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1.
AAPS PharmSciTech ; 22(5): 169, 2021 Jun 02.
Artigo em Inglês | MEDLINE | ID: mdl-34080086

RESUMO

Lutein has been used as a dietary supplement for the treatment of eye diseases, especially age-related macular degeneration. For oral formulations, we investigated lutein stability in artificial set-ups mimicking different physiological conditions and found that lutein was degraded over time under acidic conditions. To enhance the stability of lutein upon oral intake, we developed enteric-coated lutein solid dispersions (SD) by applying a polymer, hydroxypropyl methylcellulose acetate succinate (HPMCAS-LF), through a solvent-controlled precipitation method. The SD were characterized in crystallinity, morphology, and drug entrapment. In the dissolution profile of lutein SD, a F80 formulation showed resistance toward the acidic environment under simulated gastric conditions while exhibiting a bursting drug release under simulated intestinal conditions. Our results highlight the potential use of HPMCAS-LF as an effective matrix to enhance lutein bioavailability during oral delivery and to provide novel insights into the eye-care supplement industry, with direct benefits for the health of patients.


Assuntos
Luteína/síntese química , Luteína/farmacocinética , Metilcelulose/análogos & derivados , Disponibilidade Biológica , Cromatografia Líquida de Alta Pressão/métodos , Liberação Controlada de Fármacos , Estabilidade de Medicamentos , Humanos , Metilcelulose/síntese química , Metilcelulose/farmacocinética , Polímeros/síntese química , Polímeros/farmacocinética , Solubilidade , Solventes , Difração de Raios X/métodos
2.
Molecules ; 26(6)2021 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-33802031

RESUMO

Nimesulide (NIM, N-(4-nitro-2-phenoxyphenyl)methanesulfonamide) is a relatively new nonsteroidal anti-inflammatory analgesic drug. It is practically insoluble in water (<0.02 mg/mL). This very poor aqueous solubility of the drug may lead to low bioavailability. The objective of the present study was to investigate the possibility of improving the solubility and the bioavailability of NIM via complexation with polysaccharide arabinogalactan (AG), disodium salt of glycyrrhizic acid (Na2GA), hydroxypropyl-ß-cyclodextrin (HP-ß-CD) and MgCO3. Solid dispersions (SD) have been prepared using a mechanochemical technique. The physical properties of nimesulide SD in solid state were characterized by differential scanning calorimetry and X-ray diffraction studies. The characteristics of the water solutions which form from the obtained solid dispersions were analyzed by reverse phase and gel permeation HPLC. It was shown that solubility increases for all complexes under investigation. These phenomena are obliged by complexation with auxiliary substances, which was shown by 1H-NMR relaxation methods. The parallel artificial membrane permeability assay (PAMPA) was used for predicting passive intestinal absorption. Results showed that mechanochemically obtained complexes with polysaccharide AG, Na2GA, and HP-ß-CD enhanced permeation of NIM across an artificial membrane compared to that of the pure NIM. The complexes were examined for anti-inflammatory activity on a model of histamine edema. The substances were administered per os to CD-1 mice. As a result, it was found that all investigated complexes dose-dependently reduce the degree of inflammation. The best results were obtained for the complexes of NIM with Na2GA and HP-ß-CD. In noted case the inflammation can be diminished up to 2-fold at equal doses of NIM.


Assuntos
Galactanos/farmacologia , Sulfonamidas/química , Sulfonamidas/farmacologia , 2-Hidroxipropil-beta-Ciclodextrina/química , Animais , Anti-Inflamatórios não Esteroides/química , Disponibilidade Biológica , Varredura Diferencial de Calorimetria/métodos , Química Farmacêutica/métodos , Galactanos/química , Ácido Glicirrízico/química , Magnésio/química , Masculino , Camundongos , Permeabilidade , Preparações Farmacêuticas , Solubilidade , Difração de Raios X/métodos , beta-Ciclodextrinas/química
3.
Molecules ; 26(5)2021 Mar 08.
Artigo em Inglês | MEDLINE | ID: mdl-33800455

RESUMO

The thin film of N-doped ZnO/CNT nanocomposite was successfully fabricated on soda lime glass substrate by a simple sol-gel drop-coating method. The structural, morphological, chemical, and optical properties of as prepared samples were characterized by a variety of tools such as X-ray Diffraction (XRD), Field Emission Scanning Electron Microscopy (FE-SEM), Fourier Transform Infrared spectroscopy (FT-IR), and UV-visible spectroscopy. The hexagonal crystalline structure was confirmed from XRD measurement without any other impurity phase detection in samples. The N-doped ZnO/CNT composite showed excellent photo-catalytic activity towards cationic methylene blue (MB) dye degradation with 100% removal rate under UV light irradiation as compared to N-doped ZnO (65%) and pure ZnO (47.36%). The convincing performance has also been observed for the case of visible light irradiation. The enhancement of that photocatalytic activity might be due to narrowing the band gap as well as the reduction of electron-hole pair recombination in ZnO matrix with the incorporation of dopant nitrogen and CNT. It is assumed from the obtained results that N-doped ZnO/CNT nanocomposite thin film can be employed as an economically achievable and ecofriendly method to degrade dye with UV and visible light irradiation. Additionally, density functional theory (DFT) calculations were applied to explore the effect of N-doping on electronic structure of ZnO. The computational study has supported the experimental results of significant band gap contraction, which leads to the maximum absorption towards higher wavelength and no appreciable change of lattice parameters after doping. A conceivable photocatalytic mechanism of N-doped ZnO/CNT nanocomposite has been proposed as well.


Assuntos
Nanocompostos/química , Nitrogênio/química , Óxido de Zinco/química , Catálise , Contaminação de Medicamentos , Luz , Microscopia Eletrônica de Varredura/métodos , Processos Fotoquímicos , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Raios Ultravioleta , Difração de Raios X/métodos
4.
Molecules ; 26(5)2021 Mar 09.
Artigo em Inglês | MEDLINE | ID: mdl-33803244

RESUMO

Cellulose, as a natural polymer with an abundant source, has been widely used in many fields including the electric field responsive medium that we are interested in. In this work, cellulose micron particles were applied as an electrorheological (ER) material. Because of the low ER effect of the raw cellulose, a composite particle of cellulose and Laponite was prepared via a dissolution-regeneration process. Scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy (FT-IR) and X-ray diffraction (XRD) were used to observe the morphologies and structures of the composite particles, which were different from pristine cellulose and Laponite, respectively. The ER performances of raw cellulose and the prepared composite were measured by an Anton Paar rotational rheometer. It was found that the ER properties of the composite were more superior to those of raw cellulose due to the flake-like shapes of the composite particles with rough surface. Moreover, the sedimentation stability of composite improves drastically, which means better suspension stability.


Assuntos
Celulose/química , Eletricidade , Silicatos/química , Microscopia Eletrônica de Varredura/métodos , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Suspensões/química , Difração de Raios X/métodos
5.
Molecules ; 26(6)2021 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-33799864

RESUMO

Green synthesis of nanomaterials is advancing due to its ease of synthesis, inexpensiveness, nontoxicity and renewability. In the present study, an eco-friendly biogenic method was developed for the green synthesis of nickel oxide nanoparticles (NiONPs) using phytochemically rich Berberis balochistanica stem (BBS) extract. The BBS extract was rich in phenolics, flavonoids and berberine. These phytochemicals successfully reduced and stabilised the NiNO3 (green) into NiONPs (greenish-gray). BBS-NiONPs were confirmed by using UV-visible spectroscopy (peak at 305 nm), X-ray diffraction (size of 31.44 nm), Fourier transform infrared spectroscopy (identified -OH group and Ni-O formation), energy dispersive spectroscopy (showed specified elemental nature) and scanning electron microscopy (showed rhombohedral agglomerated shape). BBS-NiONPs were exposed to multiple in vitro bioactivities to ascertain their beneficial biological applications. They exhibited strong antioxidant activities: total antioxidant capacity (64.77%) and 2, 2-diphenyl-1-picrylhydrazyl (71.48%); and cytotoxic potential: Brine shrimp cytotoxicity assay with IC50 (10.40 µg/mL). BBS-NiONPs restricted the bacterial and fungal pathogenic growths at 1000, 500 and 100 µg/mL. Additionally, BBS-NiONPs showed stimulatory efficacy by enhancing seed germination rate and seedling growth at 31.25 and 62.5 µg/mL. In aggregate, BBS extract has a potent antioxidant activity which makes the green biosynthesis of NiONPs easy, economical and safe. The biochemical potential of BBS-NiONPs can be useful in various biomedical and agricultural fields.


Assuntos
Berberis/metabolismo , Química Verde/métodos , Nanopartículas Metálicas/química , Antibacterianos/química , Antioxidantes/química , Bactérias , Berberis/fisiologia , Testes de Sensibilidade Microbiana , Nanotecnologia/métodos , Níquel/química , Tamanho da Partícula , Compostos Fitoquímicos/química , Extratos Vegetais/química , Espectrometria por Raios X/métodos , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Difração de Raios X/métodos
6.
Int J Mol Sci ; 22(9)2021 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-33925955

RESUMO

The FTO protein is involved in a wide range of physiological processes, including adipogenesis and osteogenesis. This two-domain protein belongs to the AlkB family of 2-oxoglutarate (2-OG)- and Fe(II)-dependent dioxygenases, displaying N6-methyladenosine (N6-meA) demethylase activity. The aim of the study was to characterize the relationships between the structure and activity of FTO. The effect of cofactors (Fe2+/Mn2+ and 2-OG), Ca2+ that do not bind at the catalytic site, and protein concentration on FTO properties expressed in either E. coli (ECFTO) or baculovirus (BESFTO) system were determined using biophysical methods (DSF, MST, SAXS) and biochemical techniques (size-exclusion chromatography, enzymatic assay). We found that BESFTO carries three phosphoserines (S184, S256, S260), while there were no such modifications in ECFTO. The S256D mutation mimicking the S256 phosphorylation moderately decreased FTO catalytic activity. In the presence of Ca2+, a slight stabilization of the FTO structure was observed, accompanied by a decrease in catalytic activity. Size exclusion chromatography and MST data confirmed the ability of FTO from both expression systems to form homodimers. The MST-determined dissociation constant of the FTO homodimer was consistent with their in vivo formation in human cells. Finally, a low-resolution structure of the FTO homodimer was built based on SAXS data.


Assuntos
Dioxigenase FTO Dependente de alfa-Cetoglutarato/genética , Dioxigenase FTO Dependente de alfa-Cetoglutarato/metabolismo , Dioxigenase FTO Dependente de alfa-Cetoglutarato/fisiologia , Catálise , Domínio Catalítico , Dioxigenases/genética , Humanos , Ácidos Cetoglutáricos/metabolismo , Processamento de Proteína Pós-Traducional/genética , RNA Mensageiro/genética , Espalhamento a Baixo Ângulo , Relação Estrutura-Atividade , Difração de Raios X/métodos
7.
AAPS PharmSciTech ; 22(3): 106, 2021 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-33719021

RESUMO

Masking the unpleasant taste of the pharmaceutically active ingredients plays a critical role in patient acceptance, particularly for children. This work's primary objective was the preparation of taste-masked ibuprofen microparticles using cocoa butter with the assistance of supercritical fluid technology. Microparticles were prepared by dissolving ibuprofen in melted cocoa butter at 40 °C. The solution was then introduced into a supercritical fluid unit and processed at 10 MPa CO2 pressure for 30 min. The product was collected after depressurizing the system. The effect of the drug to cocoa butter ratio and the supercritical fluid units' configuration on product quality was evaluated and compared with the sample prepared by a conventional method. Physicochemical characterization of the prepared product, including particle size, crystallinity, entrapment efficiency, in vitro drug release, and product taste using a human volunteer panel was conducted. The produced microparticles were in the range of 1.42 to 15.28 µm. The entrapment efficiency of the formulated microparticles ranged from 66 to 81%. The drug:polymer ratio, the configuration of the supercritical fluid unit, and the method of preparation were found to have a critical role in the formulation of ibuprofen microparticles. Taste evaluation using human volunteers showed that microparticles containing 20% drug and processed with supercritical fluid technology were capable of masking the bitter taste of ibuprofen. In conclusion, the dispersion of ibuprofen in cocoa butter using supercritical fluid technology is a a promising innovative method to mask the bitter taste of ibuprofen.


Assuntos
Dióxido de Carbono/química , Cromatografia com Fluido Supercrítico/métodos , Gorduras na Dieta/síntese química , Desenvolvimento de Medicamentos/métodos , Ibuprofeno/síntese química , Paladar/efeitos dos fármacos , Anti-Inflamatórios não Esteroides/administração & dosagem , Anti-Inflamatórios não Esteroides/síntese química , Anti-Inflamatórios não Esteroides/farmacocinética , Gorduras na Dieta/administração & dosagem , Gorduras na Dieta/farmacocinética , Humanos , Ibuprofeno/administração & dosagem , Ibuprofeno/farmacocinética , Tamanho da Partícula , Paladar/fisiologia , Difração de Raios X/métodos
8.
AAPS PharmSciTech ; 22(3): 113, 2021 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-33751276

RESUMO

The present research focuses on the development of a nanoparticulate (nanocrystals-loaded) rapidly dissolving (orodispersible) tablet with improved solubility and bioavailability. The nanosuspension (NS) was prepared by antisolvent sonoprecipitation technique and the optimized NS was lyophilized to obtain nanocrystals (NCs), which were evaluated for various parameters. The nitrendipine (NIT) nanoparticulate orodispersible tablet (N-ODT) was prepared by direct compression method. The optimized N-ODT was evaluated for pre and post compression characteristics, in vivo pharmacokinetic and stability profile. The optimized NS showed a particle size of 505.74 ± 15.48 nm with a polydispersity index (PDI) of 0.083 ± 0.006. The % NIT content in the NCs was found to be 78.4 ± 2.3%. The saturation solubility of NIT was increased remarkably (26.14 times) in comparison to plain NIT, post NCs development. The DSC and p-XRD analysis of NCs revealed the perseverance of the integrity and crystallinity of NIT on lyophilization. The results of micromeritic studies revealed the good flow-ability and compressibility of NCs blend. All the post-compression properties of N-ODT were observed within the standard intended limit. The dispersion, wetting, and disintegration time of the optimized batch of N-ODT was found to be 39 ± 1.13 s, 44.66 ± 1.52 s, and 33.91 ± 0.94 s respectively. The in vitro dissolution study displayed 100.28 ± 2.64% and 100.61 ± 3.3% of NIT released from NCs (in 8 min) and N-ODT (in 6 min) respectively, while conventional NIT tablet took 30 min to release 99.94 ± 1.57% of NIT. The in vivo pharmacokinetic study in rabbits demonstrated significantly (p < 0.05) higher bioavailability of NIT on release from N-ODT than the conventional NIT tablet. Thus, N-ODT could be a promising tool for improving the solubility and bioavailability of NIT and to treat cardiovascular diseases effectively.


Assuntos
Sistemas de Liberação de Medicamentos/métodos , Desenvolvimento de Medicamentos/métodos , Nanopartículas/química , Nanopartículas/metabolismo , Nitrendipino/síntese química , Nitrendipino/farmacocinética , Administração Oral , Animais , Disponibilidade Biológica , Liberação Controlada de Fármacos/fisiologia , Masculino , Nanopartículas/administração & dosagem , Nitrendipino/administração & dosagem , Tamanho da Partícula , Coelhos , Solubilidade , Difração de Raios X/métodos
9.
Molecules ; 26(4)2021 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-33578637

RESUMO

The conformations of the title compounds were determined in solution (NMR and UV-Vis spectroscopy) and in the solid state (FT-IR and XRD), complemented with density functional theory (DFT) in the gas phase. The nonequivalence of the amide protons of these compounds due to the hindered rotation of the C(O)-NH2 single bond resulted in two distinct resonances of different chemical shift values in the aromatic region of their 1H-NMR spectra. Intramolecular hydrogen bonding interactions between the carbonyl oxygen and the sulfonamide hydrogen atom were observed in the solution phase and solid state. XRD confirmed the ability of the amide moiety of this class of compounds to function as a hydrogen bond acceptor to form a six-membered hydrogen bonded ring and a donor simultaneously to form intermolecular hydrogen bonded complexes of the type N-H···O=S. The distorted tetrahedral geometry of the sulfur atom resulted in a deviation of the sulfonamide moiety from co-planarity of the anthranilamide scaffold, and this geometry enabled oxygen atoms to form hydrogen bonds in higher dimensions.


Assuntos
Benzamidas/química , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Sulfonas/química , Compostos de Tosil/química , Difração de Raios X/métodos , Cristalografia por Raios X , Teoria da Densidade Funcional , Ligação de Hidrogênio , Modelos Moleculares , Conformação Molecular , Estrutura Molecular
10.
Life Sci ; 271: 119070, 2021 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-33465388

RESUMO

AIMS: In vivo biodistribution of radio labeled ZrO2 nanoparticles is addressed for better imaging, therapy and diagnosis. Nanoparticles are synthesized by microwave assisted sol-gel technique using Fe3O4 as a stabilizer. Antioxidant assay, hemolytic activity in human blood and biodistribution in rabbits was explored to study the therapeutical as well as in vivo targeted diagnostic applications of as synthesized nanoparticles. MAIN METHODS: Fe3O4 stabilized zirconia nanoparticles are synthesized using microwave assisted sol-gel method. Microwave (MW) powers are varied in the range of 100 to 1000 W. As synthesized nanoparticles are evaluated using different characterizations such as X-ray diffractometer, scanning electron microscope, Raman spectroscopy, impedance analyzer, Vickers micro hardness indenter, FTIR, and UV-Vis spectroscopy. In vitro activity of synthesized nanoparticles is checked in freshly extracted human blood serum. To study biodistribution of Fe3O4 stabilized zirconia nanoparticles in rabbit, technetium-99 m was used for labeling purpose. The labeling efficacy and stability of labeled nanoparticles are also measured with instant thin layer chromatography (ITLC) method. Intravenous injection of 99mTc-Fe3O4 stabilized zirconia nanoparticles (0.2 ml), containing 110 MBq of radioactivity, is performed to study the biodistribution; nanoparticles are injected into the ear vein of animal (rabbit). KEY FINDINGS: Zirconia (ZrO2) nanoparticles (NPs) are stabilized using Fe3O4 that were prepared by means of microwave assisted sol-gel method. Crystallite size (~20 nm) agrees well with the values required to stabilize tetragonal zirconia (t-ZrO2). Volume shrinkage results in high value of hardness (~1369). Dielectric constant values, compatible for biomedical application, are observed for tetragonally stabilized samples. Low value of hemolytic response is observed for Fe3O4 stabilized ZrO2 NPs. 99mTc radio labeled ZrO2 NPs proved to be potential candidate to study biodistribution. Biodistribution studies show stability of radiolabeled NPs in the original suspension as well as in blood serum. CT scan of rabbit is performed for several times to check the biodistribution of NPs with time and survival of rabbit. Results suggest that these NPs can also be used as targeted nanoparticles as well as variants of drug payload carrier. SIGNIFICANCE: Results signify that Fe3O4 stabilized ZrO2 nanoparticles synthesized by microwave assisted sol-gel method may be considered as "all-rounder" nanoplatform and are safe enough to be used in diagnostic as well as therapeutic purposes.


Assuntos
Compostos Férricos/metabolismo , Sequestradores de Radicais Livres/metabolismo , Micro-Ondas , Nanopartículas/metabolismo , Radioimunodetecção/métodos , Zircônio/metabolismo , Animais , Compostos Férricos/síntese química , Humanos , Nanopartículas/química , Coelhos , Tecnécio/metabolismo , Distribuição Tecidual/fisiologia , Difração de Raios X/métodos , Zircônio/química
11.
Nat Chem Biol ; 17(4): 485-491, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33462497

RESUMO

Tryptophan 2C methyltransferase (TsrM) methylates C2 of the indole ring of L-tryptophan during biosynthesis of the quinaldic acid moiety of thiostrepton. TsrM is annotated as a cobalamin-dependent radical S-adenosylmethionine (SAM) methylase; however, TsrM does not reductively cleave SAM to the universal 5'-deoxyadenosyl 5'-radical intermediate, a hallmark of radical SAM (RS) enzymes. Herein, we report structures of TsrM from Kitasatospora setae, which are the first structures of a cobalamin-dependent radical SAM methylase. Unexpectedly, the structures show an essential arginine residue that resides in the proximal coordination sphere of the cobalamin cofactor, and a [4Fe-4S] cluster that is ligated by a glutamyl residue and three cysteines in a canonical CXXXCXXC RS motif. Structures in the presence of substrates suggest a substrate-assisted mechanism of catalysis, wherein the carboxylate group of SAM serves as a general base to deprotonate N1 of the tryptophan substrate, facilitating the formation of a C2 carbanion.


Assuntos
Metiltransferases/metabolismo , Metiltransferases/ultraestrutura , Arginina/química , Catálise , Coenzimas , Proteínas Ferro-Enxofre/metabolismo , Metilação , S-Adenosilmetionina , Streptomycetaceae/genética , Streptomycetaceae/metabolismo , Tioestreptona/biossíntese , Triptofano/metabolismo , Vitamina B 12/química , Difração de Raios X/métodos
12.
Molecules ; 26(3)2021 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-33498952

RESUMO

Herein, we report the syntheses of two lithium-vanadium oxide-fluoride compounds crystallized from the same reaction mixture through a time variation experiment. A low temperature hydrothermal route employing a viscous paste of V2O5, oxalic acid, LiF, and HF allowed the crystallization of one metastable phase initially, Li2VO0.55(H2O)0.45F5⋅2H2O (I), which on prolonged heating transforms to a chemically similar yet structurally different phase, Li3VOF5 (II). Compound I crystallizes in centrosymmetric space group, I2/a with a = 6.052(3), b = 7.928(4), c = 12.461(6) Å, and ß = 103.99(2)°, while compound II crystallizes in a non-centrosymmetric (NCS) space group, Pna21 with a = 5.1173(2), b = 8.612(3), c = 9.346(3) Å. Synthesis of NCS crystals are highly sought after in solid-state chemistry for their second-harmonic-generation (SHG) response and compound II exhibits SHG activity albeit non-phase-matchable. In this article, we also describe their magnetic properties which helped in unambiguous assignment of mixed valency of V (+4/+5) for Li2VO0.55(H2O)0.45F5⋅2H2O (I) and +4 valency of V for Li3VOF5 (II).


Assuntos
Flúor/química , Óxidos/química , Compostos de Vanádio/química , Vanádio/química , Cristalização/métodos , Lítio/química , Difração de Raios X/métodos
13.
Methods Mol Biol ; 2208: 189-202, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32856264

RESUMO

Integral membrane proteins are important drug targets that are critical in supporting many biological processes. Despite that, the study of their structure-function relationships remains a major goal in structural biology, yet progress has been hampered by inherent challenges in the production for stable and homogeneous protein samples. Dynamic light scattering provides a straightforward probe of protein quality in solution, particularly in relation to stability and aggregation. However, the necessity to use large amounts of sample and the low-throughput nature of the analysis remain as major bottlenecks of the technique.Here, we present a protocol for dynamic light scattering measurements that are executed in a fully automated fashion for low-volume samples, in situ. The protocol offers a generic pre-screening method for downstream structural studies of biomolecules using higher-resolution approaches such as X-ray crystallography, electron microscopy, small-angle X-ray scattering, and NMR .


Assuntos
Difusão Dinâmica da Luz/métodos , Proteínas de Membrana/metabolismo , Peptídeos/química , Luz , Proteínas de Membrana/química , Espalhamento a Baixo Ângulo , Difração de Raios X/métodos
14.
Methods Mol Biol ; 2225: 125-162, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33108661

RESUMO

The myxoma virus has become of interest in human medicine in the last two decades as it has the ability to infect many types of human cancer cells and is being used as a platform to develop viro-therapeutic agents that suppress aggressive and damaging immune responses and inflammation. Furthermore, the myxoma virus encodes proteins that have strong immunosuppressive effects, and several of the myxoma virus-encoded immunomodulators are being developed to treat systemic inflammatory syndromes such as cardiovascular disease and transplant rejection. Myxoma virus encodes the M-T7 protein, the most abundantly secreted protein expressed in myxoma virus-infected cells, originally identified as a rabbit species-specific interferon-gamma (IFN-γ) receptor homolog and as a chemokine-modulating protein binding a wide range of mammalian chemokines. M-T7 is a critical virulence factor for viral pathogenesis that increases virus lethality when expressed. Although M-T7 has been extensively studied using biochemical and biophysical techniques and its interactome map is well known, its three-dimensional (3D) structure remains elusive. Obtaining the 3D structure of M-T7 would be greatly beneficial and is a crucial step toward advancing M-T7 research through understanding the molecular function and activity of M-T7 as a novel therapeutic reagent and to rationally develop this protein as a drug. This chapter provides an overview of the structural determination techniques, especially X-ray crystallography, that can be applied toward the goal of achieving the first high-resolution structure of M-T7. In addition, details of up-and-coming methods are discussed, including X-ray diffraction at X-ray free electron lasers (XFELs), nuclear magnetic resonance (NMR), cryo-electron microscopy (cryo-EM), Micro-electron diffraction (Micro-ED), and small-angle X-ray scattering (SAXS), and their potential applications to M-T7 structural biology.


Assuntos
Cristalização/métodos , Cristalografia por Raios X/métodos , Myxoma virus/química , Receptores de Interferon/ultraestrutura , Proteínas Virais/ultraestrutura , Fatores de Virulência/genética , Difração de Raios X/métodos , Sequência de Aminoácidos , Clonagem Molecular , Microscopia Crioeletrônica , Cristalografia por Raios X/instrumentação , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Modelos Moleculares , Receptores de Interferon/química , Receptores de Interferon/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/ultraestrutura , Espalhamento a Baixo Ângulo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Proteínas Virais/química , Proteínas Virais/genética , Fatores de Virulência/metabolismo , Difração de Raios X/instrumentação
15.
Carbohydr Polym ; 251: 117056, 2021 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-33142608

RESUMO

We describe a method for permitting efficient modification by transglucosidase (TGA), from glycoside hydrolase family 31 (GH31), sequentially after the pre-treatment by maltogenic α-amylases (MA) from GH13. TGA treatment without MA pre-treatment had negligible effects on native starch, while TGA treatment with MA pre-treatment resulted in porous granules and increased permeability to enzymes. MA→TGA treatments lead to decreased molecular size of amylopectin molecules, increased α-1,6 branching, and increased amounts of amylopectin chains with the degree of polymerization (DP)<10 and decreased amounts of DP 10-28 after debranching. Wide-angle X-ray scattering (WAXS) data showed a general decrease in crystallinity except for a long term (20 h) TGA post-treatment which increased the relative crystallinity back to normal. MA→TGA treatment significantly lowered the starch retrogradation of starch and retarded the increase of storage- and loss moduli during storage. This work demonstrates the potential of sequential addition of starch active enzymes to obtain granular starch with improved functionality.


Assuntos
Glucosidases/química , Glicosídeo Hidrolases/química , Amido/química , Zea mays/química , Amilopectina/química , Glucosidases/metabolismo , Glicosídeo Hidrolases/metabolismo , Hidrólise , Porosidade , Difração de Raios X/métodos
16.
Int J Biol Macromol ; 169: 67-74, 2021 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-33338525

RESUMO

The large agglomeration of starch paste in hot water, and fast retrogradation tendency and low transparency of starch gel restrict widespread application of kudzu starch. To improve the above defects, kudzu starch was modified with sequentially α-amylase (AA) and transglucosidase (TG), the latter for varying times. The results indicated that, compared to kudzu starch, amylose content and molecular weight of AA/TG-treated starches reduced by 20.07% and 69.50%, respectively. The proportion of A chain increased by 68.68%, whereas B1, B2 and B3 chains decreased by 14.28%, 48.29% and 23.44%, respectively. The degree of branching dramatically increased by 128.3%. After AA→TG treatment, the changes of starch structure enhanced the functional properties of kudzu starch. The solubility, paste clarity and gelatinization temperature increased, whereas the relative crystallinity, viscosity, storage and loss moduli decreased. Overall, the AA→TG modification would be desirable to improve the functional properties of kudzu starch to expand more large-scale application.


Assuntos
Pueraria/química , Pueraria/enzimologia , Amido/química , Amilases/química , Amilose/química , Glucosidases/química , Glucosidases/metabolismo , Peso Molecular , Solubilidade , Temperatura , Viscosidade , Difração de Raios X/métodos , alfa-Amilases/química , alfa-Amilases/metabolismo
17.
Int J Biol Macromol ; 170: 317-325, 2021 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-33373633

RESUMO

In this work, the influence of Sodium Acetate Trihydrate (SAT) on the gelling stage of a chitin hydrogel was studied. Characterization techniques, such as FTIR, Raman, solid-state NMR, Dielectric Spectroscopy, Small-angle X-ray scattering (SAXS), Wide-angle X-ray scattering (WAXS), and X-ray diffraction (XRD) were used to study the effect of SAT on the micro and nanostructure of the material in the wet, dry and freeze-dried states. It was demonstrated that the amount of SAT in the gelling solution can induce a variation in the supramolecular interaction among the polysaccharide chains, which leads to a change in the structural characteristics. In addition, it was observed that the polymer-water interactions are also altered by this structural ordering. Also, the affinity interaction with lysozyme was evaluated and an influence on the adsorption capacity was evidenced with the use of SAT. This could be an advance for biotechnological, biomedical, and food applications.


Assuntos
Quitina/química , Géis/química , Acetato de Sódio/química , Acetatos/química , Adsorção , Coloides , Liofilização/métodos , Espectroscopia de Ressonância Magnética/métodos , Muramidase/metabolismo , Nanoestruturas/química , Espalhamento a Baixo Ângulo , Difração de Raios X/métodos
18.
Int J Biol Macromol ; 168: 105-115, 2021 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-33309654

RESUMO

A chitosan-based (CS) film was developed with nanosized TiO2 and red apple pomace extract (APE). The intermolecular interactions of CS, TiO2 and APE were evaluated by Fourier transform infrared spectroscopy, scanning electron microscopy and X-ray diffraction. TiO2 nanoparticles remarkably improved the water vapor and UV-Vis light barrier properties, mechanical strength and thermal stability of CS-APE films. The strong antioxidant abilities of CS-APE and CS-TiO2-APE films were characterized. Nano-TiO2 and APE showed a synergistic enhancement of the antimicrobial activity in CS matrix. The addition of TiO2 nano-particles into CS-APE films resulted the sensitive color variations, which applied successfully as an indicator to monitor the freshness of salmon fillets. Consequently, the development of CS-APE-TiO2 film provides a new solution to convert rad apple pomace to an active and multifunctional food packaging material with considerable mechanical, antibacterial, antioxidant and pH-responsive color-changing properties.


Assuntos
Quitosana/química , Manipulação de Alimentos/métodos , Malus/química , Antibacterianos/química , Antioxidantes/química , Embalagem de Alimentos/métodos , Frutas/química , Nanopartículas Metálicas/química , Extratos Vegetais/química , Polifenóis/química , Solubilidade , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Titânio/química , Difração de Raios X/métodos
19.
Carbohydr Polym ; 254: 117293, 2021 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-33357862

RESUMO

Cellulose-lignin composite carbon fibers have shown to be a potential environmentally benign alternative to the traditional polyacrylonitrile precursor. With the associated cost reduction, cellulose-lignin carbon fibers are an attractive light-weight material for, e.g. wind power and automobile manufacturing. The carbon fiber tenacity, tensile modulus and creep resistance is in part determined by the carbon content and the molecular orientation distribution of the precursor. This work disassociates the molecular orientation of different components in cellulose-lignin composite fibers using rotor-synchronized solid-state nuclear magnetic resonance spectroscopy and X-ray scattering. Our results show that lignin is completely disordered, in a mechanically stretched cellulose-lignin composite fiber, while the cellulose is ordered. In contrast, the native spruce wood raw material displays both oriented lignin and cellulose. The current processes for fabricating a cellulose-lignin composite fiber cannot regain the oriented lignin as observed from the native wood.


Assuntos
Fibra de Carbono/química , Celulose/química , Lignina/química , Espectroscopia de Ressonância Magnética/métodos , Difração de Raios X/métodos , Estrutura Molecular , Picea/química , Solubilidade , Resistência à Tração , Viscosidade , Madeira/química
20.
PLoS One ; 15(12): e0236681, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33315857

RESUMO

Mechanical loading affects tendon healing and recovery. However, our understanding about how physical loading affects recovery of viscoelastic functions, collagen production and tissue organisation is limited. The objective of this study was to investigate how different magnitudes of loading affects biomechanical and collagen properties of healing Achilles tendons over time. Achilles tendon from female Sprague Dawley rats were cut transversely and divided into two groups; normal loading (control) and reduced loading by Botox (unloading). The rats were sacrificed at 1, 2- and 4-weeks post-injury and mechanical testing (creep test and load to failure), small angle x-ray scattering (SAXS) and histological analysis were performed. The effect of unloading was primarily seen at the early time points, with inferior mechanical and collagen properties (SAXS), and reduced histological maturation of the tissue in unloaded compared to loaded tendons. However, by 4 weeks no differences remained. SAXS and histology revealed heterogeneous tissue maturation with more mature tissue at the peripheral region compared to the center of the callus. Thus, mechanical loading advances Achilles tendon biomechanical and collagen properties earlier compared to unloaded tendons, and the spatial variation in tissue maturation and collagen organization across the callus suggests important regional (mechano-) biological activities that require more investigation.


Assuntos
Tendão do Calcâneo/fisiopatologia , Fenômenos Biomecânicos/fisiologia , Traumatismos dos Tendões/fisiopatologia , Cicatrização/fisiologia , Tendão do Calcâneo/efeitos dos fármacos , Animais , Fenômenos Biomecânicos/efeitos dos fármacos , Toxinas Botulínicas Tipo A/fisiologia , Colágeno/farmacologia , Modelos Animais de Doenças , Feminino , Ratos , Ratos Sprague-Dawley , Espalhamento a Baixo Ângulo , Estresse Mecânico , Traumatismos dos Tendões/tratamento farmacológico , Cicatrização/efeitos dos fármacos , Difração de Raios X/métodos
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