Fenofibrate Colon-Targeted Nanosponges Development, Optimization, and In-Vivo Anti-Inflammatory Efficacy of Fenofibrate Nanosponges: A Box–Behnken Design Approach to Formulation, Pharmacokinetics, and Experimental Colitis

Abstract

Background: Fenofibrate, a PPAR-α agonist with anti-inflammatory, lipid-modulating and lipid-lowering properties, is limited clinically by poor aqueous solubility and low oral bioavailability, restricting its use for inflammatory bowel disease (IBD). This study aimed to develop and optimize fenofibrate nanosponge formulation to improve drug solubility, achieve sustained colonic delivery, and enhance pharmacokinetic performance, and subsequently to evaluate the in-vivo anti-inflammatory and lipid-modulating efficacy of this optimized platform in dextran sulfate sodium (DSS)-induced experimental colitis.
Methods: Fenofibrate-loaded β-cyclodextrin nanosponges were prepared by the solvent evaporation method using diphenyl carbonate as crosslinker. A three-factor, three-level Box–Behnken design (BBD) was employed to optimize polymer concentration, crosslinker:polymer ratio, and stirring time, using particle size, entrapment efficiency, drug release, and zeta potential as responses. The optimized formulation was characterized by FTIR, DSC, XRD and scanning electron microscopy, and evaluated for in-vitro release and release kinetics, in-vivo pharmacokinetics in Wistar rats, and accelerated/long-term stability. The same optimized formulation was then evaluated for in-vivo anti-inflammatory and lipid-modulating efficacy in a DSS-induced colitis model in male BALB/c mice (normal control, DSS control, fenofibrate suspension 100 mg/kg, and fenofibrate nanosponge 100 mg/kg groups, treated on days 8–17), assessing serum lipid parameters, colonic inflammatory cytokines (TNF-α, IL-1β, IL-6, IL-10), and colonic histopathology, with the in-vivo design and reporting aligned with the ARRIVE 2.0 guidelines.
Results: The quadratic models for all four formulation responses were statistically significant (p ≤ 0.0004), with the crosslinker:polymer ratio identified as the predominant factor. The optimized formulation showed a particle size of 183.42 nm, entrapment efficiency of 89.76%, drug release of 78.63% at 10 h, and zeta potential of −29.88 mV, with prediction errors below 1%. FTIR, DSC and XRD confirmed drug–polymer compatibility with reduced fenofibrate crystallinity, and SEM revealed a porous nanosponge architecture. The formulation exhibited pH-dependent sustained release (Korsmeyer–Peppas, Super Case II transport) and improved pharmacokinetic parameters (higher Cmax, AUC and mean residence time) relative to pure drug suspension, with no significant change in physicochemical properties over six months of stability testing. In the DSS-colitis model, DSS administration induced significant dyslipidemia and elevated the pro-inflammatory cytokines TNF-α, IL-1β and IL-6 while reducing IL-10, together with marked crypt distortion, epithelial erosion, and inflammatory infiltration on histopathology. Treatment with the fenofibrate nanosponge restored serum lipid parameters toward normal, significantly reduced TNF-α, IL-1β and IL-6, restored IL-10, and preserved colonic mucosal architecture with near-normal crypts and goblet cells, with effects comparable to or exceeding plain fenofibrate suspension and mesalamine.
Conclusion: A colon-targeted fenofibrate nanosponge formulation was successfully optimized using Box–Behnken design, demonstrating nanoscale particle size, high entrapment efficiency, pH-dependent sustained release, favorable stability, and improved oral bioavailability compared with pure fenofibrate. This optimized delivery platform translated into promising in-vivo anti-inflammatory and lipid-modulating efficacy in DSS-induced experimental colitis, supporting its therapeutic potential as a colon-targeted delivery system for inflammatory bowel disease.