ALTERNATING CURRENT BIOSUSCEPTOMETRY TO EVALUATE THE BIOAVAILABILITY OF MAGNETIC NANOPARTICLES IN CACO-2 CELLS AFTER SIMULATED IN VITRO DIGESTION

Oral drug administration faces several challenges due to the low solubility, permeability, and stability of these substances in the gastrointestinal tract. In this scenario, nanotechnology, especially nanostructured delivery systems, represents a promising strategy to overcome these obstacles, increasing bioavailability and therapeutic efficacy. To investigate the absorption of substances by the gastrointestinal tract, the Caco-2 cell model is widely used to simulate the intestinal epithelium. However, its association with systems that simulate in vitro digestion provides an approach of greater physiological relevance. In this context, magnetic nanoparticles (MNPs) can be used as markers to monitor the interaction and internalization of nanostructured systems by Caco-2 cells. For the detection and quantification of these nanoparticles, biomagnetic techniques are required, such as Alternating Current Biosusceptometry (ACB), which enables non-invasive monitoring of the magnetic response of MNPs during the cellular internalization process. In this work, we evaluated the internalization and bioavailability of citrate-coated manganese-doped iron oxide MNPs (MnFe2O4) in Caco-2 cells previously subjected to simulated digestion. Cell culture was performed in medium supplemented with fetal bovine serum, incubated at 37 °C and 5% CO₂. Upon reaching confluence, cells were trypsinized, counted, and seeded for exposure to the already digested nanoparticles. The digestive simulation comprised three consecutive phases (oral, gastric, and intestinal), following the protocol by Minekus et al. (2014). Following exposure at different time intervals, samples were collected and divided into supernatant (non-internalized MNPs) and trypsinized fractions (cell-associated MNPs). Quantification via ACB demonstrated significant variations in internalization profiles over time and across the different digestive stages. In conclusion, in vitro digestion directly influenced the bioavailability of the nanoparticles, highlighting a significant impact of the simulated gastrointestinal environment on the interactions established between the nanostructures and the cellular system.