An In Silico Analysis of Genetic Variants and Structural Modeling of the Human Frataxin Protein in Friedreich's Ataxia.

Friedreich's Ataxia (FRDA) stands out as the most prevalent form of hereditary ataxias, marked by progressive movement ataxia, loss of vibratory sensitivity, and skeletal deformities, severely affecting daily functioning. To date, the only medication available for treating FRDA is Omaveloxolone (Skyclarys®), recently approved by the FDA. Missense mutations within the human frataxin (FXN) gene, responsible for intracellular iron homeostasis regulation, are linked to FRDA development. These mutations induce FXN dysfunction, fostering mitochondrial iron accumulation and heightened oxidative stress, ultimately triggering neuronal cell death pathways. This study amalgamated 226 FXN genetic variants from the literature and database searches, with only 18 previously characterized. Predictive analyses revealed a notable prevalence of detrimental and destabilizing predictions for FXN mutations, predominantly impacting conserved residues crucial for protein function. Additionally, an accurate, comprehensive three-dimensional model of human FXN was constructed, serving as the basis for generating genetic variants I154F and W155R. These variants, selected for their severe clinical implications, underwent molecular dynamics (MD) simulations, unveiling flexibility and essential dynamic alterations in their N-terminal segments, encompassing FXN42, FXN56, and FXN78 domains pivotal for protein maturation. Thus, our findings indicate potential interaction profile disturbances in the FXN42, FXN56, and FXN78 domains induced by I154F and W155R mutations, aligning with the existing literature.

Physiologically based pharmacokinetic modelling of semaglutide in children and adolescents with healthy and obese body weights.

AIMS: To develop paediatric physiologically based pharmacokinetic modelling (PBPK) models of semaglutide to estimate the pharmacokinetic profile for subcutaneous injections in children and adolescents with healthy and obese body weights. METHODS: Pharmacokinetic modelling and simulations of semaglutide subcutaneous injections were performed using the Transdermal Compartmental Absorption & Transit model implemented in GastroPlus v.9.5 modules. A PBPK model of semaglutide was developed and verified in the adult population, by comparing the simulated plasma exposure with the observed data, and further scaled to the paediatric populations with normal and obese body weight. RESULTS: The semaglutide PBPK model was successfully developed in adults and scaled to the paediatric population. Our paediatric PBPK simulations indicated a significant increase in maximum plasma concentrations for the 10-14 years' paediatric population with healthy body weights, which was higher than the observed values in adults at the reference dose. Since gastrointestinal adverse events are related to increased semaglutide concentrations, peak concentrations outside the target range may represent a safety risk for this paediatric age group. Besides, paediatric PBPK models indicated that body weight was inversely related to semaglutide maximum plasma concentration, corroborating the consensus on the influence of body weight on semaglutide PK in adults. CONCLUSION: Paediatric PBPK was successfully achieved using a top-down approach and drug-related parameters. The development of unprecedented PBPK models will support paediatric clinical therapy for applying aid-safe dosing regimens for the paediatric population in diabetes treatment.

Linear and areal surface roughness assessments for the study of tooth wear in human enamel.

OBJECTIVES: To test 8 models of linear surface roughness assessment in characterizing surface profile description and to correlate these models with equivalent areal parameters over sound human enamel in vitro. MATERIALS AND METHODS: Thirty enamel blocks were randomly selected. The roughness data (2D-Rp; Rv; Rz; Rc; Rt; Ra; Rq; Rsk; Rku/3D-Sp; Sv; Sz; Sa; Sq; Ssk; Sku) was obtained in duplicate in a non-contact 3D optical profilometer. The models were composed by 1 single vertical trace (model 1) until 8 traces (model 8 composed by three vertical traces, three horizontal traces, and two diagonal). RESULTS: The addition of linear sampling traces to the enamel blocks did not result in Rp, Rv, Rz, Rc, Rt, Ra, Rq, Rsk, and Rku value changes (low power-from 5 to 72%). Significant Spearman's correlation coefficients were obtained in most correlation analysis (Rp ↔ Sp; Rv ↔ Sv; Rz ↔ Sz; Ra ↔ Sa; Rq ↔ Sq; and Rku ↔ Sku). CONCLUSIONS: A single vertical trace in the middle of the sample was representative of the overall enamel surface roughness (Rp, Rv, Rz, Rc, Rt, Ra, Rq, Rsk, and Rku) models. The majority of the assessed models in the correlation evaluation presented significant and positive association. CLINICAL RELEVANCE: The findings highlight the applicable model for roughness analysis over human enamel recommended for research and in situ trials assessments.

Development of SEDDS formulation containing caffeine for dermal delivery.

OBJECTIVE: The objective of this work was to develop a self-emulsifying drug delivery system (SEDDS) containing caffeine for the treatment of cellulite. METHODS: SEDDS were prepared using the solution method. 0.5% (w/v) caffeine was added to the previously selected excipients. The system was characterized by droplet size, zeta potential, emulsification time and long-term stability. In vitro release and skin permeation were investigated using Franz-type diffusion cells. The cytotoxicity was evaluated on normal human keratinocytes. RESULTS: Caffeine SEDDS were thermodynamically stable, with a zeta potential less than - 22 mV and droplet size around 30 nm, and were long-term stable. The permeation study showed that the formulation promoted caffeine accumulation in the skin layers, suggesting an increase in local circulation. Cytotoxicity studies on HaCaT cells were not conclusive as the surfactant used indicated false-positive results due to its high molar mass. CONCLUSION: It was possible to obtain a stable SEDDS that could cause an increase in blood flow in the applied area, resulting in cellulite reduction.

OBJECTIF: L'objectif de ce travail était de développer un système d'administration de médicaments auto-émulsifiants (SEDDS) contenant de la caféine pour le traitement de la cellulite. MÉTHODES: Les SEDDS ont été préparés par la méthode en solution. 0,5 % (p/v) de caféine a été ajouté aux excipients préalablement sélectionnés. Le système a été caractérisé par la taille des gouttelettes, le potentiel zêta, le temps d'émulsification et la stabilité à long terme. La libération in vitro et la perméation cutanée ont été étudiées dans des cellules de diffusion de type Franz. La cytotoxicité était évaluée sur des kératinocytes humains normaux. RÉSULTATS: Les SEDDS de caféine étaient thermodynamiquement stables, avec un potentiel Zeta inférieur à -22 mV et une taille de gouttelettes d'environ 30 nm, et stables à long terme. L'étude de perméation a montré que les formulations favorisent l'accumulation de caféine dans les couches de la peau, suggérant une augmentation de la circulation locale. Les études de cytotoxicité sur les cellules HaCaT n'ont pas été concluantes car le surfactant utilisé indique des résultats faussement positifs dus à une masse molaire élevée. CONCLUSION: Il a été possible d'obtenir un SEDDS stable qui peut provoquer une augmentation du flux sanguin dans la zone appliquée, entraînant une réduction de la cellulite.

Antimycobacterial and anti-inflammatory activities of thiourea derivatives focusing on treatment approaches for severe pulmonary tuberculosis.

Tuberculosis (TB) remains a serious public health problem and one of the main concern is the emergence of multidrug-resistant and extensively resistant TB. Hyper-reactive patients develop inflammatory necrotic lung lesions that aggravate the pathology and facilitate transmission of mycobacteria. Treatment of severe TB is a major clinical challenge that has few effective solutions and patients face a poor prognosis, years of treatment and different adverse drug reactions. In this work, fifteen novel and thirty-one unusual thiourea derivatives were synthesized and evaluated in vitro for their antimycobacterial and anti-inflammatory potential and, in silico for ADMET parameters and for structure-activity relationship (SAR). Thioureas derivatives 10, 15, 16, 28 and 29 that had shown low cytotoxicity and high activities were selected for further investigation, after SAR study. These five thioureas derivatives inhibited Mtb H37Rv growth in bacterial culture and in infected macrophages, highlighting thiourea derivative 28 (MIC50 2.0 ± 1.1 and 2.3 ± 1.1 µM, respectively). Moreover, these compounds were active against the hypervirulent clinical Mtb strain M299, in bacterial culture, especially 16, 28 and 29, and in extracellular clumps, highlighting 29, with MIC50 5.6 ± 1.2 µM. Regarding inflammation, they inhibited NO through the suppression of iNOS expression, and also inhibited the production of TNF-α and IL-1ß. In silico studies were carried out suggesting that these five compounds could be administered by oral route and have low toxicological effects when compared to rifampicin. In conclusion, our data show that, at least, thiourea derivatives 16, 28 and 29 are promising antimycobacterial and anti-inflammatory agents, and candidates for further prospective studies aiming new anti-TB drugs, that can be used on a dual approach for the treatment of severe TB cases associated with exacerbated inflammation.

Novel rivaroxaban-loaded poly(lactic-co-glycolic acid)/poloxamer nanoparticles: preparation, physicochemical characterization, in vitro evaluation of time-dependent anticoagulant activity and toxicological profile.

Rivaroxaban (RXB), an oral direct factor Xa inhibitor, presents innovative therapeutic profile. However, RXB has shown adverse effects, mainly due to pharmacokinetic limitations, highlighting the importance of developing more effective formulations. Therefore, this work aims at the preparation, physicochemical characterization and in vitro evaluation of time-dependent anticoagulant activity and toxicology profile of RXB-loaded poly(lactic-co-glycolic acid) (PLGA)/poloxamer nanoparticles (RXBNps). RXBNp were produced by nanoprecipitation method and physicochemical characteristics were evaluated. In vitro analysis of time-dependent anticoagulant activity was performed by prothrombin time test and toxicological profile was assessed by hemolysis and MTT reduction assays. The developed RXBNp present spherical morphology with average diameter of 205.5 ± 16.95 nm (PdI 0.096 ± 0.04), negative zeta potential (-26.28 ± 0.77 mV), entrapment efficiency of 91.35 ± 2.40%, yield of 41.81 ± 1.68% and 3.72 ± 0.07% of drug loading. Drug release was characterized by an initial fast release followed by a sustained release with 28.34 ± 2.82% of RXB available in 72 h. RXBNp showed an expressive time-dependent anticoagulant activity in human and rat blood plasma and non-toxic profile. Based on the results presented, it is possible to consider that RXBNp may be able to assist in the development of promising new therapies for treatment of thrombotic disorders.

In silico bioavailability for BCS class II efavirenz tablets using biorelevant dissolution media for IVIVR and simulation of formulation changes.

OBJECTIVE: This work aims to evaluate the ability of biorelevant dissolution media to simulate the bioavailability of efavirenz tablets, establish an in vitro-in vivo relationship (IVIVR) based on in vivo data using GastroPlus® and simulate formulation changes using DDDPlus™. METHODS: Solubility and drug release profiles were conducted in SLS 0.5% and biorelevant media, such as FaSSIF, FeSSIF, FaSSIF-V2, and FeSSIF-V2. The efavirenz physicochemical properties were used to simulate the plasma concentration profile and compare the simulated pharmacokinetic parameters in fasted and fed states. An IVIVR was developed using Loo-Riegelman as the deconvolution method to estimate drug bioavailability. DDDPlus™ was used to perform virtual trials of formulations to evaluate whether formulations changes and the efavirenz particle size could influence the bioavailability. RESULTS: The drug dissolution displayed higher levels in the biorelevant media that simulated gut-fed state (FeSSIF and FeSSIF-V2). The absorption model successfully predicted the efavirenz pharmacokinetics, and FeSSIF-V2 was chosen as the predictive dissolution media, while an IVIVR was established using the Loo-Riegelman deconvolution method. CONCLUSIONS: The present work provides valuable information about efavirenz solubility and kinetics in the gastrointestinal tract, allowing an IVIVR to support future formulation changes. This understanding is essential for rational science-driven formulation development. At least, this study also showed the validity and applicability of in vitro and in silico tools in the regulatory scenario helping on drug development.

Vancomycin-loaded nanoparticles against vancomycin intermediate and methicillin resistant Staphylococcus aureus strains.

Bacterial infections represent one of the leading causes of mortality in the world. Among causative pathogens, S. aureus is prominently known as the underlying cause of many multidrug resistant infections that are often treated with the first-line choice antibiotic vancomycin (VCM). Loading antibiotics into polymeric nanoparticles (Np) displays promise as an alternative method to deliver therapy due to the greater access and accumulation of the antibiotic at the site of the infection as well as reducing toxicity, irritation and degradation. The aim of this work was to prepare, characterize and evaluate VCM-loaded nanoparticles (VNp) for use against S. aureus strains. Moreover, conjugation of Nps with holo-transferrin (h-Tf) was investigated as an approach for improving targeted drug delivery. VNp were prepared by double emulsion solvent evaporation method using PLGA and PVA or DMAB as surfactants. The particles were characterized for size distribution, Zeta Potential, morphology by transmission electron microscopy, encapsulation yield and protein conjugation efficiency. Process yield and drug loading were also investigated along with an in vitro evaluation of VNp antimicrobial effects against S. aureus strains. Results showed that Np were spontaneously formed with a mean diameter lower than 300 nm in a narrow size distribution that presented a spherical shape. The bioconjugation with h-Tf did not appear to increase the antimicrobial effect of VNp. However, non-bioconjugated Np presented a minimal inhibitory concentration lower than free VCM against a MRSA (Methicillin-resistant S. aureus) strain, and slightly higher against a VISA (VCM intermediate S. aureus) strain. VNp without h-Tf showed potential to assist in the development of new therapies against S. aureus infections.

Effect of xylitol tablets with and without red propolis on salivary parameters, dental biofilm and sensory acceptability of adolescents: a randomized crossover clinical trial.

The effect of xylitol tablets with (XPT) and without (XT) red propolis on salivary parameters, dental biofilm and acceptability of adolescents was evaluated through a blinded randomized crossover clinical trial. Healthy volunteers were allocated in the XPT and XT groups with a 30-day washout period for consumption of two tablets/dayfor seven days. An increase in salivary parameters was only observed immediately after tablet consumption, without differences between XPT and XT. The results for total microorganisms were similar, but XT was better in controlling Streptococcus spp. Rothia dentocariosa and Streptococcus salivarius were the most frequent in the biofilm and saliva, respectively. XPT and XT showed similar acceptability, with the highest purchase intention for XT. Although propolis did not enhance the properties of XT, further studies testing different protocols and follow-up are necessary; XT controlled Streptococcus spp. in biofilms, which demonstrate its potential for clinical application.

Forced degradation studies of norepinephrine and epinephrine from dental anesthetics: Development of stability-indicating HPLC method and in silico toxicity evaluation.

Injectable solutions containing epinephrine (EPI) and norepinephrine (NE) are not stable, and their degradation is favored mainly by the oxidation of catechol moiety. As studies of these drugs under forced degradation conditions are scarce, herein, we report the identification of their degradation products (DP) in anesthetic formulations by the development of stability-indicating HPLC method. Finally, the risk assessment of the major degradation products was evaluated using in silico toxicity approach. HPLC method was developed to obtain a higher selectivity allowing adequate elution for both drugs and their DPs. The optimized conditions were developed using a C18 HPLC column, sodium 1-octanesulfonate, and methanol (80:20, v/v) as mobile phase, with a flow rate of 1.5 mL/min, UV detection at 199 nm. The analysis of standard solutions with these modifications resulted in greater retention time for EPI and NE, which allow the separation of these drugs from their respective DPs. Then, five DPs were identified and analyzed by in silico studies. Most of the DPs showed important alerts as hepatotoxicity and mutagenicity. To the best of our acknowledgment, this is the first report of a stability-indicating HPLC method that can be used with formulations containing catecholamines.

In Vitro-In Vivo Correlation for Desvenlafaxine Succinate Monohydrate Extended Release Tablets.

The objective of this study was to develop a dissolution test in order to establish an in vitro-in vivo correlation (IVIVC) model for desvenlafaxine succinate monohydrate (DVSM) extended release (ER) tablets. The in vitro release characteristics of the drug were determined using USP apparatus 1 at 75 rpm, with volume of HCl pH 1.2, acetate buffer solution (ABS) pH 4.5, or phosphate buffer solution (PBS) pH 6.8. In vivo plasma concentrations and pharmacokinetic parameters in healthy volunteers were obtained from a bioequivalence study. The similarity factors f1 and f2 were used to compare the dissolution data. The IVIVC model was developed using fraction dissolved and fraction absorbed of the reference product. For predictability, the results showed that the percentage prediction error (%PE) value of Cmax was 7.63%. The observed low prediction error for Cmax demonstrated that the IVIVC model was valid for this parameter.

Sensitive detection of topiramate degradation products by high-performance liquid chromatography/electrospray ionization mass spectrometry using ion-pairing reagents and polarity switching.

RATIONALE: The chromatographic analysis of topiramate and its degradation products is challenging due to the absence of chromophoric moieties in their structures, the wide polarity range of the compounds and their ionization differences. This work proposes two new mass spectrometry approaches for evaluating these analytes. METHODS: Based on the calculated experimental limit of detection (LOD), a highly sensitive high-performance liquid chromatography (HPLC) paired-ion electrospray ionization mass spectrometry (PIESI-MS) method was developed for the determination of topiramate inorganic degradation products. The influence of different solvent systems on the LODs for topiramate and its main degradation products was determined in both positive/negative ionization modes. In addition, a HPLC method to analyze both organic and inorganic degradation products was proposed by mass spectrometry with positive/negative ion switching electrospray ionization. RESULTS: A sensitive HPLC/PIESI-MS method was achieved for the efficient separation of topiramate inorganic degradation products. Both sulfate and sulfamate were detected in the positive selected ion monitoring (SIM) mode with an increased sensitivity compared with the negative SIM mode. The HPLC/ESI-MS analysis with positive/negative ion switching allowed the simultaneous separation and detection of the major degradation products of topiramate in a 10-min run using a single column and a single detector. CONCLUSIONS: Two new alternative MS approaches for analyzing the main degradation products of topiramate were developed. The proposed methods are considered advantageous over the existing methods and can be applied to quality control studies of topiramate.

Oral pentamidine-loaded poly(d,l-lactic-co-glycolic) acid nanoparticles: an alternative approach for leishmaniasis treatment.

Leishmaniasis is a group of diseases caused by a protozoa parasite from one of over 20 Leishmania species. Depending on the tissues infected, these diseases are classified as cutaneous, mucocutaneous and visceral leishmaniasis. For the treatment of leishmaniasis refractory to antimony-based drugs, pentamidine (PTM) is a molecule of great interest. However, PTM displays poor bioavailability through oral routes due to its two strongly basic amidine moieties, which restricts its administration by a parenteral route and limits its clinical use. Among various approaches, nanotechnology-based drug delivery systems (nano-DDS) have potential to overcome the challenges associated with PTM oral administration. Here, we present the development of PTM-loaded PLGA nanoparticles (NPs) with a focus on the characterization of their physicochemical properties and potential application as an oral treatment of leishmaniasis. NPs were prepared by a double emulsion methodology. The physicochemical properties were characterized through the mean particle size, polydispersity index (PdI), zeta potential, entrapment efficiency, yield process, drug loading, morphology, in vitro drug release and in vivo pharmacological activity. The PTM-loaded PLGA NPs presented with a size of 263 ± 5 nm (PdI = 0.17 ± 0.02), an almost neutral charge (-3.2 ± 0.8 mV) and an efficiency for PTM entrapment of 91.5%. The release profile, based on PTM dissolution, could be best described by a zero-order model, followed by a drug diffusion profile that fit to the Higuchi model. In addition, in vivo assay showed the efficacy of orally given PTM-loaded PLGA NPs (0.4 mg kg-1) in infected BALB/c mice, with significant reduction of organ weight and parasite load in spleen (p-value < 0.05). This work successfully reported the oral use of PTM-loaded NPs, with a high potential for the treatment of visceral leishmaniasis, opening a new perspective to utilization of this drug in clinical practice.

Development of USP Apparatus 3 Dissolution Method with IVIVC for Extended Release Tablets of Metformin Hydrochloride and Development of a Generic Formulation.

Metformin is a euglycemic drug for the treatment of type 2 diabetes mellitus. To date, there are 13 dissolution methodologies described in the U.S. Pharmacopoeia (USP) to evaluate the release profile of metformin from extended-release tablets utilizing either a USP apparatus 1 (basket) or 2 (paddle). In the absence of a protocol for a USP apparatus 3 (reciprocating cylinder), the goal of this work was to develop an in vitro dissolution method for metformin extended-release tablets based on an in vivo-in vitro correlation (IVIVC). Following a systematic evaluation, a final dissolution method, M4, was defined. It applied 30 dips per minute (dpm) over a total period of 10 h into a series of solutions that included 2 h in HCl media (pH 1.2), 1 h in an acetate buffer solution (pH 4.5), 1 h in phosphate buffer solution (PBS) (pH 5.8) and 6 h in PBS (pH 6.8). This method showed a significant IVIVC with a calculated R2 > 0.98 (point-to-point correlation, Level A) and it was successfully used as a tool to assist in the development of generic extended release formulations for metformin consisting of a lipophilic matrix system.

In Situ Cocrystallization of Dapsone and Caffeine during Fluidized Bed Granulation Processing.

Different pharmaceutical manufacturing processes have been demonstrated to represent feasible platforms for the production of pharmaceutical cocrystals. However, new methods are needed for the manufacture of cocrystals on a large scale. In this work, the suitability of the use of a fluidized bed system for granulation and concomitant cocrystallization was investigated. Dapsone (DAP) and caffeine (CAF) have been shown to form a stable cocrystal by simple solvent evaporation. DAP is the active pharmaceutical ingredient (API) and CAF is the coformer. In the present study, DAP-CAF cocrystals were produced through liquid-assisted milling and the product obtained was used as a cocrystal reference. The granulation of DAP and CAF was carried out using four different experimental conditions. The solid-state properties of the constituents of the granules were characterised by differential scanning calorimetry (DSC) and x-ray powder diffraction (PXRD) analysis while the granule size distribution and morphology were investigated using laser diffraction and scanning electron microscopy (SEM), respectively. DAP-CAF cocrystal granules were successfully produced during fluidized bed granulation. The formation of cocrystals was possible only when the DAP and CAF were dissolved in the liquid phase and sprayed over the fluidized solid particles. Furthermore, the presence of polymers in solution interferes with the cocrystallization, resulting in the amorphization of the DAP and CAF. Cocrystallization via fluidized bed granulation represents a useful tool and a feasible alternative technique for the large scale manufacture of pharmaceutical cocrystals for solid dosage forms.

A promising oral fucoidan-based antithrombotic nanosystem: Development, activity and safety.

Fucoidan-loaded nanoparticles emerge as great candidates to oral anticoagulant therapy, due to increasing of bioavailability and circulation time of this natural anticoagulant. Crosslink between chitosan chains are performed using glutaraldehyde to confer higher gastric pH resistance to nanoparticle matrices. In this work, chitosan-fucoidan nanoparticles, without (NpCF) and with glutaraldehyde crosslink (NpCF 1% and NpCF 2%), were prepared to evaluate their anticoagulant, antithrombotic and hemorrhagic profile. Nanoparticles were characterized by average diameter, polydispersity index, zeta potential, Fourier transform infrared spectroscopy and fucoidan in vitro release. Anticoagulant and antithrombotic activities were determined by in vitro and in vivo models, respectively. Hemorrhagic profile was in vivo evaluated by tail bleeding assay. Preparations showed nanometric and homogeneous average diameters. Zeta potentials of NpCF and NpCF 1% were stable over gastrointestinal pH range, which was confirmed by low fucoidan release in gastric and enteric media. In pH 7.4, NpCF and NpCF 1% demonstrated fucoidan release of 65.5% and 60.6%, respectively, within the first 24 hours. In comparison to fucoidan, NpCF and NpCF 1% showed increased in vitro anticoagulant activity. A significant difference on oral antithrombotic profile of NpCF 1% was found in comparison to fucoidan. Bleeding profile of NpCF and NpCF 1% showed no differences to control group, indicating the safety of these systems. Surprisingly, oral antithrombotic profile of commercially available fucoidan, from Fucus vesiculosus, has not been previously determined, which reveals new possibilities. In this work, significant advances were observed in anticoagulant and antithrombotic profiles of fucoidan through the preparation of NpCF 1%.

Development and validation of a stability-indicating HPLC method for topiramate using a mixed-mode column and charged aerosol detector.

The analysis of topiramate in the presence of its main degradation products is challenging due to the absence of chromophore moieties and their wide range of polarity. Mixed-mode chromatography has been used in such cases because it combines two or more modes of separation. Charged aerosol detector is also an alternative since its detection is independent of optical properties and analyte ionization. This study is aimed to develop and validate two new stability-indicating methods by high-performance liquid chromatography for the main degradation products of topiramate using mixed-mode chromatography and a charged aerosol detector. Method 1 employed an Acclaim Trinity P1® column (3.0 mm × 150 mm, 2.7 µm) with a mobile phase comprising of 80% ammonium acetate buffer (20 mM, pH 4.0) and 20% methanol at a flow rate of 0.5 mL/min at 35°C. Method 2 utilized a C18 Acclaim 120® column (4.6 mm × 250 mm; 5 µm) with ACN/water (50:50) at a flow rate of 0.6 mL/min at 50°C. Validation of the two methods demonstrated excellent performance with respect to linearity, precision, accuracy, and selectivity. The limits of detection for topiramate, fructose, sulfate, sulfamate, and compound A were 2.97, 12.08, 4.02, 13.91, and 3.94 µg/mL, respectively.

Development of a Dissolution Method for Gliclazide Modified-Release Tablets Using USP Apparatus 3 with in Vitro-in Vivo Correlation.

Gliclazide (GLZ) is a second generation hypoglycemic drug used for the treatment of Type 2 diabetes mellitus. The low solubility of GLZ has been described as the rate limiting step for drug dissolution and absorption, thus a prediction of its in vivo behavior based on a discriminative dissolution test should lead to a relevant in vitro-in vivo correlation (IVIVC). The aim of this study was to develop a dissolution method for GLZ modified-release (MR) tablets using an United States Pharmacopeia (USP) apparatus 3 through its evaluation by an IVIVC analysis. Various dissolution parameters were evaluated to establish an in vitro method for GLZ tablets. The final dissolution conditions, referred to as method 3, utilized a 400 µm mesh and 30 dips per minute over a total period of 10 h that included 1h in HCl media (pH 1.2), 2h in acetate buffer solution (pH 4.5), 1 h in phosphate buffer solution (PBS; pH 5.8), 5h in PBS (pH 6.8) and finally 1h in PBS (pH 7.2). The calculated point-to-point IVIVC (R2=0.9970) was significantly greater than other methods. The robustness of method 3 suggests it could be applied to pharmaceutical equivalence studies and for quality control analyses of GLZ.

Development and characterization of poly(lactic-co-glycolic) acid nanoparticles loaded with copaiba oleoresin.

Copaiba oleoresin (CPO), obtained from Copaifera landgroffii, is described as active to a large number of diseases and more recently in the endometriosis treatment. In this work, poly(lactic-co-glycolic acid) (PLGA) nanoparticles containing CPO were obtained using the design of experiments (DOE) as a tool to optimize the production process. The nanoparticles optimized by means of DOE presented an activity in relation to the cellular viability of endometrial cells. The DOE showed that higher amounts of CPO combined with higher surfactant concentrations resulted in better encapsulation efficiency and size distribution along with good stability after freeze drying. The encapsulation efficiency was over 80% for all produced nanoparticles, which also presented sizes below 300 nm and spherical shape. A decrease in viability of endometrial stromal cells from ectopic endometrium of patients with endometriosis and from eutopic endometriotic lesions was demonstrated after 48 h of incubation with the CPO nanoparticles. The nanoparticles without CPO were not able to alter the cell viability of the same cells, indicating that this material was not cytotoxic to the tested cells and suggesting that the effect was specific to CPO. The results indicate that the use of CPO nanoparticles may represent a promising alternative for the treatment of endometriosis.

Development and Characterization of Dapsone Cocrystal Prepared by Scalable Production Methods.

In this study, the formation of caffeine/dapsone (CAF/DAP) cocrystals by scalable production methods, such as liquid-assisted grinding (LAG) and spray drying, was investigated in the context of the potential use of processed cocrystal powder for pulmonary delivery. A CAF/DAP cocrystal (1:1 M ratio) was successfully prepared by slow evaporation from both acetone and ethyl acetate. Acetone, ethyl acetate, and ethanol were all successfully used to prepare cocrystals by LAG and spray drying. The powders obtained were characterized by X-ray diffractometry (XRD), differential scanning calorimetry (DSC), thermogravimetry (TGA), and Fourier transform infrared spectroscopy (FTIR). Laser diffraction analysis indicated a median particle size (D50) for spray-dried powders prepared from acetone, ethanol, and ethyl acetate of 5.4 ± 0.7, 5.2 ± 0.1, and 5.1 ± 0.0 µm respectively, which are appropriate sizes for pulmonary delivery by means of a dry powder inhaler. The solubility of the CAF/DAP cocrystal in phosphate buffer pH 7.4, prepared by spray drying using acetone, was 506.5 ± 31.5 µg/mL, while pure crystalline DAP had a measured solubility of 217.1 ± 7.8 µg/mL. In vitro cytotoxicity studies using Calu-3 cells indicated that the cocrystals were not toxic at concentrations of 0.1 and of 1 mM of DAP, while an in vitro permeability study suggested caffeine may contribute to the permeation of DAP by hindering the efflux effect. The results obtained indicate that the CAF/DAP cocrystal, particularly when prepared by the spray drying method, represents a possible suitable approach for inhalation formulations with applications in pulmonary pathologies.