Enhancing Therapeutic Efficacy against Brucella canis Infection in a Murine Model Using Rifampicin-Loaded PLGA Nanoparticles.

The in vivo efficacy of rifampicin encapsulated in poly(lactic-co-glycolic acid) (PLGA) nanoparticles was evaluated for the treatment of BALB/c mice experimentally infected with Brucella canis. The PLGA nanoparticles loaded with rifampicin (RNP) were prepared using the single emulsification-solvent evaporation technique, resulting in nanoparticles with a hydrodynamic diameter of 138 ± 6 nm. The zeta potential and polydispersity index values indicated that the system was relatively stable with a narrow size distribution. The release of rifampicin from the nanoparticles was studied in phosphate buffer at pH 7.4 and 37 °C. The release profile showed an initial burst phase, followed by a slower release stage attributed to nanoparticle degradation and relaxation, which continued for approximately 30 days until complete drug release. A combined model of rifampicin release, accounting for both the initial burst and the degradation-relaxation of the nanoparticles, effectively described the experimental data. The efficacy of RNP was studied in vivo; infected mice were treated with free rifampicin at concentrations of 2 mg per kilogram of mice per day (C1) and 4 mg per kilogram of mice per day (C2), as well as equivalent doses of RNP. Administration of four doses of the nanoparticles significantly reduced the B. canis load in the spleen of infected BALB/c mice. RNP demonstrated superior effectiveness compared to the free drug in the spleen, achieving reductions of 85.4 and 49.4%, respectively, when using C1 and 93.3 and 61.8%, respectively, when using C2. These results highlight the improved efficacy of the antibiotic when delivered through nanoparticles in experimentally infected mice. Therefore, the RNP holds promise as a potential alternative for the treatment of B. canis.

Spatiotemporal Temperature Distribution of NIR Irradiated Polypyrrole Nanoparticles and Effects of pH.

The spatiotemporal temperature distributions of NIR irradiated polypyrrole nanoparticles (PPN) were evaluated by varying PPN concentrations and the pH of suspensions. The PPN were synthesized by oxidative chemical polymerization, resulting in a hydrodynamic diameter of 98 ± 2 nm, which is maintained in the pH range of 4.2-10; while the zeta potential is significantly affected, decreasing from 20 ± 2 mV to -5 ± 1 mV at the same pH range. The temperature profiles of PPN suspensions were obtained using a NIR laser beam (1.5 W centered at 808 nm). These results were analyzed with a three-dimensional predictive unsteady-state heat transfer model that considers heat conduction, photothermal heating from laser irradiation, and heat generation due to the water absorption. The temperature profiles of PPN under laser irradiation are concentration-dependent, while the pH increase only induces a slight reduction in the temperature profiles. The model predicts a value of photothermal transduction efficiency (η) of 0.68 for the PPN. Furthermore, a linear dependency was found for the overall heat transfer coefficient (U) and η with the suspension temperature and pH, respectively. Finally, the model developed in this work could help identify the exposure time and concentration doses for different tissues and cells (pH-dependent) in photothermal applications.

Synthesis and Characterization of a Bioconjugate Based on Oleic Acid and L-Cysteine.

One of the main challenges facing materials science today is the synthesis of new biodegradable and biocompatible materials capable of improving existing ones. This work focused on the synthesis of new biomaterials from the bioconjugation of oleic acid with L-cysteine using carbodiimide. The resulting reaction leads to amide bonds between the carboxylic acid of oleic acid and the primary amine of L-cysteine. The formation of the bioconjugate was corroborated by Fourier transform infrared spectroscopy (FTIR), Raman spectroscopy, and nuclear magnetic resonance (NMR). In these techniques, the development of new materials with marked differences with the precursors was confirmed. Furthermore, NMR has elucidated a surfactant structure, with a hydrophilic part and a hydrophobic section. Ultraviolet-visible spectroscopy (UV-Vis) was used to determine the critical micellar concentration (CMC) of the bioconjugate. Subsequently, light diffraction (DLS) was used to analyze the size of the resulting self-assembled structures. Finally, transmission electron microscopy (TEM) was obtained, where the shape and size of the self-assembled structures were appreciated.

PLGA nanoparticle preparations by emulsification and nanoprecipitation techniques: effects of formulation parameters.

This study presents the influence of the primary formulation parameters on the formation of poly-dl-lactic-co-glycolic nanoparticles by the emulsification-solvent evaporation, and the nanoprecipitation techniques. In the emulsification-solvent evaporation technique, the polymer and tensoactive concentrations, the organic solvent fraction, and the sonication amplitude effects were analyzed. Similarly, in the nanoprecipitation technique the polymer and tensoactive concentrations, the organic solvent fraction and the injection speed were varied. Additionally, the agitation speed during solvent evaporation, the centrifugation speeds and the use of cryoprotectants in the freeze-drying process were analyzed. Nanoparticles were characterized by dynamic light scattering, laser Doppler electrophoresis, and scanning electron microscopy, and the results were evaluated by statistical analysis. Nanoparticle physicochemical characteristics can be adjusted by varying the formulation parameters to obtain specific sizes and stable nanoparticles. Also, by adjusting these parameters, the nanoparticle preparation processes have the potential to be tuned to yield nanoparticles with specific characteristics while maintaining reproducible results.

Thermo-Magnetic Properties of Fe3O4@Poly(N-Isopropylacrylamide) Core-Shell Nanoparticles and Their Cytotoxic Effects on HeLa and MDA-MB-231 Cell Lines.

In this study, we performed a systematic evaluation of the synthesis parameters of a multiresponsive core-shell nanoplatform (Fe3O4 nanoparticles coated by poly(N-isopropylacrylamide). These nanocomposites allow the possibility of controlling specific properties, such as particle size and morphology. The polymer coating was performed by in-situ free-radical polymerization of N-isopropylacrylamide on Fe3O4 nanoparticles to obtain a nanocomposite with a core-shell structure. We monitored the size and morphology of the nanocomposite by scanning transmission electron microscopy. Electrophoretic mobility determined the evolution of ζ-potentials during coating. We assessed the functionalization of the Fe3O4 nanoparticles by Fourier-transform infrared spectroscopy and ultraviolet-visible spectroscopy. The core-shell systems exhibited a collapsed structure when heated above the lower critical solution temperature, which was determined by dynamic light scattering. Squid based vibrating sample magnetometer tests were performed to verify superparamagnetic behavior at room temperature. The platform was approved as biocompatible for the HeLa and MDA-MB-231 cell lines by MTT assays, and it shows the desired properties for potential use in localized and controlled drug delivery.

Differential Response of BEAS-2B and H-441 Cells to Methylene Blue Photoactivation.

BACKGROUND/AIM: Cancer incidence and mortalities are growing worldwide, therefore research and development of more effective and less invasive treatments, such as photodynamic therapy, are needed. Herein, we investigated the methylene blue (MB) photoactivation effects in lung epithelial cells (BEAS-2B) and lung adenocarcinoma cells (H-441). MATERIALS AND METHODS: The reactive oxygen species (ROS) produced by the laser photoactivation of MB in aqueous solutions and cell cultures were measured with probes, and the cell viability was evaluated with a colorimetric assay. RESULTS: MB up to 31.26 µM did not induce detectable effects in BEAS-2B cells. However, H-441 cells presented adverse effects below that concentration in the same range of fluencies studied. These results are in concordance with the ROS production in H-441 cells, while in BEAS-2B cells the production of ROS was less significant compared to the control. CONCLUSION: Photoactivation of MB at concentrations below 31.26 µM could be used for the selective treatment of H-441 cells over non-cancer cells.

Mathematical modeling and parametrical analysis of the temperature dependency of control drug release from biodegradable nanoparticles.

In this study we describe a mathematical analysis that considers the temperature effects of the controlled drug release process from biodegradable poly-d,l-lactide-co-glycolide (PLGA) nanoparticles. Temperature effects are incorporated and applied to two drug release models. The first one consists of a two-stage release process that considers only simultaneous contributions of initial burst and nanoparticle degradation-relaxation (BR model). The second one is a three release stage model that considers, additionally, a simultaneous drug diffusion (BRD model) step. In these models, the temperature dependency of the release parameters, initial burst constant, k b, the rate of degradation-relaxation constant, k r, time to achieve 50% of release, t max, and effective diffusion coefficient constant (D e), are determined using mathematical expressions analogous to the Arrhenius equation. The temperature dependent models are used to analyze the release of previously encapsulated Rhodamine 6G dye as a model drug in polyethylene glycol modified PLGA nanoparticles. The experimental data used to develop the mathematical model was obtained from release studies carried out in phosphate buffer pH 7.4 at 37 °C, 47 °C, and 57 °C. Multiphasic release behaviors with an overall increase rate associated with the incubation temperature were observed. The study incorporates a parametrical analysis that can evaluate diverse temperature variation effects of the controlled release parameters for the two models.

Nanoparticle encapsulation and controlled release of a hydrophobic kinase inhibitor: Three stage mathematical modeling and parametric analysis.

A mathematical model of drug release that incorporates the simultaneous contributions of initial burst, nanoparticle degradation-relaxation and diffusion was developed and used to effectively describe the release of a kinase inhibitor and anticancer drug, PHT-427. The encapsulation of this drug into PLGA nanoparticles was performed by following the single emulsion-solvent evaporation technique and the release was determined in phosphate buffer pH 7.4 at 37 °C. The size of nanoparticles was obtained in a range of 162-254 nm. The experimental release profiles showed three well defined phases: an initial fast drug release, followed by a nanoparticle degradation-relaxation slower release and then a diffusion release phase. The effects of the controlled release most relevant parameters such as drug diffusivity, initial burst constant, nanoparticle degradation-relaxation constant, and the time to achieve a maximum rate of drug release were evaluated by a parametrical analysis. The theoretical release studies were corroborated experimentally by evaluating the cytotoxicity effectiveness of the inhibitor AKT/PDK1 loaded nanoparticles over BxPC-3 pancreatic cancer cells in vitro. These studies show that the encapsulated inhibitor AKT/PDK1 in the nanoparticles is more accessible and thus more effective when compared with the drug alone, indicating their potential use in chemotherapeutic applications.

Nanoparticle delivery of an AKT/PDK1 inhibitor improves the therapeutic effect in pancreatic cancer.

The K-ras mutation in pancreatic cancer can inhibit drug delivery and increase drug resistance. This is exemplified by the therapeutic effect of PH-427, a small molecule inhibitor of AKT/PDK1, which has shown a good therapeutic effect against a BxPC3 pancreatic cancer model that has K-ras, but has a poor therapeutic effect against a MiaPaCa-2 pancreatic cancer model with mutant K-ras. To increase the therapeutic effect of PH-427 against the MiaPaCa-2 pancreatic cancer model with mutant K-ras, we encapsulated PH-427 into poly(lactic-co-glycolic acid) nanoparticles (PNP) to form drug-loaded PH-427-PNP. PH-427 showed a biphasic release from PH-427-PNP over 30 days during studies in sodium phosphate buffer, and in vitro studies revealed that the PNP was rapidly internalized into MiaPaCa-2 tumor cells, suggesting that PNP can improve PH-427 delivery into cells harboring mutant K-ras. In vivo studies of an orthotopic MiaPaCa-2 pancreatic cancer model showed reduced tumor load with PH-427-PNP as compared with treatment using PH-427 alone or with no treatment. Ex vivo studies confirmed the in vivo results, suggesting that PNP can improve drug delivery to pancreatic cancer harboring mutant K-ras.

Pre-exposure to nitrite in the absence of ammonium strongly inhibits anammox.

Anaerobic ammonium oxidizing bacteria (Anammox) are known to be inhibited by their substrate, nitrite. However, the mechanism of inhibition and the physiological conditions under which nitrite impacts the performance of anammox bioreactors are still unknown. This study investigates the role of pre-exposing anammox bacteria to nitrite alone on their subsequent activity and metabolism after ammonium has been added. Batch experiments were carried out with anammox granular biofilm pre-exposed to nitrite over a range of concentrations and durations in the absence of ammonium. The effect of pre-exposure to nitrite alone compared to nitrite simultaneously fed with ammonium was evaluated by measuring the anammox activity and the accumulation of the intermediate, nitric oxide. The results show that the inhibitory effect was more dramatic when bacteria were pre-exposed to nitrite in absence of ammonium, as revealed by the lower activity and the higher accumulation of nitric oxide. The nitrite concentration causing 50% inhibition was 53 and 384 mg N L(-1) in the absence or the presence of ammonium, respectively. The nitrite inhibition was thus 7.2-fold more severe in the absence of ammonium. Biomass exposure to nitrite (25 mg N L(-1)), in absence of ammonium, led to accumulation of nitric oxide. On the other hand when the biomass was exposed to nitrite in presence of ammonium, accumulation of nitric oxide was only observed at much higher nitrite concentrations (500 mg N L(-1)). The inhibitory effect of nitrite in the absence of ammonium was very rapid. The rate of decay of the anammox activity was equivalent to the diffusion rate of nitrite up to 46% of activity loss. The results taken as a whole suggest that nitrite inhibition is more acute when anammox cells are not actively metabolizing. Accumulation of nitric oxide in the headspace most likely indicates disruption of the anammox biochemistry by nitrite inhibition, caused by an interruption of the hydrazine synthesis step.

Breakthrough performance of large proteins on ion-exchange membrane columns.

Protein adsorption of large proteins on ion-exchange membrane columns was theoretically and experimentally investigated using batch and fixed-bed systems. Thyroglobulin was used as the model protein. The study strongly suggests that part of the protein is physically retained inside the column during frontal mode operation. These experimental results were used to obtain a filtration function of the chromatographic system. In the theoretical analysis of the frontal protein adsorption, a model was integrated by the serial coupling of the membrane-transport model, the filtration model and the system-dispersion model. Two different techniques were employed in the estimation of the maximum adsorption capacity, the equilibrium desorption constant and the forward interaction rate constant, which are the parameters of the membrane-transport model. The fit of the model to the experimental data was not possible using the equilibrium parameters obtained in the batch experiments. The parameter estimation using a simplex optimization routine coupled to the solution of the partial differential model equations yields full prediction of the adsorption phenomena.