69 research outputs found
Structural and functional stabilization of glycomacropeptide via encapsulation within multiple emulsions
Bovine glycomacropeptide (GMP), derived from whey proteins, has been demonstrated to possess an interesting bioactivity that has attracted a lot of attention over the last few years. In particular, its ability to bind Vibrio cholerae and Escherichia coli enterotoxins, inhibit bacterial and viral adhesion, suppress gastric secretions, promote bifidobacterial growth and modulate immune system responses. Of these, protection against toxins, bacteria and viruses, and modulation of the immune system, are the most promising applications for this bioactive dairy macropeptide. The development of strategies that may allow its structural and functional stabilization via nanoencapsulation within multiple emulsions may increase its food and biopharmaceutical applicabilities. In this research effort, bovine GMP was (thermodynamically) stabilized via entrapment within water-in-oil-in-water (W/O/W) multiple emulsions aiming at mimicking the multifunctional design of biology, with several lipid matrices, and stabilizing layer compositions. Due to their compartimentalized internal structure, multiple emulsions are ideal for encapsulation since they can carry both polar and non-polar (bio)molecules. The composition of the stabilizing layer of the nanosystem was changed by using different poloxamers and proportions of lecithin. Physicochemical characterization of the optimized GMP-encasing nanovesicle formulations encompassed determination of Zeta potential and particle hydrodynamic size over storage time, surface morphology via CRYO-SEM, and microcalorimetric analysis via DSC
Structural and functional stabilization of protein entities
XI ReuniĂŁo Regional Nordeste da SBBq | 4th International Symposium in Biochemistry of Macromolecules and BiotechnologyStabilization of protein and protein-like molecules translates into preservation of both
structure and functionality during storage and/or targeting, and such stabilization is
mostly attained through establishment of a thermodynamic equilibrium with the
(micro)environment. The basic thermodynamic principles that govern protein structural
transitions and the interactions of the protein and/or peptide molecule with its
(micro)environment will, therefore, be tackled. Protein stabilization is based upon
dampening the molecular motions and, therefore, eliminating conformational transitions
while the molecule is still in the native 3D (folded) state. The 3D structure of a protein
molecule depends mostly on two types of interactions: intramolecular interactions
between aminoacid moieties and intermolecular interactions with solute and/or solvent
molecules present in its microenvironment. Stabilizing a biomolecule (aiming at
preserving its function) involves dampening its molecular motions, and this can be
achieved by reducing the chemical activity of the water present in its microenvironment,
thus stabilizing both its structure and functionality. Recently, the simultaneous
entrapment-stabilization of proteins and enzymes based on nanoencapsulation in a
nanoemulsion (W/O/W) matrix with an hydrophilic core has started to gain momentum.
Similarly to the stabilization mechanism of osmolytes, in nanoencapsulation the water
activity is altered thus affecting the molecular motions of the proteins. Highlights will also
be given to structural and functional stabilization of protein entities (viz. enzymes,
(macro)peptides, (recombinant) proteins, and bacteriophages) by chemical
methodologies. Modification of the biomolecules microenvironment via multipoint
covalent attachment onto a solid surface followed by hydrophylic polymer coimmobilization,
are some of the (latest) strategies that will be discussed.info:eu-repo/semantics/publishedVersio
Biopolymeric matrices for structural and functional stabilization of bacteriophages
In the recent past years, bacteriophage research has experienced a renaissance due to their potential application in the pharmaceutical field, especially with the increase of bacterial resistance to antibiotics and the possibility to take part in new methods of early detection and diagnosis of bacterial infections. In that context, the structural and functional stabilization of bacteriophages using biopolymeric microporous hydrogels represents a promising research focus with a broad potential biomedical/ biopharmaceutical application. The scope of this work was to develop biopolymeric non-toxic phage-hydrogels of agar and sodium alginate, obtained at neutral pH and mild polymerization conditions, in order to offer adequate characteristics to the maintenance of phageâs lytic activity. Disc-like phage-hydrogels were prepared, with a phage and polymer concentration of 1.3x108 PFU/ml and 1.5% (w/v), respectively. Regarding the alginate hydrogels, CaCO3 (22.5 mM) and GDL (48 mM) were also included in the formulation. Agar hydrogels were prepared naturally by jellification, as a function of temperature lowering, and alginate hydrogels were prepared by internal gelation. The matrices were inoculated with a suspension of susceptible (host) bacteria and incubated at 37 ÂșC for 24h. Observation of bacterial lawnâs lysis demonstrated that bacteriophages kept their lytic activity, being the method of physical entrapment able to promote their stabilization. Cryo-SEM analysis revealed that both types of phage-hydrogels present interconnective microporous network, which guaranties a facilitated access of the phages to the bacteria, ensuring an efficient lysis of the host bacteria present in the surface of the hydrogels.The developed hydrogels also present appropriate physical and chemical properties for a wider variety of applications in the field of pharmaceutical sciences, such as controlled release of (macro)molecules, cell immobilization and 3D support for tissue regeneration
BacteriĂłfagos no tratamento de feridas
As infecçÔes bacterianas, particularmente as causadas
por bactérias resistentes aos antibióticos, permanecem a
principal causa de morte entre pacientes hospitalizados com
queimaduras e feridas. Para alĂ©m da terapĂȘutica sistĂ©mica, um
elemento-chave na gestão de feridas infectadas é a aplicação
local de antimicrobianos eficazes. Os bacteriĂłfagos (ou fagos)
tĂȘm demonstrado um elevado potencial de cura no tratamento
de feridas infectadas com estirpes bacterianas resistentes aos
antibiĂłticos.Bacterial infections, particularly the ones caused by antibiotic
resistant bacteria, remain as the main cause of death among
hospitalized patients with burns and ulcers. Besides systemic
therapy, a key element on wound management is the local
application of effective antimicrobial agents. Bacteriophages
(or phages) have been shown as promising candidates (used
alone or as complementary to antibiotic therapy) to target
antibiotic-resistant bacteria on wound treatment
Kinetics of b-galactosidase immobilized on polysiloxanepolyvinyl alcohol magnetic composite â POS-PVAM
b-Galactosidase is an enzyme with a wide industrial application, mostly in the
hydrolysis of lactose and, more recently, in the synthesis of oligosaccharides.
Several advantages are associated with the application of immobilized enzymes.
In this work, b-Galactosidase was covalently immobilized onto a POS-PVAM using
glutaraldehyde as activating agent and its hydrolytic properties evaluated. For both
soluble and immobilized b-Galactosidase, the optimal temperature and pH were
found to be 50 ÂșC and 6.5, respectively. The immobilized enzyme showed to be
more resistant than the soluble form when hydrolysis experiments were performed
out within the above optimal condition, being the observed difference in activity
more pronounced for temperatures higher than 50ÂșC. An enhancement of the
thermal stability of the immobilized enzyme was also observed. The apparent Km
and Ea for both soluble (7.377 ± 1.303mM and 25.51 ± 8.72Kj mol-1) and
immobilized enzyme (7.841 ± 1.189mM and 32.61 ± 5.82Kj mol-1) showed to be
not significantly different. The immobilization also proved to be advantageous as,
after twenty reutilizations, the immobilized enzyme retained about 52% of its initial
activity. These results clearly demonstrate that POS-PVAM may be used for b-
Galactosidase immobilization since, besides improving the enzyme hydrolytic
properties, its separation from the obtained reaction products is easier to
accomplish
Deformulation of a solid pharmaceutical form using computed tomography and X-ray fluorescence
Deformulation of medicines is of undeniable importance, since it can be utilized both to unravel the chemical composition of the excipients integrating a pharmaceutical formulation of a specific medicine and as an important tool to conduct morphometric studies of the formulation under study. Such strategy may be utilized in analytical studies aiming at quantifying the components of reference drugs, or in the identification of putative counterfeit pharmaceuticals. Deformulation makes use of physicochemical analysis tools to characterize, from the chemical point of view, the components integrating medicine pharmaceutical formulations and from the physical point of view, the morphological part of the pharmaceutical formulation. The techniques of computer tomography (SkyScan 1174 - Bruker microCT) and X-ray fluorescence analyses (using an X-ray source with W-anode from Hammatsu Photonics and Silicon Drift detector from Amptek) were successfully used in performing a process of deformulation of a solid pharmaceutical formulation of tablets, utilized herein as a model medicine for controlled drug release. The analytical methods used in this work, proved their effectiveness for the main goal of this study, which aimed to characterize a pharmaceutical formulation via its deconstruction.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP, São Paulo, Brasil)
(FAPESP Ref. No. 2012/15651-4, TomĂłgrafo para AplicaçÔes na Ărea das CiĂȘncias FarmacĂȘutica;
FAPESP Ref. No. 2013/03181-6, Project PneumoPhageKill; FAPESP Ref. No. 2012/15651-4 and FAPESP Ref. No. 2013/19300-4, AuxĂlio
Ă Pesquisa - Reserva TĂ©cnica para Infra-estrutura
Institucional de Pesquisa) and by FINEP â
Financiadora de Estudos e Projetos (FINEP, Rio de Janeiro,
Brasil) (FINEP Ref. No. 01.13.0286.00
Galactooligosaccharides production by ÎČ-galactosidase immobilized onto magnetic polysiloxaneâpolyaniline particles
Magnetized polysiloxane coated with polyaniline (mPOSâPANI) was used as a support for ÎČ-galactosidase immobilization via glutaraldehyde. The galactooligosaccharides (GOS) production by this derivative was investigated under different initial lactose concentrations (5â50%) and temperatures (30â60 °C). The initial lactose concentration in the reaction media affected the total amounts of produced GOS and their time course production was described as a âbell-shapedâ curve as a result of the balance between transgalactosylation and hydrolysis. No significative difference was observed for the free and immobilized enzymes. The reaction rates for lactose hydrolysis and GOS formation increased with increasing temperature from 30 °C to 60 °C, but GOS production at all lactose conversion levels was almost unchanged with changing temperature. The mPOSâPANI matrix was also characterized by scanning electronic microscopy (SEM), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), vibrating sample magnetometry (VSM), thermomagnetization, differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA).European Union Programme of High Level
Scholarships for Latin America - Programme
Alban ((Scholarship No. E05D057787BR)Brazilian National Research Council (CNPq
Development and characterization of a hydrogel containing nitrofurazone for antimicrobial topical applications
The goal of the research work entertained herein was the development and characterization of a poly-(vinyl alcohol)
(PVA) hydrogel cross-linked with glutaraldehyde and impregnated with 0.2% (w/w) nitrofurazone (NTZ), for topical
applications. To verify the active principle release capability, one has determined (i) swelling profile, (ii) in vitro release
of NTZ via UV-VIS spectrophotometry, and (iii) antimicrobial activity via exposure to the hydrogel of ATCC
strains of Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa. The optimized hydrogel was further
characterized via scanning electron microscopy (SEM), infrared spectroscopy with Fourier transform, moisture content
determinations and thermal analyses via thermal gravimetry (TGA). Swelling tests revealed a mass increase from 100±5%
up to 350±11%. Incorporated NTZ displayed bactericidal activity, as expected, being released in a linearly controlled
fashion above 6 ÎŒg/mL during experiment timeframes of 14 h. SEM analyses allowed verification of a homogeneous surface
morphology, while infrared spectra showed that NTZ did not bind strongly to the cross-linked polymer. Furthermore,
results from thermal analyses suggested a loss of thermal stability arising from incorporation of NTZ in the hydrogel. The
optimized hydrogel exhibited characteristics with high potential for (antimicrobial) treatment of skin lesions.Financial support to Victor M. Balcao, via an Invited Research Scientist fellowship (FAPESP Ref. No. 2011/51077-8) by Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP, Sao Paulo, Brazil), is hereby gratefully acknowledged. Sebastiao Coelho de Lima gratefully acknowledges financial support from the Centro Universitario da Barra Mansa (Rio de Janeiro/RJ, Brazil)
Development and characterization of a gel formulation integrating microencapsulated nitrofurazone
Nitrofurazone (NTZ) is usually employed in the topical treatment of infected wounds and lesions of both skin and mucosa. Microencapsulation is a process utilized in the incorporation of active ingredients within polymers aiming at, among other objectives, the prolonged release of pharmaceutical compounds and protection from atmospheric agents (viz. moisture, light, heat and/or oxidation). With the goal of utilizing the microparticles containing encapsulated NTZ in pharmaceutical formulations, one prepared microparticles containing NTZ via ionotropic gelation of sodium alginate. The microparticles were characterized via scanning electron microscopy analyses, Fourier transform infrared spectroscopy (FTIR) analyses, via determination of encapsulation efficiency, and via thermal analyses (both TGA and DSC). The final gel formulation was also characterized rheologically. The extrusion/solidification technique employed to obtain the calcium alginate microparticles with encapsulated NTZ was found to be adequate, and produced an NTZ encapsulation efficiency of ca. 97.8% ± 1.1%. The calcium alginate microparticles thus obtained, with encapsulated NTZ, exhibited an oval shape and hydrodynamic diameters between 500 Όm and 800 Όm. From the thermal analyses performed, together with information from the infrared spectra, one may conclude that NTZ did not strongly bind to the polymer, which may be favorable for the release of the active ingredient. From the results obtained in the present research effort, one may conclude that the microparticles produced possess the potential to be utilized as carriers for NTZ in pharmaceutical formulations such as gels, ointments, and solutions.Financial support to Victor M. Balcao, via an Invited Research Scientist fellowship (FAPESP Ref. No. 2011/51077-8) by Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP, Sao Paulo, Brazil), is hereby gratefully acknowledged
Immobilization of b-galactosidase from kluyveromyces lactis onto a polysiloxaneâpolyvinyl alcohol magnetic (mPOSâPVA) composite for lactose hydrolysis
ÎČ-Galactosidase from Kluyveromyces lactis was covalently immobilized onto a mPOSâPVA, using glutaraldehyde as activating agent and its properties were evaluated. The enzymatic water insoluble derivative displayed the same optimum pH (6.5) and optimum temperature (50 °C) of the soluble enzyme. The apparent Km app and activation energy for both soluble and immobilized enzyme derivative were found to be not significantly different. The mPOSâPVA ÎČ-galactosidase preparation presented a higher operational and thermal stability than the soluble enzyme. This immobilized ÎČ-galactosidase also was effective in hydrolyzing lactose from milk. Hence, one can conclude that mPOSâPVA is an attractive and efficient support for ÎČ-galactosidase immobilization.Alban, the European Union Programme of High Level
Scholarships for Latin America; Brazilian National
Research Council (CNPq)
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