Reshaping Insulin Fibrillation at the Nano-Bio Interface Using Carbon Quantum Dots: Implications for Amyloid...

Insulin (Ins), a 51-residue α-helical hormone, is prone to forming insoluble aggregates during administration, production, and storage, which poses significant challenges to its stability, effectiveness, and formulation. In this regard, nanoparticles have attracted tremendous interest owing to their unique physicochemical properties and high affinity for biomolecules. Carbon quantum dots are one of the newly emerged nanoparticles found effective against amyloid fibrillation due to aromaticity, hydrophobicity, and strong H-bonding ability. Herein, we studied the effect of hydrothermally synthesized CQDs on the Ins fibrillation under normal conditions. The 2-3 nm CQDs with graphitic-core and oxygenic surface functionalities were synthesized using l-ascorbic acid and their effect on Ins fibrillation kinetics was studied. Thioflavin-T (ThT) fluorescence assay showed that CQDs can reduce Ins fibrillation in a dose-dependent manner. Transmission electron microscopy indicated significant inhibition of fibril formation in 1.5 mg/mL Ins with 0.15 mg/mL CQDs at physiological pH. This was further validated by the disappearance of the β-sheet signature peak in CD spectra and lower β-sheet content in the amide-I region of the Raman spectra. Thermodynamic analysis suggested that the interaction between CQDs and Ins relies largely on H-bonding and dispersion forces, leading to ground-state complexation and static fluorescence quenching of Ins. Importantly, CQDs helped maintain the secondary structure of Ins and improved the retention of tertiary structure as revealed by the ANS binding assay. These findings highlight the potential of CQDs as effective nanomaterial-based inhibitors of insulin fibrillation while offering insights into their binding interactions. However, further research regarding their long-term compatibility and safe pharmacological formulation is required for clinical translation. This work will help develop next-generation nanotherapeutic strategies aiming at improved Ins stability and storage.