Browsing by Author "Pinheiro, Ana C."
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- Assessing the in vitro digestion of lactoferrin-curcumin nanoparticles using the realistic gastric modelPublication . Madalena, Daniel A.; Araújo, João F.; Ramos, Óscar L.; Vicente, António A.; Pinheiro, Ana C.Nanosized delivery systems have been the subject of research and discussion in the scientific community due to their unique properties and functionality. However, studies reporting the behaviour of nanodelivery systems under dynamic in vitro digestion conditions are still very scarce. To address this gap, this study aims to assess the dynamic in vitro gastric digestion of lactoferrin/curcumin nanoparticles in the realistic gastric model (RGM). For this purpose, the INFOGEST standard semi-dynamic digestion protocol was used. The nanosystems were characterized in terms of hydrodynamic size, size distribution, polydispersity index (PdI), and zeta potential using dynamic light scattering (DLS), before and during the digestion process. Confocal laser scanning microscopy (CLSM) was also used to examine particle aggregation. In addition, the release of curcumin was evaluated spectroscopically and the intrinsic fluorescence of lactoferrin was measured throughout the digestion process. The protein hydrolysis was also determined by UV-VIS-SWNIR spectroscopy to estimate, in real-time, the presence of free NH2 groups during gastric digestion. It was possible to observe that lactoferrin/curcumin nanoparticles were destabilized during the dynamic digestion process. It was also possible to conclude that low sample volumes can pose a major challenge in the application of dynamic in vitro digestion models.
- Emerging challenges in assessing bio-based nanosystems’ behaviour under in vitro digestion focused on food applications – a critical view and future perspectivesPublication . Madalena, Daniel; Fernandes, Jean; Avelar, Zita; Gonçalves, Raquel; Ramos, Óscar L.; Vicente, António A.; Pinheiro, Ana C.The current consumers’ demand for high quality food products together with the growing awareness regarding the link between health and nutrition has led to the development of novel food products with added functionality. Such functionality can be modulated by adding bio-based nanosystems that can improve the bioaccessibility of bioactive compounds and facilitate nutrient absorption. However, these functional properties can be significantly affected by the adverse conditions (e.g., low pH, presence of enzymes, salts) of the gastrointestinal tract. As such, understanding the behaviour of such delivery systems under digestion conditions is of utmost importance and several analytical tools and in vitro digestion models have been used for this purpose. This review summarizes the latest updates on nanosystems’ performance under in vitro digestion and provides critical insights related to important and complementary analytical tools (e.g., rheology, Raman spectroscopy, x-ray scattering) used to assess their performance throughout digestion. Furthermore, the most prominent and frequent challenges associated with such in vitro analyses are also described, together with the current trends regarding the development of in vitro digestion models and some considerations that should be undertaken for their validation. Efforts must be made towards developing reliable and standard in vitro digestion models that use sophisticated analytical techniques to further expand the knowledge regarding nanosystems’ behaviour under in vitro digestion conditions.
- Omega-3 fatty acids from fish by-products: innovative extraction and application in food and feedPublication . Rodrigues, Matilde; Rosa, Ana; Almeida, André; Martins, Rui; Ribeiro, Tânia; Pintado, Manuela; Gonçalves, Raquel F. S.; Pinheiro, Ana C.; Fonseca, António J. M.; Maia, Margarida R. G.; Cabrita, Ana R. J.; Barros, Lillian; Caleja, CristinaOmega-3 fatty acids (O3FA) are essential nutrients that play a crucial role in maintaining human and animal health. They are known for their numerous health claims, including cardiovascular benefits, contributing to both the prevention and treatment of immunological, neurological, reproductive, and cardiovascular complications, and supporting overall well-being. Fish, especially oily fish, comprise rich source of O3FA. In the fish industry, significant amounts of by-products and waste are generated during processing which are often discarded or used for lower-value applications. However, there is recognition of the potential value of extracting O3FA from these by-products. Various extraction techniques can be used, but the goal is to efficiently extract and concentrate the O3FA while minimizing the loss of nutritional value. To prevent oxidation and maintain the stability of O3FA, natural antioxidants can be added. Antioxidants like polyphenolic compounds and plant extracts help to protect the O3FA from degradation caused by exposure to oxygen, light, and heat. By stabilizing the O3FA, the shelf life and nutritional value of the extracted product can be extended. In summary, this work presents a forward-looking strategy for transforming fish by-products into high-quality oils, which hold great potential for application in food and feed.
- β-lactoglobulin micro- and nanostructures as bioactive compounds vehicle: In vitro studiesPublication . Simões, Lívia S.; Martins, Joana T.; Pinheiro, Ana C.; Vicente, António A.; Ramos, Oscar. L.β-Lactoglobulin (β-Lg) is known to be capable to bind hydrophilic and hydrophobic bioactive compounds. This research aimed to assess the in vitro performance of β-Lg micro- (diameter ranging from 200 to 300 nm) and nano (diameter < 100 nm) structures associated to hydrophilic and hydrophobic model compounds on Caco-2 cells and under simulated gastrointestinal (GI) conditions. Riboflavin and quercetin were studied as hydrophilic and hydrophobic model compounds, respectively. Cytotoxicity experiment was conducted using in vitro cellular model based on human colon carcinoma Caco-2 cells. Moreover, the digestion process was simulated using the harmonized INFOGEST in vitro digestion model, where samples were taken at each phase of digestion process - oral, gastric and intestinal - and characterized in terms of particle size, polydispersity index (PDI), surface charge by dynamic light scattering (DLS); protein hydrolysis degree by 2,4,6-trinitrobenzene sulfonic acid (TNBSA) assay and native polyacrylamide gel electrophoresis; and bioactive compound concentration. Caco-2 cell viability was not affected up to 21 × 10−3 mg mL−1 of riboflavin and 16 × 10−3 mg mL−1 quercetin on β-Lg micro- and nanostructures. In the oral phase, β-Lg structures’ particle size, PDI and surface charge values were not changed comparing to the initial β-Lg structures (i.e., before being subjected to in vitro GI digestion). During gastric digestion, β-Lg structures were resistant to proteolytic enzymes and to acid environment of the stomach – confirmed by TNBSA and native gel electrophoresis. In vitro digestion results indicated that β-Lg micro- and nanostructures protected both hydrophilic and hydrophobic compounds from gastric conditions and deliver them to target site (i.e., intestinal phase). In addition, β-Lg structures were capable to enhance riboflavin and quercetin bioaccessibility and bioavailability potential compared to bioactive compounds in their free form. This study indicated that β-Lg micro- and nanostructures were capable to enhance hydrophilic and hydrophobic compounds bioavailability potential and they can be used as oral delivery systems.
- Κ-carrageenan/chitosan nanolayered coating for controlled release of a model bioactive compoundPublication . Pinheiro, Ana C.; Bourbon, Ana I.; Quintas, Mafalda A.C.; Coimbra, Manuel A.; Vicente, António A.Multilayer nanocoatings composed of κ-carrageenan, a sulphated anionic polysaccharide, and chitosan, a cationic polysaccharide, were produced by layer-by-layer deposition. The model cationic compound Methylene Blue (MB) was incorporated in different positions of the nanolayered coating and its loading and release behavior was evaluated. UV–VIS spectroscopy and quartz crystal microbalance analysis showed that the amount of MB loaded increased with the distance from the first layer, suggesting that the MB was able to diffuse into the κ-carrageenan/chitosan nanolayered coating and not only adhered to the surface of the layer immediately below it. For most of the tested conditions, the MB release from the κ-carrageenan/chitosan nanolayered coatings was successfully described by the linear superimposition model, which allowed concluding that MB transport is due to both concentration gradient and the polymer relaxation of the nanolayers. However, depending on temperature and pH of the medium and on the position of MB incorporated on the nanolayered coatings, different mechanisms prevail. Industrial relevance: The development of novel edible coatings with improved functionality and performance for e.g. fresh and minimally processed fruits is one of the challenges of the post-harvest industry. This work contributes to the understanding of the loading and release phenomena involved in structures at the nanoscale, which is useful for the development of bioactive compounds release systems for application in food industry. Moreover, the κ-carrageenan/chitosan nanolayered coatings represent a promising platform from which the controlled release of different bioactive compounds may be explored.