Greenness-Assessed RP-HPLC Method for Budesonide Quantification in Layer-by-Layer Polymeric Nanoparticles: Validation, Stability, and Controlled Release in Colonic Delivery

Abstract

Robust and sustainable analytical tools are essential for the design, manufacture, and evaluation of advanced drug delivery systems. This study reports the first validated reversed-phase high-performance liquid chromatography (RP-HPLC) method for quantifying budesonide encapsulated within poly(lactic-co-glycolic acid) (PLGA)-based, layer-by-layer (LbL) coated nanoparticles intended for colonic delivery in inflammatory bowel disease (IBD). Using response surface methodology with a central composite design, optimal conditions were identified as acetonitrile:water (80:20, v/v) under isocratic mode, achieving complete separation within 5 min at a flow rate of 0.34 mL/min and detection at 244 nm. The method is buffer-free, rapid, and solvent-efficient, resulting in a favorable greenness profile, further confirmed by GAPI and NEMI assessments. Validation in line with ICH Q2(R2) demonstrated excellent linearity (R{superscript 2} > 0.999), precision (%RSD 75% degradation occurred in alkaline and oxidative environments. In vitro release profiling under simulated gastrointestinal conditions demonstrated sustained, pH-responsive release (20.6% over 48 h), consistent with colonic targeting. This validated, green, and stability-indicating method integrates controlled release assessment with comprehensive performance evaluation, providing a versatile platform for quality control of nanoparticulate drug delivery systems and supporting their progression from formulation development to clinical translation