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Proposed mechanisms for the production of calcium in the first stars (population III stars)-primordial stars that formed out of the matter of the Big Bang-are at odds with observations1. Advanced nuclear burning and supernovae were thought to be the dominant source of the calcium production seen in all stars2. Here we suggest a qualitatively different path to calcium production through breakout from the 'warm' carbon-nitrogen-oxygen (CNO) cycle through a direct experimental measurement of the 19F(p, γ)20Ne breakout reaction down to a very low energy point of 186 kiloelectronvolts, reporting a key resonance at 225 kiloelectronvolts. In the domain of astrophysical interest2, at around 0.1 gigakelvin, this thermonuclear 19F(p, γ)20Ne rate is up to a factor of 7.4 larger than the previous recommended rate3. Our stellar models show a stronger breakout during stellar hydrogen burning than previously thought1,4,5, and may reveal the nature of calcium production in population III stars imprinted on the oldest known ultra-iron-poor star, SMSS0313-67086. Our experimental result was obtained in the China JinPing Underground Laboratory7, which offers an environment with an extremely low cosmic-ray-induced background8. Our rate showcases the effect that faint population III star supernovae can have on the nucleosynthesis observed in the oldest known stars and first galaxies, which are key mission targets of the James Webb Space Telescope9.
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Osimertinib is a third-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) for lung adenocarcinoma (LUAD) harboring activating mutations, but patients ultimately develop acquired resistance. Circular RNAs are involved in EGFR-TKI resistance, while the role of hsa_circ_0005576 in the osimertinib resistance of LUAD remains unknown. In this study, we demonstrated that hsa_circ_0005576 could facilitate osimertinib-resistant LUAD cells. Briefly, knockdown of hsa_circ_0005576 not only suppressed the proliferation and promoted the apoptosis of resistant LUAD cells, but also increased their sensitivity to osimertinib. Mechanistically, hsa_circ_0005576, serving as an miRNA sponge, could directly interact with miR-512-5p and subsequently upregulate the miR-512-5p-targeted insulin-like growth factor 1 receptor. Rescue assays indicated that miR-512-5p inhibition could reverse the effects of hsa_circ_0005576 knockdown in LUAD cells resistant to osimertinib. Overall, our study revealed that hsa_circ_0005576 regulates proliferation and apoptosis through miR-512-5p/IGF1R signaling, which contributes further to the resistance of LUAD cells to osimertinib. In addition, this study provides a novel insight into the mechanisms underlying osimertinib resistance of LUAD.
Assuntos
Adenocarcinoma de Pulmão/patologia , Resistencia a Medicamentos Antineoplásicos , Neoplasias Pulmonares/patologia , MicroRNAs/genética , RNA Circular/genética , Receptor IGF Tipo 1/genética , Acrilamidas/farmacologia , Acrilamidas/uso terapêutico , Adenocarcinoma de Pulmão/tratamento farmacológico , Adenocarcinoma de Pulmão/genética , Compostos de Anilina/farmacologia , Compostos de Anilina/uso terapêutico , Animais , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Camundongos , Transplante de Neoplasias , Regulação para CimaRESUMO
BACKGROUND: Emulsification is important for food quality and processing functionality. Most emulsification occurs under high-fat conditions that eventually cause health concerns. Protein emulsifiers also have drawbacks such as lower dispersity. This study considered the effects of different high-speed shear homogenization (HSH) speeds on the emulsifying and structural properties of myofibrillar proteins (MPs) under low-fat conditions. RESULTS: High-speed shear homogenization significantly increased the emulsifying activity and emulsifying stability of MPs at lower speeds (8000 to 14 500 rpm). The primary structure of MP was not altered significantly by HSH, whereas its secondary, tertiary, and quaternary structures were changed. Particle size decreased first and then increased significantly, and reached a minimum when the HSH speed was 14 500 rpm. The absolute zeta potential values increased significantly and the dendritic fibrous structure of sample was destroyed when the speed exceeded 14 500 rpm. High-speed shear homogenization (14 500 rpm) decreased the particle size and unfolded the protein, which improved the emulsifying properties of MPs. Excessive HSH speeds (20 500 rpm or higher) caused an aggregation of MP molecules, which was not conducive to improving their emulsifying properties. CONCLUSION: Optimal HSH speed was achieved at 14 500 rpm to modify MPs' emulsifying and structural properties under low-fatconditions. © 2019 Society of Chemical Industry.
Assuntos
Gorduras/análise , Manipulação de Alimentos/métodos , Proteínas Musculares/química , Animais , Galinhas , Emulsões/química , Emulsões/isolamento & purificação , Manipulação de Alimentos/instrumentação , Carne/análise , Proteínas Musculares/isolamento & purificação , Tamanho da Partícula , Pressão , Dobramento de ProteínaRESUMO
A quick and specific pretreatment method based on a series of extraction clean-up disks, consisting of molecularly imprinted polymer monoliths and C18 adsorbent, was developed for the specific enrichment of salbutamol and clenbuterol residues in food. The molecularly imprinted monolithic polymer disk was synthesized using salbutamol as a template through a one-step synthesis process. It can simultaneously and specifically recognize salbutamol and clenbuterol. The monolithic polymer disk and series of C18 disks were assembled with a syringe to form a set of tailor-made devices for the extraction of target molecules. In a single run, salbutamol and clenbuterol can be specifically extracted, cleaned, and eluted by methanol/acetic acid/H2 O. The target molecules, after a silylation derivatization reaction were detected by gas chromatography-mass spectrometry. The parameters including solvent desorption, sample pH, and the cycles of reloading were investigated and discussed. Under the optimized extraction and clean-up conditions, the limits of detection and quantitation were determined as 0.018-0.022 and 0.042-0.049 ng/g for salbutamol and clenbuterol, respectively. The assay described was convenient, rapid, and specific; thereby potentially efficient in the high-throughput analysis of ß2 -agonists residues in real food samples.
Assuntos
Agonistas Adrenérgicos beta/isolamento & purificação , Albuterol/isolamento & purificação , Clembuterol/isolamento & purificação , Contaminação de Alimentos/análise , Carne/análise , Extração em Fase Sólida/métodos , Agonistas Adrenérgicos beta/análise , Adsorção , Albuterol/análise , Animais , Clembuterol/análise , Cromatografia Gasosa-Espectrometria de Massas , Impressão Molecular , Polímeros/síntese química , Polímeros/química , Extração em Fase Sólida/instrumentação , SuínosRESUMO
Recent studies have identified that ErbB3 binding protein 1 (EBP1) is broadly expressed in various cancer tissues and critically involved in plenty of biological processes in this regard. However, the functional role of EBP1 in pancreatic ductal adenocarcinoma (PDAC) has never been elucidated. In this study, we found that EBP1 could serve as a prognostic biomarker of PDAC. Western blot analysis revealed that EBP1 was remarkably upregulated in PDAC tissues and cell lines. Using immunohistochemical analysis, we showed that the expression of EBP1 was correlated with tumor size (P = 0.004), histological differentiation (P = 0.041), and tumor node metastasis (TNM) stage (P = 0.000). Notably, Kaplan-Meier curve showed that high expression of EBP1 predicted significantly worsened prognosis of PDAC patients (P = 0.001). In addition, knockdown of EBP1 expression suppressed PDAC cell proliferation and retarded cell cycle progression. Furthermore, depletion of EBP1 induced the apoptosis of Panc-1 cells. Of great interest, we found that EBP1 interacted with anti-apoptotic protein, Bcl-xL, and promoted its accumulation. In summary, our results suggest that EBP1 is a novel prognostic indicator and potential therapeutic target of PDAC, shedding new insights into the important role of EBP1 in cancer development.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/biossíntese , Adenocarcinoma/genética , Biomarcadores Tumorais/biossíntese , Carcinoma Ductal Pancreático/genética , Proteínas de Ligação a RNA/biossíntese , Proteínas Adaptadoras de Transdução de Sinal/genética , Adenocarcinoma/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose , Biomarcadores Tumorais/genética , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Proteínas de Ligação a RNA/genéticaRESUMO
Herein, an ultrasensitive and selective colorimetric assay for antibiotics, using chloramphenicol (CAP) as the model analyte, was developed based on magnetic aptamer-HRP-platinum composite probes and exonuclease-assisted target recycling. The composite probes were prepared through immunoreactions between the double stranded DNA antibody (anti-DNA) labeled on core-shell Fe3O4@Au nanoparticles (AuMNP-anti-DNA) as the capture probe, and the double stranded aptamer (aptamer hybrid with its complementary oligonucleotides) labeled on Pt@HRP nanoparticles as the nanotracer (ds-Apt-HRP-PtNPs). When the CAP samples were incubated with the probes for 30 min at room temperature, they could be captured by the aptamer to form a nanotracer-CAP complex, which was then released into the supernatant after magnetic separation. This is because the anti-DNA on the capture probes cannot recognize the single strand aptamer-CAP complex. The exonuclease I (Exo I) added into the supernatant can further digest the aptamer-CAP from the 3'-end of the aptamer and the CAP in the aptamer-CAP complex can be released again, which can further participate in a new cycling process to react with the probes. Pt and HRP in the nanotracer could both catalyze and dual amplify the absorbance at 650 nm ascribed to the 3,3',5,5'-tetramethylbenzidine (TMB)-H2O2 system. Moreover, Exo I can assist the target recycling, which can further amplify the signal. Thus, the triple amplified signal can be quantified by ultraviolet-visible spectroscopy. The experimental results showed that the CAP detection possessed a linear range of 0.001-10 ng mL(-1) and a detection limit of 0.0003 ng mL(-1) (S/N = 3). The assay was successfully employed to detect CAP in milk, which is much more facile, time saving, and sensitive than the commercial ELISA kits.
Assuntos
Antibacterianos/análise , Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais/métodos , Cloranfenicol/análise , Nanopartículas de Magnetita/química , Leite/química , Platina/química , Animais , Antibacterianos/metabolismo , Aptâmeros de Nucleotídeos/metabolismo , Benzidinas/química , Cloranfenicol/metabolismo , Colorimetria/métodos , Exodesoxirribonucleases/metabolismo , Ouro/química , Peróxido de Hidrogênio/química , Limite de Detecção , Nanopartículas de Magnetita/ultraestruturaRESUMO
The study elucidates that the pH shifting treatment unfolds the conformation of soybean protein isolate (SPI), enabling it to intertwine with bacterial cellulose (BC) and form SPI/BC co-assemblies. Results from intrinsic fluorescence spectroscopy and surface hydrophobicity indicate that the SPI with pH shifting treatment shows a notable blue shift in maximum emission wavelength and increased surface hydrophobicity. It demonstrates that pH shifting treatment facilitates the unfolding of SPI's molecular conformation, promoting its entanglement with high aspect ratio BC. Particle size distribution and microstructural analysis further demonstrate that the pH shifting treatment facilitates the formation of SPI/BC co-assemblies. Evaluation of processing properties reveals that the SPI/BC co-assemblies exhibited exceptional gel and emulsification properties, with gel strength and emulsifying activity respectively six and two times higher than natural SPI. This enhancement is attributed to the thickening properties of BC with a high aspect ratio and the superior hydrophobicity of SPI in its molten globule state.
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To improve prediction performance and reduce artifacts in Raman spectra, we developed an eXtreme Gradient Boosting (XGBoost) preprocessing method to preprocess the Raman spectra of glucose, glycerol and ethanol mixtures. To ensure the robustness and reliability of the XGBoost preprocessing method, an X-LR model (which combined XGBoost preprocessing and a linear regression (LR) model) and a X-MLP model (which combined XGBoost preprocessing and a multilayer perceptron (MLP) model) were developed. These two models were used to quantitatively analyze the concentrations of glucose, glycerol and ethanol in the Raman spectra of mixed solutions. The proportion map of hyperparameters was firstly used to narrow down the search range of hyperparameters in the X-LR and the X-MLP models. Then the correlation coefficients (R2), root mean square of calibration (RMSEC), and root mean square error of prediction (RMSEP) were used to evaluate the models' performance. Experimental results indicated that the XGBoost preprocessing method achieved higher accuracy and generalization capability, with better performance than those of other preprocessing methods for both LR and MLP models.
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The purpose of this study is to explain the mechanism of porcine myofibrillar protein gel properties modulated by κ-carrageenan. The textural properties results showed that the stress at fracture of the composite gel with 0.4% κ-carrageenan had the highest value (91.33 g), which suggested that the 0.4% κ-carrageenan addition was the limitation. The strain at fracture was significantly reduced with κ-carrageenan addition. The composite gel with 0.4% κ-carrageenan had the lowest proportion of T22 (7.85%) and the shortest T21 relaxation time (252.81 ms). The paraffin section showed that the phase separation behavior of the composite gel transformed from single-phase behavior to dispersed phase behavior to bi-continuous phase behavior, and the ratio of CG/MP phase area significantly increased from 0.06 to 1.73. The SEM showed that the three-dimensional network of myofibrillar protein transformed from a loose structure to a compact structure to an unaggregated structure with κ-carrageenan addition. The myofibrillar protein network of the treatment with 0.4% κ-carrageenan had the highest DF value (1.7858) and lowest lacunary value (0.452). The principal component analysis was performed on the data of microstructure and textural properties, and the results showed that the dispersed phase behavior and moisture stabilization promoted the aggregation of myofibrillar protein and the composite gel had better water holding capacity and textural properties, while bi-continuous phase behavior hindered the aggregation of myofibrillar protein and the composite gel had worse water holding capacity and textural properties.
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The 25Mg(p, γ)26Al reaction plays an important role in the study of cosmic 1.809 MeV γ-ray as a signature of ongoing nucleosynthesis in the Galaxy. At astrophysical temperature around 0.1 GK, the 25Mg(p, γ)26Al reaction rates are dominated by the 92 keV resonance capture process. We report a precise measurement of the 92 keV 25Mg(p, γ)26Al resonance in the day-one experiment at Jinping Underground Nuclear Astrophysics experiment (JUNA) facility in the China Jinping Underground Laboratory (CJPL). The resonance strength and ground state feeding factor are determined to be 3.8±0.3 ×10-10 eV and 0.66±0.04, respectively. The results are in agreement with those reported in the previous direct underground measurement within uncertainty, but with significantly reduced uncertainties. Consequently, we recommend new 25Mg(p, γ)26Al reaction rates which are by a factor of 2.4 larger than those adopted in REACLIB database at the temperature around 0.1 GK. The new results indicate higher production rates of 26gAl and the cosmic 1.809 MeV γ-ray. The implication of the new rates for the understanding of other astrophysical situations is also discussed.
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The effect and mechanism of myofibrillar protein (MP) gelation influenced by konjac glucomannan (KG) addition were studied. The KG addition significantly improved gel strength and water holding capability (WHC) of MP-KG composite gel, but it had additive limitation at 1.0%. The SEM showed that KG (<1.0%) reduced the appearance of moisture channels and promoted the formation of an integral MP gel network. Raman spectroscopy showed that KG addition (<1.0%) promoted the protein unfolding and the interaction of hydrophobic groups during thermal processing. However, the KG (>1.0%) would form continuous viscous hydrogel and interpenetrate with the MP solution, which hindered the interaction of hydrophobic groups during thermal process, and the MP formed a loose and degraded final structure. Hence, MP gels produced with the addition of KG underwent a transformation from a loose structure to a compact structure to an unaggregated structure, which was influenced by moisture stability and phase separation behavior.
Assuntos
Mananas/química , Proteínas Musculares/química , Géis , Interações Hidrofóbicas e Hidrofílicas , Estabilidade Proteica , Desdobramento de Proteína , Reologia , Água/químicaRESUMO
The effects of three polysaccharides on the textural properties and microstructure of myofibrillar protein (MP) gels were studied. The gel strength and rheological properties of composite MP gels were significantly improved with insoluble dietary fibre (DF) and modified starch (MS) addition, while konjac glucomannan (KG) had limited effects at 1% addition. The SEM images indicated that moisture extrusion formed moisture channels and deteriorated the aggregation of MP gel networks during the thermal process. The polysaccharides stabilized moisture and reduced the appearance of moisture channels in the gel network, thereby promoting the formation of compact and integral gel networks. The MP-polysaccharide mixture is a thermally incompatible system and presented two main forms after the thermal process: 1) the "trapped" structure and 2) the "interpenetrated" structure. In the "trapped" structure, the MP was the dominant structure of the composite gel network. In the "interpenetrated" structure, the continuous polysaccharide hydrogel substantially hindered the aggregation of MP gel networks. Principal component analysis showed that the phase behaviour and moisture stability of polysaccharides significantly influenced the textural quality and microstructure of composite MP gelation. The study indicated that polysaccharides that contribute to moisture stability and form a "trapped structure" (phase behaviour) are ideal fat replacements for improving composite gel properties, especially DF.
Assuntos
Produtos da Carne/análise , Miofibrilas/química , Polissacarídeos/química , Animais , Fibras na Dieta , Géis/química , Mananas , Proteínas Musculares/química , Miofibrilas/ultraestrutura , Amido , SuínosRESUMO
In this study, the effect and mechanism of myofibrillar protein (MP) gelation influenced by the hydration characteristic of three polysaccharides were studied through puncture test, paraffin section, SEM and Raman spectroscopy. The gel strength and water holding capability reflect that MP gelation only significantly improves until modified starch (MS) addition beyond 1.0%. The MS granule improves MP gel property through simply physical swelling effect. At gelatinization temperature, MS absorbs the moisture nearby to compress the MP three-dimensional networks, but the swelling effect is limited. The insoluble dietary fiber (IDF) improves MP gelation property through moisture stability. The IDF addition could lessen the appearance of moisture channel in MP gel networks and promote the interaction of hydrophobic groups. The MP gelation with 2.0% IDF addition has the highest gel strength (279 g) and water holding capability (91.87%). The konjac glucomannan (KG) (>1.0%) could degrade gel property of MP gelation through interpenetrate structure, because the KG hydrogel hinders the aggregation of the MP gel networks. In conclusion, the IDF, which has strong water-holding capability at room temperature and distribute individually, is the best polysaccharides-based fat replacement in low-fat restructured products.
Assuntos
Fibras na Dieta , Hidrogéis/química , Mananas/química , Proteínas Musculares/metabolismo , Miofibrilas/química , Amido/química , Animais , Substitutos da Gordura/química , Manihot/química , Proteínas Musculares/química , Conformação Proteica/efeitos dos fármacos , Estrutura Secundária de Proteína/efeitos dos fármacos , Saccharum/química , SuínosRESUMO
Herein, a smart single-stranded DNA binding protein (SSB)-assisted fluorescence aptamer switch based on fluorescence resonance energy transfer (FRET) was designed. The FRET switch was synthesized by connecting SSB labeled quantum dots (QDs@SSB) as donor with aptamer (apt) labeled gold nanoparticles (AuNPs@apt) as acceptor, and it was employed for detecting chloramphenicol (CAP) in a homogenous solution. In the assay, the interaction between core-shell QDs@SSB and AuNPs@apt leads to a dramatic quenching (turning off). After adding CAP in the detection system, AuNPs@apt can bind the target specifically then separate QDs@SSB with AuNPs@apt-target, resulting in restoring the fluorescence intensity of QDs (turning on). Consequently, the fluorescence intensity recovers and the recovery extent can be used for detection of CAP in homogenous phase via optical responses. Under optimal conditions, the fluorescence intensity increased linearly with increasing concentrations of CAP from 0.005 to 100ngmL-1. The limit of this fluorescence aptamer switch was around 3pgmL-1 for CAP detection. When the analyte is changed, the assay can be applied to detect other targets only by changing relative aptamer in AuNPs@apt probe. Furthermore, it has potential to be served as a simple, sensitive and portable platform for antibiotic contaminants detection in biological and environmental samples.
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Antibacterianos/análise , Aptâmeros de Nucleotídeos/química , Cloranfenicol/análise , Proteínas de Ligação a DNA/química , Transferência Ressonante de Energia de Fluorescência/métodos , Nanopartículas Metálicas/química , Pontos Quânticos/química , Animais , Técnicas Biossensoriais/métodos , Ouro/química , Limite de Detecção , Nanopartículas Metálicas/ultraestrutura , Leite/química , Pontos Quânticos/ultraestruturaRESUMO
In this work, a novel aptamer functionalized magnetic adsorbent was developed and combined with magnetic dispersive solid phase extraction (MDSPE) for selective enrichment of several amphenicol antibiotics residues (chloramphenicol(CAP), thiamphenicol(TAP) and florphenicol(FF)) in foodstuff then determined by High Performance Liquid Chromatography (HPLC)-Diode array detector(DAD). Firstly, a magnetic silica-coated Fe3O4 microsphere(Fe3O4@SiO2) was synthetized by sol-gel method, then it was functionalized by amino groups through 3-Aminopropyltriethoxysilane (APTES) reagent to form Fe3O4@SiO2-NH2; Thirdly, the amino group on Fe3O4@SiO2-NH2 was transferred to carboxylic group via the succinic anhydride to form Fe3O4@SiO2-COOH. Finally a kind of DNA aptamer with amino group which can simultaneously recognize CAP, TAP, FF, was functionalized on Fe3O4@SiO2-COOH through adding the EDC/sulfo-NHS coupling reagent (Fe3O4@SiO2@Apt). Therefore, a very stable and useful adsorbent was synthesized for the detection of chloramphenicol series antibiotics. This adsorbent can specifically and simultaneously recognize and enrich CAP, TAP, and FF with high adsorption amount from some complicated food matrix, e.g. milk based on the high affinity of aptamer towards the analytes. The saturated extraction capacities for CAP, TAP and FF by the adsorbent were 2.82, 2.56, 2.72µg/g (mass of target/adsorbent)respectively and the enrichment folds were more than 100 times. Afterwards, the target analytes were washed away by pH 8.5 0.1M Tris-HCl buffer solution and detected by HPLC-DAD. The parameters including extraction temperature, extraction capacity, extraction & desorption pH, extraction & desorption time were investigated. With the optimized conditions, the limits of detection (LOD) and limits of quantitation (LOQ) were 0.12-0.17ng/ml and 0.40-0.55ng/ml for the amphenicols in milk. The adsorbent also has good reproducibility for extraction which can be reused at least for 60 cycles with the recovery over 80% (Fig. S2). The MDSPE combined with HPLC-DAD detection possessed the advantages of high selectivity, extraction capacity and very convenient for magnetic separation. In addition, this method is environment friendly and employed no organic solution in the period of pretreatment and extracting. It is a universal platform which can be extended to selective enrichment other organic pollutants residues if changing the modified aptamers.
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Cloranfenicol/análise , Cloranfenicol/isolamento & purificação , Cromatografia Líquida de Alta Pressão/métodos , Resíduos de Drogas/análise , Resíduos de Drogas/isolamento & purificação , Análise de Alimentos/métodos , Animais , Cloranfenicol/química , Resíduos de Drogas/química , Limite de Detecção , Modelos Lineares , Imãs , Leite/química , Reprodutibilidade dos Testes , Extração em Fase Sólida/métodosRESUMO
An effective electrochemical aptasensor has been developed for the detection of multiplex antibiotics using Y-shaped DNA probes. These probes-based metal ions encoded the nanoscale metal-organic frameworks (NMOF) as a substrate, and circular strand-replacement DNA polymerization (CSRP) target triggered the amplification strategy. The Y-DNA probes (Y-DNA) were assembled using an assisted DNA probe (assisted DNA labeled with magnetic gold nanoparticles) which can hybridize to the captured DNA probe (consisting of aptamer and primer recognition region), and signal tags (NMOF encapsulating signal DNAs and different metal ions such as Pb2+ or Cd2+). Notably, NMOF was employed as the developed platform with a large specific area to load abundant metal ions that can produce distinguishable signals. In the presence of targets, chloramphenicol (CAP) and oxytetracycline (OTC) as models, the conformational change of the captured DNA can disassemble the Y-DNA probes that can consequently release the signal tags in the supernatant due to the high affinity of targets towards the aptamer domain than its complementary sequences. Subsequently, the exposed sequences of captured DNA serve as the initiators for triggering the target cyclic-induced polymerization with the assistance of Bst DNA polymerase. Thus, numerous signal tags could be detected by square wave voltammetry in the supernatant after magnetic separation, thereby amplifying the electrochemical signals. The proposed strategy exhibited a high sensitivity to antibiotics with a detection limit of 33 and 48 fM (S/N = 3) towards CAP and OTC, respectively. Moreover, this aptasensor showed promising applications for the detection of other analytes.
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Antibacterianos/análise , Aptâmeros de Nucleotídeos , DNA Circular , Técnicas Eletroquímicas , Sondas de DNA , Ouro , Limite de Detecção , Nanopartículas MetálicasRESUMO
A novel label-free, universal, and high throughput aptasensor was developed based on a microchip electrophoresis (MCE) platform for automatic detection of antibiotic residues in food. Firstly, chloramphenicol (CAP) was employed as a model to be captured by its aptamer probe (Apt). Then, the partial complementary oligonucleotide of CAP's aptamer (C-DNA) was introduced into the reaction system. Because the Apt-CAP complex can't further hybrid with free C-DNA, the amount of hybrid Apt-C-DNA double strand DNA (dsDNA) was less than that without adding the target. Finally, the above mixture was introduced into the microchip electrophoresis (MCE) platform for detection, both dsDNA and Apt-CAP can be separated and produce different fluorescence signals in the MCE. In a certain concentration range, the ratio of signal between dsDNA and Apt-CAP (IdsDNA/I Apt-CAP) was proportional to the concentration of targets. Under the optimum conditions, the ratio showed a satisfactory linearity range from 0.008 to 1ng/mL of CAP with a detection limit of 0.003ng/mL. Thus, a universal MCE-based assay was developed for quantifying CAP automatically. The method was also successfully applied in the different food samples for CAP detection, which showed a good recovery (Milk: 91.1-108%, Fish: 86.1-114%) and the results were consistent with that of ELISA. This method owned many merits as follows: firstly, MCE was a high throughput screening platform and the detection time is limited to 3min for each sample. Secondly, the aptamer probes can be directly used for detection without labeling any signal tag which can facilitate the preparation procedures of probes. Thirdly, the operation was easy just by the following steps: firstly, the mixture of aptamer probes were incubated followed adding C-DNA; then measurement was performed. Moreover, the assay with MCE platform can be used to detect other targets just by changing the corresponding aptamer probe; it can even realize simultaneous detection when the targets have aptamers with different number of base pairs. Above all, it's a high- throughput and prospective method which can be applied in high throughput screening of antibiotics in food safety.
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Antibacterianos/análise , Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais/métodos , Cloranfenicol/análise , Eletroforese em Microchip/métodos , Análise de Alimentos/métodos , Contaminação de Alimentos/análise , Animais , Antibacterianos/química , Técnicas Biossensoriais/instrumentação , Bovinos , Cloranfenicol/química , Eletroforese em Microchip/instrumentação , Peixes , Análise de Alimentos/instrumentação , LeiteRESUMO
In order to investigate the mechanism between flavor binding and proteins degradation during meat processing, the influence of different trypsin contents on the structure of myosin and the adsorption capacity on aldehydes and ketones was determined. The 1% treatment produced subfragment 2 (S2), light meromyosin (LMM) and decreased 18 and 16kDa light chains; 5% and 10% treatments produced 100 and 65kDa new bands and more S2, LMM and cleaned light chains. With the rising trypsin contents, ß-sheet, ß-turn, random coil, hydrophobicity and total sulfydryl content increased; solubility, α-helix and free percentages of aldehydes and ketones decreased. The increase of absorbing capacity could be attributed to the increased hydrophobicity and total sulphydryl and the unfolding of secondary structures by exposing reactive amino and thiol groups and hydrophobic sites; the decreased solubility was related to the increased hydrophobicity. The trypsin-dose dependent proteolysis of myosin increased the retention of volatile compounds.
Assuntos
Subfragmentos de Miosina/química , Miosinas/química , Tripsina/química , Aldeídos/química , Concentração de Íons de Hidrogênio , Hidrólise , Interações Hidrofóbicas e Hidrofílicas , Cetonas/química , Estrutura Secundária de Proteína , Solubilidade , Análise Espectral Raman , Compostos de Sulfidrila/químicaRESUMO
Novel label-free and multiplex aptasensors have been developed for simultaneous detection of several antibiotics based on a microchip electrophoresis (MCE) platform and target catalyzed hairpin assembly (CHA) for signal amplification. Kanamycin (Kana) and oxytetracycline (OTC) were employed as models for testing the system. These aptasensors contained six DNA strands termed as Kana aptamer-catalysis strand (Kana apt-C), Kana inhibit strand (Kana inh), OTC aptamer-catalysis strand (OTC apt-C), OTC inhibit strand (OTC inh), hairpin structures H1 and H2 which were partially complementary. Upon the addition of Kana or OTC, the binding event of aptamer and target triggered the self-assembly between H1 and H2, resulting in the formation of many H1-H2 complexes. They could show strong signals which represented the concentration of Kana or OTC respectively in the MCE system. With the help of the well-designed and high-quality CHA amplification, the assay could yield 300-fold amplified signal comparing that from non-amplified system. Under optimal conditions, this assay exhibited a linear correlation in the ranges from 0.001ngmL-1 to 10ngmL-1, with the detection limits of 0.7pgmL-1 and 0.9pgmL-1 (S/N=3) toward Kana and OTC, respectively. The platform has the following advantages: firstly, the aptamer probes can be fabricated easily without labeling signal tags for MCE detection; Secondly, the targets can just react with probes and produce the amplified signal in one-pot. Finally, the targets can be simultaneously detected within 10min in different channels, thus high-throughput measurement can be achieved. Based on this work, it is estimated that this detection platform will be universally served as a simple, sensitive and portable platform for antibiotic contaminants detection in biological and environmental samples.
Assuntos
Antibacterianos/análise , Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais/métodos , Resíduos de Drogas/análise , Eletroforese em Microchip/métodos , Canamicina/análise , Oxitetraciclina/análise , Animais , Técnicas Biossensoriais/instrumentação , Eletroforese em Microchip/instrumentação , Desenho de Equipamento , Análise de Alimentos/instrumentação , Análise de Alimentos/métodos , Contaminação de Alimentos/análise , Limite de Detecção , Leite/químicaRESUMO
Twelve pieces of longissimus dorsi were processed into Chinese traditional dry-cured loins. The changes in the proteolylic enzymes activities, myofibrillar proteins degradation, and free amino acids content were investigated during processing. Compared with fresh piece (0 day), the cathepsin B + L and calpains activities decreased after dry-curing and maintained potential activities values of 23.25 and 15.04% in the final products, respectively. The myosin heavy chain (MHC) and C protein were intensely degraded at the dry-ripened stage; the 50 kDa desmin increased at day 2 and then disappeared at day 11. The total free amino acids content increased from 333.18 mg/100 g in the raw to 1096.54 mg/100 g at the end of the dry-ripening. This work provided a mechanism for the accumulation of free amino acids and predicted the proteolysis extent of myofibrillar proteins by monitoring the changes of three marker proteins (MHC, C protein and 50 kDa desmin) during Chinese traditional dry-cured loins processing.