Poly(3-Hydroxybutyrate), P(3HB) production and its biomedical applications

In this project enhancement of poly(3-hydroxybutyrate), P(3HB) production using a Gram positive bacteria, Bacillus cereus SPV and sucrose as the main carbon source was successfully achieved. Different modes of fermentation including shaken flask, batch, fedbatch and two-stage fermentation were investigated in the study. A modified G-medium was formulated and used throughout the study. Potassium and sulphate were identified as the main limiting factor for P(3HB) accumulation in Bacillus cereus SPV. By limiting the potassium phosphate concentration to 0.5 g/L K2HPO4 in the production medium, the dry cell weight, P(3HB) yield and P(3HB) concentration improved to, 7.21 g/L, 82 % dcw and 5.95 g/L respectively (i.e. 236, 115.8 and 830 % increase in dry cell weight, P(3HB) yield and P(3HB) concentration respectively). In addition, economic production of P(3HB) using agricultural/industrial waste (molasses) as the main carbon substrate was achieved. The study was also carried out in both shaken flask and 2L fermenter. The maximum P(3HB) yield achieved was 61.07 % dcw in 1L shaken flask and 51.37 % dcw in 2L fermenter. A novel wet cell PHA extraction was successfully developed in this project leading to high purity of the PHA produced with reduced crystallinity and efficient recovery. This is expected to save time, cost and enhanced continuous PHA production. Furthermore, a novel inexpensive and sustainable ‘compression moulding/particulate leaching’ technique for tissue engineering scaffold fabrication was developed. The novel technique enabled the production of biodegradable composite scaffolds of P(3HB)/microfibrillated cellulose and magnetic P(3HB) nanocomposite for possible applications in cartilage and bone tissue engineering, respectively. Detailed studies on the 2D and 3D composites showed that the inclusion of microfibrillated cellulose into the P(3HB) matrix enhanced the mechanical properties, hydrophilicity, introduced microtopography features, enhanced surface chemistry and biocompatibility of the composite material while inclusion of magnetic nanoparticles and ferrofluid, in addition to the above features added magnetic properties and microhardness to the composite materials. A unique controlled drug delivery system was developed with potential application in multiple drug delivery. The release study confirmed that the delivery system was able to control the release of BSA, a model protein. Finally, composite magnetic microspheres were also produced and characterised for their efficient use in the delivery of cancer drug and analyses performed showed that the composite constructs have superparamagnetic properties which would be useful for targeted delivery

P(3HB) Based Magnetic Nanocomposites: Smart Materials for Bone Tissue Engineering

The objective of this work was to investigate the potential application of Poly(3-hydroxybutyrate)/magnetic nanoparticles, P(3HB)/MNP, and Poly(3-hydroxybutyrate)/ferrofluid (P(3HB)/FF) nanocomposites as a smart material for bone tissue repair. The composite films, produced using conventional solvent casting technique, exhibited a good uniform dispersion of magnetic nanoparticles and ferrofluid and their aggregates within the P(3HB) matrix. The result of the static test performed on the samples showed that there was a 277% and 327% increase in Young's modulus of the composite due to the incorporation of MNP and ferrofluid, respectively. The storage modulus of the P(3HB)MNP and P(3HB)/FF was found to have increased to 186% and 103%, respectively, when compared to neat P(3HB). The introduction of MNP and ferrofluid positively increased the crystallinity of the composite scaffolds which has been suggested to be useful in bone regeneration. The total amount of protein absorbed by the P(3HB)/MNP and P(3HB)/FF composite scaffolds also increased by 91% and 83%, respectively, with respect to neat P(3HB). Cell attachment and proliferation were found to be optimal on the P(HB)/MNP and P(3HB)/FF composites compared to the tissue culture plate (TCP) and neat P(3HB), indicating a highly compatible surface for the adhesion and proliferation of the MG-63 cells. Overall, this work confirmed the potential of using P(3HB)/MNP and P(3HB)/FF composite scaffolds in bone tissue engineering

Composite scaffolds for cartilage tissue engineering based on natural polymers of bacterial origin, thermoplastic poly(3-hydroxybutyrate) and micro-fibrillated bacterial cellulose

Cartilage tissue engineering is an emerging therapeutic strategy that aims to regenerate damaged cartilage caused by disease, trauma, ageing or developmental disorder. Since cartilage lacks regenerative capabilities, it is essential to develop approaches that deliver the appropriate cells, biomaterials and signalling factors to the defect site. Materials and fabrication technologies are therefore critically important for cartilage tissue engineering in designing temporary, artificial extracellular matrices (scaffolds), which support 3D cartilage formation. Hence, this work aimed to investigate the use of poly(3-hydroxybutyrate)/microfibrillated bacterial cellulose (P(3HB)/MFC) composites as 3D-scaffolds for potential application in cartilage tissue engineering. The compression moulding/particulate leaching technique employed in the study resulted in good dispersion and a strong adhesion between the MFC and the P(3HB) matrix. Furthermore, the composite scaffold produced displayed better mechanical properties than the neat P(3HB) scaffold. On addition of 10, 20, 30 and 40 wt% MFC to the P(3HB) matrix, the compressive modulus was found to have increased by 35%, 37%, 64% and 124%, while the compression yield strength increased by 95%, 97%, 98% and 102% respectively with respect to neat P(3HB). Both cell attachment and proliferation were found to be optimal on the polymer-based 3D composite scaffolds produced, indicating a non-toxic and highly compatible surface for the adhesion and proliferation of mouse chondrogenic ATDC5 cells. The large pores sizes (60 - 83 µm) in the 3D scaffold allowed infiltration and migration of ATDC5 cells deep into the porous network of the scaffold material. Overall this work confirmed the potential of P(3HB)/MFC composites as novel materials in cartilage tissue engineering

Production of polyhydroxyalkanoates: the future green materials of choice

Polyhydroxyalkanoates (PHAs) have recently been the focus of attention as a biodegradable and biocompatible substitute for conventional non degradable plastics. The cost of large-scale production of these polymers has inhibited its widespread use. Thus, economical, large-scale production of PHAs is currently being studied intensively. Various bacterial strains, either wild-type or recombinant have been utilized with a wide spectrum of utilizable carbon sources. New fermentation strategies have been developed for the efficient production of PHAs at high concentration and productivity. With the current advances, PHAs can now be produced to a concentration of 80 g L−1 with productivities greater than 4 g PHA L−1 h−1. These advances will further lower the production cost of PHAs and allow this family of polymers to become a leading biodegradable polymer in the near future. This review describes the properties of PHAs, their uses, the various attempts towards the production of PHAs, focusing on the utilization of cheap substrates and the development of different fermentation strategies for the production of these polymers, an essential step forward towards their widespread use. Copyright © 2010 Society of Chemical Industry