RESUMEN
The in-depth study of single cells requires the dynamically molecular information in one particular nanometer-sized organelle in a living cell, which is difficult to achieve using current methods. Due to high efficiency of click chemistry, a new nanoelectrode-based pipette architecture with dibenzocyclooctyne at the tip is designed to realize fast conjugation with azide group-containing triphenylphosphine, which targets mitochondrial membranes. The covalent binding of one mitochondrion at the tip of the nanopipette allows a small region of the membrane to be isolated on the Pt surface inside the nanopipette. Therefore, the release of reactive oxygen species (ROS) from the mitochondrion is monitored, which is not interfered by the species present in the cytosol. The dynamic tracking of ROS release from one mitochondrion reveals the distinctive "ROS-induced ROS release" within the mitochondria. Further study of RSL3-induced ferroptosis using nanopipettes provides direct evidence for supporting the noninvolvement of glutathione peroxidase 4 in the mitochondria during RSL3-induced ROS generation, which has not previously been observed at the single-mitochondrion level. Eventually, this established strategy should overcome the existing challenge of the dynamic measurement of one special organelle in the complicated intracellular environment, which opens a new direction for electroanalysis in subcellular analysis.
Asunto(s)
Mitocondrias , Supervivencia Celular , Mitocondrias/química , Mitocondrias/metabolismo , Química Clic/instrumentación , Química Clic/métodos , Especies Reactivas de Oxígeno/metabolismoRESUMEN
This chapter provides a brief overview of the applications of ZINClick virtual library. In the last years, we have investigated the click-chemical space covered by molecules containing the triazole ring and generated a database of 1,2,3-triazoles called ZINClick, starting from literature reported alkynes and azides synthesizable in no more than three synthetic steps from commercially available products. This combinatorial database contains millions of 1,4-disubstituted 1,2,3-triazoles that are easily synthesizable. The library is regularly updated and can be freely downloaded from http://www.ZINClick.org . This virtual library is a good starting point to explore a new portion of chemical space.
Asunto(s)
Química Clic/métodos , Descubrimiento de Drogas/métodos , Triazoles/química , Alquinos/química , Azidas/química , Química Clic/instrumentación , Bases de Datos de Compuestos Químicos , Diseño de Fármacos , Bibliotecas de Moléculas Pequeñas/química , Programas InformáticosRESUMEN
We report a covalent chemistry-based hepatocellular carcinoma (HCC)-specific extracellular vesicle (EV) purification system for early detection of HCC by performing digital scoring on the purified EVs. Earlier detection of HCC creates more opportunities for curative therapeutic interventions. EVs are present in circulation at relatively early stages of disease, providing potential opportunities for HCC early detection. We develop an HCC EV purification system (i.e., EV Click Chips) by synergistically integrating covalent chemistry-mediated EV capture/release, multimarker antibody cocktails, nanostructured substrates, and microfluidic chaotic mixers. We then explore the translational potential of EV Click Chips using 158 plasma samples of HCC patients and control cohorts. The purified HCC EVs are subjected to reverse-transcription droplet digital PCR for quantification of 10 HCC-specific mRNA markers and computation of digital scoring. The HCC EV-derived molecular signatures exhibit great potential for noninvasive early detection of HCC from at-risk cirrhotic patients with an area under receiver operator characteristic curve of 0.93 (95% CI, 0.86 to 1.00; sensitivity = 94.4%, specificity = 88.5%).
Asunto(s)
Biomarcadores de Tumor/aislamiento & purificación , Carcinoma Hepatocelular/diagnóstico , Detección Precoz del Cáncer/métodos , Vesículas Extracelulares/genética , Neoplasias Hepáticas/diagnóstico , Anciano , Biomarcadores de Tumor/genética , Carcinoma Hepatocelular/sangre , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Estudios de Casos y Controles , Química Clic/instrumentación , Química Clic/métodos , Química Computacional , Simulación por Computador , Diagnóstico Diferencial , Dimetilpolisiloxanos/química , Progresión de la Enfermedad , Detección Precoz del Cáncer/instrumentación , Femenino , Células Hep G2 , Humanos , Dispositivos Laboratorio en un Chip , Biopsia Líquida/instrumentación , Biopsia Líquida/métodos , Cirrosis Hepática/sangre , Cirrosis Hepática/patología , Neoplasias Hepáticas/sangre , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , Masculino , Técnicas Analíticas Microfluídicas/instrumentación , Técnicas Analíticas Microfluídicas/métodos , Persona de Mediana Edad , Nanoestructuras/química , Nanocables/química , Estadificación de Neoplasias , ARN Mensajero/genética , ARN Mensajero/aislamiento & purificación , Curva ROC , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/instrumentación , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodosRESUMEN
Glycan microarrays have become a powerful technology to study biological processes, such as cell-cell interaction, inflammation, and infections. Yet, several challenges, especially in multivalent display, remain. In this introductory lecture we discuss the state-of-the-art glycan microarray technology, with emphasis on novel approaches to access collections of pure glycans and their immobilization on surfaces. Future directions to mimic the natural glycan presentation on an array format, as well as in situ generation of combinatorial glycan collections, are discussed.
Asunto(s)
Análisis por Micromatrices/métodos , Polisacáridos/análisis , Animales , Bioimpresión/instrumentación , Bioimpresión/métodos , Química Clic/instrumentación , Química Clic/métodos , Diseño de Equipo , Glicómica/instrumentación , Glicómica/métodos , Humanos , Análisis por Micromatrices/instrumentaciónRESUMEN
Homology-directed DNA repair (HDR) is an evolutionary conserved mechanism that is required for genome integrity and organismal fitness across species. While a myriad of different factors and mechanisms are able to execute HDR, all forms necessitate common steps of DNA damage recognition, homology search and capture, and assembly of a DNA polymerase complex to conduct templated DNA synthesis. The central question of what determines HDR mechanism utilization in mammalian cells has been limited by an inability to directly monitor the DNA damage response and products of repair as they arise from a defined genomic lesion. In this chapter, we describe several methodologies to delineate major steps of HDR during alternative lengthening of telomeres in human cells. This includes procedures to visualize interchromosomal telomere homology searches in real time and quantitatively detect HDR synthesis of nascent telomeres emanating from synchronous activation of telomere DNA double-strand breaks. We highlight the critical details of these methods and their applicability to monitoring HDR at telomeres in a broad variety of mammalian cell types.
Asunto(s)
ADN/análisis , Desoxiuridina/análisis , Microscopía Intravital/métodos , Reparación del ADN por Recombinación , Telómero/metabolismo , Técnicas de Cultivo de Célula/instrumentación , Técnicas de Cultivo de Célula/métodos , Línea Celular Tumoral , Química Clic/instrumentación , Química Clic/métodos , ADN/química , ADN/metabolismo , Roturas del ADN de Doble Cadena , Replicación del ADN , Humanos , Microscopía Intravital/instrumentación , Microscopía Fluorescente/instrumentación , Microscopía Fluorescente/métodos , Imagen Individual de Molécula/instrumentación , Imagen Individual de Molécula/métodosRESUMEN
Genomics is a disruptive technology, having revealed that cancers are tremendously complex and differ from patient to patient. Therefore, conventional treatment approaches fit poorly with genomic reality. Furthermore, it is likely that this type of complexity will also be observed in other illnesses. Precision medicine has been posited as a way to better target disease-related aberrations, but developing drugs and tailoring therapy to each patient's complicated problem is a major challenge. One solution would be to match patients to existing compounds based on in silico modeling. However, optimization of complex therapy will eventually require designing compounds for patients using computer modeling and just-in-time production, perhaps achievable in the future by three-dimensional (3D) printing. Indeed, 3D printing is potentially transformative by virtue of its ability to rapidly generate almost limitless numbers of objects that previously required manufacturing facilities. Companies are already endeavoring to develop affordable 3D printers for home use. An attractive, but as yet scantily explored, application is to place chemical design and production under digital control. This could be accomplished by utilizing a 3D printer to initiate chemical reactions, and print the reagents and/or the final compounds directly. Of interest, the Food and Drug Administration (FDA) has recently approved a 3D printed drug-levetiracetam-indicated for seizures. Further, it is now increasingly clear that biologic materials-tissues, and eventually organs-can also be "printed." In the near future, it is plausible that high-throughput computing may be deployed to design customized drugs, which will reshape medicine.
Asunto(s)
Antineoplásicos/síntesis química , Química Clic/métodos , Simulación por Computador , Neoplasias/tratamiento farmacológico , Medicina de Precisión/métodos , Impresión Tridimensional , Anticonvulsivantes/síntesis química , Química Clic/instrumentación , Genómica/métodos , Humanos , Imagenología Tridimensional , Levetiracetam , Piracetam/análogos & derivados , Piracetam/síntesis químicaRESUMEN
A novel fluoresencent immunosensor for determination of cancer biomarkers such as alpha-fetoprotein (AFP) was designed by utilizing both the high specificity of antigen-antibody sandwich structure and the high sensitivity of the click chemistry based fluorescence detection. Instead of an enzyme or fluorophore, the CuO nanoparticles are labeled on the detection antibody, which was not susceptible to the change of the external environments. The CuO nanoparticles which were modified on the sandwich structure can be dissolved to produce Cu(2+) ions with the help of HCl and then the Cu(2+) ions were reduced by sodium ascorbate to produce Cu(+) ions which triggered the Cu(+) catalyzed alkyne-azide cycloaddition (CuAAC) reaction between the weak fluorescent compound (3-azido-7-hydroxycoumarin) and propargyl alcohol to form a strong fluorescent compound. A good linear relationship was observed between the fluorescence increase factor of the system and the concentration of AFP in the range of 0.025-5.0 ng/mL with a detection limit of 12 pg/mL (S/N=3). The proposed fluorescent sensor had been applied to detect AFP in the human serum samples and gave satisfactory results.
Asunto(s)
Técnicas Biosensibles/instrumentación , Química Clic/instrumentación , Inmunoensayo/instrumentación , Espectrometría de Fluorescencia/instrumentación , alfa-Fetoproteínas/análisis , Diseño de Equipo , Análisis de Falla de Equipo , Humanos , Nanopartículas del Metal/químicaRESUMEN
Prevention of the occurrence and development of inflammation is a vital therapeutic strategy for treating acute lung injury (ALI). Increasing evidence has shown that a wealth of ingredients from natural foods and plants have potential anti-inflammatory activity. In the present study, mangiferin, a natural C-glucosyl xanthone that is primarily obtained from the peels and kernels of mango fruits and the bark of the Mangifera indica L. tree, alleviated the inflammatory responses in lipopolysaccharide (LPS)-induced ALI mice. Mangiferin-modified magnetic microspheres (MMs) were developed on the basis of click chemistry to capture the target proteins of mangiferin. Mass spectrometry and molecular docking identified 70 kDa heat-shock protein 5 (Hspa5) and tyrosine 3-monooxygenase (Ywhae) as mangiferin-binding proteins. Furthermore, an enzyme-linked immunosorbent assay (ELISA) indicated that mangiferin exerted its anti-inflammatory effect by binding Hspa5 and Ywhae to suppress downstream mitogen-activated protein kinase (MAPK) signaling pathways. Thoroughly revealing the mechanism and function of mangiferin will contribute to the development and utilization of agricultural resources from M. indica L.
Asunto(s)
Lesión Pulmonar Aguda/tratamiento farmacológico , Lesión Pulmonar Aguda/metabolismo , Mangifera/química , Extractos Vegetales/administración & dosificación , Xantonas/administración & dosificación , Proteínas 14-3-3/genética , Proteínas 14-3-3/metabolismo , Lesión Pulmonar Aguda/genética , Animales , Química Clic/instrumentación , Química Clic/métodos , Chaperón BiP del Retículo Endoplásmico , Frutas/química , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Proteínas Quinasas Activadas por Mitógenos/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismoRESUMEN
We describe a protocol for the rapid labeling of cell-surface proteins in living mammalian cells using click chemistry. The labeling method is based on strain-promoted alkyne-azide cycloaddition (SPAAC) and strain-promoted inverse-electron-demand Diels-Alder cycloaddition (SPIEDAC) reactions, in which noncanonical amino acids (ncAAs) bearing ring-strained alkynes or alkenes react, respectively, with dyes containing azide or tetrazine groups. To introduce ncAAs site specifically into a protein of interest (POI), we use genetic code expansion technology. The protocol can be described as comprising two steps. In the first step, an Amber stop codon is introduced--by site-directed mutagenesis--at the desired site on the gene encoding the POI. This plasmid is then transfected into mammalian cells, along with another plasmid that encodes an aminoacyl-tRNA synthetase/tRNA (RS/tRNA) pair that is orthogonal to the host's translational machinery. In the presence of the ncAA, the orthogonal RS/tRNA pair specifically suppresses the Amber codon by incorporating the ncAA into the polypeptide chain of the POI. In the second step, the expressed POI is labeled with a suitably reactive dye derivative that is directly supplied to the growth medium. We provide a detailed protocol for using commercially available ncAAs and dyes for labeling the insulin receptor, and we discuss the optimal surface-labeling conditions and the limitations of labeling living mammalian cells. The protocol involves an initial cloning step that can take 4-7 d, followed by the described transfections and labeling reaction steps, which can take 3-4 d.
Asunto(s)
Aminoácidos/química , Química Clic/métodos , Colorantes Fluorescentes/química , Proteínas/química , Alquinos/química , Aminoacil-ARNt Sintetasas/genética , Aminoacil-ARNt Sintetasas/metabolismo , Animales , Azidas/química , Carbocianinas/química , Química Clic/instrumentación , Codón de Terminación , Reacción de Cicloadición , Proteínas Fluorescentes Verdes/genética , Células HEK293 , Humanos , Mamíferos , Mutagénesis Sitio-Dirigida , Proteínas/genética , Receptor de Insulina/química , Receptor de Insulina/genéticaRESUMEN
Herein, we report the "ene-thiol" photografting of 1-octadecanethiol onto vinyl pre-functionalized silica monolith to prepare clicked reversed-phase silica monolithic columns with high permeability and performances. The experimental conditions (concentration of thiol in solution, irradiation duration) are optimized with respect to highest retention and methylene selectivity, i.e. to the highest surface coverage of the monolith. It is demonstrated that an irradiation duration of 5min is enough with a 0.8M concentration of 1-octadecanethiol in solution or that it may be replaced by successive irradiations at a lower ODT concentration (0.19M) with renewing of the solution between the illuminations. Retention factors as high as those obtained with standard silanization are reached while keeping the intrinsic monolith permeability and efficiency (160,000plates/m in nano-LC at 0.7mm/s). The absence of polymerization, in the "ene-thiol" version, is demonstrated. Indeed, the steric selectivity of our clicked-material is characteristic of monolayer-like functionalized silica and significantly lower than the steric selectivity measured on polymeric-like functionalized silica.
Asunto(s)
Química Clic/instrumentación , Dióxido de Silicio/química , Química Clic/métodos , Polimerizacion , Compuestos de Sulfhidrilo/químicaRESUMEN
A novel glutathione (GSH)-silica hybrid monolithic column synthesized via a combination of thiol-ene click reaction and one-pot process was described, where thiol-end GSH organic monomer and 2,2-azobisisobutyronitrile (AIBN) were mixed with hydrolyzed tetramethyloxysilane (TMOS) and γ-methacryloxypropyltrimethoxysilane (γ-MAPS) and then introduced into a fused-silica capillary for simultaneous polycondensation and "thiol-ene" click reaction to form the GSH-silica hybrid monolith. The effects of the molar ratio of TMOS/γ-MAPS, the amount of GSH, and the volume of porogen on the morphology, permeability and pore properties of the prepared GSH-silica hybrid monoliths were studied in detail. A uniform monolithic network with high porosity was obtained. A series of test compounds including alkylbenzenes, amides, and anilines were used to evaluate the retention behaviors of the GSH-silica hybrid monolithic column. The results demonstrated that the prepared GSH-silica hybrid monolith exhibited multiple interactions including hydrophobicity, hydrophilicity, as well as cation exchange interaction. The run-to-run, column-to-column and batch-to-batch reproducibilities of the GSH-silica hybrid monolith for phenols' retention were satisfactory with the relative standard deviations (RSDs) less than 1.3% (n=5), 2.6% (n=3) and 3.2% (n=3), respectively, indicating the effectiveness and practicability of the proposed method. In addition, the GSH-silica hybrid monolith was applied to the separation of nucleotides, peptides and protein tryptic digests, respectively. The successful applications suggested the potential of the GSH-silica hybrid monolith in complex sample analysis.
Asunto(s)
Cromatografía Líquida de Alta Presión/métodos , Química Clic/métodos , Glutatión/química , Dióxido de Silicio/química , Cromatografía Líquida de Alta Presión/instrumentación , Química Clic/instrumentación , Interacciones Hidrofóbicas e Hidrofílicas , Metacrilatos/química , Microscopía Electrónica de Rastreo , Péptidos/aislamiento & purificación , Proteínas/aislamiento & purificación , Silanos/química , Compuestos de Sulfhidrilo/químicaRESUMEN
The analysis of protein glycosylation is important for biomedical and biopharmaceutical research. Recent advances in LC-MS analysis have enabled the identification of glycosylation sites, the characterisation of glycan structures and the identification and quantification of glycoproteins and glycopeptides. However, this type of analysis remains challenging due to the low abundance of glycopeptides in complex protein digests, the microheterogeneity at glycosylation sites, ion suppression effects and the competition for ionisation by co-eluting peptides. Specific sample preparation is necessary for comprehensive and site-specific glycosylation analyses using MS. Therefore, researchers continue to pursue new columns to broaden their applications. The current manuscript covers recent literature published from 2008 to 2013. The stationary phases containing various chemical bonding methods or ligands immobilisation strategies on solid supports that selectively enrich N-linked or sialylated N-glycopeptides are categorised with either physical or chemical modes of binding. These categories include lectin affinity, hydrophilic interactions, boronate affinity, titanium dioxide affinity, hydrazide chemistry and other separation techniques. This review should aid in better understanding the syntheses and physicochemical properties of each type of stationary phases for enriching glycoproteins and glycopeptides.
Asunto(s)
Cromatografía , Glicopéptidos , Glicoproteínas , Animales , Ácidos Borónicos , Cromatografía/instrumentación , Cromatografía/métodos , Química Clic/instrumentación , Química Clic/métodos , Glicopéptidos/análisis , Glicopéptidos/aislamiento & purificación , Glicoproteínas/análisis , Glicoproteínas/aislamiento & purificación , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Lectinas , RatonesRESUMEN
A novel fluorescent sensor for DNA sequence has been designed by taking advantages of copper nanoparticles (CuNPs) selectively formed on double stranded (ds) DNA template and Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) reaction. Copper(II) is derived from CuNPs which previously formed on the dsDNA template, and then copper(II) is reduced to copper(I) by ascorbate, which in turn induced CuAAC reaction between the weak-fluorescent compound (3-azido-7-hydroxycoumarin) and propargyl alcohol to form strong fluorescence compounds (1,2,3-triazole compounds). Since CuNPs are accumulated efficiently in the major groove of dsDNA and ssDNA has no groove, it indicates that the proposed sensor owns the merits of low detection limit, high sensitivity and selectivity for mutational p53 sequence detection. Additionally, the method has been successfully applied to recognize the sequence which contains a single-base mismatch in the short human p53 gene fragment. Furthermore, it has also been applied to detect DNA sequence in complex medium (hela cellular homogenate) with satisfactory results.
Asunto(s)
Técnicas Biosensibles/instrumentación , Química Clic/instrumentación , Análisis Mutacional de ADN/instrumentación , ADN/genética , Análisis de Secuencia de ADN/instrumentación , Espectrometría de Fluorescencia/instrumentación , Proteína p53 Supresora de Tumor/genética , Secuencia de Bases , Diseño de Equipo , Análisis de Falla de Equipo , Datos de Secuencia Molecular , Polimorfismo de Nucleótido Simple/genética , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
A successful combination of anti-biofouling and ion-interaction for endotoxin (ET) selective removal from protein solutions is realized via the immobilization of two ordinary polymers, PEG and PLL, which have protein resistance and endotoxin adsorption properties, respectively, together onto azide-functional polystyrene (PS-N3 ) microspheres (MSs) using a surface click reaction. Toxic copper ions residu is successfully minimized by the addition of 2,2'-bipyridine (BiPy).
Asunto(s)
Endotoxinas/aislamiento & purificación , Ácido Láctico/química , Polietilenglicoles/química , Polímeros/química , Proteínas/química , Proteínas/aislamiento & purificación , Ultrafiltración/instrumentación , Ultrafiltración/métodos , Absorción , Química Clic/instrumentación , Química Clic/métodos , Mezclas Complejas/aislamiento & purificación , Endotoxinas/química , Diseño de Equipo , Análisis de Falla de Equipo , Iones , Poliésteres , SolucionesRESUMEN
A novel colorimetric copper(II) biosensor has been developed based on the high specificity of alkyne-azide click reaction to the catalysis of copper ions and unmodified gold nanoparticles (AuNPs) as the signal reporter. The clickable DNA probe consists of two parts: an azide group-modified double-stranded DNA (dsDNA) hybrid with an elongated tail and a short alkyne-modified single-stranded DNA (ssDNA). Because of low melting temperature of the short ssDNA, these two parts are separated in the absence of Cu(2+). Copper ion-induced azide-alkyne click ligation caused a structural change of probe from the separated form to entire dsDNA form. This structural change of probe can be monitored by the unmodified AuNPs via mediating their aggregation with a red-to-blue colorimetric read-out because of the differential ability of ssDNA and dsDNA to protect AuNPs against salt-induced aggregation. Under the optimum conditions, this biosensor can sensitively and specifically detect Cu(2+) with a low detection limit of 250 nM and a linear range of 0.5-10 µM. The method is simple and economic without dual-labeling DNA and AuNPs modification. It is also highly selective for Cu(2+) in the presence of high concentrations of other environmentally relevant metal ions because of the great specificity of the copper-caused alkyne-azide click reaction, which potentially meets the requirement of the detection in real samples.
Asunto(s)
Técnicas Biosensibles/instrumentación , Química Clic/instrumentación , Colorimetría/instrumentación , Cobre/análisis , ADN/química , Oro/química , Nanopartículas del Metal/química , Monitoreo del Ambiente/instrumentación , Diseño de Equipo , Análisis de Falla de Equipo , Iones , Nanopartículas del Metal/ultraestructura , Nanotecnología/instrumentación , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Contaminantes Químicos del Agua/análisisRESUMEN
An important prerequisite for molecular shuttle-based functional devices is the development of adequate linker chemistries to load and transport versatile cargoes. Copper-free "click chemistry" has not been applied before to covalently load cargo onto molecular shuttles propelled by biological motors such as kinesin. Due to the high biocompatibility and bioorthogonality of the strain-promoted azide-alkyne cycloaddition, this approach has pronounced advantages compared to previous methods.
Asunto(s)
Química Clic/métodos , Proteínas Motoras Moleculares/química , Adenosina Trifosfato/metabolismo , Alquinos/química , Azidas/química , Transporte Biológico , Química Clic/instrumentación , Cobre , Hidrólisis , Cinesinas/química , Ensayo de Materiales/métodos , Microscopía Fluorescente , Microtúbulos/metabolismoRESUMEN
L-histidine is a promising alternative to expensive protein ligands for the adsorption of IgG due to its high selectivity, no toxicity and low cost; while click chemistry can improve the reaction selectivity between the ligands and the support matrix under mild reaction conditions. Thus, using L-histidine as a ligand and original sepharose gel as a support, a novel immunoadsorbent possessing pseudo-biospecific affinity for IgG from human plasma, Sep-triazole-His, was designed and prepared according to the principle of Click-reaction between alkyne and azide functional groups; while both sepharose-based control samples Sep-His and Sep-PA were prepared by a conventional method using L-histidine and protein A as a ligand, respectively. The ligand density and IgG adsorption performance of Sep-triazole-His from human plasma were measured and evaluated. The influences of click chemistry on the preparation, structure and performance of sepharose-based immunoadsorbent were also investigated. The results indicate that the ligand density immobilized on Sep-triazole-His is 319.1 µmol/g sepharose gel, almost 4-fold as high as that on Sep-His; the IgG adsorption capacity of Sep-triazole-His from human plasma reaches 16.49 mg/g at pH 7.0, or increases 5.72-fold with respect to Sep-His, and does not decrease noticeably after being repeatedly used for 10 times; and Sep-triazole-His can exhibit high adsorption selectivity for IgG comparable to Sep-PA. The further studies prove that the 1,2,3-triazole ring in the spacer-arm of Sep-triazole-His, can facilitate the binding of IgG without non-specific adsorption.
Asunto(s)
Cromatografía de Afinidad/métodos , Química Clic/métodos , Inmunoglobulina G/química , Inmunoadsorbentes/química , Adsorción , Cromatografía de Afinidad/instrumentación , Química Clic/instrumentación , Histidina/química , Humanos , Inmunoglobulina G/aislamiento & purificación , Cinética , Ligandos , Especificidad de la EspecieRESUMEN
"Click chemistry" is defined as a class of robust and selective chemical reactions affording high yields and is tolerant to a variety of solvents (including water), functional groups, and air. In this study, click chemistry was used as an effective strategy for coupling three alkyne-carboxylic acids onto the azide-silica to obtain three novel stationary phases of weak cation exchange chromatography, which were characterized with FTIR and elemental analysis. Six kinds of standard proteins, such as myoglobin, RNase A, RNase B, cytochrome C, α-chymotrypsin A, and lysozyme, were separated completely with the three novel weak cation exchange chromatography stationary phases. Compared with commercial weak cation exchange chromatography columns, the three kinds of novel weak cation exchange chromatography packings prepared by click chemistry approach have better resolution and selectivity. The mass recovery of more than 97% was obtained for all the tested proteins, and the bioactivity recovery of lysozyme on the prepared column was determined to be 96%. In addition, lysozyme was purified successfully from egg white with the novel weak cation exchange chromatography column by one step. The purity was more than 97% and a high specific activity was achieved to be 81 435 U/mg. The results illustrate the potential of click chemistry for preparing stationary phase for ion-exchange chromatography.
Asunto(s)
Resinas de Intercambio de Catión/química , Cromatografía por Intercambio Iónico/métodos , Química Clic/instrumentación , Proteínas/aislamiento & purificación , Adsorción , Animales , Cromatografía por Intercambio Iónico/instrumentación , Química Clic/métodos , Humanos , Proteínas/químicaRESUMEN
A simple colorimetric method for the detection of copper ions in water is described. This method is based on the 'click' copper(I)-catalyzed azide-alkyne cycloaddition reaction and its use in promoting the aggregation of azide-tagged gold nanoparticles by a dialkyne cross-linker is described. Nanoparticle cross-linking, evidenced as a colour change, is used for the detection of copper ions. The lowest detected concentration by the naked eye was 1.8 µM, with the response linear with log(concentration) between 1.8-200 µM. The selectivity relative to other potentially interfering ions was evaluated.
Asunto(s)
Técnicas Biosensibles/métodos , Química Clic/métodos , Cobre/análisis , Oro/química , Nanopartículas del Metal/química , Alquinos/química , Azidas/química , Técnicas Biosensibles/instrumentación , Catálisis , Cationes Bivalentes/análisis , Química Clic/instrumentación , Colorimetría/métodos , Reactivos de Enlaces Cruzados/química , Ciclización , Límite de Detección , Microscopía Electrónica de Rastreo/métodos , Abastecimiento de Agua/análisis , Abastecimiento de Agua/normasRESUMEN
Ultrasound (US), a nonconventional energy source, has become a very popular and useful technology in organic chemistry. Several examples of US-assisted reaction have indicated that high yields and short reaction times are reasonable, and applications of this energy transfer process on click reactions have been published. Stable, water-soluble copper nanoparticles (Cu NPs) were synthesized using a double-hydrophilic block copolymer as a template. The prepared Cu NPs were effective as catalysts for the [3+2] cycloaddition of azides with terminal alkynes to provide products in good yields and with high regioselectivity. An environment-friendly process of synthesizing regioselective triazoles was developed from a variety of azides with terminal alkynes, in which the water-soluble Cu NPs were utilized, under 10 min sonication. The Cu NPs can be reused three times under the present reaction conditions, without any loss of catalytic activity. Block copolymer nanoreactors were shown to be promising in creating a number of water-soluble catalytic metal NPs for a wide variety of organic transformations.