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Methanotrophs are crucial in keeping environmental CH4 emissions in check. However, the contributions of different groups of methanotrophs at terrestrial CH4-oxidation hotspots, such as the oxic-anoxic interface of rice paddies, have shown considerable inconsistency across observations. To address the knowledge gap regarding this inconsistency, methanotrophic microbiomes were enriched from paddy soils in well-mixed CH4-fed batch reactors under six different incubation conditions, prepared as combinations of two CH4 mixing ratios (0.5 and 10%) and three supplemented Cu2+ concentrations (0, 2, and 10 µM). Monitoring of temporal community shifts in these cultures revealed a dominance of Methylocystis spp. in all 0.5%-CH4 cultures, while methanotrophs affiliated to Gammaproteobacteria dominated the 10%-CH4 cultures that were less consistent both temporally and across conditions. The shotgun metagenome analyses of the 0.5%-CH4 cultures corroborated the Methylocystis dominance and, interestingly, showed that copper deficiency did not select for mmoXYZ-possessing methanotrophs. Instead, a mbn cluster, accounting for approximately 5% of the Methylocystis population, was identified, suggesting the ecological significance of methanobactin in Cu-deficient methanotrophy. These findings underscore the important role of Methylocystis spp. in mitigating emissions from terrestrial CH4 hotspots and suggest the feasibility of directed enrichment and/or isolation of Methylocystis spp. for utilization in, for example, methanobactin and polyhydroxybutyrate production.
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Metano , Methylococcaceae , Methylocystaceae , Metano/metabolismo , Methylococcaceae/metabolismo , Methylocystaceae/metabolismo , Microbiologia do Solo , MicrobiotaRESUMO
Unique means of copper scavenging have been identified in proteobacterial methanotrophs, particularly the use of methanobactin, a novel ribosomally synthesized, post-translationally modified polypeptide that binds copper with very high affinity. The possibility that copper sequestration strategies of methanotrophs may interfere with copper uptake of denitrifiers in situ and thereby enhance N2O emissions was examined using a suite of laboratory experiments performed with rice paddy microbial consortia. Addition of purified methanobactin from Methylosinus trichosporium OB3b to denitrifying rice paddy soil microbial consortia resulted in substantially increased N2O production, with more pronounced responses observed for soils with lower copper content. The N2O emission-enhancing effect of the soil's native mbnA-expressing Methylocystaceae methanotrophs on the native denitrifiers was then experimentally verified with a Methylocystaceae-dominant chemostat culture prepared from a rice paddy microbial consortium as the inoculum. Finally, with microcosms amended with various cell numbers of methanobactin-producing Methylosinus trichosporium OB3b before CH4 enrichment, microbiomes with different ratios of methanobactin-producing Methylocystaceae to gammaproteobacterial methanotrophs incapable of methanobactin production were simulated. Significant enhancement of N2O production from denitrification was evident in both Methylocystaceae-dominant and Methylococcaceae-dominant enrichments, albeit to a greater extent in the former, signifying the comparative potency of methanobactin-mediated copper sequestration, while implying the presence of alternative copper abstraction mechanisms for Methylococcaceae. These observations support that copper-mediated methanotrophic enhancement of N2O production from denitrification is plausible where methanotrophs and denitrifiers cohabit. IMPORTANCE Proteobacterial methanotrophs-groups of microorganisms that utilize methane as a source of energy and carbon-have been known to employ unique mechanisms to scavenge copper, namely, utilization of methanobactin, a polypeptide that binds copper with high affinity and specificity. Previously the possibility that copper sequestration by methanotrophs may lead to alteration of cuproenzyme-mediated reactions in denitrifiers and consequently increase emission of potent greenhouse gas N2O has been suggested in axenic and coculture experiments. Here, a suite of experiments with rice paddy soil slurry cultures with complex microbial compositions were performed to corroborate that such copper-mediated interplay may actually take place in environments cohabited by diverse methanotrophs and denitrifiers. As spatial and temporal heterogeneity allows for spatial coexistence of methanotrophy (aerobic) and denitrification (anaerobic) in soils, the results from this study suggest that this previously unidentified mechanism of N2O production may account for a significant proportion of N2O efflux from agricultural soils.
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Cobre/metabolismo , Imidazóis/metabolismo , Consórcios Microbianos , Óxido Nitroso , Oligopeptídeos/metabolismo , Proteobactérias/metabolismo , Óxido Nitroso/metabolismo , Solo/química , Microbiologia do SoloRESUMO
DNA methyltransferases (DNMTs) are enzymes responsible for establishing and maintaining DNA methylation in cells. DNMT inhibition is actively pursued in cancer treatment, dominantly through the formation of irreversible covalent complexes between small molecular compounds and DNMTs that suffers from low efficacy and high cytotoxicity, as well as no selectivity towards different DNMTs. Herein, we discover aptamers against the maintenance DNA methyltransferase, DNMT1, by coupling Asymmetrical Flow Field-Flow Fractionation (AF4) with Systematic Evolution of Ligands by EXponential enrichment (SELEX). One of the identified aptamers, Apt. #9, contains a stem-loop structure, and can displace the hemi-methylated DNA duplex, the native substrate of DNMT1, off the protein on sub-micromolar scale, leading for effective enzymatic inhibition. Apt. #9 shows no inhibition nor binding activity towards two de novo DNMTs, DNMT3A and DNMT3B. Intriguingly, it can enter cancer cells with over-expression of DNMT1, colocalize with DNMT1 inside the nuclei, and inhibit the activity of DNMT1 in cells. This study opens the possibility of exploring the aptameric DNMT inhibitors being a new cancer therapeutic approach, by modulating DNMT activity selectively through reversible interaction. The aptamers could also be valuable tools for study of the functions of DNMTs and the related epigenetic mechanisms.
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Aptâmeros de Nucleotídeos/química , DNA (Citosina-5-)-Metiltransferase 1/antagonistas & inibidores , Metilação de DNA/genética , Neoplasias/genética , Aptâmeros de Nucleotídeos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Desoxicitidina/análogos & derivados , Desoxicitidina/análise , Epigênese Genética/genética , Células HEK293 , Células HeLa , Humanos , Neoplasias/tratamento farmacológicoRESUMO
Humulus japonicus is an annual plant belonging to the Cannabacea family, and it has been traditionally used to treat pulmonary tuberculosis, dysentery, chronic colitis, and hypertension. We investigated the active components against Parkinson's disease from H. japonicus fraction (HJF) using high performance liquid chromatography (HPLC) coupled with quadruple-time-of-flight mass spectroscopy (qTOF-MS) and NMR. Fourteen compounds were isolated from HJF, including one new compound, using HPLC-qTOF-MS and NMR. The major compounds of HJF were luteolin-7-O-glucoside and apigenin-7-O-glucoside, and there was approximately 12.57- and 9.68-folds increase in the contents of these flavonoids compared to those of the 70% EtOH extract. Apigenin and luteolin exhibited the strongest inhibitory effects on monoamine oxidase (MAO) B enzyme activity. In animal studies, limb-use behavior was significantly reduced by unilateral 6-OHDA lesion and ipsilateral rotations. These results indicated that oral administration of 300 mg/kg HJF resulted in the improvement of motor asymmetry and motor impairment in unilateral 6-OHDA-lesioned mice. HJF, including active components leads to an improvement of motor behavior in a Parkinson's disease mouse model.
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Humulus/química , Atividade Motora/efeitos dos fármacos , Doença de Parkinson Secundária/tratamento farmacológico , Extratos Vegetais/química , Administração Oral , Animais , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Flavonas/administração & dosagem , Flavonas/química , Regulação da Expressão Gênica/efeitos dos fármacos , Glucosídeos/administração & dosagem , Glucosídeos/química , Humanos , Espectroscopia de Ressonância Magnética , Camundongos , Monoaminoxidase/genética , Atividade Motora/genética , Oxidopamina/toxicidade , Doença de Parkinson Secundária/induzido quimicamente , Doença de Parkinson Secundária/genética , Doença de Parkinson Secundária/patologia , Extratos Vegetais/administração & dosagem , Espectrometria de Massas em TandemRESUMO
Nanoparticles (NPs) tend to adsorb matrix molecules like proteins and lipids incubated with biological fluids, forming a biological corona. While the formation and functions of protein corona have been studied extensively, little attention has been paid to lipid adsorption on NPs. However, lipids are also abundantly present in biological fluids and play important roles in processes like cell signaling and angiogenesis. Therefore, in this study, we established the analytical procedure for study of lipid adsorption on three different types of NPs in two matrices: human serum and heavy cream, using nanoflow liquid chromatography-mass spectrometry (nanoflowLC-MS). Serum was chosen to represent the common environment the NPs would be present once entering human body, and heavy cream was the representative food matrix NPs may be added to improve the color or taste. Steps of liquid-liquid extraction were established and optimized to achieve maximum recovery of the adsorbed, standard lipids from the NPs. Then, the LC-MS/MS method was developed to attain base-line separation of the standard lipids that represent the major lipid classes. At last, the lipid adsorption profiles of the three NPs were compared. We found that the lipid adsorption profile on the same type of NP was significantly different between the two matrices. The established method will help us investigate lipid adsorption on additional NPs and reveal how it could be affected by the physiochemical properties of NPs and the presence of proteins and other components in the biological matrix.
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Laticínios/análise , Lipídeos/sangue , Lipídeos/química , Nanopartículas/química , Titânio/química , Adsorção , Cromatografia Líquida , Humanos , Extração Líquido-Líquido , Espectrometria de Massas em TandemRESUMO
Oxidized phospholipids (Ox-PLs) are oxidatively modified PLs that are produced during the oxidation of lipoproteins; oxidation of low density lipoproteins especially is known to be associated with the development of coronary artery disease (CAD). In this study, different lipoprotein classes (high density, low density, and very low density lipoproteins) from pooled plasma of CAD patients and pooled plasma from healthy controls were size-sorted on a semipreparative scale by multiplexed hollow fiber flow field-flow fractionation (MxHF5), and Ox-PLs that were extracted from each lipoprotein fraction were quantified by nanoflow liquid chromatography-tandem mass spectrometry (nLC-ESI-MS/MS). The present study showed that oxidation of lipoproteins occurred throughout all classes of lipoproteins with more Ox-PLs identified from CAD patient lipoproteins: molecular structures of 283 unique PL species (including 123 Ox-PLs) from controls and 315 (including 169 Ox-PLs) from patients were identified by data-dependent collision-induced dissociation experiments. It was shown that oxidation of PLs occurred primarily with hydroxylation of PL; in particular, a saturated acyl chain such as 16:0, 18:0, or even 18:1 at the sn-1 location of the glycerol backbone along with sn-2 acyl chains with at least two double bonds were identified. The acyl chain combinations commonly found for hydroxylated Ox-PLs in the lipoproteins of CAD patients were 16:0/18:2, 16:0/20:4, 18:0/18:2, and 18:0/20:4.
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Doença da Artéria Coronariana/sangue , Lipoproteínas/sangue , Lipoproteínas/química , Fosfolipídeos/análise , Fosfolipídeos/sangue , Adulto , Cromatografia Líquida/métodos , Doença da Artéria Coronariana/metabolismo , Desenho de Equipamento , Fracionamento por Campo e Fluxo/instrumentação , Fracionamento por Campo e Fluxo/métodos , Humanos , Lipoproteínas/metabolismo , Pessoa de Meia-Idade , Oxirredução , Fosfolipídeos/metabolismo , Espectrometria de Massas por Ionização por Electrospray/métodos , Espectrometria de Massas em Tandem/métodosRESUMO
Separation/isolation of subcellular species, such as mitochondria, lysosomes, peroxisomes, Golgi apparatus, and others, from cells is important for gaining an understanding of the cellular functions performed by specific organelles. This study introduces a high speed, semipreparative scale, biocompatible size sorting method for the isolation of subcellular organelle species from homogenate mixtures of HEK 293T cells using flow field-flow fractionation (FlFFF). Separation of organelles was achieved using asymmetrical FlFFF (AF4) channel system at the steric/hyperlayer mode in which nuclei, lysosomes, mitochondria, and peroxisomes were separated in a decreasing order of hydrodynamic diameter without complicated preprocessing steps. Fractions in which organelles were not clearly separated were reinjected to AF4 for a finer separation using the normal mode, in which smaller sized species can be well fractionated by an increasing order of diameter. The subcellular species contained in collected AF4 fractions were examined with scanning electron microscopy to evaluate their size and morphology, Western blot analysis using organelle specific markers was used for organelle confirmation, and proteomic analysis was performed with nanoflow liquid chromatography-tandem mass spectrometry (nLC-ESI-MS/MS). Since FlFFF operates with biocompatible buffer solutions, it offers great flexibility in handling subcellular components without relying on a high concentration sucrose solution for centrifugation or affinity- or fluorescence tag-based sorting methods. Consequently, the current study provides an alternative, competitive method for the isolation/purification of subcellular organelle species in their intact states.
Assuntos
Fracionamento Celular/métodos , Fracionamento por Campo e Fluxo , Organelas/química , Western Blotting , Células Cultivadas , Células HEK293 , Humanos , Tamanho da Partícula , Proteínas/análise , Espectrometria de Massas por Ionização por Electrospray , Propriedades de Superfície , Espectrometria de Massas em TandemRESUMO
Lifetime-reconfigurable soft robots have emerged as a new class of robots, emphasizing the unmet needs of futuristic sustainability and security. Trigger-transient materials that can both actuate and degrade on-demand are crucial for achieving life-reconfigurable soft robots. Here, we propose the use of transient and magnetically actuating materials that can decompose under ultraviolet light and heat, achieved by adding photo-acid generator (PAG) and magnetic particles (Sr-ferrite) to poly(propylene carbonate) (PPC). Chemical and thermal analyses reveal that the mechanism of PPC-PAG decomposition occurs through PPC backbone cleavage by the photo-induced acid. The self-assembled monolayer (SAM) encapsulation of Sr-ferrite preventing the interaction with the PAG allowed the transience of magnetic soft actuators. We demonstrate remotely controllable and degradable magnetic soft kirigami actuators using blocks with various magnetized directions. This study proposes novel approaches for fabricating lifetime-configurable magnetic soft actuators applicable to diverse environments and applications, such as enclosed/sealed spaces and security/military devices.
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Three-dimensional (3D) printing has various applications in many fields, such as soft electronics, robotic systems, biomedical implants, and the recycling of thermoplastic composite materials. Three-dimensional printing, which was only previously available for prototyping, is currently evolving into a technology that can be utilized by integrating various materials into customized structures in a single step. Owing to the aforementioned advantages, multi-functional 3D objects or multi-material-designed 3D patterns can be fabricated. In this study, we designed and fabricated 3D-printed expandable structural electronics in a substrateless auxetic pattern that can be adapted to multi-dimensional deformation. The printability and electrical conductivity of a stretchable conductor (Ag-RTV composite) were optimized by incorporating a lubricant. The Ag-RTV and RTV were printed in the form of conducting voxels and frame voxels through multi-nozzle printing and were arranged in a negative Poisson's ratio pattern with a missing rib structure, to realize an expandable passive component. In addition, the expandable structural electronics were embedded in a soft actuator via one-step printing, confirming the possibility of fabricating stable interconnections in expanding deformation via a missing rib pattern.
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Iontronic sensors have garnered significant attention as wearable sensors due to their exceptional mechanical performance and the ability to maintain electrical performance under various mechanical stimuli. Iontronic sensors can respond to stimuli like mechanical stimuli, humidity, and temperature, which has led to exploration of their potential as versatile sensors. Here, a comprehensive review of the recent researches and developments on several types of iontronic sensors (e.g., pressure, strain, humidity, temperature, and multi-modal sensors), in terms of their sensing principles, constituent materials, and their healthcare-related applications is provided. The strategies for improving the sensing performance and environmental stability of iontronic sensors through various innovative ionic materials and structural designs are reviewed. This review also provides the healthcare applications of iontronic sensors that have gained increased feasibility and broader applicability due to the improved sensing performance. Lastly, outlook section discusses the current challenges and the future direction in terms of the applicability of the iontronic sensors to the healthcare.
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Regeneration of over 10 mm long peripheral nerve defects remains a challenge due to the failure of regeneration by prolonged axotomy and denervation occurring in long-term recovery. Recent studies reveal that conductive conduits and electrical stimulation accelerate the regeneration of long nerve defects. In this study, an electroceutical platform combining a fully biodegradable conductive nerve conduit and a wireless electrical stimulator is proposed to maximize the therapeutic effect on nerve regeneration. Fully biodegradable nerve conduit fabricated using molybdenum (Mo) microparticles and polycaprolactone (PCL) can eliminate the unwanted effects of non-degradable implants, which occupy nerve paths and need to be removed through surgery increasing the risk of complications. The electrical and mechanical properties of Mo/PCL conduits are optimized by controlling the amounts of Mo and tetraglycol lubricant. The dissolution behavior and electrical conductivity of biodegradable nerve conduits in the biomimetic solutions are also evaluated. In in vivo experiments, the integrated strategy of a conductive Mo/PCL conduit with controlled therapeutic electrical stimulation shows accelerated axon regeneration for long sciatic nerve defects in rats compared to the use of the Mo/PCL conduit without stimulation and has a significant therapeutic effect based on the results obtained from the functional recovery test.
Assuntos
Axônios , Regeneração Nervosa , Ratos , Animais , Regeneração Nervosa/fisiologia , Próteses e Implantes , Nervo Isquiático/fisiologia , Condutividade ElétricaRESUMO
Hexagonal boron nitride (h-BN) is an excellent support material for nanocatalysts due to its two-dimensional (2D) architectural morphology and physicochemical stability. In this study, a chemically stable, recoverable, eco-friendly, and magnetic h-BN/Pd/Fe2O3 catalyst was prepared by a one-step calcination process, in which Pd and Fe2O3 nanoparticles (NPs) were uniformly decorated on the surface of h-BN via a typical adsorption-reduction procedure. In detail, nanosized magnetic (Pd/Fe2O3) NPs were derived from a Prussian blue analogue prototype, a well-known porous metal-organic framework, and then further surface-engineered to produce magnetic BN nanoplate-supported Pd nanocatalysts. The structural and morphological features of h-BN/Pd/Fe2O3 were investigated by spectroscopic and microscopic characterization techniques. Moreover, the h-BN nanosheets endow it with stability and appropriate chemical anchoring sites which solve the problems of inefficient reaction rate and high consumption caused by the inevitable agglomeration of precious metal NPs. Under mild reaction conditions, the developed nanostructured h-BN/Pd/Fe2O3 as the catalyst shows high yield and efficient reusability in reducing nitroarenes into the corresponding anilines using sodium borohydride (NaBH4) as a reductant.
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Developing soft robots that can control their own life cycle and degrade on-demand while maintaining hyperelasticity is a notable research challenge. On-demand degradable soft robots, which conserve their original functionality during operation and rapidly degrade under specific external stimulation, present the opportunity to self-direct the disappearance of temporary robots. This study proposes soft robots and materials that exhibit excellent mechanical stretchability and can degrade under ultraviolet light by mixing a fluoride-generating diphenyliodonium hexafluorophosphate with a silicone resin. Spectroscopic analysis revealed the mechanism of SiâOâSi backbone cleavage using fluoride ion (F-) and thermal analysis indicated accelerated decomposition at elevated temperatures. In addition, we demonstrated a robotics application by fabricating electronics integrated gaiting robot and a fully closed-loop trigger disintegration robot for autonomous, application-oriented functionalities. This study provides a simple yet novel strategy for designing life cycle mimicking soft robotics that can be applied to reduce soft robotics waste, explore hazardous areas, and ensure hardware security with on-demand destructive material platforms.
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The efficiencies of four different methods for the extraction of phospholipids (PLs) and lysophospholipids (LPLs) from human plasma samples were examined by comparing extraction recovery values using nanoflow liquid chromatography-electrospray ionization-mass spectrometry (nLC-ESI-MS). For recovery measurements, six PL and six LPL standards of different head groups were spiked into a human plasma sample, and the peak areas of each individual species after extraction were measured from the chromatograms of the nLC-ESI-MS runs. Recovery was calculated by comparing the peak area of an extracted standard species with that of the same species' spike after extraction of the same plasma sample. For lipid extraction, four different extraction methods were examined: three based on the Folch method with different organic solvents such as CHCl(3), methyl-tert-butyl ether (MTBE), and MTBE/CH(3)OH, and one relatively fast method involving CH(3)OH only. Evaluations of recovery showed that the modified Folch method with MTBE/CH(3)OH proposed in this study was effective for extracting most PL and LPL standards. Then, the four extraction methods were compared with the identified numbers of plasma PLs and LPLs, of which molecular structures can be confirmed by data-dependent, collision-induced dissociation experiments during nLC-ESI-MS-MS. These results demonstrated that the proposed method yielded the identification of 54 LPLs and 66 PLs from a plasma sample, which was the highest identification rate among the four methods.
Assuntos
Cromatografia Líquida , Lisofosfolipídeos/sangue , Lisofosfolipídeos/isolamento & purificação , Nanotecnologia , Fosfolipídeos/sangue , Fosfolipídeos/isolamento & purificação , Espectrometria de Massas por Ionização por Electrospray , HumanosRESUMO
Brain metastases (BM) are common in patients with non-small cell lung cancer (NSCLC). However, the pure economic burden of BM is unknown. This study aimed to evaluate the impact of BM on healthcare costs and resource utilization in patients with NSCLC by comparing patients with and without BM. This was a retrospective cohort analysis of South Korean health insurance review and assessment claims data. Patients with stage IIIB or IV NSCLC were identified (March 1, 2013 to February 28, 2018). We compared their two-year and per-patient-per-month (PPPM) healthcare costs and resource utilization with 1:3 propensity score-matched patients without the condition. A generalized linear model was used to estimate the impact of BM and other covariates on healthcare costs. After propensity score matching with the 33â 402 newly diagnosed cases of stage IIIB or IV NSCLC, 3435 and 10â 305 patients were classified as having or not having BM, respectively. Mean healthcare costs were significantly greater in patients with BM for both the two years (US$ 44â 692 vs. US$ 32â 230, p < .0001) and PPPM (US$ 3510 vs. US$ 2573, p < .0001). The length of hospital stay was longer in patients with BM (79.15 vs. 69.41 days for two years, p < .0001; 7.69 vs. 6.86 days PPPM, p < .0001), and patients with BM had more outpatient visits (50.61 vs. 46.43 times for two years, p < .0001; 3.64 vs. 3.40 times PPPM costs, p < .0001). The costs of drugs, radiology/radiotherapy, and admission comprised the majority of PPPM costs and were higher in patients with BM. The generalized linear model analysis suggested that patients with BM had significantly increased healthcare costs (by 1.29-fold, 95% confidence interval 1.26-1.32). BM is a significant economic burden for patients with NSCLC. Therefore, it is important to prevent BM in patients with NSCLC to reduce their economic burden.
Assuntos
Neoplasias Encefálicas , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Neoplasias Encefálicas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Estresse Financeiro , Custos de Cuidados de Saúde , Recursos em Saúde , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Estudos RetrospectivosRESUMO
Effects of protein denaturation and formation of protein-sodium dodecyl sulfate (SDS) complexes on protein separation and identification were investigated using hollow fiber flow field-flow fractionation (HF5) and nanoflow liquid chromatography-electrospray ionization-tandem mass spectrometry (nLC-ESI-MS-MS). Denaturation and formation of protein-SDS complexes prior to HF5 separation resulted an increase in the retention of few protein standards due to unfolding of the protein structures and complexation, yielding ~30% increase in hydrodynamic diameter. In addition, low molecular weight proteins which could be lost from the HF membrane due to the pore size limitation showed an increase of peak recovery about 2-6 folds for cytochrome C and carbonic anhydrase. In the case of proteins composed of a number of subunits, denaturation resulted in a decrease in retention due to dissociation of protein subunits. A serum proteome sample, denatured with dithiothreitol and SDS, was fractionated by HF5, and the eluting protein fractions after tryptic digestion were analyzed for protein identification using nLC-ESI-MS-MS. The resulting pools of identified proteins were found to depend on whether the serum sample was treated with or without denaturation prior to the HF5 run due to differences in the aqueous solubility of the proteins. The enhancement of protein solubility by SDS also increased the number of identified membrane proteins (54 vs. 31).
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Fracionamento por Campo e Fluxo/métodos , Proteínas/isolamento & purificação , Dodecilsulfato de Sódio/química , Animais , Ditiotreitol/química , Humanos , Desnaturação Proteica , Proteínas/análise , Proteínas/química , Soro/química , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas em TandemRESUMO
In this study, an analytical method for the simultaneous separation and characterization of various molecular species of lysophospholipids (LPLs) and phospholipids (PLs) is introduced by employing nanoflow liquid chromatography-electrospray ionization tandem mass spectrometry (nLC-ESI-MS/MS). Since LPLs and PLs in human plasma are potential biomarkers for cancer, development of a sophisticated analytical method for the simultaneous profiling of these molecules is important. Standard species of LPLs and PLs were examined to establish a separation condition using a capillary LC column followed by MS scans and data-dependent collision-induced dissociation (CID) analysis for structural identification. With nLC-ESI-MS/MS, regioisomers of each category of LPLs were completely separated and identified with characteristic CID spectra. It was applied to the comprehensive profiling of LPLs and PLs from a human blood plasma sample and yielded identifications of 50 LPLs (each regioisomer pair of 6 lysophosphatidylcholines (LPCs), 7 lysophosphatidylethanolamines (LPEs), 9 lysophosphatidic acid (LPAs), 2 lysophosphatidylglycerols (LPGs), and 1 lysophosphatidylserine (LPS)) and 62 PLs (19 phosphatidylcholines (PCs), 11 phosphatidylethanolamines (PEs), 3 phosphatidylserines (PSs), 16 phosphatidylinositols (PIs), 8 phosphatidylglycerols (PGs), and 5 phosphatidic acids (PAs)).
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Cromatografia Líquida/métodos , Lisofosfolipídeos/sangue , Fosfolipídeos/sangue , Espectrometria de Massas em Tandem/métodos , Adulto , Feminino , Humanos , Lisofosfolipídeos/química , Fosfolipídeos/química , Espectrometria de Massas por Ionização por Electrospray/métodosRESUMO
Food adulteration is a serious issue that requires verification and strict management due to healthcare, morality, and social value problems. In the context of fat, food manufacturers blend lard with vegetable oils or animal fats for convenience and gaining economic benefits. Thus, we herein report the classification of 4 animal fats, e.g., beef tallow, pork lard, chicken fat, duck oil, using Raman spectroscopy combined with simple calculation of intensity ratios of Raman signal at vibrational modes corresponding to unsaturated fatty acids and total fatty acids. Various calculated values of the species were compared to find a feature that is able to classify each fats using Raman peak ratio. As a result, we suggested "Oil gauge (OG)" as a standard feature for classification of the fats in Raman analysis field. Furthermore, a quantification of the lard in other fat was accomplished with good linear correlation using the OG values.
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Gorduras na Dieta/análise , Gorduras/análise , Contaminação de Alimentos/análise , Análise Espectral Raman/métodos , Animais , Bovinos , Galinhas , Patos , Ácidos Graxos/análise , Ácidos Graxos Insaturados , Óleos/análise , SuínosRESUMO
Melicope ptelefolia has been traditionally used to treat rheumatism and fever. The present study aimed to investigate the therapeutic effect of 3,5diCßDglucopyranosyl phloroacetophenone (ßGP), a main component of M. ptelefolia, on rheumatoid arthritis (RA). A model of collageninduced arthritis (CIA) was established in mice using the RAW 264.7 murine macrophage cell line and mouse embryonic fibroblasts (MEFs). The clinical scores of arthritis, swelling, histopathological findings, and microcomputed tomography in CIA mouse paws were assessed. The levels of antitype II collagen antibody and cytokines were determined in the plasma and cell culture supernatant, respectively. Protein and gene expression levels were analyzed by western blot and reverse transcriptionquantitative polymerase chain reaction analyses. ßGP significantly decreased the gross arthritic scores of CIA mice and joint swelling, and decreased articular inflammation, cartilage degradation and bone erosion. However, ßGP did not exert any effect on antitype II collagen immunoglobulin G plasma levels or inflammatory cytokine expression in macrophages. ßGP significantly suppressed the expression of interleukin6 and leukemia inhibitory factor and decreased the phosphorylation of signal transducer and activator of transcription 3, and expression of receptor activator of nuclear factorκB ligand in tumor necrosis factorαstimulated MEFs and in CIA mouse paws. Osteoclastrelated gene expression was significantly reduced in CIA mouse paws. Taken together, ßGP suppressed the development of RA by regulating the activation of synovial fibroblasts.
Assuntos
Acetofenonas/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Artrite Experimental/tratamento farmacológico , Artrite Reumatoide/tratamento farmacológico , Fibroblastos/efeitos dos fármacos , Glucosídeos/uso terapêutico , Acetofenonas/química , Animais , Anti-Inflamatórios/química , Artrite Experimental/patologia , Artrite Reumatoide/patologia , Citocinas/análise , Fibroblastos/patologia , Glucosídeos/química , Masculino , Camundongos , Osteoclastos/efeitos dos fármacos , Osteoclastos/patologia , Células RAW 264.7 , Rutaceae/química , Microtomografia por Raio-XRESUMO
BACKGROUND: A few bioactivities of constituents from Gynostemma laxum, which has been collected in Vietnam, have been reported until now. There is no report about the effects of constituents from G. laxum although the nuclear factor (erythroid-derived 2)-like 2 (Nrf2)-mediated heme oxygenase-1 (HO-1) antioxidant defense system is involved in neuroprotection in the brain. Therefore, we investigated whether quercetin (2), benzoic acid (10) and their analogues (1, 3-9 and 11) from G. laxum have the antioxidant and neuroprotective activities and also their underlying mechanism. METHODS: To examine their neuroprotective and antioxidant activities, oxytosis, total oxidant scavenging capacity (TOSC), 2,7-dichlorofluorescein (DCFDA), dihydroethidium (DHE), antioxidant response element (ARE)-luciferase reporter gene assays, Western blot analysis, real time-PCR, immunocytochemistry and in silico 3D molecular docking simulation were performed. RESULTS: The study of constituents using chromatographic techniques and spectroscopic analysis showed that G. laxum contained an abundance of quercetin (2), benzoic acid (10) and their analogues (1, 3-9 and 11). Our data demonstrated that quercetin (2) and its analogue (4) among the constituents from G. laxum showed the strongest neuroprotective effect against oxytosis triggered by the excessive amount of glutamate. Compounds 2, 4, 6 and 11 exhibited reactive oxygen species (ROS) inhibitory and ARE transcriptional activities in immortalized hippocampal HT22 cell line. Among them, compound 4, a second active compound, induced Nrf2/HO-1 activation. They were also fit stable onto the Tramtrack and Bric-à-Brac (BTB) domain of Kelch-like ECH-associated protein 1 (Keap1), a known Nrf2 inhibitor protein, based on the results of docking and interaction energies. Overall, these data suggest that -OH and -OCH3 groups of quercetin and its analogues are responsible for their neuroprotective effect. CONCLUSIONS: In summary, the major constituents of G. laxum had strong antioxidant and neuroprotective activities so that they could consider as a natural antioxidant supplement. Furthermore, G. laxum might be used beneficially in reducing oxidative complications with the further deep investigation in vivo.