RESUMEN

Mass spectrometers are widely used to identify protein phosphorylation sites. The process usually involves selective isolation of phosphoproteins and subsequent fragmentation to identify both the peptide sequence and phosphorylation site. Immunoprecipitation could capture and purify the protein of interest, greatly reducing sample complexity before submitting it for mass spectrometry analysis. This chapter describes a method to identify an abnormal phosphorylated site of the adaptor protein by a viral kinase through immunoprecipitation followed by LC-MS/MS.

Asunto(s)

Inmunoprecipitación , Fosfoproteínas , Espectrometría de Masas en Tándem , Fosforilación , Espectrometría de Masas en Tándem/métodos , Inmunoprecipitación/métodos , Cromatografía Liquida/métodos , Humanos , Fosfoproteínas/metabolismo , Fosfoproteínas/análisis , Espectrometría de Masas/métodos

RESUMEN

BACKGROUND-AIM: Methylmalonic acid (MMA) is currently the best biomarker of functional vitamin B12 deficiency. However, for a correct interpretation of the patient's results it is necessary to know its biological variation (BV). No BV data are available for urine MMA values, as measured by mass spectrometry. Hence, the aim of this study was to estimate the within- and between-person coefficients of variation (CVw, CVg) for MMA in a healthy population, and the associated index of individuality (II), as well as to define quality specifications based on BV and the reference change value (RCV). METHODS: Random urine samples from 34 healthy volunteers were collected over four consecutive weeks. Samples were stored at -80 °C until analysis in a single analytical run. MMA excretion was quantified by tandem liquid chromatography coupled to mass spectrometry (HPLC-MS/MS). Results were normalized to urine creatinine. The coefficients of variation were estimated by CV-ANOVA. Confidence intervals (95 %) were calculated. Quality specifications were defined according to international recommendations. RESULTS: A total of 128 samples were included. The coefficients of variation were CVw = 35.7 % (26.1-45.3) and CVg = 67.7 % (58.3-77.0). The associated II was 0.5 and the RCV was 88.1 %. CONCLUSION: Considering the II obtained, MMA in urine has high individuality, therefore, RCV is better to evaluate serial clinical results. Our results will contribute to a better clinical interpretation of this biomarker and will represent a great aid when defining analytical performance specifications for this magnitude.

Asunto(s)

Ácido Metilmalónico , Humanos , Ácido Metilmalónico/orina , Masculino , Adulto , Femenino , España , Espectrometría de Masas en Tándem , Persona de Mediana Edad , Adulto Joven , Voluntarios Sanos , Cromatografía Líquida de Alta Presión , Biomarcadores/orina

RESUMEN

BACKGROUND AND AIMS: Current laboratory methods for opioid detection involve an initial screening with immunoassays which offers efficient but non-specific results and a subsequent liquid chromatography-tandem mass spectrometry (LC-MS/MS) confirmation which offers accurate results but requires extensive sample preparation and turnaround time. Direct Analysis in Real Time (DART) tandem mass spectrometry is evaluated as an alternative approach for accurate opioid detection with efficient sample preparation and turnaround time. MATERIALS AND METHODS: DART-MS/MS was optimized by testing the method with varying temperatures, operation modes, extraction methods, hydrolysis times, and vortex times. The method was evaluated for 12 opioids by testing the analytical measurement range, percent carryover, precision studies, stability, and method-to-method comparison with LC-MS/MS. RESULTS: DART-MS/MS shows high sensitivity and specificity for the detection of 6-acetylmorphine, codeine, hydromorphone, oxymorphone, hydrocodone, naloxone, buprenorphine, norfentanyl, and fentanyl in urine samples. However, its performance was suboptimal for norbuprenorphine, morphine and oxycodone. CONCLUSION: In this proof-of-concept study, DART-MS/MS is evaluated for its rapid quantitative definitive testing of opioids drugs in urine. Further research is needed to expand its application to other areas of drug testing.

Asunto(s)

Analgésicos Opioides , Espectrometría de Masas en Tándem , Humanos , Espectrometría de Masas en Tándem/métodos , Analgésicos Opioides/orina , Cromatografía Liquida/métodos , Factores de Tiempo

RESUMEN

ETHNOPHARMACOLOGICAL RELEVANCE: In accordance with the tenets of traditional Chinese medicine, sepsis is categorized into three distinct syndromes: heat syndrome, blood stasis syndrome, and deficiency syndrome. Xiaochaihu decoction (XCHD) has many functions, including the capacity to protect the liver, cholagogue, antipyretic, anti-inflammatory, and anti-pathogenic microorganisms. XCHD exerts the effect of clearing heat and reconciling Shaoyang. The XCHD contains many efficacious active ingredients, yet the mechanism of sepsis-induced cardiomyopathy (SIC) remains elusive. AIM OF THE STUDY: To investigate the molecular mechanisms underlying the protective effects of XCHD against SIC using an integrated approach combining network pharmacology and molecular biology techniques. MATERIALS AND METHODS: Network pharmacology methods identified the active ingredients, target proteins, and pathways affected by XCHD in the context of SIC. We conducted in vivo experiments using mice with lipopolysaccharide-induced SIC, evaluating cardiac function through echocardiography and histology. XCHD-containing serum was analyzed to determine its principal active components using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The effects of XCHD-containing serum on SIC were further tested in vitro in LPS-treated H9c2 cardiac cells. Protein expression levels were quantified via Western blotting and enzyme-linked immunosorbent assay (ELISA). Additionally, molecular docking was performed between the active components and ZBP1, a potential target protein. Overexpression of ZBP1 in H9c2 cells allowed for a deeper exploration of its role in modulating SIC-associated gene expression. RESULTS: UPLC-MS/MS identified 31 shared XCHD and XCHD-containing serum components. These included organic acids, terpenoids, and flavonoids, which have been identified as the active components of XCHD. Our findings revealed that XCHD alleviated LPS-induced myocardial injury, improved cardiac function, and preserved cardiomyocyte morphology in mice. In vitro studies, we demonstrated that XCHD-containing serum significantly suppressed the expression of inflammatory cytokines (IL-6, IL-1ß, and TNF-α) in LPS-induced H9c2 cells. Mechanistic investigations showed that XCHD downregulated genes associated with PANoptosis, a novel cell death pathway, suggesting its protective role in sepsis-damaged hearts. Conversely, overexpression of ZBP1 abolished the protective effects of XCHD and amplified PANoptosis-related gene expression. CONCLUSIONS: Our study provides the first evidence supporting the protective effects of XCHD against SIC, both in vitro and in vivo. The underlying mechanism involves the inhibition of ZBP1-initiated PANoptosis, offering new insights into treating SIC using XCHD.

Asunto(s)

Cardiomiopatías , Medicamentos Herbarios Chinos , Sepsis , Animales , Medicamentos Herbarios Chinos/farmacología , Sepsis/tratamiento farmacológico , Sepsis/complicaciones , Cardiomiopatías/tratamiento farmacológico , Cardiomiopatías/metabolismo , Ratones , Masculino , Línea Celular , Ratones Endogámicos C57BL , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Lipopolisacáridos/toxicidad , Farmacología en Red , Ratas , Modelos Animales de Enfermedad , Espectrometría de Masas en Tándem

RESUMEN

ETHNOPHARMACOLOGICAL RELEVANCE: Modified Danzhi Xiaoyao San (MDXS) is an effective clinical prescription for depression in China, which was deprived of Danzhi Xiaoyao San in the Ming Dynasty. MDSX has significant implications for the development of new antidepressants, but its pharmacological mechanism has been rarely studied. AIM OF THE STUDY: To reveal the active components and molecular mechanism of MDXS in treating depression through network pharmacology and experimental verification in vivo and in vitro. MATERIALS AND METHODS: UPLC-Q-TOF-MS/MS was used to identify the chemical components in the MDXS freeze-dried powder, drug-containing serum, and cerebrospinal fluid (CSF). Based on the analysis of prototype components in the CSF, the major constituents, potential therapeutic targets and possible pharmacological mechanisms of MDXS in treating depression were investigated using network pharmacological and molecular docking. Then corticosterone (CORT)-induced mice model of depression was established to investigate the antidepressant effects of MDXS. HT22 cells were cultured to verify the neuroprotective effects and core targets of the active components. RESULTS: There were 81 compounds in MDXS freeze-dried powder, 36 prototype components in serum, and 13 prototype components in CSF were identified, respectively. Network pharmacology analysis showed that these 13 prototype components in the CSF shared 190 common targets with depression, which were mainly enriched in MAPK and PI3K/AKT signaling pathways. PPI analysis suggested that AKT1 and MAPK1 (ERK1/2) were the core targets. Molecular docking revealed that azelaic acid (AA), senkyunolide A (SA), atractylenolide III (ATIII), and tokinolide B (TB) had the highest binding energy with AKT1 and MAPK1. Animal experiments verified that MDXS could reverse CORT-induced depression-like behaviors, improve synaptic plasticity, alleviate neuronal injury in hippocampal CA3 regions, and up-regulate the protein expression of p-ERK1/2 and p-AKT. In HT22 cells, azelaic acid, senkyunolide A, and atractylenolide III significantly protected the cell injury caused by CORT, and up-regulated the protein levels of p-ERK1/2 and p-AKT. CONCLUSIONS: These results suggested that MDXS may exert antidepressant effects partially through azelaic acid, senkyunolide A, and atractylenolide III targeting ERK1/2 and AKT.

Asunto(s)

Antidepresivos , Depresión , Medicamentos Herbarios Chinos , Simulación del Acoplamiento Molecular , Farmacología en Red , Animales , Antidepresivos/farmacología , Depresión/tratamiento farmacológico , Medicamentos Herbarios Chinos/farmacología , Ratones , Masculino , Línea Celular , Modelos Animales de Enfermedad , Ratones Endogámicos C57BL , Corticosterona/sangre , Espectrometría de Masas en Tándem , Conducta Animal/efectos de los fármacos

RESUMEN

Perilla leaf oil (PLO) is a global premium vegetable oil with abundant nutrients and substantial economic value, rendering it susceptible to potential adulteration by unscrupulous entrepreneurs. The addition of cinnamon oil (CO) is one of the main adulteration avenues for illegal PLOs. In this study, new and real-time ambient mass spectrometric methods were developed to detect CO adulteration in PLO. First, atmospheric solids analysis probe tandem mass spectrometry combined with principal component analysis and principal component analysis-linear discriminant analysis was employed to differentiate between authentic and adulterated PLO. Then, a spectral library was established for the instantaneous matching of cinnamaldehyde in the samples. Finally, the results were verified using the SRM mode of ASAP-MS/MS. Within 3 min, the three methods successfully identified CO adulteration in PLO at concentrations as low as 5% v/v with 100% accuracy. The proposed strategy was successfully applied to the fraud detection of CO in PLO.

Asunto(s)

Cinnamomum zeylanicum , Contaminación de Alimentos , Hojas de la Planta , Aceites de Plantas , Contaminación de Alimentos/análisis , Aceites de Plantas/química , Aceites de Plantas/análisis , Hojas de la Planta/química , Cinnamomum zeylanicum/química , Perilla/química , Espectrometría de Masas en Tándem/métodos , Espectrometría de Masas/métodos

RESUMEN

To improve the adsorption affinity and selectivity of fipronils (FPNs), including fipronil, its metabolites and analogs, a magnetic covalent organic framework (Fe3O4@COF-F) with copious fluorine affinity sites was innovatively designed as an adsorbent of magnetic solid-phase extraction (MSPE). The enhanced surface area, pore size, crystallinity of Fe3O4@COF-F and its exponential adsorption capacities (187.3-231.5 mg g-1) towards fipronils were investigated. Combining MSPE with high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), an analytical method was established for the selective determination of fipronils in milk and milk powder samples. This method achieved high sensitivity (LODs: 0.004-0.075 ng g-1), satisfactory repeatability and accuracy with spiked recoveries ranging from 89.9% to 100.3% (RSDs≤5.1%). Overall, the constructed Fe3O4@COF-F displayed great potential for the selective enrichment of fipronils, which could be ascribed to fluorine­fluorine interaction. This method proposed a feasible and promising strategy for the development of functionalized COF and broadened its application in fluorine containing hazards detection.

Asunto(s)

Flúor , Contaminación de Alimentos , Estructuras Metalorgánicas , Leche , Pirazoles , Extracción en Fase Sólida , Espectrometría de Masas en Tándem , Pirazoles/química , Contaminación de Alimentos/análisis , Flúor/química , Leche/química , Animales , Estructuras Metalorgánicas/química , Adsorción , Cromatografía Líquida de Alta Presión , Insecticidas/química , Insecticidas/análisis , Límite de Detección

RESUMEN

The use of direct injection ion mobility mass spectrometry (DI-IM-MS) to detect and identify betacyanin pigments in A. hortensis 'rubra' extracts was explored for the first time, with results compared to conventional LC-MS/MS analysis. The anti-inflammatory activities of leaf and seed extracts, alongside purified amaranthin and celosianin pigments, were investigated using a model of lipopolysaccharide (LPS)-activated murine macrophages. Extracts and purified pigments significantly inhibited the production of prostaglandin E2 and NO by up to 90% and 70%, respectively, and reduced the expression of Il6, Il1b, Nos2, and Cox2. Leaf and seed extracts also decreased secretion of Il6 and Il1b cytokines and reduced protein levels of Nos2 and Cox2. Furthermore, extracts and purified pigments demonstrated potent dose-dependent radical scavenging activity in a cellular antioxidant activity assay (CAA) without any cytotoxic effects. Our research highlights the promising biological potential of edible, climate-resilient A. hortensis 'rubra' as a valuable source of bioactive compounds.

Asunto(s)

Lipopolisacáridos , Macrófagos , Estrés Oxidativo , Extractos Vegetales , Ratones , Animales , Extractos Vegetales/farmacología , Extractos Vegetales/química , Extractos Vegetales/aislamiento & purificación , Células RAW 264.7 , Estrés Oxidativo/efectos de los fármacos , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Lipopolisacáridos/farmacología , Antiinflamatorios/farmacología , Antiinflamatorios/química , Antiinflamatorios/aislamiento & purificación , Ciclooxigenasa 2/genética , Ciclooxigenasa 2/inmunología , Ciclooxigenasa 2/metabolismo , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Espectrometría de Masas en Tándem

RESUMEN

This study presents the contents of α-methylenecyclopropylglycine, a potentially toxic amino acid, in the peel, pulp and seed fractions of two well-known litchi varieties, namely Shahi and China, over a span of three harvest-seasons. For analysing α-methylenecyclopropylglycine, an LC-MS/MS-based method was validated. The method-accuracies fell within 75-110 % (RSD, <15 %) at 0.1 mg/kg (LOQ) and higher levels. A comparative evaluation of the results in peel, pulp and seed at 30 days before harvest (DBH), 15-DBH, and edible-ripe stage revealed that α-methylenecyclopropylglycine content increased as the litchi seeds grew towards maturity, regardless of the cultivar. In arils, at maturity, the concentration of α-methylenecyclopropylglycine ranged from not-detected to 11.7 µg/g dry weight. The Shahi cultivar showed slightly higher α-methylenecyclopropylglycine content in comparison to China litchi. This paper presents the first known analysis of combined seasonal data on different fruit components at various growth stages for the two chosen litchi cultivars grown in India.

Asunto(s)

Frutas , Litchi , Semillas , Espectrometría de Masas en Tándem , Litchi/química , Litchi/crecimiento & desarrollo , Litchi/metabolismo , Frutas/química , Frutas/crecimiento & desarrollo , China , Semillas/química , Semillas/crecimiento & desarrollo , Glicina/análogos & derivados , Glicina/análisis , Cromatografía Líquida de Alta Presión , Ciclopropanos/análisis

RESUMEN

Fortification of human milk (HM) is often necessary to meet the nutritional requirements of preterm infants. The present experiment aimed to establish whether the supplementation of HM with either an experimental donkey milk-derived fortifier containing whole donkey milk proteins, or with a commercial bovine milk-derived fortifier containing hydrolyzed bovine whey proteins, affects peptide release differently during digestion. The experiment was conducted using an in vitro dynamic system designed to simulate the preterm infant's digestion followed by digesta analysis by means of LC-MS-MS. The different fortifiers did not appear to influence the cumulative intensity of HM peptides. Fortification had a differential impact on the release of either donkey or bovine bioactive peptides. Donkey milk peptides showed antioxidant/ACE inhibitory activities, while bovine peptides showed opioid, dipeptil- and propyl endo- peptidase inhibitory and antimicrobial activity. A slight delay in peptide release from human lactoferrin and α-lactalbumin was observed when HM was supplemented with donkey milk-derived fortifier.

Asunto(s)

Digestión , Equidae , Proteínas de la Leche , Leche Humana , Péptidos , Humanos , Animales , Leche Humana/química , Leche Humana/metabolismo , Proteínas de la Leche/química , Proteínas de la Leche/metabolismo , Proteínas de la Leche/análisis , Bovinos , Péptidos/química , Péptidos/metabolismo , Alimentos Fortificados/análisis , Espectrometría de Masas en Tándem , Modelos Biológicos , Proteína de Suero de Leche/química , Proteína de Suero de Leche/metabolismo

RESUMEN

A modified QuEChERS method was developed to determine multi-class pesticide and veterinary residues in aquatic products. Chitosan microspheres were conveniently synthesized and utilized as the cleanup adsorbent in the QuEChERS procedure, showcasing rapid filtration one-step pretreatment ability for the determination of drug multi-residues in aquatic products. Compared to conventional synthetic sorbents, chitosan microspheres not only have good purification performance, but also have renewable and degradable properties. This novel sorbent worked well in the simultaneous determination of 95 pesticides and veterinary drug residues in aquatic products after being combined with an improved one-step vortex oscillating cleanup method. We achieved recoveries ranging from 64.0% to 115.9% for target drugs in shrimp and fish matrix. The limits of detection and quantification were 0.5-1.0 and 1.0-2.0 µg kg-1, respectively. Notably, hydrocortisone was detected with considerable frequency and concentration in the tested samples, underscoring the necessity for stringent monitoring of this compound in aquatic products.

Asunto(s)

Quitosano , Peces , Microesferas , Espectrometría de Masas en Tándem , Drogas Veterinarias , Animales , Quitosano/química , Cromatografía Líquida de Alta Presión , Drogas Veterinarias/análisis , Drogas Veterinarias/aislamiento & purificación , Contaminación de Alimentos/análisis , Residuos de Medicamentos/análisis , Residuos de Medicamentos/aislamiento & purificación , Residuos de Medicamentos/química , Plaguicidas/aislamiento & purificación , Plaguicidas/análisis , Plaguicidas/química , Residuos de Plaguicidas/aislamiento & purificación , Residuos de Plaguicidas/análisis , Residuos de Plaguicidas/química , Adsorción , Extracción en Fase Sólida/métodos , Extracción en Fase Sólida/instrumentación , Contaminantes Químicos del Agua/aislamiento & purificación , Contaminantes Químicos del Agua/química , Contaminantes Químicos del Agua/análisis , Alimentos Marinos/análisis , Mariscos/análisis , Cromatografía Líquida con Espectrometría de Masas

RESUMEN

Aromatic amino acid oxidation products (AAAOPs) are newly discovered risk substances of thermal processes. Due to its significant polarity and trace level in food matrices, there are no efficient pre-treatment methods available to enrich AAAOPs. Herein, we proposed a magnetic cationic covalent organic framework (Fe3O4@EB-iCOF) as an adsorbent for dispersive magnetic solid-phase extraction (DMSPE). Benefiting from the unique charged characteristics of Fe3O4@EB-iCOF, AAAOPs can be enriched through electrostatic interaction and π-π interactions. Under the optimal DMSPE conditions, the combined HPLC-MS/MS method demonstrated good linearity (R2 ≥ 0.990) and a low detection limit (0.11-7.5 µg·kg-1) for AAAOPs. In addition, the method was applied to real sample and obtained satisfactory recoveries (86.8 % âˆ¼ 109.9 %). Especially, we applied this method to the detection of AAAOPs in meat samples and conducted a preliminarily study on its formation rules, which provides a reliable basis for assessing potential dietary risks.

Asunto(s)

Aminoácidos Aromáticos , Oxidación-Reducción , Extracción en Fase Sólida , Extracción en Fase Sólida/métodos , Aminoácidos Aromáticos/química , Aminoácidos Aromáticos/análisis , Aminoácidos Aromáticos/aislamiento & purificación , Espectrometría de Masas en Tándem , Estructuras Metalorgánicas/química , Calor , Contaminación de Alimentos/análisis , Cromatografía Líquida de Alta Presión , Animales , Adsorción , Carne/análisis , Alimentos Procesados

RESUMEN

Dried citrus peel (DCP), also called "Chen Pi", has edible and medicinal value. However, the specific differences among various sources remain unknown. Herein, we collected six DCP species, namely, one Citrus reticulata 'Chachi' (CZG) and five Citrus reticulata Blanco (CRB). Targeted high-performance liquid chromatography and untargeted ultra-high-performance liquid chromatography-tandem mass spectrometry were employed to comprehensively compare the phenolic compounds and metabolites in DCP. Interestingly, 13 different phenolic compounds were noted in DCP. The total phenolic compound content in all CRB samples (58.86-127.65 mg/g) was higher than that of CZG (39.47 mg/g). Untargeted metabolomic revealed 1495 compounds, with 115 differentially expressed metabolites for CRBs and CZG, particularly flavonoids (38), terpenoids (15), and phenolic acids and derivatives (9). Lastly, antioxidant assays revealed that all CRB samples exhibited higher antioxidant activities compared with CZG. Therefore, our study results provide a theoretical basis for the high-value utilization of citrus peels and their metabolites.

Asunto(s)

Antioxidantes , Citrus , Frutas , Metabolómica , Extractos Vegetales , Espectrometría de Masas en Tándem , Citrus/química , Citrus/metabolismo , Antioxidantes/química , Antioxidantes/metabolismo , Antioxidantes/análisis , Cromatografía Líquida de Alta Presión , Frutas/química , Frutas/metabolismo , Extractos Vegetales/química , Extractos Vegetales/metabolismo , Fenoles/metabolismo , Fenoles/química , Fenoles/análisis , Flavonoides/metabolismo , Flavonoides/química , Flavonoides/análisis

RESUMEN

Toxic cyanobacterial blooms impose a health risk to recreational users, and monitoring of cyanobacteria and associated toxins is required to assess this risk. Traditionally, monitoring for risk assessment is based on cyanobacterial biomass, which assumes that all cyanobacteria potentially produce toxins. While these methods may be cost effective, relatively fast, and more widely accessible, they often lead to an overestimation of the health risk induced by cyanotoxins. Monitoring methods that more directly target toxins, or toxin producing genes, may provide a better risk assessment, yet these methods may be more costly, usually take longer, or are not widely accessible. In this study, we compared six monitoring methods (fluorometry, microscopy, qPCR of 16S and mcyE, ELISA assays, and LC-MS/MS), of which the last three focussed on the most abundant cyanotoxin microcystins, across 11 lakes in the Netherlands during the bathing water season (May-October) of 2019. Results of all monitoring methods significantly correlated with LC-MS/MS obtained microcystin levels (the assumed 'golden standard'), with stronger correlations for methods targeting microcystins (ELISA) and microcystin genes (mcyE). The estimated risk levels differed substantially between methods, with 78 % and 56 % of alert level exceedances in the total number of collected samples for fluorometry and microscopy-based methods, respectively, while this was only 16 % and 6 % when the risk assessment was based on ELISA and LC-MS/MS obtained toxin concentrations, respectively. Integrating our results with earlier findings confirmed a strong association between microcystin concentration and the biovolume of potential microcystin-producing genera. Moreover, using an extended database consisting of 4265 observations from 461 locations across the Netherlands in the bathing water seasons of 2015 - 2019, we showed a strong association between fluorescence and the biovolume of potentially toxin-producing genera. Our results indicate that a two-tiered approach may be an effective risk assessment strategy, with first a biomass-based method (fluorometry, biovolume) until the first alert level is exceeded, after which the risk level can be confirmed or adjusted based on follow-up toxin or toxin gene analyses.

Asunto(s)

Cianobacterias , Monitoreo del Ambiente , Floraciones de Algas Nocivas , Lagos , Microcistinas , Medición de Riesgo , Monitoreo del Ambiente/métodos , Microcistinas/análisis , Lagos/microbiología , Lagos/química , Países Bajos , Espectrometría de Masas en Tándem/métodos , Cromatografía Liquida , Ensayo de Inmunoadsorción Enzimática

RESUMEN

To meet the needs of developing efficient extractive materials alongside the evolution of miniaturized sorbent-based sample preparation techniques, a mesoporous structure of g-C3N4 doped with sulfur as a heteroatom was achieved utilizing a bubble template approach while avoiding the severe conditions of other methods. In an effort to increase the number of adsorption sites, the resultant exfoliated structure was then modified with thymol-coumarin NADES as a natural sorbent modifier, followed by introduction into a nylon 6 polymer via an electrospinning process. X-ray diffraction (XRD), Fourier transform infrared (FT-IR) spectroscopy, field-emission scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), and Brunauer-Emmett-Teller (BET) surface area analysis validated S-doped g-C3N4 and composite production. The prepared electrospun fiber nanocomposite, entailing satisfactory processability, was then successfully utilized as a sorbent in on-chip thin film micro-solid-phase extraction of non-steroidal anti-inflammatory drugs (NSAIDs) from saliva samples prior to liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Utilizing a chip device, a thin film µ-SPE coupled with LC-MS/MS analysis yielded promising outcomes with reduced sample solution and organic solvents while extending lifetime of a thin film sorbent. The DES-modified S-doped g-C3N4 amount in electrospun was optimized, along with adsorption and desorption variables. Under optimal conditions, selected NSAIDs were found to have a linear range of 0.05-100.0 ng mL-1 with an R2 ≥ 0.997. The detection limits were ranged between 0.02 and 0.2 ng mL-1. The intra-day and inter-day precisions obtained were less than 6.0%. Relative recoveries were between 93.3 and 111.4%.

Asunto(s)

Antiinflamatorios no Esteroideos , Disolventes Eutécticos Profundos , Grafito , Límite de Detección , Nanofibras , Saliva , Espectrometría de Masas en Tándem , Saliva/química , Espectrometría de Masas en Tándem/métodos , Grafito/química , Nanofibras/química , Humanos , Adsorción , Antiinflamatorios no Esteroideos/análisis , Porosidad , Disolventes Eutécticos Profundos/química , Cromatografía Liquida/métodos , Compuestos de Nitrógeno/química , Microextracción en Fase Sólida/métodos , Extracción en Fase Sólida/métodos

RESUMEN

In the mirabegron (MIR) synthesis, the N-nitroso mirabegron (NNM) is obtained during synthetic process of MIR; water is being used in reaction under acidic condition. Nitrite source is from water, and secondary amine source is from MIR as it has secondary amine; NNM is generated as an impurity during the synthesis of MIR. The presence of NNM in MIR could potentially affect its effectiveness. The purpose of this study was to establish a Ultra-performance liquid chromatography-mass spectrometry/mass spectrometry (UPLC-MS/MS) methodology to identify NNM in MIR samples. The method for NNM analysis was developed on Acquity HSS T3 (100*2.1) mm 1.8 µm column with gradient elution using mobile phase consisted of 0.1% formic acid in water (mobile phase A) and 0.1% formic acid in methanol (mobile phase B). Mass spectrometer with electrospray ionization operated in the MRM mode was used in the analysis of NNM (m/ z 426.20 → 170.00). The UPLC-MS/MS methodology proposed showed a good linearity (0.02 to 0.72 ppm), good system precision (RSD = 0.57%), good method precision (RSD = 0.87%), acceptable accuracy (94.5-116.5%), low detection limit (0.006 ppm) and low quantification limit (0.02 ppm) for NNM. The UPLC-MS/MS methodology proposed can be utilized to assess the quality of MIR sample for the presence of NNM impurity.

Asunto(s)

Acetanilidas , Espectrometría de Masas en Tándem , Tiazoles , Espectrometría de Masas en Tándem/métodos , Acetanilidas/análisis , Acetanilidas/química , Cromatografía Líquida de Alta Presión/métodos , Tiazoles/análisis , Tiazoles/química , Reproducibilidad de los Resultados , Límite de Detección , Modelos Lineales , Contaminación de Medicamentos , Compuestos Nitrosos/análisis , Compuestos Nitrosos/química , Cromatografía Líquida con Espectrometría de Masas

RESUMEN

Ripened pu-erh tea is known to have beneficial hypoglycemic properties. However, it remains unclear whether the bioactive peptides produced during fermentation are also related to hypoglycemic potential. This study aimed to identify hypoglycemic peptides in ripened pu-erh tea and to elucidate their bioactive mechanisms using physicochemical property prediction, molecular docking, molecular dynamics simulations, and cell experiments. Thirteen peptides were identified by liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). Among them, AADTDYRFS (AS-9) and AGDGTPYVR (AR-9) exhibited high α-glucosidase inhibitory activity, with half-maximal inhibitory concentration (IC50) values of 0.820 and 3.942 mg/mL, respectively. Molecular docking and dynamics simulations revealed that hydrogen bonding, hydrophobic interactions, and van der Waals forces assist peptides AS-9 and AR-9 in forming stable and tight complexes with α-glucosidase. An insulin-resistance (IR)-HepG2 cell model was established. AS-9 was non-toxic to IR-HepG2 cells and significantly increased the glucose consumption capacity, hexokinase, and pyruvate kinase activities of IR-HepG2 cells (p < 0.05). AS-9 alleviated glucose metabolism disorders and ameliorated IR by activating the IRS-1/PI3K/Akt signaling pathway and increasing the expression levels of MDM2, IRS-1, Akt, PI3K, GLUT4, and GSK3ß genes. In addition, no hemolysis of mice red blood cells red blood cells occurred at concentrations below 1 mg/mL. This work first explored hypoglycemic peptides in ripened pu-erh tea, providing novel insights for enhancing its functional value.

Asunto(s)

Inhibidores de Glicósido Hidrolasas , Hipoglucemiantes , Simulación del Acoplamiento Molecular , Péptidos , Té , Hipoglucemiantes/farmacología , Hipoglucemiantes/química , Animales , Té/química , Humanos , Células Hep G2 , Péptidos/química , Péptidos/farmacología , Inhibidores de Glicósido Hidrolasas/farmacología , Inhibidores de Glicósido Hidrolasas/química , Ratones , Simulación de Dinámica Molecular , Resistencia a la Insulina , Transducción de Señal/efectos de los fármacos , Proteínas Sustrato del Receptor de Insulina/metabolismo , Espectrometría de Masas en Tándem , alfa-Glucosidasas/metabolismo , Fermentación

RESUMEN

The fate of Alternaria toxin tenuazonic acid (TeA) during the processing chain of wheat flour products was systemically evaluated. TeA was analyzed by liquid chromatography-mass spectrometry (LC-MS/MS) in wheat grains and the corresponding wheat flour products produced throughout the whole chain. The results indicated that TeA contamination in wheat grains largely determines the level of TeA toxin present in byproducts, semi-finished products, and finished products of the processing of four types of simulated processed wheat flour products (e.g., dry noodles, steamed breads, baked breads, and biscuits). The different food processing techniques had different effects on the fate of TeA. Wheat flour processing can reduce the TeA content in wheat grains by 58.7-83.2 %, indicating that wheat flour processing is a key step in reducing the TeA content in the food chain. Among the four types of wheat flour products, the decreases in TeA content in biscuits (69.8-76.7 %) were greater than those in dry noodles (15.5-22.3 %) and steamed breads (24.9-43.6 %). In addition, the decreasing effect of TeA was especially obvious in the wheat flour product chain with a high level of contamination. The processing factors (PFs) for TeA were as low as 0.20 for the four wheat processing methods and as high as 1.24 for the dry noodle processing method. At the average and 95th percentiles, dietary exposure to TeA in Chinese consumers including infants and young children did not exceed the relevant threshold value of toxicological concern (TTC) of TeA (1.5 µg/kg body weight per day), indicating an acceptable health risk for Chinese consumers via wheat flour products. These findings provide new insight into the fate of TeA in the food chain and mycotoxin control on the safety of wheat flour products and public health.

Asunto(s)

Alternaria , Harina , Contaminación de Alimentos , Manipulación de Alimentos , Espectrometría de Masas en Tándem , Ácido Tenuazónico , Triticum , Ácido Tenuazónico/análisis , Harina/análisis , Triticum/química , Contaminación de Alimentos/prevención & control , Contaminación de Alimentos/análisis , Manipulación de Alimentos/métodos , Micotoxinas/análisis , Humanos , Cromatografía Liquida , Pan/análisis

RESUMEN

Donepezil (DPZ), a piperidine-based reversible cholinesterase inhibitor, finds extensive use in treating Alzheimer's disease (AD). Originally designed as an oral formulation, DPZ encounters drawbacks such as a brief duration of action and reduced treatment effectiveness in elderly patients with memory impairment or difficulty swallowing medications. To address these issues and improve patient compliance, researchers are actively exploring alternative DPZ formulations. Consequently, reliable methods are necessary to quantitate DPZ in biological samples for in vivo assessment. Therefore, we propose an efficient, sensitive, wide-dynamic, and cost-effective method for quantitating DPZ in rat plasma. Our method employs liquid-liquid extraction (LLE) followed by liquid chromatography and tandem mass spectrometry, enabling in vivo evaluation of novel DPZ formulations. Notably, our method requires only 20 µL of rat plasma and employs icopezil as the internal standard-a cost-effective compound with chemical similarity to DPZ. We meticulously optimized LLE conditions, taking into account factor interactions through design of experiments (DOE). Our rapid and straightforward extraction and purification involved using 500 µL of pure methyl tert-butyl ether to extract DPZ from the sample within five minutes. The dynamic range of the method extends from 0.5 ng/mL to 1,000 ng/mL, demonstrating excellent sensitivity and suitability for pharmacokinetic studies across diverse DPZ formulations. Following the FDA guidelines, we rigorously validated the developed method, evaluating selectivity, linearity (with a coefficient of determination ≥0.9999), accuracy (ranging from 96.0% to 109.6%), precision (≤13.9%), matrix effect (92.2% to 103.8%), recovery (98.5% to 106.8%), the lower limit of quantitation (0.5 ng/mL), and stability. Finally, we effectively employed the validated method for the long-term pharmacokinetic assessment of a DPZ formulation. We expect that this approach will make a substantial contribution to the advancement of new DPZ formulations, ultimately benefiting individuals afflicted by AD.

Asunto(s)

Donepezilo , Extracción Líquido-Líquido , Piperidinas , Espectrometría de Masas en Tándem , Donepezilo/sangre , Donepezilo/farmacocinética , Animales , Espectrometría de Masas en Tándem/métodos , Extracción Líquido-Líquido/métodos , Ratas , Cromatografía Liquida/métodos , Piperidinas/sangre , Piperidinas/farmacocinética , Piperidinas/química , Inhibidores de la Colinesterasa/sangre , Inhibidores de la Colinesterasa/farmacocinética , Indanos/sangre , Indanos/farmacocinética , Masculino , Reproducibilidad de los Resultados , Ratas Sprague-Dawley , Cromatografía Líquida con Espectrometría de Masas

RESUMEN

Understanding the human milk metabolome can help inform infant nutrition and health. Untargeted metabolomics was used to study breast milk from 31 healthy participants to assess the shared metabolites in milk from participants with various backgrounds and understand how different demographic, health, and environmental factors impact the milk metabolome. Breast milk samples were analyzed by four separate UPLC-MS/MS methods. Metabolite Set Enrichment Analysis was used to study the most and least variable metabolites. The associations between participant factors and the metabolome were assessed with redundancy analyses. Among all 31 participants and between each untargeted UPLC-MS/MS method, 731 metabolites were detected, of which 389 were shared among all participants. Of the shared metabolites, lactose was the least and lactobionate the most variable metabolite. In the biological super pathway analysis, xenobiotics were the most variable metabolites. Infant age, maternal age, number of live births, and pre-pregnancy BMI were associated with the milk metabolome. In conclusion, the most variable metabolites originate from environmental exposures while the well-conserved core metabolites are linked to cell metabolism or are crucial for infant nutrition and osmoregulation. Understanding the variability of the breast milk metabolome can help identify components that are crucial for infant nutrition, growth, and development.

Asunto(s)

Metaboloma , Metabolómica , Leche Humana , Humanos , Leche Humana/metabolismo , Leche Humana/química , Femenino , Metabolómica/métodos , Adulto , Madres , Espectrometría de Masas en Tándem , Lactante , Adulto Joven , Lactosa/metabolismo , Lactosa/análisis