Browsing by Author "Sarmento, Bruno"
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- Antioxidants in the prevention and treatment of diabetic retinopathy – a reviewPublication . Silva, Sara Baptista da; Costa, João Paulo; Pintado, Manuela Estevez; Ferreira, Domingos de Carvalho; Sarmento, BrunoDiabetic retinopathy is a disease resulting from diabetic chronic hyperglycemia characterized by microvascular complications in the retina, where neuronal elements responsible for vision are located. It is the main cause of adult blindness in developed countries. Oxidative stress has been widely regarded as the key factor for the emergence of ocular disease and has been involved in increased vascular permeability, disruption of blood-retinal barrier, apoptotic loss of retinal capillary cells, microvascular abnormalities and retinal neovascularization. Dietary supplementation with antioxidants has been related with inhibition of diabetes-induced abnormalities of retinal metabolism, reduction of apoptosis and partial restoration of pericytes. Moreover, the use of topical antioxidants to treat or delaying oxidative stress-related ocular manifestations is still poorly explored, while current diabetic retinopathy therapy includes invasive methods, like surgery. Ocular antioxidant potential therapy represents a non-invasive, safe and less painful methodology, which slows the natural progress of the disease and improves the effectiveness of treatment without significant systemic toxicity. This review underlines the innovative medicines exploited for ocular conditions, a further insight on ocular delivery, benefiting from the advantages of the eye for drug delivery and, additionally, offering new potential applications of antioxidants for the prevention, treatment and control of diabetic retinopathy.
- Bioadhesive polymeric nanoparticles as strategy to improve the treatment of yeast infections in oral cavity: in-vitro and ex-vivo studiesPublication . Roque, Luís; Castro, Pedro; Molpeceres, Jesus; Viana, Ana S.; Roberto, Amilcar; Reis, Claúdia; Rijo, Patrícia; Tho, Ingunn; Sarmento, Bruno; Reis, CatarinaCandidiasis is by far the most common fungal infection in the oral cavity and locally antifungal drugs are generally considered the first line therapy. However, some antifungal drugs like Nystatin (Nys) have very short time in the target tissue, which generally it requires several daily applications. One of the solutions to overcome those issues could be the incorporation of Nys into small, protective and polymeric nanoparticles (NPs). In this work, unloaded NPs were prepared and fully characterized and implemented in toothpaste and an oral gel. The three-best unloaded system (smallest, stable, safe and bioadhesive) was selected for further studies. After loading, NPs presented a size range from 300 to 800 nm, high encapsulation efficiency (above 70%), prolonged release and high adhesion capacity to oral mucosa when compared with free Nys. Free Nys lost its activity when it was not encapsulated. The presented results suggested that these new bioadhesive systems could be considered as good candidates to improve the time residence of Nys in buccal mucosa, reducing the number of applications during the day, using alcohol free-formulations and easily included in the daily lives of the patients.
- Characterization of solid lipid nanoparticles produced with carnauba wax for rosmarinic acid oral deliveryPublication . Madureira, Ana Raquel; Campos, Débora A.; Fonte, Pedro; Nunes, Sara; Reis, Flávio; Gomes, Ana Maria; Sarmento, Bruno; Pintado, Maria ManuelaIn the last decade, research studies have increased on the development of delivery systems for polyphenols, for protection, improvement of stability and increase of their bioavailability. Rosmarinic acid is a polyphenol with described bioactivities, such as antioxidant, anti-mutagenic, anti-bacterial and anti-viral capabilities. Thus, the aim of this research work was to produce stable solid lipid nanoparticles (SLN) using carnauba wax as lipidic matrix, for delivery of rosmarinic acid, to be further incorporated into food matrices. Hence, different concentrations of wax (0.5, 1 and 1.5%, w/v) and percentages of surfactant (1, 2 and 3%, v/v) were tested. Physical properties, surface morphology and association efficiencies were studied at time of production and after 28 day at refrigerated storage. Thermal properties and the nature of the chemical interactions between the lipids waxes and rosmarinic acid were also evaluated. The particles showed range size between 35-927 nm and zeta potentials of ca. -38 to 40, showing high stability, with no risk of aggregation due to electric repulsion of SLN. High association efficiencies % (ca. 99%) were obtained. FTIR analyses proved the association of rosmarinic acid and lipidic matrix. The low lipid and high surfactant concentrations leads to small SLN. The surfactant, polysorbate 80 decreases the interfacial tension in the SLN surfaces, preventing aggregation, leading to the development of small particles. These properties were maintained throughout the 28 day of refrigerated storage, and no rosmarinic acid was released by the particles during refrigeration, indicating good compatibility between rosmarinic acid and the waxy core of SLN. The optimum range values to obtain the desirable features for incorporation in a functional food suggest formulations containing 1.0 and 1.5% (w/v) of lipid and 2% (v/v) of surfactant.
- Chitosan nanoparticles for daptomycin delivery in ocular treatment of bacterial endophthalmitisPublication . Silva, Nádia C.; Silva, Sara; Sarmento, Bruno; Pintado, ManuelaContext: Chitosan nanoparticles were prepared to encapsulate daptomycin and proposed as a delivery system of this antibiotic to the eye for the treatment of bacterial endophthalmitis. Objective: The aim of this study was to develop daptomycin-loaded nanoparticles to apply directly to the eye, as a possible non-invasive and less painful alternative for the treatment of endophthalmitis, increasing the effectiveness of treatment and reducing toxicity associated with systemic administration.Materials and methods: Nanoparticles were obtained by ionotropic gelation between chitosan and sodium tripolyphosphate (TPP). Physicochemical and morphological characteristics of nanoparticles were evaluated, as well as determination of antimicrobial efficiency of encapsulated daptomycin and stability of the nanoparticles in the presence of lysozyme and mucin.Results: Loaded nanoparticles presented mean particle sizes around 200 nm, low polydispersity index, and positive zeta potential. Morphological examination by scanning electron microscopy (SEM) confirmed their small size and round-shaped structure. Encapsulation efficiency ranged from 80 to 97%. Total in vitro release of daptomycin was obtained within 4 h. Determination of minimum inhibitory concentrations (MICs) showed that bacteria were still susceptible to daptomycin encapsulated into the nanoparticles. Incubation with lysozyme did not significantly affect the integrity of the nanoparticles, although mucin positively affected their mucoadhesive properties. Discussion and conclusion: The obtained nanoparticles have suitable characteristics for ocular applications, arising as a promising solution for the topical administration of daptomycin to the eye.
- Chitosan-based nanoparticles for rosmarinic acid ocular delivery-in vitro testsPublication . Silva, Sara; Ferreira, Domingos; Pintado, M. E.; Sarmento, BrunotIn this study, chitosan nanoparticles were used to encapsulate antioxidant rosmarinic acid, Salvia offici-nalis (sage) and Satureja montana (savory) extracts as rosmarinic acid natural vehicles. The nanoparticleswere prepared by ionic gelation using chitosan and sodium tripolyphosphate (TPP) in a mass ratio of7:1, at pH 5.8. Particle size distribution analysis and transmission electron microscopy (TEM) confirmedthe size ranging from 200 to 300 nm, while surface charge of nanoparticles ranged from 20 to 30 mV.Nanoparticles demonstrate to be safe without relevant cytotoxicity against retina pigment epithelium(ARPE-19) and human cornea cell line (HCE-T). The permeability study in HCE monolayer cell line showedan apparent permeability coefficient Pappof 3.41 ± 0.99 × 10−5and 3.24 ± 0.79 × 10−5cm/s for rosmarinicacid loaded chitosan nanoparticles and free in solution, respectively. In ARPE-19 monolayer cell line thePappwas 3.39 ± 0.18 × 10−5and 3.60 ± 0.05 × 10−5cm/s for rosmarinic acid loaded chitosan nanoparti-cles and free in solution, respectively. Considering the mucin interaction method, nanoparticles indicatemucoadhesive proprieties suggesting an increased retention time over the ocular mucosa after instilla-tion. These nanoparticles may be promising drug delivery systems for ocular application in oxidative eyeconditions.
- Combination of PLGA nanoparticles with mucoadhesive guar-gum films for buccal delivery of antihypertensive peptidePublication . Castro, Pedro M.; Baptista, Patrícia; Madureira, Ana Raquel; Sarmento, Bruno; Pintado, Manuela E.Oral administration of proteins and peptides still is a challenging task to overcome due to low permeability through absorptive epithelia, degradation and metabolism that lead to poor bioavailability. Attempting to overcome such limitations, an antihypertensive peptide derived from whey protein, with KGYGGVSLPEW sequence, was incorporated for the first time into polymeric nanoparticles. An experimental design was followed in order to optimize drug-loading, association efficiency, mean particle size, zeta-potential and polydispersity index of a formulation of poly(lactic-co-glycolic acid) (PLGA) nanoparticles as carriers for bioactive peptides. In sequence, peptide-loaded PLGA nanoparticles were incorporated in a guar-gum film matrix, resulting in a combined delivery system aiming to promote slow release and permeation across buccal epithelium. Neither PLGA nanoparticles, guar-gum films nor the conjugation of PLGA nanoparticles and guar-gum films (GfNp) significantly compromised in vitro TR146 human buccal carcinoma cell line viability after 12 h contact, as assessed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide reduction assay (MTT). In vitro release assay for developed formulations allowed to conclude that the combination of orodispersible film and nanoparticles granted a slower release of AhP when compared with PLGA or guar-gum films alone or with control. GfNp offered more effective, synergistic, in vitro permeation of TR146 cell multilayer in comparison with guar-gum films or PLGA nanoparticles alone. The combination of PLGA nanoparticles with guar-gum films represent a suitable alternative to conventional per os delivery systems, leading to an increased buccal permeability of carried antihypertensive peptide.
- Delivery systems for antimicrobial peptides and proteins: towards optimization of bioavailability and targetingPublication . Costa, Joana R.; Sarmento, Bruno; Silva, Nádia C.; Pintado, ManuelaAntimicrobial peptides (AMPs) and proteins are produced by a wide range of organisms as important elements of their defense mechanisms, forming a large number of antimicrobial compounds that can be used to treat several human infections. The potential for the use of AMPs and antimicrobial proteins in therapeutics is growing, but their application is often limited, due to their poor physical and/or chemical properties. In recent years, several drug delivery systems have been proposed to carry such molecules, in an attempt to overcome the difficulties regarding their properties. However, no review has yet systematized the most relevant information on this subject. Therefore, this review summarizes the work that has been conducted to develop delivery systems for the transport and protection of AMPs and antimicrobial proteins, including their description and potential applications, while highlighting the opportunities for future research in this field.
- Development and characterization of chitosan microparticles-in-films for buccal delivery of bioactive peptidesPublication . Batista, Patrícia; Castro, Pedro; Madureira, Ana Raquel; Sarmento, Bruno; Pintado, ManuelaNowadays, bioactive peptides are used for therapeutic applications and the selection of a carrier to deliver them is very important to increase the efficiency, absorption, release, bioavailability and consumer acceptance. The aim of this study was to develop and characterize chitosan-based films loaded with chitosan microparticles containing a bioactive peptide (sequence: KGYGGVSLPEW) with antihypertensive properties. Films were prepared by the solvent casting method, while the microparticles were prepared by ionic gelation. The final optimized chitosan microparticles exhibited a mean diameter of 2.5 m, a polydispersity index of 0.46, a zeta potential of +61 mV and a peptide association efficiency of 76%. Chitosan films were optimized achieving the final formulation of 0.79% (w/v) of chitosan, 6.74% (w/v) of sorbitol and 0.82% (w/v) of citric acid. These thin ( 0.100 mm) and transparent films demonstrated good performance in terms of mechanical and biological properties. The oral films developed were flexible, elastic, easy to handle and exhibited rapid disintegration (30 s) and an erosion behavior of 20% when they came into contact with saliva solution. The cell viability (75–99%) was proved by methylthiazolydiphenyl-tetrazolium bromide (MTT) assay with TR146 cells. The chitosan mucoadhesive films loaded with peptide–chitosan microparticles resulted in an innovative approach to perform administration across the buccal mucosa, because these films present a larger surface area, leading to the rapid disintegration and release of the antihypertensive peptide under controlled conditions in the buccal cavity, thus promoting bioavailability.
- Development and validation method for simultaneous quantification of phenolic compounds in natural extracts and nanosystemsPublication . Silva, Sara Baptista da; Oliveira, Ana; Ferreira, Domingos; Sarmento, Bruno; Pintado, ManuelaIntroductionSage and savoury (Salvia sp. and Satureja montana, respectively) are plants used in traditional medicine. The quality control of their herbal formulations is of paramount concern to guarantee the expected biological activity of their anti-oxidant compounds. Objective To establish a simple and effective high-performance liquid chromatographic (HPLC) method to evaluate simultaneously quercetin and rosmarinic acid, in a pure form, in natural extracts (sage and savoury), and encapsulated into chitosan nanoparticles. Methods Chromatography was performed on an RP C-18-column, in a gradient mode with a mobile phase comprising methanol:formic acid:water 92.5:2.5:5 (v/v) at a flow rate of 0.75mL/min and at wavelength of 280nm. Results The method was specific, linear in the range of 0.05-1mg/mL (R-2=1.00), precise at the intraday and interday levels, accurate (recovery rate 90.50.6%), and robust to changes in equipment conditions. Conclusion The method established was effective for quercetin and rosmarinic acid characterisation in natural extracts and in chitosan nanoparticles, allowing the loading capacity determination, the association efficiency as well as the in vitro release. This study shows a new HPLC method validated to simultaneous quantification of rosmarinic acid and quercetin in natural extracts and chitosan nanoparticles. This method is also able for detection and quantification of other phenolic and flavanoid compounds in natural matrix.
- Edible films as oral delivery systems for xanthines extracted from medicinal plants: an experimental design approachPublication . Castro, Pedro M.; Oliveira, Ana; Fonte, Pedro; Madureira, Ana Raquel; Sarmento, Bruno; Pintado, ManuelaTwo formulations of edible films intended for oral delivery of therapeutic xanthines were developed, following an experimental design approach. Gelatin type A and sodium carboxymethylcellulose were used as polymeric matrices with different physico-chemical nature. Caffeine, a well-known methylxhanthine, was used as model bioactive molecule, representing overall xanthines (e.g. caffeine, theophylline, theobromine) extracted from medicinal plants. Fourier-transform infrared spectroscopy (FTIR) analysis was performed to outwit the formation of covalent bonds between caffeine and the matrix of edible films. Scanning electron microscopy (SEM) was performed to assess if caffeine was homogeneously dispersed on the matrix of edible films. Simulation of gastrointestinal tract and ex vivo permeability studies across intestinal mucosa were performed to predict the delivery profile of caffeine from developed formulations of edible films. Gelatin type A-based edible films offered a slow release of caffeine whereas sodium carboxymethylcellulose-based edible films promote an immediate release of caffeine.