Quality by Design-Driven Green RP-HPLC Method for QuantifyingSyringic Acid in Nanocarriers and Biological Fluids

Introduction: A Quality by Design (QbD) methodology was used to develop and optimize a sustainable reversed-phase high-performance liquid chromatography (RP-HPLC) method for the quantification of Syringic Acid (SA) in transferosomes and human plasma, with potential use in diabetic wound healing therapy. Methods: SA-loaded transferosomes were prepared via the ethanol injection method, using soyalecithin as the lipid and Tween 80 as the surfactant. Using a mobile phase of methanol and 0.1% formic acid (38:62 v/v) and UV detection at 272 nm, chromatographic separation was accomplished on a C18 column. The organic phase composition and flow rate were found to be crucial chromatographic variables, optimized using a central composite response surface design. Results: The optimized approach showed excellent sensitivity, repeatability, and selectivity with a 38% organic phase and a 1.0 mL/min flow rate. Excellent linearity (R2 = 0.999), precision, accuracy, and robustness were established by validation in accordance with ICH Q2 (R1 & R2) requirements. 1.22 μg/mL was the detection limit, while 3.71 μg/mL was the quantification limit. The method's eco-friendliness was verified through thorough evaluations of sustainability and greenness using ComplexGAPI, AGREE, the Analytical Eco-Scale, and the Blue Applicability Grade Index. Discussion: The developed approach aligns with green analytical chemistry principles and meets regulatory validation requirements. Its versatility and applicability to pharmacokinetic research are demonstrated by its effectiveness with plasma samples and nanoformulations. Additionally, the approach is a promising tool for the development and assessment of SA-based therapies for diabetic wound healing, owing to its low environmental impact and strong analytical performance. Conclusion: The therapeutic promise of SA in diabetic wound care is supported by this validated, environmentally friendly RP-HPLC technology, which provides a reliable, sustainable analytical platform for quantifying SA in complex matrices.