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1.
Food Chem ; 304: 125444, 2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-31491712

RESUMO

Poor aqueous solubility of some minor steviol glycosides (SGs) has prevented their potential widespread usage as non-nutritional high intensity sweeteners in beverage industry. Rebaudioside B (reb B) is one of the minor SGs found in stevia leaf, and has a better taste quality than many of the major SGs. However, reb B suffers from poor aqueous solubility and low dissolution rate, which greatly limits its application, especially in beverages. In our effort to enhance its solubility by using natural means, we discovered that under certain conditions reb B forms hemihydrate crystal, which has much lower solubility and dissolution rate than commercial powder reb B product. The crystal was characterized by Fourier Transform Infrared spectroscopy (FT-IR), Scanning Electron Microscopy (SEM), and X-ray Diffraction (XRD). This may offer more insight into the interaction of SGs with water at molecular level, and therefore provide new guidance on current efforts to enhance the solubility of SGs.

2.
Yao Xue Xue Bao ; 47(7): 930-3, 2012 Jul.
Artigo em Chinês | MEDLINE | ID: mdl-22993860

RESUMO

The aim of the study is to establish a new method of quality evaluation and validate its feasibilities by the simultaneous quantitative assay of four lignanoids in Schisandra chinensis. A new quality evaluation method, quantitative analysis of multi-components by single marker (QAMS), was established and validated with Schisandra chinensis. Four main lignanoids, schisandrin, schisantherin A, deoxyschizandrin and gamma-schizandrin, were selected as analytes and schisandrin as internal reference substance to evaluate the quality. Their contents in 13 different batches of samples, collected from different bathes, were determined by both external standard method and QAMS. The method was evaluated by comparison of the quantitative results between external standard method and QAMS. No significant differences were found in the quantitative results of four lignanoids in 13 batches of S. chinensis determined by external standard method and QAMS. QAMS is feasible for determination of four lignanoids simultaneously when some authentic standard substances were unavailable, and the developed method can be used for quality control of S. chinensis.


Assuntos
Ciclo-Octanos/análise , Dioxóis/análise , Lignanas/análise , Compostos Policíclicos/análise , Schisandra/química , Cromatografia Líquida de Alta Pressão/métodos , Medicamentos de Ervas Chinesas/química , Frutas/química , Plantas Medicinais/química , Controle de Qualidade
3.
Technol Cancer Res Treat ; 11(5): 467-73, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22775334

RESUMO

Solid tumors such as hepatocellular carcinoma are very often not amenable to chemotherapy and radiotherapy. Local ablation methods, including chemical ablation with absolute ethanol, are therefore an option for treatment but lack of information about the mechanism of devitalization leading to cell death is a hindrance to further adoption. Systemic toxicity also has limited the amount of ethanol that can be used in a single treatment session. Therefore we evaluated the mechanism of urea, a denaturant with little or no systemic toxicity, for potential use in chemical ablation. In this study we report on the use of three methods to analyze the effects in cell culture with a view towards eventual clinical application. Human hepatoma HuH-7 cells were analyzed at several time points after treatment using FTIR, DSC, and Raman microspectroscopy based on MTT and PI-exclusion viability assays. Time course fractional denaturation data plotted against viability show that a 50% viability drop occurs after only a 10-20% drop in overall protein denaturation. Other methods of cell death such as apoptosis may also be operative, but this result implies that protein denaturation is one of the major mechanisms of cell death. This is in line with what has been previously suggested for purely thermal methods, and opens the way to mechanism-based improvements in chemical ablation of solid tumors.


Assuntos
Técnicas de Ablação , Sobrevivência Celular/efeitos dos fármacos , Desnaturação Proteica , Ureia/farmacologia , Varredura Diferencial de Calorimetria , Linhagem Celular Tumoral , Humanos , Neoplasias/terapia , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Espectroscopia de Infravermelho com Transformada de Fourier , Análise Espectral Raman , Temperatura Ambiente
4.
Gene ; 487(1): 52-61, 2011 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-21839154

RESUMO

The mu opioid receptor (MOR) is the principle molecular target of opioid analgesics. The polypyrimidine/polypurine (PPy/u) motif enhances the activity of the MOR gene promoter by adopting a non-B DNA conformation. Here, we report that the PPy/u motif regulates the processivity of torsional stress, which is important for endogenous MOR gene expression. Analysis by topoisomerase assays, S1 nuclease digests, and atomic force microscopy showed that, unlike homologous PPy/u motifs, the position- and orientation-induced structural strains to the mouse PPy/u element affect its ability to perturb the relaxation activity of topoisomerase, resulting in polypurine strand-nicked and catenated DNA conformations. Raman spectrum microscopy confirmed that mouse PPy/u containing-plasmid DNA molecules under the different structural strains have a different configuration of ring bases as well as altered Hoogsteen hydrogen bonds. The mouse MOR PPy/u motif drives reporter gene expression fortyfold more effectively in the sense orientation than in the antisense orientation. Furthermore, mouse neuronal cells activate MOR gene expression in response to the perturbations of topology by topoisomerase inhibitors, whereas human cells do not. These results suggest that, interestingly among homologous PPy/u motifs, the mouse MOR PPy/u motif dynamically responds to torsional stress and consequently regulates MOR gene expression in vivo.


Assuntos
DNA/genética , Regiões Promotoras Genéticas/genética , Receptores Opioides mu/genética , Sequências Reguladoras de Ácido Nucleico/genética , Animais , Sítios de Ligação/genética , Camptotecina/análogos & derivados , Camptotecina/farmacologia , Linhagem Celular Tumoral , DNA/química , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Irinotecano , Luciferases/genética , Luciferases/metabolismo , Camundongos , Microscopia de Força Atômica , Conformação de Ácido Nucleico , Nucleotídeos de Purina/química , Nucleotídeos de Purina/genética , Nucleotídeos de Pirimidina/química , Nucleotídeos de Pirimidina/genética , Receptores Opioides mu/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise Espectral Raman , Inibidores da Topoisomerase I/farmacologia
5.
Gene ; 471(1-2): 27-36, 2011 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-20946943

RESUMO

The mu opioid receptor (MOR) is the principle molecular target of opioid analgesics. An appropriate understanding of MOR gene expression across species is critical for understanding its analgesic functions in humans. Here, we undertake a cross-species analysis of the polymorphic polypyrimidine/polypurine (PPy/u) motif, a key enhancer of MOR gene expression. The mouse PPy/u motif is highly homologous to those of rat (67%) and human (83%), but drives reporter gene expression tenfold and fivefold more effectively than those of rat and human, respectively. Circular dichroism profiles of PPy/u oligonucleotides from different species showed that they are primarily different in structure. Conformational studies of reporter plasmids using confocal Raman spectra, S1 nuclease and restriction enzymes demonstrated that the structural difference is the result of changes in the phosphodiester backbone. Furthermore, these conformational disparities produce differences in torsional stress, as shown by topoisomerase II relaxation and activation of different levels of gene expression under hypertonic conditions. This study demonstrates that homologous PPy/u motifs adopt unique species-specific conformations with different mechanisms and activities for gene expression. We further discuss how structural aspects of transcription regulatory elements, rather than the sequence itself, are significant when studying functional gene expression regulatory elements.


Assuntos
Regiões Promotoras Genéticas , Receptores Opioides mu/genética , Animais , Sequência de Bases , Dicroísmo Circular , Regulação da Expressão Gênica , Genes Reporter , Humanos , Camundongos , Dados de Sequência Molecular , Mutação , Polimorfismo Genético , Ratos , Mapeamento por Restrição/métodos , Alinhamento de Sequência , Homologia de Sequência do Ácido Nucleico , Especificidade da Espécie
6.
Mol Pharm ; 8(1): 176-84, 2011 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-21053973

RESUMO

This work demonstrates the use of confocal Raman microscopy (CRM) to measure the dynamics of cellular uptake and localization of gold nanoparticles (GNP) with nanoscale resolution. This is important as nanoparticle cellular interactions are increasingly under investigation to support applications as diverse as drug delivery, gene transfection and a variety of heat and radiation based therapeutics. At the heart of these applications is a need to know the dynamics of nanoparticle cellular uptake and localization (i.e., cell membrane, cytoplasm or nucleus). This process can change dramatically based on size, charge, shape and ligand attached to the nanoparticle. While electron microscopy, atomic emission spectroscopy and histology can be used to assess cellular uptake, they are labor intensive and post-mortem and can miss critical dynamics of the process. For this reason investigators are increasingly turning to optically active nanoparticles that allow direct microscopic interrogation of uptake. Here we show that CRM adds to this evolving armamentarium as a fast, noninvasive, and label-free technique to dynamically study cellular uptake of GNPs with subcellular detail in cancer. Raman laser interaction with GNPs inside cells shows unique spectroscopic features corresponding to the intracellular localization of GNPs over 2 to 24 h at the membrane, cytoplasm or nucleus that are separately verified by histology (silver staining) and electron microscopy. These results show that CRM has the potential to facilitate high-throughput study of the dynamics and localization of a variety of GNPs in multiple cell types.


Assuntos
Ouro/química , Nanopartículas Metálicas/química , Microscopia Confocal/métodos , Neoplasias/metabolismo , Linhagem Celular Tumoral , Humanos , Nanopartículas Metálicas/ultraestrutura , Microscopia Eletrônica de Transmissão , Neoplasias/ultraestrutura , Espectrofotometria Atômica
7.
Biophys J ; 99(8): 2453-9, 2010 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-20959085

RESUMO

We describe direct determination of the state of intracellular water, measurement of the intercellular concentration of a cryoprotectant agent (dimethylsulfoxide), and the distribution of organic material in frozen mammalian cells. Confocal Raman microspectroscopy was utilized at cryogenic temperatures with single live cells to conduct high spatial resolution measurements (350 × 350 × 700 nm), which yielded two, we believe, novel observations: 1), intracellular ice formation during fast cooling (50°C/min) causes more pronounced intracellular dehydration than slow cooling (1°C/min); and 2), intracellular dimethylsulfoxide concentration is lower (by as much as 50%) during fast cooling, decreasing the propensity for intracellular vitrification. These observations have a very significant impact for developing successful biopreservation protocols for cells used for therapeutic purposes and for cellular biofluids.


Assuntos
Fibroblastos/metabolismo , Congelamento , Água/metabolismo , Crioprotetores/metabolismo , Dimetil Sulfóxido/metabolismo , Fibroblastos/citologia , Humanos , Espaço Intracelular/metabolismo , Análise de Célula Única , Análise Espectral Raman
8.
Conf Proc IEEE Eng Med Biol Soc ; 2009: 6006-9, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19963916

RESUMO

The ElectroNanospray process (Nanocopoeia, Inc) transforms drugs and polymers into many nanoscale material states including powders, liquids, encapsulated particles, and coatings. This enabling technology platform allows application of polymers and drugs to the surface of medical devices such as coronary stents in a single-stage process. Modification of ElectroNanospray process parameters resulted in surface coatings with rich morphologies ranging in appearance from smooth and heterogeneous to highly porous and rough (open matrix). The traditional approach of measuring percent release over time by HPLC shows that the drug release profiles change significantly with coating morphology. In this study, we employed high resolution imaging techniques such as SEM, Atomic Force Microscopy (AFM) and Confocal Raman Microscopy to elucidate the drug release process on these coatings in situ, indicating a correlation of release kinetics with coating morphology.


Assuntos
Materiais Revestidos Biocompatíveis/química , Nanocápsulas/química , Nanocápsulas/ultraestrutura , Preparações Farmacêuticas/química , Polímeros/química , Difusão , Composição de Medicamentos/métodos , Teste de Materiais , Propriedades de Superfície
9.
Conf Proc IEEE Eng Med Biol Soc ; 2009: 6304-6, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19963921

RESUMO

Drug delivery systems incorporated onto the end of cardiac leads are used to reduce inflammation and fibrosis at the lead-tissue interface and enable optimal lead performance. In this research, confocal Raman microscopy was used to capture chemical images of the drug delivery system on pacemaker leads in different elution media in real-time. Raman images in ambient air showed that drug was dispersed in the polymer matrix as discrete particles with size ranging from 1 to 3 microm. Upon immersion into an aggressive elution medium, drug near the surface dissolved immediately and solvent started to penetrate into the polymer matrix through channels from which drug was eluted. The drug depletion depth was a function of time, which was consistent with the drug release profiles obtained by HPLC. Comparing the drug elution in aggressive solvent and biorelevant solvent, a mechanism of drug release is proposed.


Assuntos
Implantes de Medicamento/química , Eletrodos Implantados , Microscopia Confocal/métodos , Marca-Passo Artificial , Análise Espectral Raman/métodos , Sistemas de Computação , Difusão , Avaliação Pré-Clínica de Medicamentos/métodos , Implantes de Medicamento/análise
10.
J Phys Chem B ; 113(30): 10081-7, 2009 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-19580250

RESUMO

Amid decades of research, the basic mechanisms of lyo-/cryostabilization of proteins and more complex organisms have not yet been fully established. One major bottleneck is the inability to probe into and control the molecular level interactions. The molecular interactions are responsible for the significant differences in the outcome of the preservation processes. (1) In this communication, we have utilized confocal Raman microspectroscopy to quantify the freezing-induced microheterogeneity and phase separation (solid and liquid) in a frozen solution composed of a model protein (lysozyme) and a lyo-/cryoprotectant (trehalose), which experienced different degrees of supercooling. Detailed quantitative spectral analysis was performed across the ice, the freeze-concentrated liquid (FCL) phases, and the interface region between them. It was established that the characteristics of the microstructures observed after freezing depended not only on the concentration of trehalose in the solution but also on the degree of supercooling. It was shown that, when samples were frozen after high supercooling, small amounts of lysozyme and trehalose were occluded in the ice phase. Lysozyme preserved its native-like secondary structure in the FCL region but was denatured in the ice phase. Also, it was observed that induction of freezing after a high degree of supercooling of high trehalose concentrations resulted in aggregation of the sugar and the protein.


Assuntos
Congelamento , Proteínas/química , Animais , Crioprotetores/farmacologia , Relação Dose-Resposta a Droga , Muramidase/química , Muramidase/metabolismo , Ligação Proteica/efeitos dos fármacos , Estrutura Secundária de Proteína/efeitos dos fármacos , Proteínas/metabolismo , Soluções , Termodinâmica , Trealose/farmacologia
11.
J Phys Chem B ; 113(30): 10189-95, 2009 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-19572658

RESUMO

Migration of surfactants in water-based, pressure-sensitive adhesive (PSA) films exposed to static and cyclic relative humidity conditions was investigated using confocal Raman microscopy (CRM) and atomic force microscopy (AFM). Studied PSA films contain monomers n-butyl acrylate, vinyl acetate, and methacrylic acid and an equal mass mixture of anionic and nonionic nonylphenol ethoxylate emulsifiers. A leveling of surfactant concentration distributions is observed via CRM after films stored at 50% relative humidity (RH) are exposed to a 100% RH for an extended time period, while relatively small increases in surface enrichment occur when films are stored at 0% RH. Use of CRM for binary mixtures containing anionic or nonionic surfactant and latex that has undergone dialysis to remove nonpolymeric components indicates that surfactant-polymer compatibility governs to a great extent surface enrichment, but not changes observed with humidity variations. AFM images show that upon drying latex coatings, surfactant and other additives collect in large aggregation regions, which protrude from film surfaces. These structures are absent at high humidity, which appears to result from lateral spreading across the polymer surface. When humidity is reduced, aggregation regions reform but appear to be smaller and more evenly dispersed, and by cycling humidity between 0 and 100% RH, interfacial enrichment can be seen to diminish. Presented results provide greater insights into the distribution behavior of surfactants in latex films and potential mechanisms for observed issues arising for these systems.


Assuntos
Umidade , Látex/química , Tensoativos/química , Adesivos/química , Microscopia de Força Atômica , Movimento (Física) , Pressão , Propriedades de Superfície
12.
Langmuir ; 25(10): 5442-5, 2009 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-19432489

RESUMO

Drug release from therapeutic biomedical films such as drug-polymer composite coatings on drug eluting stents is a highly complex and poorly understood process. The dynamics of drug release and the evolution of surface morphology during release have direct impact on the performance of the device. This information is not easily accessible, and there have been few systematic studies to investigate drug release from biomedical coatings in real time. In this study, the complementary analytical techniques of confocal Raman microscopy, in-liquid atomic force microscopy, scanning electron microscopy, and high performance liquid chromatography were used to examine real-time mobilization and release of the drug rapamycin from polyisobutylene-block-polystyrene thin films, during immersion in buffered saline for 12 h. Each technique was found to have distinct limitations in either temporal or spatial resolution; in combination, however, the overlapping techniques provided a level of detail that is not available using any single approach.


Assuntos
Antibióticos Antineoplásicos/química , Materiais Revestidos Biocompatíveis/química , Preparações de Ação Retardada/química , Membranas Artificiais , Modelos Químicos , Sirolimo/química , Microscopia de Força Atômica , Microscopia Confocal , Microscopia Eletrônica de Varredura , Polienos/química , Polímeros/química , Poliestirenos/química
13.
Artigo em Inglês | MEDLINE | ID: mdl-20049810

RESUMO

This paper reviews the development of coronary stents from a polymer scientist's view point, and presents the first results of an interdisciplinary team assembled for the development of new stent systems. Poly(styrene-b-isobutylene-b-styrene) block copolymer (SIBS), a nanostructured thermoplastic elastomer, is used in clinical practice as the drug-eluting polymeric coating on the Taxus coronary stent (trademark of Boston Scientific Co.). Our group has been developing new architectures comprising of arborescent (dendritic) polyisobutylene cores (D_SIBS), which were shown to be as biocompatible as SIBS. ElectroNanospray (Nanocopoeia Inc.) was used to coat test coupons and coronary stents with selected D(S)IBS polymers loaded with dexamethasone, a model drug. The surface topology varied from smooth to nanosized particulate coating. This paper will demonstrate how drug release profiles were influenced by both the molecular weight of the polyisobutylene core and spraying conditions of the polymer-drug mixture.


Assuntos
Materiais Biocompatíveis/administração & dosagem , Materiais Biocompatíveis/química , Stents Farmacológicos , Polímeros/administração & dosagem , Polímeros/química , Humanos , Nanotecnologia/métodos
14.
J Phys Chem B ; 112(38): 11907-14, 2008 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-18767777

RESUMO

Surfactant distributions in model pressure-sensitive adhesive (PSA) films were investigated using atomic force microscopy (AFM) and confocal Raman microscopy (CRM). The PSAs are water-based acrylics synthesized with n-butyl acrylate, vinyl acetate, and methacrylic acid and two commercially available surfactants, disodium (nonylphenoxypolyethoxy)ethyl sulfosuccinate (anionic) and nonylphenoxypoly(ethyleneoxy) ethanol (nonionic). The ratio of these surfactants was varied, while the total surfactant content was held constant. AFM images demonstrate the tendency of anionic surfactant to accumulate at the film surfaces and retard latex particle coalescence. CRM, which was introduced here as a means of providing quantitative depth profiling of surfactant concentration in latex adhesive films, confirms that the anionic surfactant tends to migrate to the film interfaces. This is consistent with its greater water solubility, which causes it to be transported by convective flow during the film coalescence process. The behavior of the nonionic surfactant is consistent with its greater compatibility with the polymer, showing little enrichment at film interfaces and little lateral variability in concentration measurements made via CRM. Surfactant distributions near film interfaces determined via CRM are well fit by an exponential decay model, in which concentrations drop from their highs at interfaces to plateau values in the film bulk. It was observed that decay constants are larger at the film-air interface compared with those obtained at the film-substrate side indicating differences in the mechanism involved. In general, it is shown here that CRM acts as a powerful compliment to AFM in characterizing the distribution of surfactant species in PSA film formation.


Assuntos
Adesivos/química , Etilenoglicóis/química , Tensoativos/química , Água/química , Emulsificantes/química , Microscopia de Força Atômica , Microscopia Confocal , Pressão
15.
Environ Sci Technol ; 41(7): 2636-42, 2007 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-17438827

RESUMO

Several fullerene-based nanomaterials generate reactive oxygen species that can damage cells. In this study, we investigated the effect of buckminsterfullerene (C60) introduced as colloidal aggregates in water (nC60) on bacterial membrane lipid composition and phase behavior. Pseudomonas putida (Gram-negative) and Bacillus subtilis (Gram-positive) responded to nC60 by altering membrane lipid composition, phase transition temperature, and membrane fluidity. P. putida decreased its levels of unsaturated fatty acids and increased the proportions of cyclopropane fatty acids in the presence of nC60, possibly to protect the bacterial membrane from oxidative stress. Fourier transform infrared spectroscopy measurement of intact bacterial cells showed slightly increased phase transition temperatures (Tm) and increased membrane fluidity for cells grown in the presence of high, growth-inhibiting concentrations (0.5 mg L(-1)) of nC60. B. subtilis responded to a low dose of nC6o (0.01 mg L(-1)) by significantly increasing the levels of iso- and anteiso-branched fatty acids (from 5.8 to 31.5% and 12.9 to 32.3% of total fatty acids, respectively) and to a high, growth-inhibiting concentration of nC60 (0.75 mg L-1) by increasing synthesis of monounsaturated fatty acids. In contrast to P. putida, B. subtilis response was a decrease in Tm and an increase in membrane fluidity. These findings represent the first demonstrated physiological adaptation response of bacteria to a manufactured nanomaterial, and they showthat response inlipid composition and membrane phase behavior depends on both the nC60 concentration and the cell wall morphology.


Assuntos
Adaptação Fisiológica/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Fulerenos/toxicidade , Lipídeos de Membrana/metabolismo , Bacillus subtilis , Coloides/toxicidade , Relação Dose-Resposta a Droga , Pseudomonas putida , Espectroscopia de Infravermelho com Transformada de Fourier , Temperatura Ambiente
16.
Langmuir ; 20(7): 2695-700, 2004 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-15835139

RESUMO

The self-associating structures at the solid-liquid interface of three nonionic trisiloxane surfactants ((CH3)3SiO)2Si(CH3)(CH2)3(OCH2CH2)n OH (n = 6, 8, and 12), or BEn, are studied as a function of substrate properties by atomic force microscopy (AFM) imaging and force measurement. These trisiloxane surfactants are known as superwetters, which promote rapid spreading of dilute aqueous solutions on low-energy surfaces. This study also attempts to relate the BEn surface aggregate structures at the solid-liquid interface to their superwetting behavior. Four substrates are used in the study: muscovite mica, highly oriented pyrolytic graphite, and oxidized silicon wafer with and without a full monolayer of self-assembled n-octadecyltrichlorosilane (OTS). The concentration of BEn is fixed at 2 times the critical aggregation concentration (CAC). The BEn surfactants are only weakly attracted to hydrophilic surfaces, more on oxidized silicon than on mica. All three form ordinary planar monolayers on HOPG and OTS-covered oxidized silicon. The significance of surfactant adsorption on the AFM tip is investigated by comparing the force curves obtained by tips with and without thiol modification. The surface aggregate structures of the BEn surfactants correlate with their bulk structures and do not exhibit anomalous adsorption behavior. The adsorption behavior of the BEn superwetters is similar to that of the CmEn surfactants. Thus, our results confirm previous work showing that superwetting shares its main features with other classes of surfactants.


Assuntos
Siloxanas/química , Tensoativos/química , Silicatos de Alumínio/química , Grafite/química , Interações Hidrofóbicas e Hidrofílicas , Microscopia de Força Atômica , Modelos Moleculares , Nanotecnologia , Transição de Fase , Dióxido de Silício/química , Propriedades de Superfície
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