Optimizing PHB recovery from Cupriavidus necator under mild acidic and alkaline digestion conditions

Polyhydroxybutyrate (PHB), a biopolymer produced by microbes, has gained attention as a sustainable alternative to traditional fossil-based plastics. Although solvent extraction is commonly used for PHB recovery, its environmental and health risks underline the need for safer and greener methods. Acidic and alkaline digestion techniques have gained interest as affordable options, but most reported processes rely on harsh chemical and thermal conditions that can degrade the polymer and hinder scalability. This study applies Response Surface Methodology (RSM) with Central Composite Design (CCD) to optimize PHB recovery under mild digestion conditions using NaOH (0.02–0.2 M, 1–5 h, 30–70 °C) and H₂SO₄ (0.02–0.5 M, 1–5 h at 50 °C). For Cupriavidus necator H16 (∼40 wt% PHB), optimal acidic digestion (0.5 M H₂SO₄, 2 h) yielded ∼92 % recovery and 60 % purity, while optimized alkaline digestion (0.02 M NaOH, 1 h, 55 °C) achieved ∼82 % recovery and 75 % purity. Notably, higher PHB content (>65 wt%) under mild alkaline conditions enabled complete recovery (100 %) and > 97 % purity, with the same protocol also effective for Zobelella denitrificans. Compared to chloroform extraction (∼41 % recovery, ∼69 % purity), the mild digestion protocols substantially improved both yield and environmental profile. Structural and thermal characterization (FTIR, ¹H/¹³C MAS NMR, GPC, TG-DSC) confirmed minimal polymer degradation, highlighting the potential of these mild, scalable methods for biomedical and packaging applications.