Production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) using agro-industrial effluents with tunable proportion of 3-hydroxyvalerate monomer units.

With increasing concerns about future oil depletion and plastic pollution, bioplastics saw an increasing interest from scientists and industrials. Among bioplastics, the polyhydroxyalkanoates (PHA) are a promising family of polyester which are both biosourced and biodegradable. Biosynthesized by microorganisms, especially bacteria, control of their monomeric composition, and thus their thermal and mechanical properties, is still a challenge to really make tailor-made syntheses. Moreover, one way to decrease the high cost of production is to use waste as substrates for the microorganisms. In this study, a marine bacteria, Halomonas sp. SF2003 was grown on agro-industrial effluents as the sole carbon sources and was able to produce poly(3-hydroxybutyrate) (PHB) with a productivity of 1.3 g·L-1 in 40 h of culture and a number-average molar weight of 342,000 g·mol-1. With the addition of valeric acid in the substrates, poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBHV) with controlled proportion of hydroxyvalerate (HV) monomers were obtained. Their thermal and mechanical characteristics were investigated as a function of HV amount and showed a decrease of the glass transition and melting temperatures and in Young modulus with the HV content increase.

Photoinduced modification of the natural biopolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate) microfibrous surface with anthraquinone-derived dextran for biological applications.

A straightforward and versatile method for immobilizing polysaccharides on the surface of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBHV) electrospun fibers is developed with the objective of designing a new functional biomaterial having a significant effect on cell proliferation. The approach relies on a one-step procedure: UV grafting of a photosensitive dextran (AQ-Dext) on the surface of PHBHV fibers according to a "grafting onto" method, with the use of an anthraquinone derivative. The photografting is conducted through a photoinduced free radical process employing an anthraquinone-based photosensitizer in aqueous medium. Under appropriate conditions, AQ-Dext reacts with C-H σ-bonds of the polymer substrate (PHBHV) to produce a semianthraquinone radical according to an H-abstraction reaction. This radical recombines together with the alkylradical (R˙) formed at the surface of PHBHV fibers via the oxygen atom of the anthraquinone photolinker. The photochemical mechanism of the AQ-Dext photolysis is entirely described for the first time by an electron spin resonance technique and laser flash photolysis. The modified PHBHV microfibrous scaffolds are extensively characterized by water contact angle measurements, XPS analysis and atomic force microscopy, confirming the covalent grafting of dextran on PHBHV fibers. Finally, a primary investigation demonstrates that dextran modified PHBHV fibers are permissive for optimized cell colonization and proliferation. The cell morphologies are described by SEM micrographs, revealing a significant affinity and favorable interactions for adherence of human mesenchymal stem cells (hMSCs) on scaffolds provided by dextran chemical structure. Moreover, the proliferation rate of hMSCs increases on this new functionalized biomaterial associated with a higher extra-cellular matrix production after 5 days of culture in comparison with native PHBHV fibers.

Biosynthesis and characterization of polyhydroxyalkanoates by Pseudomonas guezennei from alkanoates and glucose.

The biosynthesis of medium chain length poly(3-hydroxyalkanoates) mcl PHAs by Pseudomonas guezennei using glucose, sodium octanoate, and 10-undecenoic acid as sole or mixed carbon sources was investigated. Chemical composition of polyesters was analyzed by GCMS and NMR. The copolyester produced by P. guezennei from glucose mainly consisted of 3-hydroxyoctanoate and 3-hydroxydecanoate, and the presence of 3-hydroxydodec-5-enoate was demonstrated. Using sodium octanoate as the sole nutrient, the microorganism produced a poly(3-hydroxyoctanoate) (PHO) polymer containing up to 94 mol% 3-hydroxyoctanoate. Biosynthesis of poly[(3-hydroxyoctanoate)-co-(3-hydroxyundecenoate)] (PHOU) copolymers bearing terminal reactive double bonds on its side chains with unsaturation degree ranging from 8.8% to 78.2% was obtained by tuning the ratio of sodium octanoate/10-undecenoic acid in the medium. Thermal analysis indicated semi-crystalline polymers with melting temperatures (T(m)) ranging from 46 to 55°C, fusion enthalpy (ΔH) comprised between 3 and 35 J/g and glass transition temperature (T(g)) from -36 to -44°C, except for the highly amorphous 78.2% unsaturated PHOU with a low T(g) (-50°C). Molecular weights determined by GPC ranged from 119000 and 530000 g/mol. The biosynthesis of natural polyesters with controlled ratio of vinyl-terminated side chains is of great interest for further chemical modifications.

Development of a three-steps derivatization assay for the localization of double bond in monounsaturated monomers of poly-beta-hydroxyalkanoates by GC-MS.

A new gas chromatography-mass spectrometry (GC-MS) method for the localization of double bond in monounsaturated 3-hydroxyalkenoic acids monomers has been developed. A three steps derivation assay was used including a methanolysis, then acetylation and dimethyldisulfide (DMDS) addition to alkene groups. Electron impact GC-MS analysis of such derivatives offers characteristic fragments allowing the unambiguous determination of double bond position in side chain. This novel method is well-suited for the routine analysis of poly-beta-hydroxyalkanoates (PHAs), and was used to characterize monounsaturated monomers in both 3-hydroxyalkenoic acids standards as well as in mcl-PHAs and poly(3-hydroxyoctanoate-co-3-hydroxyundecenoate) (PHOU) produced by bacterial strain Pseudomonas guezennei from glucose or a mixture of sodium octanoate plus 10-undecenoic acid, respectively.