Thermodynamic Mapping of C5‐Substituent Autoxidation Pathways in 1,5‐Dimethylcytosine: A Quantum Chemical Study

Abstract

The autoxidation pathways of the methyl substituent at position 5 in 1,5‐dimethylcytosine (1,5dmC) were investigated using DFT and DLPNO‐CCSD(T) calculations with implicit SMD(H2O) solvation. A radical reaction network was constructed to assess nonenzymatic pathways leading to hydroxymethyl, formyl, and carboxyl oxidation products. The results show that the reactivity of methylated cytosine derivatives is governed not by isolated CH bond strengths alone, but by the coupled thermodynamics of radical formation, oxygen addition, hydroperoxide formation, and hydroperoxide decomposition. Hydrogen abstraction from the 5‐methyl substituent is the most accessible radical‐forming step in 1,5dmC, whereas protonation increases BDE(CH) values and suppresses radical formation. Hydroperoxide intermediates play an important role within the considered propagation network, with OO cleavage being more accessible than OH cleavage. The calculated reaction free energies rationalize the possible accumulation of the formyl derivative 1m5fC: this formyl derivative is formed through strongly exergonic pathways, whereas its further conversion to the carboxyl derivative 1m5caC is more thermodynamically constrained. Representative transition‐state calculations provide a kinetic consistency check, showing only moderate substrate‐dependent variations in H‐abstraction barriers within each model‐radical class. Overall, the results identify thermodynamically relevant radical intermediates, limiting steps, and product‐forming routes within the considered reaction network for the nonenzymatic autoxidation of methylated cytosine derivatives.