An innovative azo mutual prodrug combining procaine and doxycycline intended for colon-specific targeting was successfully synthesised and characterised. This study presents the synthesis and assessment of a new mutual azo prodrug (dox-proc) including doxycycline (antibacterial, anti-inflammatory) and procaine (to mitigate hyperactive autonomic nerves and uncontrolled immunological responses), specifically designed to target ulcerative colitis in the colon. In silico ADME profiling indicated unfavorable physicochemical properties for intestinal absorption, attributable to the compound’s high molecular weight, hydrophilic log P, and elevated topological polar surface area (TPSA). Molecular docking studies revealed high binding affinity to azoreductase, with a docking score superior to that of sulfasalazine, suggesting efficient colonic bioactivation. Dox-proc was synthesized with a good yield (75 %) and purity (m.p: 258–260°C); The structure was validated by FTIR and ¹H-NMR spectroscopy. Pharmacokinetic study demonstrated low intestinal absorption potential (experimental log P = –1.65), dox-proc is high stabile in acidic/basic conditions. Fecal hydrolysis studies verified efficient enzymatic activation in the presence of azoreductase, with hydrolysis up to 81%. Biological studies show that dox-proc group had the lowest score (1.2 ± 0.310) in clinical activity score system, this indicated that dox-proc has the highest clinical activity. Histopathological study confirms a good sign of mucosal healing. These findings align with the in silico predictions and validate the compound's applicability as a colon-targeted therapy.

An innovative azo mutual prodrug combining procaine and doxycycline intended for colon-specific targeting was successfully synthesised and characterised. This study presents the synthesis and assessment of a new mutual azo prodrug (dox-proc) including doxycycline (antibacterial, anti-inflammatory) and procaine (to mitigate hyperactive autonomic nerves and uncontrolled immunological responses), specifically designed to target ulcerative colitis in the colon. In silico ADME profiling indicated unfavorable physicochemical properties for intestinal absorption, attributable to the compound’s high molecular weight, hydrophilic log P, and elevated topological polar surface area (TPSA). Molecular docking studies revealed high binding affinity to azoreductase, with a docking score superior to that of sulfasalazine, suggesting efficient colonic bioactivation. Dox-proc was synthesized with a good yield (75 %) and purity (m.p: 258–260°C); The structure was validated by FTIR and ¹H-NMR spectroscopy. Pharmacokinetic study demonstrated low intestinal absorption potential (experimental log P = –1.65), dox-proc is high stabile in acidic/basic conditions. Fecal hydrolysis studies verified efficient enzymatic activation in the presence of azoreductase, with hydrolysis up to 81%. Biological studies show that dox-proc group had the lowest score (1.2 ± 0.310) in clinical activity score system, this indicated that dox-proc has the highest clinical activity. Histopathological study confirms a good sign of mucosal healing. These findings align with the in silico predictions and validate the compound's applicability as a colon-targeted therapy.