Special Issue "Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2"

A special issue of Bioengineering (ISSN 2306-5354).

Deadline for manuscript submissions: closed (31 October 2019) | Viewed by 78833

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Special Issue Editor

Office of Research Management and Service, c/o Institute of Chemistry, University of Graz, 8010 Graz, Austria
Interests: polyhydroxyalkanoates (PHA); biopolymers; fermentation technology; downstream processing; sustainable process development
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Special Issue Information

Dear Colleagues,

Nowadays, we are witnessing highly dynamic research activities related to the intriguing field of biodegradable materials with plastic-like properties. These activities are provoked by a strengthened public awareness of prevailing ecological issues connected to growing piles of plastic waste and increasing greenhouse gas emissions; this goes hand-in-hand with the ongoing depletion of fossil feedstocks, which are traditionally used to produce full carbon backbone polymers. To a steadily increasing extend, polyhydroxyalkanoate (PHA) biopolyesters, a family of plastic-like materials with versatile material properties, are considered a future-oriented solution for diminishing these concerns. PHA production is based on renewable resources, and occurs in a bio-mediated fashion by the action of living organisms. If accomplished in an optimized way, PHA production and the entire PHA lifecycle are embedded into nature´s closed cycles of carbon.

You as a highly recognized expert in this field are aware of the fact that sustainable and efficient PHA production requires the understanding and improvement of all individual process steps. Holistic improvement of PHA production, applicable on an industrially relevant scale, calls for inter alia: consolidated knowledge about the enzymatic and genetic particularities of PHA accumulating organisms, in-depth understanding of the kinetics of the bioprocess, the selection of appropriate inexpensive fermentation feedstocks, tailoring the composition of PHA on the level of the monomeric constituents, optimized biotechnological engineering, and novel strategies for PHA recovery from biomass characterized by minor energy and chemical requirement.

In order to provide a comprehensive compilation of articles addressing all these individual aspects, we are contacting globally recognized experts like you. We are convinced that a contribution based on your special expertise in the PHA arena will be of major benefit to make this Special Issue even more attractive to the scientific community.

Dr. Martin Koller
Guest Editor

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Keywords

  • Polyhydroxyalkanoate (PHA)
  • Biopolyesters
  • Bioprocess Design
  • Sustainable Development

Published Papers (12 papers)

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Editorial

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Editorial
Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2
Bioengineering 2020, 7(1), 24; https://doi.org/10.3390/bioengineering7010024 - 04 Mar 2020
Cited by 12 | Viewed by 4869
Abstract
During the two years that have passed since the first volume of “Advances in Polyhydroxyalkanoate (PHA) production” was published, the progress in PHA-related research was indeed tremendous, calling for the next, highly bioprocess- and bioengineering-oriented volume. This editorial paper summarizes and puts into [...] Read more.
During the two years that have passed since the first volume of “Advances in Polyhydroxyalkanoate (PHA) production” was published, the progress in PHA-related research was indeed tremendous, calling for the next, highly bioprocess- and bioengineering-oriented volume. This editorial paper summarizes and puts into context the contributions to this second volume of the Bioengineering Special Issue; it covers highly topical fields of PHA-related R&D activities, covering, beside the pronounced bioengineering-related articles, the fields of the microbiology of underexplored, but probably emerging, PHA production strains from the groups of Pseudomonas, cyanobacteria, methanotrophs, and from the extremophilic domain of haloarchaea. Moreover, novel second-generation lignocellulose feedstocks for PHA production from agriculture to be used in biorefinery concepts, new approaches for fine-tuning the composition of PHA co- and terpolyesters, process simulation for PHA production from methane-rich natural gas, the challenges associated with rheology-governed oxygen transfer in high cell density cultivations, rapid spectroscopic in-line analytics for process monitoring, and the biomedical application of PHA biopolyesters after appropriate advanced processing are the subjects of the presented studies. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)

Research

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Article
Tequila Agave Bagasse Hydrolysate for the Production of Polyhydroxybutyrate by Burkholderia sacchari
Bioengineering 2019, 6(4), 115; https://doi.org/10.3390/bioengineering6040115 - 17 Dec 2019
Cited by 19 | Viewed by 5081
Abstract
Tequila agave bagasse (TAB) is the fibrous waste from the Tequila production process. It is generated in large amounts and its disposal is an environmental problem. Its use as a source of fermentable sugars for biotechnological processes is of interest; thus, it was [...] Read more.
Tequila agave bagasse (TAB) is the fibrous waste from the Tequila production process. It is generated in large amounts and its disposal is an environmental problem. Its use as a source of fermentable sugars for biotechnological processes is of interest; thus, it was investigated for the production of polyhydroxybutyrate (PHB) by the xylose-assimilating bacteria Burkholderia sacchari. First, it was chemically hydrolyzed, yielding 20.6 g·L−1 of reducing sugars, with xylose and glucose as the main components (7:3 ratio). Next, the effect of hydrolysis by-products on B. sacchari growth was evaluated. Phenolic compounds showed the highest toxicity (> 60% of growth inhibition). Then, detoxification methods (resins, activated charcoal, laccases) were tested to remove the growth inhibitory compounds from the TAB hydrolysate (TABH). The highest removal percentage (92%) was achieved using activated charcoal (50 g·L−1, pH 2, 4 h). Finally, detoxified TABH was used as the carbon source for the production of PHB in a two-step batch culture, reaching a biomass production of 11.3 g·L−1 and a PHB accumulation of 24 g PHB g−1 dry cell (after 122 h of culture). The polymer structure resulted in a homopolymer of 3-hydroxybutyric acid. It is concluded that the TAB could be hydrolyzed and valorized as a carbon source for producing PHB. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Article
Rheological Behavior of High Cell Density Pseudomonas putida LS46 Cultures during Production of Medium Chain Length Polyhydroxyalkanoate (PHA) Polymers
Bioengineering 2019, 6(4), 93; https://doi.org/10.3390/bioengineering6040093 - 09 Oct 2019
Cited by 3 | Viewed by 5342
Abstract
The rheology of high-cell density (HCD) cultures is an important parameter for its impact on mixing and sparging, process scale-up, and downstream unit operations in bioprocess development. In this work, time-dependent rheological properties of HCD Pseudomonas putida LS46 cultures were monitored for microbial [...] Read more.
The rheology of high-cell density (HCD) cultures is an important parameter for its impact on mixing and sparging, process scale-up, and downstream unit operations in bioprocess development. In this work, time-dependent rheological properties of HCD Pseudomonas putida LS46 cultures were monitored for microbial polyhydroxyalkanoate (PHA) production. As the cell density of the fed-batch cultivation increased (0 to 25 g·L−1 cell dry mass, CDM), the apparent viscosity increased nearly nine-fold throughout the fed-batch process. The medium behaved as a nearly Newtonian fluid at lower cell densities, and became increasingly shear-thinning as the cell density increased. However, shear-thickening behavior was observed at shearing rates of approximately 75 rad·s−1 or higher, and its onset increased with viscosity of the sample. The supernatant, which contained up to 9 g·L−1 soluble organic material, contributed more to the observed viscosity effect than did the presence of cells. Owing to this behavior, the oxygen transfer performance of the bioreactor, for otherwise constant operating conditions, was reduced by 50% over the cultivation time. This study has shown that the dynamic rheology of HCD cultures is an important engineering parameter that may impact the final outcome in PHA cultivations. Understanding and anticipating this behavior and its biochemical origins could be important for improving overall productivity, yield, process scalability, and the efficacy of downstream processing unit operations. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Article
Development of High Cell Density Cultivation Strategies for Improved Medium Chain Length Polyhydroxyalkanoate Productivity Using Pseudomonas putida LS46
Bioengineering 2019, 6(4), 89; https://doi.org/10.3390/bioengineering6040089 - 26 Sep 2019
Cited by 15 | Viewed by 5215
Abstract
High cell density (HCD) fed-batch cultures are widely perceived as a requisite for high-productivity polyhydroxyalkanoate (PHA) cultivation processes. In this work, a reactive pulse feed strategy (based on real-time CO2 or dissolved oxygen (DO) measurements as feedback variables) was used to control [...] Read more.
High cell density (HCD) fed-batch cultures are widely perceived as a requisite for high-productivity polyhydroxyalkanoate (PHA) cultivation processes. In this work, a reactive pulse feed strategy (based on real-time CO2 or dissolved oxygen (DO) measurements as feedback variables) was used to control an oxygen-limited fed-batch process for improved productivity of medium chain length (mcl-) PHAs synthesized by Pseudomonas putida LS46. Despite the onset of oxygen limitation half-way through the process (14 h post inoculation), 28.8 ± 3.9 g L−1 total biomass (with PHA content up to 61 ± 8% cell dry mass) was reliably achieved within 27 h using octanoic acid as the carbon source in a bench-scale (7 L) bioreactor operated under atmospheric conditions. This resulted in a final volumetric productivity of 0.66 ± 0.14 g L−1 h−1. Delivering carbon to the bioreactor as a continuous drip feed process (a proactive feeding strategy compared to pulse feeding) made little difference on the final volumetric productivity of 0.60 ± 0.04 g L−1 h−1. However, the drip feed strategy favored production of non-PHA residual biomass during the growth phase, while pulse feeding favored a higher rate of mcl-PHA synthesis and yield during the storage phase. Overall, it was shown that the inherent O2-limitation brought about by HCD cultures can be used as a simple and effective control strategy for mcl-PHA synthesis from fatty acids. Furthermore, the pulse feed strategy appears to be a relatively easy and reliable method for rapid optimization of fed-batch processes, particularly when using toxic substrates like octanoic acid. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Article
In-Line Monitoring of Polyhydroxyalkanoate (PHA) Production during High-Cell-Density Plant Oil Cultivations Using Photon Density Wave Spectroscopy
Bioengineering 2019, 6(3), 85; https://doi.org/10.3390/bioengineering6030085 - 19 Sep 2019
Cited by 18 | Viewed by 7990
Abstract
Polyhydroxyalkanoates (PHAs) are biodegradable plastic-like materials with versatile properties. Plant oils are excellent carbon sources for a cost-effective PHA production, due to their high carbon content, large availability, and comparatively low prices. Additionally, efficient process development and control is required for competitive PHA [...] Read more.
Polyhydroxyalkanoates (PHAs) are biodegradable plastic-like materials with versatile properties. Plant oils are excellent carbon sources for a cost-effective PHA production, due to their high carbon content, large availability, and comparatively low prices. Additionally, efficient process development and control is required for competitive PHA production, which can be facilitated by on-line or in-line monitoring devices. To this end, we have evaluated photon density wave (PDW) spectroscopy as a new process analytical technology for Ralstonia eutropha (Cupriavidus necator) H16 plant oil cultivations producing polyhydroxybutyrate (PHB) as an intracellular polymer. PDW spectroscopy was used for in-line recording of the reduced scattering coefficient µs’ and the absorption coefficient µa at 638 nm. A correlation of µs’ with the cell dry weight (CDW) and µa with the residual cell dry weight (RCDW) was observed during growth, PHB accumulation, and PHB degradation phases in batch and pulse feed cultivations. The correlation was used to predict CDW, RCDW, and PHB formation in a high-cell-density fed-batch cultivation with a productivity of 1.65 gPHB·L−1·h−1 and a final biomass of 106 g·L−1 containing 73 wt% PHB. The new method applied in this study allows in-line monitoring of CDW, RCDW, and PHA formation. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Article
Polyhydroxybutyrate Production from Natural Gas in A Bubble Column Bioreactor: Simulation Using COMSOL
Bioengineering 2019, 6(3), 84; https://doi.org/10.3390/bioengineering6030084 - 16 Sep 2019
Cited by 8 | Viewed by 5263
Abstract
In this study, the simulation of microorganism ability for the production of poly-β-hydroxybutyrate (PHB) from natural gas (as a carbon source) was carried out. Based on the Taguchi algorithm, the optimum situations for PHB production from natural gas in the columnar bubble reactor [...] Read more.
In this study, the simulation of microorganism ability for the production of poly-β-hydroxybutyrate (PHB) from natural gas (as a carbon source) was carried out. Based on the Taguchi algorithm, the optimum situations for PHB production from natural gas in the columnar bubble reactor with 30 cm length and 1.5 cm diameter at a temperature of 32 °C was evaluated. So, the volume ratio of air to methane of 50:50 was calculated. The simulation was carried out by COMSOL software with two-dimensional symmetric mode. Mass transfer, momentum, density-time, and density-place were investigated. The maximum production of biomass concentration reached was 1.63 g/L, which shows a 10% difference in contrast to the number of experimental results. Furthermore, the consequence of inlet gas rate on concentration and gas hold up was investigated Andres the simulation results were confirmed to experimental results with less than 20% error. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Article
Biotechnological Production of Poly(3-Hydroxybutyrate-co-4-Hydroxybutyrate-co-3-Hydroxyvalerate) Terpolymer by Cupriavidus sp. DSM 19379
Bioengineering 2019, 6(3), 74; https://doi.org/10.3390/bioengineering6030074 - 26 Aug 2019
Cited by 8 | Viewed by 5101
Abstract
The terpolymer of 3-hydroxybutyrate (3HB), 3-hydroxyvalerate (3HV), and 4-hydroxybutyrate (4HB) was produced employing Cupriavidus sp. DSM 19379. Growth in the presence of γ-butyrolactone, ε-caprolactone, 1,4-butanediol, and 1,6-hexanediol resulted in the synthesis of a polymer consisting of 3HB and 4HB monomers. Single and two-stage [...] Read more.
The terpolymer of 3-hydroxybutyrate (3HB), 3-hydroxyvalerate (3HV), and 4-hydroxybutyrate (4HB) was produced employing Cupriavidus sp. DSM 19379. Growth in the presence of γ-butyrolactone, ε-caprolactone, 1,4-butanediol, and 1,6-hexanediol resulted in the synthesis of a polymer consisting of 3HB and 4HB monomers. Single and two-stage terpolymer production strategies were utilized to incorporate the 3HV subunit into the polymer structure. At the single-stage cultivation mode, γ-butyrolactone or 1,4-butanediol served as the primary substrate and propionic and valeric acid as the precursor of 3HV. In the two-stage production, glycerol was used in the growth phase, and precursors for the formation of the terpolymer in combination with the nitrogen limitation in the medium were used in the second phase. The aim of this work was to maximize the Polyhydroxyalkanoates (PHA) yields with a high proportion of 3HV and 4HB using different culture strategies. The obtained polymers contained 0–29 mol% of 3HV and 16–32 mol% of 4HB. Selected polymers were subjected to a material properties analysis such as differential scanning calorimetry (DSC), thermogravimetry, and size exclusion chromatography coupled with multi angle light scattering (SEC-MALS) for determination of the molecular weight. The number of polymers in the biomass, as well as the monomer composition of the polymer were determined by gas chromatography. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Review

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Review
Biomedical Processing of Polyhydroxyalkanoates
Bioengineering 2019, 6(4), 108; https://doi.org/10.3390/bioengineering6040108 - 29 Nov 2019
Cited by 36 | Viewed by 7661
Abstract
The rapidly growing interest on polyhydroxyalkanoates (PHA) processing for biomedical purposes is justified by the unique combinations of characteristics of this class of polymers in terms of biocompatibility, biodegradability, processing properties, and mechanical behavior, as well as by their great potential for sustainable [...] Read more.
The rapidly growing interest on polyhydroxyalkanoates (PHA) processing for biomedical purposes is justified by the unique combinations of characteristics of this class of polymers in terms of biocompatibility, biodegradability, processing properties, and mechanical behavior, as well as by their great potential for sustainable production. This article aims at overviewing the most exploited processing approaches employed in the biomedical area to fabricate devices and other medical products based on PHA for experimental and commercial applications. For this purpose, physical and processing properties of PHA are discussed in relationship to the requirements of conventionally-employed processing techniques (e.g., solvent casting and melt-spinning), as well as more advanced fabrication approaches (i.e., electrospinning and additive manufacturing). Key scientific investigations published in literature regarding different aspects involved in the processing of PHA homo- and copolymers, such as poly(3-hydroxybutyrate), poly(3-hydroxybutyrate-co-3-hydroxyvalerate), and poly(3-hydroxybutyrate-co-3-hydroxyhexanoate), are critically reviewed. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Review
Production of Polyhydroxyalkanoates and Extracellular Products Using Pseudomonas Corrugata and P. Mediterranea: A Review
Bioengineering 2019, 6(4), 105; https://doi.org/10.3390/bioengineering6040105 - 14 Nov 2019
Cited by 19 | Viewed by 6105
Abstract
Some strains of Pseudomonas corrugata (Pco) and P. mediterranea (Pme) efficiently synthesize medium-chain-length polyhydroxyalkanoates elastomers (mcl-PHA) and extracellular products on related and unrelated carbon sources. Yield and composition are dependent on the strain, carbon source, fermentation process, and any [...] Read more.
Some strains of Pseudomonas corrugata (Pco) and P. mediterranea (Pme) efficiently synthesize medium-chain-length polyhydroxyalkanoates elastomers (mcl-PHA) and extracellular products on related and unrelated carbon sources. Yield and composition are dependent on the strain, carbon source, fermentation process, and any additives. Selected Pco strains produce amorphous and sticky mcl-PHA, whereas strains of Pme produce, on high grade and partially refined biodiesel glycerol, a distinctive filmable PHA, very different from the conventional microbial mcl-PHA, suitable for making blends with polylactide acid. However, the yields still need to be improved and production costs lowered. An integrated process has been developed to recover intracellular mcl-PHA and extracellular bioactive molecules. Transcriptional regulation studies during PHA production contribute to understanding the metabolic potential of Pco and Pme strains. Data available suggest that pha biosynthesis genes and their regulations will be helpful to develop new, integrated strategies for cost-effective production. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Review
Recent Advances in the Use of Polyhydroyalkanoates in Biomedicine
Bioengineering 2019, 6(3), 82; https://doi.org/10.3390/bioengineering6030082 - 12 Sep 2019
Cited by 51 | Viewed by 6796
Abstract
Polyhydroxyalkanoates (PHAs), a family of natural biopolyesters, are widely used in many applications, especially in biomedicine. Since they are produced by a variety of microorganisms, they possess special properties that synthetic polyesters do not have. Their biocompatibility, biodegradability, and non-toxicity are the crucial [...] Read more.
Polyhydroxyalkanoates (PHAs), a family of natural biopolyesters, are widely used in many applications, especially in biomedicine. Since they are produced by a variety of microorganisms, they possess special properties that synthetic polyesters do not have. Their biocompatibility, biodegradability, and non-toxicity are the crucial properties that make these biologically produced thermoplastics and elastomers suitable for their applications as biomaterials. Bacterial or archaeal fermentation by the combination of different carbohydrates or by the addition of specific inductors allows the bioproduction of a great variety of members from the PHAs family with diverse material properties. Poly(3-hydroxybutyrate) (PHB) and its copolymers, such as poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHVB) or poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (PHB4HB), are the most frequently used PHAs in the field of biomedicine. PHAs have been used in implantology as sutures and valves, in tissue engineering as bone graft substitutes, cartilage, stents for nerve repair, and cardiovascular patches. Due to their good biodegradability in the body and their breakdown products being unhazardous, they have also been remarkably applied as drug carriers for delivery systems. As lately there has been considerable and growing interest in the use of PHAs as biomaterials and their application in the field of medicine, this review provides an insight into the most recent scientific studies and advances in PHAs exploitation in biomedicine. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Review
Polyhydroxyalkanoate Biosynthesis at the Edge of Water Activity-Haloarchaea as Biopolyester Factories
Bioengineering 2019, 6(2), 34; https://doi.org/10.3390/bioengineering6020034 - 16 Apr 2019
Cited by 65 | Viewed by 7757
Abstract
Haloarchaea, the extremely halophilic branch of the Archaea domain, encompass a steadily increasing number of genera and associated species which accumulate polyhydroxyalkanoate biopolyesters in their cytoplasm. Such ancient organisms, which thrive in highly challenging, often hostile habitats characterized by salinities between 100 and [...] Read more.
Haloarchaea, the extremely halophilic branch of the Archaea domain, encompass a steadily increasing number of genera and associated species which accumulate polyhydroxyalkanoate biopolyesters in their cytoplasm. Such ancient organisms, which thrive in highly challenging, often hostile habitats characterized by salinities between 100 and 300 g/L NaCl, have the potential to outperform established polyhydroxyalkanoate production strains. As detailed in the review, this optimization presents due to multifarious reasons, including: cultivation setups at extreme salinities can be performed at minimized sterility precautions by excluding the growth of microbial contaminants; the high inner-osmotic pressure in haloarchaea cells facilitates the recovery of intracellular biopolyester granules by cell disintegration in hypo-osmotic media; many haloarchaea utilize carbon-rich waste streams as main substrates for growth and polyhydroxyalkanoate biosynthesis, which allows coupling polyhydroxyalkanoate production with bio-economic waste management; finally, in many cases, haloarchaea are reported to produce copolyesters from structurally unrelated inexpensive substrates, and polyhydroxyalkanoate biosynthesis often occurs in parallel to the production of additional marketable bio-products like pigments or polysaccharides. This review summarizes the current knowledge about polyhydroxyalkanoate production by diverse haloarchaea; this covers the detection of new haloarchaea producing polyhydroxyalkanoates, understanding the genetic and enzymatic particularities of such organisms, kinetic aspects, material characterization, upscaling and techno-economic and life cycle assessment. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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Review
Bioprocess Engineering Aspects of Sustainable Polyhydroxyalkanoate Production in Cyanobacteria
Bioengineering 2018, 5(4), 111; https://doi.org/10.3390/bioengineering5040111 - 18 Dec 2018
Cited by 27 | Viewed by 11020
Abstract
Polyhydroxyalkanoates (PHAs) are a group of biopolymers produced in various microorganisms as carbon and energy reserve when the main nutrient, necessary for growth, is limited. PHAs are attractive substitutes for conventional petrochemical plastics, as they possess similar material properties, along with biocompatibility and [...] Read more.
Polyhydroxyalkanoates (PHAs) are a group of biopolymers produced in various microorganisms as carbon and energy reserve when the main nutrient, necessary for growth, is limited. PHAs are attractive substitutes for conventional petrochemical plastics, as they possess similar material properties, along with biocompatibility and complete biodegradability. The use of PHAs is restricted, mainly due to the high production costs associated with the carbon source used for bacterial fermentation. Cyanobacteria can accumulate PHAs under photoautotrophic growth conditions using CO2 and sunlight. However, the productivity of photoautotrophic PHA production from cyanobacteria is much lower than in the case of heterotrophic bacteria. Great effort has been focused to reduce the cost of PHA production, mainly by the development of optimized strains and more efficient cultivation and recovery processes. Minimization of the PHA production cost can only be achieved by considering the design and a complete analysis of the whole process. With the aim on commercializing PHA, this review will discuss the advances and the challenges associated with the upstream processing of cyanobacterial PHA production, in order to help the design of the most efficient method on the industrial scale. Full article
(This article belongs to the Special Issue Advances in Polyhydroxyalkanoate (PHA) Production, Volume 2)
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