Oscar Edgardo Perez

The mechanisms underlying the health benefits of probiotics remain incompletely understood. Among the proposed mediators, bacterial extracellular vesicles (EVs) have been proposed as mediators of interkingdom communication between probiotic bacteria and their hosts. In this study, we show that Lacticaseibacillus casei BL23 releases higher EV levels during late stationary phase, with a predominant population of ∼113 nm. Scanning electron microscopy (SEM) confirmed the ultrastructural morphology of the EV preparations. Proteomic analysis revealed enrichment of proteins associated with probiotic function, including P40 and P75, as well as adhesion‐related moonlighting proteins such as glyceraldehyde 3‐phosphate dehydrogenase (GAPDH) and enolase implicated in interactions with intestinal mucus and epithelial cells. L. casei BL23–derived EVs showed dose‐dependent interactions with mucin and with differentiated Caco‐2 epithelial monolayers in vitro. Using Transwell systems, we further demonstrated that intact EVs cross these polarized monolayers within 240 min. Super‐resolution microscopy further revealed time‐dependent uptake and intracellular trafficking of fluorescently‐labeled EVs. Together, these findings, obtained in vitro, are consistent with a role for probiotic‐derived EVs as postbiotics capable of mediating microbiota–host interactions at the intestinal barrier, providing the experimental basis for future validation in more complex physiological models.

The objective of this work was to generate and characterize complexes of quinoa proteins (QP) with either resveratrol (RSV) or tocopherol (TOC) whose dimensions fell into the nanoscale. The QP‐based nanocomplexes characterization was carried out jointly with encapsulation efficiency determination. Fourier‐Transform Infrared Spectroscopy (FTIR) showed no covalent bond formation and indicated that the protein's secondary structure was modified. Transmission Electron Microscopy (TEM) images and Dynamic Light Scattering (DLS) measurements confirmed that the addition of bioactive compounds did not substantially alter the protein's original size. A consistent particle diameter of 100 nm was observed in both types of nanocomplexes with and without RSV and TOC. The antioxidant capacity of nanocomplexes was analyzed after an in vitro simulated digestion process, showing an increase in this property postdigestion. These nanocomplexes hold great potential as functional ingredients.

The objective of this work was to generate and characterize complexes of quinoa proteins (QP) with either resveratrol (RSV) or tocopherol (TOC) whose dimensions fell into the nanoscale. The QP-based nanocomplexes characterization was carried out jointly with encapsulation efficiency determination. Fourier-Transform Infrared Spectroscopy (FTIR) showed no covalent bond formation and indicated that the protein’s secondary structure was modified. Transmission Electron Microscopy (TEM) images and Dynamic Light Scattering (DLS) measurements confirmed that the addition of bioactive compounds did not substantially alter the protein’s original size. A consistent particle diameter of 100 nm was observed in both types of nanocomplexes with and without RSV and TOC. The antioxidant capacity of nanocomplexes was analyzed after an in vitro simulated digestion process, showing an increase in this property post-digestion. These nanocomplexes hold great potential as functional ingredients.

Background: Chitosan (CS) crosslinked with genipin (GNP) provides a mild, non-toxic route to generate nanogels (NGs) with enhanced integrity and colloidal stability. Objectives: To develop and characterise CS-GNP NG as a novel platform for targeted cellular delivery, optimising design through physicochemical characterisation and biocompatibility evaluation. Methods: NGs were synthesised under optimised conditions by adjusting the pH of the CS solution, followed by high-intensity ultrasound (HIUS) to achieve disaggregation. Physicochemical characterisation was carried out using UV-Vis spectroscopy, FTIR, dynamic light scattering (DLS), and scanning electron microscopy (SEM). Rheological studies and SAXS analysis assessed structural properties. Biocompatibility was evaluated via MTT assay, and internalisation was monitored by fluorescence microscopy on mammalian cell lines. Results: NG formation was highly pH-dependent, with optimal configuration at pH 4.5, yielding stable, uniformly sized particles (~200 nm, ζ-potential +29 mV). Kinetic modelling showed a sigmoidal formation pattern, suggesting nucleation, growth, and stabilisation. FTIR confirmed covalent bonding between CS and GNP via primary amide bonds and Schiff bases. Rheology indicated pseudoplastic behaviour, and SAXS revealed a compact network formation. Biocompatibility assays confirmed non-cytotoxicity below 100 µg/mL and efficient cellular uptake. Conclusions: This study presents a rapid, reproducible protocol for generating colloidally stable, biocompatible NGs suitable for drug delivery.

Background: the crosslinking of chitosan (CS) with genipin (GNP) offers a mild, non-toxic strategy to improve the structural integrity and stability of nanogels (NGs). However, generating homogeneous, colloidally stable systems still requires optimisation of parameters such as CS and GNP concentrations to ensure functional performance for cellular uptake. Objectives: To develop and characterise CS-based NGs crosslinked with GNP as a novel platform for targeted delivery at the cellular level. The aim was to optimise NG synthesis and evaluate their physicochemical properties and biocompatibility. Methods: NGs were synthesised under optimised conditions by adjusting the pH of the CS solution, followed by high-intensity ultrasound (HIUS) to achieve disaggregation. Physicochemical characterisation was carried out using UV-Vis spectroscopy, FTIR, dynamic light scattering (DLS), and scanning electron microscopy (SEM). Rheological studies and SAXS analysis assessed structural properties. Biocompatibility was evaluated via MTT assay, and internalisation was monitored by fluorescence microscopy on mammalian cell lines. Results: NG formation was highly pH-dependent, with optimal configuration at pH 4.5, yielding stable, uniformly sized particles (~200 nm, ζ-potential +29 mV). Kinetic modelling showed a sigmoidal formation pattern, suggesting nucleation, exponential growth, and stabilisation. FTIR confirmed covalent bonding between CS and GNP via primary amide bonds and Schiff bases. Rheology indicated pseudo-plastic behaviour, and SAXS revealed a compact network. Biocompatibility assays confirmed non-cytotoxicity below 50 µg/mL and efficient cellular uptake. Conclusions: This study presents a rapid, reproducible protocol for generating colloidally stable, biocompatible NGs suitable for drug delivery and biomedical applications requiring cellular internalisation.