Melt electrowriting (MEW) enables precise scaffold fabrication for biomedical applications. With a limited number of processable materials with short and tunable degradation times, polyhydroxyalkanoates (PHAs) present an interesting option. Here, poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) and a blend of PHBV and poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (PHBV+P34HB) are successfully melt electrowritten into scaffolds with various architectures. PHBV+P34HB exhibits greater thermal stability, making it a superior printing material compared to PHBV in MEW. The PHBV+P34HB scaffolds subjected to enzymatic degradation show tunable degradation times, governed by enzyme dilution, incubation time, and scaffold surface area. PHBV+P34HB scaffolds seeded with human dermal fibroblasts (HDFs), demonstrate enhanced cell adherence, proliferation, and spreading. The HDFs, when exposed to the enzyme solutions and enzymatic degradation residues, show good viability and proliferation rates. Additionally, HDFs grown on enzymatically pre-incubated scaffolds do not show any difference in behavior compared those grown on control scaffolds. It is concluded that PHAs, as biobased materials with enzymatically tunable degradability rates, are an important addition to the already limited set of materials available for MEW technology.
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Multi-layer cell constructs produced in vitro are an innovative treatment option to support the growing demand for therapy in regenerative medicine. Our research introduces a novel construct integrating organ-derived decellularised extracellular matrix (dECM) hydrogels and 3D-printed biodegradable polymer meshes composed of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) and poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (P34HB) to support and maintain multiple layers of different cell types. We achieved that by integrating the mechanical stability of PHBV+P34HB, commonly used in the food storage industry, with a dECM hydrogel, which replicates organ stiffness and supports cellular survival and function. The construct was customised by adjusting the fibre arrangement and pore sizes, making it a suitable candidate for a personalised design. We showed that the polymer is degradable after precoating it with PHB depolymerase (PhaZ), with complete degradation achieved in 3–5 days and delayed by adding the hydrogel to 10 days, enabling tuneable degradation for regenerative medicine applications. Finally, as a proof of concept, we composed a three-layered tissue in vitro; each layer represented a different tissue type: epidermal, vascular, and subcutaneous layers. Possible future applications include wound healing and diabetic ulcer paths, personalised drug delivery systems, and personalised tissue implants.
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The transition to a biobased economy necessitates utilizing renewable resources as a sustainable alternative to traditional fossil fuels. Bioconversion is a way to produce many green chemicals from renewables, e.g., biopolymers like PHAs. However, fermentation and bioconversion processes mostly rely on expensive, and highly refined pure substrates. The utilization of crude fractions from biorefineries, especially herbaceous lignocellulosic feedstocks, could significantly reduce costs. This presentation shows the microbial production of PHA from such a crude stream by a wild-type thermophilic bacterium Schlegelella thermodepolymerans [1]. Specifically, it uses crude xylose-rich fractions derived from a newly developed biorefinery process for grassy biomasses (the ALACEN process). This new stepwise mild flow-through biorefinery approach for grassy lignocellulosic biomass allows the production of various fractions: a fraction containing esterified aromatics, a monomeric xylose-rich stream, a glucose fraction, and a native-like lignin residue [2]. The crude xylose-rich fraction was free of fermentation-inhibiting compounds meaning that the bacterium S.thermodepolymerans could effectively use it for the production of one type of PHA, polyhydroxybutyrate. Almost 90% of the xylose in the refined wheat straw fraction was metabolized with simultaneous production of PHA, matching 90% of the PHA production per gram of sugars, comparable to PHA yields from commercially available xylose. In addition to xylose, S. thermodepolymerans converted oligosaccharides with a xylose backbone (xylans) into fermentable xylose, and subsequently utilized the xylose as a source for PHA production. Since the xylose-rich hydrolysates from the ALACEN process also contain some oligomeric xylose and minor hemicellulose-derived sugars, optimal valorization of the C5-fractions derived from the refinery process can be obtained using S. thermodepolymerans. This opens the way for further exploration of PHA production from C5-fractions out of a variety of herbaceous lignocellulosic biomasses using the ALACEN process combined with S. thermodepolymerans. Overall, the innovative utilization of renewable resources in fermentation technology, as shown herein, makes a solid contribution to the transition to a biobased economy.[1] W. Zhou, D.I. Colpa, H. Permentier, R.A. Offringa, L. Rohrbach, G.J.W. Euverink, J. Krooneman. Insight into polyhydroxyalkanoate (PHA) production from xylose and extracellular PHA degradation by a thermophilic Schlegelella thermodepolymerans. Resources, Conservation and Recycling 194 (2023) 107006, ISSN 0921-3449, https://doi.org/10.1016/j.resconrec.2023.107006. [2] S. Bertran-Llorens, W.Zhou. M.A.Palazzo, D.I.Colpa, G.J.W.Euverink, J.Krooneman, P.J.Deuss. ALACEN: a holistic herbaceous biomass fractionation process attaining a xylose-rich stream for direct microbial conversion to bioplastics. Submitted 2023.
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It is known that several bacteria in sewage treatment plants can produce attractive quantities of biodegradable polymers within their cell walls (up to 80% of the cell weight). These polymers may consist of polyhydroxyalkanoates (PHA), a bioplastic which exhibits interesting characteristics like excellent biodegradation, low melting point and good environmental footprint. PHA bioplastics or PHBV are still quite expensive because cumbersome downstream processing steps of the PHAcontaining bacteria are needed before PHA can be applied in products. In this proposal, the consortium investigates the possibilities for eliminating these expensive and environmentally intensive purification steps, and as a result contribute to speeding up the up-take of PHA production of residual streams by the market. The objective of the project is to investigate the possibilities of direct extrusion of PHAcontaining bacteria and the application opportunities of the extruded PHA. The consortium of experienced partners (Paques Biomaterials, MAAN Group, Ecoras and CoEBBE) will investigate and test the extrusion of different types of PHA-containing biomass, and analyse the products on composition, appearance and mechanical properties. Moreover, the direct extrusion process will be evaluated and compared with conventional PHA extraction and subsequent extrusion. The expected result will be a proof of principle and provide an operational window for the application of direct extrusion with PHA-containing biomass produced using waste streams, either used as such or in blends with purified PHA. Both the opportunities of the direct extrusion process itself as well as the application opportunities of the extruded PHA will be mapped. If the new process leads to a cheaper, more environmentally friendly produced and applicable PHA, the proof of principle developed by the consortium could be the first step in a larger scale development that could help speeding up the implementation of the technology for PHA production from residual streams in the market.
Binnen dit project zal de initiatiefnemer Paques Biomaterials B.V., in samenwerking met de MKB-ondernemingen Maan Glueing Technologies, Senbis Polymer Innovations en de kennisinstellingen Rijksuniversiteit Groningen, Hanzehogeschool Groningen en NHL Stenden Hogeschool werken aan het verbeteren van de aantrekkelijkheid van PHA’s als biobased en biodegradeerbaar polymeer voor tal van toepassingen. Binnen dit JTF-project wordt een deze mismatch tussen vraag en aanbod opgelost waardoor het potentieel van PHA’s voor hoogwaardige toepassingen ontsloten wordt. Er zal hiervoor nieuwe kennis worden ontwikkeld en uitgewisseld op zowel labschaal als een grotere onderzoekschaal van de fermentatie- en extractieprocessen die kunnen sturen op de geselecteerde hydroxyvaloraat-gehalte (HV-gehalte) en molecuulgewicht/viscositeit voor de betreffende applicaties. Paques Biomaterials richt zich binnen dit JTF-project op een gepatenteerd groen oplosmiddelextractieproces voor PHA dat een hoge mate van kwaliteitscontrole mogelijk maakt, vooral bij het gebruik van PHA-biomassa van verschillende herkomst en met verschillende kwaliteiten. Het project zal grotendeels uitgevoerd worden op de nieuwe bedrijfslocatie van Paques Biomaterials, te Emmen. Daarnaast zullen enkele activiteiten ook plaatsvinden op de vestigingslocaties van de samenwerkingspartners, welke overwegend gevestigd zijn in het Noorden.
