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Polyhydroxyalkanoates (PHAs) signify the most promising biological substitute to petrochemical plastics. Renewable and inexpensive agro-industrial by-products can be used as potent fermentation feedstocks for sustainable PHA biosynthesis. This study aimed at using a wild type B. megaterium strain Ti3 innate hydrolytic enzyme/s for eco-friendly valorization of 16 lignocellulosic agrowastes to PHA without pretreatments. Initial hydrolytic screening PHA concentration of (0.04–0.17 g/L), highlighted the strain's metabolic versatility. Pareto ranking of Taguchi orthogonal array (TOA) established ragi husk (RH), sesame oil cake (SOC) and KH2PO4 as the most influential factors (p

Petri nets (PNs) have been widely utilized for modelling and analysing different expert systems which are characterized by concurrency, parallelism and conflicts. Due to the complex and dense nature of metabolic networks, modelling of these systems is needed to understand their topology. Petri net is one of the extensively used mathematical tools for modelling and studying metabolic pathways. Here, we have explained how analysis techniques of PNs can be exploited to simulate, validate and study the behaviour of metabolic pathways. We considered the biosynthesis of polyhydroxyalkanoates (PHAs) in the presence of krebs cycle for illustration to explain the validation and analysis process using Petri net. PHAs are synthesized by bacteria during nutrient lack conditions and are stored as energy storage materials. Krebs cycle is the competing pathway when glucose is the main source of carbon in PHA biosynthesis. The obtained results provide a valid mathematical model for confirming some of the known properties about their metabolic pathway and also provide some results on how the system will behave in the presence of krebs cycle. The model can be used in further studies to get new insights in this pathway along with other competing pathways. Analysis of the metabolic pathways can be more useful if this Petri net approach is related with some experimental approach.

This chapter gives an overview of the key elements that need to be considered for understanding and triggering the microstructure of polyhydroxyalkanoate (PHA) copolymers. This family of microbial biopolyesters offers a large potential considering its broad spectrum of possible material properties due to the diversity of comonomers. It is, therefore, of great interest to study and understand the influences on the molecular composition and composition distribution to valorize this potential. PHA biosynthesis is described considering substrates, microbial producers, the polymerizing enzyme class, and its spatial structure, as well as the kinetics of the PHA synthesis. In the second part of this chapter, a summary of in vitro approaches to synthesize sequence controlled PHA (e.g., block copolymers) is given to emphasize the importance of the molecular architecture. Furthermore, existing approaches for sequence controlled PHA syntheses in living microorganisms are described considering feed strategies, genetic modifications, and artificial genetic networks.

The aim of the current review is to analyze trends in development of an efficient technology for polyhydroxyalkanoate (PHA) biomanufacture highlighting the up-to-date progress on PHA biosynthesis and focusing on the downstream processing. Three main production pathways were identified: through microbial, enzymic, or plant routes. Microbial fermentation processes were predominant, with a wide range of microorganisms, starting materials and culture conditions reported. Largely, two schemes for recovering PHAs from the reaction medium post fermentation were identified: dissolving biomass to separate PHAs granules with strong oxidants, and extracting PHAs directly from the biomass using suitable solvents. For the valuable industrial scale biosynthesis of PHA several technological elements need to be applied such as robust whole-cell microbial catalyst with its optimal culturing conditions, suitable carbon source, proper mode of process operation, as well as economical and ecological purification methods.

Competitive polyhydroxyalkanoate (PHAs) production requires progress in microbial strain performance, feedstock selection, downstream processing, and more importantly according to the process design with process kinetics of the microbial growth phase and the phase of product formation. The multistage continuous production in a bioreactor cascade was described for the first time in a continuously operated, flexible five-stage bioreactor cascade that mimics the characteristics involved in the engineering process of tubular plug flow reactors. This process was developed and used for Cupriavidus necator-mediated PHA production at high volumetric and specific PHA productivity (up to 2.31 g/(Lh) and 0.105 g/(gh), respectively). Based on the experimental data, formal kinetic and high structured kinetic models were established, accompanied by footprint area analysis of binary imaged cells. As a result of the study, there has been an enhanced understanding of the long-term continuous PHA production under balanced, transient, and nutrient-deficient conditions that was achieved on both the micro and the macro kinetic level. It can also be concluded that there were novel insights into the complex metabolic occurrences that developed during the multistage- continuous production of PHA as a secondary metabolite. This development was essential in paving the way for further process improvement. At the same time, a new method of specific growth rate and specific production rate based on footprint area analysis was established by using the electron microscope.

The Swiss Priority Program in Biotechnology of the Swiss National Science Foundation that lasted between 1992 and 2001 had a boosting effect on many biotech disciplines and on the developments of polyhydroxyalkanoates (PHAs) in Switzerland in particular. The funding organization led by Prof. Oreste Ghisalba enabled a better understanding of the PHA biosynthesis and the development, as well as the implementation of novel bioprocesses (e.g. two-phase fermentations, multiple nutrient limited growth conditions, multi-stage chemostats, and product formation in different host organisms). However, production of PHA in Switzerland appeared to be impossible for cost reasons due to the strong competition from cheaper, petrol-based plastics. The recent reports on environmental issues with non-degradable plastics has triggered a general change in the perception of biodegradable plastics, giving them an added value and thus justifying a higher price. Ongoing research focuses on the sustainable production of PHAs using carbon waste streams, synthesis gas or even CO2

Together with other so-called "bio-plastics", Polyhydroxyalkanoates (PHAs) are expected to soon replace established polymers on the plastic market. As a prerequisite, optimized process design is needed to make PHAs attractive in terms of costs and quality. Nowadays, large-scale PHA production relies on discontinuous fed-batch cultivation in huge bioreactors. Such processes presuppose numerous shortcomings such as nonproductive time for reactor revamping, irregular product quality, limited possibility for supply of certain carbon substrates, and, most of all, insufficient productivity. Therefore, single- and multistage continuous PHA biosynthesis is increasingly investigated for production of different types of microbial PHAs; this goes for rather crystalline, thermoplastic PHA homopolyesters as well as for highly flexible PHA copolyesters, and even blocky-structured PHAs consisting of alternating soft and hard segments. Apart from enhanced productivity and constant product quality, chemostat processes can be used to elucidate kinetics of cell growth and PHA formation under constant process conditions. Furthermore, continuous enrichment processes constitute a tool to isolate novel powerful PHA-producing microbial strains adapted to special environmental conditions. The article discusses challenges, potential and case studies for continuous PHA production, and shows up new strategies to further enhance such processes economically by developing unsterile open continuous processes combined with the application of inexpensive carbon feedstocks.

The hazardous effects of synthetic polymers encouraged investigators to find about biodegradable polymers among which microbial derived polyhydroxyalkanoate (PHA) have got attention during the two last decades. In the present study, isolation and identification of a PHA producing bacterial strain and optimization of cultural conditions for PHA biosynthesis were aimed. Several soil and spring samples were collected and screened for PHA producing bacterial strain according to the reported protocols. The most efficient isolate was identified using biochemical, morphological, and 16S rDNA gene sequencing methods. The related PHA production was then optimized using D-optimal statistical design. The obtained results of screening step using Nile blue containing culture plate together with biochemical and morphological studies as well as 16S rDNA gene analysis introduced Pseudomonas pseudoalcaligenes strain Te as the most efficient PHA producer. Evaluation of PHA productivity profile of P. pseudoalcaligenes strain Te revealed that maximum PHA biosynthesis was achieved at the stationary phase of growth. Assessment the influence of five parameters on PHA productivity of the selected strain revealed the significant effect of three factors including pH, K2HPO4, and temperature. The optimum level of these three factors (pH 8.6; K2HPO4, 4.7 g/L; and temperature of 25 °C) enhanced the PHA production to 5412.9 mM. Although, the capability of P. pseudoalcaligenes for PHA production was confirmed in the present study, it merits further investigations to find about the related pathways involved in PHA biosynthesis by this strain.

Microbially produced polyhydroxyalkanoates (PHAs) are fully biodegradable biopolyesters that have attracted much attention recently as alternative polymeric materials that can be produced from biorenewable and biowaste resources. The properties of these biological polymers are affected by the same fundamental principles as those of fossil-fuel derived polyolefins, with a broad range of compositions available based on the incorporation of different monomers into the PHA polymer structure, and with this broad range tailoring subsequent properties. This review comprehensively covers current understanding with respect to PHA biosynthesis and crystallinity, and the effect of composition, microstructure and supramacromolecular structures on chemomechanical properties. While polymer composition and microstructure are shown to affect these properties, the review also finds that a key driver for determining polymer performance properties is compositional distribution. From this review it follows that PHA–PHA blend compositions are industrially important, and the performance properties of such blends are discussed. A particular need is identified for further research into the effect of chemical compositional distribution on macromolecular structure and end-use properties, advanced modeling of the PHA accumulation process and chain growth kinetics for better process control.

Cupriavidus sp. USMAA1020, USMAA2-4, and USMAHM13 are capable of producing polyhydroxyalkanoate (PHA). This biopolymer is an alternative solution to synthetic plastics, whereby polyhydroxyalkanoate synthase is the key enzyme involved in PHA biosynthesis. Here, we report the complete genomes of three Cupriavidus sp. strains: USMAA1020, USMAA2-4, and USMAHM13.

Traditional plastics pose a serious threat to the environment due to their non-degradable nature. As an alternative, a family of naturally occurring biodegradable polymer known as Polyhydroxyalkanoates (PHAs) have been produced by a range of bacterial species. These PHAs are polyesters of R-hydroxyalkanoic acids and exhibit a wide range of properties depending on the number of carbon atoms in their monomer units. This book chapter discusses the discovery, occurrence, and mechanisms of PHA biosynthesis in bacteria. Additionally, the wide-ranging potential of PHAs and their future trends have been discussed.

A novel description of mcl -PHA biosynthesis by Ps. chlororaphis from tallow-based biodiesel as an inexpensive carbon feed stock is presented. Fermentation protocols, kinetic analysis, an efficient product recovery strategy, and product characterization are included. Maximum specific growth rates (μ max. ) of 0.08 h −1 , 0.10 h −1 and 0.13 h −1 , respectively, were achieved in three different fermentation set-ups. Volumetric productivity for mcl -PHA amounted to 0.071 g/L h, 0.094 g/L h and 0.138 g/L h, final intracellular PHA contents calculated from the sum of active biomass and PHA from 22.1 to 29.4 wt.-%, respectively. GC-FID analysis showed that the obtained biopolyester predominantly consists of 3-hydroxyoctanoate and 3-hydroxydecanoate, and, to a minor extent, 3-hydroxydodecanoate, 3-hydroxynonanoate, 3-hydroxyhexanoate, and 3-hydroxyheptanoate monomers. The overall distribution of the monomers remained similar, regardless to working volumes, biodiesel concentrations and pre-treatment of the inoculum.

This paper deals with the microbial production of polyhydroxyalkanoates (PHAs), biodegradable thermoplastics which perform excellently as a material, from inexpensive renewable carbon sources. To date, with the help of genetic engineering techniques, it has become possible to design several types of PHAs with different compositions and to enhance the productivities of PHAs. In addition, molecular breeding of PHA biosynthesis enzymes has been demonstrated to improve polymer production. Mutant PHA synthases generated by an in vitro evolution technique have allowed the enhanced production and quality alteration of PHAs. Furthermore, use of inexpensive renewable carbon sources, such as plant oils, waste materials, and carbon dioxide, would be a key for a reduction in PHA production cost.

Bacterial poly(hydroxyalkanoate)s (PHA) are a complex class of polyesters, and almost one hundred different constituents are currently known. Unfortunately, most of these constituents only occur in the polyesters if specific precursor substrates are provided as carbon source. Recently, however, bacteria and mutants were isolated, which synthesize PHA exhibiting an interesting composition from simple unrelated substrates obtainable from renewable resources provided by the agriculture. This contribution will provide an example how PHA based on 4-hydroxyvaleric acid can be obtained from bulk chemicals. In addition, some aspects of our current knowledge on the physiological and molecular basis of PHA biosynthesis will be provided.

This paper studies the effect of organic load rate (OLR) and nutrient ratio on operation stability of the moving bed bioreactor (MBBR) for kraft mill wastewater treatment, analyzing the incidence of polyhydroxyalkanoate (PHA) production. The MBBR operating strategy was to increase OLR from 0.25 ± 0.05 to 2.41 ± 0.19 kg COD m−3 d−1 between phases I and IV. The BOD5:N:P ratio (100:5:1 and 100:1:0.2) was evaluated as an operation strategy for phases IV to V. A stable MBBR operation was found when the OLR was increased during 225 days in five phases. The maximum absolute fluorescence against the proportion of cells accumulating PHA was obtained for an OLR of 2.41 ± 0.19 kg COD m−3d−1 and a BOD5:N:P relationship of 100:1:0.2. The increase of PHA biosynthesis is due to the increased OLR and is not attributable to the increased cell concentration, which is maintained constant in stationary status during bioreactor biosynthesis.

Polyhydroxyalkanoates (PHAs) are natural polyesters that accumulate in numerous microorganisms as a carbon- and energy-storage material under the nutrient-limiting condition in the presence of an excess carbon source. PHAs are considered to be one of the potential alternatives to petrochemically derived plastics owing to their versatile material properties. Over the past few decades, extensive detailed biochemical, molecular-biological, and metabolic studies related to PHA biosynthesis have been carried out. Advances in our knowledge of PHA biosynthesis led to the development of engineered strains and fermentation processes for the production of PHAs with high efficiency. Even though the traditional metabolic engineering based on our rational thinking and trial-and-error type approaches allowed development of improved strains, further enhancement in the performance is expected through systems metabolic engineering, which is metabolic engineering integrated with systems-biological approaches. In this chapter, the strategies taken for the metabolic engineering of PHA producers are briefly reviewed. Then, genomic and proteomic studies performed to understand the PHA biosynthesis in the context of whole cell metabolism as well as to develop further engineered strains are reviewed. Finally, the strategies for the systems metabolic engineering of PHA producers are suggested; these will make it possible to produce PHAs with higher efficiencies and to develop tailor-made PHAs by systems-level optimization of the metabolic network and establishment of novel pathways.

Bacteria are able to synthesize a wide variety of different thermoplastic polyhydroxyalkanoic acids (PHA). The physiology of PHA metabolism has been studied in most detail in the aerobic hydrogen bacterium Alcaligenes eutrophus. Strains of this bacterium are already used for the production of biodegradable polyesters on a technological scale. The current knowledge on the pathways for PHA synthesis, on the genes essential for PHA biosynthesis and on genes affecting the accumulation of PHA in this bacterium will be reviewed.

Introduction to PHAs Diversity of PHAs Biosynthesis of PHAs Short-chain-length PHAs Medium-chain-length PHAs Organization of Genes Required for PHA Biosynthesis Degradation of Poly(3HB) Intracellular Degradation of Poly(3HB) Properties of Native Poly(3HB) Granules Extracellular Degradation of PHA PHA Granule-associated Proteins Conversion of 3HB to Acetyl-CoA Regulation of PHA metabolism Keywords: polyhydroxyalkanoic (PHA); poly(3-hydroxybutyric acid) (poly(3HB)); PHA metabolism; PHA biosynthesis; gene organization; intracellular poly(3HB) degradation; extracellular PHA degradation; regulation of PHA metabolism; native poly(3HB) granules

Polyhydroxyalkanoic acids (PHA) represent a storage compound for carbon and/or energy and occur as insoluble inclusions in the cells of a wide range of different bacteria (Steinbuchel 1991) mostly during cultivation of the cells in the presence of an excess of carbon source and if one other essential nutrient limits growth (Anderson, Dawes 1990). In addition to poly(3-hydroxybutyric acid), poly(3HB), which is the most well-known representative of PHA, 90 different hydroxyalkanoic acids have been detected as constituents of PHA from bacterial origin (Steinbuchel, Valentin 1995). These polyesters have been attracted much attention during the last years since they are biodegradable thermoplastics and/or elastomers which exhibit many other interesting properties and which therefore lend themselves for a wide range of different technical applications (Hocking, Marchessault 1994).