Titanium dioxide nanoparticles (TiO₂ NPs), designated as E171 in food applications, have become a subject of intense scientific scrutiny due to their potential to induce oxidative stress, inflammation, and cellular damage in the gastrointestinal tract. While initial toxicological assessments focused on the intrinsic properties of the particles, recent evidence suggests that the biological fate of these nanomaterials is heavily influenced by their interaction with biomolecules present in the digestive environment. This study investigates how protein adsorption—specifically from gelatin and β-lactoglobulin—affects the colloidal stability, aggregation behavior, and intracellular penetration of TiO₂ NPs using human intestinal epithelial cells (HT-29) as a model system.
The experimental design involved coating rutile-phase TiO₂ nanoparticles with either gelatin or β-lactoglobulin at interfacial concentrations of 0.32 mg/mg and 0.25 mg/mg, respectively. Dynamic light scattering (DLS) analysis revealed that uncoated TiO₂ NPs underwent rapid aggregation in aqueous solution, with particle size increasing from ~100 nm to over 480 nm within five days. In contrast, both protein-coated formulations remained stable, maintaining average diameters of approximately 91 nm (gelatin) and 90 nm (β-lactoglobulin), indicating that the protein corona effectively prevents particle agglomeration. The autocorrelation functions further confirmed reduced hydrodynamic radius and improved dispersion stability upon protein adsorption.
Fourier-transform infrared spectroscopy (FTIR) under D₂O provided insight into structural changes induced by surface binding. For β-lactoglobulin, an increase in the 1455 cm⁻¹ band intensity—associated with N–D stretching—was observed, suggesting enhanced solvent accessibility of amide groups and conformational rearrangement upon adsorption. This indicates that β-lactoglobulin undergoes structural reorganization, likely facilitating stronger interactions with the TiO₂ surface. Gelatin, in contrast, exhibited minimal spectral shifts, implying limited conformational adaptation, which may explain its relatively lower performance in stabilizing dispersions.
To evaluate cellular uptake, confocal Raman microscopy was employed to track nanoparticle localization without fixation, labeling, or genetic modification. The characteristic Raman signals of TiO₂ (275 cm⁻¹, 450 cm⁻¹, 610 cm⁻¹) were detected exclusively within HT-29 cells when protein-coated NPs were used. After one hour of incubation, only 35% of uncoated NPs entered the cells, whereas this proportion increased to 42% with gelatin coating and 55% with β-lactoglobulin.Vimentin Antibody Biological Activity Moreover, the percentage of cells containing internalized NPs rose from 28% (control) to 48% (gelatin) and 70% (β-lactoglobulin), demonstrating a significant enhancement in bioavailability.Cyclin D2 Antibody MedChemExpress
Spatial mapping revealed that protein-coated NPs were distributed throughout the cytoplasm, often localized near mitochondria and lysosomes—organelles frequently targeted by nanoparticles. The high spatial resolution (300 nm) of Raman microscopy enabled precise tracking of individual particles, confirming that smaller, non-aggregated entities were preferentially internalized. This aligns with the principle that nanoscale dimensions are essential for effective cellular entry.
These results demonstrate that protein adsorption fundamentally alters the biological behavior of TiO₂ NPs.PMID:34757449 By preventing aggregation and maintaining a stable dispersion, protein coronas enhance the likelihood of cellular uptake. Furthermore, the nature of the protein—its secondary structure, charge, and conformational flexibility—plays a critical role in determining the efficiency of penetration. The superior performance of β-lactoglobulin may be attributed to its ability to undergo structural changes that optimize surface binding despite its positive charge at physiological pH.
This study underscores the importance of considering dietary proteins in risk assessment models for nanomaterials. The formation of a protein corona is not merely a passive phenomenon but an active determinant of nanoparticle fate in biological systems. Future research should explore the long-term consequences of enhanced intracellular delivery, including organelle dysfunction, metabolic disruption, and potential genotoxicity. These findings call for updated regulatory frameworks that account for dynamic interactions between food additives and biomolecules in the human body.MedChemExpress (MCE) offers a wide range of high-quality research chemicals and biochemicals (novel life-science reagents, reference compounds and natural compounds) for scientific use. We have professionally experienced and friendly staff to meet your needs. We are a competent and trustworthy partner for your research and scientific projects.Related websites: https://www.medchemexpress.com