Optimisation of verapamil hydrochloride loaded polyhydroxyalkanoate nano and microparticles using response surface methodology

Polyhydroxyalkanoates (PHAs) are biodegradable polymers with significant potential for drug delivery. However, a challenge remains in their application for encapsulating highly water-soluble drugs. This study aims to develop and optimise PHA-based nanoparticles (NPs) and microparticles (MPs) for the delivery of verapamil hydrochloride (VRP·HCl) using a systematic statistical approach. A quaternary PHA copolymer, poly(3-hydroxybutyrate-co-4-hydroxybutyrate-co-5-hydroxyvalerate-co-3-hydroxyhexanoate) [P(3HB-co-4HB-co-5HV-co-3HHx)], was biosynthesised using genetically modified Cupriavidus necator. Polymer characterisation by Nuclear Magnetic Resonance (1H NMR), Gas Chromatography (GC), Gel Permeation Chromatography (GPC), and Limulus Amebocyte Lysate (LAL) assay confirmed a high-purity (97 ± 5 wt%) and endotoxin-free P(3HB-co-4HB-co-5HV-co-3HHx) copolymer with a monomer composition of 69 mol% 3HB, 14 mol% 4HB, 12 mol% 5HV, and 5 mol% 3HHx. VRP·HCl-loaded NPs and MPs were prepared via a double-emulsion solvent evaporation method and optimised using response surface methodology (RSM) based on a central composite design (CCD) to predict and evaluate the influence of polymer mass, drug mass, and stabiliser concentration on particle size, drug loading (DL), and encapsulation efficiency (EE). Particle size, polydispersity index (PDI), zeta potential (ZP), DL, and EE were evaluated experimentally. The optimised NP and MP formulations achieved mean particle sizes of 245.06 ± 0.01 nm and 2.23 ± 1.50 µm, with maximum EE of 38.95 ± 20.37% and 45.23 ± 2.85%, respectively. The corresponding DL values for NPs and MPs were 23.37 ± 12.22% and 18.09 ± 1.14%, respectively. Statistical analysis demonstrated good model predictability within the explored design space, enabling the identification of formulation regions governed primarily by polymer mass, drug distribution, and emulsion characteristics. In conclusion, this study demonstrates the feasibility of encapsulating a hydrophilic drug within a fully hydrophobic, biodegradable PHA matrix while establishing a systematic optimisation framework for the development of PHA-based NP and MP drug delivery systems.