Design, Synthesis, and Characterization of <i>N</i> -Doped Carbon Dots from a Ternary System of Citric Acid, Urea, and ( <i>E</i> )-2-(2,5-Dimethoxyphenyl)methylenebutane-1,4-dioic Acid

Synthesizing carbon dots (CDs) using therapeutic compounds as precursors has emerged as a promising strategy for biomedical applications, enabling simultaneous utilization of the beneficial properties of both nanoparticles and bioactive molecules without additional modification. In this work, we report the synthesis of novel CDs (ARI-NCDs) using citric acid, urea, and (E)-2-(2,5-dimethoxyphenyl)methylenebutane-1,4-dioic acid via a green, one-step hydrothermal process at 200 °C for 10 h. Control CDs prepared from citric acid and urea (U-NCDs) were synthesized identically. Comprehensive characterization by TEM, FT-IR, Raman, XRD and XPS confirmed nitrogen-doped, oxygen-rich surfaces and successful aryl functionalization. ARI-NCDs exhibited bright, nearly excitation-independent blue emission and a higher quantum yield than U-NCDs. TGA revealed the enhanced thermal stability of ARI-NCDs compared with U-NCDs. Zeta potential measurements across pH 4–10 indicated a single well-defined negative surface charge for ARI-NCDs at neutral pH, consistent with superior colloidal stability and dispersion homogeneity, while U-NCDs displayed charge heterogeneity. In vitro assays using HeLa cells demonstrated efficient cellular internalization of ARI-NCDs and low cytotoxicity, highlighting their suitability for live cell imaging and other biomedical applications. Additionally, ARI-NCDs displayed pronounced antibacterial activity against both Gram-positive and Gram-negative bacteria with larger inhibition zones and lower minimum inhibitory concentrations relative to U-NCDs, attributable to the functional groups and structural attributes imparted by the aryl itaconic acid precursor. These findings underscore the versatility of ARI-NCDs for multifunctional biomedical applications, combining imaging capability with antimicrobial efficacy.