Enhanced intracellular polyhydroxybutyrate accumulation by Cupriavidus necator using sunflower oil-based waste cooking oil

• SF-WCO serves as an effective carbon source for PHB production by C. necator. • Yeast extract with 2% SF-WCO under phosphorus limitation raised PHB accumulation to 93% • C. necator preferentially consumed linoleic and palmitic acids from SF-WCO. • FTIR, DSC, and NMR showed the extracted PHB matched commercial PHB properties. Polyhydroxybutyrate (PHB), a biodegradable polymer, offers a promising eco-friendly alternative to petroleum-based plastic. Cupriavidus necator is a well-known PHB producer that can accumulate about 80% of PHB of its dry cell weight. However, the high cost of carbon sources remains a major barrier to commercial-scale production. Waste cooking oil (WCO), being rich in carbon and abundantly available, represents a low-cost substrate for PHB biosynthesis. In this study, C. necator DSM428 was used to convert sunflower oil-based waste cooking oil (SF-WCO) to PHB, and process optimization was performed by examining the effects of phosphorus limitation, SF-WCO concentration (2–10%), nitrogen source (ammonium sulfate, sodium nitrate, ammonium nitrate, ammonium chloride, peptone, urea, and yeast extract), and incubation time (24–120 h) on PHB production. Results demonstrated that C. necator DSM428 could efficiently metabolize SF-WCO, achieving a PHB accumulation of 22.88% under initial conditions. Optimization revealed that the best conditions for PHB production were 0.04% of phosphorus, 2% SF-WCO, yeast extract as a nitrogen source, and 72 to 96 h of incubation. Under these conditions, PHB concentration reached 6.75 g/L, biomass 7.42 g/L, and the maximum PHB accumulation was 93%, the highest reported for this strain. Characterization analysis of the produced PHB showed similar properties to the standard PHB. Overall, the findings confirm that the SF-WCO is an effective substrate for PHB production, with potential for industrial-scale application following further optimization. Abbreviations: PHB, Polyhydroxybutyrate; WCO, Waste cooking oil; SF-WCO, Sunflower oil-based waste cooking oil; FAME, Fatty acid methyl ester; PV, Peroxide value; AV, Acid value; GC–MS, Gas Chromatography-Mass Spectrometry; FTIR, Fourier Transform Infrared Spectroscopy; NMR, Nuclear magnetic resonance; DSC, Differential Scanning Calorimetry.