Gellan gum-coated gold nanorods: an intracellular nanosystem for bone tissue engineering

Gold nanorods (AuNRs) have emerged as an exceptional nanotool for a myriad of applications ranging from cancer therapy to tissue engineering. However, their surface modification with biocompatible and stabilizing biomaterials is crucial to allow their use in a biological environment. Herein, low-acyl gellan gum (GG) was used to coat AuNRs surface, taking advantage of its stabilizing, biocompatible and gelling features. The layer-by-layer based strategy implied the successive deposition of poly(acrylic acid), poly(allylamine hydrochloride) and GG, which allowed the formation of a GG hydrogel-like shell with 7 nm thickness around individual AuNRs. Stability studies in a wide range of pH and salt concentrations showed that the polysaccharide coating can prevent AuNRs aggregation. Moreover, a reversible pH-responsive feature of the nanoparticles was observed. Cytocompatibility and osteogenic ability of GG-coated AuNRs was also addressed. After 14 days of culturing within SaOS-2, an osteoblast-like cell line, in vitro studies revealed that AuNRs-GG exhibit no cytotoxicity, were internalized by the cells and localized inside lysosomes. AuNRs-GG combined with osteogenic media enhanced the mineralization capacity two-fold, as compared to cells exposed to osteogenic media alone. The proposed system has shown interesting features for osteogenesis, and further insights might be relevant for drug delivery, tissue engineering and regenerative medicine.

Evaluation of two immunoassays for the measurement of human chorionic gonadotropin in urine for anti-doping purposes.

Background: Urinary human chorionic gonadotropin (hCG) concentration is routinely measured in all anti-doping laboratories to exclude the misuse of recombinant or urinary hCG preparations. In this study, extended validation of two commercial immunoassays for hCG measurements in urine was performed. Both tests were initially designed for hCG determination in human serum/plasma. Methods: Access (R) and Elecsys (R) 1010 are two automated immunoanalysers for central laboratories. The limits of detection and quantification, as well as intra-laboratory and inter-technique correlation, precision, and accuracy, were determined. Stability studies of hCG in urine following freezing and thawing cycles (n = 3) as well as storage conditions at room temperature, 4 degrees C and 20 degrees C, were performed. Results: Statistical evaluation of hCG concentrations in male urine samples (n = 2429) measured with the Elecsys (R) 1010 system enabled us to draw a skewed frequency histogram and establish a far outside value equal to 2.3 IU/L. This decision limit corresponds to the concentration at which a sportsman will be considered positive for hCG. Intra-assay precision for the Access (R) analyser was less than 4.0 A, whereas the inter-assay precision was closer to 4.5 % (concentrations of the official external controls contained between 5.5 and 195.0 IU/L). Intra and inter-assay precision for the Elecsys (R) 1010 analyser was slightly better. A good inter-technique correlation was obtained when measuring various urine samples (male and female). No urinary hCG loss was observed after two freeze/thaw cycles. On the other hand, time and inappropriate storage conditions, such as temperatures above 10 degrees C for more than 5 days, can deteriorate urinary hCG. Conclusions: Both analysers showed acceptable performances and are suitable for screening urine for anti-doping analyses. Each laboratory should validate and establish its own reference values because hCG concentrations measured in urine can be different from one immunoassay to another. The time delay between urine collection and analysis should be reduced as much as possible, and urine samples should be transported in optimal conditions to avoid a loss of hCG immunoreactivity.

Fate of Hydrophilic Nanoparticles in Biological Environments

The nanoparticles developed are based on chitosan, a biocompatible and biodegradable polysaccharide. The chitosan nanoparticles are formed in an entirely water-based process by electrostatic interactions with other biocompatible molecules. As a prerequisite to understand the fate of such nanoparticles in cells, comprehensive characterization and stability studies serve to identify quantitatively the impact of the raw material characteristics and preparation conditions on the nanoparticle characteristics. Methods included H-1 NMR spectroscopy, dilution viscometry, particle size analysis and electron microscopy. Cytotoxicity and cell uptake experiments on RAW 264.7 murine macrophages and p23 murine endothelial cells were performed to investigate the correlation with nanoparticle characteristics and effect of surface decoration with alginate. Cytotoxicity was assessed by the MTT survival test; cell uptake was monitored by fluorescent microscopy using labeled polymers.

Determination of etoricoxib in human plasma using automated on-line solid-phase extraction coupled with LC-APCI/MS/MS

A liquid chromatography-tandem mass spectrometry method with atmospheric pressure chemical ionization (LC-APCI/MS/MS) was validated for the determination of etoricoxib in human plasma using antipyrin as internal standard, followed by on-line solid-phase extraction. The method was performed on a Luna C18 column and the mobile phase consisted of acetonitrile:water (95:5, v/v)/ammonium acetate (pH 4.0; 10 mM), run at a flow rate of 0.6 mL/min. The method was linear in the range of 1-5000 ng/mL (r²>0.99). The lower limit of quantitation was 1 ng/mL. The recoveries were within 93.72-96.18%. Moreover, method validation demonstrated acceptable results for the precision, accuracy and stability studies.

Determination of phenobarbital in human plasma by a specific liquid chromatography method: application to a bioequivalence study

A liquid chromatography method was developed and validated for the determination of phenobarbital in human plasma using phenytoin as internal standard. The drugs were extracted from plasma by liquid-liquid extraction and separated isocratically on a C12 analytical column, maintained at 35 ºC, with water:acetonitrile:methanol (58.8:15.2:26, v/v/v) as mobile phase, run at a flow rate of 1.2 mL/min with detection at 205 nm. The method was linear in the range of 0.1-4 μg/mL (r²=0.9999) and demonstrated acceptable results for the precision, accuracy and stability studies. The method was successfully applied for the bioequivalence study of two tablet formulations (test and reference) of phenobarbital 100 mg after single oral dose administration to healthy human volunteers.

Desenvolvimento e validação de método analítico por cromatografia líquida de alta eficiência (CLAE) para determinação de associação de ampicilinas em pó liofilizado para injeção

An analytical method has been developed and validated for the determination of an association of ampicillins in a lyophilized powder for injection by HPLC. The advantage of chromatographic method other than the microbiological one is that, it is possible to monitor precisely, out-of-specification results in quality control processes and also during stability studies, in which an association of ampicillins is present. The proposed HPLC method was developed by using forced degraded samples, in order to reach a selective analysis of ampicillins when in the presence of their degradation products. It was possible to detect benzatine and through indirect calculation, to determine the ampicillin sodium in the drug sample. The method showed to be selective, accurate, precise, robust and linear (from 45.92 to 36.04 μg mL-1 of total ampicillin and from 14.53 to 43.28 μg mL-1 of benzatine). The accuracy determined from recovery test, gave results in the range of 99.41% of total ampicillin to 100.31% of benzatine. Hence, it can be concluded that the proposed HPLC method is applicable for ampicillins determination.

Definição de shelf life para produtos químicos: a importância de um guia de estabilidade específico para o segmento

The chemical industries worldwide are passing through a very particular moment of re-adaptation due to the implementation of an European regulation called, Registration, Evaluation, Authorization and Restriction of Chemicals (REACH). In Brazil, the Brazilian Chemical Industry needs urgently a specific guide of chemical products stability. The main purpose of this work is to present a proposal of a guide of stability for chemical products based on the reference guides of the International Conference on Harmonization (ICH). Thus, this work proposes an innovation in terms of methodology which will be useful for shelf life definition purpose for chemical industry products.

Simultaneous determination of four phenolic compounds in extracts of aerial parts of Ipomoea pes-caprae (L.) R. Br. (Convolvulaceae) by HPLC-UV

Among other applications, Ipomoea pes-caprae is popularly used to treat jellyfish stings, supporting the development of a product for dermatological use. Hydroethanolic spray-dried extract was chosen for the further development of phytomedicines, and a stability-indicative HPLC-UV method was developed and validated for the determination of isoquercitrin and isochlorogenic acids A, B and C. The method was developed using a C18 column (250 x 4.6 mm, 5 µm) with an acetonitrile:water mobile phase at pH 3.0 in a gradient run. The four constituents and other unidentified components of the extract were appropriately resolved without interference of degradation products after stress tests (acid, alkali, neutral, oxidant, photolysis). The method showed linearity in the isoquercitrin concentration range from 5.0-50.0 µg mL-1, with adequate precision (RSD% < 2.5% for the intra- and inter-day studies), accuracy (recovery of 100.0 ± 2.0%), and robustness. Both the herbal drug and spray-dried extract of I. pes-caprae were subjected to stability studies in accelerated and long-term conditions over four months. The samples maintained their characteristics and marker contents (< 10% of variation).

Soursop juice stabilized with soy fractions: a rheologial approach

The potential use of soybean soluble polysaccharide (SSPS) as a stabilizer in acidic beverages was evaluated using rheological and stability studies. For this purpose, soy-based beverages were formulated with soy protein isolate (SPI) and soursop juice due to the low stability of this kind of dispersion. The influences of the concentrations of soybean soluble polysaccharide, calcium chloride, and soy protein isolate on the stability and rheology of soursop juice were evaluated using a factorial experimental design. Interactions between the concentrations of soybean soluble polysaccharide and soy protein isolate exerted a positive effect on the maximum Newtonian viscosity. The stability was positively influenced by the soybean soluble polysaccharide and soy protein isolate concentrations, but the interactions between soy protein isolate and CaCl2 also affected the sedimentation index. These results suggest that soybean soluble polysaccharide is effective in stabilizing fibers and proteins in acidic suspensions due to the increase in viscosity and steric effect caused by the formation of complexes between the soybean soluble polysaccharide and soy protein isolate.

Investigation of single tryptophan proteins encapsulated in TEOS-derived sol-gel matrices by fluorescence spectroscopy /

In the work reported here, optically clear, ultrathin TEOS derived sol-gel slides which were suitable for studies of tryptophan (Trp) fluorescence from entrapped proteins were prepared by the sol-gel technique and characterized. The monitoring of intrinsic protein fluorescence provided information about the structure and environment of the entrapped protein, and about the kinetics of the interaction between the entrapped protein and extemal reagents. Initial studies concentrated on the single Trp protein monellin which was entrapped into the sol-gel matrices. Two types of sol-gel slides, termed "wet aged", in which the gels were aged in buffer and "dry-aged", in which the gels were aged in air , were studied in order to compare the effect of the sol-gel matrix on the structure of the protein at different aging stages. Fluorescence results suggested that the mobility of solvent inside the slides was substantially reduced. The interaction of the entrapped protein with both neutral and charged species was examined and indicated response times on the order of minutes. In the case of the neutral species the kinetics were diffusion limited in solution, but were best described by a sum of first order rate constants when the reactions occurred in the glass matrix. For charged species, interactions between the analytes and the negatively charged glass matrix caused the reaction kinetics to become complex, with the overall reaction rate depending on both the type of aging and the charge on the analyte. The stability and conformational flexibility of the entrapped monellin were also studied. These studies indicated that the encapsulation of monellin into dry-aged monoliths caused the thermal unfolding transition to broaden and shift upward by 14°C, and causedthe long-term stability to improve by 12-fold (compared to solution). Chemical stability studies also showed a broader transition for the unfolding of the protein in dry-aged monoliths, and suggested that the protein was present in a distribution of environments. Results indicated that the entrapped proteins had a smaller range of conformational motions compared to proteins in solution, and that entrapped proteins were not able to unfold completely. The restriction of conformational motion, along with the increased structural order of the internal environment of the gels, likely resulted in the improvements in themial and long-term stability that were observed. A second protein which was also studied in this work is the metal binding protein rat oncomodulin. Initially, the unfolding behavior of this protein in aqueous solution was examined. Several single tryptophan mutants of the metal-binding protein rat oncomodulin (OM) were examined; F102W, Y57W, Y65W and the engineered protein CDOM33 which had all 12 residues of the CD loop replaced with a higher affinity binding loop. Both the thermal and the chemical stability were improved upon binding of metal ions with the order apo < Ca^^ < Tb^"^. During thermal denaturation, the transition midpoints (Tun) of Y65W appeared to be the lowest, followed by Y57W and F102W. The placement of the Trp residue in the F-helix in F102W apparently made the protein slightly more thermostable, although the fluorescence response was readily affected by chemical denaturants, which probably acted through the disruption of hydrogen bonds at the Cterminal end of the F-helix. Under both thermal and chemical denaturation, the engineered protein showed the highest stability. This indicated that increasing the number of metal ligating oxygens in the binding site, either by using a metal ion with a higher coordinatenumber (i.e. Tb^*) which binds more carboxylate ligands, or by providing more ligating groups, as in the CDOM33 replacement, produces notable improvements in protein stability. Y57W and CE)OM33 OM were chosen for further studies when encapsulated into sol-gel derived matrices. The kinetics of interaction of terbium with the entrapped proteins, the ability of the entrapped protein to binding terbium, as well as thermal stability of these two entrapped protein were compared with different levels of Ca^"*^ present in the matrix and in solution. Results suggested that for both of the proteins, the response time and the ability to bind terbium could be adjusted by adding excess calcium to the matrix before gelation. However, the less stable protein Y57W only retained at most 45% of its binding ability in solution while the more stable protein CDOM33 was able to retain 100% binding ability. Themially induced denaturation also suggested that CDOM33 showed similar stability to the protein in solution while Y57W was destabilized. All these results suggested that "hard" proteins (i.e. very stable) can easily survive the sol-gel encapsulation process, but "soft" proteins with lower thermodynamic stability may not be able to withstand the sol-gel process. However, it is possible to control many parameters in order to successfully entrap biological molecules into the sol-gel matrices with maxunum retention of activity.

Topological Invariants in Hydrodynamics and Hydromagnetics

In this thesis we are studying possible invariants in hydrodynamics and hydromagnetics. The concept of flux preservation and line preservation of vector fields, especially vorticity vector fields, have been studied from the very beginning of the study of fluid mechanics by Helmholtz and others. In ideal magnetohydrodynamic flows the magnetic fields satisfy the same conservation laws as that of vorticity field in ideal hydrodynamic flows. Apart from these there are many other fields also in ideal hydrodynamic and magnetohydrodynamic flows which preserves flux across a surface or whose vector lines are preserved. A general study using this analogy had not been made for a long time. Moreover there are other physical quantities which are also invariant under the flow, such as Ertel invariant. Using the calculus of differential forms Tur and Yanovsky classified the possible invariants in hydrodynamics. This mathematical abstraction of physical quantities to topological objects is needed for an elegant and complete analysis of invariants.Many authors used a four dimensional space-time manifold for analysing fluid flows. We have also used such a space-time manifold in obtaining invariants in the usual three dimensional flows.In chapter one we have discussed the invariants related to vorticity field using vorticity field two form w2 in E4. Corresponding to the invariance of four form w2 ^ w2 we have got the invariance of the quantity E. w. We have shown that in an isentropic flow this quantity is an invariant over an arbitrary volume.In chapter three we have extended this method to any divergence-free frozen-in field. In a four dimensional space-time manifold we have defined a closed differential two form and its potential one from corresponding to such a frozen-in field. Using this potential one form w1 , it is possible to define the forms dw1 , w1 ^ dw1 and dw1 ^ dw1 . Corresponding to the invariance of the four form we have got an additional invariant in the usual hydrodynamic flows, which can not be obtained by considering three dimensional space.In chapter four we have classified the possible integral invariants associated with the physical quantities which can be expressed using one form or two form in a three dimensional flow. After deriving some general results which hold for an arbitrary dimensional manifold we have illustrated them in the context of flows in three dimensional Euclidean space JR3. If the Lie derivative of a differential p-form w is not vanishing,then the surface integral of w over all p-surfaces need not be constant of flow. Even then there exist some special p-surfaces over which the integral is a constant of motion, if the Lie derivative of w satisfies certain conditions. Such surfaces can be utilised for investigating the qualitative properties of a flow in the absence of invariance over all p-surfaces. We have also discussed the conditions for line preservation and surface preservation of vector fields. We see that the surface preservation need not imply the line preservation. We have given some examples which illustrate the above results. The study given in this thesis is a continuation of that started by Vedan et.el. As mentioned earlier, they have used a four dimensional space-time manifold to obtain invariants of flow from variational formulation and application of Noether's theorem. This was from the point of view of hydrodynamic stability studies using Arnold's method. The use of a four dimensional manifold has great significance in the study of knots and links. In the context of hydrodynamics, helicity is a measure of knottedness of vortex lines. We are interested in the use of differential forms in E4 in the study of vortex knots and links. The knowledge of surface invariants given in chapter 4 may also be utilised for the analysis of vortex and magnetic reconnections.

The impact of deregulation and re-regulation on bank efficiency: evidence from Asia

Following the 1997 crisis, banking sector reforms in Asia have been characterised by the emphasis on prudential regulation, associated with increased financial liberalisation. Using a panel data set of commercial banks from eight major Asian economies over the period 2001-2010, this study explores how the coexistence of liberalisation and prudential regulation affects banks’ cost characteristics. Given the presence of heterogeneity of technologies across countries, we use a stochastic frontier approach followed by the estimation of a deterministic meta-frontier to provide ‘true’ estimates of bank cost efficiency measures. Our results show that the liberalization of bank interest rates and the increase in foreign banks' presence have had a positive and significant impact on technological progress and cost efficiency. On the other hand, we find that prudential regulation might adversely affect bank cost performance. When designing an optimal regulatory framework, policy makers should combine policies which aim to foster financial stability without hindering financial intermediation.

Desenvolvimento de sistemas de pastas leves para aplicação em cimentação de poços petrolíferos com baixo gradiente de fratura

All around the world, naturally occurring hydrocarbon deposits, consisting of oil and gas contained within rocks called reservoir rocks , generally sandstone or carbonate exists. These deposits are in varying conditions of pressure and depth from a few hundred to several thousand meters. In general, shallow reservoirs have greater tendency to fracture, since they have low fracture gradient, ie fractures are formed even with relatively low hydrostatic columns of fluid. These low fracture gradient areas are particularly common in onshore areas, like the Rio Grande do Norte basin. During a well drilling, one of the most favorable phases for the occurrence of fractures is during cementing, since the cement slurry used can have greater densities than the maximum allowed by the rock structure. Furthermore, in areas which are already naturally fractured, the use of regular cement slurries causes fluid loss into the formation, which may give rise to failures cementations and formation damages. Commercially, there are alternatives to the development of lightweight cement slurries, but these fail either because of their enormous cost, or because the cement properties were not good enough for most general applications, being restricted to each transaction for which the cement paste was made, or both reasons. In this work a statistical design was made to determine the influence of three variables, defined as the calcium chloride concentration, vermiculite concentration and nanosilica concentration in the various properties of the cement. The use of vermiculite, a low density ore present in large amounts in northeastern Brazil, as extensor for cementing slurries, enabled the production of stable cements, with high water/cement ratio, excellent rheological properties and low densities, which were set at 12.5 lb / gal, despite the fact that lower densities could be achieved. It is also seen that the calcium chloride is very useful as gelling and thickening agent, and their use in combination with nanosilica has a great effect on gel strength of the cement. Hydrothermal Stability studies showed that the pastes were stable in these conditions, and mechanical resistance tests showed values of the order of up to 10 MPa

Nitric oxide release using natural rubber latex as matrix

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Synthesis, Characterization and Pharmacological Evaluation of 1-(2-Chloro-6-Fluorophenyl)-5-Methylindolin-2-One: A New Anti-Inflammatory Compound with Reduced Gastric Ulceration Properties

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)