Exploring Lemon Industry By-Products for Polyhydroxyalkanoate Production: Comparative Performances of Haloferax mediterranei PHBV vs. Commercial PHBV

This study investigates the valorisation of lemon industry by-products as carbon sources to produce poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) using the halophilic archaeon Haloferax mediterranei. The resulting polymer (HFX PHBV) was supplemented with nucleating agents (orotic acid, boron nitride, and theobromine) and compared with a commercial PHBV grade (Enmat Y1000) under identical conditions. Fermentation strategies were optimised by varying the lemon by-product concentration, inoculum size, and nutrient stoichiometry (C:N:P ratios), followed by scaleup in a 2 L bioreactor. A 11% (v/v) lemon by-product combined with a 5% (v/v) inoculum yielded the highest productivity under minimal medium conditions (2.127 g/L PHBV), while enriched media further enhanced the polymer accumulation (up to 3.250 g/L PHBV). A comparative characterisation of HFX PHBV and Enmat Y1000, using NMR, TGA, MFR, DSC, Raman spectroscopy, XRD, and DMA, revealed that HFX PHBV exhibited lower crystallinity, increased flexibility, and a high hydroxyvalerate content (27.4%), which conferred improved ductility. Investigation of nucleating agents demonstrated that orotic acid was the most effective at enhancing the crystallisation kinetics. Overall, this study demonstrates an efficient PHBV production process based on waste valorisation, yielding a biopolymer with competitive physicochemical properties relative to a commercial standard, and provides integrated solutions to the global challenges of plastic pollution and food waste.