Wearable bioelectronics are gaining extraordinary attention due to their capabilities to achieve continuous monitoring of human health status. However, mainstream manufacturing technologies, including photolithography and printing technology, limit current wearable bioelectronics on 2D planar structures with little surface area in contact with the body. It thus limits the amount of physiological information that current wearable bioelectronics could obtain. Furthermore, they need to be firmly attached to the body, affecting the wearing comfort. In this study, we leveraged the versatile thermal drawing process and developed a flexible microelectronic fiber with bioanalytical functions that could be woven into textiles as a new form of wearable bioelectronics. Within a single strand of fiber, we successfully integrated all-in-one multiplexed electrochemical sensing capabilities, with the sweat as the primary object. Adopting the laser micromachining technique, we developed biosensing functions on the longitudinal surface of the fiber with two sensing electrodes for Na+ and uric acid (UA), respectively, together with a pseudo reference electrode (p-RE). We carefully characterized the all-in-one multiplexed sensing performance of the fiber and demonstrated its successful application in sweat sensing based on its textile forms. The results show significant potential for application in wearable textiles for monitoring key health signals of humans.

Proteins have been considered key building blocks of life. In particular, the protein content of an organism and a cell offers significant information for the in-depth understanding of the disease and biological processes. Single-molecule protein detection/sequencing tools will revolutionize clinical (proteomics) research, offering ultrasensitivity for low-abundance biomarker (protein) detection, which is important for the realization of early-stage disease diagnosis and single-cell proteomics. This improved detection/measurement capability delivers new sets of techniques to explore new frontiers and address important challenges in various interdisciplinary areas including nanostructured materials, molecular medicine, molecular biology, and chemistry. Importantly, fluorescence-based methods have emerged as indispensable tools for single protein detection/sequencing studies, providing a higher signal-to-noise ratio (SNR). Improvements in fluorescent dyes/probes and detector capabilities coupled with advanced (image) analysis strategies have fueled current developments for single protein biomarker detections. For example, in comparison to conventional ELISA (i.e., based on ensembled measurements), single-molecule fluorescence detection is more sensitive, faster, and more accurate with reduced background, high-throughput, and so on. In comparison to MS sequencing, fluorescence-based single-molecule protein sequencing can achieve the sequencing of peptides themselves with higher sensitivity. This review summarizes various typical single-molecule detection technologies including their methodology (modes of operation), detection limits, advantages and drawbacks, and current challenges with recent examples. We describe the fluorescence-based single-molecule protein sequencing/detection based on five kinds of technologies such as fluorosequencing, N-terminal amino acid binder, nanopore light sensing, and DNA nanotechnology. Finally, we present our perspective for developing high-performance fluorescence-based sequencing/detection techniques.

Primary amines are the target of many bioanalytical analyses, as they are ubiquitous in biological systems and responsible for numerous important processes including neurotransmission and cell signaling. Primary amines can be sensitively detected via fluorescence after their reaction with the fluorogenic reagent naphthalene-2,3-dicarboxaldehyde (NDA) in the presence of cyanide through the formation of fluorescent N-substituted 1-cyanobenz[f]isoindole (CBI) derivatives. While fluorogenic reagents such as NDA can be advantageous for sensitive detection, improvements in both long-term stability and speed of reaction are necessary to enable practical and reproducible quantitative analysis. In this work, various CBI-amines were interrogated for their fluorescence characteristics over time under previously reported conditions (75:25 aqueous buffer:acetonitrile). An amine-specific decline in fluorescence and delay to reach maximum fluorescence were observed. Based on structural characteristics, we hypothesized that these effects were due to the solvents employed enabling analyte intermolecular interactions that resulted in fluorescence quenching over time. To mitigate fluorescence-quenching intermolecular interactions, we developed two strategies to improve the fluorescence of the CBI-product over long time periods: (1) the addition of the complexation reagent β-cyclodextrin to the reaction matrix and (2) the substitution of acetonitrile with dimethyl sulfoxide. Both strategies improved fluorescence stability over time, and the incorporation of dimethyl sulfoxide also enabled more rapid attainment of maximum fluorescence and a higher absolute fluorescence when compared to initial conditions. When employed in combination, these two approaches further improve fluorescence stability over time for the most hydrophobic analytes. In the future, these strategies can be employed for the practical and reproducible quantitative analysis of primary amines in biological systems, including those related to neurological disorders and disease states.

As two main types of liquid biopsy markers, both circulating tumor cells (CTCs) and small extracellular vesicles (sEVs) play important roles in the diagnosis and prognosis of cancers. CTCs are malignant cells that detach from the original tumor tissue and enter the circulation of body fluids. sEVs are nanoscale vesicles secreted by normal cells or pathological cells. However, CTCs and sEVs in body fluids are scarce, leading to great difficulties in the accurate analysis of related diseases. For the sensitive detection of CTCs and sEVs in body fluids, various types of nucleic acid and nanomaterial-assisted signal amplification strategies have been developed. In this review, we summarize the recent advances in fluorescent detection of CTCs and sEVs in liquid biopsy based on nucleic acid and nanomaterial-assisted signal amplification strategies. We also discuss their advantages, challenges, and future prospects.

Electrical asymmetric-flow field-flow fractionation (EAF4) is a new and interesting analytical technique recently proposed for the characterization of metallic nanoparticles (NPs). It has the potential to simultaneously provide relevant information about size and electrical parameters, such as electrophoretic mobility (μ) and zeta-potential (ζ), of individual NP populations in an online instrumental setup with an array of detectors. However, several chemical and instrumental conditions involved in this technique are definitely influential, and only few applications have been proposed until now. In the present work, an EAF4 system has been used with different detectors, ultraviolet-visible (UV-vis), multi-angle light scattering (MALS), and inductively coupled plasma with triple quadrupole mass spectrometry (ICP-TQ-MS) for the characterization of gold, silver, and platinum NPs with both citrate and phosphate coatings. The behavior of NPs has been studied in terms of retention time and signal intensity under both positive and negative current with results depending on the coating. Carrier composition, particularly ionic strength, was found to be critical to achieve satisfactory recoveries and a reliable measurement of electrical parameters. Dynamic light scattering (DLS) has been used as a comparative technique for these parameters. The NovaChem surfactant mix (0.01%) showed a quantitative recovery (93 ± 1%) of the membrane, but the carrier had to be modified by increasing the ionic strength with 200 μM of Na2CO3 to achieve consistent μ values. However, ζ was one order of magnitude lower in EAF4-UV-vis-MALS than in DLS, probably due to different electric processes in the channel. From a practical point of view, EAF4 technique is still in its infancy and further studies are necessary for a robust implementation in the characterization of NPs.

The proposal of high-throughput platforms in microextraction-based approaches is important to offer sustainable and efficient tools in analytical chemistry. Particularly, automated configurations exhibit enormous potential because they provide accurate and precise results in addition to less analyst intervention. Recently, significant achievements have been obtained in proposing affordable platforms for microextraction techniques capable of being integrated with different analytical instrumentations. Considering the evolution of these approaches, this article describes innovative high-throughput platforms that have recently been proposed for the analysis of varied matrices, with special attention to laboratory-made devices. Additionally, some challenges, opportunities, and trends regarding these experimental workflows are pointed out.

The deployment of metal-organic frameworks (MOFs) in a plethora of analytical and bioanalytical applications is a growing research area. Their unique properties such as high but tunable porosity, well-defined channels or pores, and ease of post-synthetic modification to incorporate additional functional units make them ideal candidates for sensing applications. This is possible because the interaction of analytes with a MOF often results in a change in its structure, eventually leading to a modification of the intrinsic physicochemical properties of the MOF which is then transduced into a measurable signal. The high porosity allows for the adsorption of analytes very efficiently, while the tunable pore sizes/nature and/or installation of specific recognition groups allow modulating the affinity towards different classes of compounds, which in turn lead to good sensor sensitivity and selectivity, respectively. Some figures are given to illustrate the potential of MOF-based sensors in the most relevant application fields, and future challenges and opportunities to their possible translation from academia (i.e., laboratory testing of MOF sensing properties) to industry (i.e., real-world analytical sensor devices) are critically discussed.

Organic field-effect transistors (OFETs) have been proposed beyond three decades while becoming a research hotspot again in recent years because of the fast development of flexible electronics. Many novel flexible OFETs-based devices have been reported in these years. Among these devices, flexible OFETs-based sensors made great strides because of the extraordinary sensing capability of FET. Most of these flexible OFETs-based sensors were designed for biological applications due to the advantages of flexibility, reduced complexity, and lightweight. This paper reviews the materials, fabrications, and applications of flexible OFETs-based biosensors. Besides, the challenges and opportunities of the flexible OFETs-based biosensors are also discussed.

Erhuangquzhi granules (EQG) have been clinically proven to be effective in nonalcoholic steatohepatitis (NASH) treatment. However, the active components and molecular mechanisms remain unknown. This study aimed to screen active components targeting tumor necrosis factor α (TNF-α) in EQG for the treatment of NASH by a surface plasmon resonance (SPR) biosensor-based active ingredient recognition system (SPR-AIRS). The amine-coupling method was used to immobilize recombinant TNF-α protein on an SPR chip, the specificity of the TNF-α-immobilized chip was validated, and nine medicinal herbs in EQG were prescreened. Nuciferine (NF), lirinidine (ID), and O-nornuciferine (NNF) from lotus leaves were found and identified as TNF-α ligands by UPLC‒MS/MS, and the affinity constants of NF, ID, and NNF to TNF-α were determined by SPR experiments (Kd = 61.19, 31.02, and 20.71 µM, respectively). NF, ID, and NNF inhibited TNF-α-induced apoptosis in L929 cells, the levels of secreted IL-6 and IL-1β were reduced, and the phosphorylation of IKKβ and IκB was inhibited in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. In conclusion, a class of new active small-molecule TNF-α inhibitors was discovered, which also provides a valuable reference for the material basis and mechanism of EQG action in NASH treatment.

The Fenton reaction is one of the most effective methods for treating organic wastewater, which is extremely harmful to humans but difficult to treat. However, finding simple, low-cost, and efficient catalysts for the Fenton reaction remains a challenge. In this study, a BSA-Cu3(PO4)2 hybrid nanoflower (NF) was synthesized to investigate its peroxidase-like activity for the treatment of organic wastewater. Its morphology, composition, and crystallization had been fully studied and the results confirmed that the NFs were successfully prepared. Subsequently, the origin of the peroxidase-like activity of the NFs was further analyzed, with the results suggesting two reasons: (i) the transformation between Cu(I) and Cu(II) and (ii) nano-effects. Additionally, Congo red was selected as the organic pollutant to simulate the decolorization of wastewater. After 3 h, the decolorization efficiency reached 96%. Furthermore, the NFs exhibited good storage performance, maintaining approximately 90% relative activity after storage for 30 days. In summary, the NFs have great application prospects in the treatment of organic wastewater.

The blood levels of cancer antigen 125 (CA125) and human epididymal secretory protein 4 (HE4) are measured in the diagnosis and progression monitoring of ovarian cancer (OC), and the Risk of Ovarian Malignancy Algorithm (ROMA) score% values are calculated for cancer risk assessment. For the first time, disposable dual screen-printed carbon electrodes modified with reduced graphene oxide, polythionine, and gold nanoparticles were used to fabricate label-free electrochemical dual CA125-HE4 immunosensors for the sensitive, fast, and practical simultaneous determination of CA125 and HE4. DPV and SWV methods were used to simultaneously determine antigens in two different linear ranges (1-100 pg mL-1 and 1-50 ng mL-1). High sensitivity, low LOD, and LOQ were obtained for two linear ranges with a correlation coefficient above 0.99. The application stability of the dual CA125-HE4 immunosensors was determined as 60 days, and the storage stability was determined as 16 weeks. The dual immunosensors exhibited high selectivity in eight different antigen mixtures. The reusability of the dual immunosensors has been tested up to 9 cycles. ROMA score% values for pre-menopausal and post-menopausal status were calculated using the concentration of CA125 and HE4 in the blood serum and assessing OC risk. The disposable dual immunosensors can be used in point-of-care tests for rapid and practical simultaneous determination of CA125 and HE4 with high selectivity, sensitivity, and repeatability.

Emerging insights from metabolomic-based studies of major depression disorder (MDD) are mainly related to biochemical processes such as energy or oxidative stress, in addition to neurotransmission linked to specific metabolite intermediates. Hub metabolites represent nodes in the biochemical network playing a critical role in integrating the information flow in cells between metabolism and signaling pathways. Limited technical-scientific studies have been conducted to understand the effects of ayahuasca (Aya) administration in the metabolism considering MDD molecular context. Therefore, this work aims to investigate an in vitro primary astrocyte model by untargeted metabolomics of two cellular subfractions: secretome and intracellular content after pre-defined Aya treatments, based on DMT concentration. Mass spectrometry (MS)-based metabolomics data revealed significant hub metabolites, which were used to predict biochemical pathway alterations. Branched-chain amino acid (BCAA) metabolism, and vitamin B6 and B3 metabolism were associated to Aya treatment, as "housekeeping" pathways. Dopamine synthesis was overrepresented in the network results when considering the lowest tested DMT concentration (1 µmol L-1). Building reaction networks containing significant and differential metabolites, such as nicotinamide, L-DOPA, and L-leucine, is a useful approach to guide on dose decision and pathway selection in further analytical and molecular studies.

Herein, we subtly engineered an amplified colorimetric biosensor for the cyclic detection of African swine fever virus DNA (ASFV-DNA), which associated the branched catalytic hairpin assembly (bCHA) amplification with G-quadruplex DNAzyme activity through triplex DNA formation. Firstly, a Y-shaped hairpin trimer was constructed for the dynamic self-assembly of DNA dendrimers. Then, in the presence of ASFV-DNA, the signal strand CP was opened, exposing the toehold regions, which would trigger the CHA cascade reaction between hairpin trimers. In the CHA cascade reaction, H1, H2, and H3 opened and bound in sequence, eventually forming the structure of DNA dendrimers. Subsequently, the obtained bCHA product was specifically recognized by the GGG repeat sequences of L1 and L2, then amplified by the synergistic effect of triplex DNA and the formation of asymmetric split G-quadruplex. Benefiting from the amplification properties of bCHA and the high peroxidase-like catalytic activity of asymmetrically split G-quadruplex DNAzymes, it could achieve effective colorimetric signal output in the presence of ASFV-DNA by means of triplex DNA formation. Under the optimal experimental conditions, this biosensor exhibited excellent sensitivity with a detection limit of 1.8 pM. Further, it was applied to the content detection of simulated samples of African swine fever, and the recoveries were 98.9 ~ 103.2%. This method has the advantages of simple operation, good selectivity, and high sensitivity, which is expected to be used for highly sensitive detection of actual samples of African swine fever virus.

The etiology of neurodegenerative diseases is poorly understood; however, studies have shown that heavy metals, such as copper, play a critical role in neurotoxicity, thus, adversely affecting the development of these diseases. Because of the limitations associated with classical metal detection tools to obtain accurate speciation information of ultra-low concentrations of heavy metals in the brain, analysis is primarily performed in blood, urine, or postmortem tissues, limiting the translatability of acquired knowledge to living systems. Inadequate and less accurate data obtained with such techniques provide little or no information for developing efficient therapeutics that aid in slowing down the deterioration of brain cells. In this study, we developed a biocompatible, ultra-fast, low-cost, and robust surface-modified electrode with carbon fibers by electrodepositing dopamine via fast-scan cyclic voltammetry (FSCV) to detect Cu2+ in modified tris buffer. We studied the surface morphology of our newly introduced sensors using high-resolution images by atomic force microscopy under different deposition conditions. The limit of detection (LOD) of our surface-modified sensor was 0.01 µM (0.64 ppb), and the sensitivity was 11.28 nA/µM. The LOD and sensitivity are fifty and two times greater, respectively, compared to those of a bare electrode. The sensor's response is not affected by the presence of dopamine in the matrix. It also exhibited excellent stability to multiple subsequent injections and repeated measurements of Cu2+ over a month, thus showing its strength to be developed into an accurate, fast, robust electrochemical tool to monitor ultra-low concentrations of heavy metals in the brain in real time.

Emitter tip arrays for electrospray ionization have been used for a variety of MS sample introduction purposes, including detection of multiple sample eluent streams and improved accuracy through parallel infusion of an internal standard. User control is typically required for targeted application of high voltage to specific channels to maximize analyte signal and minimize other background signals. In this communication, an automated approach to applying electrospray voltage only when a detectable analyte is present is described. An in-line absorbance detector is used to identify the presence of an analyte in the fluidic path between the sample introduction valve and the mass spectrometer. A Raspberry Pi-controlled system is then used to apply high voltage to a downstream emitter tip at the MS inlet following a delay volume between the detectors. Demonstration of this technique on two parallel sample channels is reported, including a pulsed voltage application to maximize signal when analytes elute on each channel simultaneously.

Fluorescence imaging has been widely employed for biomedical research and clinical diagnostics. With ease of synthesis and excellent photophysical properties, D-A type fluorophores are widely designed for fluorescence imaging. However, traditional D-A type fluorophores are solvatochromic which reduces the fluorescence brightness in the biological system. To solve this problem and build on our previous work, we devised a novel HIEE fluorophore MTC with typical anti-solvatochromic fluorescence. Furthermore, the activated fluorescent probe designed based on MTC showed excellent imaging performance. We believe that the strategy based on the fluorophores with typical anti-solvatohromic fluorescence can be a useful platform for designing fluorescent probes for high-brightness imaging in the biological system.

The discovery of new molecular biomarkers of cancer during the last decades and the development of new diagnostic devices exploiting those have significantly contributed to the clinical analysis of cancer and to improve the outcomes. Among those, liquid biopsy sensors exploiting aptamers for the detection of cancer biomarkers in body fluids are useful and accurate tools for a fast and inexpensive non-invasive screening of population. The incorporation of aptamers in electrochemical sandwich biosensors using enzyme labels, a so-called ELASA, has demonstrated its utility to improve the detection schemes. In this review, we overview the existing ELASA assays for numerous cancer biomarkers as alternatives to the traditional ELISA and discuss their possibilities to reach the market, currently dominated by optical immunoassays.

Aroma represents an important quality aspect for wine. The aroma of different grapes and wines is formed by the varying composition and concentrations of numerous aroma compounds, which result in different sensory impressions. The analysis of aroma compounds is usually complex and time-consuming, which requires the development of rapid alternative methods. In this study, grape mash samples were examined for aroma compounds, which were released under tasting conditions. A selection of the determined aroma compounds was grouped according to their sensory characteristics and calibration models were developed for the determination of sensory attributes by near-infrared (NIR) spectroscopy. The calibration models for the selected sensory attributes "fruity," "green," "floral" and "microbiological" showed very high prediction accuracies (0.979 < R2C < 0.996). Moreover, four different grape model solutions, whose compositions were based on the results from GC-MS-based analysis of the grape mash samples, were examined in a sensory evaluation. Despite large variation of the single values, the averaged values of the given scores for intensity of odour and taste showed differences between the model solutions for most of the evaluated sensory attributes. Sensory analysis remains essential for the evaluation of the overall aroma; however, NIR spectroscopy can be used as an additional and more objective method for the estimation of possible desired or undesired flavour nuances of grape mash and the quality of the resulting wine.

An assembled-aptasensor based on Fe3O4@Au@Ag nanocomposites grafting onto the gold foil was prepared, which can be developed into a universal approach for sensitive and rapid detection of various pathogenic bacteria, such as Escherichia coli (E. coli), Salmonella typhimurium (S. typhimurium), Staphylococcus aureus (S. aureus), Listeria monocytogenes (L. monocytogenes), Pseudomonas aeruginosa (P. aeruginosa), and Shigella flexneri (S. flexneri). Firstly, the gold foil paper was modified with thiolated capture probe and SERS tag in proportion, and at the same time, the specific thiolated aptamer probe for corresponding pathogenic bacteria was fixed with Fe3O4@Au@Ag nanocomposites. An obvious Raman signal can be subsequently increased about 106 times by the external electromagnetic field enhancement at the "hot spots" caused by the hybridization of aptamer and capture probe. But in the presence of target pathogenic bacteria, Raman intensity will decrease as Fe3O4@Au@Ag nanocomposites are dissociated from gold foil. Thus, all of the concentrations of the six kinds of pathogenic bacteria both in PBS and liquorice extract showed an obvious negative linear correlation with the Raman intensity of SERS tag in the range of 10-107 CFU/mL with detection limits were all lower than 10 CFU/mL. And there was no significant difference between our method and the plate counting method. Besides, the assembled-aptasensor had superior specific recognition ability even in the mixed interfering bacteria. Our study showed that this assembled-aptasensor had good specific detection ability to a variety of foodborne pathogens based on magnetic field-assisted SERS technique, which can be used for rapid and sensitive detection of a variety of pathogens in complex substrates.

COVID-19 is caused by SARS-CoV-2 infection and remains one of the biggest pandemics around the world since 2019. Vaccination has proved to be an effective way of preventing SARS-CoV-2 infection and alleviating the hospitalization burden. Among different forms of COVID-19 vaccine design, the spike protein of SARS-CoV-2 virus is widely used as a candidate vaccine antigen. As a surface protein on the virus envelop, the spike was reported to be heavily N-glycosylated and glycosylation had a great impact on its immunogenicity and efficacy. Besides, N-glycosylation might vary greatly on different expression systems and sequence variant designs. Therefore, comprehensive analysis of spike N-glycosylation is of great significance for better vaccine understanding and quality control. In this study, full characterization of N-glycosylation was performed for a Chinese Hamster Ovary (CHO) cell expressed variant-designed spike protein. The spike protein featured the latest six-proline substitution design together with the incorporation of a combination of mutation sites. Trypsin and Glu-C digestion coupled with PNGase F strategies were adopted, and effective LC-MS/MS methods were applied to analyze samples. As a result, a total of 19 N-glycosites were identified in the recombinant pike protein at intact N-glycopeptide level. Quantitative analysis of released glycan by LC-MS/MS was also performed, and 31 high-abundance N-glycans were identified. Sequencing analysis of glycan was further provided to assist glycan structure confirmation. Moreover, all of the analyses were performed on three consecutive manufactured batches and the glycosylation results on both glycosite and glycans showed good batch-to-batch consistency. Thus, the reported analytical strategy and N-glycosylation information may well facilitate studies on SARS-CoV-2 spike protein analysis and quality studies.

In this study, the coulometric method with titratable impurity analysis and the mass balance method were successfully applied in the quantification of the certified reference material of potassium hydrogen phthalate (KHP) with accurate metrological traceability of chemical purity value (99.983% with an expanded uncertainty of 0.024%, k = 2). In contrast to the general coulometric titration method, the coulometric method with titratable impurity analysis enables the conversion of acidimetric purity to chemical purity: The acidimetric purity was determined by coulometric titration method, and then the impurities that may be titrated as principal components were found as far as possible using various methods and the result of deducting these impurities from the acidimetric purity can be considered as chemical purity. The mass balance method also accounted for all possible types of impurities as much as possible to improve the accuracy of the determined result. The accuracy and reliability of the purity results were subsequently verified by a two-step quantitative nuclear magnetic resonance (qNMR) method. This KHP certified reference material was the first hydrophilic internal standard of qNMR (applied in polar solvents) with an expanded uncertainty lower than 0.03%, which will become a major reference standard in the organic chemistry traceable calibration chain, especially when evaluating hydrophilic organic compounds to obtain purity values with very low uncertainty.

Trastuzumab is known to be heterogeneous in terms of charge. Stressing trastuzumab under physiological conditions (pH 7.4 and 37 °C) increases charge heterogeneity further. Separation of charge variants of stressed trastuzumab at the intact protein level is challenging due to increasing complexity making it difficult to obtain pure charge variants for further characterization. Here we report an approach for revealing charge heterogeneity of stressed trastuzumab at the subunit level by pH gradient cation-exchange chromatography. Trastuzumab subunits were generated after limited proteolytic cleavage with papain, IdeS, and GingisKHAN®. The basic pI of Fab and F(ab)2 fragments allowed to use the same pH gradient for intact protein and subunit level analysis. Baseline separation of Fab subunits was obtained after GingisKHAN® and papain digestion and the corresponding modifications were determined by LC-MS/MS peptide mapping and middle-down MALDI-ISD FT-ICR MS. The described approach allows a comprehensive charge variant analysis of therapeutic antibodies that have two or more modification sites in the Fab region.

A new analytical method based on the use of dispersive magnetic solid-phase extraction (DMSPE) is described for the preconcentration of capsaicin (CAP), dihydrocapsaicin (DCAP), and N-vanillylnonanamide (PCAP) from human serum samples. The influence of several experimental factors affecting the adsorption (nature and amount of magnetic material, adsorption time, and pH) and desorption (nature of solvent, its volume and desorption time) steps was studied. Among seven different nanomaterials studied, the best results were obtained using magnetic multiwalled carbon nanotubes, which were characterized by means of spectrometry- and microscopy-based techniques. Analyses were performed by ultra-high-performance liquid chromatography with quadrupole-time-of-flight mass spectrometry using electrospray ionization in positive mode (UHPLC-ESI-Q-TOF-MS). The developed method was validated by obtaining several parameters, including linearity (0.3-300 μg L-1 range), and limits of detection which were 0.1, 0.15, and 0.17 μg L-1 for CAP, DCAP, and PCAP, respectively. The repeatability of the method, expressed as relative standard deviation (RSD, n = 7), varied from 3.4 to 11%. The serum samples were also studied through a non-targeted approach in a search for capsaicinoid metabolites and related compounds. With this objective, the fragmentation pathway of this family of compounds was initially studied and a strategy was established for the identification of novel or less studied capsaicinoid-derived compounds.

Glufosinate is widely used to control various weeds. Glufosinate and its main metabolites have become the focus of attention because of their high water solubility and persistence in aquatic systems. Quantification of the agrochemical product and its metabolite residues is essential for the safety of agricultural products. In this study, a highly specific, simple method was developed to directly determine glufosinate and its metabolite residues in 21 plant origin foods by liquid chromatography with tandem mass spectrometry (LC-MS/MS), and it was validated on 11 foods in five laboratories. Finally, the repeatability limit, reproducibility limit, and uncertainty of the method were calculated based on these validated data and used to support the more accurate detection results. Four different chromatographic columns were used to analyze three target compounds, and the anionic polar pesticide column showed the optimum separation and peak shape. Composition of the mobile phase, extraction solvent, and the clean-up procedure were optimized. The developed method was validated on 21 plant origin foods. The average recoveries were 74-115% for all matrices. The validation results of five laboratories showed this method had a good repeatability (RSDr < 9.5%) and reproducibility (RSDR < 18.9%). The method validation parameters met the requirements of guidance established by the European Union (EU) and China for pesticide residue analysis. This methodology can be used for a routine monitoring that performs well for glufosinate and its metabolite residues.

Parkinson's disease is a health-threatening neurodegenerative disease of the elderly with clinical manifestations of motor and non-motor deficits such as tremor palsy and loss of smell. Alpha-synuclein (α-Syn) is the pathological basis of PD, it can abnormally aggregate into insoluble forms such as oligomers, fibrils, and plaques, causing degeneration of nigrostriatal dopaminergic neurons in the substantia nigra in the patient's brain and the formation of Lewy bodies (LBs) and Lewy neuritis (LN) inclusions. As a result, achieving α-Syn aggregate detection in the early stages of PD can effectively stop or delay the progression of the disease. In this paper, we provide a brief overview and analysis of the molecular structures and α-Syn in vivo and in vitro detection methods, such as mass spectrometry, antigen-antibody recognition, electrochemical sensors, and imaging techniques, intending to provide more technological support for detecting α-Syn early in the disease and intervening in the progression of Parkinson's disease.

Hexavalent chromium is a highly toxic substance, which will pose a serious threat to human life and health and the entire ecosystem. Therefore, it is crucial to establish a simple and rapid detection method for hexavalent chromium. In this work, we fabricated bovine serum albumin-stabilized silver nanocluster (BSA-Ag13 NC) which exhibited photoresponsive oxidase-like activity, catalyzing the oxidation of colorless 3,3',5,5'-tetramethylbenzidine (TMB) to the blue oxidized state TMB (oxTMB) in a short time. Interestingly, 8-hydroxyquinoline (8-HQ) can significantly inhibit the color reaction of TMB oxidation while Cr(VI) can interact specifically with 8-HQ to restore this chromogenic reaction. Based on the above facts, a colorimetric sensing system for detecting Cr(VI) was developed. The sensing system shows a wide linear range, and good selectivity, with a low detection limit of 2.32 nM. Moreover, this sensing system could be successfully applied to the detection of Cr(VI) in lake water, tap water, and sewage with satisfactory results.

Bioanalytical and biomedical applications often require nanoparticles that exhibit narrow size distributions and biocompatibility. Here, we demonstrate how different synthesis methods affect gold nanoparticle (AuNPs) monodispersity and cytotoxicity. Using single particle inductively coupled plasma mass spectrometry (SP-ICP-MS), we found that the size distribution of AuNPs synthesized with a cetyltrimethylammonium chloride (CTAC) cap was significantly improved compared to AuNPs synthesized with citrate capping agents. We determined an up to 4× decrease in the full width at half maximum (FWHM) value of the normal distributions of AuNP diameter and up to a 12% decrease in relative standard deviation (RSD). While the CTAC-capped AuNPs exhibit narrow nanoparticle size distributions, they are cytotoxic, which limits safe and effective bioanalytical and biomedical applications. We sought to impart biocompatibility to CTAC-capped AuNPs through a PEGylation-based surface ligand exchange. We developed a unique ligand exchange method driven by physical force. We demonstrated the successful PEGylation using various PEG derivatives and used these PEGylated nanoparticles to further bioconjugate nucleic acids and peptides. Using cell viability quantification, we confirmed that the monodisperse PEGylated AuNPs were biocompatible. Our monodisperse and biocompatible nanoparticles may advance safe and effective bioanalytical and biomedical applications of nanomaterials.

Histones are the major proteinaceous components of chromatin in eukaryotic cells and an important part of the epigenome. The broad-spectrum herbicide atrazine (2-chloro-4-[ethylamino]-6-[isopropylamino]-1, 3, 5-triazine) and its metabolites are known to form protein adducts, but the formation of atrazine-histone adducts has not been studied. In this study, a bottom-up proteomics analysis method was optimized and applied to identify histone adduction by atrazine in vitro. Whole histones of calf thymus or human histone H3.3 were incubated with atrazine. After solvent-based protein precipitation, the protein was digested by trypsin/Glu-C and the resulting peptides were analyzed by high-resolution mass spectrometry using an ultra-high-performance liquid chromatograph interfaced with a quadrupole Exactive-Orbitrap mass spectrometer. The resulting tryptic/Glu-C peptide of DTNLCAIHAK from calf thymus histone H3.1 or human histone H3.3 was identified with an accurate mass shift of  +179.117 Da in atrazine incubated samples. It is deduced that a chemical group with an elemental composition of C8H13N5 (179.1171 Da) from atrazine adducted with calf thymus histone H3.1 or human histone H3.3. It was confirmed by MS/MS analysis that the adduction position was at its cysteine 110 residue. Time- and concentration-dependent assays also confirmed the non-enzymatic covalent modification of histone H3.3 by atrazine in vitro. Thus, the potential exists that atrazine adduction may lead to the alteration of histones that subsequently disturbs their normal function.

Organoid culture is a promising biomedical technology that requires specialized growth factors. Recently, a recombinant L-WRN cell line has been extensively used to generate conditioned medium (L-CM) for organoid culture. Nevertheless, methods for evaluating the stability of the L-WRN cells have been limited. In this study, a novel proteomics-based approach was developed to analyze the secretome of the cells. Serum-free L-CM was lyophilized, precipitated by trichloroacetic acid, and desalted prior to analysis by liquid chromatography-tandem mass spectrometry. Data-dependent acquisition (DDA) was conducted for the untargeted secretome profiling of the cells, and parallel reaction monitoring (PRM) was applied for the targeted quantification of the Wnt3A, R-spondin3, and noggin proteins (WRNs). This study also compared the performance of two types of PRM methods, namely MS1-independent PRM and MS1-dependent PRM, that can be executed on an Orbitrap instrument. The results showed that the growth of mouse intestinal organoids was closely related to the use of L-CM. The composition of L-CM could be markedly affected by the medium collection scheme. A total of 1725, 2302, and 2681 proteins were identified from the L-CM collected on day 5, day 9, and day 13, respectively. The MS1-independent PRM outperformed the MS1-dependent PRM and effectively quantified the WRNs with high repeatability and specificity. In conclusion, by integrating untargeted and targeted proteomics, this study develops a mass spectrometry-based method for the secretome analysis and quality control of the L-WRN cells. The methodology and findings of the present work will benefit future studies on organoids and secretomes.

One convenient strategy to reduce environmental impact and pollution involves the reuse and revalorization of waste produced by modern society. Nowadays, global plastic production has reached 367 million tons per year and because of their durable nature, their recycling is fundamental for the achievement of the circular economy objective. In closing the loop of plastics, advanced recycling, i.e., the breakdown of plastics into their building blocks and their transformation into valuable secondary raw materials, is a promising management option for post-consumer plastic waste. The most valuable product from advanced recycling is a fluid hydrocarbon stream (or pyrolysis oil) which represents the feedstock for further refinement and processing into new plastics. In this context, gas chromatography is currently playing an important role since it is being used to study the pyrolysis oils, as well as any organic contaminants, and it can be considered a high-resolution separation technique, able to provide the molecular composition of such complex samples. This information significantly helps to tailor the pyrolysis process to produce high-quality feedstocks. In addition, the detection of contaminants (i.e., heteroatom-containing compounds) is crucial to avoid catalytic deterioration and to implement and design further purification processes. The current review highlights the importance of molecular characterization of waste stream products, and particularly the pyrolysis oils obtained from waste plastics. An overview of relevant applications published recently will be provided, and the potential of comprehensive two-dimensional gas chromatography, which represents the natural evolution of gas chromatography into a higher-resolution technique, will be underlined.

A stopped-flow microfluidic system to monitor glutathione peroxidase (GPx) activity and evaluate potential inhibitors of the enzyme has been developed based on the integration of the microfluidic chip in the reaction/detection zone. This integration supposes the physical alignment at the optimal location of the microfluidic channel, both the magnetically retained enzyme microreactor (MREµR) and the remote luminescence detection using a focused bifurcated fiber optic bundle (BFOB) connected to a conventional spectrofluorometer detector. The method is based on the coupling of two competitive oxidative chemical reactions, in which glutathione (GSH) and homovanillic acid (HVA) competed for their interaction with hydrogen peroxide in the presence of the magnetically retained GPx-MNPs. The biocatalytic reaction was followed by monitoring the fluorescence of the biphenyl-HVA dimer formed. The dynamic range of the calibration graph was 0.45-10 µmol L-1, expressed as GSH concentration with a detection limit of 0.1 µmol L-1 (r2 = 0.9954, n = 10, r = 3). The precision expressed as the relative standard deviation (RSD%) was between 0.5 and 3.9%. The stopped-flow microfluidic system showed a sampling frequency of 25 h-1. The method was applied to the study of GPx inhibition provided by three inhibitory compounds, two metallic ions Hg(II) and Cu(II) and t-butyl hydroperoxide, and their presence in liquid samples, as water, milk, and edible oil. Recovery values between 88.7 and 99.4% were achieved in all instances.

To systematically study the influence of host-guest interactions on the analytical performance of direct analysis in real time mass spectrometry (DART-MS), the interactions between cyclodextrins (CDs) and different Sudan dyes were investigated. The results showed that the host-guest interaction between CDs and Sudan dyes did not affect qualitative analysis of the target compounds, but led to a lower signal intensity for Sudan dyes, thus affecting quantitative analysis of the target compounds. The stronger the host-guest interaction, the weaker the signal intensity of target compound on DART-MS. The results also show that both in solution and in solid-phase microextraction (SPME), the addition of organic solvents can weaken the host-guest interaction between CDs and Sudan dyes, thus improving the signal intensity in DART-MS. In SPME, adding organic solvents has a certain practical value and can improve the efficiency of Sudan dye analysis. This study suggests that appropriate sample pretreatment is needed to weaken noncovalent interactions prior to DART-MS analysis to obtain more accurate quantitative results. The data provide some insight into the effects of other noncovalent interactions on the efficiency of DART-MS as an analytical tool, as well as the potential to study intermolecular interactions with DART-MS.

Polybrominated diphenyl ethers (PBDEs) are considered emerging organic contaminants that attract more attention in the environment. Herein, online coupling of solid-phase microextraction and ultrahigh-resolution mass spectrometry was developed for rapid screening of eight PBDEs in water samples. This procedure was completed in 22 min, about 6 times faster than the routine workflow such as solid-phase extraction coupled with gas chromatography-mass spectrometry. Thermal desorption and solvent-assisted atmospheric pressure chemical ionization were developed for the effective coupling of solid-phase microextraction (SPME) with ultrahigh-resolution mass spectrometry (UHRMS), which contributed to the signal enhancement and made the methodology feasible for environmental screening. The limits of detection and quantification were 0.01-0.50 ng/mL and 0.05-4.00 ng/mL, respectively. The recoveries were 57.2-75.2% for quality control samples at spiking levels of 0.8-10 ng/mL (4-50 ng/mL for BDE209), with relative standard deviation less than 19.0%. Twelve water samples from different river sites near industrial areas were screened using the developed method. The results showed that BDE-209 was the dominant PBDE (1.02-1.28 ng/mL in positive samples), but its amount was lower than the human health ambient water quality criteria. Consequently, the developed method provides a rapid and reliable way of evaluating contamination status and risks of PBDEs in aqueous environment.

Since its discovery in 2007, polydopamine nanofilms have been widely used in many areas for surface functionalization. The simple and low-cost preparation method of the nanofilms with tunable thickness can incorporate amine and oxygen-rich chemical groups in virtually any interface. The remarkable advantages of this route have been successfully used in the field of electrochemical sensors. The self-adhesive properties of polydopamine are used to attach nanomaterials onto the electrode's surface and add chemical groups that can be explored to immobilize recognizing species for the development of biosensors. Thus, the combination of 2D materials, nanoparticles, and other materials with polydopamine has been successfully demonstrated to improve the selectivity and sensitivity of electrochemical sensors. In this review, we highlight some interesting properties of polydopamine and some applications where polydopamine plays an important role in the field of electrochemical sensors.

The so-coined fourth paradigm in science has reached the sensing area, with the use of machine learning (ML) toward data-driven improvements in sensitivity, reproducibility, and accuracy, along with the determination of multiple targets from a single measurement using multi-output regression models. Particularly, the use of supervised ML models trained on large data sets produced by electrical and electrochemical bio/sensors has emerged as an impacting trend in the literature by allowing accurate analyses even in the presence of usual issues such as electrode fouling, poor signal-to-noise ratio, chemical interferences, and matrix effects. In this trend article, apart from an outlook for the coming years, we present examples from the literature that demonstrate how helpful ML algorithms can be for dispensing the adoption of experimental methods to address the aforesaid interfering issues, ultimately contributing to translate testing technologies into on-site, practical, and daily applications.

In this paper, we proposed an efficient method for mercury(II) and glutathione detection using a fluorescent nanoprobe as a sensor. Carbon dots were synthesized from polyethyleneimine and ammonium citrate via a one-step hydrothermal method. The fluorescence of carbon dots was quenched since electron transfer occurred due to the interaction between mercury(II) and functional groups on the surface of carbon dots. Adding glutathione to the carbon dots-mercury(II) system, the fluorescence was recovered due to the stronger binding ability of glutathione to mercury(II). Based on the above-mentioned principle, this "off-on" fluorescent sensor can easily achieve the detection of mercury(II) and glutathione, which provided limits of detection of 22.45 nM and 61.89 nM, respectively. In this paper, the proposed method has been applied to detect mercury(II) and glutathione in real lake water and serum, respectively, and a logic gate for sensing glutathione was presented. The developed "off-on" fluorescent sensor with high sensitivity and selectivity has shown great potential for mercury(II) and glutathione detection in environmental and biosensing fields.

Nanozymes exhibit their great potential as alternatives to natural enzymes. In addition to catalytic activity, nanozymes also need to have biologically relevant catalytic reactions at physiological pH to fit in the definition of an enzyme and to achieve efficient analytical applications. Previous reviews in the nanozyme field mainly focused on the catalytic mechanisms, activity regulation, and types of catalytic reactions. In this paper, we discuss efforts made on the substrate-dependent catalytic activity of nanozymes at neutral pH. First, the discrepant catalytic activities for different substrates are compared, where the key differences are the characteristics of substrates and the adsorption of substrates by nanozymes at different pH. We then reviewed efforts to enhance reaction activity for model chromogenic substrates and strategies to engineer nanomaterials to accelerate reaction rates for other substrates at physiological pH. Finally, we also discussed methods to achieve efficient sensing applications at neutral pH using nanozymes. We believe that the nanozyme is catching up with enzymes rapidly in terms of reaction rates and reaction conditions. Designing nanozymes with specific catalysis for efficient sensing remains a challenge.

Comprehensive two-dimensional gas chromatography with time-of-flight mass spectrometry (GC × GC-TOFMS) is one the most powerful analytical platforms for chemical investigations of complex biological samples. It produces large datasets that are rich in information, but highly complex, and its consistency may be affected by random systemic fluctuations and/or changes in the experimental parameters. This study details the optimization of a data processing strategy that compensates for severe 2D pattern misalignments and detector response fluctuations for saliva samples analyzed across 2 years. The strategy was trained on two batches: one with samples from healthy subjects who had undergone dietary intervention with high/low-Maillard reaction products (dataset A), and the second from healthy/unhealthy obese individuals (dataset B). The combined untargeted and targeted pattern recognition algorithm (i.e., UT fingerprinting) was tuned for key process parameters, the signal-to-noise ratio (S/N), and MS spectrum similarity thresholds, and then tested for the best transform function (global or local, affine or low-degree polynomial) for pattern realignment in the temporal domain. Reliable peak detection achieved its best performance, computed as % of false negative/positive matches, with a S/N threshold of 50 and spectral similarity direct match factor (DMF) of 700. Cross-alignment of bi-dimensional (2D) peaks in the temporal domain was fully effective with a supervised operation including multiple centroids (reference peaks) and a match-and-transform strategy using affine functions. Regarding the performance-derived response fluctuations, the most promising strategy for cross-comparative analysis and data fusion included the mass spectral total useful signal (MSTUS) approach followed by Z-score normalization on the resulting matrix.

In this work, a solid-phase microextraction (SPME) Arrow method combined with comprehensive two-dimensional gas chromatography-mass spectrometry (GC × GC-MS) was developed for the elucidation of the volatile composition of honey samples. The sample preparation protocol was optimized to ensure high extraction efficiency of the volatile organic compounds (VOCs) which are directly associated with the organoleptic properties of honey and its acceptance by the consumers. Following its optimization, SPME Arrow was compared to conventional SPME in terms of sensitivity, precision, and number of extracted VOCs. The utilization of SPME Arrow fibers enabled the determination of 203, 147, and 149 compounds in honeydew honey, flower honey, and pine honey, respectively, while a significantly lower number of compounds (124, 94, and 111 for honeydew honey, flower honey, and pine honey, respectively) was determined using conventional SPME. At the same time, the utilization of SPME Arrow resulted in enhanced sensitivity and precision. All things considered, SPME Arrow and GC × GC-MS can be considered as highly suitable for the elucidation of the volatile composition of complex food samples resulting in high sensitivity and separation efficiency.

Microbial metabolomics allows understanding and to comprehensively analyse metabolites, and their related cellular and metabolic processes, that are produced and released to the extracellular environment under specific conditions. In that regard, the main objective of this research is to understand the impact of culture media changes in the metabolic profile of Pedobacter lusitanus NL19 (NL19) and Pedobacter himalayensis MTCC 6384 (MTCC6384) and respective influence on the production of biotechnologically relevant compounds. Solid-phase microextraction combined with comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry with time-of-flight analyser (GC × GC-ToFMS) was applied to comprehensively study the metabolites produced by NL19 and MTCC6384 both in tryptic soy broth 100% (TSB100) and tryptic soy broth with 25% casein peptone (PC25). A total of 320 metabolites were putatively identified, which belong to different chemical families: alcohols, aldehydes, esters, ethers, hydrocarbons, ketones, nitrogen compounds, sulphur compounds, monoterpenes, and sesquiterpenes. Metabolites that were statistically different from the control (sterile medium) were selected allowing for the construction of the metabolic profile of both strains. A set of 80 metabolites was tentatively associated to the metabolic pathways such as the metabolism of fatty acids, branched-chain aminoacids, phenylalanine, methionine, aromatic compounds, and monoterpene and sesquiterpene biosynthesis. This study allowed to better understand how slight changes of the culture media and thus the composition of nutrients impair the metabolic profile of bacteria, which may be further explored for metabolomics pipeline construction or biotechnological applications.

The separation and identification of lipids in complex mixtures are critical to deciphering their cellular functions. Failure to resolve isobaric compounds (e.g., via high mass resolution or tandem mass spectrometry) can result in incorrect identifications in mass spectrometry experiments. In imaging mass spectrometry, unresolved peaks can also result in composite images of multiple compounds, giving inaccurate depictions of molecular distributions. Gas-phase ion/ion reactions can be used to selectively react with specific chemical functional groups on a target analyte, thereby extracting it from a complex mixture and shifting its m/z value to an unobstructed region of the mass range. Herein, we use selective Schiff base formation via a novel charge inversion ion/ion reaction to purify phosphatidylserines from other isobaric (i.e., same nominal mass) lipids and reveal their singular distributions in imaging mass spectrometry. The selective Schiff base formation between singly deprotonated phosphatidylserine (PS) lipid anions and doubly charged N,N,N',N'-tetramethyl-N,N'-bis(6-oxohexyl)hexane-1,6-diaminium (TMODA) cations is performed using a modified commercial dual source hybrid Fourier transform ion cyclotron resonance (FTICR) mass spectrometer. This process is demonstrated using the isobaric lipids [PS 40:6 - H]- (m/z 834.528) and [SHexCer d38:1 - H]- (m/z 834.576), which produces [PS 40:6 + TMODA - H - H2O]+ (m/z 1186.879), and [SHexCer d38:1 + TMODA - H]+ (m/z 1204.938) product ions following the gas-phase charge inversion reaction. These product ions differ by roughly 18 Da in mass and are easily separated by low mass resolution analysis, while the isobaric precursor ions require roughly 45,000 mass resolving power (full-width at half maximum) to separate. Imaging mass spectrometry using targeted gas-phase ion/ion reactions shows distinct spatial distributions for the separated lipid product ions relative to the composite images of the unseparated precursor ions.

Sensitive detection of the minute and yet pathologically significant pH variation is important and in fact challenging for the conventional pH probes following the Henderson-Hasselbalch equation, i.e., HH-type probes. A paradigm shift to Hill-type pH probes is ongoing. Bestowed by their positive cooperative acid-base chemistry, their pH-responsive profile follows the Hill equation, which exhibits a narrower acid/base transition width than HH-type probes and warrants a higher detection sensitivity. A polymer-based Hill-type pH-responsive material was first developed. More recently, there emerged several distinct small-molecular approaches to achieve Hill-type pH-responsive profiles. They complement the polymer-based sensing materials in applications where membrane permeability is a concern. In this trends article, we rationalize the molecular origins of their positive cooperativity in pH sensing and highlight some interesting proof-of-concept applications. We also discussed future directions of this dynamic research area.