Exploring the Toxicity, Lung Distribution, and Cellular Uptake of Rifampicin and Ascorbic Acid-Loaded Alginate Nanoparticles as Therapeutic Treatment of Lung Intracellular Infections.

Nanotechnology is a very promising technological tool to combat health problems associated with the loss of effectiveness of currently used antibiotics. Previously, we developed a formulation consisting of a chitosan and tween 80-decorated alginate nanocarrier that encapsulates rifampicin and the antioxidant ascorbic acid (RIF/ASC), intended for the treatment of respiratory intracellular infections. Here, we investigated the effects of RIF/ASC-loaded NPs on the respiratory mucus and the pulmonary surfactant. In addition, we evaluated their cytotoxicity for lung cells in vitro, and their biodistribution on rat lungs in vivo after their intratracheal administration. Findings herein demonstrated that RIF/ASC-loaded NPs display a favorable lung biocompatibility profile and a uniform distribution throughout lung lobules. RIF/ASC-loaded NPs were mainly uptaken by lung macrophages, their primary target. In summary, findings show that our novel designed RIF/ASC NPs could be a suitable system for antibiotic lung administration with promising perspectives for the treatment of pulmonary intracellular infections.

Promising Chitosan-Coated Alginate-Tween 80 Nanoparticles as Rifampicin Coadministered Ascorbic Acid Delivery Carrier Against Mycobacterium tuberculosis.

The aim of this study was to design a nanocarrier system for inhalation delivery of rifampicin (RIF) in combination with ascorbic acid (ASC), namely constituted of sodium alginate coated with chitosan and Tween 80 (RIF/ASC NPs) as a platform for the treatment of pulmonary tuberculosis infection. A Box-Behnken experimental design and response surface methodology (RSM) were applied to elucidate and evaluate the effects of several factors on the nanoparticle properties. On the other hand, it was found that RIF/ASC NPs were less cytotoxic than the free RIF, showing a significantly improved activity against nine clinical strains of Mycobacterium tuberculosis (M. tb) in comparison with the free drug. RIF/ASC NPs had an average particle size of 324.0 ± 40.7 nm, a polydispersity index of 0.226 ± 0.030, and a zeta potential of - 28.52 ± 0.47 mV and the surface was hydrophilic. The addition of sucrose (1% w/v) to the nanosuspension resulted in the formation of a solid pellet easily redispersible after lyophilization. RIF/ASC NPs were found to be stable at different physiological pH values. In summary, findings of this work highlight the potential of the RIF/ASC NP-based formulation development herein to deliver RIF in combination with ASC through pulmonary route by exploring a non-invasive route of administration of this antibiotic, increasing the local drug concentrations in lung tissues, the primary infection site, as well as reducing the risk of systemic toxicity and hence improving the patient compliance.

Effect of Complexes and Microemulsions on the Permeability of Drugs: Determination Using a New Biomimetic Artificial Membrane.

The aim of this work was to predict the permeability of two model drugs, sulfamerazine (SMR) and indomethacin (INM), and to determine the effect on their apparent permeabilities by complexation with cyclodextrins and/or meglumine or incorporation in microemulsions. Permeation experiments were performed using two-chamber diffusion cells with a new composition of bio-mimetic membrane composed of 80% of Lipoid® S100 and 20% of cholesterol in n-octanol 10% w/w solution, at 37 ± 0.5°C and 14,000 rpm. The predictive capacity of the permeability of passive diffusion absorbed compounds was evaluated using 20 drug standards and showed an exponential correlation between the apparent permeability coefficients (Papp) and the fraction absorbed percentages in humans (Fa%), with an R2 value of 0.67942 and a constant value of - 4.1 ± 0.8. SMR and INM were classified as Class II and I, respectively, according to the Biopharmaceutical Classification System. These drugs were complexed and incorporated in microemulsions. The Fa% from all the drug products was higher than 90%. SMR in the complexes and both drugs in microemulsions were classified as highly soluble. Thus, SMR and INM incorporated in these pharmaceutical products could be classified as Class I.

Rate of vision loss in neovascular age-related macular degeneration explored.

PURPOSE: To explore decline in visual acuity in patients with neovascular age-related macular degeneration (n-AMD) awaiting intravitreal bevacizumab or ranibizumab treatment following initial diagnosis and after disease reactivation. METHODS: Retrospective analysis of 74 treatment-naïve patients (84 eyes) in two centers in Córdoba, Argentina. The time between treatment indication and intravitreal injection, and the changes in BCVA produced during this delay were studied in both periods. A linear regression model to search the impact of time on progression visual impairment was conducted. RESULTS: In both periods, a significant reduction in vision occurred awaiting intravitreal injection. The longer the delay, the greater the vision loss (R2 = 0.55 p < 0.01) and the less improvement following treatment (Pearson coefficient -0.26). The result of the model shows that the change in vision as a function of initial delay were best described by a polynomic model with a mean loss of 5 letters in the first 3 weeks, a slowdown in the rate of change of VA, and a dependence of visual acuity at the moment of diagnosis . The loss of visual acuity after reactivation shows the same behavior as at the onset of the disease but independent of visual acuity prior to reactivation. CONCLUSION: Visual loss awaiting injection intravitreal anti-VEGF is clinically significant and with an asymptotic pattern, with early rapid loss of vision in both the onset of the disease and the reactivation. Initiation of anti-VEGF treatment must be undertaken urgently, as should retreatment of disease activation to reduce visual loss.

Synergistic bactericidal combinations between gentamicin and chitosan capped ZnO nanoparticles: A promising strategy for repositioning this first-line antibiotic.

Gentamicin (GEN), a widely used broad-spectrum antibiotic, faces challenges amid the global emergency of antimicrobial resistance. This study aimed to explore the synergistic effects of zinc oxide nanoparticles (ZnO NPs) in combination with GEN on the bactericidal activity against various bacterial strains. Results showed ZnO NPs with MICs ranging from 0.002 to 1.5 µg/mL, while the precursor salt displayed a MIC range of 48.75-1560 µg/mL. Chitosan (CS)-capped ZnO NPs exhibited even lower MICs than their uncapped counterparts, with the CS-capped synthesized ZnO NPs demonstrating the lowest values. Minimal bactericidal concentrations (MBC) aligned with MIC trends. Combinations of CS-capped synthesized ZnO NPs and GEN proved highly effective, inhibiting bacterial growth at significantly lower concentrations than GEN or ZnO NPs alone. This phenomenon may be attributed to the conformation of CS on the ZnO NPs' surface, enhancing the positive particle surface charge. This possibly facilitates a more effective interaction between ZnO NPs and microorganisms, leading to increased accumulation of zinc and GEN within bacterial cells and an overproduction of reactive oxygen species (ROS). It's crucial to note that, while this study did not specifically involve resistant strains, its primary focus remains on enhancing the overall antimicrobial activity of gentamicin. The research aims to contribute to addressing the global challenge of antimicrobial resistance, recognizing the urgent need for effective strategies to combat this critical issue. The findings, particularly the observed synergy between ZnO NPs and GEN, hold significant implications for repositioning the first-line antibiotic GEN.

Magnetic layered double hydroxides with carbamazepine for breast cancer treatment.

Current cancer chemotherapy is associated with many side effects and, in some cases, drug resistance, which makes the search for new active molecules and drug delivery strategies imperative. Carbamazepine is an antiepileptic compound that has shown efficacy against breast cancer cell lines. In this study, it was incorporated into layered double hydroxide nanoclays, the percentage of drug loading was increased compared to previous research, and the clays were impregnated with magnetic Fe3O4 nanoparticles. The goal of the magnetic Fe3O4-impregnation was to direct the nanocomposites to the therapeutic target with an external magnetic field. The nanoclay-carbamazepine composites had a carbamazepine loading of 51 %, and the nanoclay-carbamazepine-nanoparticles had a drug loading of 13 % due to the addition of more ingredients. The structure of the composites was analyzed by X-ray diffraction and Scherrer equation, showing a layered double hydroxide organization with crystal sizes of 9-15 nm; from transmission electron microscopy, the final compounds showed a particle size of 97-158 nm, small enough for systemic circulation. In vibrating sample magnetization studies, the composites showed a superparamagnetic behavior with high magnetic saturation (9-17 emu/gr), which should allow a good material attraction by an external magnetic field located near the tumor. In vitro drug release studies were done in Franz cells and measured by UV/Vis spectrophotometry; they showed that carbamazepine release from the nanocomposites responds to the media pH: a good drug release at the lysosome pH and slow release at the blood pH. Finally, the efficacy was tested in vitro in MDA-MB-231 breast cancer cells, and the composites showed an enhanced efficacy in comparison with that produced by the free drug (96 % and 62 % of cell inhibition respectively). Carbamazepine administered with magnetic clays as a carrier is a promising treatment for breast cancer, and further studies should be done to measure the arrival time and the efficacy in vivo.

Synthesis, characterization and in vitro release studies of a new acetazolamide-HP-beta-CD-TEA inclusion complex.

The aim of our work was to develop a multicomponent inclusion complex of acetazolamide (ACZ) in order to investigate the combined effect of hydroxypropyl-beta-cyclodextrin (HP-beta-CD) and triethanolamine (TEA) on the solubility of ACZ and its possibility of ophthalmic delivery. Phase solubility study was used to evaluate the complexation in solution at 25 degrees C. Complex formation was also evaluated by comparing the infrared (FT-IR) spectra of the solid complexes with a simple physical mixture containing the same amount of ACZ. FT-IR experiments provided data indicating that the carbonamido group of ACZ is involved in the inclusion process. In vitro release data showed that both formulations, containing the freeze-dried ternary complex and the corresponding simple physical mixture of ACZ with HP-beta-CD and TEA presented the fastest release rate of ACZ. These results suggest that the ACZ-HP-beta-CD-TEA complex represents an effective novel formulation to enhance ACZ solubility in water, turning it promising for ophthalmic administration.

Possibility of enterohepatic recycling of ketoprofen in dogs.

Ketoprofen is mainly cleared by glucuronidation. The rate of glucuronidation of this compound has been demonstrated to be greater in dog than in human liver microsomes. Dog is the most common secondary nonprimate species used in drug metabolism studies in the pharmaceutical industry. Therefore, this study was undertaken to provide valuable information to pharmaceutical companies using dog as a model species for pharmacokinetic analyses when differences in glucuronidation occur across species for therapeutic drugs known to be extensively glucuronidated. The pharmacokinetics of ketoprofen was investigated after intravenous (0.27, 0.57 and 1.10 mg/kg) and oral administration of ketoprofen ( approximately 10 mg/100 ml) of the racemate in dogs. Serial blood samples were collected at timed intervals for 7 and 24h following intravenous and oral administration of the dose, respectively, and concentrations in plasma were determined by a sensitive and specific HPLC method. By comparing the AUC0-infinity following oral and intravenous administrations, ketoprofen bioavailability was approximately 100%. A possibility of enterohepatic cycling of ketoprofen in dogs was proposed because of multiple peak phenomenon in the concentration-time profiles after intravenous and oral dosing was observed.

Biosynthesized silver nanoparticles: Decoding their mechanism of action in Staphylococcus aureus and Escherichia coli.

The oxidative stress generation in bacteria by the presence of antibiotics (in this case silver nanoparticles (AgNPs)) is already widely known. Previously, we demonstrated that AgNPs generate oxidative stress in Staphylococcus aureus and Escherichia coli mediated by the increase of reactive oxygen species (ROS). In this work we are demonstrating the consequences of the oxidative stress by the presence of AgNPs; these bacterial strains increased the levels of oxidized proteins and lipids. In addition, it was possible to determine which reactive oxygen species are mainly responsible for the oxidative damage to macromolecules. Also, we found that the bacterial DNA was fragmented and the membrane potential was modified. This increase in the levels of ROS found in both, S. aureus and E. coli, was associated with the oxidation of different types of important macromolecules for the normal functioning of cell, so the oxidative stress would be one of the mechanisms by which the AgNPs would exert their toxicity in both strains, one Gram positive and the other Gram negative of great clinical relevance.

Stability of valacyclovir: implications for its oral bioavailability.

The absolute bioavailability of the prodrug valacyclovir, the l-valyl ester of acyclovir, after oral administration is approximately 54.5%. Since premature hydrolysis of this prodrug in the intestinal lumen may be a possible reason for its incomplete bioavailability and the chemical and enzymatic stability of the valacyclovir has been investigated. Release rates were investigated in both phosphate buffers with varying pH as well as in human and dog gastrointestinal fluids. The stability of the prodrug was found to be dependent on pH. This prodrug is chemically stable along the acidic pH side (under 4), while the prodrug degrades in alkaline medium through a base-catalyzed pseudo-first-order kinetics. The degradation of the prodrug valacyclovir progressed faster in intestinal fluid than in phosphate buffer at the same pH. There was no appreciable release of valacyclovir neither in the human and dog stomach contents nor in phosphate buffers at pHs fewer than 4, although its degradation was fastest in the human and dog stomach contents. In light of this result, we can conclude that the degradation of the valacyclovir in the upper intestinal lumen is probably one of the causes of its poor bioavailability.

Intestinal uptake and toxicity evaluation of acetazolamide and its multicomponent complexes with hidroxypropyl-ß-cyclodextrin in rats.

Large oral doses of ACZ lower the intraocular pressure (IOP), but usually lead to a multitude of systemic side effects, including gastrointestinal upset. The present study was undertaken to evaluate the effect of ACZ on the histological structure of rat duodenal mucosa and to assess a possible protective role of the complex formation of ACZ with HP-ß-CD, either separately or in combination with a third compound, on the gut epithelial layer by histological and ultrastructural examinations of sections of rat duodenum exposed to ACZ or its formulations. In addition, the transport process of ACZ and its binary or ternary complexes across the duodenal mucosa by means of the single-pass intestinal perfusion (SPIP) method in rats was evaluated. Evidence was found that ACZ alters intestinal permeability and induces damage to the rat small intestine. In contrast, ACZ-induced intestinal injury may be abrogated by ACZ complexation. In addition, the complexation of ACZ with HP-ß-CD, alone or in combination with a third compound, facilitated significant levels of ACZ uptake across the rat duodenal segment. Ternary complexes of ACZ with HP-ß-CD in combination with TEA (triethanolamine) or calcium ions were found to provide an excellent approach that enabled an increased apparent permeability of ACZ across the duodenal epithelium, with a concomitant ability to preserve the integrity of the gut epithelium from ACZ-induced injury. These results could be useful for the design and development of novel ACZ formulations that can reduce GI toxicity, while still maintaining their essential therapeutic efficacies.

Characterization, dissolution and in vivo evaluation of solid acetazolamide complexes.

The effects of binary and ternary systems of acetazolamide (ACZ) with hydroxypropyl-ß-cyclodextrin (HP-ß-CD) alone or with triethanolamine (TEA) on the crystalline properties, dissolution and intraocular pressure (IOP)-lowering effect were investigated. It was found that the crystal structure of ACZ powder could be modified by the processing conditions. Freeze-drying ACZ powder affected not only the particle morphology but also its polymorphic form and the starting ACZ was converted to pure form A upon freeze-drying treatment. Results provided by DSC/TGA, XRPD, SEM and FT-IR suggested the formation of inclusion complexes between ACZ with HP-ß-CD alone or with TEA, obtained by the freeze-drying method and the conversion of the drug into the amorphous state. Binary and ternary systems of ACZ obtained by freeze-drying exhibited significantly enhanced ACZ dissolution rates. The IOP-lowering effects of ACZ and its complexes with HP-ß-CD alone or with TEA were studied in normotensive rabbits. Whereas the maximum IOP-lowering effect (~4 mmHg, ~33%), obtained with these binary and ternary lyophilized ACZ systems occurred at around 90 min, the ternary system exhibited a longer maximum IOP-lowering effect peak compared with that of the binary system. These results are in line with those obtained from the dissolution studies, where the ternary system exhibited longer dissolution times compared to the lyophilized binary one. Results obtained from the dissolution studies, also showed that freeze-drying the native crystalline form of ACZ significantly increased the dissolution rate of ACZ, thus improving the IOP-lowering effect of this drug.

Accessibility as a conditioning factor in treatment for exudative age-related macular degeneration.

PURPOSE: Ranibizumab and bevacizumab coexist as the main therapeutic strategies for the treatment of neovascular age-related macular degeneration (NV-AMD). In Argentina, the access pathways to the drugs are different. Patients with different pathways and gatekeepers to access may experience different outcomes. The purpose of this work was to estimate the impact on therapeutic effects and visual outcome of the different accessibilities to NV-AMD treatment. 
 METHODS: A retrospective analysis of the charts of 78 patients with previously untreated exudative AMD, who were treated with ranibizumab or bevacizumab between January 2009 and December 2011, was conducted. The main outcomes measured included time delay and change in mean best-corrected visual acuity (BCVA) between diagnosis and treatment and mean BCVA change at 1-year follow-ups.
 RESULTS: The delay between diagnosis and treatment and decrease in visual acuity over this time was significantly higher for patients treated with ranibizumab. At 1 year after the initiation of treatment, BCVA had a mean increase from baseline of 0.11 letters in the bevacizumab group with a mean of 4.71 injections, compared with a decrease of 8.87 letters with a mean of 2.98 injections in the ranibizumab group.
 CONCLUSIONS: Access to treatment can be a key factor for success of therapy. Waiting times and availability of doses are crucial in the treatment of NV-AMD. Solving the problems related to delayed initiation of therapy and the difficulties in the maintenance phase are more important than define whether bevacizumab or ranibizumab is used.

Characterization of the hydrochlorothiazide: ß-cyclodextrin inclusion complex. Experimental and theoretical methods.

Hydrochlorothiazide (HCT) is one of the most commonly prescribed antihypertensive drugs. In an attempt to gain an insight into the physicochemical and molecular aspects controlling the complex architecture of native ß-cyclodextrin (ß-CD) with HCT, we performed multiple-temperature-pH isothermal titration calorimetric measurements of the HCT:ß-CD system, together with proton nuclear magnetic resonance spectroscopy ((1)H NMR), phase solubility analysis, and molecular modeling methods. The A(L)-type diagrams, obtained at different pH values and temperatures, suggested the formation of soluble 1:1 inclusion complexes of ß-CD with HCT. The corresponding stability constants (K(1:1)) were determined by phase solubility studies and compared with those obtained by ITC, with good agreement between these two techniques being found. The three-dimensional array of the complex was studied by (1)H NMR and molecular modeling methods. Both techniques confirmed the formation of the inclusion complex, with good agreement between the experimental and theoretical techniques regarding the HCT binding mode to ß-CD. Also, the forces involved in the association process were determined, both from the thermodynamic parameters obtained by ITC (association enthalpy, binding constant, Gibbs free energy, and entropy) and from energetic decomposition analyses derived from computational methods. We concluded that the formation of the HCT:ß-CD complex was enthalpy driven, with the inclusion mode of HCT being highly dependent on its ionization state. In all cases, sustained hydrogen bond interactions with hydroxyl groups of ß-CD were identified, with the solvation energy limiting the affinity. Regarding the pH and temperature dependence, lower affinity constants were found at higher HCT ionization states and temperatures.

Complex formation of chlorhexidine gluconate with hydroxypropyl-ß-cyclodextrin (HPßCD) by proton nuclear magnetic resonance spectroscopy ((1)H NMR).

The complex formation of chlorhexidine digluconate (CHX-G(2)) with hydroxypropyl-ß-cyclodextrin (HPßCD) was studied using NMR spectroscopy. The results revealed that this surfactant agent shows an monomer/aggregate equilibrium, which is dependent on the concentration of this drug. This equilibrium can be modified by the presence of HPßCD, which reduces the aggregation of the CHX-G(2) molecules. An inclusion process of the CHX-G(2) aromatic residue within the cyclodextrin cavity was confirmed by 2D ROESY spectroscopy. (1)H NMR titration studies of CHX-G(2) with HPßCD in D(2)O confirmed the formation of higher order complexes between CHX-G(2) and HPßCD. Moreover, the addition of HPßCD into CHX-G(2) solutions forms insoluble aggregates. Such insoluble aggregates may result in the stacking of CHX-G(2) molecules on the surface of the CHX-G(2):HPßCD complexes.

Promising complexes of acetazolamide for topical ocular administration.

IMPORTANCE OF THE FIELD: Acetazolamide (ACZ), a carbonic anhydrase inhibitor (CAI), and other oral CAIs have been an integral part of antiglaucoma therapy for > 40 years. ACZ is used orally for the reduction of intraocular pressure in patients suffering from glaucoma. However, this treatment leads to unpleasant systemic side effects. The answer to the undesirable effects of ACZ is the topical delivery of this drug into the eye, where it could elicit its physiological action. However, the development of a topical formulation of ACZ is limited by its poor ocular bioavailability, which can be largely attributed to its poor penetration coefficient and poor biphasic solubility. AREAS COVERED IN THIS REVIEW: This review offers an overview of different approaches to delivering ACZ to the eye, highlighting the potential of the ternary system ACZ:HP-beta-CD:TEA as a tool for formulating aqueous ACZ eye drop solutions. WHAT THE READER WILL GAIN: A critical analysis is provided to highlight the key issues to design formulations containing hydrophilic cyclodextrins. TAKE HOME MESSAGE: The ACZ:HP-beta-CD:TEA complex is an important new approach to improve the ocular bioavailability of this drug. This approach may be applied to other CAIs in the future.

Synthesis and characterization of binary and ternary complexes of diclofenac with a methyl-beta-CD and monoethanolamine and in vitro transdermal evaluation.

Here, we describe the chemical characterization of the inclusion complex between diclofenac (DCF) and methyl-beta-cyclodextrin (M-beta-CD) in the presence or absence of monoethanolamine (MEA). Several techniques were used to analyze the complex both in solution and in the solid state. Solubility of DCF was increased by the addition of M-beta-CD. However, the DCF solubility increase was more significant by the addition of M-beta-CD in the presence of MEA. In vitro permeation experiments through excised human skin revealed that DCF was enhanced by M-beta-CD. Nevertheless, further improvement in the flux and the permeability coefficient of DCF was obtained by the ternary system DCF-M-beta-CD-MEA.

An efficient ternary complex of acetazolamide with HP-ss-CD and TEA for topical ocular administration.

In order to enhance the ocular bioavailability of acetazolamide (ACZ), a multicomponent complex with hydroxypropyl-ss-cyclodextrin (HP-ss-CD) and triethanolamine (TEA) was prepared to be applied topically. In vitro corneal permeation across isolated rabbit cornea of proposed ACZ formulations and the marketed AZOPT(R) formulation (1% w/v brinzolamide) was studied. Formulations were also tested for their effect on the intraocular pressure (IOP) in rabbits. (1)H- and (13)C-NMR experiments were undertaken to verify the real inclusion of ACZ in the ACZ-HP-ss-CD-TEA multicomponent complex. The binding of ACZ to HP-ss-CD in the presence of TEA is described. The increase of TEA concentration decreases the apparent equilibrium constant for the ACZ-HP-ss-CD complex. The ternary system ACZ-HP-ss-CD-TEA seemed to be able to reduce IOP in about 30%. This effect was sustained for 4 h after instillation. In vitro corneal permeation studies demonstrated that the ACZ permeation was increased. RMN experiments indicated that TEA can weaken the association between ACZ and HP-ss-CD increasing the drug ocular hypotensive effect by increasing the free drug available for absorption. Our formulations were considered practically non-irritant. These results indicate that the ternary system ACZ-HP-ss-CD-TEA might be a useful tool for formulating aqueous ACZ eye drop solutions.

Dissolution and solubility behavior of fenofibrate in sodium lauryl sulfate solutions.

The solubility of fenofibrate in pH 6.8 McIlvaine buffers containing varying concentrations of sodium lauryl sulfate was determined. The dissolution behavior of fenofibrate was also examined in the same solutions with rotating disk experiments. It was observed that the enhancement in intrinsic dissolution rate was approximately 500-fold and the enhancement in solubility was approximately 2000-fold in a pH 6.8 buffer containing 2% (w/v) sodium lauryl sulfate compared to that in buffer alone. The micellar solubilization equilibrium coefficient (k*) was estimated from the solubility data and found to be 30884+/-213 L/mol. The diffusivity for the free solute, 7.15x10(-6) cm2/s, was calculated using Schroeder's additive molal volume estimates and Hayduk-Laurie correlation. The diffusivity of the drug-loaded micelle, estimated from the experimental solubility and dissolution data and the calculated value for free solute diffusivity, was 0.86x10(-6) cm2/s. Thus, the much lower enhancement in dissolution of fenofibrate compared to its enhancement in solubility in surfactant solutions appears to be consistent with the contribution to the total transport due to enhanced micellar solubilization as well as a large decrease (approximately 8-fold) in the diffusivity of the drug-loaded micelle.