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1.
Food Chem Toxicol ; 135: 110975, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31747619

RESUMO

This paper presents innovative packaging materials made of environmentally friendly biodegradable polymers (polylactide and polyhydroxybutyrate) with the addition of natural colorants commonly used in the food industry. Colorants fulfilled the role of indicator, changing colour under the influence of external factors, and gave the materials the characteristics of intelligent packaging, where colour changes indicated the life time of the materials. The paper gives the mechanical and thermal properties of the materials obtained, and describes changes in the colour of the samples under the influence of thermooxidation, UV and weathering, as well as the biodegradability of the materials. The packaging materials presented are in line with current trends in the packaging market and legal requirements. The samples, in addition to the basic functions of packaging materials, are pro-ecological and fully biodegradable new generation materials.


Assuntos
Plásticos Biodegradáveis/química , Compostos Cromogênicos/química , Embalagem de Alimentos , /química , Plásticos Biodegradáveis/metabolismo , Chaetomium/metabolismo , Clorofila/química , Curcumina/química , Luteína/química , Fungos Mitospóricos/metabolismo , Oxirredução , Poliésteres/química , Poliésteres/metabolismo , Temperatura , beta Caroteno/química
2.
Biochem Med (Zagreb) ; 30(1): 010702, 2020 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-31839722

RESUMO

Introduction: Clinical application of rivaroxaban and apixaban does not require therapeutic monitoring. Commercial anti-activated factor X (anti-FXa) inhibition methods for all anti-FXa drugs are based on the same principle, so there are attempts to evaluate potential clinical application of heparin-calibrated anti-FXa assay as an alternative method for direct FXa inhibitors. We aimed to evaluate relationship between anti-FXa methods calibrated with low molecular weight heparin (LMWH) and with drug specific calibrators, and to determine whether commercial LMWH anti-FXa assay can be used to exclude the presence of clinically relevant concentrations of rivaroxaban and apixaban. Materials and methods: Low molecular weight heparin calibrated reagent (Siemens Healthineers, Marburg, Germany) was used for anti-FXa activity measurement. Innovance heparin (Siemens Healthineers, Marburg, Germany) calibrated with rivaroxaban and apixaban calibrators (Hyphen BioMed, Neuville-sur-Oise, France) was used for quantitative determination of FXa inhibitors. Results: Analysis showed good agreement between LMWH calibrated and rivaroxaban calibrated activity (κ = 0.76) and very good agreement with apixaban calibrated anti-Xa activity (κ = 0.82), respectively. Low molecular weight heparin anti-FXa activity cut-off values of 0.05 IU/mL and 0.1 IU/mL are suitable for excluding the presence of clinically relevant concentrations (< 30 ng/mL) of rivaroxaban and apixaban, respectively. Concentrations above 300 ng/mL exceeded upper measurement range for LMWH anti-FXa assay and cannot be determined by this method. Conclusion: Low molecular weight heparin anti-FXa assay can be used in emergency clinical conditions for ruling out the presence of clinically relevant concentrations of rivaroxaban and apixaban. However, use of LMWH anti-FXa assay is not appropriate for their quantitative determination as an interchangeable method.


Assuntos
Anticoagulantes/química , Testes de Coagulação Sanguínea/métodos , Heparina de Baixo Peso Molecular/química , Pirazóis/química , Piridonas/química , Rivaroxabana/química , Anticoagulantes/metabolismo , Área Sob a Curva , Testes de Coagulação Sanguínea/normas , Calibragem , Compostos Cromogênicos/química , Fator Xa/química , Fator Xa/metabolismo , Inibidores do Fator Xa/química , Inibidores do Fator Xa/metabolismo , Heparina de Baixo Peso Molecular/metabolismo , Humanos , Pirazóis/metabolismo , Piridonas/metabolismo , Curva ROC
3.
Talanta ; 206: 120211, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31514873

RESUMO

Urinary glucose determination using a glucose test strip is simple and convenient in daily self-monitoring of diabetes. However, diabetic patients exhibit acquired impaired color vision (ICV), which results in the inability to discriminate between hues. Even with the assistance of a color chart, it is still not easy for these patients to read the urinary glucose results with the naked eye. In this study, a smartphone camera using an image-based colorimetric detection method was successfully developed for quantitative analysis of urine glucose. A horseradish peroxidase-hydrogen peroxide-3,3'5,5'-tetramethylbenzidine (HRP-H2O2-TMB) system was optimized for a reliable and gradual color fading process via a glucose oxidase (GOD) catalyzed oxidation reaction. The color changes of the peroxidase-H2O2 enzymatic reactions in the 96-well microplate were captured by a smartphone RGB camera with subsequent detection of red, green, and blue (RGB) intensities decreasing at each image pixel. The highly quantitative relationships between the glucose concentrations and the color characteristic values of the blue channel of the captured images were successfully established. The high accuracy of this method was demonstrated in urine glucose measurements with a linear response over the 0.039 mg mL-1 to 10.000 mg mL-1 glucose concentration range and a 0.009 mg mL-1 detection limit. The method has great potential as a point-of-need platform for diabetic patients with defective color vision and features high accuracy and low cost.


Assuntos
Diabetes Mellitus/urina , Glucose/análise , Glicosúria/diagnóstico , Smartphone , Armoracia/enzimologia , Benzidinas/química , Compostos Cromogênicos/química , Colorimetria/métodos , Glucose/química , Glucose Oxidase/química , Peroxidase do Rábano Silvestre/química , Humanos , Peróxido de Hidrogênio/química , Limite de Detecção , Fotografação/instrumentação , Testes Imediatos
4.
Talanta ; 204: 278-284, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31357294

RESUMO

Two-dimensional WO3 nanosheets were prepared by ultrasonic exfoliation of bulk WO3·2H2O in water and characterized by transmission electron microscopy, atomic force microscopy, X-ray diffraction, X-ray photoelectron spectroscopy, Raman, dynamic light scattering. The nanosheets were discovered to possess the peroxidase-like catalytic activity, which can catalyze the oxidation of 3, 3', 5, 5'-tetramethylbenzidine by H2O2. The catalytic mechanism was also investigated by the scavenger experiments. Taking advantage of the peroxidase-like activity of WO3 nanosheets, a facile colorimetric method for xanthine was developed by combining the oxidation reaction of xanthine catalyzed by xanthine oxidase. The linear range for xanthine was ranged from 25 to 200 µmol L-1. The limit of detection for xanthine was 1.24 µmol L-1. The colorimetric method was applied to determine xanthine in urine samples.


Assuntos
Colorimetria/métodos , Nanoestruturas/química , Óxidos/química , Tungstênio/química , Xantina/urina , Animais , Benzidinas/química , Materiais Biomiméticos/química , Catálise , Bovinos , Compostos Cromogênicos/química , Humanos , Peróxido de Hidrogênio/química , Limite de Detecção , Oxirredução , Peroxidase/química , Xantina/química , Xantina Oxidase/química
5.
Talanta ; 204: 542-547, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31357331

RESUMO

The repeatable immobilization of molecular recognition elements onto particle surfaces has a strong impact on the outcomes of affinity-based assays. In this work, an automatic method for the immobilization of immunoglobulin G (IgG) onto protein A-Sepharose microbeads was established through the flow programming features of the portable lab-on-valve platform using micro-bead injection spectroscopy. The reproducible packing of protein A-microbeads between two optic fibers was feasible, allowing on-column probing of IgG retention. The automation of solutions handling and the precise control of time of IgG interaction with the beads rendered repeatable immobilization cycles, within a short timeframe (<2 min). The proposed method featured the preparation of disposable immunosorbents for downstream analytical applications, such as immunosensing or microenrichment of target analytes. In-situ quantification of IgG@protein A-microbeads was carried out using a horseradish peroxidase-labeled detection IgG. The colorimetric oxidation of 3,3',5,5'-tetramethylbenzidine was monitored on-column. Quantitation of mouse and human IgG immobilized@protein A-microbeads was achieved for loading masses between 0.1 and 0.4 µg per ca. 5.5 mg of sorbent. The implemented detection strategy allowed the quantification of human IgG in certified human serum (ERM®- DA470k/IFCC) and spiked saliva, yielding recoveries of 102-108% and requiring minimal volume (1-15 µL) from serum and saliva.


Assuntos
Cromatografia de Afinidade/métodos , Proteínas Imobilizadas/química , Imunoglobulina G/sangue , Sefarose/análogos & derivados , Proteína Estafilocócica A/química , Animais , Armoracia/enzimologia , Benzidinas/química , Compostos Cromogênicos/química , Colorimetria/métodos , Peroxidase do Rábano Silvestre/química , Humanos , Camundongos , Microesferas , Oxirredução , Saliva/química , Sefarose/química
6.
Talanta ; 204: 833-839, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31357372

RESUMO

The peroxidase-like activity of ficin is relatively low, which limits its application. It was found that thiol groups of ficin could inhibit its peroxidase-like activity. So, two procedures, i.e., direct blocking with N-ethylmaleimide (NEM), or using tris (2-carboxyethyl) phosphine hydrochloride (TCEP) to interrupt disulfide bonds then blocking thiol groups with NEM, were applied to block thiol groups of ficin, ficin-NEM (ficin-N) and ficin-TCEP-NEM (ficin-TN) were produced, respectively. The blocking of thiol groups accelerated the peroxidase activity dramatically. The peroxidase catalytic activity of ficin-N and ficin-TN toward the peroxidase substrate 3,3',5,5'-tetramethylbenzidine (TMB) oxidation by H2O2 was about 2.5-fold and 5-fold increase compared with ficin, respectively, which accompanied a color change from colorless to blue and followed classic Michaelis-Menten model. The kinetic parameters indicated that higher affinity of ficin-N (Km = 0.31) and ficin-TN (Km = 0.39) to H2O2 compared with ficin (Km = 0.58), and ficin-TN had the highest Kcat which increased by 6.5 times and 4.5 times for TMB and H2O2, respectively. According to these findings, a colorimetric method with high sensitivity for the detection of biothiols was developed due to sulfhydryl compounds inhibited the peroxidase activity of ficin. Comparing with ficin and ficin-N, ficin-TN had the widest detection range (0.01-16 µM) and the lowest detection limit (3 nM). The practical applications of ficin-TN for biothiol determination in human serum samples have been demonstrated with satisfactory results. Ficin-N and ficin-TN are promising to apply to the bioanalysis.


Assuntos
Cisteína/sangue , Ficina/química , Glutationa/sangue , Homocisteína/sangue , Peroxidases/química , Benzidinas/química , Compostos Cromogênicos/química , Colorimetria/métodos , Etilmaleimida/química , Humanos , Peróxido de Hidrogênio/química , Indicadores e Reagentes/química , Cinética , Limite de Detecção , Fosfinas/química
7.
Parasit Vectors ; 12(1): 282, 2019 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-31159851

RESUMO

BACKGROUND: Avian haemosporidian parasites can cause severe disease in their hosts due to excessive exo-erythrocytic merogony and anaemia caused by blood stages. Notably, the development of megalomeronts by species of Haemoproteus and Leucocytozoon has been associated with mortalities in birds. Diagnosis of lethal infections is currently accomplished by the detection of parasites' tissue stages in histological sections combined with PCR and sequencing. However, sequences frequently are not reliably obtained and the generic discrimination of exo-erythrocytic tissue stages based on morphological characters is challenging. Therefore, the present study aimed at developing specific molecular probes for the identification of Haemoproteus spp. and Leucocytozoon spp. in histological sections using chromogenic in situ hybridization. METHODS: Parasite subgenus-specific oligonucleotide probes were designed to target the 18S ribosomal RNA of Haemoproteus species (subgenus Parahaemoproteus) and Leucocytozoon spp. (subgenus Leucocytozoon) and were in situ hybridized to sections from formalin-fixed, paraffin-embedded tissue samples determined positive for these parasites by PCR and histopathology. To confirm the presence of parasites at sites of probe hybridization, consecutive sections were stained with haematoxylin-eosin and examined. RESULTS: Parahaemoproteus- and Leucocytozoon-specific probes labelled erythrocytic and exo-erythrocytic stages of Haemoproteus spp. and Leucocytozoon spp., respectively. Binding of probes to parasites was consistent with detection of the same exo-erythrocytic meronts in consecutive haematoxylin-eosin-stained sections. Cross-reactivity of the probes was ruled out by negative chromogenic in situ hybridization when applied to samples positive for a parasite of a genus different from the probes' target. CONCLUSIONS: Chromogenic in situ hybridization using 18S ribosomal RNA-specific oligonucleotide probes reliably identifies and discriminates Haemoproteus and Leucocytozoon parasites in tissue sections and enables unequivocal diagnosis of haemosporidioses.


Assuntos
Aves/parasitologia , Haemosporida/genética , Sondas Moleculares , Infecções Protozoárias em Animais/diagnóstico , Animais , Doenças das Aves/diagnóstico , Compostos Cromogênicos/química , DNA de Protozoário/genética , Haemosporida/isolamento & purificação , Hibridização In Situ , Filogenia , Reação em Cadeia da Polimerase , RNA Ribossômico 18S/genética
8.
Spectrochim Acta A Mol Biomol Spectrosc ; 222: 117228, 2019 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-31212194

RESUMO

A sensitive and visible colorimetric strategy was proposed for Hg2+ detection by thymine-Hg2+-thymine (T-Hg2+-T) coordination chemistry and entropy driven catalytic reaction. The entropy driven catalytic reaction is induced by T-Hg2+-T coordination chemistry, resulting the releasing of G-riched sequence. Hemin/G-quadruplex-HRP-mimicking DNAzyme can be formed with the help of hemin, catalyzing TMB to TMB+ with a color change from colorless to blue. The sensitivity of this strategy can be reached to 2 pM, which is significantly improved by entropy driven catalytic reaction. In addition, entropy driven catalytic reaction provides a more reliable and accurate results. This method shows great promise for on-site analysis and in-house diagnosis of Hg2+ in water.


Assuntos
Quadruplex G , Hemina/química , Mercúrio/análise , Timina/análogos & derivados , Poluentes Químicos da Água/análise , Benzidinas/química , Técnicas Biossensoriais/métodos , Catálise , Compostos Cromogênicos/química , Colorimetria/métodos , Complexos de Coordenação/química , DNA Catalítico/química , Entropia , Limite de Detecção , Água/química
9.
Int J Biol Macromol ; 136: 994-999, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31229547

RESUMO

The non-catalytic conversion of chitin into N-acetyl-ᴅ-glucosamine (GlcNAc) derivatives such as 2-acetamido-2,3-dideoxy-ᴅ-erythro-hex-2-enofuranose (Chromogen I) was investigated in high-temperature water at 290-390 °C and 25 MPa with a reaction time of 0-180 min. High-temperature water treatment is a promising method for chitin conversion as it does not require the use of any additional organic solvents or ionic liquids. A semi-batch reactor was developed to control the reaction temperature and time. It was found that the chitin powder could be converted into a water-soluble fraction in ~90% yield, with Chromogen I being obtained in a maximum yield of 2.6%. Furthermore, a kinetic model was developed to estimate the reaction rate for the conversion of the chitin powder to the water-soluble fraction.


Assuntos
Quitina/química , Compostos Cromogênicos/química , Temperatura Alta , Água/química , Hidrólise , Cinética , Modelos Químicos , Solubilidade
10.
Ann Biol Clin (Paris) ; 77(3): 350-352, 2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-31145076

RESUMO

CHROMmagar Orientation media (Becton Dickinson) was developed and validated for the culture of urinary samples. It allows a direct identification of E. coli colonies without additional tests. As CHROMmagar Orientation media is superior to non-chromogenic media for the distinction of enterobacterial colonies, it is used for the inoculation of a large variety of samples in clinical laboratories. Direct identification of E. coli colonies cultured from these samples is not validated by the manufacturer. The difference in microbial ecology and the nature of the sample may impact CHROMagar Orientation performances for this use. We evaluated these media for the direct identification of E. coli colonies from 410 samples (excluding urine). Its sensitivity of 99% allows a direct identification of E. coli colonies cultured from a wide variety of samples. On-site testing using a large number of representative samples, allows laboratories to assess agar media performance and adapt their uses. Suppliers who are aware of frequent and non-recommended use of their culture media should perform tests and if conclusive, adapt their technical instructions.


Assuntos
Técnicas Bacteriológicas/métodos , Compostos Cromogênicos/química , Meios de Cultura/química , Escherichia coli/isolamento & purificação , Hemocultura/métodos , Humanos , Testes de Sensibilidade Microbiana , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
11.
RNA ; 25(8): 1038-1046, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31064786

RESUMO

Visualization of gene expression at single RNA molecular level represents a great challenge to both imaging technologies and molecular engineering. Here we show a single molecule chromogenic in situ hybridization (smCISH) assay that enables counting and localizing individual RNA molecules in fixed cells and tissue under bright-field microscopy. Our method is based on in situ padlock probe assays directly using RNA as a ligation template and rolling circle amplification combined with enzyme catalyzed chromogenic reaction for amplification product visualization. We show potential applications of our method by detecting gene expression variations in single cells, subcellular localization information of expressed genes, and gene expression heterogeneity in formalin-fixed, paraffin-embedded tissue sections. This facile and straightforward method can in principle be applied to any type of RNA molecules in different samples.


Assuntos
Compostos Cromogênicos/química , RNA Mensageiro/análise , Imagem Individual de Molécula/métodos , Animais , Expressão Gênica , Humanos , Hibridização in Situ Fluorescente , RNA Mensageiro/química , Inclusão do Tecido , Fixação de Tecidos
12.
Analyst ; 144(9): 2936-2941, 2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-30920552

RESUMO

Imidacloprid (IMD) is one of the most used pesticides worldwide as a systemic insecticide as well as for pest control and seed treatment. The toxic and potential carcinogenic character of IMD makes its monitoring of great relevance in the field of agriculture and environment, so sensitive methodologies for in field analysis are strongly required. In this context, we have developed a competitive immunoassay for the determination of IMD using specific monoclonal antibodies followed by electrochemical detection on screen-printed carbon electrodes (SPCE). The optimized immunosensor exhibited a good reproducibility (RSD of 9%) and a logarithmic response in the range 50-10 000 pM of IMD, with an estimated detection limit (LOD) of 24 pM, which was below the maximum levels allowed by the legislation. High-Performance Liquid Chromatography-Mass Spectrometry-Mass Spectrometry (HPLC-MSMS) and Enzyme-Linked Immunosorbent Assay (ELISA) analysis were also performed for comparison purposes, where the electrochemical immunosensor exhibited a wider range of response and a lower detection limit. Matrix effects below 6.5% were obtained using tap water samples. All these characteristics make our electrochemical immunosensor a valid and advantageous tool for the in field determination of IMD.


Assuntos
Anticorpos Monoclonais/imunologia , Técnicas Eletroquímicas/métodos , Imunoensaio/métodos , Inseticidas/análise , Neonicotinoides/análise , Nitrocompostos/análise , Animais , Armoracia/enzimologia , Benzidinas/química , Técnicas Biossensoriais/instrumentação , Técnicas Biossensoriais/métodos , Carbono/química , Bovinos , Compostos Cromogênicos/química , Água Potável/análise , Técnicas Eletroquímicas/instrumentação , Eletrodos , Peroxidase do Rábano Silvestre/química , Inseticidas/imunologia , Limite de Detecção , Neonicotinoides/imunologia , Nitrocompostos/imunologia , Oxirredução , Coelhos , Reprodutibilidade dos Testes , Soroalbumina Bovina/química
13.
Mikrochim Acta ; 186(4): 257, 2019 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-30915577

RESUMO

A highly sensitive colorimetric assay is described for the determination of glutathione (GSH). It is based on the use of tungsten disulfide (WS2) which is a peroxidase mimic. It catalyzes the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) by hydrogen peroxide to form a blue-colored product (oxTMB) at near neutral pH values. In the presence of the analyte GSH, it is transformed to its oxidized form (GSSG), while the blue oxTMB is increasingly reduced and eventually is converted to colorless TMB. This can readily be detected with bare eyes. GSH can be determined spectroscopically (at 652 nm) by this method at concentrations down to 61 pM, and the response is linear in the 100 pM to 10 nM GSH concentration range. The assay is cost-effective and simple. Conceivably, it provides a promising tool for the determination of GSH in food and medical samples. Graphical abstract Schemaric of a sensitive method for colorimetric detection of glutathione (GSH) based on WS2-catalyzed oxidation of TMB by H2O2 and the reduction of blue oxTMB by GSH.


Assuntos
Colorimetria/métodos , Glutationa/análise , Nanoestruturas/química , Sulfetos/química , Compostos de Tungstênio/química , Benzidinas/química , Catálise , Compostos Cromogênicos/química , Peróxido de Hidrogênio/química , Limite de Detecção , Espectrofotometria/métodos
14.
Mikrochim Acta ; 186(4): 250, 2019 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-30888507

RESUMO

It is shown that metallothionein-stabilized copper nanoclusters (MT-CuNCs) display catalase-like activity. In the presence of either lead(II) or mercury(II), the catalase-like activity is converted to a peroxidase-like activity. On addition of Pb(II) or Hg(II), the inhibitory effect of MT-CuNCs on the chromogenic reaction of 3,3',5,5'-tetramethylbenzidine (TMB) with H2O2 is weakened. On the other hand, the catalytic effect of the nanoclusters on the chromogenic reaction is increased. The system MT-CuNCs-Pb(II)/Hg(II) exhibits high affinity for the substrates TMB and H2O2. Their catalytic behavior follows Michaelis-Menten kinetics. Based on these findings, a method was developed for visual detection (via the blue coloration formed) and spectrophotometric determination (at 450 nm) of Pb(II) and Hg(II). The linear range for Pb(II) extends from 0.7 to 96 µM, and the linear ranges for Hg(II) from 97 nM to 2.3 µM and from 3.1 µM to 15.6 µM. The detection limits are 142 nM for Pb(II) and 43.8 nM for Hg(II). Graphical abstract Metallothionein-stabilized copper nanoclusters (MT-CuNCs) display catalase-like activity. On addition of Pb(II) or Hg(II), the catalase-like activity is converted to a peroxidase-like activity. The latter catalyzes the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) by H2O2.


Assuntos
Colorimetria/métodos , Cobre/química , Chumbo/análise , Mercúrio/análise , Nanopartículas Metálicas/química , Metalotioneína/química , Benzidinas/química , Catálise , Compostos Cromogênicos/química , Peróxido de Hidrogênio/química , Cinética , Limite de Detecção , Espectrometria de Fluorescência/métodos
15.
Mikrochim Acta ; 186(4): 234, 2019 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-30859332

RESUMO

A nanocomposite was hydrothermally prepared from C-dots and V2O5 nanowires, and characterized by TEM, FTIR and XRD. Due to the synergistic effects between C-dots and V2O5 nanowires, the nanocomposite is found to possess peroxidase-mimicking activity. This finding was exploited to design colorimetric methods for determination of H2O2 and glucose (via glucose oxidase) by using of 3,3',5,5'-tetramethylbenzidine (TMB) as the chromogenic substrate. The C-dot/V2O5 nanocomposite catalyzes hydrogen peroxide to oxidize TMB and the resultant product, i.e., TMB* produces a blue color in the solution. Also for glucose determination, at first glucose reacts with dissolved oxygen in the presence of glucose oxidase and generates H2O2. Then, produced H2O2 was monitored by the C-dot/V2O5 nanozyme in the presence of TMB. Intensity of the blue color in the solution at wavelength of 650 nm is an indication of H2O2 or glucose concentration. The response to H2O2 is linear in the 0.5-520 µM concentration ranges, and that for glucose from 0.7 µM to 300 µM. Graphical abstract Schematic presentation of peroxidase mimicking activity of C-dot/V2O5 nanocomposite and its application as sensitive colorimetric H2O2/glucose assay by using of 3,3',5,5'-tetramethylbenzidine (TMB) as chromogenic substrate to induce a typical blue color reaction.


Assuntos
Carbono/química , Glucose/análise , Peróxido de Hidrogênio/análise , Nanofios/química , Pontos Quânticos/química , Compostos de Vanádio/química , Benzidinas/química , Materiais Biomiméticos/química , Técnicas Biossensoriais/métodos , Compostos Cromogênicos/química , Colorimetria/métodos , Glucose/química , Glucose Oxidase/química , Limite de Detecção , Nanocompostos/química , Oxirredução , Peroxidase/química
16.
Bioorg Med Chem ; 27(7): 1444-1448, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30795989

RESUMO

We designed a conjugated molecule bearing an O-nitrobenzoxadiazole (O-NBD) unit and an acetylated trimethyl lock as a chromogenic and fluorogenic probe for the detection of esterase activity. The designed molecule was briefly synthesized from a commercially available compound in two steps. Several experiments revealed that the conjugated molecule serves as a sensitive chromogenic and fluorogenic probe for the detection of porcine liver esterase activity. Mechanistic studies indicated that an intramolecular O- to N-NBD migration is involved in the chromogenic/fluorogenic phenomena. The results here would be helpful for designing other O-NBD-based chromogenic/fluorogenic probes in future.


Assuntos
Compostos Cromogênicos/química , Esterases/análise , Corantes Fluorescentes/química , Nitrobenzenos/química , Oxidiazóis/química , Animais , Compostos Cromogênicos/síntese química , Esterases/metabolismo , Corantes Fluorescentes/síntese química , Fígado/enzimologia , Estrutura Molecular , Nitrobenzenos/síntese química , Oxidiazóis/síntese química , Suínos
17.
Org Biomol Chem ; 17(9): 2467-2478, 2019 02 27.
Artigo em Inglês | MEDLINE | ID: mdl-30747188

RESUMO

The neuropeptide Y (NPY) Y4 receptor is a G protein coupled receptor, which is targeted by pancreatic polypeptide, a homologue of NPY. Selective Y4R agonists were suggested as potential therapeutics for the treatment of obesity. Highly potent dimeric peptidic Y4R agonists, constituting two pentapeptide moieties connected through an aliphatic linker, represent an interesting class of Y4R ligands. Based on this compound class, photoresponsive Y4R ligands, containing an azobenzene, azopyrazole, diethienylethene or a fulgimide chromophore were prepared to explore structural requirements of such Y4R agonists on Y4R binding. The synthesized Y4R ligands, containing a non-aliphatic rigid photochromic linker, switch reversibly in aqueous buffer and exhibited high Y4R affinity throughout. This demonstrated that the replacement of the highly flexible aliphatic linker by a considerably less flexible photochromic linker was well tolerated with respect to Y4R binding. Differences in Y4R affinity and activity between the individual photoisomers (varying in spatial orientation and flexibility) were marginal suggesting that the linking element in the dimeric ligands is less critical for the adaptation of high-affinity binding modes at the receptor.


Assuntos
Compostos Cromogênicos/química , Compostos Cromogênicos/farmacologia , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Receptores de Neuropeptídeo Y/metabolismo , Animais , Compostos Azo/química , Compostos Azo/farmacologia , Células CHO , Cricetulus , Humanos , Ligantes , Ligação Proteica , Receptores de Neuropeptídeo Y/agonistas , Relação Estrutura-Atividade
18.
Methods Enzymol ; 614: 87-106, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30611434

RESUMO

Isotopic labeling of recombinantly expressed proteins is generally required for investigation by modern nuclear magnetic resonance (NMR) methods. Purification strategies of the labeled proteins often include the use of a polyhistidine affinity tag (His-tag) and immobilized metal ion affinity chromatography (IMAC). Described herein are rapid and inexpensive qualitative and quantitative assays to determine the concentration of paramagnetic Ni2+ in protein samples purified by IMAC. Both qualitative and quantitative colorimetric methods detect the amount of Ni2+ via the color change produced when a [Ni(PAR)n]2+ (PAR=4-(2-pyridylazo)resorcinol, n=1, 2) complex is formed. The qualitative assay provides a rapid visual test for the presence of Ni2+ in the low micromolar range in a sample of interest. The usefulness of the spectroscopic quantitative assay is illustrated by: (i) detecting a 12µM Ni2+ contamination in an NMR sample containing 950µM of the 7.5kDa α3W protein purified by a standard His-tag Ni2+/IMAC approach and (ii) showing that the 15N-HSQC spectrum of the α3W NMR sample, containing 1 paramagnetic Ni2+ ion per 80 protein molecules, displays clear line broadening of both water and protein spectral lines. We also (iii) measured Ni2+ release during the equilibration, wash, and elution steps of three commonly used Ni2+/IMAC resins when following manufacturer's protocols. The concentration of Ni2+ detected in elutes of the three resins ranged from 2µM to nearly 1mM.


Assuntos
Compostos Cromogênicos/química , Colorimetria/métodos , Complexos de Coordenação/análise , Espectroscopia de Ressonância Magnética/métodos , Níquel/análise , Resorcinóis/química , Cátions Bivalentes , Cromatografia de Afinidade , Complexos de Coordenação/química , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Níquel/química , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Sensibilidade e Especificidade , Tiorredoxinas/química , Tiorredoxinas/genética , Tiorredoxinas/metabolismo
19.
Haemophilia ; 25(1): 162-169, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30488994

RESUMO

BACKGROUND: Both one-stage (OSA) and chromogenic substrate assays (CSA) are used to measure factor VIII (FVIII) activity. Factors explaining analytical variation in FVIII activity levels are still to be completely elucidated. AIM: The aim of this study was to investigate and quantify the analytical variation in OSA and CSA. METHODS: Factors determining analytical variation were studied in sixteen lyophilized plasma samples (FVIII activity <0.01-1.94 IU/mL) and distributed by the ECAT surveys. To elucidate the causes of OSA variation, we exchanged deficient plasma between three company set-ups. RESULTS: On average, 206 (range 164-230) laboratories used the OSA to measure FVIII activity and 30 (range 12-51) used CSA. The coefficient of variation of OSA and CSA increased with lower FVIII levels (FVIII <0.05 IU/mL). This resulted in misclassification of a severe haemophilia A sample into a moderate or mild haemophilia A sample in 4/30 (13.3%) of CSA measurements, while this was 37/139 (26.6%) for OSA. OSA measurements performed with reagents and equipment from Werfen showed slightly lower FVIII activity (0.93, IQR 0.88-0.98 IU/mL) compared to measurements with Stago (1.07, IQR 1.02-1.14 IU/mL) and Siemens (1.03, IQR 0.97-1.07 IU/mL). Part of this difference is explained by the value of the calibrator. For CSA, the measured FVIII levels were similar using the different kits. CONCLUSIONS: In the lower range (<0.05 IU/mL), analytical variation of FVIII measurements is high in both OSA and CSA measurements. The variation in FVIII activity levels was partly explained by specific manufacturers. Further standardization of FVIII measurements and understanding of analytical variation is required.


Assuntos
Testes de Coagulação Sanguínea/métodos , Fator VIII/análise , Hemofilia A/patologia , Testes de Coagulação Sanguínea/normas , Calibragem , Compostos Cromogênicos/química , Fator VIII/normas , Humanos , Plasma/química , Índice de Gravidade de Doença
20.
Molecules ; 23(10)2018 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-30314330

RESUMO

We report experimental tests of whether non-rigid, π-conjugated luminophores in the photoexcited (S1) and ground (S0) states dissolved in achiral liquids are mirror symmetrical by means of circularly polarized luminescence (CPL) and circular dichroism (CD) spectroscopy. Herein, we chose ten oligofluorenes, eleven linear/cyclic oligo-p-arylenes, three binaphthyls and five fused aromatics, substituted with alkyl, alkoxy, phenyl and phenylethynyl groups and also with no substituents. Without exception, all these non-rigid luminophores showed negative-sign CPL signals in the UV-visible region, suggesting temporal generation of energetically non-equivalent non-mirror image structures as far-from equilibrium open-flow systems at the S1 state. For comparison, unsubstituted naphthalene, anthracene, tetracene and pyrene, which are achiral, rigid, planar luminophores, did not obviously show CPL/CD signals. However, camphor, which is a rigid chiral luminophore, showed mirror-image CPL/CD signals. The dissymmetry ratio of CPL (glum) for the oligofluorenes increased discontinuously, ranging from ≈ -(0.2 to 2.0) × 10-3, when the viscosity of the liquids increased. When the fluorene ring number increased, the glum value extrapolated at [η] = 0 reached -0.8 × 10-3 at 420 nm, leading to (⁻)-CPL signals predicted in the vacuum state. Our comprehensive CPL and CD study should provide a possible answer to the molecular parity violation hypothesis arising due to the weak neutral current mediated by the Z°-boson.


Assuntos
Compostos Cromogênicos/química , Dicroísmo Circular , Fluorenos/química , Espectrometria de Fluorescência , Algoritmos , Modelos Químicos , Estrutura Molecular
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