Polystyrene-type resin used for peptide synthesis: application for anion-exchange and affinity chromatography

This paper deals with an unusual application for a copolymer of styrene-1 % divinylbenzene bearing high amount of aminomethyl groups for anion-exchange and affinity chromatography. The so-called aminomethyl resin (AMR), to date only employed for peptide synthesis, swelled appreciably in water and was used successfully to purify negatively charged peptides. By correlating swelling degree of beads with pH of the media, it was possible to estimate that the AMR amino group pK(a) is approximately 5.5. In addition, the synthesized acetyl-(NANP)(3)-AMR succeeded in the affinity interaction with large antibody molecules related to malaria transmission and raised previously against this dodecapeptide sequence. (C) 2004 Elsevier B.V. All rights reserved.

Determination of low-aliphatic aldehydes indoors by micellar electrokinetic chromatography using sample dissolution manipulation for signal enhancement

This work describes a novel approach for the analysis of selected aldehydes (formaldehyde, acetaldehyde, propionaldehyde, and acrolein) and acetone in environmental samples using micellar electrokinetic chromatography (MEKC). The method is based on the reaction of carbonyl compounds with 3-methyl-2-benzothiazoline hydrazone (MBTH) that gives an azine intermediate with maximum absorbance at 216 nm. A systematic evaluation of sample dissolution medium was conducted as a means to enhancing sensitivity. In the best condition, samples were dissolved in 0.030 mol.L-1 tetraborate solution. This condition presented enhancement factors in the range of 35-54 for the aldehydes under investigation, computed as the improvement of the concentration limits of detection (LODs) with reference to the sample dissolved in pure water. The running buffer was 0.020 mol.L-1 tetraborate, pH 9.3, containing 0.050 mol-L-1 sodium dodecyly sulfate (SIDS). The overall methodology presented several advantages over established methods for aldehydes. Worthy mentioning that MBTH is available in high purity degree, dispensing laborious reagent purification procedures. A few method validation parameters were determined revealing good migration time repeatability (< 2.5% coefficient of variation, CV) and area repeatability (< 4% CV), excellent linearity (20-120 mug/L, r > 0.995) and adequate sensitivity for environmental applications. The LODs with respect to each single aldehyde were in the range of 0.54-4.0 mug.L-1 and 11 mug.L-1 for acetone. The methodology was applied to the determination of aldehydes indoors. Samples were collected in an impinger flask containing 0.05% MBTH solution, at a flow rate of 0.80 L.min(-1), during 2.5 h, at different times during the day. The most abundant carbonyls in the samples were acetone, followed by formaldehyde and acetaldehyde, with estimate peak concentrations of 452, 5.2 and 2.2 ppbv, respectively.

Purification and partial characterization of cathepsin D from porcine (Sus scrofa) liver using affinity chromatography

Cathepsin D, a lysosomal aspartic protease, has been purified from porcine liver using a combination of pepstatin-A agarose and Affi-Gel Blue affinity chromatography, followed by size-exclusion chromatography. The purified protein consists of two polypeptide chains of 15 and 30 kDa, and has an isoelectric point of 6.8. Porcine liver cathepsin D has maximum activity at pH 2.5-3.0 as determined by its activity against hemoglobin, with a K-cat of 14.3 s(-1) and a k(cat)/K-M of 2.70 x 10(6) s(-1) M-1 as determined by the hydrolysis of a fluorogenic peptide substrate.

Validation of an immunoassay method for the determination of traces of carbaryl in vegetable and fruit extracts by liquid chromatography with photodiode array and mass spectrometric detection

A competitive enzyme-linked immunosorbent assay (ELISA) method for carbaryl quantitation in crop extracts was validated by liquid chromatography (LC) with diode array detection (DAD). For this purpose, six crops (banana, carrot, green bean, orange, peach and potato) were chosen for recovery and reproducibility studies. The general sample preparation included extraction with methanol followed by liquid-liquid partitioning and clean-up on Celite-charcoal adsorbent column of the vegetable extracts. ELISA samples consisted of a diluted LC extract in assay phosphate buffer (pH 7.5). The potential effect of methanol in these samples was evaluated. It was observed that a maximum content of 10% methanol present in the assay buffer could be tolerated without expressive losses in the ELISA performance. Under these conditions, a IC50 similar to 1.48 mu g l(-1) was obtained. A minimum matrix effect with a 1:50 dilution of the methanolic extracts in assay buffer was noticed, except for green bean samples that inhibited completely the assay. For the vegetable extracts, the ELISA sensitivities varied from 3.9 to 5.7 mu g l(-1), and good recoveries (82-96%) with R.S.D.s ranging from 5.7 to 12.1% were found. An excellent correlation between the LC-DAD and ELISA techniques was obtained. The confirmation of the carbaryl in less concentrated samples was achieved by LC-mass spectrometry interfaced with atmospheric pressure chemical ionisation. The [M + H](+)= 202 and [M + H-57](+)=145 ions, equivalent to the protonated molecular and l-naphthol ions, respectively, were used to carbaryl identification in these samples. (C) 1998 Elsevier B.V. B.V. All rights reserved.

Synthesis and characterization of phosphorylcholine-substituted chitosans soluble in physiological pH conditions

A polymer analogous synthesis involving the reductive amination of phosphorylcholine (PC)-glyceraldehyde with primary amines of deacetylated chitosan (M-w approximate to 57000 g mol(-1)) was used to prepare phosphorylcholine-substituted chitosans (PC-CH) with a degree of substitution (DS) ranging from similar to 11 to similar to 53 mol% PC-substituted glucosamine residues. The PC-CH derivatives were characterized by H-1 NMR spectroscopy, FTIR spectroscopy, and multiangle laser light scattering gel permeation chromatography (MALLS-GPC). The pKa of the PC-substituted amine groups (pKa approximate to 7.20) was determined by H-1 NMR titration. The PC-CH samples (1.0 g L-1) were shown to be nontoxic using an MTT assay performed with human KB cells. Aqueous solutions of PC-CH samples (4.0 g L-(1)) of DS g 22 mol% PC-substituted glucosamine residues remained clear, independently of pH (4.0 < pH < 11.0). The remarkable water solubility and nontoxicity displayed by the new PC-CH samples open up new opportunities in the design of chitosan-based biomaterials and nanoparticles.

Cytotoxic effects of current dental adhesive systems on immortalized odontoblast cell line MDPC-23

Objectives: Evaluate the cytotoxic effect of the three dental adhesive systems. Methods: The immortalized mouse odontoblast cell line (MDPC-23) was plated (30,000 cell/cm 2) in 24 well dishes, allowed to grow for 72 h, and counted under inverted light microscopy. Uncured fresh adhesives were added to culture medium to simulate effects of unset adhesive. Three adhesives systems were applied for 120 min to cells in six wells for each group: Group 1) Single Bond (3M), Group 2) Prime & Bond 2.1 (Dentsply), and Group 3) Syntac Sprint (Vivadent). In the control group, PBS was added to fresh medium. The cell number was counted again and the cell morphology was assessed under SEM. In addition, the adhesive systems were applied to circles of filter paper, light-cured for 20 s, and placed in the bottom of 24 wells (six wells for each experimental materials and control group). MDPC-23 cells were plated (30,000 cell/cm 2) in the wells and allowed to incubate for 72 h. The zone of inhibition around the filter papers was measured under inverted light microscopy; cell morphology was evaluated under SEM; and the MTT assay was performed for mitochondrial respiration. Results: The fresh adhesives exhibited more toxic (cytopathic effects) to MDPC-23 cells than polymerized adhesives on filter papers, and as compared to the control group. The cytopathic effect of the adhesive systems occurred in the inhibition zone around the filter papers, which was confirmed by the MTT assay and statistical analysis (ANOVA) combined with Fisher's PLSD test. In the control group, MDPC-23 cells were dense on the plastic substrate and were in contact with the filter paper. In the experimental groups, when acid in the adhesive systems was removed by changing the culture medium, or when the adhesives were light-cured, some cells grew in the wells in spite of the persistent cytotoxic effect. Significance: All dentin adhesive systems were cytotoxic odontoblast-like cells. Both acidity and non-acidic components of these systems were responsible for the high cytopathic effect of those dental materials. © 1999 Academy of Dental Materials. Published by Elsevier Science Ltd. All rights reserved.

Longitudinal distribution of Copepoda populations in the transition zone of Paranapanema River and Jurumirim Reservoir (São Paulo, Brazil) and interchange with two lateral lakes.

Longitudinal changes in composition, abundance, and distribution of copepods were studied at the transition zone of Paranapanema River-Jurumirim Reservoir (SP, Brazil). The interchange of biotic material between marginal lakes and the river system was also examined. Water samples were obtained from 6 stations along a stretch of 13 km of the Paranapanema River, from an upstream reach with high water velocity up to the river mouth into Jurumirim Reservoir. Two other sites in lateral lakes were also sampled. Nine copepod taxa were identified: 3 calanoids (Argyrodiaptomus furcatus Sars, Notodiaptomus iheringi Wright, and N. conifer Sars) and 6 cyclopoids (Eucyclops Claus, Microcyclops Claus, Mesocyclops longisetus Thiébaud, Thermocyclops decipiens Fischer, T. minutus Lowndes, and Paracyclops Claus). Harpacticoids were also collected. Calanoid and cyclopoid nauplii and copepodids, and harpacticoids were the most abundant organisms. In general, there was a longitudinal decrease in copepod abundance, whereas an increase was detected near the lakes. The abundance of most copepods was inversely correlated with current velocity and suspended solids. Higher abundance was observed in the river main course during the rainy season, during which there is a higher connectivity between the lakes and the main river. This promotes exportation of biologic material from marginal lakes to the river system, a biotic exchange reflecting the importance of marginal lakes to the river community structure.

Search for the flavor-changing neutral current decay Bs 0 → μ+ μ- pp̄ Collisions at √s = 1.96 TeV with the DO Detector

We present the results of a search for the flavor-changing neutral current decay Bs 0 → μ+ μ-. using a data set with integrated luminosity of 240 pb-1 of pp̄ collisions at √s = 1.96 TeV collected with the D0 detector in run II of the Fermilab Tevatron collider. We find the upper limit on the branching fraction to be B(Bs 0 → μ+ π-) ≤ 5.0 × 10-7 at the 95% C.L. assuming no contributions from the decay Bd 0 → μ+ μ- in the signal region. This limit is the most stringent upper bound on the branching fraction Bs 0 → μ+ μ- to date. © 2005 The American Physical Society.

Determination of flutamide in tablets by high-performance liquid chromatography

Flutamide is a potent antiandrogen used for the treatment of prostatic cancer. A simple, sensitive and accurate high-performance liquid Chromatographic (HPLC) method is presented for quantitative determination of flutamide in tablets, using a reversed-phase technique and UV detection at 240 nm. The isocratic elution was used to quantify the analyte. The samples were chromatographed on Luna-C18 column and the mobile phase was 0.05 M phosphate buffer pH 4.0 - acetonitrile (50:50, v/v). The method was linear between 2.9 - 11.6 mg L -1. Over the tested concentration range the intra-day relative standard deviation for replicate analysis in tablets ranged from 0.44 to 0.78%. It was also found that the excipients in the commercial tablets did not interfere with the method.

Influence of radiopacifying agents on the solubility, pH and antimicrobial activity of Portland cement

The aim of this study was to evaluate the interference of the radiopacifiers bismuth oxide (BO), bismuth carbonate (BC), bismuth subnitrate (BS), and zirconiun oxide (ZO) on the solubility, alkalinity and antimicrobial properties of white Portland cement (WPC). The substances were incorporated to PC, at a ratio of 1:4 (v/v) and subjected to a solubility test. To evaluate the pH, the cements were inserted into retrograde cavities prepared in simulated acrylic teeth and immediately immersed in deionized water. The pH of the solution was measured at 3, 24, 72 and 168 h. The antimicrobial activity was evaluated by a radial diffusion method against the microorganisms S. aureus (ATCC 25923), P. aeruginosa (ATCC 27853), E. faecalis (ATCC 29212) and C. albicans (ATCC 10231). The zone of microbial growth inhibition was measured after 24 h. The addition of BS and BC increased the solubility of the cement. The pH values demonstrated that all materials produced alkaline levels. At 3 h, BS showed lower pH than WPC (p<0.05). At 168 h, all materials showed similar pHs (p>0.05). The materials did not present antimicrobial activity for S. aureus, P. aeruginosas and E. faecalis (p>0.05). With regards to C. albicans, all materials formed an inhibition zone, mainly the mixture of WPC with ZO (p<0.05). The type of radiopacifier incorporated into WPC interfered with its physical and antimicrobial properties. ZO was found to be a viable radiopacifier that can be used with WPC.

Determination of sildenafil and vardenafil by capillary zone electrophoresis using capacitively coupled contactless conductivity detection

A method based on capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-C4D) for determination of two important phosphodiesterase type-5 inhibitors (sildenafil and vardenafil) is introduced. The background electrolyte (BGE) consisted of an aqueous solution of 500 mmol L-1 acetic acid, and the capillary was previously treated with polybrene solution to prevent cationic analytes from adsorbing onto the inner surface. Although the analytes migrate in the counter flow, the total time is short. An instrument with two C4D detectors allowed a seamless transition from a fast method (less than one minute) but of low-efficiency using the first detector to a more efficient method using the second detector. The analysis of commercial tablets showed no significant difference between CE-C4D and HPLC methods. Conductivity detection is a well-known low selectivity detection scheme, which in conjunction with the high mobility of the co-ion in the BGE (hydroxonium) allows one to predict that other cationic analogues of sildenafil can also be detected. This is an interesting feature given the increasing number of compounds in this class. © 2013 The Royal Society of Chemistry.

Evidence for euphotic zone anoxia during the deposition of Aptian source rocks based on aryl isoprenoids in petroleum, Sergipe-Alagoas Basin, northeastern Brazil

Four crude oil samples from the Sergipe-Alagoas Basin, northeastern Brazil, were analyzed using full scan gas chromatography-quadrupole mass spectrometry (GC-qMS) for biomarkers, in order to correlate them using aromatic carotenoids thereby enhancing knowledge about the depositional environment of their source rocks. The geochemical parameters derived from saturated fractions of the oils show evidence of little or no biodegradation and similar thermal maturation (Ts/(Ts+Tm) for terpanes, C29 αββ/(αββ+ααα), C27, and C29 20S/(20S+20R) for steranes). Low pristane/phytane ratios and the abundance of gammacerane and β-carotane are indicative of an anoxic and saline depositional environment for the source rocks. Moreover, we identified a large range of diagenetic and catagenetic products of the aromatic carotenoid isorenieratene, including C40, C33, and C32 diaryl isoprenoids and aryl isoprenoid derivatives with short side chains and/or additional rings. These results indicate anoxia in the photic zone during the deposition of the source rocks. © 2013 The Authors.

Síntese, caracterização e estudos em solução de terpolímeros antififílicos termo-sensíveis ao pH: uma perspectiva para obtenção de sistemas transportadores de fármacos

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Condições sócio-ambientais da bacia hidrográfica do rio Caeté (Pará, Brasil)

A ausência de gerenciamento em bacias hidrográficas, na região Amazônica, tem afetado seus recursos hídricos nos últimos anos. Por esta razão, esta tese visa estudar os aspectos sociais e ambientais de dezoito comunidades rurais ao longo da Bacia Hidrográfica do Caeté, bem como o efeito da descarga de água residual lançada no estuário do Caeté e no rio Cereja, na cidade de Bragança. A metodologia adotada para o levantamento do perfil socioeconômico e das condições de vida e moradia foi baseada na aplicação de questionários estruturados e semi-estruturados. Em algumas comunidades, a produção doméstica de lixo foi estimada e catalogada, em 20% das residências. Quanto à qualidade da água subterrânea, análises das variáveis físicoquímicas e microbiológicas (turbidez, cor real, cor aparente, pH, temperatura, ferro dissolvido, nutrientes dissolvidos, coliformes termotolerantes e coliformes totais) foram realizadas nos principais poços que abastecem as comunidades rurais estudadas. Coletas oceanográficas (com medidas de variáveis hidrológicas e hidrodinâmicas) como também análises microbiológicas foram realizadas nas áreas mais urbanizadas do estuário do Caeté e do rio Cereja, para caracterizar a qualidade das águas nos setores estudados. Os principais impactos ambientais foram identificados, georreferênciados, fotografados e mapeados. Diretrizes de gerenciamento foram propostas para minimizar os problemas sócio-ambientais encontrados. Os resultados obtidos mostraram que das 2.207 famílias rurais (~9.573 habitantes), a maioria possui baixa-renda, baixo grau de escolaridade e precárias condições de vida e moradia. Os serviços e infraestrutura disponíveis são ineficientes ou ausentes, como evidenciados pela falta abastecimento de água potável, coleta de lixo, coleta de esgoto, escolas, atendimento médico etc. Por outro lado, a cidade de Bragança tem mais de 72.621 habitantes, vivendo sobre uma área de aproximadamente 16 km², e é uma das mais antigas cidades da região amazônica. Entretanto, os dois rios estudados recebem influência da descarga de esgoto por residências, hospitais, comércio, fábricas etc. e tem apresentado elevado índice de coliformes termotolerantes. A falta de políticas públicas eficazes tem acentuado os problemas ambientais e sócio-econômico. Desta forma, os autores sugerem várias medidas para melhorar a situação atual, incluindo: (i) a regulamentação do uso da terra para reduzir o impacto ambiental do setor econômico; (ii) a implementação de programas de gestão para a exploração sustentável dos recursos naturais (peixes, caranguejos, argila, madeira); (iii) a instalação de serviços públicos, em especial água encanada e saneamento e (iv) o controle e penalização da exploração ilegal dos recursos naturais.

Condições oceanográficas, ocupação territorial e problemas ambientais na praia do Atalaia (nordeste do Pará, Brasil)

A conservação e gestão da zona costeira da região amazônica merecem atenção especial, devido à riqueza de seus recursos naturais. O presente estudo visa avaliar os impactos dos eventos naturais e atividades humanas na praia de Atalaia, situada no estado do Pará (Brasil), e o desenvolvimento de diretrizes para a implementação de programas de gestão costeira. Os dados foram coletados entre novembro/2008 e novembro/2010. Quatro conjuntos de variáveis foram avaliados: (i) variáveis físicas (climatologia, hidrodinâmica e morfodinâmica), (ii) variáveis hidrológicas (temperatura da água, salinidade, pH, turbidez, oxigênio dissolvido e nutrientes inorgânicos dissolvidos, clorofila a e níveis de coliformes termotolerantes), (iii) desenvolvimento urbano e (iv) distribuição espacial de serviços e infraestrutura. Os resultados indicam que o clima e as condições hidrodinâmicas foram os principais fatores responsáveis pelas flutuações na qualidade de água, turbidez, oxigênio dissolvido, nutrientes inorgânicos dissolvidos e concentrações de clorofila a. A descarga de esgoto doméstico não tratado foi responsável pela contaminação bacteriológica, embora a rápida turbulência decorrente da alta energia hidrodinâmica do ambiente tenha limitado a contaminação por coliformes termotolerantes. Esta alta energia hidrodinâmica, principalmente durante as marés equinociais de sizígia e a falta de planejamento urbano gera outros problemas, tais como a erosão costeira. A área de estudo é caracterizada por altas taxas pluviométricas (> 1900 mm durante a estação chuvosa), ventos de NE com velocidades médias mensais superiores a 4,36 m/s na estação seca e 3,06 m/s na estação chuvosa, condições de macromaré (alcance da maré > 4,0 m), velocidades moderadas de correntes de maré (superior a 0,5 m/s) e alturas de ondas significantes superior a 1,5 m. Em março e junho (meses chuvosos), a corrente de maré vazante alcançou um máximo de 0,4 m/s. O ciclo de maré foi fracamente assimétrico com a maré vazante durando mais de 6 horas e 40 minutos. A energia das ondas foram fracamente moduladas pela maré baixa devido à atenuação das ondas em bancos de areia. A temperatura da água foi relativamente homogênea (27,4°C a 29,3°C). A salinidade variou de 5,7 (junho) a 37,4 (novembro). A água foi bem oxigenada (superior a 9,17 mg/L), turva (superior a 118 NTU), alcalina (acima de 8,68) e eutrófica (máximo de 2,36 μmol/L para nitrito, 24,34 μmol/L para nitrato, 0,6 μmol/L para fosfato e 329,7 μmol/L para silicato), além de apresentar altas concentrações de clorofila a (acima de 82 mg/m³). As condições naturais observadas no presente estudo indicam a necessidade de uma revisão dos critérios hidrológicos usados para avaliação de praias por agências nacionais e internacionais e sua adaptação para a realidade da costa amazônica. A falta de sistema de saneamento público levou a contaminação bacteriológica e a perda da qualidade da água. Com relação ao estado morfodinâmico, as condições dissipativas foram encontradas durante alta a moderada energia hidrodinâmica (condições equinociais e não-equinociais), porém em novembro as maiores alturas de ondas geraram características de barred dissipative, enquanto nos outros meses características nonbarred foram dominantes. Desta forma, o modelo proposto por Masselink & Short (1993) parece não ser ideal para ser aplicado em praias com características similares a praia de Atalaia, na qual a energia das ondas é modulada pela presença de bancos de areia durante algumas fases da maré.