synthesized Au@Pt exhibited excellent ethanol oxidation activity under alkaline conditions (8.4 times that of commercial Pt/C). This approach is also successfully applied to the synthesize of Au@Pd core/shell nanomaterials, thus demonstrating its generality.

Formation of Protein Nanoparticles in Microdroplet Flow Reactors.

Nanoparticles are increasingly being used for biological applications, such as drug delivery and gene transfection. Different biological and bioinspired building blocks have been used for generating such particles, including lipids and synthetic polymers. Proteins are an attractive class of material for such applications due to their excellent biocompatibility, low immunogenicity, and self-assembly characteristics. Stable, controllable, and homogeneous formation of protein nanoparticles, which is key to successfully delivering cargo intracellularly, has been challenging to achieve using conventional methods. In order to address this issue, we employed droplet microfluidics and utilized the characteristic of rapid and continuous mixing within microdroplets in order to produce highly monodisperse protein nanoparticles. We exploit the naturally occurring vortex flows within microdroplets to prevent nanoparticle aggregation following nucleation, resulting in systematic control over the particle size and monodispersity. Through combination of simulation and experiment, we find that the internal vortex velocity within microdroplets determines the uniformity of the protein nanoparticles, and by varying parameters such as protein concentration and flow rates, we are able to finely tune nanoparticle dimensional properties. Finally, we show that our nanoparticles are highly biocompatible with HEK-293 cells, and through confocal microscopy, we determine that the nanoparticles fully enter into the cell with almost all cells containing them. Due to the high throughput of the method of production and the level of control afforded, we believe that the approach described in this study for generating monodisperse protein-based nanoparticles has the potential for intracellular drug delivery or for gene transfection in the future.

Sensitive electrochemical measurement of nitric oxide released from living cells based on dealloyed PtBi alloy nanoparticles.

Nitric oxide (NO), as a vital signaling molecule related to different physiological and pathological processes in living systems, is closely associated with cancer and cardiovascular disease. However, the detection of NO in real-time remains a difficulty. Here, PtBi alloy nanoparticles (NPs) were synthesized, dealloyed, and then fabricated to NP-based electrodes for the electrochemical detection of NO. Transmission electron microscopy (TEM), small-angle X-ray scattering (SAXS), and nitrogen physical adsorption/desorption show that dealloyed PtBi alloy nanoparticles (dPtBi NPs) have a porous nanostructure. Electrochemical impedance spectroscopy and cyclic voltammetry results exhibit that the dPtBi NP electrode possesses unique electrocatalytic features such as low charge transfer resistance and large electrochemically active surface area, which lead to its excellent NO electrochemical sensing performance. Owing to the higher density of catalytical active sites formed PtBi bimetallic interface, the dPtBi NP electrode displays superior electrocatalytic activity toward the oxidation of NO with a peak potential at 0.74 V vs. SCE. The dPtBi NP electrode shows a wide dynamic range (0.09-31.5 µM) and a low detection limit of 1 nM (3σ/k) as well as high sensitivity (130 and 36.5 µA µM-1 cm-2). Moreover, the developed dPtBi NP-based electrochemical sensor also exhibited good reproducibility (RSD 5.7%) and repeatability (RSD 3.4%). The electrochemical sensor was successfully used for the sensitive detection of NO produced by live cells. This study indicates a highly effective approach for regulating the composition and nanostructures of metal alloy nanomaterials, which might provide new technical insights for developing high-performance NO-sensitive systems, and have important implications in enabling real-time detection of NO produced by live cells.

Quantitative Analysis of Nanoplastics in Single Cells by Subcellular Chromatography.

The accumulation and spatial distribution of intracellular nanoplastic particles provide useful information about their spatiotemporal toxicological effects mediated by the physicochemical parameters of nanoplastics in living cells. In this study, a sample injection-transfer method was designed with an accuracy of up to femtoliters to attoliters to match the volume required for ultranarrow-bore open-tubular liquid chromatography. The separation and concentration quantification of mixed polystyrenes in different regions in living cells were achieved by directly transferring picoliter/femtoliter volumes of intracellular cytoplasm to an ultranarrow-bore open-tubular chromatographic column. The measurement of pollutant concentration in different areas of a small-volume target (single cell) was realized. This method is expected to be used in the qualitative and quantitative analyses of complex, mixed, and label-free nanoplastics (a few nm in size) in the subregions of living cells.

Dealloying of Pt1Bi2 intermetallic toward optimization of electrocatalysis on a Bi-continuous nanoporous core-shell structure.

Noble metal nanoporous materials hold great potential in the field of catalysis, owing to their high open structures and numerous low coordination surface sites. However, the formation of porous nanoparticles is restricted by particle size. Herein, we utilized a Pt1Bi2 intermetallic nanocatalyst to develop a dealloying approach for preparing nanoparticles with a bi-continuous porous and core-shell structure and proposed a mechanism for the formation of pores. The particle size used to form the porous structure can be <10 nm, which enhances the nanocatalyst's performance for the oxygen reduction reaction (ORR). This study provides a new understanding of the formation of porous materials via a dealloying approach.

Highly Selective Activation of C-H Bond and Inhibition of C-C Bond Cleavage by Tuning Strong Oxidative Pd Sites.

Improving the product selectivity meanwhile restraining deep oxidation still remains a great challenge over the supported Pd-based catalysts. Herein, we demonstrate a universal strategy where the surface strong oxidative Pd sites are partially covered by the transition metal (e. g., Cu, Co, Ni, and Mn) oxide through thermal treatment of alloys. It could effectively inhibit the deep oxidation of isopropanol and achieve the ultrahigh selectivity (>98%) to the target product acetone in a wide temperature range of 50-200 °C, even at 150-200 °C with almost 100% isopropanol conversion over PdCu1.2/Al2O3, while an obvious decline in acetone selectivity is observed from 150 °C over Pd/Al2O3. Furthermore, it greatly improves the low-temperature catalytic activity (acetone formation rate at 110 °C over PdCu1.2/Al2O3, 34.1 times higher than that over Pd/Al2O3). The decrease of surface Pd site exposure weakens the cleavage for the C-C bond, while the introduction of proper CuO shifts the d-band center (εd) of Pd upward and strengthens the adsorption and activation of reactants, providing more reactive oxygen species, especially the key super oxygen species (O2-) for selective oxidation, and significantly reducing the barrier of O-H and ß-C-H bond scission. The molecular-level understanding of the C-H and C-C bond scission mechanism will guide the regulation of strong oxidative noble metal sites with relatively inert metal oxide for the other selective catalytic oxidation reactions.

Single-Cell Metabolomics-Based Strategy for Studying the Mechanisms of Drug Action.

Studying the mechanisms of drug antitumor activity at the single-cell level can provide information about the responses of cell subpopulations to drug therapy, which is essential for the accurate treatment of cancer. Due to the small size of single cells and the low contents of metabolites, metabolomics-based approaches to studying the mechanisms of drug action at the single-cell level are lacking. Herein, we develop a label-free platform for studying the mechanisms of drug action based on single-cell metabolomics (sMDA-scM) by integrating intact living-cell electro-launching ionization mass spectrometry (ILCEI-MS) with metabolomics analysis. Using this platform, we reveal that non-small-cell lung cancer (NSCLC) cells treated by gefitinib can be clustered into two cell subpopulations with different metabolic responses. The glutathione metabolic pathway of the subpopulation containing 14.4% of the cells is not significantly affected by gefitinib, exhibiting certain resistance characteristics. The presence of these cells masked the judgment of whether cysteine and methionine metabolic pathway was remarkably influenced in the analysis of overall average results, revealing the heterogeneity of the response of single NSCLC cells to gefitinib treatment. The findings provide a basis for evaluating the early therapeutic effects of clinical medicines and insights for overcoming drug resistance in NSCLC subpopulations.

Spontaneous Phase Segregation Enabling Clogging Aversion in Continuous Flow Microfluidic Synthesis of Nanocrystals Supported on Reduced Graphene Oxide.

Eliminating clogging in capillary tube reactors is critical but challenging for enabling continuous-flow microfluidic synthesis of nanoparticles. Creating immiscible segments in a microfluidic flow is a promising approach to maintaining a continuous flow in the microfluidic channel because the segments with low surface energy do not adsorb onto the internal wall of the microchannel. Herein we report the spontaneous self-agglomeration of reduced graphene oxide (rGO) nanosheets in polyol flow, which arises because the reduction of graphene oxide (GO) nanosheets by hot polyol changes the nanosheets from hydrophilic to hydrophobic. The agglomerated rGO nanosheets form immiscible solid segments in the polyol flow, realizing the liquid-solid segmented flow to enable clogging aversion in continuous-flow microfluidic synthesis. Simultaneous reduction of precursor species in hot polyol deposits nanocrystals uniformly dispersed on the rGO nanosheets even without surfactant. Cuprous oxide (Cu2O) nanocubes of varying edge lengths and ultrafine metal nanoparticles of platinum (Pt) and palladium (Pd) dispersed on rGO nanosheets have been continuously synthesized using the liquid-solid segmented flow microfluidic method, shedding light on the promise of microfluidic reactors in synthesizing functional nanomaterials.

NaCl Micro-Crystal as a Molecular Mold for Enhanced Synthesis of Planar Phenazines and Their Applications on Chemosensing and a Full-Color Fluorescent Material.

NaCl has been successfully used as a template for the synthesis of 2D nanomaterials, but it is seldom used for the construction of flat small organic molecules. Herein, a simple, low-cost, and highly efficient synthesis of phenazines with planar main frames, such as 5-phenyl-5,14-dihydro-5,7,12,14-tetraazapentacene, in the presence of NaCl micro-crystal as a kind of molecular mold is described. The reactants were mixed with NaCl powder and heated to 320 °C for 5 min. Yields >90% were readily achieved after a simple precipitation in water. The effectiveness of NaCl crystal as a mold with HCl was confirmed by comparison with common inorganic salts, SiO2, and γ-Al2O3 with HCl together with combinations including NaNO3 + HNO3, Na2SO4 + H2SO4, NaH2PO4 + H3PO4, and NaH2PO4 + polyphosphoric acid. The mechanism was deduced with the aid of computer simulation, which confirms the stabilization of 5,14-dihydro-5,7,12,14-tetraazapentacene by the NaCl surface. DMSO solution of a product, 1,3-dihydro-imidazo[4,5-b]phenazin-2-one, showed enhanced fluorescence in H2O, and it was used as a fluorescent probe for pH and Hg2+. A full-color material was prepared by mixing precursors of epoxy resin and phenazines, and its fluorescent color could be adjusted by the ratio of phenazines.

Intact living-cell electrolaunching ionization mass spectrometry for single-cell metabolomics.

While single-cell mass spectrometry can reveal cellular heterogeneity and the molecular mechanisms of intracellular biochemical reactions, its application is limited by the insufficient detection sensitivity resulting from matrix interference and sample dilution. Herein, we propose an intact living-cell electrolaunching ionization mass spectrometry (ILCEI-MS) method. A capillary emitter with a narrow-bore, constant-inner-diameter ensures that the entire living cell enters the MS ion-transfer tube. Inlet ionization improves sample utilization, and no solvent is required, preventing sample dilution and matrix interference. Based on these features, the detection sensitivity is greatly improved, and the average signal-to-noise (S/N) ratio is about 20 : 1 of single-cell peaks in the TIC of ILCEI-MS. A high detection throughput of 51 cells per min was achieved by ILCEI-MS for the single-cell metabolic profiling of multiple cell lines, and 368 cellular metabolites were identified. Further, more than 4000 primary single cells digested from the fresh multi-organ tissues of mice were detected by ILCEI-MS, demonstrating its applicability and reliability.

Analytical methods for obtaining binding parameters of drug-protein interactions: A review.

The study of drug-protein interactions can reveal the corresponding binding mechanisms, providing valuable information for the early phase drug development and development of new drugs. This article reviews the methods used for obtaining the binding parameters of drug-protein systems. The methods include equilibrium dialysis, high-performance affinity chromatography, capillary electrophoresis, spectroscopy, calorimetry, competition and displacement, mass spectrometry, fluorescence resonance energy transfer, and thermal stability shift analysis. Relevant parameters include the association constant, number of binding sites, thermodynamic properties, binding force types, binding site types, binding distances, changes in protein conformation, and changes in protein stability. In addition, the review also summarizes the principles, advantages, and limitations of each method in detail. The comparison of parameter information can not only guide method selection but also provide valuable reference information for in-depth exploration of drug-protein interaction mechanisms.

Displacement Reaction-Assisted Synthesis of Sub-Nanometer Pt/Bi Boost Methanol-Tolerant Fuel Cells.

The development of new synthetic methods for methanol-tolerant catalysts with improved performance is of fundamental importance for the commercialization of fuel cells. Herein, we reported a facile displacement reaction-assisted synthesis of graphene-supported sub-nanometer Pt/Bi catalysts (Pt/Bi/rGO). Bismuth (0) nanoparticles produced by NH3BH3 reduction can be further dissolved into the ethylene glycol, implying Bi(0) has a strong interaction with the hydroxyl group. That is the key interaction between Bi(0) and the functional group on the rGO to form the ultra-small Bi/rGO catalyst. Furthermore, Pt clusters are obtained by the displacement between Bi(0) and HPtCl4 and are directly anchored to the rGO surface. The as-synthesized Pt/Bi/rGO catalyst exhibits high oxygen reduction mass activity and high tolerance to methanol poisoning. In the presence of 0.5 mol/L CH3OH, the initial potential and activity of ORR were almost unchanged, which demonstrated great potential in the application of direct methanol fuel cells.

Synergy in Au-CuO Janus Structure for Catalytic Isopropanol Oxidative Dehydrogenation to Acetone.

The controlled oxidation of alcohols to the corresponding ketones or aldehydes via selective cleavage of the ß-C-H bond of alcohols under mild conditions still remains a significant challenge. Although the metal/oxide interface is highly active and selective, the interfacial sites fall far behind the demand, due to the large and thick support. Herein, we successfully develop a unique Au-CuO Janus structure (average particle size=3.8 nm) with an ultrathin CuO layer (0.5 nm thickness) via a bimetal in situ activation and separation strategy. The resulting Au-CuO interfacial sites prominently enhance isopropanol adsorption and decrease the energy barrier of ß-C-H bond scission from 1.44 to 0.01 eV due to the strong affinity between the O atom of CuO and the H atom of isopropanol, compared with Au sites alone, thereby achieving ultrahigh acetone selectivity (99.3 %) over 1.1 wt % AuCu0.75 /Al2 O3 at 100 °C and atmospheric pressure with 97.5 % isopropanol conversion. Furthermore, Au-CuO Janus structures supported on SiO2 , TiO2 or CeO2 exhibit remarkable catalytic performance, and great promotion in activity and acetone selectivity is achieved as well for other reducible oxides derived from Fe, Co, Ni and Mn. This study should help to develop strategies for maximized interfacial site construction and structure optimization for efficient ß-C-H bond activation.

Evaluation of the effect of dexmedetomidine combined with ropivacaine in epidural labor analgesia: a systematic review and meta-analysis.

INTRODUCTION: This review and meta-analysis comprehensively elaborated the analgesic and sedative effects of ropivacaine combined with dexmedetomidine in epidural labor analgesia, and its possible resulting motor block and adverse reactions. EVIDENCE ACQUISITION: PubMed, Web of science, Cochrane, China National Knowledge Infrastructure and WANFANG DATA were searched for randomized controlled trials (RCTs) on the efficacy and safety of ropivacaine combined with dexmedetomidine in epidural labor analgesia. Standard mean difference (SMD) and 95% confidence interval (CI) were calculated. EVIDENCE SYNTHESIS: As of September 10th, 17 articles were finally included for meta-analysis. The results showed that compared with the control group (ropivacaine alone or ropivacaine + sufentanil), singleton full-term pregnant patients treated with ropivacaine + dexmedetomidine had better analgesic and sedative effects, with lower VAS scores at 15 min (T1), 30 min (T2), 1 h (T3), 2 h (T4) after labor analgesia, and the moment of delivery (T5) (T1: SMD=-1.90, 95% CI: -2.39, -1.40; T2: SMD=-1.43, 95% CI: -1.92, -0.93; T3: SMD=-0.91, 95% CI: -1.41, -0.41; T4: SMD=-0.94, 95% CI: -1.54, -0.34; T5: SMD=-0.56, 95% CI: -1.02, -0.10) and higher Ramsay scores at 15 min (T1) and 30 min (T2) after labor analgesia (T1: SMD=1.17, 95% CI: 0.80, 1.53; T2: SMD=2.17, 95% CI: 1.30, 3.03). CONCLUSIONS: In patients with singleton full-term pregnancy, ropivacaine + dexmedetomidine have better analgesic and sedative effects than in the control group. Both groups have no significant motor block and neonatal asphyxia and hypoxia.

Controllable Fabrication of Small-Size Holding Pipets for the Nondestructive Manipulation of Suspended Living Single Cells.

The capture and manipulation of single cells are an important premise and basis for intracellular delivery, which provides abundant molecular and omics information for biomedical development. However, for intracellular delivery of cargos into/from small-size suspended living single cells, the capture methods are limited by the lack of small-size holding pipets, poor cell activity, and the low spatial accuracy of intracellular delivery. To solve these problems, a method for the controllable fabrication of small-size holding pipets was proposed. A simple, homemade microforge instrument including an imaging device was built to cut and melt the glass capillary tip by controlling the heat production of a nichrome wire. The controllable fabrication of small-size holding pipets was realized by observing the fabrication process in real time. Combined with an electroosmotic drive system and a micromanipulation system with high spatial resolution, the holding pipet achieved the active capture, movement, and sampling of suspended living single cells. Moreover, solid-phase microextraction was performed on captured single pheochromocytoma cells, and the extracted dopamine was successfully detected using an electrochemical method. The homemade microforge instrument overcame the limitations of traditional microforges, resulting in holding pipets that were sufficiently small for small-size suspended single living cells (5-30 µm). This proactive capture method overcame the shortcomings of existing methods to achieve the multiangle, high-precision manipulation of single cells, thereby allowing the intracellular delivery of small-size single cells in suspension with high spatiotemporal resolution.

Investigation of metformin hydrochloride-bovine serum albumin interaction by narrow-bore capillary zone electrophoresis.

A narrow-bore capillary (inner diameter = 2 µm) zone electrophoresis method was developed to study the interaction between metformin hydrochloride and bovine serum albumin. Free metformin hydrochloride, free bovine serum albumin, and their complex were completely separated in a volume of tens of picoliters. The association constant (Ka = 1.04 × 103 L mol-1) and the number of binding sites (n = 0.9789) were then obtained.