Novel unsymmetrically N-substituted 1,3,5-triazacyclohexanes: Synthesis, X-ray structural characterization, In vitro antioxidant evaluation and structure-activity relationships supported by DFT and Molecular Docking studies

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In this work, two novel unsymmetrical 1,3,5-triazacyclohexane derivatives, 1,3-bis(4-methylphenyl)-5-cyclohexyl-1,3,5-triazinane (1) and 1,3-bis(4-iodophenyl)-5-cyclohexyl-1,3,5-triazinane (2) , were efficiently synthesized via a mild one-pot condensation of mixed primary amines with formalin. The compounds were obtained in high yields and characterized by FT-IR, 1 H/ 13 C NMR spectroscopy, and single-crystal X-ray diffraction. Compound (1) crystallizes in the orthorhombic space group Pnma with an eea conformation, while compound (2) crystallizes in P2 1 /c with an aae conformation. Hirshfeld surface analysis revealed the predominance of long-range H···H contacts in both crystal packings. DFT calculations, including geometry optimization, electronic structure, NBO, and vibrational frequency analyses, provided further insight into their structural and electronic properties. The antioxidant activity of the synthesized compounds was evaluated using DPPH•, ABTS•⁺, FRAP, o-phenanthroline, and NO• scavenging assays. Although weak activity was observed in the DPPH test, the compounds exhibited significant electron-transfer antioxidant behavior in the ABTS and iron-reducing assays. The methyl-substituted derivative showed the highest ABTS•⁺ scavenging activity, comparable to BHT and BHA. Preliminary structure–activity analysis suggested that the electron-donating methyl substituent was associated with enhanced antioxidant capacity, whereas replacement by iodoaryl groups reduced the antioxidant response under the experimental conditions investigated. Molecular docking studies were performed to explore the potential interactions of the synthesized triazacyclohexane derivatives with enzymes associated with oxidative stress, including MAO-B, NOX2, XO, and COX-1. The results suggested that compound (2) displayed the most favorable predicted binding affinities toward MAO-B, NOX2, and XO, while weaker interactions were observed with COX-1. These differences were associated with the nature of the substituents, where the presence of iodo-substituted aromatic groups enhanced interaction diversity and binding stability compared to the methyl-substituted analogue. While these findings provide preliminary molecular insights into possible enzyme-binding behavior, they did not directly parallel the experimentally observed antioxidant ranking, which appears mainly governed by electron-transfer mechanisms rather than enzyme-binding affinity. Therefore, these results should be considered as complementary molecular insights into possible enzyme-binding behavior.