In the absence of external electron donors, oxidized bovine cytochrome c oxidase (CcO) exhibits the ability to decompose excess H₂O₂. Depending on the concentration of peroxide, two mechanisms of degradation were identified. At submillimolar peroxide concentrations, decomposition proceeds with virtually no production of superoxide and oxygen. In contrast, in the millimolar H₂O₂ concentration range, CcO generates superoxide from peroxide. At submillimolar concentrations, the decomposition of H₂O₂ occurs at least at two sites. One is the catalytic heme a₃–Cu_B center where H₂O₂ is reduced to water. During the interaction of the enzyme with H₂O₂, this center cycles back to oxidized CcO via the intermediate presence of two oxoferryl states. We show that at pH 8.0 two molecules of H₂O₂ react with the catalytic center accomplishing one cycle. In addition, the reactions at the heme a₃–Cu_B center generate the surface-exposed lipid-based radical(s) that participates in the decomposition of peroxide. It is also found that the irreversible decline of the catalytic activity of the enzyme treated with submillimolar H₂O₂ concentrations results specifically from the decrease in the rate of electron transfer from heme a to the heme a₃–Cu_B center during the reductive phase of the catalytic cycle. The rates of electron transfer from ferrocytochrome c to heme a and the kinetics of the oxidation of the fully reduced CcO with O₂ were not affected in the peroxide-modified CcO.

Due to the simple one-step preparation method and a promising application in biomedical research, amphiphilic gradient copoly(2-oxazoline)s are gaining more and more interest compared to their analogous block copolymers. In this work, the curcumin solubilization ability was tested for a series of amphiphilic gradient copoly(2-oxazoline)s with different lengths of hydrophobic side-chains, consisting of 2-ethyl-2-oxazoline as a hydrophilic monomer and 2-(4-alkyloxyphenyl)-2-oxazoline as a hydrophobic monomer. It is shown that the length of the hydrophobic side-chain in the copolymers plays a crucial role in the loading of curcumin onto the self-assembled nanoparticles. The kinetic stability of self-assembled nanoparticles studied using FRET shows a link between their integrity and cellular uptake in human glioblastoma cells. The present study demonstrates how minor changes in the molecular structure of gradient copoly(2-oxazoline)s can lead to significant differences in the loading, stability, cytotoxicity, cellular uptake, and pharmacokinetics of nano-formulations containing curcumin. The obtained results on the behavior of the complex of gradient copoly(2-oxazoline)s and curcumin may contribute to the development of effective next-generation polymeric nanostructures for biomedical applications.