Vitexin and isovitexin are natural flavone C-glucosides that have numerous benefits for human health. However, their low oral bioavailability and poor gastrointestinal absorption dramatically restrict their potential medicinal uses. To overcome this challenge, chitosan-coated alginate microcapsules were prepared for intragastrical administration to rabbits. An LC-MS/MS method was developed and validated for the simultaneous determination of vitexin and isovitexin in the plasma of treated rabbits, using salicylic acid as the internal standard. Raw rabbit plasma samples were deproteinized using acetonitrile as a precipitation agent. Chromatographic separation was performed on a reversed-phase C18 column (100 mm × 4.6 mm, 3.5 µm), with an isocratic mobile solvent system comprising methanol and 0.1% acetic acid (40:60) as the mobile phase. All the analytes and the internal standard were ionized on a triple quadrupole mass spectrometer and electrospray ionization, operating in negative mode and multiple reaction monitoring. The analytical approach developed underwent validation in terms of system suitability, specificity, selectivity, LLOQ of 2 ng/mL, linearity (2.0–200 ng/mL, R² > 0.99), accuracy (the intra- and inter-day from 94 to 110% with the relative standard deviations no more than 8.7%, precision with the recoveries from 97% to 102%, matrix effect (90–100%), carry-over, dilution integrity (2 times), and stability at room and frozen temperature for up to 1 month, and all the parameters met FDA and EMA requirements for bioanalytical methods. The validated procedure was applied to measure the absorption of vitexin and isovitexin from encapsulated extracts in a pilot pharmacokinetic study on rabbit plasma. Compared to the raw traditional extracts, the microcapsules enhanced the bioavailability of vi-texin and isovitexin regarding C_max and AUC values.

Background/Objectives: Vitexin and isovitexin are bioactive flavonoids with promising pharmacological effects; however, they have poor bioavailability. Microencapsulation with biodegradable polymers is a promising strategy for improving their stability, bioavailability, and biocompatibility. This study aimed to optimize the formulation parameters to obtain microspheres with desired properties in terms of size, loading ratio, and vitexin–isovitexin release. Methods: Microspheres were prepared using alginate as the core matrix and a chitosan outer layer. A Design of Experiment approach using response surface methodology was employed. The hypoglycemic effects of the obtained microspheres were evaluated. Results: The formulation using 1.17% low-viscosity alginate, 7.60% calcium chloride, 5.78% Tween 80, and 5.00% Span 80 resulted in microspheres with optimal mean size (10.78 µm), high loading ratio (22.45%) and encapsulation efficiency (68.92%). The in vitro release of vitexin–isovitexin from microspheres was completed within 24 h in controlled manner. The microspheres were found to be non-toxic in vivo and exhibited hypoglycemic effects after 21 days at doses equivalent to 30 and 60 mg/kg of vitexin–isovitexin. The potential mechanisms might involve increasing the size of Islets of Langerhans and improving pancreatic β-cell function and insulin resistance, as observed in alloxan-induced diabetic mice. Conclusions: This work successfully developed alginate–chitosan-based microspheres for the controlled release of vitexin–isovitexin while maintaining their bioactivities.

Background/Objectives: Vitexin and isovitexin are bioactive flavonoids with promising pharmacological effects; however, they have poor bioavailability. Microencapsulation with biodegradable polymers is promising strategy for improving their stability, bioavailability, and biocompatibility. This study aimed to optimize the formulation parameters to obtain microspheres with desired properties in terms of size, loading ratio, and vitexin-isovitexin release. Methods: Microspheres were prepared using alginate as core matrix and a chitosan outer layer. Design of experiments approach using response surface methodology was employed. The hypoglycemic effects of obtained microspheres were evaluated. Results: The formulation using 1.17% low-viscosity alginate, 7.60% calcium chloride, 5.78% Tween 80, and 5.00% Span 80 resulted in microspheres with optimal mean size (10.78 µm), high loading ratio (22.45%) and encapsulation efficiency (68.92%). The in vitro release of vitexin-isovitexin from microspheres was completed within 24 hours in controlled manner. The microspheres were found to be non-toxic in vivo and exhibited hypoglycemic effects after 21 days at doses equivalent to 30 and 60 mg/kg of vitexin-isovitexin. These effects were attributed to increasing in the size of Islets of Langerhans, improving in pancreatic β-cell function and insulin resistance, as observed in alloxan-induced diabetic mices. Conclusions: This work successfully developed alginate-chitosan-based microspheres for the controlled release of vitexin-isovitexin while maintaining their bioactivities.

Interleukin-33 (IL-33) is a therapeutic target in inflammatory diseases, yet inhibitor screening is hindered by high commercial protein costs. While expressing IL-33 as a SUMO fusion protein overcomes yield limitations in Escherichia coli, whether this intact chimera can directly serve as a screening surrogate remains unresolved. Here, we combined in silico simulations with empirical bioprocess optimization to establish a high-yield pipeline for native human IL-33 production. AI-based structure predictions (AlphaFold and RoseTTAFold) revealed distinct inter-domain dynamics mediated by a flexible linker. Molecular dynamics simulations of the compact RoseTTAFold model suggested potential steric occlusion near receptor-binding interfaces, whereas AlphaFold models exhibited extended conformations, highlighting model-dependent variations that warrant further structural verification. To restore accessibility, tag cleavage was optimized using 1.0% (mol/mol) SUMO protease at 4 °C for 1 h, achieving near-complete digestion. Sandwich ELISA demonstrated an 87.6% molar epitope recognition recovery for cleaved IL-33 versus only 12.0% for the intact fusion, providing empirical evidence of substantial functional epitope masking by the SUMO tag. Two-step IMAC purification yielded ~19% mass recovery (~32.6% molar recovery). These findings demonstrate that tag removal is operationally essential for IL-33 functional availability in screening applications.